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11 pages, 898 KiB  
Article
Epithelial Antimicrobial Peptide/Protein and Cytokine Expression Profiles Obtained from Nasopharyngeal Swabs of SARS-CoV-2-Infected and Non-Infected Subjects
by Thilo Gambichler, Silke Goesmann, Marina Skrygan, Laura Susok, Christian Schütte, Nahza Hamdani and Wolfgang Schmidt
Viruses 2024, 16(9), 1471; https://doi.org/10.3390/v16091471 (registering DOI) - 15 Sep 2024
Viewed by 143
Abstract
Immune responses of the epithelia of the upper respiratory tract are likely crucial in early inhibition of the viral replication and finally clearance of SARS-CoV-2. We aimed to compare the expression profiles of antimicrobial peptides/proteins (AMPs) and related cytokines observed in the nasopharynx [...] Read more.
Immune responses of the epithelia of the upper respiratory tract are likely crucial in early inhibition of the viral replication and finally clearance of SARS-CoV-2. We aimed to compare the expression profiles of antimicrobial peptides/proteins (AMPs) and related cytokines observed in the nasopharynx of SARS-CoV-2-infected patients and non-infected controls and to assess the associations between these parameters and COVID-19 patients’ outcomes. We included 45 subjects who had tested positive for SARS-CoV-2 and 22 control subjects who had tested negative for SARS-CoV-2. Biomaterial for SARS-CoV-2 detection, as well as gene and protein expression studies, was obtained from all subjects using nasopharyngeal swabs which were performed a maximum of 7 days before inclusion in the study. Univariable and multivariable statistics were performed. When compared to the controls, the mRNA expression levels of human β-defensin 1 (hBD-1), LL-37, and trappin-2 were significantly higher in specimens of nasopharyngeal swabs from COVID-19 patients. Protein expression of hBD-1 was also increased in the COVID-19 group. mRNA expression levels of interferon-ɣ (IFN-ɣ), tumor necrosis factor- ɑ (TNF-ɑ), and interleukin-6 (IL-6) measured in SARS-CoV-2-infected patients were significantly higher than those observed in the controls, which could also be confirmed in the protein levels of IFN-ɣ and IL-6. A significant correlation between mRNA and protein levels could be observed only for IL-6. Univariable analysis revealed that low IFN-ɣ mRNA levels were associated with severe/fatal outcomes. The occurrence of COVID-19 pneumonia was significantly associated with lower expression levels of IL-6 mRNA, IFN-ɣ mRNA, and TNF-ɑ mRNA. Concerning the severe/fatal outcomes, the multivariable logistic regression model revealed that none of the aforementioned parameters remained significant in the model. However, the logistic regression model revealed that higher TNF-ɑ mRNA expression was a significant independent predictor of absence of pneumonia [odds ratio: 0.35 (95% CI 0.14 to 0.88, p = 0.024)]. In conclusion, nasopharyngeal expression of AMPs (hBD-1, LL-37, and trappin-2) and cytokines (IL-6, IFN-ɣ, and TNF-ɑ) is upregulated in response to early SARS-CoV-2 infection, indicating that these AMPs and cytokines play a role in the local host defense against the virus. Upregulated nasopharyngeal TNF-ɑ mRNA expression during the early phase of SARS-CoV-2 infection was a significant independent predictor of the absence of COVID-19 pneumonia. Hence, high TNF-ɑ mRNA expression in the nasopharynx appears to be a protective factor for lung complications in COVID-19 patients. Full article
(This article belongs to the Special Issue Coronaviruses Pathogenesis, Immunity, and Antivirals)
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<p>Showing an ROC analysis indicating that lower tumor necrosis factor-α mRNA levels (criterion: ≤1.64, AUC 0.75, <span class="html-italic">p</span> = 0.0008, Youden index 0.45) obtained by nasopharyngeal swabs are associated with the occurrence of COVID-19 pneumonia.</p>
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18 pages, 6280 KiB  
Systematic Review
In Vitro Bond Strength of Dentin Treated with Sodium Hypochlorite: Effects of Antioxidant Solutions
by Guillermo Grazioli, Elisa de León Cáceres, Romina Tessore, Rafel Guerra Lund, Ana Josefina Monjarás-Ávila, Monika Lukomska-Szymanska, Louis Hardan, Rim Bourgi and Carlos Enrique Cuevas-Suárez
Antioxidants 2024, 13(9), 1116; https://doi.org/10.3390/antiox13091116 (registering DOI) - 14 Sep 2024
Viewed by 294
Abstract
This systematic review aims to evaluate whether the application of antioxidant solutions can enhance the bond strength of resin-based materials to sodium hypochlorite (NaOCl)-treated dentin. This study follows the PICOT strategy: population (sodium hypochlorite-treated dentin), intervention (application of antioxidants), control (distilled water), outcome [...] Read more.
This systematic review aims to evaluate whether the application of antioxidant solutions can enhance the bond strength of resin-based materials to sodium hypochlorite (NaOCl)-treated dentin. This study follows the PICOT strategy: population (sodium hypochlorite-treated dentin), intervention (application of antioxidants), control (distilled water), outcome (bond strength), and type of studies (in vitro studies). The systematic review and meta-analysis were conducted following PRISMA guidelines. Electronic databases were searched for in vitro studies evaluating the effects of antioxidants on bond strength to sodium hypochlorite-treated dentin. Two independent reviewers screened articles, extracted data, and assessed risk of bias. Meta-analyses were performed using a random-effects model to compare standardized mean differences in bond strength between antioxidant pretreatment and control groups. Inclusion criteria consisted of in vitro studies that examined the bond strength of resin-based materials to NaOCl-treated dentin with antioxidant application, while exclusion criteria included studies with incomplete data, those not using a control group, or those that did not directly measure bond strength. From 3041 initial records, 29 studies were included in the qualitative analysis and 25 in the meta-analysis. Ascorbic acid, sodium ascorbate, grape seed extract, green tea, and rosmarinic acid significantly improved bond strength to sodium hypochlorite-treated dentin (p < 0.05). The effectiveness of grape seed extract varied with adhesive system type. Hesperidin, p-toluene sulfonic acid, and sodium thiosulfate did not significantly improve bond strength. Most studies had a high risk of bias. This suggests that the conclusions drawn from these studies should be interpreted with caution, and further research with more robust methodologies may be needed to confirm the findings. In conclusion, this systematic review implies that certain antioxidants can improve bond strength to sodium hypochlorite-treated dentin, with efficacy depending on the specific agent and adhesive system used. Further standardized studies are needed to optimize protocols and confirm these findings. Full article
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<p>PRISMA flow diagram of study selection process.</p>
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<p>Forest plot showing the effect of the application of ascorbic acid as pretreatment on the bond strength to sodium hypochlorite-treated dentin.</p>
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<p>Forest plot showing the effect of the application of grape seed as pretreatment on the bond strength to sodium hypochlorite-treated dentin.</p>
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<p>Forest plot showing the effect of the application of green tea as pretreatment on the bond strength to sodium hypochlorite-treated dentin.</p>
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<p>Forest plot showing the effect of the application of rosmarinic acid as pretreatment on the bond strength to sodium hypochlorite-treated dentin.</p>
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<p>Forest plot showing the effect of the application of hesperidin as pretreatment on the bond strength to sodium hypochlorite-treated dentin.</p>
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<p>Forest plot showing the effect of the application of <span class="html-italic">p-toluene</span> sulfonic acid as pretreatment on the bond strength to sodium hypochlorite-treated dentin.</p>
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<p>Forest plot showing the effect of the application of sodium thiosulfate as pretreatment on the bond strength to sodium hypochlorite-treated dentin.</p>
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<p>Forest plot showing the effect of the application of sodium ascorbate as pretreatment on the bond strength to sodium hypochlorite-treated dentin.</p>
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12 pages, 4220 KiB  
Article
Experimental Investigation of Full Hole Embedment Behavior of Bamboo Scrimber with Dowel-Type Fasteners
by Yanyan Liu, Xiaoyu Huang and Siyuan Tang
Buildings 2024, 14(9), 2909; https://doi.org/10.3390/buildings14092909 (registering DOI) - 14 Sep 2024
Viewed by 188
Abstract
A comprehensive understanding of the embedment behavior is of great importance in the design of contemporary bamboo constructions with connections utilizing dowel-type fasteners. The objective of this research was to assess the embedment behavior of bamboo scrimber using full-hole embedment tests. To investigate [...] Read more.
A comprehensive understanding of the embedment behavior is of great importance in the design of contemporary bamboo constructions with connections utilizing dowel-type fasteners. The objective of this research was to assess the embedment behavior of bamboo scrimber using full-hole embedment tests. To investigate the effect of the loading angle and bolt diameter, a series of tests were performed using bolts of varying diameters (16 mm, 18 mm, and 20 mm) and loading angles (0° to 90°, with an increment of 15°). The experimental results demonstrated that the loading angle has a considerable influence on the embedment behavior. As the loading angle was increased, the failure mode underwent a change from a brittle failure mode, which was dominated by shear mechanisms, to a ductile failure mode, which was dominated by fiber crushing. The yield and ultimate embedment strengths showed an M-shaped response to changes in the loading angle, with the lowest values being 0°, 45°, and 90°. The bolt diameter was found to have no impact on the failure mode of the specimen. However, an increase in bolt diameter resulted in a reduction in the embedment strength when the specimen was loaded at 90°. Full article
(This article belongs to the Special Issue Research on Seismic Performance of Timber/Bamboo Buildings)
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<p>Test set-up for the evaluation of the embedment strength: (<b>a</b>) half-hole compression test; (<b>b</b>) full-hole compression test. The symbol ‘d’ is used to denote the diameter of the fastener.</p>
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<p>Schematic diagram of the full-hole embedment test.</p>
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<p>Schematic illustration of the yield and ultimate embedment strengths.</p>
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<p>Failure modes of all specimens with a dowel diameter of 18 mm: (<b>a</b>) loading angle of 0°; (<b>b</b>) loading angle of 15°; (<b>c</b>) loading angle of 30°; (<b>d</b>) loading angle of 45°; (<b>e</b>) loading angle of 60°; (<b>f</b>) loading angle of 75°; (<b>g</b>) loading angle of 90°.</p>
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<p>Embedment load–displacement curves for different bolt diameters: (<b>a</b>) <span class="html-italic">d</span> = 16 mm and <math display="inline"><semantics> <mi>θ</mi> </semantics></math> = 0°; (<b>b</b>) <span class="html-italic">d</span> = 16 mm and <math display="inline"><semantics> <mi>θ</mi> </semantics></math> = 90°; (<b>c</b>) <span class="html-italic">d</span> = 18 mm and <math display="inline"><semantics> <mi>θ</mi> </semantics></math> = 0°; (<b>d</b>) <span class="html-italic">d</span> = 18 mm and <math display="inline"><semantics> <mi>θ</mi> </semantics></math> = 90°; (<b>e</b>) <span class="html-italic">d</span> = 20 mm and <math display="inline"><semantics> <mi>θ</mi> </semantics></math> = 0°; (<b>f</b>) <span class="html-italic">d</span> = 20 mm and <math display="inline"><semantics> <mi>θ</mi> </semantics></math> = 90°.</p>
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<p>Embedment load–displacement curves for varying load-to-grain angles (<span class="html-italic">d</span> = 18 mm).</p>
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<p>Embedment strength for varying load-to-grain angles: (<b>a</b>) yield embedment strength; (<b>b</b>) ultimate embedment strength.</p>
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<p>Schematic diagram of the stress occurring in specimens under different loading angles.</p>
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21 pages, 3419 KiB  
Article
Novel Bioplastic Based on PVA Functionalized with Anthocyanins: Synthesis, Biochemical Properties and Food Applications
by Giuseppe Tancredi Patanè, Antonella Calderaro, Stefano Putaggio, Giovanna Ginestra, Giuseppina Mandalari, Santa Cirmi, Davide Barreca, Annamaria Russo, Teresa Gervasi, Giovanni Neri, Meryam Chelly, Annamaria Visco, Cristina Scolaro, Francesca Mancuso, Silvana Ficarra, Ester Tellone and Giuseppina Laganà
Int. J. Mol. Sci. 2024, 25(18), 9929; https://doi.org/10.3390/ijms25189929 (registering DOI) - 14 Sep 2024
Viewed by 215
Abstract
Over the last ten years, researchers’ efforts have aimed to replace the classic linear economy model with the circular economy model, favoring green chemical and industrial processes. From this point of view, biologically active molecules, coming from plants, flowers and biomass, are gaining [...] Read more.
Over the last ten years, researchers’ efforts have aimed to replace the classic linear economy model with the circular economy model, favoring green chemical and industrial processes. From this point of view, biologically active molecules, coming from plants, flowers and biomass, are gaining considerable value. In this study, firstly we focus on the development of a green protocol to obtain the purification of anthocyanins from the flower of Callistemon citrinus, based on simulation and on response surface optimization methodology. After that, we utilize them to manufacture and add new properties to bioplastics belonging to class 3, based on modified polyvinyl alcohol (PVA) with increasing amounts from 0.10 to 1.00%. The new polymers are analyzed to monitor morphological changes, optical properties, mechanical properties and antioxidant and antimicrobial activities. Fourier transform infrared spectroscopy (FTIR) spectra of the new materials show the characteristic bands of the PVA alone and a modification of the band at around 1138 cm−1 and 1083 cm−1, showing an influence of the anthocyanins’ addition on the sequence with crystalline and amorphous structures of the starting materials, as also shown by the results of the mechanical tests. These last showed an increase in thickening (from 29.92 μm to approx. 37 μm) and hydrophobicity with the concomitant increase in the added anthocyanins (change in wettability with water from 14° to 31°), decreasing the poor water/moisture resistance of PVA that decreases its strength and limits its application in food packaging, which makes the new materials ideal candidates for biodegradable packaging to extend the shelf-life of food. The functionalization also determines an increase in the opacity, from 2.46 to 3.42 T%/mm, the acquisition of antioxidant activity against 2,2-diphenyl-1-picrylhdrazyl and 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) radicals and, in the ferric reducing power assay, the antimicrobial (bactericidal) activity against different Staphylococcus aureus strains at the maximum tested concentration (1.00% of anthocyanins). On the whole, functionalization with anthocyanins results in the acquisition of new properties, making it suitable for food packaging purposes, as highlighted by a food fresh-keeping test. Full article
(This article belongs to the Section Materials Science)
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<p>Representative pictogram for the green index of the extraction of anthocyanin from <span class="html-italic">Callistemon citrinus</span> using two different methods. (<b>A</b>) The index for the extraction with accelerated solvent extraction using methanol solution; (<b>B</b>) The index for the extraction with MAE using the new ethanol (EtOH) solution. In both pictograms, the colour scale (red-yellow-green) indicates the performance at each stage of the procedure. The less chemically ‘green’ the process, the more red it appears.</p>
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<p>Pareto chart diagram for the extraction of total anthocyanin content (TAC) from <span class="html-italic">Callistemon citrinus</span>. A = microwave power (W); B = extraction time (min); C = EtOH% in the extraction solution.</p>
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<p>Response surface plots analysis for the total anthocyanin content yield (TAC) from <span class="html-italic">Callistemon citrinus</span> powder with microwave assisted extraction (MAE) with the same solid-liquid ratio, 1:10 (<span class="html-italic">w</span>/<span class="html-italic">v</span>). (<b>A</b>) microwave power and EtOH % in the reaction mix; (<b>B</b>) microwave power and minutes; (<b>C</b>) EtOH % in the reaction mix and minutes.</p>
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<p>Representative FTIR spectra of the new PVA-based bioplastics produced by the addition of increasing amount of anthocyanins (0.0–1.0%). (<b><span style="color:lime">―</span></b>) Anthocyanins powder; (<b>―</b>) PVA alone; (<b><span style="color:red">―</span></b>) PVA plus 0.1% anthocyanins; (<b><span style="color:#2E74B5">―</span></b>) PVA plus 0.25% anthocyanins; (<b><span style="color:#FF66FF">―</span></b>) PVA plus 0.5% anthocyanins; (<b><span style="color:#66FFFF">―</span></b>) PVA plus 1.0% anthocyanins.</p>
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<p>Release for short- and long-term migration of anthocyanins from PVA films in different food simulants. (<b>A</b>) PVA plus 0.10% of anthocyanins; (<b>B</b>) PVA plus 0.25% of anthocyanins; (<b>C</b>) PVA plus 0.50% of anthocyanins; (<b>D</b>) PVA plus 1.00% of anthocyanins. (<span style="color:#3366CC">⬤</span>) H<sub>2</sub>O; (<span style="color:red">■</span>) ethanol 10%; (<span style="color:green">▲</span>) ethanol 50%; (<span style="color:#7100E2">▼</span>) acetic acid 3%.</p>
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<p>Evaluation of antioxidant activity of PVA-based bioplastics with different % of anthocyanins (0.10, 0.25, 0.50, 1%) in the most common antioxidant assays. (<b>A</b>) ABTS assay; (<b>B</b>) DPPH assay; (<b>C</b>) ferric reducing power (FRAP) assay. The letters in the different graph indicate: a, control sample; b, PVA alone; c, PVA plus 0.10% of anthocyanins; d, PVA plus 0.25% of anthocyanins; e, PVA plus 0.50% of anthocyanins; f, PVA plus 1.00% of anthocyanins. *** <span class="html-italic">p</span> &lt; 0.001.</p>
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<p>Preparation of bags for food packaging produced with PVA plus 1.0% of anthocyanins and its utilization for apple samples.</p>
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<p>Analysis of the changes in apple samples packed or not packed after different intervals of time (0 and 72 h). (<b>A</b>) Changes in the browning of the samples monitored at 420 nm. The letters in the graph indicate: a, apple samples not packed after 0 h; b, apple samples not packed after 72 h; c, apple sample packed with PVA film alone after 72 h; d, apple samples packed with PVA plus 1.00% of anthocyanins after 72 h. (<b>B</b>) Changes in the antioxidant potential monitored by DPPH assay. The letters in the graph indicate: a, control without samples; b, apple samples not packed after 0 h; c, apple samples not packed after 72 h; d, apple sample packed with PVA film alone after 72 h; e, apple samples packed with PVA plus 1.00% of anthocyanins after 72 h. The ** indicates significant changes with respect to the control at <span class="html-italic">p</span> &gt; 0.05.</p>
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14 pages, 2997 KiB  
Article
Lactic Acid Bacterial Fermentation of Esterified Agave Fructans in Simulated Physicochemical Colon Conditions for Local Delivery of Encapsulated Drugs
by Carmen Miramontes-Corona, Abraham Cetina-Corona, María Esther Macías-Rodríguez, Alfredo Escalante, Rosa Isela Corona-González and Guillermo Toriz
Fermentation 2024, 10(9), 478; https://doi.org/10.3390/fermentation10090478 (registering DOI) - 14 Sep 2024
Viewed by 179
Abstract
Understanding drug release in the colon is fundamental to developing efficient treatments for colon-related diseases, while unraveling the relationship between the colonic microbiota and excipients is crucial to unveiling the effect of biomaterials on the release of drugs. In this contribution, the bio-release [...] Read more.
Understanding drug release in the colon is fundamental to developing efficient treatments for colon-related diseases, while unraveling the relationship between the colonic microbiota and excipients is crucial to unveiling the effect of biomaterials on the release of drugs. In this contribution, the bio-release of ibuprofen (encapsulated in acetylated and palmitoylated agave fructans) was evaluated by fermentation with lactic acid bacteria in simulated physicochemical (pH and temperature) colon conditions. It was observed that the size of the acyl chain (1 in acetyl and 15 in palmitoyl) was critical both in the growth of the microorganisms and in the release of the drug. For example, both the bacterial growth and the release of ibuprofen were more favored with acetylated fructan microspheres. Among the microorganisms evaluated, Bifidobacterium adolescentis and Lactobacillus brevis showed great potential as probiotics useful to release drugs from modified fructans. The production of short-chain fatty acids (lactic, acetic, and propionic acids) in the course of fermentations was also determined, since such molecules have a positive effect both on colon-related diseases and on the regulation of the intestinal microbiota. It was found that a higher concentration of acetate is related to a lower growth of bacteria and less release of ibuprofen. Full article
(This article belongs to the Special Issue Fermentation: 10th Anniversary)
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<p>(<b>A</b>) A model of agave fructan according to [<a href="#B33-fermentation-10-00478" class="html-bibr">33</a>]; in native fructan, R is OH; in palmitoylated fructan, 4 Rs are substituted with palmitoyl moieties; for acetylated fructan, about 62 Rs should be acetyl groups. (<b>B</b>) <sup>1</sup>H NMR spectra: (<b>a</b>) native fructan analyzed in D<sub>2</sub>O; H-C* denotes the proton at the anomeric carbon in glucose (<b>b</b>) palmitoylated fructan obtained in CDCl<sub>3</sub>; (<b>c</b>) acetylated fructans analyzed in <sup>d6</sup>DMSO.</p>
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<p>Scanning Electron Micrographs of (<b>a</b>) acetylated (7920X) and (<b>b</b>) palmitoylated fructan (1750X).</p>
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<p>Growth profile of strains and their consortium obtained by optical density of (<b>a</b>) acetylated fructan microspheres and (<b>b</b>) palmitoylated fructan microspheres: <span class="html-italic">B. adolescentis</span> (●); <span class="html-italic">Weisella paramesenteroides</span> (◆); <span class="html-italic">Enterococcus mundtii</span> (◼); <span class="html-italic">Lactobacillus brevis</span> (▲); <span class="html-italic">consortium</span> (∗).</p>
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<p>Percentage of ibuprofen release as function of fermentation time from (<b>a</b>) acetylated and (<b>b</b>) palmitoylated fructan microspheres with <span class="html-italic">B. adolescentis</span> (●); <span class="html-italic">Weisella paramesenteroides</span> (◆); <span class="html-italic">Enterococcus mundtii</span> (◼); <span class="html-italic">Lactobacillus brevis</span> (▲); and <span class="html-italic">consortium</span> (∗).</p>
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<p>Production of SCFA (g/L) by lactic acid bacteria at 48 h of fermentation: (<b>a</b>) acetylated fructan microspheres and (<b>b</b>) palmitoylated fructan microspheres, (■) lactic acid, (□) acetic acid, and (<span class="html-fig-inline" id="fermentation-10-00478-i001"><img alt="Fermentation 10 00478 i001" src="/fermentation/fermentation-10-00478/article_deploy/html/images/fermentation-10-00478-i001.png"/></span>) propionic acid. BA: <span class="html-italic">B. adolescentis</span>; WP: <span class="html-italic">Weisella paramesenteroides</span>; EL: <span class="html-italic">Enterococcus mundtii</span>; LB: <span class="html-italic">Lactobacillus brevis</span>; C: <span class="html-italic">consortium</span>.</p>
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<p>Phylogenetic tree for identification of <span class="html-italic">Weissella paramesenteroides</span> (Jal1), <span class="html-italic">Enterococcus mundtii</span> (BT5inv), and <span class="html-italic">Lactobacillus brevis</span> (Col18) isolated from carposphere of tomato. The red boxes indicate the identified strains in the phylogenetic tree.</p>
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3 pages, 163 KiB  
Editorial
Functional Biomaterials and Biomaterial Composites with Antimicrobial Properties
by John H. T. Luong
J. Funct. Biomater. 2024, 15(9), 267; https://doi.org/10.3390/jfb15090267 (registering DOI) - 14 Sep 2024
Viewed by 179
Abstract
AMR occurs when bacteria, viruses, fungi, and parasites no longer respond to antimicrobial medicines, including antibiotics, antivirals, antifungals, and antiparasitics [...] Full article
19 pages, 10442 KiB  
Article
Comparison of Bioengineered Scaffolds for the Induction of Osteochondrogenic Differentiation of Human Adipose-Derived Stem Cells
by Elena Fiorelli, Maria Giovanna Scioli, Sonia Terriaca, Arsalan Ul Haq, Gabriele Storti, Marta Madaghiele, Valeria Palumbo, Ermal Pashaj, Fabio De Matteis, Diego Ribuffo, Valerio Cervelli and Augusto Orlandi
Bioengineering 2024, 11(9), 920; https://doi.org/10.3390/bioengineering11090920 (registering DOI) - 14 Sep 2024
Viewed by 353
Abstract
Osteochondral lesions may be due to trauma or congenital conditions. In both cases, therapy is limited because of the difficulty of tissue repair. Tissue engineering is a promising approach that relies on designed scaffolds with variable mechanical attributes to favor cell attachment and [...] Read more.
Osteochondral lesions may be due to trauma or congenital conditions. In both cases, therapy is limited because of the difficulty of tissue repair. Tissue engineering is a promising approach that relies on designed scaffolds with variable mechanical attributes to favor cell attachment and differentiation. Human adipose-derived stem cells (hASCs) are a very promising cell source in regenerative medicine with osteochondrogenic potential. Based on the assumption that stiffness influences cell commitment, we investigated three different scaffolds: a semisynthetic animal-derived GelMA hydrogel, a combined scaffold made of rigid PEGDA coated with a thin GelMA layer and a decellularized plant-based scaffold. We investigated the role of different biomechanical stimulations in the scaffold-induced osteochondral differentiation of hASCs. We demonstrated that all scaffolds support cell viability and spontaneous osteochondral differentiation without any exogenous factors. In particular, we observed mainly osteogenic commitment in higher stiffness microenvironments, as in the plant-based one, whereas in a dense and softer matrix, such as in GelMA hydrogel or GelMA-coated-PEGDA scaffold, chondrogenesis prevailed. We can induce a specific cell commitment by combining hASCs and scaffolds with particular mechanical attributes. However, in vivo studies are needed to fully elucidate the regenerative process and to eventually suggest it as a potential approach for regenerative medicine. Full article
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<p>GelMA scaffold characterization and biomechanical test. (<b>A</b>) Hydrogels made with 10% and 15% of GelMA. (<b>B</b>,<b>C</b>) SEM imaging of 10% and 15% GelMA hydrogel, respectively. (<b>D</b>) Stress and strain graph showing a representative elastic modulus for 10% and 15% GelMA hydrogels.</p>
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<p>PEGDA scaffold production. (<b>A</b>) The final PEGDA scaffold with GelMA coating. (<b>B</b>) SEM images of PEGDA scaffold (GelMA 0.5%) showing its ultrastructure and porosity. (<b>C</b>) Stress and strain graph showing a representative elastic modulus for PEGDA-0,5% GelMA and PEGDA-5% GelMA.</p>
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<p>(<b>A</b>) Celery-based scaffold after decellularization and sterilization in 70% ethanol. (<b>B</b>) SEM images of decellularized celery-based scaffolds (24 h and 72 h SDS protocol). (<b>C</b>) Representative trends of stress and strain (0–5% strain) comparing different decellularized protocols (24 h and 72 h) and cut orientation (longitudinally cut and transversal cut).</p>
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<p>Degradation tests. (<b>A</b>) Representative images of 10% and 15% GelMA hydrogel degradation with significant differences at T3 and T4. (<b>B</b>) Graph showing the degradation trend of 10% and 15% GelMA hydrogel in 0.5% collagenase at different points (T, 15 min intervals). (<b>C</b>) Representative images of PEGDA-0.5% GelMA scaffold degradation at T0 (baseline) and T2 (20 h of incubation). (<b>D</b>) Graphs showing the degradation trend of PEGDA scaffold in H2O, 1N NAOH and 1N HCL, and (<b>E</b>) the comparison between PEGDA-0.5% GelMA and PEGDA-5% GelMA in 1N NAOH. The different time points are: T0 (baseline), T1 (18 h of incubation), T2 (20 h of incubation) and T3 (44 h of incubation). (<b>F</b>,<b>G</b>) Degradation test graphs of celery-based scaffolds, (<b>F</b>) cut longitudinally or (<b>G</b>) transversally, incubated with H2O, 1N NAOH and 1N HCL, respectively. T0, T1, T2, T3 and T4 stands for baseline, 2, 4, 6 and 8 weeks. Results are reported as mean ± SEM of <span class="html-italic">n</span> = 3 samples/groups. T test: * <span class="html-italic">p</span> &lt; 0.05 and ** <span class="html-italic">p</span> &lt; 0.001; § <span class="html-italic">p</span> &lt; 0.0001 and §§ <span class="html-italic">p</span> &lt; 0.00001.</p>
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<p>Swelling tests. (<b>A</b>) Representative images of 10% and 15% GelMA hydrogel before and after the swelling test at T0 (baseline) and T4 (20 min). (<b>B</b>) Graph showing the swelling trend of 10% and 15% GelMA hydrogel, each time point (T) is a 5 min interval. (<b>C</b>) Representative images of PEGDA-0.5% GelMA scaffold before and after the swelling degree at T0 (baseline) and T4 (20 min). (<b>D</b>) Graph showing the swelling trend of PEGDA-0.5% GelMA, PEGDA-5% GelMA and PEGDA-0% GelMA scaffold for each time point (T, 5 min intervals). (<b>E</b>) Representative images of celery-based scaffolds cut longitudinally and transversally dried (T0, baseline) and hydrated (T4, 40 min). (<b>F</b>) Graph showing the swelling trend of celery-based scaffolds cut longitudinally or transversally at different time points (T, 10 min-intervals). Since the standard size of scaffold was too light and did not weigh enough to permit a precise measurement, a bigger scaffold was made purposely to perform this test. Results are reported as mean ± SEM of <span class="html-italic">n</span> = 3 samples/group.</p>
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<p>hASC survival and differentiation in GelMA hydrogels. (<b>A</b>) CCK8 assay of hASCs encapsulated in the GelMA hydrogel at different time points (1 week intervals). Results are reported as mean ± SEM of <span class="html-italic">n</span> = 3 samples/group. (<b>B</b>) Representative confocal imaging of Dead/live fluorescence staining of hASCs in the GelMA hydrogel after 3 weeks of culture. Red cells are dead while green cells are alive. (<b>C</b>) Alcian blue staining of GelMA hydrogel after 24 h and 3 weeks of culture. (<b>D</b>) Alizarin red staining of GelMA hydrogel after 24 h and 3 weeks of culture. (<b>E</b>,<b>F</b>) Confocal imaging of immunofluorescence for COL2A1 (green) and OCN (red) in hASCs encapsulated in GelMA hydrogel after 3 weeks of culture.</p>
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<p>hASC survival and differentiation in PEGDA scaffold. (<b>A</b>) CCK8 assay of hASCs seeded in the PEGDA scaffold at different time points (1 week intervals). Results are reported as mean ± SEM of <span class="html-italic">n</span> = 3 samples/group. T test: * <span class="html-italic">p</span> &lt; 0.05. (<b>B</b>) Representative confocal imaging of Dead/live fluorescence staining of hASCs seeded in the PEGDA scaffold after 3 weeks of culture. (<b>C</b>) Confocal imaging of immunofluorescence for COL2A1 (green) and OCN (red) in hASCs seeded in the PEGDA scaffold after 3 weeks of culture.</p>
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<p>hASC survival and differentiation in celery-based scaffold. (<b>A</b>) SEM imaging of a single hASC inside a niche of the celery-based scaffold. (<b>B</b>) CCK8 assay of hASCs seeded in the celery-based scaffold at different time points (1 week intervals). Results are reported as mean ± SEM of <span class="html-italic">n</span> = 3 samples/group. T test: * <span class="html-italic">p</span> &lt; 0.05. (<b>C</b>) Representative confocal imaging of Dead/live fluorescence staining of hASCs seeded in the scaffold after 3 weeks of culture. (<b>D</b>) Confocal 3D stack and (<b>E</b>) the projection of hASC distribution inside the scaffold. (<b>F</b>) Confocal imaging of immunofluorescence for COL2A1 (green) and OCN (red) in hASCs seeded in the PEGDA scaffold after 3 weeks of culture.</p>
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23 pages, 4268 KiB  
Article
In Vitro Wound-Healing Potential of Phenolic and Polysaccharide Extracts of Aloe vera Gel
by Andreea Iosageanu, Elena Mihai, Ana-Maria Seciu-Grama, Elena Utoiu, Alexandra Gaspar-Pintiliescu, Florentina Gatea, Anisoara Cimpean and Oana Craciunescu
J. Funct. Biomater. 2024, 15(9), 266; https://doi.org/10.3390/jfb15090266 (registering DOI) - 13 Sep 2024
Viewed by 250
Abstract
The present study aimed to conduct a comparative investigation of the biological properties of phenolic and polysaccharide extracts obtained using an ultrasound-assisted technique from Aloe vera gel and their effects on each stage of the wound healing process in in vitro experimental models. [...] Read more.
The present study aimed to conduct a comparative investigation of the biological properties of phenolic and polysaccharide extracts obtained using an ultrasound-assisted technique from Aloe vera gel and their effects on each stage of the wound healing process in in vitro experimental models. HPLC analysis showed that the phenolic extract contained aloin, ferulic, and caffeic acid, as well as quercetin dihydrate, as major compounds. Capillary zone electrophoresis indicated the prevalence of mannose and glucose in the polysaccharide extract. Cell culture testing revealed the anti-inflammatory properties of the phenolic extract at a concentration of 0.25 mg/mL through significant inhibition of pro-inflammatory cytokines—up to 28% TNF-α and 11% IL-8 secretion—in inflamed THP-1-derived macrophages, while a pro-inflammatory effect was observed at 0.5 mg/mL. The phenolic extract induced 18% stimulation of L929 fibroblast proliferation at a concentration of 0.5 mg/mL, enhanced the cell migration rate by 20%, and increased collagen type I synthesis by 18%. Moreover, the phenolic extract exhibited superior antioxidant properties by scavenging free DPPH (IC50 of 2.50 mg/mL) and ABTS (16.47 mM TE/g) radicals, and 46% inhibition of intracellular reactive oxygen species (ROS) production was achieved. The polysaccharide extract demonstrated a greater increase in collagen synthesis up to 25%, as well as antibacterial activity against Staphylococcus aureus with a bacteriostatic effect at 25 mg/mL and a bactericidal one at 50 mg/mL. All these findings indicate that the phenolic extract might be more beneficial in formulations intended for the initial phases of wound healing, such as inflammation and proliferation, while the polysaccharide extract could be more suitable for use during the remodeling stage. Moreover, they might be combined with other biomaterials, acting as efficient dressings with anti-inflammatory, antioxidant, and antibacterial properties for rapid recovery of chronic wounds. Full article
(This article belongs to the Special Issue Biomaterials for Wound Healing and Tissue Repair)
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<p>Scheme of the preparation of phenolic and polysaccharide extracts from <span class="html-italic">Aloe vera</span> gel using UAE.</p>
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<p>Quantification of TNF-α (<b>a</b>) and IL-8 (<b>b</b>) levels in LPS-stimulated THP-1-derived macrophages (light grey) and after treatment with phenolic and polysaccharide extract of <span class="html-italic">Aloe vera</span> gel. Untreated cells served as the control group (dark grey). * <span class="html-italic">p</span> &lt; 0.05, # <span class="html-italic">p</span> &lt; 0.01.</p>
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<p>Viability of L929 fibroblasts at 24 (<b>a</b>) and 48 (<b>b</b>) h of treatment with phenolic and polysaccharide UAE extracts of <span class="html-italic">Aloe vera</span> gel, determined by Neutral Red assay. * <span class="html-italic">p</span> &lt; 0.05, # <span class="html-italic">p</span> &lt; 0.01.</p>
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<p>Phase contrast images showing the cell morphology of L929 fibroblasts at 48 h of treatment with phenolic and polysaccharide UAE extracts of <span class="html-italic">Aloe vera</span> gel. Scale bar = 50 μm.</p>
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<p>(<b>a</b>) Phase contrast images showing cell migration of L929 cells treated with phenolic and polysaccharide UAE extracts of <span class="html-italic">Aloe vera</span> gel for 24 h in a scratch model assay. Scale bar = 100 µm; (<b>b</b>) cell migration rate determined by ImageJ analysis. * <span class="html-italic">p</span> &lt; 0.05, # <span class="html-italic">p</span> &lt; 0.01.</p>
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<p>Collagen type I production in L929 fibroblasts after 72 h of treatment with phenolic and polysaccharide UAE extracts of <span class="html-italic">Aloe vera</span> gel, as determined by ELISA. * <span class="html-italic">p</span> &lt; 0.05, # <span class="html-italic">p</span> &lt; 0.01.</p>
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<p>(<b>a</b>) Flow cytometry histograms showing count vs. fluorescence intensity in L929 fibroblasts after 48 h of treatment with phenolic and polysaccharide UAE extracts of <span class="html-italic">Aloe vera</span> gel (red—t-BHP treated cells, green—ascorbic acid treated cells, purple - aloe extracts treated cells). (<b>b</b>) Percentage of intracellular ROS production. * <span class="html-italic">p</span> &lt; 0.05, # <span class="html-italic">p</span> &lt; 0.01.</p>
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<p>Bacterial growth of <span class="html-italic">S. aureus</span> (<b>a</b>) and <span class="html-italic">P. aeruginosa</span> (<b>b</b>) in the presence of the phenolic and polysaccharide UAE extracts of <span class="html-italic">Aloe vera</span> gel. * <span class="html-italic">p</span> &lt; 0.05, # <span class="html-italic">p</span> &lt; 0.01.</p>
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13 pages, 3015 KiB  
Article
Applicability of Paper and Pulp Industry Waste for Manufacturing Mycelium-Based Materials for Thermoacoustic Insulation
by Hugo Muñoz, Paulo Molina, Ignacio A. Urzúa-Parra, Diego A. Vasco, Magdalena Walczak, Gonzalo Rodríguez-Grau, Francisco Chateau and Mamié Sancy
Sustainability 2024, 16(18), 8034; https://doi.org/10.3390/su16188034 - 13 Sep 2024
Viewed by 407
Abstract
Cellulose and paper produce significant waste such as ash, activated sludge, and sludge from the pulp and paper industry. Depending on the raw material, legislation, and subprocesses, these sludges contain around 30–50% organic matter, mainly composed of less than 0.02 mm cellulose fibers [...] Read more.
Cellulose and paper produce significant waste such as ash, activated sludge, and sludge from the pulp and paper industry. Depending on the raw material, legislation, and subprocesses, these sludges contain around 30–50% organic matter, mainly composed of less than 0.02 mm cellulose fibers and hemicellulose and lignin. This work used sludge from the pulp and paper industry as a substrate for manufacturing mycelium-based biomaterials using the white rot fungus Trametes versicolor. Chemical and surface analyses revealed the formation of new materials. Acoustic impedance analyses revealed that these materials have a noise reduction coefficient and sound absorption average comparable to extruded polystyrene and polyurethane. In addition, the material’s thermal conductivity was near that of sheep wool. Therefore, the biomaterials fabricated using sludge and Trametes versicolor have the potential to be a game-changer in the industry as promising thermoacoustic insulators. Full article
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<p>Schematic representation of mycelium-based material fabrication.</p>
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<p>Photograph of biomaterials obtained using <span class="html-italic">Trametes versicolor</span> on PPMS.</p>
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<p>ATR FT-IR spectrum of sludge.</p>
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<p>X-ray diffraction pattern of sludge sample. The red lines highlight the diffraction peaks related to the crystalline cellulose phase, as mentioned in the previous paragraph.</p>
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<p>SEM images of mycelium-based biomaterials using <span class="html-italic">Trametes versicolor</span>. (<b>a</b>) Exterior face; (<b>b</b>) interior face.</p>
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<p>Radial extension of <span class="html-italic">Trametes versicolor</span> hyphae in MEA and PPMS. (<b>a</b>) Cumulative growth; (<b>b</b>) daily growth rate.</p>
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<p>Hyphal diameter of biomaterials obtained using <span class="html-italic">Trametes versicolor</span> on PMMS.</p>
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<p>ATR FT-IR spectrum of <span class="html-italic">Trametes versicolor</span> on PMMS.</p>
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29 pages, 1721 KiB  
Review
Medical Applications and Cellular Mechanisms of Action of Carboxymethyl Chitosan Hydrogels
by Weronika Kruczkowska, Karol Kamil Kłosiński, Katarzyna Helena Grabowska, Julia Gałęziewska, Piotr Gromek, Mateusz Kciuk, Żaneta Kałuzińska-Kołat, Damian Kołat and Radosław A. Wach
Molecules 2024, 29(18), 4360; https://doi.org/10.3390/molecules29184360 - 13 Sep 2024
Viewed by 272
Abstract
Carboxymethyl chitosan (CMCS) hydrogels have been investigated in biomedical research because of their versatile properties that make them suitable for various medical applications. Key properties that are especially valuable for biomedical use include biocompatibility, tailored solid-like mechanical characteristics, biodegradability, antibacterial activity, moisture retention, [...] Read more.
Carboxymethyl chitosan (CMCS) hydrogels have been investigated in biomedical research because of their versatile properties that make them suitable for various medical applications. Key properties that are especially valuable for biomedical use include biocompatibility, tailored solid-like mechanical characteristics, biodegradability, antibacterial activity, moisture retention, and pH stimuli-sensitive swelling. These features offer advantages such as enhanced healing, promotion of granulation tissue formation, and facilitation of neutrophil migration. As a result, CMCS hydrogels are favorable materials for applications in biopharmaceuticals, drug delivery systems, wound healing, tissue engineering, and more. Understanding the interactions between CMCS hydrogels and biological systems, with a focus on their influence on cellular behavior, is crucial for leveraging their versatility. Because of the constantly growing interest in chitosan and its derivative hydrogels in biomedical research and applications, the present review aims to provide updated insights into the potential medical applications of CMCS based on recent findings. Additionally, we comprehensively elucidated the cellular mechanisms underlying the actions of these hydrogels in medical settings. In summary, this paper recapitulates valuable data gathered from the current literature, offering perspectives for further development and utilization of carboxymethyl hydrogels in various medical contexts. Full article
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<p>Carboxymethylation and cross-linking methods of chitosan. Carboxymethylation of chitosan is a chemical modification process that produces different derivatives of chitosan. The reaction with ClCH<sub>2</sub>COOH and NaOH leads to two products, i.e., <span class="html-italic">O-</span>carboxymethyl chitosan and <span class="html-italic">N</span>,<span class="html-italic">O-</span>carboxymethyl chitosan. There are also different types of cross-linking that carboxymethyl chitosan can undergo including physical, chemical, enzymatic, and radiation cross-linking. Each cross-linking method forms connections among the polymer chains, chemical bonds, or interactions based on affinity.</p>
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<p>Properties and potential application of carboxymethyl chitosan-based hydrogels. CMCS hydrogels are known for their vast properties and application, with key examples depicted in the figure. Biocompatibility makes hydrogels applicable in the medical field, while their porous structure allows for the incorporation of drugs or bioactive molecules into their structure. This, combined with potential biodegradation, enables the controlled release of these substances. Hydrogels can also provide antimicrobial properties, which is attractive in medical applications. The major applications include wound healing, tissue regeneration, and targeted drug delivery from the hydrogel carrier.</p>
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<p>Carboxymethyl chitosan hydrogels—impact on cells. The cellular effects of carboxymethyl chitosan hydrogels include promoting collagen deposition, influencing fibroblasts, stimulating or inhibiting angiogenesis, providing antibacterial properties, and reducing inflammation. These diverse effects highlight the potential of these hydrogels in various biomedical applications, such as tissue engineering, wound healing, and disease treatment.</p>
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<p>Number of publications on carboxymethyl chitosan indexed in the Scopus database, sourced from publication “title” or “abstract” or “keyword”. The number of publications in the last ten years* and summarized number of publications until 2014, encompassing “carboxymethyl chitosan” and “a specific keyword” of the most exploited hydrogel biomaterial application and biological testing (* in 2024, only 8 months were taken into account for the date of this publication).</p>
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23 pages, 1172 KiB  
Article
Prevention of Cardiometabolic Syndrome in Children and Adolescents Using Machine Learning and Noninvasive Factors: The CASPIAN-V Study
by Hamid Reza Marateb, Mahsa Mansourian, Amirhossein Koochekian, Mehdi Shirzadi, Shadi Zamani, Marjan Mansourian, Miquel Angel Mañanas and Roya Kelishadi
Information 2024, 15(9), 564; https://doi.org/10.3390/info15090564 - 13 Sep 2024
Viewed by 251
Abstract
Cardiometabolic syndrome (CMS) is a growing concern in children and adolescents, marked by obesity, hypertension, insulin resistance, and dyslipidemia. This study aimed to predict CMS using machine learning based on data from the CASPIAN-V study, which involved 14,226 participants aged 7–18 years, with [...] Read more.
Cardiometabolic syndrome (CMS) is a growing concern in children and adolescents, marked by obesity, hypertension, insulin resistance, and dyslipidemia. This study aimed to predict CMS using machine learning based on data from the CASPIAN-V study, which involved 14,226 participants aged 7–18 years, with a CMS prevalence of 82.9%. We applied the XGBoost algorithm to analyze key noninvasive variables, including self-rated health, sunlight exposure, screen time, consanguinity, healthy and unhealthy dietary habits, discretionary salt and sugar consumption, birthweight, and birth order, father and mother education, oral hygiene behavior, and family history of dyslipidemia, obesity, hypertension, and diabetes using five-fold cross-validation. The model achieved high sensitivity (94.7% ± 4.8) and specificity (78.8% ± 13.7), with an area under the ROC curve (AUC) of 0.867 ± 0.087, indicating strong predictive performance and significantly outperformed triponderal mass index (TMI) (adjusted paired t-test; p < 0.05). The most critical selected modifiable factors were sunlight exposure, screen time, consanguinity, healthy and unhealthy diet, dietary fat type, and discretionary salt consumption. This study emphasizes the clinical importance of early identification of at-risk individuals to implement timely interventions. It offers a promising tool for CMS risk screening. These findings support using predictive analytics in clinical settings to address the rising CMS epidemic in children and adolescents. Full article
(This article belongs to the Section Artificial Intelligence)
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<p>Flow chart of the Prevalence and Dissemination of Non-Communicable Diseases in Children and Adolescents (CASPIAN-V) study design.</p>
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<p>Comparison of ROC curves between our method and the TMI index. The ROC curve for “our method” (blue, solid line) shows an AUC of 0.87. The ROC curve for the TMI index (red, dashed-dot line) is also compared, illustrating its discriminative performance (AUC = 0.75). The diagonal grey dashed line represents the line of no discrimination (AUC = 0.50).</p>
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26 pages, 3627 KiB  
Article
Unveiling the Performance of Co-Assembled Hybrid Nanocarriers: Moving towards the Formation of a Multifunctional Lipid/Random Copolymer Nanoplatform
by Efstathia Triantafyllopoulou, Diego Romano Perinelli, Aleksander Forys, Pavlos Pantelis, Vassilis G. Gorgoulis, Nefeli Lagopati, Barbara Trzebicka, Giulia Bonacucina, Georgia Valsami, Natassa Pippa and Stergios Pispas
Pharmaceutics 2024, 16(9), 1204; https://doi.org/10.3390/pharmaceutics16091204 - 13 Sep 2024
Viewed by 219
Abstract
Despite the appealing properties of random copolymers, the use of these biomaterials in association with phospholipids is still limited, as several aspects of their performance have not been investigated. The aim of this work is the formulation of lipid/random copolymer platforms and the [...] Read more.
Despite the appealing properties of random copolymers, the use of these biomaterials in association with phospholipids is still limited, as several aspects of their performance have not been investigated. The aim of this work is the formulation of lipid/random copolymer platforms and the comprehensive study of their features by multiple advanced characterization techniques. Both biomaterials are amphiphilic, including two phospholipids (1,2-dioctadecanoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC)) and a statistical copolymer of oligo (ethylene glycol) methyl ether methacrylate (OEGMA) and 2-(diisopropylamino) ethyl methacrylate (DIPAEMA). We examined the design parameters, including the lipid composition, the % comonomer ratio, and the lipid-to-polymer ratio that could be critical for their behavior. The structures were also probed in different conditions. To the best of the authors’ knowledge, this is the first time that P(OEGMA-co-DIPAEMA)/lipid hybrid colloidal dispersions have been investigated from a membrane mechanics, biophysical, and morphological perspective. Among other parameters, the copolymer architecture and the hydrophilic to hydrophobic balance are deemed fundamental parameters for the biomaterial co-assembly, having an impact on the membrane’s fluidity, morphology, and thermodynamics. Exploiting their unique characteristics, the most promising candidates were utilized for methotrexate (MTX) loading to explore their encapsulation capability and potential antitumor efficacy in vitro in various cell lines. Full article
(This article belongs to the Special Issue Polymer-Based Delivery System)
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<p>(<b>a</b>) The chemical structure of the random copolymer P(OEGMA950-co-DIPAEMA) synthesized by RAFT polymerization; (<b>b</b>) Graphic illustration of P(OEGMA-co-DIPAEMA)-1 or copolymer 1 and P(OEGMA-co-DIPAEMA)-2 or copolymer 2, respectively, with a different % comonomer ratio.</p>
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<p>Charts derived from DLS measurements at 25 °C: (<b>a</b>) The hydrodynamic radius (R<sub>h</sub>, nm); (<b>b</b>) the scattered intensity (I, kilocounts per second or kcps) of hybrid colloidal dispersions the day of their preparation, utilizing water for injection as the dispersion medium. The standard deviation (SD) is less than 10% in both diagrams. * Hybrid systems with more than one population; the predominant (higher intensity) one is presented in the graph.</p>
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<p>The hydrodynamic radius (R<sub>h</sub>, nm) of the hybrid colloidal dispersions in: (<b>a</b>) different dispersion media at body temperature (37 °C); (<b>b</b>) FBS:PBS biorelevant medium at different temperatures. The standard deviation (SD) is less than 10% in both diagrams. The DSPC:DOPC:1 9:1 hybrid system at 37 °C in both diagrams refers to a very high R<sub>h</sub> compared with the rest of the systems exceeding the scale of the graph.</p>
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<p>GP parameter vs. lipid composition of P(OEGMA<sub>950</sub>-co-DIPAEMA) hybrid systems at a steady lipid to polymer weight ratio (9:1).</p>
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<p>Cryo-TEM images of (<b>a</b>) P(OEGMA-co-DIPAEMA)-1. (<b>b</b>) P(OEGMA-co-DIPAEMA)-2 hybrid systems with different lipid compositions: (<b>i</b>) DSPC; (<b>ii</b>) DSPC:DOPC (9:1 weight ratio) and constant lipid to polymer ratio (9:1) or a constant lipid composition (DSPC) with different lipid to copolymer weight ratios: (<b>iii</b>) 7:3 and (<b>iv</b>) 5:5. The arrows represent the following: green color: spherical or irregularly shaped particles with distinct membrane; red color: “patchy” spherical- or pentagon-shaped vesicles; black color: rods; yellow color: small spherical particles.</p>
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<p>mDSC traces of (<b>a</b>) DSPC; (<b>b</b>) DSPC: DOPC (9:1 weight ratio) hybrid systems integrating P(OEGMA-co-DIPAEMA)-1 or -2 at different lipid to polymer weight ratios into aqueous medium.</p>
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<p>Thermodynamic evaluation of DSPC:P(OEGMA-co-DIPAEMA)-1 and -2 in an acidic environment (pH 4.5): (<b>a</b>) mDSC profiles; (<b>b</b>) (<b>i</b>) sound speed or (<b>ii</b>) attenuation vs. temperature from HR-US.</p>
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<p>(<b>a</b>) Size distributions from the DLS of DSPC:P(OEGMA-co-DIPAEMA)-2 hybrid systems incorporating MTX at two different lipid to polymer ratios, 9:1 (black line) and 5:5 (red line), on the day of their preparation; (<b>b</b>) the systems’ stability assessment (R<sub>h</sub> vs. time) under storage conditions (4 °C) for 21 days.</p>
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<p>Cell viability vs. different concentrations of MTX-DSPC:2 9:1 (blue line) and MTX-DSPC:2 5:5 (orange line) on (<b>a</b>) HEK293 and (<b>b</b>) HeLa cells. The concentration levels refer to MTX concentration, and the obtained data represent the means ± standard deviation from three experiments conducted in triplicates. The asterisks (*) in (<b>b</b>) correspond to p values of less than 0.05 (<span class="html-italic">p</span> &lt; 0.05) that are considered as statistically significant.</p>
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37 pages, 11194 KiB  
Review
Systematic Reversal of Drug Resistance in Cancer
by Shujie Zhu, Xuemei Wang and Hui Jiang
Targets 2024, 2(3), 250-286; https://doi.org/10.3390/targets2030015 - 13 Sep 2024
Viewed by 165
Abstract
Drug resistance in cancer is a significant contributor to high mortality, and it exists in the complex form of a multi-parameter. Here, we unravel the roles of tumor heterogeneity, intratumoral physiological barriers, and safe havens in the onset and progression of cancer drug [...] Read more.
Drug resistance in cancer is a significant contributor to high mortality, and it exists in the complex form of a multi-parameter. Here, we unravel the roles of tumor heterogeneity, intratumoral physiological barriers, and safe havens in the onset and progression of cancer drug resistance, and outline strategies for resolution. We advocate for a “three-step approach” to reverse cancer drug resistance, including the management of cancer evolution and early intervention, the normalization of intratumoral physiological barriers, and the breakage of tumor safe havens. This approach aims to effectively manage the source of drug resistance, dismantle the breeding grounds of drug resistance, and break the sanctuaries where drug resistance hides. Full article
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<p>Drug resistance in cancer. (<b>A</b>) Matrix changes in cancer [<a href="#B39-targets-02-00015" class="html-bibr">39</a>]; (<b>B</b>) schematic representation of (A) neurovascular unit; (B) paracellular transport pathway and transcellular transport pathway of BBB; (C) tight junction (TJ) associated components [<a href="#B62-targets-02-00015" class="html-bibr">62</a>]. (Figure was created by <a href="http://biorender.com" target="_blank">biorender.com</a> accessed on 30 August 2024).</p>
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<p>Management of cancer evolution and early intervention. (<b>A</b>) The heterogeneity of non-silent mutations from multiple-sample sequencing across a range of cancer types (black circles represent treatment naive tumors, with red triangles indicating tumors that have received treatment) [<a href="#B9-targets-02-00015" class="html-bibr">9</a>]; (<b>B</b>) evolutionary trees illustrating intratumor heterogeneity across cancer types [<a href="#B9-targets-02-00015" class="html-bibr">9</a>]; (<b>C</b>) phylogenetic relationships of the tumor regions [<a href="#B68-targets-02-00015" class="html-bibr">68</a>]; (<b>D</b>) clinical applications of CTC and ctDNA analyses in cancer care [<a href="#B72-targets-02-00015" class="html-bibr">72</a>]. (<b>E</b>) ctDNA-based MRD testing is predictive of survival outcomes in postsurgical patients with colorectal cancer [<a href="#B73-targets-02-00015" class="html-bibr">73</a>]. (<b>F</b>) A BEAMing analysis of circulating tumor DNA of patients with acquired resistance to cetuximab or panitumumab displays complex patterns of KRAS and NRAS mutations [<a href="#B74-targets-02-00015" class="html-bibr">74</a>]. (<b>G</b>) Resistance to EGFR therapy is reversed by pharmacological inhibition of EGFR and MEK (a combinatorial treatment with cetuximab plus pimasertib is effective in inducing tumor shrinkage in vivo) [<a href="#B74-targets-02-00015" class="html-bibr">74</a>]. (<b>H</b>) The evolution of resistance in a metastatic lesion. As the lesion (green) grows from one cell to a detectable size, new resistant subclones appear. Some of them are lost to stochastic drift (yellow and pink), while others survive (purple, red and orange triangles). Instead of looking at the time of appearance of new clones, the approach takes into account the total size of the lesion when the resistance mutation first occurred [<a href="#B75-targets-02-00015" class="html-bibr">75</a>]. (<b>I</b>) Resistant subclones in metastatic lesions [<a href="#B75-targets-02-00015" class="html-bibr">75</a>]. (<b>J</b>) The number of circulating tumor DNA (ctDNA) fragments per milliliter (Y1 to Y4) harboring different mutations associated with resistance to anti-EGFR agents in colorectal cancer patients treated with an EGFR blockade. The ratio of resistant clone sizes is given by the ratio of the ctDNA counts for any two resistance-associated mutations [<a href="#B75-targets-02-00015" class="html-bibr">75</a>]. (<b>K</b>) Heterologous vaccination with ChAd68 and samRNA induces broad, durable CD8<sup>+</sup> T cell responses in NHPs that are detectable long-term and can be boosted ≥ 2 years after their prime [<a href="#B76-targets-02-00015" class="html-bibr">76</a>]. (<b>L</b>) The removal of immunodominant epitopes and repetition of epitopes leads to increased target density and T cell response to KRAS mutant neoantigens [<a href="#B77-targets-02-00015" class="html-bibr">77</a>].</p>
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<p>Employing vascular endothelial growth factor inhibitors. (<b>A</b>) A summary of some of the major molecules implicated in angiogenesis [<a href="#B128-targets-02-00015" class="html-bibr">128</a>]. (<b>B</b>) Potential mechanisms of resistance to targeted VEGF therapy in cancer [<a href="#B128-targets-02-00015" class="html-bibr">128</a>]. In established tumours, VEGF blockade aggravates hypoxia, which upregulates the production of other angiogenic factors or increases tumour cell invasiveness (a). Other modes of tumour vascularisation, including intussusception, vasculogenic mimicry, differentiation of putative cancer stem cells into endothelial cells, vasculogenic vessel growth, and vessel co-option, might be less sensitive to VEGF blockade (b). Tumour vessels covered by pericytes are less sensitive to VEGF blockade (c). Recruited proangiogenic bone-marrow-derived cells, macrophages or activated cancer-associated fibroblasts can rescue tumour vascularisation by production of proangiogenic factors (d). (<b>C</b>) A plot of the Kaplan–Meier estimates for progression-free survival (non-small-cell lung cancer) for the (a) 7.5 mg/kg bevacizumab (Bev) arm and the (b) 15 mg/kg bevacizumab arm compared with a placebo [<a href="#B122-targets-02-00015" class="html-bibr">122</a>]. (<b>D</b>) The effect of a single injection of bevacizumab on the structural and functional markers of vascular normalization [<a href="#B123-targets-02-00015" class="html-bibr">123</a>]. (a) Microvessel density decreased. (b) Bevacizumab did not affect the density of mature vessels. (c) Fraction of vessel perimeter associated with pericytes (αSMA+ cells), a marker that distinguishes between poorly and completely covered vessels, increased. (d) Interstitial fluid pressure, which is a functional measurement of vessel leakiness and lymphatic vessel dysfunction, decreased. (e,f) Histological markers of functional vascular normalization, Ki67 for proliferation and HIF-1α, did not change significantly. In order to distinguish the figure and subfigure, the subfigures’ labels are replaced by lowercase letters in annotation.</p>
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<p>Enhancing pericyte coverage. (<b>A</b>) Neural cell adhesion molecule deficiency induces increased tumor blood vessel leakage during β tumor cell progression [<a href="#B129-targets-02-00015" class="html-bibr">129</a>]. NCAM deficiency induces increased Pancreas sections from 8-week-old RT (a) and RT<sup>NCAM+/−</sup> mice (b) were stained with H&amp;E. (b) Isolated cell clusters were specifically found inside blood-filled cavities within RT<sup>NC/KO</sup> islets (arrow). Inset in b shows a higher magnification of the isolated cell cluster. (c–h) Pancreas sections of mice perfused with FITC-dextran (green) were double-immunostained with antibodies against PECAM (red) and insulin (blue). (c,d) Islets from WT (c) and NCAM<sup>−/−</sup>(d) mice. (e–h) Angiogenic islets from 8-week-old RT (e,g) and RT<sup>NCAM+/−</sup>(f,h) mice. The islet area is indicated by dashed lines, and extravascular and intravascular FITC-dextran is indicated by arrowheads and arrows, respectively, in inset in (e). Dashed lines in g and h mark blood-filled cavities. FITC-dextran specifically leaked into RT<sup>NC/KO</sup> blood-filled cavities (h). (i) The percentage of islets containing blood-filled cavities was higher in RT<sup>NC/KO</sup> compared with RT mice at 8 weeks of age (χ<sup>2</sup> test, *** <span class="html-italic">p</span> &lt; 0.001). (j) Distribution of RT and RT<sup>NC/KO</sup> islets at 8 weeks of age according to their vessel leakage (grades 0–3, where grade 3 includes islets with most extensive leakage). (<b>B</b>) The pathological organization of periendothelial α-SMA<sup>+</sup> cells correlates with increased tumor vessel leakage in RT<sup>NC/KO</sup> angiogenic islets [<a href="#B129-targets-02-00015" class="html-bibr">129</a>]. Pancreas sections from 8-week-old mice were double-immunostained with antibodies against PECAM (red) and α-SMA (a–e, green; f, red). In WT (a) and NCAM<sup>−/−</sup> (b) islets, α-SMA<sup>+</sup> cells were closely attached to the endothelium. Premature abnormal organization of periendothelial α-SMA<sup>+</sup> cells, including detachment of α-SMA<sup>+</sup> cells from endothelial cells (arrow in c) and multiple layers of α-SMA<sup>+</sup> cells with an apparent loose attachment to the endothelium (d), and presence of fibroblastlike α-SMA<sup>+</sup> cells in RT<sup>NCAM+/−</sup> angiogenic islets (e), were observed in RT<sup>NCAM+/−</sup> islets. The dashed lines and arrows in d indicate the borders of the endothelium and α-SMA<sup>+</sup> cells stretching away from the vessel, respectively. (f) Pancreas section of an 8-week-old RT<sup>NCAM+/−</sup> mouse perfused with FITC-dextran (green), immunostained with anti–α-SMA (red). Increased leakage correlated with severely disorganized α-SMA<sup>+</sup> periendothelial cells. The inset in (c) shows coexpression of NG2 (green) and α-SMA (red). (g) Analysis of blood vessel density revealed no difference between RT and RT<sup>NC/KO</sup> islets. (<b>C</b>) The effect of host-derived Ang-2 deficiency on MVD (a,b), the diameter of intratumoral microvessels (a,b), and perfusion (c) in Lewis lung carcinoma tumors [<a href="#B130-targets-02-00015" class="html-bibr">130</a>]. (<b>D</b>) Structural changes within established primary tumor vessels invoked by vascular endothelial protein tyrosine phosphatase (VE-PTP) inhibition [<a href="#B134-targets-02-00015" class="html-bibr">134</a>]. (a) Colocalization of CD31-positive area (endothelial cells) and NG2-positive area (perivascular cells) (* <span class="html-italic">p</span> = 0.05 by two-sided t test; n = 6 per group) in 4T1 tumors. (b) Colocalization of CD31-positive area (endothelial cells) and desmin-positive area (perivascular cells) (* <span class="html-italic">p</span> = 0.01 by two-sided t test; n = 6 per group) in 4T1 tumors. (c) Mean distance between desmin-positive pericytes and CD31 = positive endothelial cells in microns (* <span class="html-italic">p</span> &lt; 0.01 by two-sided t test; n = 6 per group) in 4T1 tumors. (d) Colocalization of CD31+ area (endothelial cells) and NG2+ area (perivascular cells) (* <span class="html-italic">p</span> = 0.05 by two-sided t test; n = 6 per group) in E0771 tumors (* <span class="html-italic">p</span> &lt; 0.01 by two-sided t test; n = 6 per group). (e) Enhanced vascular pericyte coverage in AKB-9778–treated tumors (right panel) compared with control-treated tumors (left panel) (red: CD31; green: NG2; blue: DAPI). (f,g) Vessel diameter in control vs. AKB-9778–treated 4T1 (f) and E0771 (g) tumors (f: * <span class="html-italic">p</span> &lt; 0.01 by two-sided t test, n = 6 per group; g: * <span class="html-italic">p</span> &lt; 0.01 by two-sided t test, n = 6 per group). (h,i) Vessel density in control vs AKB-9778–treated 4T1 (h) and E0771 (i) tumors (h: * <span class="html-italic">p</span> = 0.05 by two-sided t test, n = 6 per group; i: * <span class="html-italic">p</span> = 0.05 by two-sided t test, n = 6 per group). (<b>E</b>) Ang2-blocking antibodies inhibit primary tumor growth, angiogenesis, and lymphangiogenesis [<a href="#B132-targets-02-00015" class="html-bibr">132</a>]. (a) Growth curves of LNM35 primary tumors in nu/nu mice treated with the Ang2-blocking antibodies or hIgG control, n = 8 in both groups. (b) Tumor weights at excision 16 days after implantation, <span class="html-italic">p</span> = 0.002. Student’s <span class="html-italic">t</span> test. (c) Representative immunohistochemical images of LYVE-1- and CD31-stained tumor sections. (d) Quantification of densities and area fractions of Lyve-1-positive lymphatic vessels and of CD31-positive blood vessels from at least five histological sections, <span class="html-italic">p</span> = 0.013 and 0.019, respectively. Student’s <span class="html-italic">t</span> test. (<b>F</b>) The Ang2-blocking antibody induces internalization of the Ang2-Tie2 complexes, leaving Ang1-Tie2 complexes intact at endothelial cell–cell junctions [<a href="#B132-targets-02-00015" class="html-bibr">132</a>]. (<b>G</b>) The overall survival in patients with ascites at baseline (trebananib plus weekly paclitaxel in recurrent ovarian cancer) [<a href="#B133-targets-02-00015" class="html-bibr">133</a>]. In order to distinguish the figure and subfigure, the subfigures’ labels are replaced by lowercase letters in annotation.</p>
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<p>Reinforcing tight junctions. (<b>A</b>) CU06-1004 reduces vascular leakage with a concomitant increase in junction integrity in tumor blood vessels [<a href="#B137-targets-02-00015" class="html-bibr">137</a>]. (a) Schematic plan for the administration of Sac-1004 or control (DMSO) to tumor-bearing mice. (b) B16F10 tumor-bearing mice (n = 5) were injected with Sac-1004 or control as in (a) and tumor vascular leakage was quantified by the Evans blue method. (c) Vascular leakage was assessed by FITC-dextran. (d) Images shown in (c) were quantified using ImageJ software. (e) Immunofluorescence staining of B16F10 tumor sections, treated with Sac-1004 or control, for CD31 and VE-cadherin. Arrows indicate discontinuity in VE-cadherin staining. (f) Quantification of immunofluorescence images shown in (e) using Multi Gauge software (n = 5). (g) LLC tumor sections, treated with Sac-1004 or control were costained for CD31, ZO-1 and DAPI. (h) Images shown in (g) were quantified using ImageJ software (n = 5). (i) Western blot analysis of B16F10 tumors treated with Sac-1004 or control for VE-cadherin. (j) VE-cadherin and actin blots from (i) were quantified using ImageJ software. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.001 (Student’s <span class="html-italic">t</span>-test). (<b>B</b>) CU06-1004 improves vascular perfusion, alleviates hypoxia, and normalizes tumor blood vessels in tumors [<a href="#B137-targets-02-00015" class="html-bibr">137</a>]. (a) Immunofluorescence staining of B16F10 tumor sections (n = 5), treated with Sac-1004 or control, for CD31 and tomato lectin. (b) Images shown in (a) were quantified using Image J software. (c) Immunohistochemical analysis of B16F10 tumor sections (n = 5) for CD31, hypoxia, and vascular perfusion (Hoechst dye) in the peritumoral and intratumoral zone. Arrows indicate non-perfused vessels. (d#x2013;f) Quantification of immunofluorescence images shown in (c) with Multi Gauge software. (g) Quantification of HIF-1α positive area using Multi Gauge software. (h) B16F10 tumor sections (n = 5), treated with Sac-1004 or control, were stained for CD31 and ColIV (up)/laminin (bottom). Arrowheads indicate the point of detachment between basement membrane and endothelial cells. (i) Quantification of basement membrane thickness in B16F10 tumor vessels shown in (h) using Multi Gauge software. (j) Immunofluorescence staining of LLC tumor sections (n = 5) for CD31 and NG2. Quantification was done using Multi Gauge software. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.001 (Student’s <span class="html-italic">t</span>-test). In order to distinguish the figure and subfigure, the subfigures’ labels are replaced by lowercase letters in annotation.</p>
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<p>Normalizing intratumoral lymphatics while inhibiting peripheral tumor lymphangiogenesis. (<b>A</b>) The inhibition of tumor lymphangiogenesis by VEGFR−3−Ig (blood and lymphatic vessels in the LNM35 and N15 tumors) [<a href="#B139-targets-02-00015" class="html-bibr">139</a>]. Immunohistochemical staining is shown for LYVE-1 (a–d,g,h) to identify lymphatic vessels and for PECAM-1 (e,f) to identify both blood and lymphatic vessels. (a,c,e) Sections of the control LNM35/pEBS7 tumors. (b,d,f) Sections from the LNM35/vascular endothelial growth factor receptor−3−Ig (VEGFR−3−Ig) tumors. (g,h) Sections of the N15/pEBS7 and N15/VEGF−C tumors, respectively. (i) LYVE-1-stained or PECAM−1−-stained vessels in three microscopic fields of the highest vessel density were counted, and the results were compared by use of the unpaired t test. (<b>B</b>) The suppression of axillary lymph node metastasis by VEGFR−3−Ig [<a href="#B139-targets-02-00015" class="html-bibr">139</a>]. (a) Typical lymph nodes in mice bearing the vascular endothelial growth factor receptor 3 Ig (VEGFR-3-Ig) tumors (upper pair) and control LNM35 tumors (lower pair). (b) Lymph node (LN) volume and 95% confidence intervals (<span class="html-italic">p</span> = 0.070). (c,d) Histologic staining of lymph node sections from mice with the VEGFR−3−Ig−overexpressing and control tumors, respectively. Arrows = tumor cells in the lymph node. (<b>C</b>) The inhibition of the metastatic spread of orthotopic gastric AZL5G tumors by the systemic administration of anti−VEGFR−3 antibodies (AFL4) [<a href="#B143-targets-02-00015" class="html-bibr">143</a>]. (a) Control mouse (P; primary tumor, arrow; metastatic lymph node). (b): AFL4−treated mouse (P; primary tumor). (<b>D</b>) An assessment of lymphatic and blood vessel density in control and AFL4−treated mice [<a href="#B143-targets-02-00015" class="html-bibr">143</a>]. Immunohistochemistry of primary tumors for LYVE−1 (a,b) and CD31 (c,d) and schema of the vessel counts (e,f). Compared with the control group, the number of LYVE−1−positive lymphatic vessels (LVD) in the primary tumors in the AFL4−treated group is dramatically decreased (e, <span class="html-italic">p</span> &lt; 0.05). In contrast, CD31−positive LYVE−1−negative microvessel density (MVD) was not significantly different between the control group and the AFL4−treated group (f, <span class="html-italic">p</span> = 0.84). In order to distinguish the figure and subfigure, the subfigures’ labels are replaced by lowercase letters in annotation.</p>
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<p>The therapeutic potential of lymphatic system immunomodulatory capacity for lymphatic normalization. (<b>A</b>) VEGF−C/VEGFR-3 signaling increases the responsiveness of melanoma to immunotherapy [<a href="#B144-targets-02-00015" class="html-bibr">144</a>]. Tumor growth and survival of three different melanoma models treated with control (Iso) or aR3-blocking antibodies receiving different immunotherapies (arrows indicate times of administration). (a,b) B16-OVA/VC tumors treated with ATT in (a) WT (n ≥ 15) and (b) K14−VEGFR−3−Ig mice that lack dermal lymphatics (n = 4). (c–f) B16−OVA/VC tumors in WT mice treated with (c) ex vivo activated DCs (DC vax; n = 6), (d) 50 mg of CpG (n = 6), (e) 10 mg of OVA + 50 mg of CpG (n ≥ 8), and (f) 2 mg of Trp2 peptide−conjugated nanoparticles (NP-Trp2) + 50 mg of CpG (n = 7). (g) B16/VC tumors treated with NP-Trp2 + 50 mg of CpG (n = 6). (h) Tamoxifen-induced tumors in Braf<sup>V600E</sup>/Pten<sup>−/−</sup> mice treated with CpG + gp100 peptide (days 8 and 12) and anti−PD−1 antibody (day 12 and every 4 days thereafter). Each panel shows data from one (b–d,f,g), two (e), or three (a) independent experiments. * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001 by two-tailed Student’s <span class="html-italic">t</span> test for growth curves and log-rank (Mantel-Cox) test for comparing survival curves. (<b>B</b>) The increased efficacy of immunotherapy in lymphangiogenic B16 melanomas depends on CCR7 signaling before therapy and the local activation and expansion of TILs after therapy [<a href="#B144-targets-02-00015" class="html-bibr">144</a>]. (a,b) B16-OVA/VC tumor–bearing mice treated with control IgG (Iso) or anti−VEGFR−3 (aR3)–blocking antibodies were euthanized 3 days after ATT, and tumor single-cell suspensions were analyzed by flow cytometry (n = 5). Quantification of overall naïve CD8<sup>+</sup> (CD45<sup>+</sup> CD8<sup>+</sup> CD44<sup>−</sup> CD62L<sup>+</sup>), effector CD8<sup>+</sup> (CD45<sup>+</sup> CD8<sup>+</sup> CD44<sup>+</sup> CD62L<sup>−</sup>), and OT-I (CD45<sup>+</sup> CD8<sup>+</sup> CD45.1<sup>+</sup>) T cells (a) in the tumor and (b) in the dLNs. (c) Tumor growth and survival curves of B16-OVA/VC tumor−bearing mice treated with anti-CCR7 (aCCR7), control IgG (Iso), or aR3 antibodies combined with ATT on day 9. CCR7 blockade was performed only before ATT (days 0, 3, and 6) (data pooled from two or more independent experiments, n ≥ 15 total). (d) Tumor growth curves of B16−OVA/VC tumor–bearing mice treated with control IgG (Iso) or aR3 antibodies received daily injections of the small molecular S1P inhibitor FTY720 starting on the same day as ATT was performed (day 9) (n ≥ 5). Statistics show differences between Iso + FTY720 and aR3 + FTY720 by one-way ANOVA. (e) Representative flow cytometry plots and (f) quantification of circulating CD4<sup>+</sup> and CD8<sup>+</sup> T cells (after B220 exclusion) in blood 26 days after tumor inoculation. * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001 by two−tailed Student’s <span class="html-italic">t</span> test or one−way ANOVA and log-rank (Mantel−Cox) test for survival curves. (<b>C</b>) Dorsal MLVs are the main route for immune cell entry to draining CLNs [<a href="#B145-targets-02-00015" class="html-bibr">145</a>]. (a) Heat map of DEGs (Up, 219; Down, 100; power &gt; 0.4). (b, c) Gene sets involved in lymphatic remodeling, fluid drainage, as well as inflammatory and immunological responses as shown by the representative upregulated pathways in GL261 tumor-associated and B16 tumor-associated MLECs compared to control MLECs (b), and heat map of DEGs enriched in the antigen processing and presentation pathway (c). (d) Left panels, treatment scheme and representative flow cytometry dot plots of DC trafficking from GL261 tumors to dCLNs in mice treated with Vehicle + Laser or Visudyne + Laser, determined by the quantity of CD11c<sup>+</sup>MHCII<sup>+</sup>FITC<sup>+</sup> cells in the dCLNs 24 h after intratumoral injection of FITC-labeled latex beads. Right panel, quantification of Bead<sup>+</sup> DCs in the dCLNs of mice treated with Vehicle + Laser or Visudyne + Laser. (e) Immunoprecipitation of secreted VEGF−C protein (arrow) in conditioned medium from GL261-Vector, GL261-VEGF-C, B16-Vector, and B16−VEGF−C cells. (f) Left panels, LYVE-1 and CCL21 staining of MLVs in mice bearing Empty and VEGF-C-overexpressing GL261 tumors in the striatum (scale bars, 100 µm in wide-fields; 50 µm in insets). Right panels, quantification of the percentage area of LYVE-1 and CCL21 (n = 10). (g) Left panels, treatment scheme and representative flow cytometry dot plots of DC trafficking in the dCLNs of mice bearing GL261 tumors overexpressing Vector or VEGF−C. Right panel, quantification of bead<sup>+</sup> DCs in dCLNs (n = 10). (h) Left panels, treatment scheme and representative flow cytometry dot plots of DC trafficking in the dCLNs of GL261 tumor-bearing mice treated with CCL21 (αCCL21)- or IgG (Iso)-blocking antibodies. Right panel, quantification of bead<sup>+</sup> DCs in dCLNs (n = 10). * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001. (<b>D</b>) A high level of tumor-derived VEGF-C improves anti-PD-1/CTLA-4 efficacy [<a href="#B145-targets-02-00015" class="html-bibr">145</a>]. (a) Survival of mice with striatal Vector- or VEGF-Coverexpressing GL2161 tumors following the administration of anti-PD-1/CTLA-4 or IgG controls (n = 15). (b) Representative T2-weighted single brain slices from mice with intracranial injection of GL261 cells overexpressing Vector or VEGF-C (n = 6). (c) Tumor volumes in mice with striatal injection of GL261 cells overexpressing Vector or VEGF-C (n = 6). (d,e) Quantification of CD8<sup>+</sup>Ki67<sup>+</sup> T cells (d) and CD4<sup>+</sup>Foxp3<sup>+</sup> T cells (e) as percentages of overall CD45<sup>+</sup> cells in tumors and in dCLNs on day 14 after inoculation (n = 12 in each). (f) Ratios of CD8<sup>+</sup>Ki67<sup>+</sup> T cells to Tregs in tumors and in dCLNs. * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001. In order to distinguish the figure and subfigure, the subfigures’ labels are replaced by lowercase letters in annotation.</p>
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<p>Normalization of the matrix. (<b>A</b>) The effect of collagenase on oncolytic viral therapy [<a href="#B147-targets-02-00015" class="html-bibr">147</a>]. (<b>B</b>) The effect of collagenase on MGH2-induced tumor growth delay [<a href="#B147-targets-02-00015" class="html-bibr">147</a>]. (<b>C</b>) The buckling of a collagen fiber in a relaxin-treated tumor [<a href="#B149-targets-02-00015" class="html-bibr">149</a>]. (<b>D</b>) A quantitative analysis of collagen fiber length and the area of colocalization between stromal cells and fibers [<a href="#B149-targets-02-00015" class="html-bibr">149</a>]. (a,b) The end-to-end fiber length (a) and the area of overlap between stromal cells and collagen fibers (b) was determined over 4 d in control groups as well as those treated with relaxin, relaxin and b1 integrin antibody (R + b1) and relaxin and GM6001 (R + GM). Shown are the averaged differences between day 1 and days 2 (D2-1), 3 (D3-1) and 4 (D4-1). * <span class="html-italic">p</span> &lt; 0.05. (<b>E</b>) Blocking TGF-β signaling improves the intratumoral distribution of doxorubicin in orthotopic mammary carcinoma models [<a href="#B153-targets-02-00015" class="html-bibr">153</a>]. (a,b) Representative images of doxorubicin intratumoral distribution in 4T1 (a) and 4T1–sTβRII (b) tumors. Green, FITC–lectin-labeled perfused vessels; red, fluorescent doxorubicin; blue, DAPI. (c) Quantification of the fraction of tumor area positive for doxorubicin (n = 12 sections, with 3 sections per tumor). * <span class="html-italic">p</span> &lt; 0.001. (<b>F</b>) Blocking TGF-β signaling decreases collagen I content and improves Doxil tissue penetration [<a href="#B153-targets-02-00015" class="html-bibr">153</a>]. (a) Representative images and quantification of collagen I immunofluorescent staining in 4T1 and 4T1–sTβRII tumors. Red, collagen I staining; blue, DAPI (×20). (b) Representative images and quantification of Doxil intratumoral distribution in 4T1 and 4T1–sTβRII tumors. Green, FITC-lectin labeled perfused vessels; red, fluorescent doxorubicin; blue, DAPI (n = 12 sections, with 3 sections per tumor). * <span class="html-italic">p</span> &lt; 0.001. (<b>G</b>) Blocking TGF-β signaling enhances Doxil efficacy in orthotopic mammary carcinoma models [<a href="#B153-targets-02-00015" class="html-bibr">153</a>]. (a and b) Primary tumor growth of 4T1 and 4T1–sTβRII tumors, with or without Doxil treatment (a) and 4T1 (b) and MDA-MB-231 (c) tumors treated with saline (control), Doxil alone (9 mg/kg, weekly), 1D11 alone (5 mg/kg, three times a week), or combined Doxil and 1D11 (n = 8 in all groups). In order to distinguish the figure and subfigure, the subfigures’ labels are replaced by lowercase letters in annotation.</p>
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<p>Biochemical therapies for breaking the safe haven. (<b>A</b>) The brain penetration of ABT-888 is limited by Abcb1 and Abcg2 [<a href="#B162-targets-02-00015" class="html-bibr">162</a>]. (a) ABT-888 plasma concentrations, brain concentrations, and brain-to-plasma ratios following intravenous administration of 10 mg/kg of ABT-888 (n = 5/time point/strain). (b) ABT-888 levels following 10 mg/kg p.o. administered to wild-type and Abcb1a/1b<sup>−/−</sup>; Abcg2<sup>−/−</sup> mice with/without co-administration of 100 mg/kg elacridar p.o. Blood samples were collected from the tail at 15 min, 1, 2, and 4 h (n = 8/strain). Brain samples were harvested at 4 h after drug administration. **** <span class="html-italic">p</span> &lt; 0.0001 compared with wild-type. (<b>B</b>) The inhibition of Abcb1 and Abcg2 improve efficacy of ABT-888 + TMZ treatment [<a href="#B162-targets-02-00015" class="html-bibr">162</a>]. (a) Efficacy of TMZ versus TMZþABT-888 treatment against intracranial p53; p16<sup>Ink4a</sup>/p19<sup>Arf</sup>; K-Ras<sup>v12</sup>; LucR GBM652457 cells injected into wild-type mice. (b) Same setup but now injected into Abcb1a/1b<sup>−/−</sup>; Abcg2<sup>−/−</sup>(KO) mice. (c) Efficacy of TMZ + ABT-888 with and without elacridar in both WT and KO mice. (d) Efficacy of TMZ or TMZ + ABT-888 with or without elacridar against (lentivirally induced) spontaneous p53; p16<sup>Ink4a</sup>/p19<sup>Arf</sup>; K-Ras<sup>v12</sup>; LucR tumors. TMZ p.o. at 100 mg/kg every day alone or concurrently with ABT-888 p.o. at 10 mg/kg twice a day and/or elacridar p.o. at 100 mg/kg every day 15 min before TMZ for 5 days. Left, relative tumor growth curves. Right, Kaplan–Meier analysis of survival. (<b>C</b>) PTEN-deficient tumors are more sensitive to ABT-888 + TMZ treatment [<a href="#B162-targets-02-00015" class="html-bibr">162</a>]. (a) Kaplan–Meier analysis of PTEN deletion (≤1.8 copies) on overall survival of patients with glioblastoma. (b) Sensitivity (in vitro) of two panels of glioblastoma cell lines of different genetic backgrounds exposed to 100 μmol/L TMZ and increasing concentrations of ABT-888 for 5 days. (c) Efficacy of ABT-888 in combination with TMZ against Pten;p16<sup>Ink4a</sup>/p19<sup>Arf</sup>;K-Ras<sup>v12</sup>;LucR GBM696677 cells injected intracranially into WT or Abcb1a/1b<sup>−/−</sup>;Abcg2<sup>−/−</sup> (KO) mice and (d) spontaneous Pten;p16<sup>Ink4a</sup>/p19<sup>Arf</sup>; K-Ras<sup>v12</sup>;LucR tumors. TMZ (100 mg/kg p.o. every day) alone or concurrently with ABT-888 (10 mg/kg p.o. twice a day) for 5 days. C and d (right), Kaplan-Meier analysis of survival. (<b>D</b>) The brain concentration of paclitaxel (10 mg/kg) in mice at 4 h after administration of paclitaxel alone and in combination with different (putative) inhibitors of Pgp (Pgp knockout mice were used as a reference for “complete” inhibition of Pgp) [<a href="#B163-targets-02-00015" class="html-bibr">163</a>]. In order to distinguish the figure and subfigure, the subfigures’ labels are replaced by lowercase letters in annotation.</p>
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<p>Physical therapies for breaking the safe haven. (<b>A</b>) FUS enhances doxorubicin (Dox) penetration and promotes convective transport in BT474-Gluc brain tumors [<a href="#B164-targets-02-00015" class="html-bibr">164</a>]. (a) Sequential intravital multiphoton microscopy of Dox autofluorescence. (b) Temporal evaluation of Dox extravasation with and without FUS-BTB disruption. Cv and Ce are the mean pixel intensity of the vessel and the extravascular space, respectively. The maximum mean fluorescence for the FUS and non-FUS was 0.52 ± 0.15 and 0.07 ± 0.02, a sevenfold difference. (c) Dox penetration from a line profile perpendicular to vessel wall (red dotted arrow in a). The plot shows the normalized maximum intensity projection (MIP) across the series of images. The dotted line shows a regression fitted to the data from four different animals for each condition (non-FUS and FUS). (d) Representative data of the temporal evolution of the normalized intensity of the line profile (red dotted arrow in a). For consistency in the notation of the experiments/modeling, Cv is the Dox intensity/concentration in the vessel, Ce is the Dox intensity/concentration in the extracellular/interstitial space. (<b>B</b>) FUS-BTB disruption increases early extravasation and penetration of T-DM1 in BT474-Gluc brain tumors [<a href="#B164-targets-02-00015" class="html-bibr">164</a>]. (a) Representative microscopy data of TDM1 extravasation with and without FUS at 4 h and 5 d. (b) Quantification of the T-DM1 extravasation (Left) and penetration (Right) with and without FUS at 4 h (Upper) and 5 d (Lower) posttreatment. Parametric Student’s <span class="html-italic">t</span> test for <span class="html-italic">p</span> &lt; 0.05. (<b>C</b>) The quantification of transvascular transport via mathematical modeling demonstrates a multifold increase in the effective diffusion coefficient (4.3-fold) and in hydraulic conductivity (4.5–fold) after FUS-BTB disruption [<a href="#B164-targets-02-00015" class="html-bibr">164</a>]. (a) Schematic illustrating the transport of the anticancer agents from the vessel to the interstitial space along with the studied model parameters and agent-specific cellular uptake model equations. (Upper) Convection–diffusion–reaction model following Michaelis–Menten kinetics with binding of doxorubicin to DNA (Vb). (Lower) Convection–diffusion–reaction model for T-DM1. Excellent fit was observed for both doxorubicin and T-DM1. (b, Upper) The time dependence of doxorubicin extravasation from the fitted and experimental data for non–FUS and FUS–BBB/BTB disruption groups. (b, Lower) Parameter fit methodology for T–DM1 and fitted data from two different experiments. The fitted vascular and interstitial effective porosity (fraction of surface area occupied by pores) from the doxorubicin model was used as input to the T-DM1 fitting (i.e., same animal model). (c) Normalized parameter fit for non-FUS and FUS-BBB/BTB disruption groups (Upper, doxorubicin; Lower, T-DM1). The values for each parameter were normalized to maximum to be displayed on the same plot. The exact numbers and their units are shown in <a href="#targets-02-00015-t001" class="html-table">Table 1</a> and <a href="#targets-02-00015-t002" class="html-table">Table 2</a> for doxorubicin and T-DM1, respectively. (<b>D</b>) The diminished interaction of intracellular scaffolding proteins ZO-1 and occludin as a result of ultrasound treatment [<a href="#B166-targets-02-00015" class="html-bibr">166</a>]. (a) Western Blot analysis on whole brain tissue lysates (WTL) shows that ZO-1 and occludin protein levels are not changed in response to ultrasound treatment. (b), Co-immunoprecipitation of occludin and ZO-1. The amount of occludin co-precipitating with ZO-1 in the presence of US-treatment is reduced when compared to non-sonicated brains (-US). (<b>E</b>) Ultrasound in the presence of microbubbles increases the activity of the Akt signaling pathway, while the activity of MAPK signaling remains unchanged [<a href="#B166-targets-02-00015" class="html-bibr">166</a>]. Brains were removed 1.5 h (a) or 24 h (b) post-sonication and snap-frozen using liquid nitrogen. Brain tissue regions with trypan blue leakage in the sonicated hemisphere (+US) and the equivalent area from opposite hemisphere (-US) were then homogenized with RIPA lysis buffer. Equal amounts of extracted proteins were analyzed by western blotting for the indicated proteins. (c) Graphical representation of three independent experiments illustrating the marked increase in pAkt (Ser473), pAkt (Thre308) and pGSK3b (Ser9) 1.5 hrs after sonication treatment. The star (*) represents <span class="html-italic">p</span> &lt; 0.05 for -US versus +US. (<b>F</b>) Increased phosphorylation of Akt and GSK3β in neuronal cells of sonicated rat brain regions [<a href="#B166-targets-02-00015" class="html-bibr">166</a>]. (a) Panel [i] shows a control section without application of any primary antibodies. Panel [ii] represents a direct staining of the astrocytes with Alexa-fluor488-conjugated GFAP (green, right arrow). Panels [iii] and [iv] represent immunofluorescence staining with pAkt (red, curved arrow) and GFAP (green, right arrow) in non-sonicated (-US) as well as sonicated (+US) hemispheres respectively. Panels [v] and [vi] illustrate co-staining of pGSK3b (red, pentagon) and GFAP (green, right arrow) in ‘-US’ and ‘+US’ brain regions respectively. (b) Panels [ii] and [iii] represent neuronal cells morphology as directly stained with Alexa-Fluor 488-conjugated NeuN (green, lightning bolt). Co-staining of pAkt (red, curved arrow) with NeuN (green) and also pGSK3b (red, pentagon) with NeuN (green) are shown in panels [iii, iv] and [v, vi] respectively. The regions of the brain sections with IgG extravasation (arrow head) and their surrounding neuronal cells with elevated levels of pAkt and pGSK3b are shown in panels [iv, vi]. (<b>G</b>) Representative Evans Blue (EB) dye staining in animal brains after FUS-BBB opening [<a href="#B167-targets-02-00015" class="html-bibr">167</a>]. (<b>H</b>) (a) The TMZ concentration (mean ± STD) at 2 h after TMZ administration obtained from plasma and brain tissues from each experimental group. (b) The estimated time (in hours) for TMZ to degrade to 50% of the peak level [<a href="#B167-targets-02-00015" class="html-bibr">167</a>]. (<b>I</b>) (a) The average fluorescence intensity and (b) average area of fluorescence for the 3, 70, 500, and 2000 kDa dextrans sonicated at 0.31, 0.51, and 0.84 MPa. The pressure threshold for significant increases in both fluorescence and the area of fluorescence was 0.51 MPa for 3 and 70 kDa dextrans. However, it increased to 0.84 MPa for the 500 and 2000 kDa dextrans [<a href="#B169-targets-02-00015" class="html-bibr">169</a>]. (<b>J</b>) Microscopic examination of (a,c) left (sonicated) and (b,d) the corresponding right (nonsonicated) hippocampi in hemotoxylin and eosin-stained, 6 μm-thick horizontal sections [<a href="#B169-targets-02-00015" class="html-bibr">169</a>].</p>
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16 pages, 3758 KiB  
Article
Properties of Skin Collagen from Southern Catfish (Silurus meridionalis) Fed with Raw and Cooked Food
by Qi Zhang, Shufang Hou, Yanmei Liu, Jia Du, Yongkang Jia, Qiushi Yang, Tingting Xu, Yasuaki Takagi, Dapeng Li and Xi Zhang
Foods 2024, 13(18), 2901; https://doi.org/10.3390/foods13182901 - 13 Sep 2024
Viewed by 268
Abstract
The southern catfish (Silurus meridionalis) is an economically important carnivorous freshwater fish in China. In this study, we compared the properties of skin collagen from southern catfish fed with raw food (RF) and cooked food (CF). The skin collagen yield in [...] Read more.
The southern catfish (Silurus meridionalis) is an economically important carnivorous freshwater fish in China. In this study, we compared the properties of skin collagen from southern catfish fed with raw food (RF) and cooked food (CF). The skin collagen yield in the RF group (8.66 ± 0.11%) was significantly higher than that of the CF group (8.00 ± 0.27%). SDS-PAGE, circular dichroism spectroscopy, and FTIR analyses revealed that the collagen extracted from southern catfish skin in both groups was type I collagen, with a unique triple helix structure and high purity. The thermal denaturation temperature of collagen in the RF group (35.20 ± 0.11 °C) was significantly higher than that of the CF group (34.51 ± 0.25 °C). The DPPH free radical scavenging rates were 68.30 ± 2.41% in the RF collagen and 61.78 ± 3.91% in the CF collagen, which was higher than that found in most fish collagen. Both the RF and CF groups had high ability to form fibrils in vitro. Under the same conditions, the CF group exhibited faster fibril formation and a thicker fibril diameter (p < 0.05). In addition, the RF group exhibited significantly higher expression of col1a1 compared to the CF group. These results indicated that feeding southern catfish raw food contributed to collagen production, and the collagen from these fish may have potential in biomaterial applications. Full article
(This article belongs to the Section Nutraceuticals, Functional Foods, and Novel Foods)
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<p>(<b>A</b>) Wet weight extraction rates of collagen from southern catfish skin under raw (RF) and cooked (CF) conditions. (<b>B</b>) Dry weight extraction rates of collagen from southern catfish skin under raw (RF) and cooked (CF) conditions. * denotes a significant difference between RF and CF (<span class="html-italic">p</span> &lt; 0.05). ** indicates highly significant differences (<span class="html-italic">p</span> &lt; 0.01).</p>
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<p>(<b>A</b>) SDS-PAGE results of skin collagen from southern catfish skin under raw food (RF) and cooked food (CF) feeding. M: Marker; 1–3: RF feeding; 4–6: CF feeding. (<b>B</b>) Relative band density levels of α1 and α2 chains.</p>
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<p>Infrared spectra of skin collagen from southern catfish skin under raw food (RF) and cooked food (CF) feeding.</p>
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<p>Collagen CD spectra of southern catfish skin under raw food (RF) and cooked food (CF) feeding.</p>
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<p>(<b>A</b>) Effects of temperature on collagen CD of catfish skin at 221 nm; (<b>B</b>) Collagen denaturation temperature of southern catfish skin. * Indicates significant differences between RF and CF (<span class="html-italic">p</span> &lt; 0.05).</p>
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<p>DPPH radical scavenging rate of skin collagen from southern catfish skin under raw food (RF) and cooked food (CF) feeding.</p>
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<p>Effect of salinity on collagen fibril formation rate of skin collagen from southern catfish skin under raw food (RF) and cooked food (CF) feeding.</p>
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<p>Scanning electron microscope images of collagen fibrils obtained from the skin of southern catfish. (<b>A</b>–<b>C</b>): raw food (RF) feeding, NaCl concentration was 0 mM, 140 mM, and 280 mM, respectively; (<b>D</b>–<b>F</b>): cooked food (CF) feeding, NaCl concentration was 0 mM, 140 mM, and 280 mM, respectively.</p>
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<p>Effect of salinity on the diameter of skin collagen fibrils from southern catfish skin under raw food (RF) and cooked food (CF) feeding (<span class="html-italic">n</span> = 150, unit in nm). Different lowercase letters indicate differences among different salinities and groups (<span class="html-italic">p</span> &lt; 0.05).</p>
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<p>Effects of different feeding modes on gene expression of type I collagen in southern catfish skin. RF, raw food feeding; CF, cooked food feeding. * Indicates significant differences among different salinities (<span class="html-italic">p</span> &lt; 0.05).</p>
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53 pages, 1743 KiB  
Review
Applications of Chitosan in Prevention and Treatment Strategies of Infectious Diseases
by Genada Sinani, Melike Sessevmez and Sevda Şenel
Pharmaceutics 2024, 16(9), 1201; https://doi.org/10.3390/pharmaceutics16091201 - 13 Sep 2024
Viewed by 357
Abstract
Chitosan is the most commonly investigated functional cationic biopolymer in a wide range of medical applications due to its promising properties such as biocompatibility, biodegradability, and bioadhesivity, as well as its numerous bioactive properties. Within the last three decades, chitosan and its derivatives [...] Read more.
Chitosan is the most commonly investigated functional cationic biopolymer in a wide range of medical applications due to its promising properties such as biocompatibility, biodegradability, and bioadhesivity, as well as its numerous bioactive properties. Within the last three decades, chitosan and its derivatives have been investigated as biomaterials for drug and vaccine delivery systems, besides for their bioactive properties. Due to the functional groups in its structure, it is possible to tailor the delivery systems with desired properties. There has been a great interest in the application of chitosan-based systems also for the prevention and treatment of infectious diseases, specifically due to their antimicrobial, antiviral, and immunostimulatory effects. In this review, recent applications of chitosan in the prevention and treatment of infectious diseases are reviewed, and possibilities and limitations with regards to technical and regulatory aspects are discussed. Finally, the future perspectives on utilization of chitosan as a biomaterial are discussed. Full article
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<p>Chemical structure of chitin (<b>a</b>) and chitosan (<b>b</b>,<b>c</b>).</p>
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<p>Potential of chitosan for prevention and treatment of infectious diseases.</p>
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