Matrix vesicles (MVs) are nano-sized extracellular vesicles that are anchored in the extracellular matrix (ECM). In addition to playing a role in biomineralization, osteoblast-derived MVs were recently suggested to have regulatory duties.... more
Matrix vesicles (MVs) are nano-sized extracellular vesicles that are anchored in the extracellular matrix (ECM). In addition to playing a role in biomineralization, osteoblast-derived MVs were recently suggested to have regulatory duties. The aims of this study were to: establish the characteristics of osteoblast-derived MVs in the context of extracellular vesicles like exosomes, assess their role in modulating osteoblast differentiation, and examine their mechanism of uptake. MVs were isolated from the ECM of MG-63 human osteoblast-like cell cultures and characterized via enzyme activity, transmission electron microscopy, nanoparticle tracking analysis, western blot, and small RNA sequencing. Osteoblasts were treated with MVs from two different culture conditions (growth media [GM]; osteogenic media [OM]) to evaluate their effects on differentiation and production of inflammatory markers and on macrophage polarization. MV endocytosis was assessed using a lipophilic, fluorescent dye...
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Reduced skeletal loading associated with many conditions, such as neuromuscular injuries, can lead to bone fragility and may threaten the success of implant therapy. Our group has developed a botulinum toxin A (botox) injection model to... more
Reduced skeletal loading associated with many conditions, such as neuromuscular injuries, can lead to bone fragility and may threaten the success of implant therapy. Our group has developed a botulinum toxin A (botox) injection model to imitate disease-reduced skeletal loading and reported that botox dramatically impaired the bone formation and osseointegration of titanium implants. Semaphorin 3A (sema3A) is an osteoprotective factor that increases bone formation and inhibits bone resorption, indicating its potential therapeutic role in improving osseointegration in vivo. We first evaluated the sema3A effect on whole bone morphology following botox injections by delivering sema3A via injection. We then evaluated the sema3A effect on the osseointegration of titanium implants with two different surface topographies by delivering sema3A to cortical bone defect sites prepared for implant insertion and above the implants after insertion using a copper-free click hydrogel that polymerizes...
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Transfection of chondrocytes with microRNA-451(miR-451), present in growth zone cartilage of the growth plate, upregulates production of enzymes association with extracellular matrix degradation. miR-451 is also present in articular... more
Transfection of chondrocytes with microRNA-451(miR-451), present in growth zone cartilage of the growth plate, upregulates production of enzymes association with extracellular matrix degradation. miR-451 is also present in articular cartilage and exacerbates IL-1β effects in articular chondrocytes. Moreover, when osteoarthritis (OA) was induced in Sprague Dawley rats via bilateral anterior cruciate ligament transection (ACLT), miR-451 expression was increased in OA cartilage compared to control, suggesting its inhibition might be used to prevent or treat OA. To examine the prophylactic and therapeutic potential of inhibiting miR-451, we evaluated treatment with miR-451 power inhibitor (451-PI) at the onset of joint trauma and treatment after OA had developed. The prophylactic animal cohort received twice-weekly intra-articular injections of either 451-PI or a negative control (NC-PI) beginning on post-surgical day 3. OA was allowed to develop for 24 days in the therapeutic cohort be...
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Laryngeal squamous cell carcinoma (LSCC) responds to 17β-estradiol via estrogen-receptor (ER, transcribed from ESR1) dependent mechanisms, but is not recognized as a hormonally responsive cancer. 17β-estradiol production by LSCC cell... more
Laryngeal squamous cell carcinoma (LSCC) responds to 17β-estradiol via estrogen-receptor (ER, transcribed from ESR1) dependent mechanisms, but is not recognized as a hormonally responsive cancer. 17β-estradiol production by LSCC cell lines UM-SCC-11A and UM-SCC-12 was examined. Wild type (WT) and ESR1-silenced LSCC cultures and xenografts were examined for 17β-estradiol responsiveness in vivo. 14 LSCC and surrounding epithelial samples at various pathological stages were obtained from patients; ERα and ERβ expression were verified using data from the total cancer genome atlas. UM-SCC-11A and UM-SCC-12 both produce 17β-estradiol, but only UM-SCC-12, not UM-SCC-11A, xenograft tumors grow larger in vivo in response to systemic 17β-estradiol treatments. ERα66 and ERα36 expression inversely correlated with clinical cancer stage and tumor burden. LSCC ERα66 expression was higher compared to surrounding epithelia in indolent samples but lower in aggressive LSCC. ERβ expression was highly v...
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Regenerative medicine treatments for severe skeletal muscle injuries are limited, resulting in persistent functional deficits. Clinical options include neglecting the wound with the expectation that fibrosis will develop or using an... more
Regenerative medicine treatments for severe skeletal muscle injuries are limited, resulting in persistent functional deficits. Clinical options include neglecting the wound with the expectation that fibrosis will develop or using an autologous muscle graft with minimal functional improvement. A regenerative matrix can be used, but muscle fiber development on these matrices remains a challenge in vivo. Here, we explored the fundamental mechanisms that mediate cell-substrate signaling and its effect on cell-cell communication during myoblast fusion and tube formation to improve outcomes following implantation of matrices used to stimulate muscle regeneration. We previously reported that integrin-α7 was increased on anisotropic biomaterials, suggesting a role for α7β1 signaling in myoblast communication via connexin 43 and M-cadherin. Our results demonstrated that α7 silencing blocked expression of myogenic differentiation factor 1 (Myod), myogenin (Myog), myogenic factor 6 (Myf6), myo...
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Custom devices supporting bone regeneration and implant placement are needed for edentulous patients with large mandibular deficiencies where endosteal implantation is not possible. We developed a novel subperiosteal... more
Custom devices supporting bone regeneration and implant placement are needed for edentulous patients with large mandibular deficiencies where endosteal implantation is not possible. We developed a novel subperiosteal titanium-aluminum-vanadium bone onlay device produced by additive manufacturing (AM) and post-fabrication osteogenic micro-/nano-scale surface texture modification. Human osteoblasts produced osteogenic and angiogenic factors when grown on laser-sintered nano-/micro-textured surfaces compared to smooth surfaces. Surface-processed constructs caused higher bone-to-implant contact, vertical bone growth into disk pores (microCT and histomorphometry), and mechanical pull-out force at 5 and 10 w on rat calvaria compared to non surface-modified constructs, even when pre-treating the bone to stimulate osteogenesis. Surface-modified wrap-implants placed around rabbit tibias osseointegrated by 6 w. Finally, patient-specific constructs designed to support dental implants produced ...
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Estrogens cause growth plate closure in both males and females, by decreasing proliferation and inducing apoptosis of postproliferative growth plate chondrocytes. In vitro studies using 17β-estradiol (E2) conjugated to bovine serum... more
Estrogens cause growth plate closure in both males and females, by decreasing proliferation and inducing apoptosis of postproliferative growth plate chondrocytes. In vitro studies using 17β-estradiol (E2) conjugated to bovine serum albumin (E2-BSA) show that rat costochondral growth plate resting zone chondrocytes also respond to E2. Moreover, they are regulated by E2-BSA via a protein kinase C and ERK MAPK signaling pathway that is functional only in female cells. To better understand how E2 regulates apoptosis of growth plate chondrocytes, rat resting zone chondrocytes cells were treated with E2 or E2-BSA. E2 caused apoptosis in male and female resting zone and growth zone chondrocytes in a dose-dependent manner, based on elevated DNA fragmentation, terminal deoxynucleotidyl transferase dUTP nick end labeling staining and caspase-3 activation. E2 also up-regulated p53 and Bax protein (Bcl-2-associated X protein) levels and induced release of cytochrome C from the mitochondria, ind...
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Research Interests: Chemistry, Dentistry, Biomedical Engineering, Osseointegration, Cell Adhesion, and 15 moreDental Materials, Medicine, Extracellular Matrix, Cell line, Cell Differentiation, Humans, Alkaline phosphatase, Lysine, Biomimetic materials, Arginine, Cell Proliferation, Materials Testing, Drug Carriers, Extracellular Matrix Proteins, and Cell count
Recent studies have shown that porcine fetal enamel matrix derivative (EMD) can enhance the osteoinductive ability of demineralized freeze-dried bone allograft (DFDBA) in a nude mouse muscle implantation model. This suggests that one or... more
Recent studies have shown that porcine fetal enamel matrix derivative (EMD) can enhance the osteoinductive ability of demineralized freeze-dried bone allograft (DFDBA) in a nude mouse muscle implantation model. This suggests that one or more components of EMD can regulate the process of endochondral ossification initiated by DFDBA. To substantiate this hypothesis, in the current study, chondrocytes in the endochondral pathway at two stages of maturation were tested for their response to EMD. Chondrocytes were isolated from the resting zone and growth zone (prehypertrophic and upper hypertrophic cell zones) of the costochondral growth plate cartilage of adolescent rats. The results showed that the relatively immature resting zone cells responded to EMD with an increase in proliferation and a decrease in differentiation as measured by alkaline-phosphatase-specific activity. In addition, EMD stimulated a fivefold increase in PGE2 production, but was without effect on collagen synthesis...
Research Interests: Endocrinology, Chemistry, Cell Biology, Medicine, Extracellular Matrix, and 15 moreCell Division, Cell Differentiation, Cartilage, Collagen, Animals, Male, Endochondral Ossification, Medical Physiology, Alkaline phosphatase, Chondrocytes, Osteogenesis, Fetus, Chondrocyte, Biochemistry and cell biology, and Cell Growth
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Research Interests: Materials Science, Biomaterials, PHOTOELECTRON SPECTROSCOPY, Titanium, Adolescent, and 15 moreMedicine, Multidisciplinary, Aluminum, Cell Differentiation, Humans, Male, Nanostructures, Osteoblast, Mesenchymal Stem Cell, Vanadium, Surface Properties, Osteoblast differentiation, Osteoblasts, Alloys, and Mesenchymal Stromal Cells
Research Interests: Chemistry, Molecular Biology, Medical Microbiology, Biology, Cell Biology, and 15 moreApoptosis, Medicine, Biological Sciences, Nitric oxide, Animals, Cell Viability, Ascorbic Acid, Nitric Oxide Synthase, Chondrocytes, Chondrocyte, Growth Plate, Biochemistry and cell biology, Gene Expression Regulation, DNA fragmentation, and MTT assay
Research Interests: Chemistry, Biomedical Engineering, Calcium, Cell Adhesion, Cell Biology, and 15 moreMedicine, Cell Culture, Cell Division, Humans, Female, Osteoblast, Glycoproteins, Osteogenesis, Osteoblasts, Growth Factor, Estradiol, Integrin, Biochemistry and cell biology, OSTEOPROTEGERIN, and Cell culture techniques
Research Interests: Materials Science, Biomaterials, Calcium, Titanium, Immunohistochemistry, and 15 moreMedicine, Multidisciplinary, Surface Roughness, Cell Differentiation, Humans, Osteoblast, Osteocalcin, Alkaline phosphatase, Surface Properties, Transforming Growth Factor Beta, Surface Energy, Surface free energy, Osteoblasts, Biocompatible Materials, and OSTEOPROTEGERIN
Background Osseointegration depends on the implant surface, bone quality and the local and systemic host environment, which can differ in male and female patients. This study was undertaken in order to determine if male and female cells... more
Background Osseointegration depends on the implant surface, bone quality and the local and systemic host environment, which can differ in male and female patients. This study was undertaken in order to determine if male and female cells respond differently to titanium surfaces that have micron-scale roughness and if interactions of calciotropic hormones [1α,25(OH)2D3 and 17β-oestradiol (E2)] and microstructured surfaces on osteoblasts are sex dependent. Methods Osteoblasts from 6-week old Sprague-Dawley rats were cultured on tissue culture polystyrene (TCPS) or on titanium (Ti) disks with two different surface topographies, a smooth pretreated (PT) surface and a coarse grit-blasted/acid-etched (SLA) surface, and treated with 1α,25(OH)2D3, E2, or E2 conjugated to bovine serum albumin (E2-BSA). Results Male and female cells responded similarly to Ti microstructure with respect to cell number and levels of osteocalcin, transforming growth factor-β1, osteoprotegerin and prostaglandin E2...
Research Interests: Endocrinology, Osseointegration, Titanium, Medicine, Sexual dimorphism, and 12 moreInternal Medicine, Tissue culture, Surface topography, Osteoblast, Osteocalcin, Surface Properties, Microstructures, Bovine Serum Albumin, Prostaglandin E2, OSTEOPROTEGERIN, Transforming Growth Factor, and Conditioned media
Efforts to improve bone response to biomaterials have focused on ligands that bind α5β1 integrins. However, antibodies to α5β1 reduce osteoblast proliferation but do not affect differentiation when cells are grown on titanium (Ti).... more
Efforts to improve bone response to biomaterials have focused on ligands that bind α5β1 integrins. However, antibodies to α5β1 reduce osteoblast proliferation but do not affect differentiation when cells are grown on titanium (Ti). β1-silencing blocks the differentiation stimulus of Ti microtopography, suggesting that other β1 partners are important. Stably α2-silenced MG63 human osteoblast-like cells were used to test whether α2β1 specifically mediates osteoblast response to Ti surface micron-scale structure and energy. WT and α2-silenced MG63 cells were cultured on tissue culture polystyrene (TCPS) and Ti disks with different surface microtopographies: machined pretreatment (PT) surfaces [mean peak to valley roughness ( R a ) < 0.02 μm], PT surfaces that were grit-blasted and acid-etched (SLA; R a = 4 μm), and SLA with high surface energy (modSLA). Alkaline phosphatase (ALP), α2 and β1 mRNA, but not α5, αv, β3, type-I collagen, or osteocalcin, increased on SLA and modSLA at 6 d...
Research Interests: Chemistry, Medicine, Multidisciplinary, Surface Roughness, Signal Transduction, and 15 moreCell Differentiation, Humans, Osteoblast, Osteocalcin, Alkaline phosphatase, Surface Structure, Surface Properties, Microstructures, Microchemistry, Surface Energy, Osteoblasts, Biocompatible Materials, Bone and Bones, Integrin, and Cell culture techniques
C-type natriuretic peptide (CNP) possesses nitric oxide-like signaling mechanisms and actions in the vasculature, including the inhibition of fibrosis and vascular remodeling through counterregulation of transforming growth factor-β... more
C-type natriuretic peptide (CNP) possesses nitric oxide-like signaling mechanisms and actions in the vasculature, including the inhibition of fibrosis and vascular remodeling through counterregulation of transforming growth factor-β (TGF-β) signaling. The leucine zipper protein transforming growth factor stimulated clone 22 domain 1 (TSC22D1), cloned via its presumed binding to a GC-rich element in the CNP promoter, was the first protein to be described as a CNP transcription factor, but the lack of supporting evidence since its discovery and its lack of a classical DNA-binding site have left in question its role in the regulation of CNP by TGF-β and other factors. To define a specific role for TSC22D1 in CNP transcription, we have examined the effects of the profibrotic growth factors TGF-β1 and PDGF-BB on CNP mRNA expression in cultured human vascular smooth muscle cells (SMC) in which TSC22D1 expression was suppressed with small interfering RNA. Results showed that TGF-β and PDGF...
Research Interests: Endocrinology, Chemistry, Cancer, Biology, Cell Cycle, and 15 moreCell Biology, Biological Chemistry, Cancer Research, Cell Culture, Endothelial Cells, American, CREB, Epidermal Growth Factor, Adenylate Kinase, Chromatin Immunoprecipitation, Biological Macromolecules, Collagenase, Biochemistry and cell biology, Cell Growth, and Cycloheximide
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Matrix vesicles are key players in the development of the growth plate during endochondral bone formation. They are involved in the turnover of the extracellular matrix and its mineralization, as well as being a vehicle for chondrocyte... more
Matrix vesicles are key players in the development of the growth plate during endochondral bone formation. They are involved in the turnover of the extracellular matrix and its mineralization, as well as being a vehicle for chondrocyte communication and regulation. These extracellular organelles are released by the cells and are anchored to the matrix via integrin binding to collagen. The exact function and makeup of the vesicles are dependent on the zone of the growth plate in which they are produced. Early studies defined their role as sites of initial calcium phosphate deposition based on the presence of crystals on the inner leaflet of the membrane and subsequent identification of enzymes, ion transporters, and phospholipid complexes involved in mineral formation. More recent studies have shown that they contain small RNAs, including microRNAs, that are distinct from the parent cell, raising the hypothesis that they are a distinct subset of exosomes. Matrix vesicles are produced...
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The use of metallic and polymeric materials for implants has been increasing over the past decade. This trend can be attributed to a variety of factors including a significant increase in basic science research focused on implant material... more
The use of metallic and polymeric materials for implants has been increasing over the past decade. This trend can be attributed to a variety of factors including a significant increase in basic science research focused on implant material characteristics and how various surface modifications may stimulate osseointegration and, ultimately, fusion. There are many interbody fusion devices and dental implants commercially available; however, detailed information about their surface properties, and the effects that various materials and surface modifications may have on osteogenesis, is lacking in the literature. While the concept of bone-implant osseointegration is a relatively recent addition to the spine fusion literature, there is a comparatively large body of literature related to dental implants. The purpose of this article is to summarize the science of surface modified bone-facing implants, focusing on biomimetic material chemistry and topography of titanium implants, to promote ...
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Decellularized tissues are biocompatible materials that engraft well, but the age of their source has not been explored for clinical translation. Advanced glycation end products (AGEs) are chemical cross-links that accrue on skeletal... more
Decellularized tissues are biocompatible materials that engraft well, but the age of their source has not been explored for clinical translation. Advanced glycation end products (AGEs) are chemical cross-links that accrue on skeletal muscle collagen in old age, stiffening the matrix and increasing inflammation. Whether decellularized biomaterials derived from aged muscle would suffer from increased AGE collagen cross-links is unknown. We characterized gastrocnemii of 1-, 2-, and 20-month-old C57BL/6J mice before and after decellularization to determine age-dependent changes to collagen stiffness and AGE cross-linking. Total and soluble collagen was measured to assess if age-dependent increases in collagen and cross-linking persisted in decellularized muscle matrix (DMM). Stiffness of aged DMM was determined using atomic force microscopy. AGE levels and the effect of an AGE cross-link breaker, ALT-711, were tested in DMM samples. Our results show that age-dependent increases in colla...
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Advanced age causes skeletal muscle to undergo deleterious changes including muscle atrophy, fast-to-slow muscle fiber transition, and an increase in collagenous material that culminates in the age-dependent muscle wasting disease known... more
Advanced age causes skeletal muscle to undergo deleterious changes including muscle atrophy, fast-to-slow muscle fiber transition, and an increase in collagenous material that culminates in the age-dependent muscle wasting disease known as sarcopenia. Advanced glycation end-products (AGEs) non-enzymatically accumulate on the muscular collagens in old age via the Maillard reaction, potentiating the accumulation of intramuscular collagen and stiffening the microenvironment through collagen cross-linking. This review contextualizes known aspects of skeletal muscle extracellular matrix (ECM) aging, especially the role of collagens and AGE cross-linking, and underpins the motor nerve’s role in this aging process. Specific directions for future research are also discussed, with the understudied role of AGEs in skeletal muscle aging highlighted. Despite more than a half century of research, the role that intramuscular collagen aggregation and cross-linking plays in sarcopenia is well accep...
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Products developed for skeletal muscle regeneration frequently incorporate allogeneic and xenogeneic materials to elicit a regenerative response to heal skeletal muscle wounds. To avoid graft rejection in preclinical studies,... more
Products developed for skeletal muscle regeneration frequently incorporate allogeneic and xenogeneic materials to elicit a regenerative response to heal skeletal muscle wounds. To avoid graft rejection in preclinical studies, immunodeficient rodents are used. Whether the immunodeficiency alters the host response to the material in skeletal muscle has not been studied. In this study, we hypothesized that an allogeneic acellular skeletal muscle grafts implanted in an immunodeficient rat (RNU, Foxn1-deficient) would exhibit better new muscle fiber formation compared to grafts implanted in immunocompetent Sprague Dawley (SD) rats. Decellularized SD skeletal muscle matrix (DMM) was implanted in the gastrocnemius (N = 8 rats/group). 56 days after surgery, animal gait was examined and animals were euthanized. Muscle force was assessed and fiber number as well as immune cell infiltrate was measured by histomorphometry and immunohistochemistry. Animal gait and percent recovery of muscle forc...
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Osseointegration is dependent on the implant surface, surrounding bone quality, and the systemic host environment, which can differ in male and female patients. Titanium (Ti) implants with microstructured surfaces exhibit greater pullout... more
Osseointegration is dependent on the implant surface, surrounding bone quality, and the systemic host environment, which can differ in male and female patients. Titanium (Ti) implants with microstructured surfaces exhibit greater pullout strength when compared to smooth-surfaced implants and exhibit enhanced osteogenic cellular responses in vitro. Previous studies showed that 1α,25-dihydroxyvitamin D3 [1α,25(OH)D] has a greater effect on rat osteoblast differentiation on microstructured Ti compared to smooth Ti surfaces and tissue culture polystyrene (TCPS). The stimulatory effect of 17β-estradiol (E) on differentiation is observed in female osteoblasts on micro-rough Ti, but it is not known if male osteoblasts behave similarly in response to E and microtopography. This study assessed whether human male and female osteoblasts exhibit sex-specific differences in response to E and 1α,25(OH)D when cultured on microstructured Ti surfaces. Osteoblasts from three male and three female hum...
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Cell-based tissue engineering can promote cartilage tissue regeneration, but cell retention in the implant site post-delivery is problematic. Alginate microbeads containing adipose stem cells (ASCs) pretreated with chondrogenic media have... more
Cell-based tissue engineering can promote cartilage tissue regeneration, but cell retention in the implant site post-delivery is problematic. Alginate microbeads containing adipose stem cells (ASCs) pretreated with chondrogenic media have been used successfully to regenerate hyaline cartilage in critical size defects in rat xiphoid suggesting that they may be used to treat defects in elastic cartilages such as the ear. To test this, we used microbeads made with low viscosity, high mannuronate medical grade alginate using a high electrostatic potential, and a calcium cross linking solution containing glucose. Microbeads containing rabbit ASCs (rbASCs) were implanted bilaterally in 3 mm critical size midcartilage ear defects of six skeletally mature male New Zealand White rabbits (empty defect; microbeads without cells; microbeads with cells; degradable microbeads with cells; and autograft). Twelve weeks post-implantation, regeneration was assessed by microCT and histology. Microencap...
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Keratin proteins derived from human hair are known to contain a large number of cysteine residues. In hydrogels composed of oxidatively extracted keratin (keratose), sulfonic acid on the cysteine residues prevents disulfide crosslinking.... more
Keratin proteins derived from human hair are known to contain a large number of cysteine residues. In hydrogels composed of oxidatively extracted keratin (keratose), sulfonic acid on the cysteine residues prevents disulfide crosslinking. Reductively extracted keratin (kerateine) has thiol groups on cysteine residues that can form disulfide crosslinks. The rates of keratin hydrogel erosion and, in some cases, drug release can be tuned by manipulating disulfide crosslinking levels via the keratose:kerateine (KOS:KTN) ratio. To investigate the use of these hydrogel mixtures as carriers for growth factors in tissue engineering applications, we fabricated 15% (w/v) hydrogels with KOS: KTN ratios of 100:0, 70:30, 50:50, 30:70, and 0:100 with and without recombinant human bone morphogenetic protein 2 (rhBMP-2). We compared the keratin rhBMP-2 carriers to the clinical system of resorbable collagen sponges, which are known to elicit problems including edema, inflammation and ectopic bone gro...
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An increasing demand to regenerate tissues from patient-derived sources has led to the development of cell-based therapies using autologous stem cells, thereby decreasing immune rejection of scaffolds coupled with allogeneic stem cells or... more
An increasing demand to regenerate tissues from patient-derived sources has led to the development of cell-based therapies using autologous stem cells, thereby decreasing immune rejection of scaffolds coupled with allogeneic stem cells or allografts. Adult stem cells are multipotent and are readily available in tissues such as fat and bone marrow. They possess the ability to repair and regenerate tissue through the production of therapeutic factors, particularly vasculogenic proteins. A major challenge in cell-based therapies is localizing the delivered stem cells to the target site. Microencapsulation of cells provides a porous polymeric matrix that can provide a protected environment, localize the cells to one area, and maintain their viability by enabling the exchange of nutrients and waste products between the encapsulated cells and the surrounding tissue. In this chapter, we describe a method to produce injectable microbeads containing a tunable number of stem cells using the b...
Research Interests: Chemistry, Tissue Engineering, Stem Cells, Calcium, Stem Cell, and 15 moreMedicine, Stem Cell Transplantation, Cell microencapsulation, Humans, Animals, Rats, Alginates, Microspheres, Drug Compounding, Delayed-Action Preparations, Equipment Design, Cell Survival, Biochemistry and cell biology, Injections, and Cell- and tissue-based therapy
Endochondral bone formation is a precise and highly ordered process whose exact regulatory framework is still being elucidated. Multiple regulatory pathways are known to be involved. In some cases, regulation impacts gene expression,... more
Endochondral bone formation is a precise and highly ordered process whose exact regulatory framework is still being elucidated. Multiple regulatory pathways are known to be involved. In some cases, regulation impacts gene expression, resulting in changes in chondrocyte phenotypic expression and extracellular matrix synthesis. Rapid regulatory mechanisms are also involved, resulting in release of enzymes, factors and micro RNAs stored in extracellular matrisomes called matrix vesicles. Vitamin D metabolites modulate endochondral development via both genomic and rapid membrane-associated signaling pathways. 1α,25-dihydroxyvitamin D3 [1α,25(OH)D] acts through the vitamin D receptor (VDR) and a membrane associated receptor, protein disulfide isomerase A3 (PDIA3). 24R,25-dihydroxyvitamin D3 [24R,25(OH)D] affects primarily chondrocytes in the resting zone (RC) of the growth plate, whereas 1α,25(OH)Daffects cells in the prehypertrophic and upper hypertrophic cell zones (GC). This includes ...
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There is growing evidence that laryngeal cancers are responsive to sex hormones, specifically 17β-estradiol (E), despite controversy regarding the presence and characterization of Ereceptors (ER). Determination of sex hormone... more
There is growing evidence that laryngeal cancers are responsive to sex hormones, specifically 17β-estradiol (E), despite controversy regarding the presence and characterization of Ereceptors (ER). Determination of sex hormone responsiveness impacts the prognosis of laryngeal cancer patients and the treatment modalities implemented by their clinicians. Discovery of membrane-associated steroid hormone receptors and rapid membrane signaling opened the possibility that cancers previously labeled 'non-hormone dependent' and 'ER negative' might in fact be susceptible to the effects of Evia these membrane receptors. ERα66 and ERβ, the classical nuclear receptors, are present in the membranes of different cancer cells via a mechanism referred to as trafficking. Novel splice variants of these traditional receptors, a key example being ERα36, have also been found in the caveolae of cancer cells. Previous work demonstrated that ERα36 has a role in the tumorigenesis of laryngeal...
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A critical stage during osseointegration of a titanium (Ti) implant is primary bone remodeling, which involves cross talk among osteoclast precursors, osteoclasts, mesenchymal stem cells (MSCs), and osteoblasts. This phase couples the... more
A critical stage during osseointegration of a titanium (Ti) implant is primary bone remodeling, which involves cross talk among osteoclast precursors, osteoclasts, mesenchymal stem cells (MSCs), and osteoblasts. This phase couples the processes of bone formation and resorption. During remodeling, osteoclasts produce factors capable of regulating MSC migration and osteogenesis. Furthermore, they degrade primary bone, creating a foundation with a specific chemistry, stiffness, and morphology for osteoblasts to synthesize and calcify their matrix. MSCs and osteoblasts receiving cues from the implant surface produce factors capable of regulating osteoclasts in order to promote net new bone formation. The purpose of this study was to determine the effects Ti implant surfaces have on bone remodeling. Human MSCs and normal human osteoblasts (NHOsts) were cultured separately on 15 mm grade 2 smooth PT, hydrophobic-microrough SLA, hydrophilic-microrough Ti (mSLA) (Institut Straumann AG, Base...
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Current strategies to treat volumetric muscle loss use primarily pedicle or free muscle transfers, but these grafts fail to adequately regenerate functional tissue. Decellularized soft tissue grafts possess physical and chemical cues to... more
Current strategies to treat volumetric muscle loss use primarily pedicle or free muscle transfers, but these grafts fail to adequately regenerate functional tissue. Decellularized soft tissue grafts possess physical and chemical cues to promote muscle regeneration, suggesting their potential for use in large muscle defects. In this study, we developed a decellularized muscle matrix (DMM) graft using rat gastrocnemius. Anisotropy and chemical components of the extracellular matrix were retained, including laminin, fibronectin, and collagen. We compared the ability of DMM, autologous muscle grafts (clinical standard), and type I collagen plugs (negative control) to support muscle regeneration. DMM supported regeneration over a 56-day period in 1x1cm and 1.5x1cm gastrocnemius defects in rats. Muscle function tests demonstrated improved muscle recovery in rats with DMM grafts when compared to collagen. Histological sections were assessed using morphometrics and immunostaining. DMM suppo...
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Chondrocytes at different maturation states in the growth plate produce matrix vesicles (MVs), membrane organelles found in the extracellular matrix, with a wide range of contents, such as matrix processing enzymes and receptors for... more
Chondrocytes at different maturation states in the growth plate produce matrix vesicles (MVs), membrane organelles found in the extracellular matrix, with a wide range of contents, such as matrix processing enzymes and receptors for hormones. We have shown that MVs harvested from growth zone (GC) chondrocyte cultures contain abundant small RNAs, including miRNAs. Here, we determined whether RNA also exists in MVs produced by less mature resting zone (RC) chondrocytes and, if so, whether it differs from the RNA in MVs produced by GC cells. Our results showed that RNA, small RNA specifically, was present in RC-MVs, and it was well-protected from RNase by the phospholipid membrane. A group of miRNAs was enriched in RC-MVs compared RC-cells, suggesting that miRNAs are selectively packaged into MVs. High throughput array and RNA sequencing showed that ~39% miRNAs were differentially expressed between RC-MVs and GC-MVs. Individual RT-qPCR also confirmed that miR-122-5p and miR-150-5p were...
Research Interests: Chemistry, RNA, MicroRNA, Biology, Cell Biology, and 4 moreMedicine, Extracellular Matrix, Vesicle, and Chondrocyte
Low-frequency ultrasound is widely used in the treatment of chronically infected wounds. To investigate its feasibility as a method for in situ restoration of metal implant surfaces in cases of peri-implantitis, we evaluated how... more
Low-frequency ultrasound is widely used in the treatment of chronically infected wounds. To investigate its feasibility as a method for in situ restoration of metal implant surfaces in cases of peri-implantitis, we evaluated how low-frequency ultrasound affected surface properties of and response of human osteoblast-like MG63 cells to titanium (Ti). Three Ti surfaces [hydrophobic/smooth (pretreatment, PT); hydrophobic/rough (sandblasted/acid-etched, SLA); and hydrophilic/rough (SLA processed and stored hydrophilicity, mSLA)] were subjected to 25 kHz ultrasound for 10 min/cm2 . Substrate roughness, chemical composition, and wettability were analyzed before and after ultrasound application. Osteoblastic maturation of cells on sonicated disks was compared to cells on untreated disks. Ultrasound treatment altered the topography of all surfaces. Contact angles were reduced, and chemical compositions were altered by ultrasound on PT and SLA surfaces. Cell response to sonicated PT was comp...
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Micro-to-nanoscale surface topographies of orthopaedic and dental implants can affect fluid wetting and biological response. Nanoscale features can be superimposed on microscale roughness of titanium (Ti) surfaces at high temperatures,... more
Micro-to-nanoscale surface topographies of orthopaedic and dental implants can affect fluid wetting and biological response. Nanoscale features can be superimposed on microscale roughness of titanium (Ti) surfaces at high temperatures, resulting in increased osteoblast differentiation. However, high temperatures can compromise mechanical properties of the bulk material. Here, we have developed a novel low-temperature microwave hydrothermal (MWHT) oxidation process for nanomodification of microrough (SLA) Ti surfaces. Nanoscale protuberances (20 -100 nm average diameter) were generated on SLA surfaces via MWHT treatment at 200°C in H2 O, or in aqueous solutions of H2 O2 or NH4 OH, for times ranging from 1 to 40 h. The size, shape, and crystalline content of the nanoprotuberances varied with the solution used and treatment time. The hydrophilicity of all MWHT-modified surfaces was dramatically enhanced. MG63 and normal human osteoblasts (NHOsts) were cultured on MWHT-treated SLA surfa...
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17β-Estradiol (E2) promotes metastasis of triple negative breast cancer cells to bone. Recent studies show many triple negative breast cancer cell lines lacking the 66 kDa estrogen receptor (ER) alpha (ERα66) or its splice variant ERα46,... more
17β-Estradiol (E2) promotes metastasis of triple negative breast cancer cells to bone. Recent studies show many triple negative breast cancer cell lines lacking the 66 kDa estrogen receptor (ER) alpha (ERα66) or its splice variant ERα46, express another splice variant, ERα36 associated with membrane-mediated rapid actions of the hormone. qPCR and western blot confirmed that MCF7 cells possessed ERα splice variants ERα66, ERα46 and ERα36, while ER-negative breast cancer cells MDA-MB-231 possessed only ERα36. MDA-MB-231 breast cancer cells were implanted into medullary canals of ovariectomized female athymic nude mice femurs (N=8 mice/treatment). To examine the effect of E2 on osteolysis, mice were treated with 0.72mg E2 or placebo via 60 day release osmotic pumps implanted subcutaneously. Legs were examined by Faxitron through the course of the study, and by microCT and histology after 8 weeks. Greater occurrence of osteolysis and pathologic fracture was observed in E2-treated animal...
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Recent studies have shown that osteoblast-like cells respond directly to ultra-high molecular weight polyethylene particles in culture, suggesting that they may be involved in aseptic loosening of endoprostheses. We tested the hypothesis... more
Recent studies have shown that osteoblast-like cells respond directly to ultra-high molecular weight polyethylene particles in culture, suggesting that they may be involved in aseptic loosening of endoprostheses. We tested the hypothesis that the state of cell maturation plays a role in the response of osteogenic cells to ultra-high molecular weight polyethylene particles. MG63 cells (immature osteoblast-like cells), OCT-1 cells (mature secretory osteoblast-like cells), and MLO-Y4 cells (osteocyte-like cells) were treated for twenty-four hours with commercial ultra-high molecular weight polyethylene particles with an average diameter of 1 mm. The effect of particle treatment on cell proliferation was assessed by measuring the number of cells, whereas the effects on differentiation and local factor production were assessed by measuring the production of osteocalcin, prostaglandin E2, and nitric oxide. The effect of particles on apoptosis was also evaluated. The addition of ultra-high molecular weight polyethylene particles increased the number of MG63 cells, did not affect the number of OCT-1 cells, and led to a decrease in the number of MLO-Y4 cells. The observed changes in cell number were not due to programmed cell death, as no more than 3% of the cells in cultures treated with the highest concentration of particles were undergoing apoptosis. Osteocalcin production was not affected by the addition of particles. Prostaglandin E2 production was increased in all three types of cultures, but the effect was greatest in OCT-1 cell cultures, as was the absolute amount of prostaglandin E2 produced. Nitric oxide production was unaffected in MG63 cell cultures, but it was stimulated in OCT-1 and MLO-Y4 cell cultures. The results of the present study support the hypothesis that osteoblast cell maturation state plays an important role in the response to ultra-high molecular weight polyethylene particles and that the terminally differentiated osteocyte may be involved in the bone response to wear debris in vivo.
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Biologics can improve bone formation, but may diffuse away from sites of therapeutic need. We developed a click-chemistry hydrogel that rapidly polymerizes in situ to control delivery of biologics during post-suturectomy resynostosis in... more
Biologics can improve bone formation, but may diffuse away from sites of therapeutic need. We developed a click-chemistry hydrogel that rapidly polymerizes in situ to control delivery of biologics during post-suturectomy resynostosis in 21-d-old male mice. Here, we used this model to determine the role of angiogenesis in post-suturectomy resynostosis and examine whether controlled release of angiogenesis inhibitors could delay bone regeneration. Hydrogels [DB-co-PEG/poly(TEGDMA)-co-(N3-TEGDMA)] were produced containing anti-angiogenic compounds (anti-VEGFA-antibody or HIF1α-inhibitor topotecan). Bioactivity in vitro was assessed by tube length and branching points of endothelial cells in hydrogel-conditioned media. In vivo effects were examined 14 d post-suturectomy, based on the temporal analysis of angiogenic mRNAs during resynostosis following posterior frontal suture removal. MicroCT was used to quantify angiogenesis in contrast-agent-perfused blood vessels and bone defect size ...
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Regulating soft tissue repair to prevent fibrosis and promote regeneration is central to creating a microenvironment conducive to soft tissue development. Macrophages play an important role in this process. Their response can be modulated... more
Regulating soft tissue repair to prevent fibrosis and promote regeneration is central to creating a microenvironment conducive to soft tissue development. Macrophages play an important role in this process. Their response can be modulated using biomaterials, altering cytokine and growth factor secretion to promote regeneration. Electrospun polydioxanone (PDO) fiber scaffolds promoted an M2 phenotype when macrophages were cultured on large diameter, highly porous scaffolds, but an M1 phenotype on smaller diameter fibers. Here we investigated whether incorporation of galectin-1, an immunosuppressive protein that enhances muscle regeneration, could promote the M2 response. Galectin-1 was incorporated into large and small fiber PDO scaffolds during electrospinning. Galectin-1 incorporation increased arginase-1 and reduced iNOS and IL-6 production in mouse bone-marrow derived macrophages compared to PDO alone for both scaffold types. Inhibition of ERK mitogen activated protein kinase did...
Research Interests:
Steroid hormones regulate a wide variety of physiological and developmental functions. Traditional steroid hormone signaling acts through nuclear and cytosolic receptors, altering gene transcription and subsequently regulating cellular... more
Steroid hormones regulate a wide variety of physiological and developmental functions. Traditional steroid hormone signaling acts through nuclear and cytosolic receptors, altering gene transcription and subsequently regulating cellular activity. This is particularly important in hormonally-responsive cancers, where therapies that target classical steroid hormone receptors have become clinical staples in the treatment and management of disease. Much progress has been made in the last decade in detecting novel receptors and elucidating their mechanisms, particularly their rapid signaling effects and subsequent impact on tumorigenesis. Many of these receptors are membrane-bound and lack DNA-binding sites, functionally separating them from their classical cytosolic receptor counterparts. Membrane-bound receptors have been implicated in a number of pathways that disrupt the cell cycle and impact tumorigenesis. Among these are pathways that involve phospholipase D, phospholipase C, and ph...