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At least 649 records · Page 36Linked to original sources

PubMed2006

Neurofibromin plays a critical role in modulating osteoblast differentiation of mesenchymal stem/progenitor cells.

Mutations in the NF1 tumor suppressor gene cause neurofibromatosis type 1, a pandemic autosomal dominant genetic disorder with an incidence of 1:3000. Individuals with NF1 have a variety of malignant and non-malignant manifestations, including skeletal manifestations, such as osteoporosis, scoliosis and short statures. However, the mechanism(s) underlying the osseous manifestations in NF1 are poorly understood. In the present study, utilizing Nf1 haploinsufficient (+/-) mice, we demonstrate that Nf1+/- mesenchymal stem/progenitor cells (MSPC) have increased proliferation and colony forming unit-fibroblast (CFU-F) capacity compared with wild-type (WT) MSPC. Nf1+/- MSPC also have fewer senescent cells and have a significantly higher telomerase activity compared with WT MSPC. Nf1+/- MSPC have impaired osteoblast differentiation as determined by alkaline phosphatase staining, and confirmed by single CFU-F replating assays. The impaired osteoblast differentiation in Nf1+/- MSPC is consistent with the reduced expression of osteoblast markers at the mRNA level, including osteocalcin and osteonectin. Importantly, re-expression of the full-length NF1 GTPase activating related domain (NF1 GAP-related domain) is sufficient to restore the impaired osteoblast differentiation in Nf1+/- MSPC. Taken together, our results suggest that neurofibromin plays a crucial role in modulating MSPC differentiation into osteoblasts, and the defect in osteoblast differentiation may contribute at least in part to the osseous abnormalities seen in individuals with NF1.

Animals↗

PubMed2002

Short-term zinc deficiency inhibits chondrocyte proliferation and induces cell apoptosis in the epiphyseal growth plate of young chickens.

The purpose of this study was to investigate the effect of zinc deficiency on chondrocyte proliferation, differentiation and apoptosis in the epiphyseal growth plate of juvenile chickens. Newly hatched broiler chickens were fed either a low zinc (10 mg/kg diet) or a zinc-adequate (68 mg/kg diet) soy protein-based purified diet. Cell proliferation in the growth plate was evaluated with bromodeoxyuridine (BrdU) labeling. Apoptosis was assessed using the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method. Chondrocyte differentiation was evaluated with immunostaining of osteonectin as a marker of maturation. As early as d 3 of feeding, zinc deficiency significantly inhibited chondrocyte proliferation, promoted cell differentiation and induced cell apoptosis in the growth plate. These effects were manifested primarily in areas remote from the blood supply. Immunostaining for local growth factors such as insulin-like growth factor-1 (IGF-1), parathyroid hormone-related protein (PTHrP) and fibroblast growth factor-2 (FGF-2) did not reveal any differences between growth plates of zinc-deficient and zinc-adequate chickens after 3 d of feeding. By d 7, severe growth plate lesions characterized by reduced cellularity and abnormally shaped cells were formed in areas remote from blood vessels. Immunoreactive IGF-1, PTHrP and FGF-2 were all greatly reduced in the lesion. However, the growth rate and food intake of zinc-deficient chickens were not different from those of the controls during the 7-d experiment. Therefore, a direct effect of zinc deficiency on proliferation, differentiation, and apoptosis of growth plate chondrocytes was indicated.

Animals↗

PubMed2006

Energy restriction lowers the expression of genes linked to inflammation, the cytoskeleton, the extracellular matrix, and angiogenesis in mouse adipose tissue.

Using high-density oligonucleotide microarrays, we examined the actions of energy restriction (ER) on the expression of >11,000 genes in epididymal white adipose tissue (WAT) of 10- to 11-mo-old male C57Bl6 mice. Four groups were studied: controls not subjected to food restriction (CO), food-restricted 18 h before being killed (FR), short-term ER for 23 d (SER), and long-term ER for 9 mo (LER). As we reported previously, compared with CO mice, FR and SER minimally influenced the gene expression profiles; however, 345 transcripts of 6,266 genes determined to be expressed in WAT were significantly altered by LER. We focus here on the 109 (31%) of these genes that were involved in either inflammation (56 genes), cytoskeleton (16 genes), extracellular matrix (23 genes), or angiogenesis (14 genes). Among these 109 genes, 104 transcripts (95%) were down regulated by LER. Western blotting for heat shock protein 47 and osteonectin, and immunohistochemical staining for hypoxia inducible factor (HIF)-1alpha), supported the microarray data that LER down regulated the expressions of these genes. Additionally, a 75% reduction in adipocyte size with LER reflected the change in the expression of genes involved in cell morphology. Our findings provide evidence that LER suppresses the expression of genes encoding inflammatory molecules in WAT while promoting structural remodeling of the cytoskeleton, extracellular matrix, and vasculature. These alterations may play an important role in the protection against WAT-derived inflammation and in lifespan extension by LER.

Adipose Tissue↗

PubMed2004

Differences in connective tissue gene expression between normally functioning, polycystic and post-menopausal ovaries.

Arrested follicular maturation is a characteristic feature of polycystic ovary syndrome (PCOS). Follicles mature in ovarian stroma composed of extracellular matrix (ECM). However, little is known of the expression of ECM genes in polycystic ovaries. The present study compares the expression levels of genes coding for collagens, matrix metalloproteinases (MMP), their inhibitors (TIMP) and cathepsins in polycystic ovaries using fertile and post-menopausal ovaries as controls. In northern analyses, the gene expression profiles of type I and III collagen of PCOS samples resembled those observed in normal follicular phase ovaries, while mRNA levels of proalpha1(IV) collagen and TIMP-3 mRNA were significantly lower in polycystic than control ovaries. During the normal menstrual cycle, an increase was observed in MMP-9 gene expression during the luteal phase. In post-menopausal ovaries, mRNA levels for type I, III and IV collagens and osteonectin were reduced, while the MMP, TIMP (excluding TIMP-3) and cathepsins did not reflect this metabolic down-regulation. Immunohistochemical staining for MMP-9 and TIMP-4 suggested differences between polycystic and normally functioning ovaries. These data demonstrate that normal ovarian functions are associated with changes in production and degradation of ECM. The alterations observed in the production and/or distribution of type IV collagen, TIMP-3 and TIMP-4 suggest involvement of basement membranes in the pathogenesis of PCOS.

Connective Tissue↗

PubMed2000

Identification of some components of basal lamina of avian ovarian follicle.

Experiments were conducted to identify components of the basal lamina of the ovarian follicle. Pure and intact basal lamina was isolated from preovulatory follicles of the chicken ovary. Some components of the basal lamina could be solubilized with guanidine-HCl (designated Fraction 1) and remaining components with beta-mercaptomethanol containing guanidine-HCl (designated Fraction 2). With Western blot analysis, monoclonal and polyclonal antibodies raised against avian, mammalian, and human proteins recognized proteins in Fractions 1 and 2 of solubilized basal lamina. Thus, antibodies raised against extracellular matrix proteins, laminin, fibronectin, entactin or nidogen, tenascin, heparan sulfate proteoglycan, osteonectin, and Type IV collagen reacted positively with basal lamina proteins. Antibodies raised against the growth factors; epidermal growth factor; acidic and basic fibroblast growth factors; platelet-derived growth factor-AA; transforming growth factor-alpha; transforming growth factor-beta1, -beta2, -beta3, and -beta5; and insulin-like growth factor-I and -II cross-reacted with basal lamina proteins. Similarly, antibodies raised against insulin-like growth factor-binding proteins-2, -3, -4, -5, -6, and -7 cross-reacted with basal lamina proteins. In addition, antibodies generated against matrix metalloproteinases-1, -2, -3, -4, -8, -9, and -13 reacted positively with basal lamina proteins. Furthermore, antibodies produced against tissue inhibitors of matrix metalloproteinases-1, -2, -3, and -4 also reacted positively with basal lamina proteins. Moreover, interleukin-3, granulocyte macrophage-colony-stimulating factor, interferon-gamma antibodies recognized proteins in basal lamina. These observations are consistent with the view that the basal lamina of avian ovarian follicle is a store or source of biologically active molecules, namely growth factors, growth factor-binding proteins, cytokines, matrix metalloproteinases, and their tissue inhibitors. The growth factors could exert major effects on ovarian cell behavior and function, and the enzymes could participate in tissue remodeling during follicular development.

Animals↗

PubMed2003

Phosphodiesterase 4 in osteoblastic osteosarcoma cells as a potential target for growth inhibition.

Reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed that HOSM-1 cells, an osteosarcoma cell line established from human mandible, expressed mRNA for osteoblastic markers, such as alkaline phosphatase, osteonectin, osteocalcin and parathyroid hormone receptor, thus exhibiting an osteoblastic phenotype. We have investigated a possible role of cyclic nucleotide phosphodiesterases (PDEs) in osteosarcoma cells. RT-PCR analysis revealed that HOSM-1 cells expressed mRNA for PDE4A, 4B and 4C. In addition, rolipram, a specific inhibitor of PDE4, inhibited HOSM-1 cell proliferation. The finding that PDE4 is involved in proliferation of osteosarcoma cells suggests the possibility that PDE4 may be a new target for antitumor therapy.

3',5'-Cyclic-AMP Phosphodiesterases↗

PubMed1991

Connective tissue metabolism including cytokines in scleroderma.

Fibroblasts within the skin of scleroderma patients constitute a phenotypically heterogeneous population with regard to expression of collagens, cytokines, and cytokine receptors. By in situ hybridization techniques, scleroderma skin is shown to contain a subpopulation of fibroblasts that are stimulated for expression of type VI collagen; the size of this subpopulation is larger than that found in normal skin. The heterogeneity in collagen production among scleroderma fibroblasts can also be demonstrated in vitro following sorting by flow cytometric analysis. An isoform of a cytokine known to be a potent modulator of collagen expression, transforming growth factor-beta 2, is overexpressed in and around inflammatory infiltrates in biopsies of skin from scleroderma patients. Scleroderma fibroblasts grown in culture express slightly elevated levels of transcripts for transforming growth factor-beta 1, demonstrated by Northern analysis. Osteonectin, or SPARC (secreted protein, acidic and rich in cysteine), messenger RNA is also clearly elevated in fibroblasts cultured from the affected skin of scleroderma patients. The affinity of epidermal growth factor receptors on fibroblasts derived from the skin of scleroderma patients is decreased compared with that of receptors on normal fibroblasts. Platelet-derived growth factor-beta receptors were detectable by immunohistochemical staining in dermal vessels and surrounding fibroblasts in 13 of 14 biopsies of skin of scleroderma patients, whereas they were absent in sections of normal skin. These studies completed within the past year allow recognition of the importance of interactions between cell types, and the possible consequences of alterations in cytokine secretory patterns and cell responsiveness.

Connective Tissue Diseases↗

PubMed1998

The cell and molecular biology of fracture healing.

Fracture healing is a complex physiologic process that involves the coordinated participation of several cell types. By using a reproducible model of experimental fracture healing in the rat, it is possible to elucidate the integrated cellular responses that signal the pathways and the role of the extracellular matrix components in orchestrating the events of fracture healing. Histologic characterization of fracture healing shows that intramembranous ossification occurs under the periosteum within a few days after an injury. Events of endochondral ossification occur adjacent to the fracture site and span a period of up to 28 days. Remodeling of the woven bone formed by intramembranous and endochondral ossification proceeds for several weeks. Spatial and temporal expression of genes for major collagens (Types I and II), minor fibrillar collagens (Types IV and XI), and several extracellular matrix components (osteocalcin, osteonectin, osteopontin, fibronectin and CD44) are detected by in situ hybridization. Immunohistochemical studies show that expression of proliferating cell nuclear antigen is both time and space dependent and differentially expressed in the callus tissues formed by the intramembranous and endochondral processes. Chondrocytes involved in endochondral ossification undergo apoptosis (programmed cell death), and early events in fracture healing may be initiated by the expression of early response genes such as c-fos. Additional characterization and elucidation of fracture healing will lay the foundation for subsequent studies aimed at identifying mechanisms for enhancing skeletal repair.

Animals↗

PubMed1997

Bone formation in vivo: comparison of osteogenesis by transplanted mouse and human marrow stromal fibroblasts.

BACKGROUND: Marrow stromal fibroblasts (MSFs) are known to contain bone precursor cells. However, the osteogenic potential of human MSFs has been poorly characterized. The aim of this study was to compare the osteogenic capacity of mouse and human MSFs after implantation in vivo. METHODS: After in vitro expansion, MSFs were loaded into a number of different vehicles and transplanted subcutaneously into immunodeficient mice. RESULTS: Mouse MSFs transplanted within gelatin, polyvinyl sponges, and collagen matrices all formed a capsule of cortical-like bone surrounding a cavity with active hematopoiesis. In transplants of MSFs from transgenic mice harboring type I procollagen-chloramphenicol acetyltransferase constructs, chloramphenicol acetyltransferase activity was maintained for up to 14 weeks, indicating prolonged bone formation by transplanted MSFs. New bone formation by human MSFs was more dependent on both the in vitro expansion conditions and transplantation vehicles. Within gelatin, woven bone was observed sporadically and only after culture in the presence of dexamethasone and L-ascorbic acid phosphate magnesium salt n-hydrate. Consistent bone formation by human MSFs was achieved only within vehicles containing hydroxyapatite/tricalcium phosphate ceramics (HA/TCP) in the form of blocks, powder, and HA/TCP powder-type I bovine fibrillar collagen strips, and bone was maintained for at least 19 weeks. Cells of the new bone were positive for human osteonectin showing their donor origin. HA/TCP powder, the HA/TCP powder-type I bovine fibrillar collagen strips, and HA/TCP powder held together with fibrin were easier to load and supported more extensive osteogenesis than HA/TCP blocks and thus may be more applicable for therapeutic use. CONCLUSIONS: In this article, we describe the differences in the requirements for mouse and human MSFs to form bone, and report the development of a methodology for the consistent in vivo generation of extensive bone from human MSFs.

Animals↗

PubMed2002

cDNA microarray analysis of vestibular schwannomas.

BACKGROUND: Vestibular schwannomas are known to harbor mutations in the neurofibromatosis type 2 tumor suppressor gene, but the mechanism of the neurofibromatosis type 2 tumor suppressor gene action is not well understood. Identification of genes differentially expressed in normal and diseased tissues through the use of a large-scale, cDNA microarray approach may lead to increased understanding of pathways that lead to tumor formation. OBJECTIVE: The objectives of this study were to evaluate the gene expression profiles in vestibular schwannomas in comparison with normal vestibular nerve tissues and to identify pathways that may be altered in schwannomas. METHODS: Total RNA was extracted from one normal vestibular nerve and seven vestibular schwannomas. The normal vestibular nerve was from one of the seven patients with small vestibular schwannomas. Radiolabeled cDNA was synthesized and hybridized to cDNA microarray filters that contained 25,920 known genes or expressed sequence tags. Expression profiles were imaged and analyzed. Selected genes that showed three-fold or greater difference in the intensity between the normal nerve and the schwannomas were further examined by real-time polymerase chain reaction and by immunohistochemical staining. RESULTS: Forty-two genes (0.2%) were upregulated 3-fold or more in at least 5 of the 7 tumors when the filter images were compared with a normal adjacent vestibular nerve. Among them, osteonectin, an angiogenesis mediator, and RhoB GTPase, which is important in cell signaling, were significantly upregulated in 5 of 7 tumors. Among genes that were downregulated, an apoptosis-related LUCA-15 gene was highly underexpressed in 6 of 7 schwannomas when compared with the normal nerve. Also, ezrin, a relative of the NF2 protein, was significantly downregulated in 5 of 7 tumors. Real-time PCR and immunohistochemistry data support the cDNA microarray findings. CONCLUSION: Our cDNA microarray analysis of schwannomas suggested several interesting and potentially important tumorigenesis pathways associated with vestibular schwannoma formation. Further in vivo study is necessary to define the roles of these identified genes and their potential relationships with the neurofibromatosis type 2 tumor suppressor gene.

Adult↗

PubMed2006

Disc distraction shows evidence of regenerative potential in degenerated intervertebral discs as evaluated by protein expression, magnetic resonance imaging, and messenger ribonucleic acid expression analysis.

STUDY DESIGN: An animal model of degeneration was used to determine the effects of disc distraction, and was evaluated with magnetic resonance imaging (MRI) as well as gene and protein expression levels. OBJECTIVE: To investigate gene expression and MRI effects of distraction. SUMMARY OF BACKGROUND DATA: Disc degeneration can result from hyper-physiologic loading. Distracted discs with degeneration showed histologic signs of tissue recovery. METHODS: There were 18 rabbits that underwent 28 days of compression (200 N) to induce moderate disc degeneration followed by 28 days of distraction (120 N; attached and loaded distraction device) or sham distraction (attached but unloaded distraction device). Comparison was performed with 56 days of compressed discs without distraction. Quantitative outcome measures were MRI signal intensity and gene expression analysis to determine: messenger ribonucleic acid levels for extracellular matrix genes, including collagen 1, collagen 2, biglycan, decorin, aggrecan, fibromodulin, and osteonectin; and matrix-regulative genes, including matrix metalloproteinase-13, tissue-inhibitor of matrix metalloproteinase-1, and bone morphogenetic protein (BMP)-2. Immunohistology was performed for collagen 2 and BMP-2 to label cells semiquantitatively by staining of the cell-surrounding matrix. RESULTS: A total of 28 days of compression decreased signal intensity. Distraction over the same period reestablished physiologic signal intensity, however, a persistent reduction was found in sham distraction. Distraction resulted in gene expression up-regulation of collagen 1 (5.4-fold), collagen 2 (5.5-fold), biglycan (7.7-fold), and decorin (3.4-fold), while expression of fibromodulin (0.16-fold), tissue-inhibitor of matrix metalloproteinase-1 (0.05-fold), and BMP-2 (0.15-fold) was decreased, as compared with 56 days compression. Distracted discs showed more BMP-2 (19.67 vs. 3.67 in 56 days compression) and collagen 2 (18.67 vs. 11.33 in 56 days compression) positive cells per field. CONCLUSIONS: Distraction results in disc rehydration, stimulated extracellular matrix gene expression, and increased numbers of protein-expressing cells.

Animals↗

PubMed2006

Osteogenic protein-1 induced gene expression: evaluation in a posterolateral spinal pseudarthrosis model.

STUDY DESIGN: Molecular study of gene expression in rabbit lumbar pseudarthrosis repairs using reverse transcriptase polymerase chain reaction. OBJECTIVE: To evaluate differential gene expression of no graft, autograft, and osteogenic protein-1 treated pseudarthroses. SUMMARY OF BACKGROUND DATA: Osteogenic protein-1 is a potential bone graft alternative that has achieved high fusion rates in a rabbit lumbar fusion model, including in the repair of nicotine-induced pseudarthroses. A previous study established a correlation between osteogenic protein-1 fusion outcomes and an enhanced level of cytokine gene expression. The expression of such cytokines is known to be decreased in nicotine-exposed rabbit fusion masses. METHODS: Messenger ribonucleic acid was isolated from nicotine-exposed New Zealand white rabbit lumbar pseudarthroses following attempted no graft, autograft, and osteogenic protein-1 pseudarthrosis repairs. Reverse transcriptase polymerase chain reaction was used to assess the expression of angiogenin, angiopoietin, intercellular adhesion molecule, platelet-derived growth factor-beta, vascular endothelial growth factor, bone morphogenetic proteins 2 and 7, type I collagen, and osteonectin. Glyceraldehyde-3-phosphate dehydrogenase was used as a constitutively expressed control. RESULTS: Levels of gene expression in the osteogenic protein-1 group were higher than those of the autograft group, which were higher than the no graft group for the majority of the genes studied. CONCLUSIONS: In the rabbit pseudarthrosis model, gene expression data supported the hypothesis that successful pseudarthrosis repair is related to the induction of osteogenic and angiogenic cytokines by osteogenic protein-1.

Animals↗

PubMed2005

Role of bovine bone morphogenetic proteins in bone matrix protein and osteoblast-related gene expression during rat bone marrow stromal cell differentiation.

Bone morphogenetic proteins (BMPs) are known to promote osteogenesis, and clinical trials are currently underway evaluating the ability of certain BMPs to promote bone graft and fracture healing. To observe the mechanism of osteoinductive and bone formation, 100 microg of bovine BMP was tested during osteogenic differentiation of rat bone marrow stromal cells (MSCs) and C2C12 line culture for 14 and 28 days. We examined alkaline phosphatase (ALP) by assay, immunohistochemical studies for bone matrix proteins, and mRNA expression of bone matrix proteins and osteoblast-related analysis by reverse-transcription polymerase chain reaction. ALP activity in MSC cultures was elevated by bovine BMP by two to fivefold (P < 0.05-0.001). DNA and protein content increased over 14 days. BMP significantly increased the mRNA expression of type I collagen, ALP, osterix, osteocalcin, osteopontin, vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF)-A, and parathyroid hormone receptor time dependently during the osteoblastic differentiation. There was no markedly enhanced mRNA expression of bone sialoprotein (BSP) and glyceraldehyde-3-phosphate dehydrogenase compared with that of control. Immunohistochemical results also showed BMP increased immunoreactive positivity of type I collagen, osteocalcin, osteonectin, osteopontin, and BSP during the C2C12 differentiation. These data indicated that BMP enhances our ability to stimulate the differentiation of osteoblast-like cells and increases osteoinductivity, bone matrix protein formation and mineralization, angiogenesis, and chondrogenesis during osteoblast progenitor cell differentiation in vitro and that the role of chondrogenic is weak.

Alkaline Phosphatase↗

PubMed1992

Haemopoietic long-term bone marrow cultures from adult mice show osteogenic capacity in vitro on 3-dimensional collagen sponges.

Adult murine bone marrow cells, cultured under conditions for long-term haemopoietic marrow cultures, produce bone matrix proteins and mineralized tissue in vitro, but only after the adherent stromal cells were loaded on a 3-dimensional collagen sponge. Provided more than 8 x 10(6) cells are loaded, mineralization as measured by 85Sr uptake from the culture medium, occurred in this 3-dimensional configuration (3-D) within 6 days. In contrast if undisrupted marrow fragments (containing more than 10(7) cells) are placed directly on a collagen sponge, then it requires more than 10 days before significant mineralization can similarly be detected. The 2-dimensional (2-D) long-term marrow culture system allows prior expansion of the stromal cells and some differentiation in an osteogenic direction within the adherent stromal layer. This is suggested by the presence of type I collagen and alkaline phosphatase positive cells. However; synthesis of osteonectin and a bone specific protein, osteocalcin, as well as calcification are only observed in 3-D cultures. Electron microscopy demonstrated hydroxyapatite mineral on collagen fibres, osteoblast-like cells, fibroblasts, cells which accumulated lipids, and macrophages which were retained on the collagen matrices. Irradiation of confluent long-term bone marrow cultures, prior to their loading on the collagen sponge showed that haemopoietic stem cells are not necessary for the mineralization.

Animals↗

PubMed1990

Formation of chondrous and osseous tissues in micromass cultures of rat frontonasal and mandibular ectomesenchyme.

Rat frontonasal and mandibular mesenchyme was isolated from day-12 1/2 (stage-22) rat embryos and cultured at high density for up to 12 days. The stage chosen was based on the observation that mandibular mesenchyme at this stage became independent of its epithelium with respect to the production of both cartilage and bone. Frontonasal cultures developed aggregates of anastomosing columns of cells within 2 days. These grew as the cells enlarged, laying down an Alcian-blue-positive matrix by day 3 of culture. Significant mineral was detected by von Kossa staining by day 5 at which time the aggregates covered a large portion of the culture, eventually covering the entire micromass by day 10-12. Mandibular cultures developed centrally located nodular aggregates by 3 days of culture. These nodules increased in number, spreading outwards as the cells enlarged, laying down an Alcian-blue-positive matrix by day 4 and mineral by days 6-7. At this time the nodules began to elongate and coalesce, but never covered the entire culture over the 12-day period. Antibody staining revealed that in both cultures the cells were initially positive for type I collagen. Subsequently, the aggregates began expressing type II collagen, followed by type X, which coincided with the onset of mineralization. At this time some cells were negative for these cartilage markers, but positive for osteoblast markers, bone sialoprotein II, osteocalcin and type I collagen. In addition osteonectin and alkaline phosphatase were demonstrable in all of the aggregate cells late in the culture period. This provided clear evidence that chondroblast and osteoblast differentiation was proceeding within these cultures. The culture of rat facial mesenchyme should prove very useful, not only for the analysis of bone and cartilage induction and lineage relationships, but also in furthering our knowledge of craniofacial differentiation, growth and pattern formation by extending our analysis to a mammalian system.

Animals↗

PubMed1989

Structure and biological activity of basement membrane proteins.

Collagen type IV, laminin, heparan sulfate proteoglycans, nidogen (entactin) and BM-40 (osteonectin, SPARC) represent major structural proteins of basement membranes. They are well-characterized in their domain structures, amino acid sequences and potentials for molecular interactions. Such interactions include self-assembly processes and heterotypic binding between individual constituents, as well as binding of calcium (laminin, BM-40) and are likely to be used for basement membrane assembly. Laminin, collagen IV and nidogen also possess several cell-binding sites which interact with distinct cellular receptors. Some evidence exists that those interactions are involved in the control of cell behaviour. These observations have provided a more defined understanding of basement membrane function and the definition of new research goals in the future.

Animals↗

PubMed2006

Effects of tacrolimus, cyclosporin A and sirolimus on MG63 cells.

The reduction in bone mineral density after organ transplantation results in increased morbidity (post-transplantation bone disease) and remains an unsolved problem. A connection with the long-term application of nonglucocorticoidal immunosuppressants is the subject of controversial discussion. We hypothesized that such substances have an influence on the skeletal system on the cellular level by modulating osteoblast differentiation. Therefore, we investigated the effects of tacrolimus, cyclosporin A and sirolimus as representative substances of nonglucocorticoidal immunosuppressants on cell proliferation and expression of bone tissue-specific genes of human osteoblasts (MG63). None of the examined substances affected cell proliferation, but all influenced the gene expression pattern towards change in cell differentiation. In detail, collagen III and XII, matrix metalloproteinase 2, SMAD2, epithelial growth factor receptor, annexin V and osteonectin expression were increased by all of the examined substances. Tacrolimus, cyclosporin A and sirolimus influence intracellular signalling pathways, transmembranous receptors and bone-specific matrix synthesis. They do not have antiproliferative or toxic effects. We postulate that the shown changes of osteoblast differentiation cause post-transplantation disease.

Bone Density↗

PubMed2005

TGF-beta3 induces ectopic mineralization in fetal mouse dental pulp during tooth germ development.

Several members of the transforming growth factor (TGF)-beta superfamily are expressed in developing teeth from the initiation stage through adulthood. Of those, TGF-beta1 regulates odontoblast differentiation and dentin extracellular matrix synthesis. However, the molecular mechanism of TGF-beta3 in dental pulp cells is not clearly understood. In the present study, beads soaked with human recombinant TGF-beta3 induced ectopic mineralization in dental pulp from fetal mouse tooth germ samples, which increased in a dose-dependent manner. Further, TGF-beta3 promoted mRNA expression, and increased protein levels of osteocalcin (OCN) and type I collagen (COL I) in dental pulp cells. We also observed that the expression of dentin sialophosphoprotein and dentin matrix protein 1 was induced by TGF-beta3 in primary cultured dental pulp cells, however, not in calvaria osteoblasts, whereas OCN, osteopontin and osteonectin expression was increased after treatment with TGF-beta3 in both dental pulp cells and calvaria osteoblasts. Dentin sialoprotein was also partially detected in the vicinity of TGF-beta3 soaked beads in vivo. These results indicate for the first time that TGF-beta3 induces ectopic mineralization through upregulation of OCN and COL I expression in dental pulp cells, and may regulate the differentiation of dental pulp stem cells to odontoblasts.

Animals↗