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Life Sciences 1996Osteonectin is an extracellular matrix (ECM) protein which is secreted by various cell types, and regulates tissue remodeling and cell proliferation. In the present...
Osteonectin is an extracellular matrix (ECM) protein which is secreted by various cell types, and regulates tissue remodeling and cell proliferation. In the present study, we have examined the expression of osteonectin in fibrotic liver. Osteonectin transcripts were undetectable in normal liver, however, the abundant expression of the osteonectin gene was detected in fibrotic liver. The transcripts of osteonectin were only detected in hepatic lipocytes and the number of lipocytes was increased in fibrotic liver. These results suggested that in fibrotic liver, enhanced osteonectin expression may play an important role in liver fibrosis.
Topics: Animals; Base Sequence; Gene Expression; Immunohistochemistry; Liver; Liver Cirrhosis; Male; Molecular Sequence Data; Osteonectin; Rats; Rats, Wistar
PubMed: 8786698
DOI: 10.1016/0024-3205(96)00035-5 -
American Journal of Physiology. Cell... Aug 2022Megakaryocyte hyperplasia associated with myeloproliferative neoplasms commonly leads to abnormal bone tissue deposition in the bone marrow, known as osteosclerosis. In...
Megakaryocyte hyperplasia associated with myeloproliferative neoplasms commonly leads to abnormal bone tissue deposition in the bone marrow, known as osteosclerosis. In this study, we aimed to synthesize the known proteomics literature describing factors released by megakaryocytes and platelets and to examine if any of the secreted factors have a known ability to stimulate the bone-forming cells, osteoblasts. Using a systematic search of Medline, we identified 77 articles reporting on factors secreted by platelets and megakaryocytes. After a full-text screening and analysis of the studies, we selected seven papers that reported proteomics data for factors secreted by platelets from healthy individuals. From 60 proteins reported in at least two studies, we focused on 23 that contained a putative signal peptide, which we searched for a potential osteoblast-stimulatory function. From nine proteins with a positive effect on osteoblast formation and function, two extracellular matrix (ECM) proteins, secreted protein acidic and rich in cysteine (SPARC) and tissue inhibitor of metalloproteinase-1 (TIMP1), and three cellular proteins with known extracellular function, the 70-kDa heat shock protein (HSP70), thymosin-β4 (TB4), and super dismutase (SOD), were identified as hypothetical candidate molecules to be examined as potential mediators in mouse models of osteomyelofibrosis. Thus, careful analysis of prior literature can be beneficial in assisting the planning of future experimental studies.
Topics: Animals; Blood Platelets; Extracellular Matrix Proteins; Mice; Osteoblasts; Osteonectin; Secretome; Tissue Inhibitor of Metalloproteinase-1
PubMed: 35675640
DOI: 10.1152/ajpcell.00187.2022 -
Experimental Cell Research Aug 2023Fibrotic scar is a severe side effect of trabeculectomy, resulting in unsatisfactory outcomes for glaucoma surgery. Accumulating evidence showed human Tenon's...
BACKGROUND
Fibrotic scar is a severe side effect of trabeculectomy, resulting in unsatisfactory outcomes for glaucoma surgery. Accumulating evidence showed human Tenon's fibroblasts (HTFs) play an important role in fibrosis formation. We previously reported that the aqueous level of secreted protein acidic and rich in cysteine (SPARC) was higher in the patients with primary angle closure glaucoma, which was associated with the failure of trabeculectomy. In this study, the potential effect and mechanism of SPARC in promoting fibrosis were explored by using HTFs.
METHODS
HTFs were employed in this study and examined under a phase-contrast microscope. Cell viability was determined by CCK-8. The expressions of SPARC-YAP/TAZ signaling and the fibrosis-related markers were examined with reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), Western blot, and immunofluorescence, subcellular fractionation was conducted to further determined the variation of YAP and phosphorylated YAP. The differential gene expressions were analyzed with RNA sequencing (RNAseq), followed by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses.
RESULTS
Exogenous SPARC induced HTFs-myofibroblast transformation, as evidenced by the increased expression of α-SMA, collagen I and fibronectin in both protein and mRNA levels. SPARC knockdown decreased the expressions of the above genes in TGF-β2-treated HTFs. KEGG analysis showed that the Hippo signaling pathway was mostly enriched. SPARC treatment increased the expressions of YAP, TAZ, CTGF and CYR61 as well as enhanced YAP translocation from cytoplasm to nucleus, and decreased the phosphorylation of YAP and LAST1/2, which was reversed by SPARC knockdown. Knockdown of YAP1 decreased the fibrosis-related markers, such as α-SMA, collagen I and Fibronectin, in SPARC-treated HTFs.
CONCLUSIONS
SPARC induced HTFs-myofibroblast transformation via activating YAP/TAZ signaling. Targeting SPARC-YAP/TAZ axis in HTFs might provide a novel strategy for inhibiting fibrosis formation after trabeculectomy.
Topics: Humans; Myofibroblasts; Fibronectins; Osteonectin; Fibroblasts; Collagen Type I; Fibrosis; Cells, Cultured
PubMed: 37225012
DOI: 10.1016/j.yexcr.2023.113649 -
The American Journal of Pathology Sep 1997Osteonectin/SPARC is a glycoprotein involved in the regulation of cell shape, adhesion, migration, and proliferation. It also has complex effects on extracellular matrix...
Osteonectin/SPARC is a glycoprotein involved in the regulation of cell shape, adhesion, migration, and proliferation. It also has complex effects on extracellular matrix synthesis and turnover. We found that osteonectin mRNA was very abundant in a human liver myofibroblast library. Using Northern and Western blot, immunoprecipitation, and radioimmunoassay, we found that cultured liver myofibroblasts actively secreted osteonectin. Myofibroblasts are very rare in normal liver but proliferate during liver fibrosis where they synthesize extracellular matrix components. Thus, we studied the distribution of osteonectin in normal and fibrotic human liver using in situ hybridization. Osteonectin mRNA expression was weak in normal liver but very high in fibrotic liver within fibrous septae and scattered sinusoidal cells. Serial sectioning and double staining experiments with an antibody to smooth muscle alpha-actin showed that osteonectin transcripts were mostly co-localized with myofibroblasts. In conclusion, osteonectin is highly expressed in human liver myofibroblasts in culture as well as in human liver fibrosis in vivo. The many biological properties of osteonectin make it a candidate effector of human liver fibrogenesis.
Topics: Actins; Blotting, Northern; Cells, Cultured; Humans; Immunoblotting; Immunohistochemistry; In Situ Hybridization; Liver; Liver Cirrhosis; Osteonectin; RNA, Messenger
PubMed: 9284812
DOI: No ID Found -
Current Protocols in Cell Biology Feb 2003SPARC is a matricellular protein that regulates cell adhesion, extracelluolar matrix production, growth factor activity and cell cycle. This unit describes the...
SPARC is a matricellular protein that regulates cell adhesion, extracelluolar matrix production, growth factor activity and cell cycle. This unit describes the purification of SPARC, also termed osteonectin and BM/40, from cultured mammalian cells. Additional information is presented on the purification of recombinant SPARC (rSPARC) from E. coli and from Sf9 cells, as well as its isolation from blood platelets. Assays for the activity of SPARC, de-adhesion and inhibition of cellular proliferation in vitro, are described. The expression of SPARC during remodeling and repair tissue in response to injury identifies it as a therapeutic target for the treatment of fibrotic disease, certain cancers and other disorders in which regulation of angiogenesis is a key factor.
Topics: Animals; Biological Assay; Cell Culture Techniques; Cell Line, Tumor; Chromatography; Culture Media; Escherichia coli; Humans; Insecta; Mice; Osteonectin; Recombinant Proteins
PubMed: 18228415
DOI: 10.1002/0471143030.cb1011s17 -
Annals of Surgery Aug 2005We sought to examine the expression and functional role of osteonectin in primary and metastatic pancreatic ductal adenocarcinoma (PDAC).
OBJECTIVE
We sought to examine the expression and functional role of osteonectin in primary and metastatic pancreatic ductal adenocarcinoma (PDAC).
BACKGROUND
The glycoprotein osteonectin plays a vital role in cell-matrix interactions and is involved in various biologic processes. Overexpression of osteonectin is present in malignant tumors and correlates with disease progression and poor prognosis.
METHODS
Expression of osteonectin was analyzed by quantitative polymerase chain reaction and immunohistochemistry in pancreatic tissues and by enzyme-linked immunosorbent assay in the serum of patients and donors. Recombinant osteonectin and specific antisense oligonucleotides were used to examine the effects of osteonectin on induction of target genes, and on proliferation and invasiveness of pancreatic cancer cells.
RESULTS
There was a 31-fold increase in osteonectin mRNA levels in PDAC and a 16-fold increase in chronic pancreatitis as compared with the normal pancreas (P < 0.01). By immunohistochemistry, faint immunoreactivity was detected in the normal pancreas. In contrast, strong staining of the cancer cells was observed in addition to extensive osteonectin immunoreactivity in surrounding fibroblasts and in the extracellular matrix. In metastatic tissues, strong immunoreactivity was observed in fibroblasts and in extracellular matrix surrounding metastatic cancer cells, whereas the signal was absent in most tumor cells. In vitro studies showed that osteonectin was able to inhibit cancer cell growth while promoting invasiveness of pancreatic tumor cells.
CONCLUSION
Osteonectin is markedly overexpressed in pancreatic cancer and has the potential to increase the invasiveness of pancreatic cancer cells.
Topics: Adenocarcinoma; Adult; Aged; Disease Progression; Enzyme-Linked Immunosorbent Assay; Extracellular Matrix; Female; Fibroblasts; Humans; Immunohistochemistry; Male; Middle Aged; Neoplasm Invasiveness; Oligonucleotides, Antisense; Osteonectin; Pancreatic Neoplasms; Pancreatitis; Polymerase Chain Reaction; Prognosis; RNA, Messenger; Recombinant Proteins; Tumor Cells, Cultured
PubMed: 16041213
DOI: 10.1097/01.sla.0000171866.45848.68 -
Investigative Ophthalmology & Visual... Aug 2000Osteonectin/SPARC is a secreted protein that has been implicated in ocular disease. Deletion of osteonectin/SPARC causes age-onset cataract in mice and the cataractous...
PURPOSE
Osteonectin/SPARC is a secreted protein that has been implicated in ocular disease. Deletion of osteonectin/SPARC causes age-onset cataract in mice and the cataractous human lens has increased expression of osteonectin/SPARC. In this study, the expression and localization of osteonectin/SPARC in the monkey retina were determined as was secretion by cultured human retinal pigment epithelial (RPE) cells.
METHODS
Adult Rhesus monkey eyes (Macaca mulatta) were dissected, and 5-mm macula and peripheral retina punches were obtained. Supernatants were collected from cultured human RPE cells. Subcellular fractionation of whole monkey retina was also performed. Osteonectin/SPARC expression and/or secretion was monitored by Northern and Western blot analyses, and localization was determined by immunocytochemistry.
RESULTS
Outside of the retina osteonectin/SPARC mRNA is broadly expressed in many human tissues. Northern blot analysis shows that in the retina osteonectin/SPARC is expressed almost exclusively by the macular RPE/choroid. Western blot analysis revealed osteonectin/SPARC in both the macula and the peripheral neural retina but only in trace amounts in the RPE/choroid. In subcellular fractions of the whole retina, osteonectin/SPARC was detected, mainly in the soluble fraction but also in the membrane and nuclear fractions. Immunohistochemical analysis localized osteonectin/SPARC specifically to the outer plexiform layer. Western blot analysis of conditioned medium from human RPE cells cultured on porous substrates indicated that osteonectin/SPARC is secreted in large amounts from both the apical and basal sides of the RPE.
CONCLUSIONS
Collectively these data provide evidence that osteonectin/SPARC is synthesized in the macular RPE, secreted, and subsequently transported to the outer plexiform layer. The expression pattern of osteonectin/SPARC in the subcellular retinal fractions is consistent with a soluble protein that is transported and internalized.
Topics: Animals; Biological Transport; Blotting, Northern; Blotting, Western; Cells, Cultured; DNA Primers; Electrophoresis, Polyacrylamide Gel; Eye Proteins; Immunoenzyme Techniques; Macaca mulatta; Osteonectin; Pigment Epithelium of Eye; RNA, Messenger; Retina; Subcellular Fractions
PubMed: 10937551
DOI: No ID Found -
Eye (London, England) Jul 2002Epiretinal and subretinal membranes are fibrocellular proliferations which form on the surfaces of the neuroretina as a sequel to a variety of ocular diseases. When... (Review)
Review
Epiretinal and subretinal membranes are fibrocellular proliferations which form on the surfaces of the neuroretina as a sequel to a variety of ocular diseases. When these proliferations complicate rhegmatogenous retinal detachment (a condition known as proliferative vitreoretinopathy or PVR), the membranes often contain numerous retinal pigment epithelial (RPE) cells and a variety of extracellular proteins. The extracellular proteins include adhesive proteins like collagen, laminin and fibronectin. In addition, several matricellular proteins with potential counter-adhesive functions are present in the membranes. Two such matricellular proteins, thrombospondin 1 and osteonectin (or SPARC: Secreted Protein Acidic and Rich in Cysteine), tend to be co-distributed with the RPE cells in PVR membranes. By virtue of their counter-adhesive properties, thrombospondin 1 and SPARC may reduce RPE cell-matrix adhesion and so permit key RPE cellular activities (for example, migration or shape change) in periretinal membrane development. Furthermore, within a 'cocktail' containing other proteins such as the metalloproteinases and growth factors like the scatter factor/hepatocyte growth factor family, matricellular proteins may play a role in the RPE cell dissociation from Bruch's membrane, which characterises early PVR.
Topics: Epiretinal Membrane; Humans; Osteonectin; Pigment Epithelium of Eye; Thrombospondin 1; Vitreoretinopathy, Proliferative
PubMed: 12101446
DOI: 10.1038/sj.eye.6700196 -
Blood Mar 1990Our laboratory has previously shown that osteonectin, an abundant noncollagenous bone protein, is contained in and secreted from human platelets. In this study, the...
Our laboratory has previously shown that osteonectin, an abundant noncollagenous bone protein, is contained in and secreted from human platelets. In this study, the distribution of osteonectin both in the supernatant and on the platelet surface after activation was measured by fluid-phase and solid-phase radioimmunoassay, respectively. Total cellular osteonectin was determined by RIA of guanidinium chloride extracted platelets and ranged from 0.65 to 2.2 micrograms/10(8) platelets or 135,000 to 457,000 molecules/platelet. Platelets treated with varying concentrations of collagen and thrombin released osteonectin in a dose-dependent fashion. Approximately 61% of the total platelet osteonectin was secreted at saturating concentrations of collagen and thrombin. A small fraction of platelet osteonectin is expressed on the surface of platelets in an activation-specific manner as evidenced by the specific and saturable binding of [125I]-anti-osteonectin monoclonal antibody, IIIA3A8, to thrombin-activated platelets. Based on a non-linear least squares regression analysis of the antibody binding, 2,200 IIIA3A8 molecules, or 0.8% of the total platelet osteonectin, is expressed on the platelet surface on activation. Platelet osteonectin was purified from the supernatant of thrombin-activated platelets by immunoaffinity chromatography. Western blotting of proteins secreted by washed, thrombin-stimulated platelets with IIIA3A8 indicated that the osteonectin molecule released from the platelet is a single chain polypeptide. Comparison of immunopurified platelet osteonectin with isolated bovine bone osteonectin and isolated human bone osteonectin by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that platelet osteonectin has a greater apparent molecular weight than bone osteonectin. The NH2-terminal sequence of immunopurified platelet osteonectin was obtained by automated Edman degradation and is identical to the sequence of human bone osteonectin derived from the cDNA of SaOS-2 cells. Collectively, these data suggest that platelet osteonectin is structurally distinct from bone osteonectin in a region of the molecule at a distance from the NH2-terminus.
Topics: Amino Acid Sequence; Antibodies, Monoclonal; Antigens, Surface; Blood Platelets; Collagen; Humans; Molecular Sequence Data; Molecular Weight; Osteonectin; Platelet Activation; Radioimmunoassay; Thrombin
PubMed: 2306517
DOI: No ID Found -
Cancer Research Aug 2005Skeletal metastases occur with high incidence in patients with breast cancer and cause long-term skeletal morbidity. Osteonectin (SPARC, BM-40) is a bone matrix factor...
Skeletal metastases occur with high incidence in patients with breast cancer and cause long-term skeletal morbidity. Osteonectin (SPARC, BM-40) is a bone matrix factor that is an in vitro chemoattractant for breast and prostate cancer cells. Increased expression of osteonectin is found in malignant breast tumors. We infected MDA-231 breast cancer cells with an adenovirus expressing osteonectin to examine the role of osteonectin expression in breast cancer cells and its effect on metastasis, in particular to bone. Expression of osteonectin did not affect MDA-231 cell proliferation, apoptosis, migration, cell aggregation, or protease cleavage of collagen IV. However, in vitro invasion of these osteonectin-infected cells through Matrigel and colony formation on Matrigel was decreased. Interestingly, high osteonectin expression in MDA-231 cells inhibited metastasis in a dose-dependent manner to many different organs including bone. The reduction in metastasis may be due to decreased platelet-tumor cell aggregation, because exogenous osteonectin inhibited platelet aggregation in vitro and the high osteonectin expression in MDA-231 cells reduced tumor cell-induced thrombocytopenia in vivo compared with control-infected cells. These studies suggest that high endogenous expression of osteonectin in breast cancer cells may reduce metastasis via reduced invasive activity and reduced tumor cell-platelet aggregation.
Topics: Adenoviridae; Animals; Blood Platelets; Breast Neoplasms; Cell Growth Processes; Cell Line, Tumor; Female; Genetic Vectors; Humans; Mice; Mice, Nude; Neoplasm Invasiveness; Neoplasm Metastasis; Osteonectin; Thrombocytopenia
PubMed: 16103089
DOI: 10.1158/0008-5472.CAN-05-0807