International Journal of Molecular Sciences

22 pages, 4078 KiB

Open AccessArticle

Dendrobium nobile Lindley Administration Attenuates Atopic Dermatitis-like Lesions by Modulating Immune Cells

by Sooyeon Hong, Eun-Young Kim, Seo-Eun Lim, Jae-Hyun Kim, Youngjoo Sohn and Hyuk-Sang Jung

Int. J. Mol. Sci. 2022, 23(8), 4470; https://doi.org/10.3390/ijms23084470 - 18 Apr 2022

Cited by 20 | Viewed by 3837

Atopic dermatitis (AD) is a chronic inflammatory skin disease that can significantly affect daily life by causing sleep disturbance due to extreme itching. In addition, if the symptoms of AD are severe, it can cause mental disorders such as ADHD and suicidal ideation. [...] Read more.

Atopic dermatitis (AD) is a chronic inflammatory skin disease that can significantly affect daily life by causing sleep disturbance due to extreme itching. In addition, if the symptoms of AD are severe, it can cause mental disorders such as ADHD and suicidal ideation. Corticosteroid preparations used for general treatment have good effects, but their use is limited due to side effects. Therefore, it is essential to minimize the side effects and study effective treatment methods. Dendrobium nobile Lindley (DNL) has been widely used for various diseases, but to the best of our knowledge, its effect on AD has not yet been proven. In this study, the inhibitory effect of DNL on AD was confirmed in a DNCB-induced Balb/c mouse. In addition, the inhibitory efficacy of inflammatory cytokines in TNF-α/IFN-γ-induced HaCaT cells and PMACI-induced HMC-1 cells was confirmed. The results demonstrated that DNL decreased IgE, IL-6, IL-4, scratching behavior, SCORAD index, infiltration of mast cells and eosinophils and decreased the thickness of the skin. Additionally, DNL inhibited the expression of cytokines and inhibited the MAPK and NF-κB signaling pathways. This suggests that DNL inhibits cytokine expression, protein signaling pathway, and immune cells, thereby improving AD symptoms in mice. Full article

(This article belongs to the Special Issue Biological Interactions of Bioactive Natural Products)

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16 pages, 3912 KiB

Open AccessArticle

Coinfection of Porcine Circovirus 2 and Pseudorabies Virus Enhances Immunosuppression and Inflammation through NF-κB, JAK/STAT, MAPK, and NLRP3 Pathways

by Xue Li, Si Chen, Liying Zhang, Guyu Niu, Xinwei Zhang, Lin Yang, Weilong Ji and Linzhu Ren

Int. J. Mol. Sci. 2022, 23(8), 4469; https://doi.org/10.3390/ijms23084469 - 18 Apr 2022

Cited by 25 | Viewed by 4372

Abstract

Porcine circovirus 2 (PCV2) and pseudorabies virus (PRV) are economically important pathogens in swine. PCV2 and PRV coinfection can cause more severe neurological and respiratory symptoms and higher mortality of piglets. However, the exact mechanism involved in the coinfection of PRV and PCV2 [...] Read more.

Porcine circovirus 2 (PCV2) and pseudorabies virus (PRV) are economically important pathogens in swine. PCV2 and PRV coinfection can cause more severe neurological and respiratory symptoms and higher mortality of piglets. However, the exact mechanism involved in the coinfection of PRV and PCV2 and its pathogenesis remain unknown. Here, porcine kidney cells (PK-15) were infected with PCV2 and/or PRV, and then the activation of immune and inflammatory pathways was evaluated to clarify the influence of the coinfection on immune and inflammatory responses. We found that the coinfection of PCV2 and PRV can promote the activation of nuclear factor-κB (NF-κB), c-Jun N-terminal protein kinases (JNK), p38, and nod-like receptor protein 3 (NLRP3) pathways, thus enhancing the expression of interferon-γ (IFN-γ), interferon-λ1 (IFN-λ1), interferon-stimulated gene (ISG15), interleukin 6 (IL6), and interleukin 1β (IL1β). Meanwhile, PCV2 and PRV also inhibit the expression and signal transduction of IFN-β, tumor necrosis factor α (TNFα), and the Janus kinase-signal transducer and activator of transcription (JAK/STAT) pathway. In addition, PCV2 and PRV infection can also weaken extracellular-signal-regulated kinase (ERK) activity. These results indicate that the regulations of cellular antiviral immune responses and inflammatory responses mediated by NF-κB, JAK/STAT, mitogen-activated protein kinase (MAPK), and NLRP3 pathways, contribute to immune escape of PCV2 and PRV and host antiviral responses. Full article

(This article belongs to the Special Issue The Interaction between Cell and Virus)

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15 pages, 3815 KiB

Open AccessArticle

The Discovery of Potent SHP2 Inhibitors with Anti-Proliferative Activity in Breast Cancer Cell Lines

by Rose Ghemrawi, Mostafa Khair, Shaima Hasan, Raghad Aldulaymi, Shaikha S. AlNeyadi, Noor Atatreh and Mohammad A. Ghattas

Int. J. Mol. Sci. 2022, 23(8), 4468; https://doi.org/10.3390/ijms23084468 - 18 Apr 2022

Cited by 2 | Viewed by 5486

Abstract

Despite available treatments, breast cancer is the leading cause of cancer-related death. Knowing that the tyrosine phosphatase SHP2 is a regulator in tumorigenesis, developing inhibitors of SHP2 in breast cells is crucial. Our study investigated the effects of new compounds, purchased from NSC, [...] Read more.

Despite available treatments, breast cancer is the leading cause of cancer-related death. Knowing that the tyrosine phosphatase SHP2 is a regulator in tumorigenesis, developing inhibitors of SHP2 in breast cells is crucial. Our study investigated the effects of new compounds, purchased from NSC, on the phosphatase activity of SHP2 and the modulation of breast cancer cell lines’ proliferation and viability. A combined ligand-based and structure-based virtual screening protocol was validated, then performed, against SHP2 active site. Top ranked compounds were tested via SHP2 enzymatic assay, followed by measuring IC50 values. Subsequently, hits were tested for their anti-breast cancer viability and proliferative activity. Our experiments identified three compounds 13030, 24198, and 57774 as SHP2 inhibitors, with IC50 values in micromolar levels and considerable selectivity over the analogous enzyme SHP1. Long MD simulations of 500 ns showed a very promising binding mode in the SHP2 catalytic pocket. Furthermore, these compounds significantly reduced MCF-7 breast cancer cells’ proliferation and viability. Interestingly, two of our hits can have acridine or phenoxazine cyclic system known to intercalate in ds DNA. Therefore, our novel approach led to the discovery of SHP2 inhibitors, which could act as a starting point in the future for clinically useful anticancer agents. Full article

(This article belongs to the Special Issue Computational Studies of Drugs and Biomolecules)

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(a) The four features of the top pharmacophore query no. 4. (b) Two known inhibitors overlaid on the said pharmacophore. (c) The measured distances and angles between the four pharmacoohoric features (d) The effect of using pharmacophore query no. 4 as pre-docking filter in a pilot showing the number of known SHP2 inhibitors retrieved (%Enrichment) at any given percentage of the top-ranked ligand library. Full article ">Figure 2
The workflow of the filter-based VS used against the SHP2 active site. Full article ">Figure 3
Remaining activity (%) of the SHP2 enzyme after treatment with the 35 tested compounds (at 100 μM) along with the control suramin. Data presented are the mean +/− SEM of three independent experiments. Full article ">Figure 4
The effect of the compounds NSC 13030, 24198, 57774, and 137420 on MCF-7 and MDA-MB-231 proliferation (a) and viability (b). Cells were incubated with increasing concentrations of compounds in culture medium for 48 h. The viability and the proliferative response were assessed. Data presented are the mean ± SEM of three independent experiments. * p < 0.05. Continuous lines are for compounds, and dashed lines are for the vehicle DMSO. Full article ">Figure 5
(a) The RMSD plot of the 500 ns MD simulations of compound 13030 along with (b) the hydrogen bonding plot, (c) the top-cluster conformation filling (blue spheres) in the SHP2 active site (gray surface), and (d) the ligand 3D binding mode showing the key interactions made with the surrounding residues. Full article ">Figure 6
(a) The RMSD plot of the 500 ns MD simulations of compound 24198 along with (b) the hydrogen bonding plot, (c) the top-cluster conformation filling (blue spheres) in the SHP2 active site (gray surface), and (d) the ligand 3D binding mode showing the key interactions made with the surrounding residues. Full article ">Figure 7
(a) The RMSD plot of the 500 ns MD simulations of compound 57774 along with (b) the hydrogen bonding plot, (c) the top-cluster conformation filling (blue spheres) in the SHP2 active site (gray surface), and (d) the ligand 3D binding mode showing the key interactions made with the surrounding residues. Full article ">

17 pages, 793 KiB

Open AccessReview

Recent Experimental Studies of Maternal Obesity, Diabetes during Pregnancy and the Developmental Origins of Cardiovascular Disease

by Stephanie M. Kereliuk and Vernon W. Dolinsky

Int. J. Mol. Sci. 2022, 23(8), 4467; https://doi.org/10.3390/ijms23084467 - 18 Apr 2022

Cited by 22 | Viewed by 8279

Abstract

Globally, cardiovascular disease remains the leading cause of death. Most concerning is the rise in cardiovascular risk factors including obesity, diabetes and hypertension among youth, which increases the likelihood of the development of earlier and more severe cardiovascular disease. While lifestyle factors are [...] Read more.

Globally, cardiovascular disease remains the leading cause of death. Most concerning is the rise in cardiovascular risk factors including obesity, diabetes and hypertension among youth, which increases the likelihood of the development of earlier and more severe cardiovascular disease. While lifestyle factors are involved in these trends, an increasing body of evidence implicates environmental exposures in early life on health outcomes in adulthood. Maternal obesity and diabetes during pregnancy, which have increased dramatically in recent years, also have profound effects on fetal growth and development. Mounting evidence is emerging that maternal obesity and diabetes during pregnancy have lifelong effects on cardiovascular risk factors and heart disease development. However, the mechanisms responsible for these observations are unknown. In this review, we summarize the findings of recent experimental studies, showing that maternal obesity and diabetes during pregnancy affect energy metabolism and heart disease development in the offspring, with a focus on the mechanisms involved. We also evaluate early proof-of-concept studies for interventions that could mitigate maternal obesity and gestational diabetes-induced cardiovascular disease risk in the offspring. Full article

(This article belongs to the Special Issue Risk Factors and Molecular Mechanisms of Gestational Diabetes III)

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11 pages, 2822 KiB

Open AccessArticle

AcidoCEST-UTE MRI Reveals an Acidic Microenvironment in Knee Osteoarthritis

by Alecio F. Lombardi, Yajun Ma, Hyungseok Jang, Saeed Jerban, Qingbo Tang, Adam C. Searleman, Robert Scott Meyer, Jiang Du and Eric Y. Chang

Int. J. Mol. Sci. 2022, 23(8), 4466; https://doi.org/10.3390/ijms23084466 - 18 Apr 2022

Cited by 23 | Viewed by 5317 | Correction

Abstract

A relationship between an acidic pH in the joints, osteoarthritis (OA), and pain has been previously demonstrated. Acidosis Chemical Exchange Saturation Transfer (acidoCEST) indirectly measures the extracellular pH through the assessment of the exchange of protons between amide groups on iodinated contrast agents [...] Read more.

A relationship between an acidic pH in the joints, osteoarthritis (OA), and pain has been previously demonstrated. Acidosis Chemical Exchange Saturation Transfer (acidoCEST) indirectly measures the extracellular pH through the assessment of the exchange of protons between amide groups on iodinated contrast agents and bulk water. It is possible to estimate the extracellular pH in the osteoarthritic joint using acidoCEST MRI. However, conventional MR sequences cannot image deep layers of cartilage, meniscus, ligaments, and other musculoskeletal tissues that present with short echo time and fast signal decay. Ultrashort echo time (UTE) MRI, on the other hand, has been used successfully to image those joint tissues. Here, our goal is to compare the pH measured in the knee joints of volunteers without OA and patients with severe OA using acidoCEST-UTE MRI. Patients without knee OA and patients with severe OA were examined using acidoCEST-UTE MRI and the mean pH of cartilage, meniscus, and fluid was calculated. Additionally, the relationship between the pH measurements and the Knee Injury and Osteoarthritis Outcome Score (KOOS) was investigated. AcidoCEST-UTE MRI can detect significant differences in the pH of knee cartilage, meniscus, and fluid between joints without and with OA, with OA showing lower pH values. In addition, symptoms and knee-joint function become worse at lower pH measurements. Full article

(This article belongs to the Special Issue New Advances in Osteoarthritis)

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Representative image examples from patients without OA (A) and with OA (B). Sagittal PD-weighted (first row), low-power acido-CEST UTE (second row), high-power acido-CEST UTE (third row), and pH pixel maps (fourth row) of cartilage, meniscus, and fluid. The pH is directly correlated with the radiofrequency power mismatch (RPM) measurements, as described in Equations (3) and (4). Note the higher pH values (yellow and red colors) in patients without OA compared with patients with OA (blue colors). Full article ">Figure 2
Boxplots of mean pH measurements versus groups for all ROIs. Significant differences are observed between the two groups. ***: p values lower than 0.001. Full article ">Figure 3
Boxplots of mean pH measurements versus each group of participants (No OA and OA) for ROIs drew in cartilage (A), meniscus (B), and fluid (C). Significant differences in pH measurements are observed between groups with lower pH in patients with OA compared with patients without OA. “***”: p values lower than 0.001; “**”: p values lower than 0.01; “*”: p values lower than 0.05. Full article ">Figure 4
Boxplots of mean pH measurements versus ROIs in patients without OA (A) and with advanced OA (B). No significant differences were found, except for the pH of cartilage and meniscus in the OA group (p = 0.024). “*”: p values lower than 0.05; “•”: outliers. Full article ">Figure 5
Scatterplots of pH versus KOOS scores and the visual analog pain score. Strong direct correlations were observed for all KOOS subscale scores (A–F). There was a strong inverse correlation between pH measurements and the visual analog pain score (G). KOOS: Knee Injury and Osteoarthritis Outcome Score; ADL: activities of daily living; Sports/Rec: sports and recreation activities; QOL: quality of life; PF: patellofemoral. Full article ">

27 pages, 4740 KiB

Open AccessArticle

Functional Characterization of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G)

by Roua Hassoun, Constanze Erdmann, Sebastian Schmitt, Setsuko Fujita-Becker, Andreas Mügge, Rasmus R. Schröder, Matthias Geyer, Mina Borbor, Kornelia Jaquet, Nazha Hamdani and Hans Georg Mannherz

Int. J. Mol. Sci. 2022, 23(8), 4465; https://doi.org/10.3390/ijms23084465 - 18 Apr 2022

Cited by 5 | Viewed by 2969

Abstract

Human wild type (wt) cardiac α-actin and its mutants p.A295S or p.R312H and p.E361G correlated with hypertrophic or dilated cardiomyopathy, respectively, were expressed by using the baculovirus/Sf21 insect cell system. The c-actin variants inhibited DNase I, indicating maintenance of their native state. Electron [...] Read more.

Human wild type (wt) cardiac α-actin and its mutants p.A295S or p.R312H and p.E361G correlated with hypertrophic or dilated cardiomyopathy, respectively, were expressed by using the baculovirus/Sf21 insect cell system. The c-actin variants inhibited DNase I, indicating maintenance of their native state. Electron microscopy showed the formation of normal appearing actin filaments though they showed mutant specific differences in length and straightness correlating with their polymerization rates. TRITC-phalloidin staining showed that p.A295S and p.R312H exhibited reduced and the p.E361G mutant increased lengths of their formed filaments. Decoration of c-actins with cardiac tropomyosin (cTm) and troponin (cTn) conveyed Ca²⁺-sensitivity of the myosin-S1 ATPase stimulation, which was higher for the HCM p.A295S mutant and lower for the DCM p.R312H and p.E361G mutants than for wt c-actin. The lower Ca²⁺-sensitivity of myosin-S1 stimulation by both DCM actin mutants was corrected by the addition of levosimendan. Ca²⁺-dependency of the movement of pyrene-labeled cTm along polymerized c-actin variants decorated with cTn corresponded to the relations observed for the myosin-S1 ATPase stimulation though shifted to lower Ca²⁺-concentrations. The N-terminal C0C2 domain of cardiac myosin-binding protein-C increased the Ca²⁺-sensitivity of the pyrene-cTM movement of bovine, recombinant wt, p.A295S, and p.E361G c-actins, but not of the p.R312H mutant, suggesting decreased affinity to cTm. Full article

(This article belongs to the Section Molecular Biology)

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19 pages, 3521 KiB

Open AccessArticle

Chaperone Hsp70 (HSPA1) Is Involved in the Molecular Mechanisms of Sleep Cycle Integration

by Valentina V. Simonova, Mikhail A. Guzeev, Irina V. Ekimova and Yuri F. Pastukhov

Int. J. Mol. Sci. 2022, 23(8), 4464; https://doi.org/10.3390/ijms23084464 - 18 Apr 2022

Cited by 5 | Viewed by 2802

Abstract

The molecular mechanisms of sleep cycle integration at the beginning and the end of the inactive period are not clear. Sleep cycles with a predominance of deep slow-wave sleep (SWS) seem to be associated with accelerated protein synthesis in the brain. The inducible [...] Read more.

The molecular mechanisms of sleep cycle integration at the beginning and the end of the inactive period are not clear. Sleep cycles with a predominance of deep slow-wave sleep (SWS) seem to be associated with accelerated protein synthesis in the brain. The inducible Hsp70 chaperone corrects protein conformational changes and has protective properties. This research explores (1) whether the Hspa1 gene encoding Hsp70 protein activates during the daily rapid-eye-movement sleep (REMS) maximum, and (2) whether a lower daily deep SWS maximum affects the Hspa1 expression level during the subsequent REMS. Combining polysomnography in male Wistar rats, RT-qPCR, and Western blotting, we reveal a three-fold Hspa1 upregulation in the nucleus reticularis pontis oralis, which regulates REMS. Hspa1 expression increases during the daily REMS maximum, 5–7 h after the natural peak of deep SWS. Using short-term selective REMS deprivation, we demonstrate that REMS rebound after deprivation exceeds the natural daily maximum, but it is not accompanied by Hspa1 upregulation. The results suggest that a high proportion of deep SWS, usually observed after sleep onset, is a necessary condition for Hspa1 upregulation during subsequent REMS. The data obtained can inform the understanding of the molecular mechanisms integrating SWS and REMS and key biological function(s) of sleep. Full article

(This article belongs to the Collection State-of-the-Art Molecular Neurobiology in Russia)

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13 pages, 1149 KiB

Open AccessReview

MAPK Cascades and Transcriptional Factors: Regulation of Heavy Metal Tolerance in Plants

by Shaocui Li, Xiaojiao Han, Zhuchou Lu, Wenmin Qiu, Miao Yu, Haiying Li, Zhengquan He and Renying Zhuo

Int. J. Mol. Sci. 2022, 23(8), 4463; https://doi.org/10.3390/ijms23084463 - 18 Apr 2022

Cited by 61 | Viewed by 5569

Abstract

In nature, heavy metal (HM) stress is one of the most destructive abiotic stresses for plants. Heavy metals produce toxicity by targeting key molecules and important processes in plant cells. The mitogen-activated protein kinase (MAPK) cascade transfers the signals perceived by cell membrane [...] Read more.

In nature, heavy metal (HM) stress is one of the most destructive abiotic stresses for plants. Heavy metals produce toxicity by targeting key molecules and important processes in plant cells. The mitogen-activated protein kinase (MAPK) cascade transfers the signals perceived by cell membrane surface receptors to cells through phosphorylation and dephosphorylation and targets various effector proteins or transcriptional factors so as to result in the stress response. Signal molecules such as plant hormones, reactive oxygen species (ROS), and nitric oxide (NO) can activate the MAPK cascade through differentially expressed genes, the activation of the antioxidant system and synergistic crosstalk between different signal molecules in order to regulate plant responses to HMs. Transcriptional factors, located downstream of MAPK, are key factors in regulating plant responses to heavy metals and improving plant heavy metal tolerance and accumulation. Thus, understanding how HMs activate the expression of the genes related to the MAPK cascade pathway and then phosphorylate those transcriptional factors may allow us to develop a regulation network to increase our knowledge of HMs tolerance and accumulation. This review highlighted MAPK pathway activation and responses under HMs and mainly focused on the specificity of MAPK activation mediated by ROS, NO and plant hormones. Here, we also described the signaling pathways and their interactions under heavy metal stresses. Moreover, the process of MAPK phosphorylation and the response of downstream transcriptional factors exhibited the importance of regulating targets. It was conducive to analyzing the molecular mechanisms underlying heavy metal accumulation and tolerance. Full article

(This article belongs to the Special Issue Heavy Metals Accumulation, Toxicity and Detoxification in Plants 2.0)

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30 pages, 1201 KiB

Open AccessReview

Bone Marrow Niches of Hematopoietic Stem and Progenitor Cells

by Oleg Kandarakov, Alexander Belyavsky and Ekaterina Semenova

Int. J. Mol. Sci. 2022, 23(8), 4462; https://doi.org/10.3390/ijms23084462 - 18 Apr 2022

Cited by 40 | Viewed by 9074

Abstract

The mammalian hematopoietic system is remarkably efficient in meeting an organism’s vital needs, yet is highly sensitive and exquisitely regulated. Much of the organismal control over hematopoiesis comes from the regulation of hematopoietic stem cells (HSCs) by specific microenvironments called niches in bone [...] Read more.

The mammalian hematopoietic system is remarkably efficient in meeting an organism’s vital needs, yet is highly sensitive and exquisitely regulated. Much of the organismal control over hematopoiesis comes from the regulation of hematopoietic stem cells (HSCs) by specific microenvironments called niches in bone marrow (BM), where HSCs reside. The experimental studies of the last two decades using the most sophisticated and advanced techniques have provided important data on the identity of the niche cells controlling HSCs functions and some mechanisms underlying niche-HSC interactions. In this review we discuss various aspects of organization and functioning of the HSC cell niche in bone marrow. In particular, we review the anatomy of BM niches, various cell types composing the niche, niches for more differentiated cells, metabolism of HSCs in relation to the niche, niche aging, leukemic transformation of the niche, and the current state of HSC niche modeling in vitro. Full article

(This article belongs to the Special Issue Hematopoietic System under Physiological Conditions and Following Hematopoietic Reconstitution or Stress)

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19 pages, 2377 KiB

Open AccessReview

Serrated Colorectal Lesions: An Up-to-Date Review from Histological Pattern to Molecular Pathogenesis

by Martino Mezzapesa, Giuseppe Losurdo, Francesca Celiberto, Salvatore Rizzi, Antonio d’Amati, Domenico Piscitelli, Enzo Ierardi and Alfredo Di Leo

Int. J. Mol. Sci. 2022, 23(8), 4461; https://doi.org/10.3390/ijms23084461 - 18 Apr 2022

Cited by 33 | Viewed by 10256

Abstract

Until 2010, colorectal serrated lesions were generally considered as harmless lesions and reported as hyperplastic polyps (HPs) by pathologists and gastroenterologists. However, recent evidence showed that they may bear the potential to develop into colorectal carcinoma (CRC). Therefore, the World Health Organization (WHO) [...] Read more.

Until 2010, colorectal serrated lesions were generally considered as harmless lesions and reported as hyperplastic polyps (HPs) by pathologists and gastroenterologists. However, recent evidence showed that they may bear the potential to develop into colorectal carcinoma (CRC). Therefore, the World Health Organization (WHO) classification has identified four categories of serrated lesions: hyperplastic polyps (HPs), sessile serrated lesions (SSLs), traditional serrated adenoma (TSAs) and unclassified serrated adenomas. SSLs with dysplasia and TSAs are the most common precursors of CRC. CRCs arising from serrated lesions originate via two different molecular pathways, namely sporadic microsatellite instability (MSI) and the CpG island methylator phenotype (CIMP), the latter being considered as the major mechanism that drives the serrated pathway towards CRC. Unlike CRCs arising through the adenoma–carcinoma pathway, APC-inactivating mutations are rarely shown in the serrated neoplasia pathway. Full article

(This article belongs to the Special Issue 25th Anniversary of IJMS: Advances in Biochemistry)

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14 pages, 8625 KiB

Open AccessReview

The Structural and the Functional Aspects of Intercellular Communication in iPSC-Cardiomyocytes

by Eva Kiss, Carolin Fischer, Jan-Mischa Sauter, Jinmeng Sun and Nina D. Ullrich

Int. J. Mol. Sci. 2022, 23(8), 4460; https://doi.org/10.3390/ijms23084460 - 18 Apr 2022

Cited by 5 | Viewed by 3677

Abstract

Recent advances in the technology of producing novel cardiomyocytes from induced pluripotent stem cells (iPSC-cardiomyocytes) fuel new hope for future clinical applications. The use of iPSC-cardiomyocytes is particularly promising for the therapy of cardiac diseases such as myocardial infarction, where these cells could [...] Read more.

Recent advances in the technology of producing novel cardiomyocytes from induced pluripotent stem cells (iPSC-cardiomyocytes) fuel new hope for future clinical applications. The use of iPSC-cardiomyocytes is particularly promising for the therapy of cardiac diseases such as myocardial infarction, where these cells could replace scar tissue and restore the functionality of the heart. Despite successful cardiogenic differentiation, medical applications of iPSC-cardiomyocytes are currently limited by their pronounced immature structural and functional phenotype. This review focuses on gap junction function in iPSC-cardiomyocytes and portrays our current understanding around the structural and the functional limitations of intercellular coupling and viable cardiac graft formation involving these novel cardiac muscle cells. We further highlight the role of the gap junction protein connexin 43 as a potential target for improving cell–cell communication and electrical signal propagation across cardiac tissue engineered from iPSC-cardiomyocytes. Better insight into the mechanisms that promote functional intercellular coupling is the foundation that will allow the development of novel strategies to combat the immaturity of iPSC-cardiomyocytes and pave the way toward cardiac tissue regeneration. Full article

(This article belongs to the Special Issue The Magnificent World of Induced Pluripotent Stem Cell-Derived Cardiomyocytes)

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19 pages, 1861 KiB

Open AccessArticle

Metabolic Alteration Analysis of Steroid Hormones in Niemann–Pick Disease Type C Model Cell Using Liquid Chromatography/Tandem Mass Spectrometry

by Ai Abe, Masamitsu Maekawa, Toshihiro Sato, Yu Sato, Masaki Kumondai, Hayato Takahashi, Masafumi Kikuchi, Katsumi Higaki, Jiro Ogura and Nariyasu Mano

Int. J. Mol. Sci. 2022, 23(8), 4459; https://doi.org/10.3390/ijms23084459 - 18 Apr 2022

Cited by 11 | Viewed by 3523

Abstract

Niemann–Pick disease type C (NPC) is an autosomal recessive disease caused by a functional deficiency of cholesterol-transporting proteins in lysosomes, and exhibits various clinical symptoms. Since mitochondrial dysfunction in NPC has recently been reported, cholesterol catabolism to steroid hormones may consequently be impaired. [...] Read more.

Niemann–Pick disease type C (NPC) is an autosomal recessive disease caused by a functional deficiency of cholesterol-transporting proteins in lysosomes, and exhibits various clinical symptoms. Since mitochondrial dysfunction in NPC has recently been reported, cholesterol catabolism to steroid hormones may consequently be impaired. In this study, we developed a comprehensive steroid hormone analysis method using liquid chromatography/tandem mass spectrometry (LC–MS/MS) and applied it to analyze changes in steroid hormone concentrations in NPC model cells. We investigated the analytical conditions for simultaneous LC–MS/MS analysis, which could be readily separated from each other and showed good reproducibility. The NPC phenotype was verified as an NPC model with mitochondrial abnormalities using filipin staining and organelle morphology observations. Steroid hormones in the cell suspension and cell culture medium were also analyzed. Steroid hormone analysis indicated that the levels of six steroid hormones were significantly decreased in the NPC model cell and culture medium compared to those in the wild-type cell and culture medium. These results indicate that some steroid hormones change during NPC pathophysiology and this change is accompanied by mitochondrial abnormalities. Full article

(This article belongs to the Special Issue Mass Spectrometry Techniques for Biomarker Discovery)

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Chemical structures of steroid hormones analyzed in this study. Full article ">Figure 2
Comparison of peak intensities in negative ion detection mode for various mobile phases. (a) Acidic conditions using mobile phase A: Formic acid/water (0.1:100, v/v) and mobile phase B: Formic acid/MeOH/ACN (0.1:50:50, v/v/v); (b) weak basic conditions using mobile phase A: Formic acid/water (0.1:100, v/v) and mobile phase B: 1 M ammonium acetate aqueous solution/MeOH/ACN (0.1:50:50, v/v/v); (c) basic conditions using mobile phase A: 28% ammonium aqueous solution/water (0.1:100, v/v) and mobile phase B: 28% aqueous ammonium solution/MeOH/ACN (0.1:50:50, v/v/v). ACN, acetonitrile; MeOH, methanol. Full article ">Figure 3
SRM chromatogram under optimized conditions. (a) Positive ion detection mode, (b) negative ion detection mode. SRM, selected reaction monitoring. Full article ">Figure 4
The amounts of steroid hormones in the cells. (a) Androgens, (b) glucocorticoids and mineralocorticoid, (c) progestins, and (d) estrogens. White and black bars indicate wild-type and NPC model cells, respectively. * indicates significant differences observed using Wilcoxon’s test (p < 0.05). NPC, Niemann–Pick disease type C; N.Q., not quantified. Full article ">Figure 5
Steroid hormone concentrations in the medium. (a) Androgens, (b) glucocorticoids and mineralocorticoid, (c) progestins, (d) estrogens. White and black bars indicate wild-type and NPC model cells, respectively. * indicates significant differences observed using Wilcoxon’s test (p < 0.05). NPC, Niemann–Pick disease type C; N.Q., not quantified. Full article ">Figure 6
The hypothesis of an altered steroid hormone metabolism in NPC. Blue boxes indicate a decrease; gray boxes indicate no change. The open text indicates the absence of analytes in this study. CYP11A1, cytochrome P450 family 11 subfamily A member 1; 3β-HSD, 3β-hydroxysteroid dehydrogenase; 17β-HSD, 17β-hydroxysteroid dehydrogenase; CYP11B1, 11β-hydroxylase; CYP11B2, aldosterone synthase; CYP17A, 17α-hydroxylase/17,20 lyase; CYP19A, aromatase; CYP21A, 21-hydroxylase; DHEA, dehydroepiandrosterone; StAR, steroidogenic acute regulatory protein. Full article ">

19 pages, 2118 KiB

Open AccessReview

Applications of Metal-Organic Frameworks as Drug Delivery Systems

by Bianca Maranescu and Aurelia Visa

Int. J. Mol. Sci. 2022, 23(8), 4458; https://doi.org/10.3390/ijms23084458 - 18 Apr 2022

Cited by 119 | Viewed by 9437

Abstract

In the last decade, metal organic frameworks (MOFs) have shown great prospective as new drug delivery systems (DDSs) due to their unique properties: these materials exhibit fascinating architectures, surfaces, composition, and a rich chemistry of these compounds. The DSSs allow the release of [...] Read more.

In the last decade, metal organic frameworks (MOFs) have shown great prospective as new drug delivery systems (DDSs) due to their unique properties: these materials exhibit fascinating architectures, surfaces, composition, and a rich chemistry of these compounds. The DSSs allow the release of the active pharmaceutical ingredient to accomplish a desired therapeutic response. Over the past few decades, there has been exponential growth of many new classes of coordination polymers, and MOFs have gained popularity over other identified systems due to their higher biocompatibility and versatile loading capabilities. This review presents and assesses the most recent research, findings, and challenges associated with the use of MOFs as DDSs. Among the most commonly used MOFs for investigated-purpose MOFs, coordination polymers and metal complexes based on synthetic and natural polymers, are well known. Specific attention is given to the stimuli- and multistimuli-responsive MOFs-based DDSs. Of great interest in the COVID-19 pandemic is the use of MOFs for combination therapy and multimodal systems. Full article

(This article belongs to the Section Molecular Pharmacology)

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MOFs synthesis—alternative reaction conditions. Features shown in blue are advantages and black are disadvantages for all described methods. Full article ">Figure 2
Number of publications from Web of Science: (A) “Metal Organic Frameworks” and (B) “Metal Organic Frameworks and Drug Delivery Systems”, from 1996 and 2009, through 21 March 2022. Full article ">Figure 3
Some examples of drugs delivered by MOFs as DDS: (a)—indocianine green, (b)—doxorubicin hydrochloride, (c)—5-fluorouracil, (d)—caffeine, (e)—cidofovir, (f)—acid folic, (g)—calcein, (h)—curcumin. Full article ">Figure 4
Encapsulation, direct assembly, and post-synthesis of cargo-loading strategies for MOFs. Reproduced with permission from [<a href="#B51-ijms-23-04458" class="html-bibr">51</a>]; Copyright (Wang, 2018), RSC J. Mater. Chem. B. Full article ">Figure 5
Synthesis of O2@UiO-66@ICG@RBC (A). Schematic mechanism of NIR-triggered O2 enhanced and discharging PDT (B). Reproduced with permission from [<a href="#B81-ijms-23-04458" class="html-bibr">81</a>]; Copyright (Gao, 2018), Elsevier Biomaterials. Full article ">Figure 6
Schematic illustration of metal–organic frameworks (MOFs)-based stimuli-responsive system for drug delivery. Reproduced from reference [<a href="#B110-ijms-23-04458" class="html-bibr">110</a>]; Copyright (Cai, 2018) Wiley Online Library. Full article ">

15 pages, 1005 KiB

Open AccessReview

Psoriasis and Systemic Inflammatory Disorders

by Tomoko Tashiro and Yu Sawada

Int. J. Mol. Sci. 2022, 23(8), 4457; https://doi.org/10.3390/ijms23084457 - 18 Apr 2022

Cited by 47 | Viewed by 6873

Abstract

Psoriasis is a representative inflammatory skin disease occupied by large surface involvement. As inflammatory cells and cytokines can systemically circulate in various organs, it has been speculated that psoriatic skin inflammation influences the systemic dysfunction of various organs. Recent updates of clinical studies [...] Read more.

Psoriasis is a representative inflammatory skin disease occupied by large surface involvement. As inflammatory cells and cytokines can systemically circulate in various organs, it has been speculated that psoriatic skin inflammation influences the systemic dysfunction of various organs. Recent updates of clinical studies and experimental studies showed the important interaction of psoriasis to systemic inflammatory diseases. Furthermore, the importance of systemic therapy in severe psoriasis is also highlighted to prevent the development of systemic inflammatory diseases. In this review, we introduced representative systemic inflammatory diseases associated with psoriasis and the detailed molecular mechanisms. Full article

(This article belongs to the Section Molecular Pathology, Diagnostics, and Therapeutics)

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6 pages, 218 KiB

Open AccessEditorial

Microgravity and Space Medicine 2.0

by Daniela Grimm

Int. J. Mol. Sci. 2022, 23(8), 4456; https://doi.org/10.3390/ijms23084456 - 18 Apr 2022

Cited by 1 | Viewed by 3188

Abstract

This Special Issue (SI), “Microgravity and Space Medicine 2 [...] Full article

(This article belongs to the Special Issue Microgravity and Space Medicine 2.0)

20 pages, 6030 KiB

Open AccessArticle

Transcriptomic Data Meta-Analysis Sheds Light on High Light Response in Arabidopsis thaliana L.

by Aleksandr V. Bobrovskikh, Ulyana S. Zubairova, Eugeniya I. Bondar, Viktoriya V. Lavrekha and Alexey V. Doroshkov

Int. J. Mol. Sci. 2022, 23(8), 4455; https://doi.org/10.3390/ijms23084455 - 18 Apr 2022

Cited by 8 | Viewed by 4050

Abstract

The availability and intensity of sunlight are among the major factors of growth, development and metabolism in plants. However, excessive illumination disrupts the electronic balance of photosystems and leads to the accumulation of reactive oxygen species in chloroplasts, further mediating several regulatory mechanisms [...] Read more.

The availability and intensity of sunlight are among the major factors of growth, development and metabolism in plants. However, excessive illumination disrupts the electronic balance of photosystems and leads to the accumulation of reactive oxygen species in chloroplasts, further mediating several regulatory mechanisms at the subcellular, genetic, and molecular levels. We carried out a comprehensive bioinformatic analysis that aimed to identify genetic systems and candidate transcription factors involved in the response to high light stress in Arabidopsis thaliana L. using resources GEO NCBI, string-db, ShinyGO, STREME, and Tomtom, as well as programs metaRE, CisCross, and Cytoscape. Through the meta-analysis of five transcriptomic experiments, we selected a set of 1151 differentially expressed genes, including 453 genes that compose the gene network. Ten significantly enriched regulatory motifs for TFs families ZF-HD, HB, C2H2, NAC, BZR, and ARID were found in the promoter regions of differentially expressed genes. In addition, we predicted families of transcription factors associated with the duration of exposure (RAV, HSF), intensity of high light treatment (MYB, REM), and the direction of gene expression change (HSF, S1Fa-like). We predicted genetic components systems involved in a high light response and their expression changes, potential transcriptional regulators, and associated processes. Full article

(This article belongs to the Special Issue Plant Biology and Biotechnology: Focus on Genomics and Bioinformatics)

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Figure 1
Interaction of data searching and processing steps in the framework large-scale systematic analysis of the high light response regulation at different levels. Full article ">Figure 2
Sets of up- and downregulated differentially expressed genes (DEGs) allowed us to identify the correspondence between enriched transcription factor families and directions of expression changes, as well as PPFD levels and duration of high light expose in treatments. (a) The distribution of DEGs between datasets (up- and downregulated genes are marked by red and blue, respectively); (b) the distribution of upregulated DEGs (marked by red) and downregulated DEGs (marked by blue) in individual experimental datasets indicated by PPFD levels and duration of high light exposure (gray color indicate the Ws-0 ecotype); (c) the distribution of the enriched transcription factor families corresponding to individual experimental datasets; (d–f) the results of a multivariate analysis between experimental conditions (B1) and the sets of transcriptional regulators (B2) by the 2B-PLS method. The percentages of covariance for each axis, as well as families of transcription factors that have more than 0.15 of load modulus, are shown in the corresponding scatter plots. The point size represents the treatment duration. The point intensity represents the PPFD. Red color corresponds to upregulated genes, blue color corresponds to downregulated genes. The main differentiator of the first axis (d) is the light intensity. The second axis (e) distinguishes between upregulating and downregulating DEGs. The third axis (f) classifies the treatment by their duration. Full article ">Figure 3
Prediction of enriched motifs in promoters of selected DEGs. Prediction of cis-regulatory elements in the upstream regions enriched with hexamers and their genetic targets based on Arabidopsis thaliana L. transcriptomic meta-analysis. (a) The general scheme of pipeline of transcriptome regulators predictions. Promoters 1500 bp upstream regions of top 1151 genes (from transcriptomic meta-analysis) were used to predict 61 overpresented hexamers (p-value < 0.05) (see <a href="#app1-ijms-23-04455" class="html-app">Supplementary Tables S5–S7</a>). After, 10 enriched motifs were identified by STREME and Tomtom tools (see <a href="#app1-ijms-23-04455" class="html-app">Supplementary File S10</a>). (b) Cluster analysis of identified motifs. (c) Venn diagrams for sets of genes for which isolated motifs are enriched in their promoter regions. Full article ">Figure 4
The reconstructed gene network split by 21 connectivity clusters corresponding to functional modules of high light stress response regulation. Full article ">Figure 5
The high light response gene network layouts for distributions of (a) genes associated with red, blue, and visible light response, photoreceptors, and transcription factors; (b) genes associated with circadian rhythm, photosynthesis, flavonoid biosynthesis, response to jasmonic acid, nitrogen compound, and ROS, and developmental processes; (c) selected motifs; (d) phylostratigraphic index (PAI), low PAI corresponds to evolutionary ancient genes. Full article ">Figure 6
General scheme of molecular genetic mechanisms involved in high light response according to our meta-analysis. (a) Major high light response regulators; (b) the functional modules of the gene network identified as a result of the meta-analysis correspond to the main response pathways according to the color coding in (a). Full article ">

27 pages, 3449 KiB

Open AccessReview

CRISPR-Based Genome Editing: Advancements and Opportunities for Rice Improvement

by Workie Anley Zegeye, Mesfin Tsegaw, Yingxin Zhang and Liyong Cao

Int. J. Mol. Sci. 2022, 23(8), 4454; https://doi.org/10.3390/ijms23084454 - 18 Apr 2022

Cited by 22 | Viewed by 6666

Abstract

To increase the potentiality of crop production for future food security, new technologies for plant breeding are required, including genome editing technology—being one of the most promising. Genome editing with the CRISPR/Cas system has attracted researchers in the last decade as a safer [...] Read more.

To increase the potentiality of crop production for future food security, new technologies for plant breeding are required, including genome editing technology—being one of the most promising. Genome editing with the CRISPR/Cas system has attracted researchers in the last decade as a safer and easier tool for genome editing in a variety of living organisms including rice. Genome editing has transformed agriculture by reducing biotic and abiotic stresses and increasing yield. Recently, genome editing technologies have been developed quickly in order to avoid the challenges that genetically modified crops face. Developing transgenic-free edited plants without introducing foreign DNA has received regulatory approval in a number of countries. Several ongoing efforts from various countries are rapidly expanding to adopt the innovations. This review covers the mechanisms of CRISPR/Cas9, comparisons of CRISPR/Cas9 with other gene-editing technologies—including newly emerged Cas variants—and focuses on CRISPR/Cas9-targeted genes for rice crop improvement. We have further highlighted CRISPR/Cas9 vector construction model design and different bioinformatics tools for target site selection. Full article

(This article belongs to the Section Molecular Plant Sciences)

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Components of CRISPR/Cas9 system: (A). Genomic structures of the CRISPR/Cas system (top) and the engineered CRISPR/Cas9 system (bottom); (B). A schematic representation of the Cas9 protein structure. Domains includes REC (large recognition lobe) and RuvC (a nuclease domain), which is linked with an arginine-rich region. HNH is a second nuclease domain. PI is PAM-interacting domain; (C). The conformation of the Cas9–sgRNA complex in the process of DNA cleavage. The Cas9 endonuclease is targeted to DNA by a guide RNA which can be supplied as a two-part system consisting of crRNA and tracrRNA or as a single guide RNA, where the crRNA and tracrRNA are connected by a linker. Target recognition is facilitated by the protospacer-adjacent motif (PAM). Cleavage occurs on both strands (scissors) 3 bp upstream of the PAM. Full article ">Figure 2
The advantages and disadvantages of the CRISPR/Cas9 system over other approaches for genome editing. (A). Conventional gene targeting. (B). ZNFs and TALENs. (C). CRISPR/SpCas9. (D). CRISPR/NmCas9. The red arrow indicates the corresponding gene editing method with its features, advantages, and disadvantages. Full article ">Figure 3
Comparison of CRISPR/Cas9 with newly emerging CRISPR/Cas GE tools. (A) In the CRISPR/Cas9 system, Cas9 is a multicomponent protein and recognizes a G-rich PAM at the 3′ end of the target site. Both tracrRNA and crRNA are required to recruit Cas9. Then, the Cas9 creates a DSB, resulting in blunt ends. (B) In the CRISPR/Cas12a System, Cas12a is a single-component protein which recognizes T-rich PAM at the 5′ end of the target sequence; tracrRNA is not required. The DSB results in a 5′ overhang sticky ends with staggered cuts. (C) In the nuclear base-editing system, cytidine deaminase fused with dCas9 is used to target the desired site. There is no DSB, cytidine deaminase converts C directly into U, and during mismatch repair a C→ T substitution can be corrected when the modified strand is used as template. (D) Base editing in RNA. In the REPAIR system, “A-to-I” editing uses dCas13 fused to ADAR2. REPAIR uses 50- nucleotide RNA with a 50-nucleotide mRNA–gRNA duplex. “A–C” mismatch in the RNA–gRNA duplex determines the target A. RESCUE system editing “C-to-U.” The optimum results are achieved with a gRNA with a 30-nucleotide spacer. The target “C” is specified by an induced “C–C” or “C–U” mismatch in the mRNA–gRNA duplex. (E) Prime editing. (a) Nicking the desired DNA sequence at the PAM strand by the fusion protein, (b) the exposed 3′ hydroxyl group primes the reverse transcription (RT) of the RT template of the prime editing gRNA (pegRNA), (c) reverse transcription, (d) the branched intermediate form containing two flaps of DNA: a 3′ flap (containing the edited sequence), and a 5′ flap (containing the dispensable, unedited DNA sequence) followed by flap cleavage, and (e) ligation and mismatch repair; either incorporating the edited strand or removing it. Full article ">Figure 4
T-DNA vector with all the components necessary for Cas9-induced mutagenesis. The 20 bp protospacer sequences of each target site are subcloned or integrated between the sgRNA scaffold and the U3 promoter by ligation of primers into an AarI-digested SK-sgRNA vector. Then, this vector is again re-ligated with a pC1300-Cas9 vector by ligation of BamHI and KnpI-digested enzymes. The whole sgRNA cassette is then delivered into a pC1300-Cas9 vector (contains the Cas9 gene under the control of the 2 × 35S promoter) for plant transformation. Full article ">

13 pages, 1486 KiB

Open AccessReview

Revisiting the Role of Astrocytic MAOB in Parkinson’s Disease

by Min-Ho Nam, Moonsun Sa, Yeon Ha Ju, Mingu Gordon Park and C. Justin Lee

Int. J. Mol. Sci. 2022, 23(8), 4453; https://doi.org/10.3390/ijms23084453 - 18 Apr 2022

Cited by 31 | Viewed by 7005

Abstract

Monoamine oxidase-B (MAOB) has been believed to mediate the degradation of monoamine neurotransmitters such as dopamine. However, this traditional belief has been challenged by demonstrating that it is not MAOB but MAOA which mediates dopamine degradation. Instead, MAOB mediates the aberrant synthesis of [...] Read more.

Monoamine oxidase-B (MAOB) has been believed to mediate the degradation of monoamine neurotransmitters such as dopamine. However, this traditional belief has been challenged by demonstrating that it is not MAOB but MAOA which mediates dopamine degradation. Instead, MAOB mediates the aberrant synthesis of GABA and hydrogen peroxide (H₂O₂) in reactive astrocytes of Parkinson’s disease (PD). Astrocytic GABA tonically suppresses the dopaminergic neuronal activity, whereas H₂O₂ aggravates astrocytic reactivity and dopaminergic neuronal death. Recently discovered reversible MAOB inhibitors reduce reactive astrogliosis and restore dopaminergic neuronal activity to alleviate PD symptoms in rodents. In this perspective, we redefine the role of MAOB for the aberrant suppression and deterioration of dopaminergic neurons through excessive GABA and H₂O₂ synthesis of reactive astrocytes in PD. Full article

(This article belongs to the Special Issue Molecular Mechanism in Movement Disorders and Neurotransmitter Diseases)

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The traditional view on MAOB as a DA- and MPTP-metabolizing enzyme. MAOA and MAOB have been traditionally believed to be responsible for dopamine (DA) metabolism. DA taken up via DA transporter (DAT) is known to be metabolized into homovanillic acid (HVA) through two different pathways. First, MAO converts DA to 3,4-dihydroxyphenylacetaldehyde (DOPAL), which is then converted to 3,4-dihydroxyphenylacetic acid (DOPAC) by aldehyde dehydrogenases (ALDH). DOPAC is finally converted to HVA through catechol-o-methyltransferase (COMT). Second, COMT converts DA to 3-methoxytyramine (3-MT), which is then converted to 3-methoxy-4-hydroxyphenylacetaldehyde (MHPA) by MAO. Then, MHPA is finally converted to HVA by ALDH. Due to the presence of these enzymes in both DAergic presynaptic terminals and astrocytes, it has been assumed that DA is degraded by MAOA and MAOB in both cell types. Full article ">Figure 2
The recent findings on MAOB as a GABA- and H2O2-synthesizing enzyme. According to recent discoveries, it is not MAOB but MAOA that is responsible for dopamine (DA) metabolism to HVA. Instead, MAOB is responsible for astrocytic GABA and H2O2 synthesis. In detail, under the PD pathogenesis, the putrescine level in reactive astrocytes is increased, which could be linked to the accumulation of misfolded α-synuclein. Putrescine is converted to N-acetylputrescine by putrescine aminotransferase (PAT). MAOB converts the N-acetylputrescine to N-acetyl-γ-aminobutyraldehyde and produces H2O2 as a byproduct. N-acetyl-γ-aminobutyraldehyde is sequentially converted to N-acetyl-γ-aminobutyrate and GABA. GABA, released via Bestrophin 1 (BEST1) from astrocytes, inhibits the excitability of neighboring DAergic neurons. On the other hand, H2O2 exacerbates DAergic neuronal degeneration. The aberrant suppression and deterioration of DAergic neurons lead to reduced expression of tyrosine hydroxylase (TH) and DA deficiency. Full article ">

23 pages, 1954 KiB

Open AccessReview

Bio-Synthesized Nanoparticles in Developing Plant Abiotic Stress Resilience: A New Boon for Sustainable Approach

by Sarika Kumari, Risheek Rahul Khanna, Faroza Nazir, Mohammed Albaqami, Himanshu Chhillar, Iram Wahid and M. Iqbal R. Khan

Int. J. Mol. Sci. 2022, 23(8), 4452; https://doi.org/10.3390/ijms23084452 - 18 Apr 2022

Cited by 49 | Viewed by 5389

Abstract

Agriculture crop development and production may be hampered in the modern era because of the increasing prevalence of ecological problems around the world. In the last few centuries, plant and agrarian scientific experts have shown significant progress in promoting efficient and eco-friendly approaches [...] Read more.

Agriculture crop development and production may be hampered in the modern era because of the increasing prevalence of ecological problems around the world. In the last few centuries, plant and agrarian scientific experts have shown significant progress in promoting efficient and eco-friendly approaches for the green synthesis of nanoparticles (NPs), which are noteworthy due to their unique physio-biochemical features as well as their possible role and applications. They are thought to be powerful sensing molecules that regulate a wide range of significant physiological and biochemical processes in plants, from germination to senescence, as well as unique strategies for coping with changing environmental circumstances. This review highlights current knowledge on the plant extract-mediated synthesis of NPs, as well as their significance in reprogramming plant traits and ameliorating abiotic stresses. Nano particles-mediated modulation of phytohormone content in response to abiotic stress is also displayed. Additionally, the applications and limitations of green synthesized NPs in various scientific regimes have also been highlighted. Full article

(This article belongs to the Special Issue Nanoparticles: From Synthesis to Applications)

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A summary of physiological and biochemical responses in plants on exogenous application of green synthesized nanoparticles by foliar application or direct administration in the soil. Full article ">Figure 2
Schematic diagram showing the preparation of green synthesized nanoparticles from different parts of the plant. Plant extract preparation from different plant parts such as roots, leaves, flower, fruits and seeds are used in the synthesis of nanoparticles (NPs). The bio-reduction mediated synthesis of NPs is controlled by several factors including concentration of plant extracts and metal ions, pH of the solution, reaction time, and temperature at which reaction is carried out. Purifications and characterization of NPs play a determinant role in the synthesis of desired NPs, which could be beneficial in plant science and research-oriented disciplines. The reaction should be restarted from the bio-reduction process if the synthesized NPs do not meet the desired morphological characteristics. Black, red and dotted arrows show the steps involved in NPs synthesis. Full article ">Figure 3
Summarization of cellular parameters regulated by the application of green synthesized nanoparticles to plants under different abiotic stresses. The presented NPs are a generalization of various green synthesized NPs administered in abiotic stressed plants, and reported to impart tolerance traits by regulating the various cellular/physiological aspects. Membrane transporters bring about nutrient uptake and efflux, the efficiency of which is hampered by various abiotic stress conditions but is restored and maintained by green synthesized NPs’ supplementation. Enzymatic antioxidants up-regulated by the action of green synthesized NPs aid in ameliorating abiotic stress-induced oxidative damages and maintain cellular homeostasis. Photosynthetic efficiency, nitrogen metabolism and osmolyte concentrations were enhanced upon NPs’ application, and thus play a crucial role in imparting abiotic stress tolerance. Genes and proteins represented in green are up-regulated, while those represented in red are down-regulated. Black arrow lines represent effects imposed by NPs and/or a further step in a natural series of cellular events. Red line with a flat head represents the repression effect. APX, ascorbate peroxidase gene; Ca2+, calcium ions; CAT, catalase gene; Cd2+, cadmium ions; Fe2+, ferrous ions; GK, glutamyl kinase; GPX, glutathione peroxidase gene; K+; potassium ions; Mn2+, manganese ions; Na+, sodium ions; NiR, nitrite reductase; NR, nitrate reductase; NPs, nanoparticles; POX, proline oxidase; ROS, reactive oxygen species; SOD, superoxide dismutase gene. Full article ">Figure 4
Potential targets for nanoparticles-mediated abiotic stress mitigation. The synthesis of nanoparticles (NPs) by green approaches is an environment-friendly method and can be efficiently used to trigger various abiotic stress-induced responses upon exogenous application to stressed plants. Chemically synthesized NPs show nanotoxicity and limit the ameliorative responses for abiotic stress alleviation (indicated by the red inhibitory arrow). Strategies focused on eliciting plant defense responses by triggering osmotic adjustment, antioxidant machinery for amelioration of cellular damage by ROS, and improving protein activity, can be efficient in conferring resistance to various abiotic stresses. Green synthesized NPs prove to be a suitable candidate for stress mitigation as they help in altering the gene expression of enzymatic antioxidants (SOD, CAT, and GR), and enhance osmotic content and nutrient homeostasis. Furthermore, green synthesized NPs favor the enhancement in protein activity, particularly of enzymes involved in N and proline metabolism which help in accomplishing the abiotic stress tolerance in crops. CAT, catalase; GR, glutathione reductase; N, nitrogen; ROS, reactive oxygen species; SOD, superoxide dismutase. Full article ">Figure 5
Applications of green synthesized nanoparticles in different fields. Black straight line represents the different applications of plant-synthesized nanoparticles in various fields of plant science and research-oriented disciplines. Full article ">

22 pages, 8971 KiB

Open AccessArticle

E2F1 Expression and Apoptosis Initiation in Crayfish and Rat Peripheral Neurons and Glial Cells after Axonal Injury

by Valentina Dzreyan, Moez Eid, Stanislav Rodkin, Maria Pitinova and Svetlana Demyanenko

Int. J. Mol. Sci. 2022, 23(8), 4451; https://doi.org/10.3390/ijms23084451 - 18 Apr 2022

Cited by 7 | Viewed by 3029

Abstract

Neurotrauma is among the main causes of human disability and mortality. The transcription factor E2F1 is one of the key proteins that determine the fate of cells. The involvement of E2F1 in the regulation of survival and death of peripheral nerve cells after [...] Read more.

Neurotrauma is among the main causes of human disability and mortality. The transcription factor E2F1 is one of the key proteins that determine the fate of cells. The involvement of E2F1 in the regulation of survival and death of peripheral nerve cells after axotomy has not been previously studied. We, for the first time, studied axotomy-induced changes in the expression and localization of E2F1 following axonal injury in rats and crayfish. Immunoblotting and immunofluorescence microscopy were used for the analysis of the expression and intracellular localization of E2F1 and its changes after axotomy. To evaluate whether this transcription factor promotes cell apoptosis, we examined the effect of pharmacological inhibition of E2F activity in axotomized rat models. In this work, axotomy caused increased expression of E2F1 as early as 4 h and even 1 h after axotomy of mechanoreceptor neurons and ganglia of crayfish ventral nerve cord (VNC), as well as rat dorsal root ganglia (DRG). The level of E2F1 expression increased both in the cytoplasm and the nuclei of neurons. Pharmacological inhibition of E2F demonstrated a pronounced neuroprotective activity against axotomized DRGs. E2F1 and downstream targets could be considered promising molecular targets for the development of potential neuroprotective agents. Full article

(This article belongs to the Special Issue Cell Apoptosis)

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16 pages, 2380 KiB

Open AccessArticle

Arabidopsis KCS5 and KCS6 Play Redundant Roles in Wax Synthesis

by Haodong Huang, Asma Ayaz, Minglü Zheng, Xianpeng Yang, Wajid Zaman, Huayan Zhao and Shiyou Lü

Int. J. Mol. Sci. 2022, 23(8), 4450; https://doi.org/10.3390/ijms23084450 - 18 Apr 2022

Cited by 34 | Viewed by 4274

Abstract

3-ketoacyl-CoA synthases (KCSs), as components of a fatty acid elongase (FAE) complex, play key roles in determining the chain length of very-long-chain fatty acids (VLCFAs). KCS6, taking a predominate role during the elongation from C26 to C28, is well known to play an [...] Read more.

3-ketoacyl-CoA synthases (KCSs), as components of a fatty acid elongase (FAE) complex, play key roles in determining the chain length of very-long-chain fatty acids (VLCFAs). KCS6, taking a predominate role during the elongation from C26 to C28, is well known to play an important role in wax synthesis. KCS5 is one paralog of KCS6 and its role in wax synthesis remains unknown. Wax phenotype analysis showed that in kcs5 mutants, the total amounts of wax components derived from carbon 32 (C32) and C34 were apparently decreased in leaves, and those of C26 to C32 derivatives were obviously decreased in flowers. Heterologous yeast expression analysis showed that KCS5 alone displayed specificity towards C24 to C28 acids, and its coordination with CER2 and CER26 catalyzed the elongation of acids exceeding C28, especially displaying higher activity towards C28 acids than KCS6. BiLC experiments identified that KCS5 physically interacts with CER2 and CER26. Wax phenotype analysis of different organs in kcs5 and kcs6 single or double mutants showed that KCS6 mutation causes greater effects on the wax synthesis than KCS5 mutation in the tested organs, and simultaneous repression of both protein activities caused additive effects, suggesting that during the wax biosynthesis process, KCS5 and KCS6 play redundant roles, among which KCS6 plays a major role. In addition, simultaneous mutations of two genes nearly block drought-induced wax production, indicating that the reactions catalyzed by KCS5 and KCS6 play a critical role in the wax biosynthesis in response to drought. Full article

(This article belongs to the Collection Feature Papers in Molecular Plant Sciences)

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Wax profile in rosette leaves and flowers of kcs5-1, kcs5-2 mutants, and Col-0. (A,D) Amount of each wax component in rosette leaves and flowers; wax coverage is expressed as wax amounts per stem surface area (μg.dm−2). Each wax constituent is designated by carbon chain length and labeled by chemical class along the x axis. (B,E) Amount of all wax components with equal carbon chain lengths in rosette leaves and flowers. (C,F) Total wax amounts. Data are means ± SE of five biological replicates. (* p <0.01; ** p < 0.05; *** p < 0.001, **** p < 0.0001). Full article ">Figure 2
The catalytic activity of KCS5 and KCS6 examined by the yeast system. KCS5, KCS6, or empty vector (EV) were alone expressed or co-expressed with CER2 and CER26 in yeast strain BY4741 Δelo3Δfah1, and the generated products are displayed in the form of VLCFA-FAMEs (A–D). FAMEs were synthesized by transmethylation before GC analysis. The values shown are mean ± SD (n = 4). * p < 0.05; ** p < 0.01; *** p < 0.001. Full article ">Figure 3
Interactions between KCS5 and CER2 or CER26 by BiLC experiments. Agrobacterium strain GV3101 containing different combinations (A–D) was transiently infiltrated into tobacco leaves. After 2–3 days, luminescence signals were captured by Tanon 5200 luminescence imaging system (Tianneng Technology Co., Ltd., Shanghai, China). The colors labeled beside the figures indicate signal intensity. Full article ">Figure 4
Wax profile in different organs of different plants. Rosette leaves (A), stems (B), and flowers (C) of Col-0, kcs5-1, kcs6-1, and kcs5-1 kcs6-1 were collected for wax analysis. Wax coverage is expressed as wax amounts per leaf surface area (μg.dm−2). Each wax constituent was designated by carbon chain length and was labeled by chemical class along the x-axis. The values shown are means ± SD (n = 4). * p < 0.05; ** p < 0.01. Full article ">Figure 5
Water loss assay of detached leaves. Excised leaves’ water loss rates were recorded over 120 min and measured as a percentage of the initial weight of fully hydrated leaves. Values are the mean of five replicate assays. Error bar = SD. The experiments were repeated once with similar results. Full article ">Figure 6
Wax production of different plants in response to water deficit conditions. Wax analysis was performed in different plants grown under normal and water deficit conditions (A). Total wax amounts of plants grown under different conditions are shown (B). The values shown are means ± SD (n = 4). The experiments were repeated once with similar results. * p < 0.05; **** p < 0.0001. Full article ">

18 pages, 4623 KiB

Open AccessArticle

The Ameliorative Effect of Dexamethasone on the Development of Autoimmune Lung Injury and Mediastinal Fat-Associated Lymphoid Clusters in an Autoimmune Disease Mouse Model

by Yaser Hosny Ali Elewa, Md Abdul Masum, Sherif Kh. A. Mohamed, Md Rashedul Islam, Teppei Nakamura, Osamu Ichii and Yasuhiro Kon

Int. J. Mol. Sci. 2022, 23(8), 4449; https://doi.org/10.3390/ijms23084449 - 18 Apr 2022

Cited by 2 | Viewed by 2707

Abstract

In our previous study, we revealed the ameliorative therapeutic effect of dexamethasone (Dex) for Lupus nephritis lesions in the MRL/MpJ-Fas ^lpr/lpr (Lpr) mouse model. The female Lpr mice developed a greater number of mediastinal fat-associated lymphoid clusters (MFALCs) and inflammatory lung lesions [...] Read more.

In our previous study, we revealed the ameliorative therapeutic effect of dexamethasone (Dex) for Lupus nephritis lesions in the MRL/MpJ-Fas ^lpr/lpr (Lpr) mouse model. The female Lpr mice developed a greater number of mediastinal fat-associated lymphoid clusters (MFALCs) and inflammatory lung lesions compared to the male mice. However, the effect of Dex, an immunosuppressive drug, on both lung lesions and the development of MFALCs in Lpr mice has not been identified yet. Therefore, in this study, we compared the development of lung lesions and MFALCs in female Lpr mice that received either saline (saline group “SG”) or dexamethasone (dexamethasone group “DG”) in drinking water as a daily dose along with weekly intraperitoneal injections for 10 weeks. Compared to the SG group, the DG group showed a significant reduction in the levels of serum anti-dsDNA antibodies, the size of MFALCs, the degree of lung injury, the area of high endothelial venules (HEVs), and the number of proliferating and immune cells in both MFALCs and the lungs. A significant positive correlation was observed between the size of MFALCs and the cellular aggregation in the lungs of Lpr mice. Therefore, this study confirmed the ameliorative effect of Dex on the development of lung injury and MFALCs via their regressive effect on both immune cells’ proliferative activity and the development of HEVs. Furthermore, the reprogramming of MFALCs by targeting immune cells and HEVs may provide a therapeutic strategy for autoimmune-disease-associated lung injury. Full article

(This article belongs to the Special Issue Human and Animal Monocytes and Macrophages in Homeostasis and Disease 3.0)

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14 pages, 1450 KiB

Open AccessPerspective

Small RNAs beyond Model Organisms: Have We Only Scratched the Surface?

by Emilie Boutet, Samia Djerroud and Jonathan Perreault

Int. J. Mol. Sci. 2022, 23(8), 4448; https://doi.org/10.3390/ijms23084448 - 18 Apr 2022

Cited by 6 | Viewed by 3310

Abstract

Small RNAs (sRNAs) are essential regulators in the adaptation of bacteria to environmental changes and act by binding targeted mRNAs through base complementarity. Approximately 550 distinct families of sRNAs have been identified since their initial characterization in the 1980s, accelerated by the emergence [...] Read more.

Small RNAs (sRNAs) are essential regulators in the adaptation of bacteria to environmental changes and act by binding targeted mRNAs through base complementarity. Approximately 550 distinct families of sRNAs have been identified since their initial characterization in the 1980s, accelerated by the emergence of RNA-sequencing. Small RNAs are found in a wide range of bacterial phyla, but they are more prominent in highly researched model organisms compared to the rest of the sequenced bacteria. Indeed, Escherichia coli and Salmonella enterica contain the highest number of sRNAs, with 98 and 118, respectively, with Enterobacteriaceae encoding 145 distinct sRNAs, while other bacteria families have only seven sRNAs on average. Although the past years brought major advances in research on sRNAs, we have perhaps only scratched the surface, even more so considering RNA annotations trail behind gene annotations. A distinctive trend can be observed for genes, whereby their number increases with genome size, but this is not observable for RNAs, although they would be expected to follow the same trend. In this perspective, we aimed at establishing a more accurate representation of the occurrence of sRNAs in bacteria, emphasizing the potential for novel sRNA discoveries. Full article

(This article belongs to the Special Issue Bacterial Non-coding RNA)

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Figure 1
Number of distinct annotated sRNAs per bacterial strain in Proteobacteria. The iceberg is intended to be a graphical representation of the knowledge we have about the prevalence of sRNAs in Proteobacteria (gray section) as opposed to what we could be missing (hatched section). The ratio of the surface versus underwater portions of the iceberg is proportional to results represented in the graph, where the gray region is what is known (i.e., the visible part of the iceberg), and the hatched area under that region is what could be left to discover (that is, the underwater section of the iceberg). Percentages also represent this ratio. This figure represents a compilation of 2629 strains. Only sRNAs with an E-value lower than 0.0005 were considered. Full article ">Figure 2
Top 20 bacterial species with the highest number of distinct annotated sRNAs in (A) Proteobacteria and in (B) bacteria from the Terrabacteria group. Species denoted with “sp.” represent instances where only the genus of the bacteria was noted. It can be observed that in (A), all species are from the same family, Enterobacteriaceae. In (B), species from different orders are emphasized by their own color. The number of distinct sRNAs considers all strains for each species. Only sRNAs with an E-value lower than 0.0005 were considered. Full article ">Figure 3
Top 20 sRNAs annotated in bacteria. Each individual occurrence of sRNAs were counted, even if some were found multiple times within the same genome. Only sRNAs with an E-value lower than 0.0005 were taken into consideration. Full article ">Figure 4
Number of annotated genes and RNAs in bacteria. Data required for the creation of this graph were taken from RiboGap [<a href="#B15-ijms-23-04448" class="html-bibr">15</a>]. (A) The number of annotated genes is graphed according to the genome size, which comprises all chromosomes and plasmids of each individual strain if applicable. (B) The number of annotated RNAs is graphed according to the “fragment size”, which considers chromosomes and plasmids separately for each individual strain. RNAs are not limited only to sRNAs but also include CRISPR RNAs, antisense RNAs, sRNAs, long non-coding RNAs (lncRNAs), rRNAs, ribozymes, tRNAs and cis-regulatory elements. Species from Terrabacteria group and Proteobacteria that were found to have the most annotated sRNAs (<a href="#ijms-23-04448-f002" class="html-fig">Figure 2</a>) are represented by black and blue dots, respectively; all other strains are shown in gray. Full article ">

12 pages, 2345 KiB

Open AccessArticle

Thrombin Preconditioning Improves the Therapeutic Efficacy of Mesenchymal Stem Cells in Severe Intraventricular Hemorrhage Induced Neonatal Rats

by So Yeon Jung, Young Eun Kim, Won Soon Park, So Yoon Ahn, Dong Kyung Sung, Se In Sung, Kyeung Min Joo, Seong Gi Kim and Yun Sil Chang

Int. J. Mol. Sci. 2022, 23(8), 4447; https://doi.org/10.3390/ijms23084447 - 18 Apr 2022

Cited by 15 | Viewed by 2948

Abstract

Severe intraventricular hemorrhage (IVH) remains a major cause of high mortality and morbidity in extremely preterm infants. Mesenchymal stem cell (MSC) transplantation is a possible therapeutic option, and development of therapeutics with enhanced efficacy is necessary. This study investigated whether thrombin preconditioning improves [...] Read more.

Severe intraventricular hemorrhage (IVH) remains a major cause of high mortality and morbidity in extremely preterm infants. Mesenchymal stem cell (MSC) transplantation is a possible therapeutic option, and development of therapeutics with enhanced efficacy is necessary. This study investigated whether thrombin preconditioning improves the therapeutic efficacy of human Wharton’s jelly-derived MSC transplantation for severe neonatal IVH, using a rat model. Severe neonatal IVH was induced by injecting 150 μL blood into each lateral ventricle on postnatal day (P) 4 in Sprague-Dawley rats. After 2 days (P6), naïve MSCs or thrombin-preconditioned MSCs (1 × 10⁵/10 μL) were transplanted intraventricularly. After behavioral tests, brain tissues and cerebrospinal fluid of P35 rats were obtained for histological and biochemical analyses, respectively. Thrombin-preconditioned MSC transplantation significantly reduced IVH-induced ventricular dilatation on in vivo magnetic resonance imaging, which was coincident with attenuations of reactive gliosis, cell death, and the number of activated microglia and levels of inflammatory cytokines after IVH induction, compared to naïve MSC transplantation. In the behavioral tests, the sensorimotor and memory functions significantly improved after transplantation of thrombin-preconditioned MSCs, compared to naïve MSCs. Overall, thrombin preconditioning significantly improves the therapeutic potential and more effectively attenuates brain injury, including progressive ventricular dilatation, gliosis, cell death, inflammation, and neurobehavioral functional impairment, in newborn rats with induced severe IVH than does naïve MSC transplantation. Full article

(This article belongs to the Special Issue New Sources, Differentiation, and Therapeutic Uses of Mesenchymal Stem Cells 2.0)

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13 pages, 6982 KiB

Open AccessArticle

Leaf Mutant 7 Encoding Heat Shock Protein OsHSP40 Regulates Leaf Size in Rice

by Fuhua Wang, Zhengbin Tang, Ya Wang, Jing Fu, Wenbo Yang, Shengxuan Wang, Yuetao Wang, Tao Bai, Zhibo Huang, Haiqing Yin and Zhoufei Wang

Int. J. Mol. Sci. 2022, 23(8), 4446; https://doi.org/10.3390/ijms23084446 - 18 Apr 2022

Cited by 9 | Viewed by 3024

Abstract

Leaf size is an important agronomic trait directly affecting yield in rice, and thus understanding the genes determining leaf size is important in breeding. In this study, one Leaf Mutant 7 (lm7) with small leaf size was isolated using ethyl methane [...] Read more.

Leaf size is an important agronomic trait directly affecting yield in rice, and thus understanding the genes determining leaf size is important in breeding. In this study, one Leaf Mutant 7 (lm7) with small leaf size was isolated using ethyl methane sulphonate (EMS) mutagenesis from the japonica Zhenggeng 1925. MutMap by whole genome resequencing of phenotypic bulks revealed that LM7 is likely located in the 133 kb region on chromosome 7 using F₂ population from a cross between lm7 and wild-type (WT) Zhenggeng 1925. The candidate gene encoding heat shock protein OsHSP40 for LM7 was functionally validated. Disruption of this gene in Oshsp40 mutants significantly reduced the leaf size compared with that of WT in rice. Microscopic examination showed that OsHSP40 modulated leaf size via regulating the veins formation and cell size/cell number. Nucleotide diversity analysis indicated that a single nucleotide polymorphism (SNP) variation of C to T in the coding region of OsHSP40 may cause small leaves among rice accessions. Therefore, the natural variation of OsHSP40 contributing to leaf size might be useful for rice breeding. Full article

(This article belongs to the Section Molecular Plant Sciences)

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Figure 1
Phenotype characterization of lm7. (a) Representative image of mature plant of wild-type (WT) Zhenggeng 1925 and lm7. Scale bar is 20 cm. Representative image of the (b) top, (c) second, and (d) third leaf in WT and lm7. Scale bar is 5 cm. Representative image of the (e) panicle, (f) grain length, and (g) grain width. Scale bar is 1 cm. The mean value of (h) plant height, (i) leaf width, (j) leaf length, (k) panicle length, (l) grain length, (m) grain width, and (n) 1000-grain weight. Significant differences compared with the WT were determined using Student’s t-test: ** p < 0.01. Full article ">Figure 2
Leaf tissue and cytological observation of lm7. Representative image of epidermal cells of (a) wild-type (WT) Zhenggeng 1925 and (b) lm7 leaves. Scale bar is 50 μm. Cross section of (c) WT and (d) lm7 leaves. Asterisks represent large veins and solid circles represent small veins. Scale bar is 1 mm. A clear show of epidermal cells of (e) WT and (f) lm7 leaves. Scale bars = 50 μm. The mean value of (g) large veins number, (h) small veins number, (i) epidermal cell width, (j) epidermal cell length, and (k) epidermal cell number along leaf-width axis. Significant differences compared with the WT were determined using Student’s t-test: ** p < 0.01. n.s. means not significant. Full article ">Figure 3
Mapping and cloning of LM7 by bulked segregant analysis using sequencing. (a) Distribution of Δ index of single nucleotide polymorphism (SNP) across 12 chromosomes. Δ (SNP-index) means the absolute value of the difference of SNP index between the bulked pool and Zhenggeng 1925. (b) SNP screening of 99 F2 individuals originating from lw7 × Zhenggeng 1925 narrowed down the location of the LW7 locus to a 133 kb region bounded by markers SNP3036 and SNP36 on chromosome 7. Numbers below the chromosome indicate the physical position of markers. White and gray indicates the lw7 and Zhenggeng 1925 background respectively. (c) Physical position of the LW7 locus. The arrows represent 13 annotated ORFs in the 133 kb fine-mapping interval according to the rice MSU 7 reference genome. LOC_Os07g09450 in red is the candidate gene for LW7. (d) CDS structure of candidate gene LW7/OsHSP40 (LOC_Os07g09450) and mutation site. Sequence analysis revealed a C-to-T nucleotide mutation, which results in forming stop codon in lw7. Full article ">Figure 4
Function verification of the candidate gene OsHSP40 for LM7. (a) Oshsp40-1 and Oshsp40-2 mutants generated in wild-type (WT) Zhenggeng 1925 by CRISPR/Cas9 approach. A total of 66 and 2 bp nucleotides were deleted in Oshsp40-1 and Oshsp40-2 respectively. The two sgRNA:Cas9 target sites are labelled in red lines; gray rectangles indicate exons. (b) Representative image of mature plant of WT and Oshsp40 mutants. Scale bar is 20 cm. (c) Representative image of the top, second, and third leaf in WT and Oshsp40 mutants. Scale bar is 1 cm. The mean value of (d) leaf length and (e) leaf width. Cross section of (f) WT, (g) Oshsp40-1, and (h) Oshsp40-2 leaves. Asterisks represent large veins and solid circles represent small veins. Scale bar is 1 mm. A clear show of epidermal cells of (i) WT, (j) Oshsp40-1, and (k) Oshsp40-2 leaves. Scale bars = 50 μm. The mean value of (l) large veins number, (m) small veins number, (n) epidermal cell width, (o) epidermal cell length, and (p) epidermal cell number along leaf-width axis. Significant differences compared with the WT were determined using Student’s t-test: ** p < 0.01. n.s. means not significant. Full article ">Figure 5
Characterization of OsHSP40 and its expression pattern. (a) Protein sequence alignment of OsHSP40 in different plants. Full-length of amino acid sequences from NCBI were used for analyses. (b) Phylogenetic tree showing the relationship between OsHSP40 homologs in monocots (Group A) and dicots (Group B). (c) Relative expression of OsHSP40 in various rice tissues determined by quantitative RT-PCR in wild type Zhenggeng 1925. R, roots; LS, leaf sheath; ML, mature leaves; P, panicles (10–15 cm); S, seeds (11–20 days after pollination); N, nodes; IN, internodes. Expression is relative to that in the root, the value of which was set as 1. OsActin gene was used as the internal control. (d) Subcellular localization of OsHSP40 tagged at the C-terminus with GFP in rice protoplasts. Full article ">Figure 6
Functional SNP and nucleotide diversity analysis of OsHSP40. (a) The position of functional SNP in the CDS region of OsHSP40. (b) Comparisons of phenotype in the accessions with different functional SNPs of OsHSP40. FLW, flag leaf width; PH, plant height; PL, panicle length; GL, grain length; GW, grain width; TGW, 1000-grain weight. (c) Nucleotide diversity of OsHSP40 in japonica, indica, and wild rice. Red box denotes the position of OsHSP40. (d) Average nucleotide diversity of the 20 kb region surrounding OsHSP40. The different letters indicate the significant differences determined using ANOVA test: p < 0.05. Full article ">

4 pages, 222 KiB

Open AccessComment

Comment on Tremmel et al. In Vitro Metabolism of Six C-Glycosidic Flavonoids from Passiflora incarnata L. Int. J. Mol. Sci. 2021, 22, 6566

by Monika Beszterda and Rafał Frański

Int. J. Mol. Sci. 2022, 23(8), 4445; https://doi.org/10.3390/ijms23084445 - 18 Apr 2022

Viewed by 2717

Abstract

In recent years, growing attention has been paid to the chemical composition of aerial parts extracts and the bioavailability of active compounds from different species of Passiflora genus [...] Full article

(This article belongs to the Section Molecular Plant Sciences)

17 pages, 3595 KiB

Open AccessArticle

HPR1 Is Required for High Light Intensity Induced Photorespiration in Arabidopsis thaliana

by Zi Wang, Yetao Wang, Yukun Wang, Haotian Li, Zhiting Wen and Xin Hou

Int. J. Mol. Sci. 2022, 23(8), 4444; https://doi.org/10.3390/ijms23084444 - 18 Apr 2022

Cited by 13 | Viewed by 2999

Abstract

High light intensity as one of the stresses could lead to generation of large amounts of reactive oxygen species (ROS) in plants, resulting in severe plant growth retardation. The photorespiration metabolism plays an important role in producing and removing a variety of ROS, [...] Read more.

High light intensity as one of the stresses could lead to generation of large amounts of reactive oxygen species (ROS) in plants, resulting in severe plant growth retardation. The photorespiration metabolism plays an important role in producing and removing a variety of ROS, maintaining the dynamic balance of the redox reaction, and preventing photoinhibition. Arabidopsis hydroxypyruvate reductase 1 (HPR1) is a primary metabolic enzyme in the photorespiration cycle. However, the role of HPR1 in plants response to high light is not clear. Here, we found that the expression of HPR1 could be induced by high light intensity. The growth and photosynthetic capacity of hpr1 mutants are seriously affected under high light intensity. The absence of HPR1 suppresses the rates of photorepair of Photosystem II (PSII), aggravates the production of ROS, and accelerates photorespiration rates. Moreover, the activity of ROS scavenging enzymes in the hpr1 mutants is significantly higher. These results indicate that HPR1 is involved in plant response to high light intensity and is essential for maintaining the dynamic balance of ROS and photorespiration. Full article

(This article belongs to the Special Issue Light Reactions and Oxidative Stress in Photosynthesis)

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15 pages, 2841 KiB

Open AccessArticle

A Non-Hazardous Deparaffinization Protocol Enables Quantitative Proteomics of Core Needle Biopsy-Sized Formalin-Fixed and Paraffin-Embedded (FFPE) Tissue Specimens

by Georgia Mitsa, Qianyu Guo, Christophe Goncalves, Samuel E. J. Preston, Vincent Lacasse, Adriana Aguilar-Mahecha, Naciba Benlimame, Mark Basik, Alan Spatz, Gerald Batist, Wilson H. Miller, Jr., Sonia V. del Rincon, René P. Zahedi and Christoph H. Borchers

Int. J. Mol. Sci. 2022, 23(8), 4443; https://doi.org/10.3390/ijms23084443 - 18 Apr 2022

Cited by 9 | Viewed by 7296

Abstract

Most human tumor tissues that are obtained for pathology and diagnostic purposes are formalin-fixed and paraffin-embedded (FFPE). To perform quantitative proteomics of FFPE samples, paraffin has to be removed and formalin-induced crosslinks have to be reversed prior to proteolytic digestion. A central component [...] Read more.

Most human tumor tissues that are obtained for pathology and diagnostic purposes are formalin-fixed and paraffin-embedded (FFPE). To perform quantitative proteomics of FFPE samples, paraffin has to be removed and formalin-induced crosslinks have to be reversed prior to proteolytic digestion. A central component of almost all deparaffinization protocols is xylene, a toxic and highly flammable solvent that has been reported to negatively affect protein extraction and quantitative proteome analysis. Here, we present a ‘green’ xylene-free protocol for accelerated sample preparation of FFPE tissues based on paraffin-removal with hot water. Combined with tissue homogenization using disposable micropestles and a modified protein aggregation capture (PAC) digestion protocol, our workflow enables streamlined and reproducible quantitative proteomic profiling of FFPE tissue. Label-free quantitation of FFPE cores from human ductal breast carcinoma in situ (DCIS) xenografts with a volume of only 0.79 mm³ showed a high correlation between replicates (r² = 0.992) with a median %CV of 16.9%. Importantly, this small volume is already compatible with tissue micro array (TMA) cores and core needle biopsies, while our results and its ease-of-use indicate that further downsizing is feasible. Finally, our FFPE workflow does not require costly equipment and can be established in every standard clinical laboratory. Full article

(This article belongs to the Special Issue New Insights on Mass Spectometry Applied to Bioscience)

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Experimental Design. Xenografts were generated from human DCIS cells and tumors were resected after 1.5 weeks, followed by formalin-fixation and paraffin-embedding, as described in [<a href="#B17-ijms-23-04443" class="html-bibr">17</a>]. One-millimeter-diameter FFPE cores were used to optimize individual steps of the FFPE sample preparation: (1) deparaffinization, (2) homogenization, (3) extraction, and (4) digestion. Peptide samples were analyzed by nano-LC-MS/MS label-free quantitation (LFQ) to compare the performance of the evaluated protocols for each step of the sample preparation workflow. Full article ">Figure 2
Water-based deparaffinization is a ‘green’ alternative. (A) Total protein extracted after deparaffinization with either water (depW) or xylene (depX) (n = 5, unpaired t-test, p = 0.54). (B) Intra-method %CVs based on quantified proteins, median %CV are given. (C) Pearson correlation plot based on all proteins quantified by both methods. (D) Volcano plot highlighting proteins significantly enriched by either method (Benjamini–Krieger multiple hypothesis testing, FDR 1%). Cytosolic ribosomal proteins significantly enriched with depX are shown in orange. EIF4E, EGFR, and AKT1S1 are highlighted in dark grey. Full article ">Figure 3
Efficient tissue homogenization using micropestles. (A) Total protein extracted from 1 mm cores with a dry-weight < 1 mg (n = 5; unpaired t-test; p = 0.70). (B) Intra-method %CVs based on all quantified proteins, median %CV are given. (C) Pearson correlation plot based on all quantified proteins. (D) Volcano plot highlighting proteins that were significantly enriched by one method (multiple hypothesis testing using the FDR-based approach by Benjamini–Krieger, FDR 1%). Full article ">Figure 4
An SDC–TCEP-based buffer improves overall protein recovery from FFPE tissues. (A) Total protein extracted from 1 mm cores with dry-weight < 1 mg (n = 5; unpaired t-test; p = 0.0341). (B) Intra-method %CVs based on all quantified proteins, median %CVs are given (unpaired t-test; p < 0.0001). (C) Pearson correlation plot based on all proteins quantified with both methods. (D) Volcano plot highlighting proteins that were significantly enriched by either method (multiple hypothesis testing using FDR-based approach by Benjamini–Krieger, FDR 1%). Full article ">Figure 5
Comparison of PAC and STRAP with FASP. (A) Efficacy of tryptic digestion shown by percentage of missed cleavages. (B) Intra-method %CVs based on all quantified proteins, median %CVs are given. (C) Hierarchical clustering [<a href="#B49-ijms-23-04443" class="html-bibr">49</a>] of all quantified proteins, colors reflect log2-normalized abundances. (D–F) Volcano plots highlighting proteins that were significantly enriched by either method (multiple hypothesis testing using FDR-based approach by Benjamini–Krieger, FDR 1%). Full article ">Figure 6
Representative size of FFPE core used in this study. After deparaffinization, the core volume was approximately 0.4 mm3. Full article ">Figure 7
Representative tubes after deparaffinization. The molten paraffin in the depW approach forms a layer on the surface of the hot water and residual paraffin ‘flakes’ precipitate to the bottom of the tube during centrifugation. The deparaffinized tissue floats in the water and can be easily transferred into a new tube for further sample preparation. Full article ">

22 pages, 1378 KiB

Open AccessReview

Linear and Circular Long Non-Coding RNAs in Acute Lymphoblastic Leukemia: From Pathogenesis to Classification and Treatment

by Yasen Maimaitiyiming, Linyan Ye, Tao Yang, Wenjuan Yu and Hua Naranmandura

Int. J. Mol. Sci. 2022, 23(8), 4442; https://doi.org/10.3390/ijms23084442 - 18 Apr 2022

Cited by 11 | Viewed by 4298

Abstract

The coding regions account for only a small part of the human genome, and the remaining vast majority of the regions generate large amounts of non-coding RNAs. Although non-coding RNAs do not code for any protein, they are suggested to work as either [...] Read more.

The coding regions account for only a small part of the human genome, and the remaining vast majority of the regions generate large amounts of non-coding RNAs. Although non-coding RNAs do not code for any protein, they are suggested to work as either tumor suppressers or oncogenes through modulating the expression of genes and functions of proteins at transcriptional, posttranscriptional and post-translational levels. Acute Lymphoblastic Leukemia (ALL) originates from malignant transformed B/T-precursor-stage lymphoid progenitors in the bone marrow (BM). The pathogenesis of ALL is closely associated with aberrant genetic alterations that block lymphoid differentiation and drive abnormal cell proliferation as well as survival. While treatment of pediatric ALL represents a major success story in chemotherapy-based elimination of a malignancy, adult ALL remains a devastating disease with relatively poor prognosis. Thus, novel aspects in the pathogenesis and progression of ALL, especially in the adult population, need to be further explored. Accumulating evidence indicated that genetic changes alone are rarely sufficient for development of ALL. Recent advances in cytogenic and sequencing technologies revealed epigenetic alterations including that of non-coding RNAs as cooperating events in ALL etiology and progression. While the role of micro RNAs in ALL has been extensively reviewed, less attention, relatively, has been paid to other non-coding RNAs. Herein, we review the involvement of linear and circular long non-coding RNAs in the etiology, maintenance, and progression of ALL, highlighting the contribution of these non-coding RNAs in ALL classification and diagnosis, risk stratification as well as treatment. Full article

(This article belongs to the Special Issue Molecular Research on Acute Lymphoblastic Leukemia 2.0)

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Schematic representation of major non-coding RNA types produced in cells. Size of each ncRNA type and main functions are included in the same panel as well. The figure is created with Biorender (<a href="http://Biorender.com" target="_blank">Biorender.com</a> (accessed on 28 January 2022)). Full article ">Figure 2
Main biological functions of lncRNAs in cells. In general, lncRNAs with multiple exons are exported to the cytoplasm similar to mRNAs, while intronic lncRNAs are retained in the nucleus. Due to the flexible structure and long size, lncRNAs interact with DNA, RNA, and protein, thereby regulating chromatin state (histone modification, DNA methylation), transcription, pre-mRNA stability as well as splicing and processing, nuclear condensate formation (i.e., paraspeckles, nuclear speckles), mRNA stability, translation, and sponging miRNAs as well as orchestrating protein complex formation. The figure is created with Biorender (<a href="http://Biorender.com" target="_blank">Biorender.com</a> (accessed on 28 January 2022)). Full article ">Figure 3
Main biogenesis pathways and functions of circRNAs in cells. (A), Biogenesis of exonic circRNA from back-splicing of pre-mRNA. RNA-binding proteins (RBPs) or orientation-opposite complementary sequences (OOCS) that form loop-like structures facilitate back-splicing and circRNAs formation; these circRNAs are generally exported to the cytoplasm and act as a miRNA sponge, protein complex coordinators as well as small peptide producers to modulate protein function; (B), Biogenesis of lncRNAs from intron lariats. A consensus RNA motif containing a 7-nt GU-rich element near the 5′ splice site and an 11-nt C-rich element near the branch point prevents detaching of the intron lariats and promotes intronic circRNA formation; these circRNAs are retained in the nucleus and mainly regulate Pol II-mediated transcription. The figure is created with Biorender (<a href="http://Biorender.com" target="_blank">Biorender.com</a> (accessed on 28 January 2022)). Full article ">

25 pages, 2347 KiB

Open AccessReview

Circulating Tumor DNA in Precision Oncology and Its Applications in Colorectal Cancer

by Maria F. Arisi, Efrat Dotan and Sandra V. Fernandez

Int. J. Mol. Sci. 2022, 23(8), 4441; https://doi.org/10.3390/ijms23084441 - 18 Apr 2022

Cited by 46 | Viewed by 10302

Abstract

Circulating tumor DNA (ctDNA) is a component of cell-free DNA (cfDNA) that is shed by malignant tumors into the bloodstream and other bodily fluids. ctDNA can comprise up to 10% of a patient’s cfDNA depending on their tumor type and burden. The short [...] Read more.

Circulating tumor DNA (ctDNA) is a component of cell-free DNA (cfDNA) that is shed by malignant tumors into the bloodstream and other bodily fluids. ctDNA can comprise up to 10% of a patient’s cfDNA depending on their tumor type and burden. The short half-life of ctDNA ensures that its detection captures tumor burden in real-time and offers a non-invasive method of repeatedly evaluating the genomic profile of a patient’s tumor. A challenge in ctDNA detection includes clonal hematopoiesis of indeterminate potential (CHIP), which can be distinguished from tumor variants using a paired whole-blood control. Most assays for ctDNA quantification rely on measurements of somatic variant allele frequency (VAF), which is a mutation-dependent method. Patients with certain types of solid tumors, including colorectal cancer (CRC), can have levels of cfDNA 50 times higher than healthy patients. ctDNA undergoes a precipitous drop shortly after tumor resection and therapy, and rising levels can foreshadow radiologic recurrence on the order of months. The amount of tumor bulk required for ctDNA detection is lower than that for computed tomography (CT) scan detection, with ctDNA detection preceding radiologic recurrence in many cases. cfDNA/ctDNA can be used for tumor molecular profiling to identify resistance mutations when tumor biopsy is not available, to detect minimal residual disease (MRD), to monitor therapy response, and for the detection of tumor relapse. Although ctDNA is not yet implemented in clinical practice, studies are ongoing to define the appropriate way to use it as a tool in the clinic. In this review article, we examine the general aspects of ctDNA, its status as a biomarker, and its role in the management of early (II–III) and late (IV; mCRC) stage colorectal cancer (CRC). Full article

(This article belongs to the Special Issue Liquid Biopsies in Oncology)

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ctDNA during cancer progression. Created with BioRender.com. Full article ">Figure 2
Potential applications of ctDNA detection assays in early and late stages of solid tumors. Adapted from Wan et al. [<a href="#B25-ijms-23-04441" class="html-bibr">25</a>]. Created with BioRender.com. Full article ">Figure 3
The number of tracked mutations impacts the ctDNA detection rate. Overview of MRD testing, including why tracking many mutations is important, particularly when there is a low fraction of cancerous cfDNA in the bloodstream. (A) Tumor tissue is available; (B) Tumor tissue is not available. WES, whole-exome sequencing. Created with BioRender.com. Full article ">Figure 4
Kaplan–Meier estimates of recurrence-free interval according to ctDNA status in patients with stage II and III colon cancer after surgery (postoperative). In (A) Recurrence-free survival (RFS) in colorectal stage II patients post-surgery not treated with adjuvant chemotherapy. Patients with ctDNA-positive status postoperative had a markedly reduced RFS compared with those with a ctDNA-negative status. From Tie et al. [<a href="#B21-ijms-23-04441" class="html-bibr">21</a>]; (B) Kaplan–Meier estimates of recurrence-free interval according to ctDNA status in patients with stage III colon cancer after surgery. From Tie et al. [<a href="#B123-ijms-23-04441" class="html-bibr">123</a>]. Full article ">Figure 5
Kaplan–Meier estimates of recurrence-free interval according to ctDNA in colon cancer patients after adjuvant chemotherapy. ctDNA status after adjuvant chemotherapy in patients with colon cancer. (A) Stage II; ctDNA positivity immediately after completion of chemotherapy was associated with poorer RFS. From Tie et al., 2016 [<a href="#B21-ijms-23-04441" class="html-bibr">21</a>]. (B) In Stage III. From Tie et al. [<a href="#B123-ijms-23-04441" class="html-bibr">123</a>]. Full article ">Figure 6
Kaplan–Meier estimates of recurrence-free interval according to ctDNA in stage III colon cancer patients after chemotherapy. (A) Postoperative positive ctDNA; (B) Postoperative negative ctDNA. From Tie et al. [<a href="#B123-ijms-23-04441" class="html-bibr">123</a>]. Full article ">

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