Biomedical subjects

Rang-Woon Park

Publications and source records attributed to Rang-Woon Park.

At least 19 recordsLinked to original sources

PubMed2006

Antiangiogenic effect of bile acid acylated heparin derivative.

PURPOSE: Chemically modified heparin-DOCA was prepared and found to have markedly lower anticoagulant activity than heparin. In the present study, we elucidated the antiangiogenic and antitumoral activities of heparin-DOCA derivative. METHODS: To evaluate the antiangiogenic and antitumoral effects of heparin-DOCA, capillary-like tube formation assay, Matrigel plug assay in vivo, western blotting for FGFR phosphorylation, ERK and p38 MAPK activities, tumor growth of SCC in vivo and immunostaining of blood vessels in tumor tissues were performed. RESULTS: Heparin-DOCA inhibited capillary-like tubular structures of endothelial cells and bFGF-induced neovascularizations in Matrigel plug assays. Signaling experiments showed that heparin-DOCA significantly inhibited angiogenesis by suppressing the phosphorylation of FGFR and its downstream signal pathways (ERK and p38 MAPK activities). The antiangiogenic activity of this heparin derivative was found to be closely associated with antitumoral activity in a mouse model. In addition, histological evaluations supported the inhibitory effect of heparin-DOCA on blood vessel formation in tumor tissues. CONCLUSION: Heparin-DOCA derivative exerted a significant antitumoral effect by inhibiting angiogenesis resulting from the disruption of FGF/FGFR and its downstream signal pathways, and could be applied to treat various angiogenic diseases.

Angiogenesis Inhibitors↗

PubMed2006

Polymorphisms in the FAS and FASL genes and risk of lung cancer in a Korean population.

BACKGROUND: The FAS and FASL system play an important role in regulating extrinsic apoptotic pathway and inappropriate regulation of this signaling pathway contributes to lung tumorigenesis. Polymorphisms in the promoter region of the FAS (-1377G>A and -670A>G) and FASL (-844C>T) have been shown to alter the transcriptional activities of these genes. In order to evaluate the contribution of these polymorphisms to the risk of lung cancer, we carried out a case-control study in a Korean population. METHODS: The FAS and FASL genotypes were determined in 582 lung cancer patients and 582 healthy control subjects who were frequency-matched for age and gender. RESULTS: The FAS and FASL genotypes and the FAS haplotypes exhibited no apparent relationship with the risk of lung cancer. In addition, there was no significant interaction between the FAS and FASL polymorphisms in the development of lung cancer. CONCLUSION: These results suggest that the FAS-1377G>A and -670A>G and FASL-844C>T polymorphisms do not significantly affect the susceptibility to lung cancer in Koreans.

Asian People↗

PubMed2006

pH- and temperature-sensitive, injectable, biodegradable block copolymer hydrogels as carriers for paclitaxel.

Paclitaxel (PTX) was loaded into synthetic pH/T-sensitive block copolymer (OSM-PCLA-PEG-PCLA-OSM) solution with various concentrations. The phase diagram of PTX-loaded block copolymer solution shifted to lower temperature region compared to net block copolymer because of the salting-out effect of PTX. Release profiles of PTX showed sustained manner regardless of loading amount of PTX. To evaluate anti-tumor effect of PTX-loaded block copolymer, solutions were injected subcutaneously to tumor-bearing mice and TUNEL assay examined. PTX-loaded block copolymer hydrogel for in vivo use showed good anti-tumor effect for 2 weeks and induced strong apoptosis in tumor tissue. Therefore, we conclude OSM-PCLA-PEG-PCLA-OSM block copolymer as an effective injectable carrier of PTX.

Animals↗

PubMed2006

Heparin-deoxycholic acid chemical conjugate as an anticancer drug carrier and its antitumor activity.

A chemically modified heparin-DOCA (HD) conjugate was developed as a drug carrier for cancer therapy. HD conjugate was found to have markedly low anticoagulant activity and to form self-assembled nanoparticles in aqueous condition. We observed that HD conjugate prevented squamous cell carcinoma (SCC) and human umbilical vascular endothelial cell (HUVEC) proliferation during BrdU incorporation assays. Here, we prepared doxorubicin-loaded heparin nanoparticles by entrapping doxorubicin into the amphiphilic HD conjugate by physical interaction and characterized the properties of these nanoparticles using Dynamic Light Scattering (DLS) and Atomic Force Microscope (AFM). In this study, doxorubicin-loaded heparin nanoparticles were designed to improve the antitumor effects of nano-sized particles (range of 180 to 210 nm) at high drug-loading efficiencies in the range 64% to 96%. These doxorubicin-loaded heparin nanoparticles displayed sustained drug release patterns. It was confirmed in vivo toxicity studies that HD conjugate did not induce unexpected side effects and that DHN 20 was safer than free DOX. An in vivo study showed that HD conjugate, doxorubicin and DHN 20 (one of doxorubicin-loaded heparin nanoparticles) induced tumor volume reductions of 43%, 56% and 74%, respectively, relative to the saline treated control. These results suggest that the drug-entrapped with heparin nanoparticles might provide a novel therapy for SCC.

Animals↗

PubMed2006

Polymorphisms in TGF-beta1 gene and the risk of lung cancer.

BACKGROUND: Transforming growth factor-beta1 (TGF-beta1) functions as a suppressor of tumor initiation by inhibiting cellular proliferation or by promoting cellular differentiation or apoptosis in the early phase of cancer development. Variations in the DNA sequence in the TGF-beta1 gene may lead to altered TGF-beta1 production and/or activity, and so this can modulate an individual's susceptibility to lung cancer. To test this hypothesis, we investigated the association of the TGF-beta1 -509C > T and 869T > C (L10P) polymorphisms and their haplotypes with the risk of lung cancer in a Korean population. METHODS: The TGF-beta1 genotypes were determined in 432 lung cancer patients and in 432 healthy control subjects who were frequency-matched for age and gender. The TGF-beta1 haplotypes were predicted using a Bayesian algorithm in the Phase program. RESULTS: Individuals with at least one -509T allele were at a significantly decreased risk of adenocarcinoma (AC) and small cell carcinoma (SM), as compared with carriers with the -509CC genotype [adjusted odds ratio (OR), 0.63; 95% confidence interval (CI), 0.42-0.96; P = 0.04; and adjusted OR, 0.45; 95% CI, 0.27-0.76; P = 0.002; respectively]. For the 869T > C polymorphism, the combined TC + CC genotype was associated with a significantly decreased risk of SM compared with the TT genotype (adjusted OR, 0.52; 95% CI, 0.31-0.88; P = 0.01). Consistent with the results of the genotyping analyses, the -509T/869C haplotype was associated with a significantly decreased risk of AC and SM as compared with the -509C/869T haplotype (adjusted OR, 0.75; 95% CI, 0.57-0.98; P = 0.04; and adjusted OR, 0.67; 95% CI, 0.47-0.96; P = 0.02; respectively). CONCLUSION: The TGF-beta1 -509C > T and 869T > C polymorphisms and their haplotypes may contribute to genetic susceptibility to AC and SM of the lung.

Adenocarcinoma↗

PubMed2006

Hydrophobically modified glycol chitosan nanoparticles as carriers for paclitaxel.

Self-assembled nanoparticles based on hydrophobically modified glycol chitosan (HGC) were prepared as a carrier for paclitaxel. HGC conjugates were prepared by chemically linking 5beta-cholanic acid to glycol chitosan chains using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide chemistry. In phosphate-buffered saline (PBS; pH 7.4), the synthesized HGC conjugates formed nano-sized particles with a diameter of 200 nm and exhibited high thermodynamic stability as reflected by their low critical aggregation concentration (0.03 mg/ml). Paclitaxel was efficiently loaded into HGC nanoparticles up to 10 wt.% using a dialysis method. The paclitaxel-loaded HGC (PTX-HGC) nanoparticles were 400 nm in diameter and were stable in PBS for 10 days. These PTX-HGC nanoparticles also showed sustained release of the incorporated of paclitaxel (80% of the loaded dose was released in 8 days at 37 degrees C in PBS). Owing to sustained release, the PTX-HGC nanoparticles were less cytotoxic to B16F10 melanoma cells than free paclitaxel formulated in Cremophor EL. Injection of PTX-HGC nanoparticles into the tail vein of tumor-bearing mice prevented increases in tumor volume for 8 days. Finally, PTX was less toxic to the tumor-bearing mice when formulated in HGC nanoparticles than when formulated with Cremophor EL.

Animals↗

PubMed2006

Self-assembled nanoparticles based on glycol chitosan bearing hydrophobic moieties as carriers for doxorubicin: in vivo biodistribution and anti-tumor activity.

Self-assembled nanoparticles, formed by polymeric amphiphiles, have been demonstrated to accumulate in solid tumors by the enhanced permeability and retention effect, following intravenous administration. In this study, hydrophobically modified glycol chitosans capable of forming nano-sized self-aggregates were prepared by chemical conjugation of fluorescein isothiocyanate or doxorubicin to the backbone of glycol chitosan. Biodistribution of self-aggregates (300 nm in diameter) was evaluated using tissues obtained from tumor-bearing mice, to which self-aggregates were systemically administered via the tail vein. Irrespective of the dose, a negligible quantity of self-aggregates was found in heart and lung, whereas a small amount (3.6-3.8% of dose) was detected in liver for 3 days after intravenous injection of self-aggregates. The distributed amount of self-aggregates gradually increased in tumor as blood circulation time increased. The concentration of self-aggregates in blood was as high as 14% of dose at 1 day after intravenous injection and was still higher than 8% even at 3 days. When self-aggregates loaded with doxorubicin were administered into the tumor-bearing mice via the tail vein, they exhibited lower toxicity than but comparable anti-tumor activity to free doxorubicin. These results revealed the promising potential of self-aggregates on the basis of glycol chitosan as a carrier for hydrophobic anti-tumor agents.

Animals↗

PubMed2006

Antimetastatic activity of insulin-like growth factor binding protein-3 in lung cancer is mediated by insulin-like growth factor-independent urokinase-type plasminogen activator inhibition.

Insulin-like growth factor binding protein-3 (IGFBP-3), a major IGF-binding protein in human serum, regulates the growth of non-small cell lung cancer (NSCLC) cells through IGF-dependent and IGF-independent mechanisms. However, the role of IGFBP-3 in lung cancer metastasis is not well known. In the present study, we showed that noncytotoxic doses of adenoviral or recombinant IGFBP-3 significantly decreased the migration and invasion of H1299 and A549 NSCLC cells. Furthermore, treatment of human lung fibroblasts with recombinant IGFBP-3 suppressed their ability to stimulate the invasion of H1299 cells. Overexpression of IGFBP-3 markedly reduced lung metastasis of A549 cells in an experimental animal model system and prolonged the survival time of the animals. Urokinase-type plasminogen activator (uPA) inhibitor treatment or uPA small interfering RNA transfection of A549 and H1299 cells resulted in a significant decrease in invasion. Corresponding ELISA, Western blot, gelatin zymogram, and semiquantitative reverse transcription-PCR analyses revealed that IGFBP-3 reduced the expression of uPA mRNA through IGF-independent mechanisms. The specific role of uPA in anti-invasive activity of IGFBP-3 was further confirmed in NSCLC cells, in which uPA expression/activity was suppressed by the transfection with synthetic small interfering RNA or by the treatment with uPA inhibitor or induced by the infection with an adenoviral vector. IGFBP-3 also decreased the expression/activity of matrix metalloproteinase-2 through IGF-dependent but uPA-independent pathways. Taken together, our data suggest that IGFPB-3 effectively block uPA- and matrix metalloproteinase-2-stimulated invasion pathways, ultimately reducing lung cancer cell metastasis. Our findings indicate that IGFBP-3 may be a promising anti-invasive and antimetastatic therapeutic agent in lung cancer.

Animals↗

PubMed2005

Regulation of tumor angiogenesis by fastatin, the fourth FAS1 domain of betaig-h3, via alphavbeta3 integrin.

We previously reported that the FAS1 domains of betaig-h3 bear motifs that mediate endothelial cell adhesion and migration via interactions with alphavbeta3 integrin and regulate angiogenesis. In the present study, we show that the fourth FAS1 domain, designated fastatin, inhibits endothelial adhesion and migration, not only to betaig-h3, but also fibronectin and vitronectin, in a RGD-dependent manner. Fastatin and other FAS1 domains suppress endothelial cell tube formation and in vivo neovascularization in a Matrigel plug assay. The antiangiogenic activity of fastatin is associated with antitumor activity in mouse tumor models. Fastatin additionally induces apoptosis in several cells expressing alphavbeta3 integrin, including endothelial cells. Binding of fastatin to alphavbeta3 integrin inhibits phosphorylation of focal adhesion kinase, Raf, extracellular signal-regulated kinase, Akt, and mammalian target of rapamycin. Fastatin is thus the first endogenous angiogenesis regulator identified that inhibits both endothelial cell migration and growth by binding to alphavbeta3 integrin. Our data suggest that FAS1 domains from all possible forms of the four human FAS1 family proteins are potential endogenous regulators for pathologic angiogenesis. Moreover, FAS1 domains such as fastatin may be developed into drugs for blocking tumor angiogenesis.

Animals↗

PubMed2005

Methyl-CpG binding domain 1 gene polymorphisms and risk of primary lung cancer.

The methyl-CpG binding domain 1 (MBD1) protein plays an important role for transcriptional regulation of gene expression. Polymorphisms and haplotypes of the MBD1 gene may have an influence on MBD1 activity on gene expression profiles, thereby modulating an individual's susceptibility to lung cancer. To test this hypothesis, we investigated the association of MBD1 -634G>A, -501delT (-501 T/T, T/-, -/-), and Pro(401)Ala genotypes and their haplotypes with the risk of lung cancer in a Korean population. The MBD1 genotype was determined in 432 lung cancer patients and in 432 healthy control subjects who were frequency matched for age and gender. The -634GG genotype was associated with a significantly increased risk of overall lung cancer compared with the -634AA genotype [adjusted odds ratio (OR), 3.10; 95% confidence interval (95% CI), 1.24-7.75; P = 0.016]. When analyses were stratified according to the tumor histology, the -634GG genotype was associated with a significantly increased risk of adenocarcinoma compared with the -634AA genotype (adjusted OR, 4.72; 95% CI, 1.61-13.82; P = 0.005). For the MBD1 -501delT and Pro(401)Ala polymorphisms, the -501 T/T genotype was associated with a marginal significantly increased risk of adenocarcinoma compared with the -501(-/-) genotype (adjusted OR, 2.07; 95% CI, 1.02-4.20; P = 0.045), and the Pro/Pro genotype was associated with a significantly increased risk of adenocarcinoma compared with the Ala/Ala genotype (adjusted OR, 3.41; 95% CI, 1.21-9.60; P = 0.02). Consistent with the genotyping analyses, the -634G/-501T/(401)Pro haplotype was associated with a significantly increased risk of overall lung cancer and adenocarcinoma compared with the -634A/-501(-)/(401)Ala haplotype (adjusted OR, 1.44; 95% CI, 1.08-1.91; P = 0.012 and P(c) = 0.048; adjusted OR, 1.75; 95% CI, 1.20-2.56; P = 0.004 and P(c) = 0.016, respectively). On a promoter assay, the -634A allele had significantly higher promoter activity compared with the -634G allele in the Chinese hamster ovary cells and A549 cells (P < 0.05 and P < 0.001, respectively), but the -501delT polymorphism did not have an effect on the promoter activity. When comparing the promoter activity of the MBD1 haplotypes, the -634A/-501(-) haplotype had a significantly higher promoter activity than the -634G/-501T haplotype (P < 0.001). These results suggest that the MBD1 -634G>A, -501delT, and Pro(401)Ala polymorphisms and their haplotypes contribute to the genetic susceptibility for lung cancer and particularly for adenocarcinoma.

Adenocarcinoma↗

PubMed2004

Beta ig-h3 promotes renal proximal tubular epithelial cell adhesion, migration and proliferation through the interaction with alpha3beta1 integrin.

Betaig-h3 (betaig-h3) is a secretory protein composed of fasciclin I-like repeats containing sequences that allows binding of integrins and glycosaminoglycans in vivo. Expression of betaig-h3 is responsive to TGF-Beta and the protein is found to be associated with extracellular matrix (ECM) molecules, implicating betaig-h3 as an ECM adhesive protein of developmental processes. We previously observed predominant expression of betaig-h3 expression in the basement membrane of proximal tubules of kidney. In this study, the physiological relevance of such localized expression of betaig-h3 was examined in the renal proximal tubular epithelial cells (RPTEC). RPTEC constitutively expressed betaig-h3 and the expression was dramatically induced by exogenous TGF-Beta1 treatment. betaig-h3 and its second and fourth FAS1 domain were able to mediate RPTEC adhesion, spreading and migration. Two known alpha3beta1 integrin-interaction motifs including aspartatic acid and isoleucine residues, NKDIL and EPDIM in betaig-h3 were responsible to mediate RPTEC adhesion, spreading, and migration. By using specific antibodies against integrins, we confirmed that alpha3beta1 integrin mediates the adhesion and migration of RPTECs on betaig-h3. In addition, it also enhanced proliferation of RPTECs through NKDIL and EPDIM. These results indicate that betaig-h3 mediates adhesion, spreading, migration and proliferation of RPTECs through the interaction with alpha3beta1 integrin and is intimately involved in the maintenance and the regeneration of renal proximal tubular epithelium.

Amino Acid Motifs↗

PubMed2004

Gene expression profile of human chondrocyte HCS-2/8 cell line by EST sequencing analysis.

Large-scale single-pass sequencing of randomly selected cDNA clones from cell type specific libraries has proven to be a powerful approach for the discovery of novel gene functions, identification of novel gene family members, and definition of gene expression profiles. HCS-2/8 chondrocyte has been used as a cell culture model to study chondrocyte differentiation. Here we performed 3350 single-pass sequencing reactions obtained from the 5' ends of cDNAs from HCS-2/8 cells. To define the expression profiles of HCS-2/8 chondrocytes, we analyzed the identity of these representative cDNA sequences using database searches (BLAST). The sequences represent 1927 unique genes with known function (i.e., unigene clusters), 38 transcripts that are similar to genes with known function, 739 expressed genes with unknown function (i.e., expressed sequence tags), and 18 cDNAs which have not previously been sequenced. Interestingly, many transcripts were expressed from chromosome 12 compared with total genes, while the fewer numbers of cDNAs were derived from genes on chromosomes 14, 18 and Y. The chondrocytic phenotype of HCS-2/8 cells is reflected by abundant expression of genes related to cell structure and motility and the 20 most frequently expressed unigenes reflect a chondrocyte-related gene expression signature. Thus, our data establish a representative set of more than 2000 genes expressed in a chondrocytic cell line. This finding provides a framework for understanding cell growth and differentiation of chondrocytes and their metabolic function in the formation and remodeling of cartilage.

Cell Line, Tumor↗

PubMed2004

Self-assembled nanoparticles based on glycol chitosan bearing 5beta-cholanic acid for RGD peptide delivery.

The synthetic peptide bearing Arg-Gly-Asp (RGD) sequence is considered to specifically bind to alpha(v)beta(3) integrin expressed on endothelial cells in the angiogenic blood vessels, which provides a potential to inhibit the tumor growth. As a carrier for the RGD peptide, hydrophobically modified glycol chitosan (HGC) capable of forming nano-sized self-aggregates was prepared by the chemical conjugation of 5beta-cholanic acid to the main backbone of glycol chitosan. The RGD peptide labeled with fluoresein isothiocyanate (FITC-GRGDS) was loaded into self-aggregates in three different conditions: simple mixing, sonication, and solvent evaporation methods. Of different methods applied, solvent evaporation method showed the most promising results for peptide loading, as judged by the yield (>70%) and loading efficiency (>75%). It was found that the presence of FITC-labeled peptides makes the self-aggregates to be compact, possibly due to the role of both hydrophobic FITC and peptides containing carboxylic acids that allow hydrogen bonding and electrostatic interaction with the primary amino groups in the main backbone of glycol chitosan. FITC-labeled peptides were released from self-aggregates in a physiological solution (pH 7.4) for up to 1 day. From the cell adhesion and migration assays, it was demonstrated that FITC labeling of peptides does not significantly deteriorate biological activity of the parent peptide drug (GRGDS). Overall, the self-aggregates loaded with FITC-GRGDS might be useful for monitoring or destroying the angiogenic vessels surrounding the tumor tissue.

Amino Acid Sequence↗

PubMed2003

Differential gene expression analysis using paraffin-embedded tissues after laser microdissection.

Recent advances in laser microdissection allow for precise removal of pure cell populations from morphologically preserved tissue sections. However, RNA from paraffin-embedded samples is usually degraded during microdissection. The purpose of this study is to determine the optimal fixative for RNA extractions from laser microdissected paraffin-embedded samples. The integrity of RNA was evaluated with the intactness of 18S and 28S ribosomal RNA by electrophoresis and by the length of individual gene transcripts using RT-PCR. The various fixatives were methacarn (a combination of methanol, chloroform, and acetic acid) and several concentrations of ethanol and isopropanol. Methacarn was the optimal fixative for RNA preservation in paraffin-embedded tissues, which included liver, lung, kidney, muscle, and limb. Based on RT-PCR analysis, methacarn fixed samples exhibited the expected RNA sizes for individual genes such as glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) and bone-related genes (e.g., alkaline phosphatase and osteonectin). The laser microdissection technique with methacarn fixation was then applied to analyze the differential gene expression between hypertrophic and proliferative chondrocytes in the growth plate of long bone. The expression of type X collagen, a specific gene for hypertrophic chondrocytes, was only observed in hypertrophic chondrocytes, while type II collagen was observed more broadly in the growth plate as anticipated. Thus, combining laser microdissection with methacarn fixation facilitates the examination of differentially expressed genes from various tissues.

Acetic Acid↗

PubMed2003

Identification of the alphavbeta3 integrin-interacting motif of betaig-h3 and its anti-angiogenic effect.

betaig-h3 is an extracellular matrix protein that mediates adhesion and migration of several cell types through interaction with integrins. In the present study, we tested whether betaig-h3 mediates endothelial cell adhesion and migration, thereby regulating angiogenesis. In this study, we demonstrate that not only betaig-h3 itself but also all four fas-1 domains of betaig-h3 mediate endothelial cell adhesion and migration through interaction with the alphavbeta3 integrin. We found that the alphavbeta3 integrin-interacting motif of the four fas-1 domains of betaig-h3 is the same YH motif that we reported previously to interact with alphavbeta5 integrin. The YH peptide inhibited endothelial cell adhesion and migration in a dose-dependent manner. We demonstrate that the YH peptide has anti-angiogenic activity in vitro and in vivo using an endothelial cell tube formation assay and a Matrigel plug assay, respectively. Our results reveal that betaig-h3 bears alphavbeta3 integrin-interacting motifs that mediate endothelial cell adhesion and migration and, therefore, may regulate angiogenesis.

Amino Acid Motifs↗

PubMed2003

RGD peptides released from beta ig-h3, a TGF-beta-induced cell-adhesive molecule, mediate apoptosis.

Beta ig-h3 is a transforming growth factor-beta (TGF-beta)-induced cell-adhesive molecule and has an RGD sequence at its C-terminus. A previous report suggested that beta ig-h3 normally undergoes carboxy-terminal processing that results in the loss of the RGD sequence. RGD peptides appear to play various roles in cell function. Here we show that the RGD peptides released from beta ig-h3 may facilitate TGF-beta-induced apoptosis. We found that carboxy-terminal cleavage of beta ig-h3 occurred after its secretion, and that overexpression of the wild-type beta ig-h3 induced apoptosis, unlike the C-terminal deleted but RGD-containing mutant beta ig-h3, which is resistant to C-terminal processing. The beta ig-h3-induced apoptosis was abolished by either deletion of the RGD sequence or mutation of RGD to RAE. Synthetic peptides of ERGDEL and GRGDSP derived from beta ig-h3 and fibronectin, respectively, also induced apoptosis, unlike ERGEEL and GRGESP. Culture supernatants of cells overexpressing beta ig-h3 filtered to isolate molecules smaller than 3 kDa also induced apoptosis. A fusion protein composed of the N-terminal 100 amino acids of fibronectin and the RGD-containing C-terminal part of beta ig-h3 was also subjected to C-terminal cleavage and overexpression resulted in apoptosis. The anti-beta ig-h3 antibody blocks TGF-beta-induced apoptosis. Thus, beta ig-h3 may be important in regulating cell apoptosis by providing soluble RGD peptides.

Amino Acid Motifs↗

PubMed2003

Expression of TGF-beta-induced matrix protein betaig-h3 is up-regulated in the diabetic rat kidney and human proximal tubular epithelial cells treated with high glucose.

BACKGROUND: betaig-h3 is an extracellular matrix protein whose expression in several cell types is greatly increased by transforming growth factor-beta (TGF-beta). TGF-beta is believed to be involved in the development of diabetic nephropathy and thus we have assessed the possibility that betaig-h3 may be a downstream molecule in this pathogenic process. METHODS: Immunoblotting and immunohistochemistry were done using an antibody against mouse betaig-h3 protein. betaig-h3 and TGF-beta concentrations were measured by enzyme-linked immunosorbent assay (ELISA). Cell adhesion and migration were assessed by measuring activity of N-acetyl-beta-d-glucosaminidase and using a transwell plate, respectively. RESULTS: Immunohistochemistry revealed that betaig-h3 occurs mainly in the basement membrane of proximal tubules, particularly the S3 segment but also to lesser extents in the basement membranes of the cortical thick ascending limb cells and the parietal glomerular epithelial cells in Bowman's capsule. Immunoblotting revealed that approximately 68 kD bands were seen only in the cortex + the outer stripe of the outer medulla. Rats with streptozotocin (STZ)-induced diabetes exhibited a marked and sustained increase in renal betaig-h3 abundance. This was mirrored by urinary betaig-h3 levels. In vitro experiments with human primary renal proximal tubular epithelial cells revealed that their expression of betaig-h3 was greatly increased by either TGF-beta or glucose. High glucose levels also stimulated TGF-beta production by renal proximal tubular epithelial cells and the high glucose-induced betaig-h3 expression was almost completely blocked by anti-TGF-beta antibody. betaig-h3 mediated renal proximal tubular epithelial cells adhesion and migration. CONCLUSION: betaig-h3 may be important in the development of diabetic nephropathy. Furthermore, the level of urinary betaig-h3 may be useful as an early marker reflecting disease onset and progression.

Animals↗

PubMed2002

Identification of motifs in the fasciclin domains of the transforming growth factor-beta-induced matrix protein betaig-h3 that interact with the alphavbeta5 integrin.

betaig-h3 is a TGF-beta-induced matrix protein known to mediate the adhesion of several cell types. In this study, we found that all four of the fas-1 domains in betaig-h3 mediate MRC-5 fibroblast adhesion and that this was specifically inhibited by a function-blocking monoclonal antibody specific for the alphavbeta5 integrin. Using deletion mutants of the fourth fas-1 domain revealed the MRC-5 cell adhesion motif (denoted the YH motif) is located in amino acids 548-614. Experiments with substitution mutants showed that tyrosine 571, histidine 572, and their flanking leucine and isoleucine amino acids, which are all highly conserved in many fas-1 domains, are essential for mediating MRC-5 cell adhesion. A synthetic 18-amino acid peptide encompassing these conserved amino acids could effectively block MRC-5 cell adhesion to betaig-h3. Using HEK293 cells stably transfected with the beta5 integrin cDNA, we confirmed that the alphavbeta5 integrin is a functional receptor for the YH motif. In conclusion, we have identified a new alphavbeta5 integrin-interacting motif that is highly conserved in the fas-1 domains of many proteins. This suggests that fas-1 domain-containing proteins may perform their biological functions by interacting with integrins.

Amino Acid Motifs↗