Growth Factor-beta Receptor Type Research Articles

Transforming growth factor-beta receptor type 1 (TGFBR1) is not associated with non-syndromic cleft lip with or without cleft palate in patients of Central European descent

Objective Transforming growth factor-beta (TGF-β) type 1 receptor (also known as activin receptor-like kinase 5, ALK5) is expressed in palatal tissue during embryogenesis. Experimental studies in transgenic mice with a genetic deletion of Alk5 showed that TGF-β type 1 receptor is required for upper lip and midline fusion of the hard and soft palate. In humans, association of TGF-β type 1 receptor gene ( TGFBR1) and the development of non-syndromic cleft lip with or without cleft palate (NSCL/P) had been observed in a multiethnic sample of Chinese, Philippine, Indian and Turkish families. In order to re-evaluate the relevance of these findings, we carried out a family-based association study among 218 NSCL/P families of Central European descent. Methods Genomic DNA was obtained from peripheral blood of 218 complete parent–offspring triads with NSCL/P. The sample comprised 14 patients with cleft lip only (CLO) and 204 patients with cleft lip and palate (CLP). Genotyping and transmission disequilibrium test (TDT) were performed on all 218 triads with a total of 17 tagging single-nucleotide polymorphisms (SNPs). We also performed testing for extended haplotypes and a log-linear model by Weinberg was used to screen parent-of-origin effects. Furthermore the use of estimates for the relative risks (RR) of Weinberg's model was obtained. Results TDT analysis revealed no significant transmission distortion, neither at the level of individual markers nor at the level of haplotypes. Similarly negative results were obtained when we restricted our analysis to the subgroup of patients with CLP ( n = 204). Relative risk calculations (RR) of the children's and mothers’ genotypes obtained negative results, after correction of p-values for multiple testing. Likewise application of Weinberg's log-linear model did not find any evidence for parent-of-origin effects in our sample. Conclusion Despite the ample evidence supporting the role of TGF-β type 1 receptor as a critically important and widespread morphogenetic regulator of craniofacial development in murine models, our results do not support TGFBR1 as major risk factor for NSCL/P in patients of Central European descent.

TGFBR1 variants TGFBR1*6A and Int7G24A are not associated with an increased familial colorectal cancer risk

Variants of the transforming growth factor-beta receptor type 1 (TGFBR1) gene, TGFBR1*6A and Int7G24A, have been suggested to act as low-penetrance tumour susceptibility alleles with TGFBR1*6A being causally responsible for some cases of familial colorectal cancer (CRC). We performed a case–control study of 262 unrelated familial CRC cases; 83 hereditary non-polyposis colorectal cancer (HNPCC) and 179 non-HNPCC. Patients were genotyped for TGFBR1*6A and Int7G24A and compared with 856 controls. Further, we screened the coding region of TGFBR1 in affected members of a large family with CRC linked to 9q22.32-31.1. TGFBR1*6A allelic frequency was not significantly different in all of the familial cases compared with controls (0.107 and 0.106, respectively; P=0.915). In a subgroup analysis allele frequencies were, however, different between HNPCC and non-HNPCC familial cases (0.157 and 0.084, respectively; P=0.013). TGFBR1*6A genotype did not influence age of onset. Int7G24A allele frequencies were similar in cases and controls. No germ-line mutation was identified in the family with CRC linked to this chromosomal region. Our study provides no substantial support for the hypothesis that the polymorphic variants TGFBR1*6A or Int7G24A contribute to familial CRC risk. We cannot, however, exclude the possibility that TGFBR1 variants have a modifying effect on inherited risk per se.

Mechanisms of decreased expression of transforming growth factor-beta receptor type I at late stages of HPV16-mediated transformation

Transforming growth factor-beta (TGF-β) signaling is disrupted in many cancers, including cervical cancer, leading to TGF-β resistance. Although initially sensitive, human papillomavirus type 16 (HPV16) immortalized human keratinocytes (HKc/HPV16) become increasingly resistant to the growth inhibitory effects of TGF-β during in vitro progression to a differentiation resistant phenotype (HKc/DR). We have previously shown that loss of TGF-β sensitivity in HKc/DR is attributed to decreased expression of TGF-β receptor type I (TGF-β RI), while the levels of TGF-β receptor type II (TGF-β RII) remain unchanged. The present study explored molecular mechanisms leading to reduced TGF-β RI expression in HKc/DR. Using TGF-β RI and TGF-β RII promoter reporter constructs, we determined that acute expression of the HPV16 oncogenes E6 and E7 decreased the promoter activity of TGF-β RI and TGF-β RII by about 50%. However, promoter activity of TGF-β RI is decreased to a greater extent than TGF-β RII as HKc/HPV16 progress to HKc/DR. Reduced TGF-β RI expression in HKc/DR was found not to be linked to mutations within the TGF-β RI promoter or to promoter methylation. Electrophoretic mobility shift and supershift assays using probes encompassing Sp1 binding sites in the TGF-β RI promoter found no changes between HKc/HPV16 and HKc/DR in binding of the transcription factors Sp1 or Sp3 to the probes. Also, Western blots determined that protein levels of Sp1 and Sp3 remain relatively unchanged between HKc/HPV16 and HKc/DR. Overall, these results demonstrate that mutations in or hypermethylation of the TGF-β RI promoter, along with altered levels of Sp1 or Sp3, are not responsible for the reduced expression of TGF-β RI we observe in HKc/DR. Rather the HPV16 oncogenes E6 and E7 themselves exhibit an inhibitory effect on TGF-β receptor promoter activity.

Intrahepatic Bile Ducts Develop According to a New Mode of Tubulogenesis Regulated by the Transcription Factor SOX9

A number of diseases are characterized by defective formation of the intrahepatic bile ducts. In the embryo, hepatoblasts differentiate to cholangiocytes, which give rise to the bile ducts. Here, we investigated duct development in mouse liver and characterized the role of the SRY-related HMG box transcription factor 9 (SOX9). We identified SOX9 as a new biliary marker and used it in immunostaining experiments to characterize bile duct morphogenesis. The expression of growth factors was determined by in situ hybridization and immunostaining, and their role was studied on cultured hepatoblasts. SOX9 function was investigated by phenotyping mice with a liver-specific inactivation of Sox9. Biliary tubulogenesis started with formation of asymmetrical ductal structures, lined on the portal side by cholangiocytes and on the parenchymal side by hepatoblasts. When the ducts grew from the hilum to the periphery, the hepatoblasts lining the asymmetrical structures differentiated to cholangiocytes, thereby allowing formation of symmetrical ducts lined only by cholangiocytes. We also provide evidence that transforming growth factor-beta promotes differentiation of the hepatoblasts lining the asymmetrical structures. In the absence of SOX9, the maturation of asymmetrical structures into symmetrical ducts was delayed. This was associated with abnormal expression of CCAAT/Enhancer Binding Protein alpha and Homolog of Hairy/Enhancer of Split-1, as well as of the transforming growth factor-beta receptor type II, which are regulators of biliary development. Our results suggest that biliary development proceeds according to a new mode of tubulogenesis characterized by transient asymmetry and whose timing is controlled by SOX9.

TGF‐β and smad signalling in fibrosis

TGF‐β and smad signalling in fibrosis

Absence of TGFBR1 and TGFBR2 Mutations in Patients With Bicuspid Aortic Valve and Aortic Dilation

Mutations in the genes encoding transforming growth factor-beta receptor types I and II (TGFBR1 and TGFBR2, respectively) are commonly identified in patients with Loeys-Dietz syndrome, as well as some patients with Marfan's syndrome or familial thoracic aortic aneurysms and dissections. This suggests that there is considerable phenotypic heterogeneity associated with mutations in these genes. Because bicuspid aortic valve (BAV) is a congenital heart defect in patients with Loeys-Dietz syndrome, this study was conducted to investigate whether variants in TGFBR1 or TGFBR2 are responsible for sporadic BAV. Analysis of these genes in 35 patients with BAVs identified only known single-nucleotide polymorphisms or novel synonymous or intronic substitutions. In conclusion, mutations in TGFBR1 and TGFBR2 rarely cause sporadic BAV.

Ulcerative Colitis-Associated Colorectal Cancer is Frequently Associated with the Microsatellite Instability Pathway

Patients with ulcerative colitis have a high risk for the development of colorectal cancer. To understand the molecular mechanisms of the carcinogenesis process of ulcerative colitis-associated colorectal cancer, the genetic alterations in inflamed or neoplastic colon epithelium in ulcerative colitis were analyzed. Fifty-seven patients with ulcerative colitis were enrolled in this study. Specimens were obtained from the patients randomly at six colonic sites. Each patient was histologically classified according to the worst pathologic finding into cancer, dysplasia, indefinite, and normal cases. Microsatellite instability, mutations of target genes, hypermethylation of the hMLH1 promoter region, and mismatch repair protein expression were analyzed. High-microsatellite instability was found in 4 of 11 cancer cases (36 percent), 5 of 15 dysplasia cases (33 percent), 5 of 11 indefinite cases (45 percent), and none of 20 normal cases (0 percent). A significant correlation was found between the malignant potential and high-microsatellite instability. A frameshift mutation of transforming growth factor beta receptor Type II (TGFbetaRII) was significantly correlated with worsening histologic grade. High-microsatellite instability was significantly associated with hMLH1 hypermethylation and loss of hMSH2 expression. The carcinogenesis process in ulcerative colitis-associated colorectal cancer was closely associated with the microsatellite instability pathway through TGFbetaRII mutation by a dysfunction of the mismatch repair system.

Growth inhibition induced by transforming growth factor-β1 in human oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is a world-wide health problem and its incidence accounts for 1.9-3.5% of all malignant tumors. Transforming growth factor beta/Smads (TGF-beta/Smads) signaling pathway plays an important role in oncogenesis, but its function and molecular mechanisms in OSCC remain unclear. Expression of transforming growth factor-beta receptor type II (TbetaRII) and Smad4 was studied by immunohistochemistry in 108 OSCC patients and 10 normal controls. Function and molecular mechanisms of TGF-beta/Smads signaling pathway was then investigated in two human tongue squamous carcinoma cell lines with high and low metastasis (Tb and Tca8113) by RT-PCR, Western Blot, immunofluorescence, cell growth curve and flow cytometry (FCM), respectively. TbetaRII and Smad4 were significantly down-regulated in tumor tissues (with or without lymph node metastasis) compared to normal oral epithelium tissues (P < 0.05). TGF-beta1 induced arrest of the cell cycle rather than cell death in Tca8113 and Tb cells, and this influence was mediated by the increasing the expression and changing the location of its downstream components of TGF-beta1/Smads signaling pathway. TGF-beta1 rapidly increased the expression of p15 and p21 in both Tca8113 and Tb cells. TGF-beta1 did not increase p27 expression in Tca8113 cells, but p27 expression was increased in Tb cells. These indicated that TGF-beta1 induced G(1) arrest of cell cycle through a different regulating pathway in Tb cells compared with Tca8113 cells. Thus, we conclude that TGF-beta/Smads signaling pathway play a important role on cell growth and metastasis potential in OSCC.

Adoptive transfer of CD8+ T cells from transforming growth factor beta receptor type II (dominant negative form) induces autoimmune cholangitis in mice

We recently reported that mice with a T cell-restricted expression of a dominant negative form of transforming growth factor beta receptor type II (dnTGFbetaRII) spontaneously develop autoimmune cholangitis that resembles human primary biliary cirrhosis (PBC), including antimitochondrial antibodies (AMAs) and extensive portal CD4(+) and CD8(+) lymphocytic infiltrates. On the basis of these data, we performed a series of experiments to determine whether the pathology was secondary to direct dnTGFbetaRII disruption of the liver and/or alternatively the appearance of autoreactive T cells. First, using dnTGFbetaRIIRag1(-/-) mice, we noted a normal hepatic and biliary structure. Hence, we performed a rigorous series of adoptive transfer studies, transferring Ly5.1(+) unfractionated spleen cell CD4(+) or CD8(+) T cells from dnTGFbetaRII mice into B6/Rag(-/-) (Ly 5.2) recipients. In unmanipulated dnTGFbetaRII mice, there was a marked increase in CD4(+) and CD8(+) T cell biliary infiltrates with AMA. Indeed, B6/Rag(-/-) recipients of dnTGFbetaRII unfractionated cells develop features of liver disease similar to PBC, suggesting that splenic loss of self-tolerance alone is sufficient to cause disease in this model and therefore that there is no specific abnormality in the biliary targets required for appearance of disease. More importantly, adoptive transfer of CD8(+) but not CD4(+) T cells into B6/Rag(-/-) mice led to liver histopathology remarkably similar to PBC, emphasizing a prominent role for CD8 T cell-mediated pathogenesis. In contrast, B6/Rag(-/-) recipients of CD4(+) T cells from dnTGFbetaRII mice predominantly developed inflammatory bowel disease associated with higher levels of serum interferon gamma and tumor necrosis factor alpha. These data suggest that in this model of PBC, autoreactive CD8(+) cells destroy bile ducts.

Mutational inactivation of TGFBR2 in microsatellite unstable colon cancer arises from the cooperation of genomic instability and the clonal outgrowth of transforming growth factor β resistant cells

The mutational inactivation of transforming growth factor beta receptor type II (TGFBR2) occurs in approximately 30% of colon cancers and promotes the formation of colon cancer by inhibiting the tumor suppressor activity of the TGFB signaling pathway. TGFBR2 mutations occur in >90% of microsatellite unstable (MSI) colon cancers and affect a polyadenine tract in exon 3 of TGFBR2, called BAT-RII, which is vulnerable to mutation in the setting of DNA mismatch repair (MMR) system deficiency. In light of the vulnerable nature of the BAT-RII tract in the setting of MMR inactivation and the favorable effects of TGFBR2 inactivation in colon cancer, analysis of TGFBR2 inactivation provides an opportunity to assess the roles of genomic instability vs. clonal selection in cells acquiring TGFBR2 BAT-RII tract mutations in MSI colon cancer formation. The contribution of genomic instability and/or clonal evolution to the mutational inactivation of TGBFR2 in MSI colon cancers has not been studied in a systematic way that would allow a determination of the relative contribution of these two mechanisms in the formation of MSI colon cancer. It has not been demonstrated whether the BAT-RII tract mutations are strictly a consequence of the BAT-RII region being hypermutable in the setting of MMR deficiency or if the mutations are rather a consequence of clonal selection pressure against the TGFB receptor. Through the use of defined cell line systems, we show that both genomic instability and clonal selection of TGFB resistant cells contribute to the high frequency of TGFBR2 mutations in MSI colon cancer.

Genetic testing in patients with aortic aneurysms/dissections: a novel genotype/phenotype correlation?☆

Mutations in the genes encoding fibrillin-1 (FBN1) and transforming growth factor beta receptor type II (TGFBR2) are known causes of Marfan syndrome (MFS) and related disorders. However, a sound correlation between the genotype and the cardiovascular phenotype has not yet been established. The objective of the present study was to identify novel mutations in FBN1 and TGFBR2 and to assess whether the type of mutation is linked to a particular clinical subtype of the cardiovascular condition. The clinical records of 36 patients referred to us for molecular genetic diagnosis were reviewed to assess the course and severity of the vascular deterioration. A semiautomatic protocol was established enabling a rapid and cost-effective screening of the genes FBN1 and TGFBR2 by direct sequencing of all coding exons and flanking intronic regions. Novel mutations in FBN1 and TGFBR2 were detected in 12 and 2 patients, respectively. Four individuals carried a recurrent mutation in FBN1. Throughout the study cohort, the incidence of aortic dissections per se did not depend on the type of mutation. However, we found that mutations affecting the calcium-binding epidermal growth factor-like domain were more frequently associated with a dissection of distal parts of the aorta than mutations that lead to a premature termination codon (chi(1)(2): p=0.013), suggesting that the spatio-temporal pattern of vascular deterioration may vary with the type of mutation. Detecting a mutation in the genes FBN1 and TGFBR2 proves the genetic origin of vascular findings and allows the identification of family members at risk who should undergo preventive checkups. Routine genetic testing of patients with suspected MFS or thoracic aortic aneurysms/dissections could provide further insight into genotype/phenotype correlations related to aortic dissection.

1256: Lower Tumor Expression Levels of the Transforming Growth Factor Beta Receptor Type II Protein are Associated with a Less Aggressive Tumor Phenotype and Improved Survival Among Patients with Clear Cell Renal Cell Carcinoma

You have accessJournal of UrologyModerated Poster 37, Tuesday, May 22, 2007, 8:00 - 10:00 am1 Apr 20071256: Lower Tumor Expression Levels of the Transforming Growth Factor Beta Receptor Type II Protein are Associated with a Less Aggressive Tumor Phenotype and Improved Survival Among Patients with Clear Cell Renal Cell Carcinoma Alexander S. Parker, Christine M. Lohse, Kevin Wu, John Copland, Michael Wehle, and John C. Cheville Alexander S. ParkerAlexander S. Parker More articles by this author , Christine M. LohseChristine M. Lohse More articles by this author , Kevin WuKevin Wu More articles by this author , John CoplandJohn Copland More articles by this author , Michael WehleMichael Wehle More articles by this author , and John C. ChevilleJohn C. Cheville More articles by this author View All Author Informationhttps://doi.org/10.1016/S0022-5347(18)31470-8AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail "1256: Lower Tumor Expression Levels of the Transforming Growth Factor Beta Receptor Type II Protein are Associated with a Less Aggressive Tumor Phenotype and Improved Survival Among Patients with Clear Cell Renal Cell Carcinoma." The Journal of Urology, 177(4S), p. 414 © 2016 by American Urological AssociationFiguresReferencesRelatedDetails Volume 177Issue 4SApril 2007Page: 414 Advertisement Copyright & Permissions© 2016 by American Urological AssociationMetricsAuthor Information Alexander S. Parker More articles by this author Christine M. Lohse More articles by this author Kevin Wu More articles by this author John Copland More articles by this author Michael Wehle More articles by this author John C. Cheville More articles by this author Expand All Advertisement Loading ...

TGFBR2 mutation is correlated with CpG island methylator phenotype in microsatellite instability-high colorectal cancer

The transforming growth factor-beta receptor type 2 gene (TGFBR2) is mutated in most microsatellite instability-high (MSI-H) colorectal cancers. Promoter methylation of RUNX3 (runt-related transcription factor 3; encoding a transcription factor downstream of the TGF-beta pathway) is observed in colorectal cancer with CpG island methylator phenotype (CIMP), which is characterized by extensive promoter methylation and is associated with MSI-H and BRAF mutations. However, no study to date has examined interrelationship between TGFBR2 mutation, RUNX3 methylation, and CIMP in colorectal cancer. Using 144 MSI-H colorectal cancers derived from 2 large prospective cohort studies, we analyzed a mononucleotide repeat of TGFBR2 and quantified DNA methylation (by MethyLight technology) in 8 CIMP-specific promoters (RUNX3, CACNA1G [calcium channel, voltage-dependent, T type alpha-1G subunit], CDKN2A [p16], CRABP1 [cellular retinoic acid binding protein 1], IGF2 [insulin-like growth factor 2], MLH1, NEUROG1 [neurogenin 1], and SOCS1 [suppressor of cytokine signaling 1]). Among the 144 MSI-H tumors, the presence of TGFBR2 mutation (overall 72% frequency) was correlated positively with CIMP-high (with >/=6/8 methylated promoters; P < .0001), RUNX3 methylation (P = .0004), BRAF mutation (P = .0006), and right colon (P = .05); inversely with KRAS mutation (P = .006); but not significantly with sex, tumor differentiation, and p53 status (assessed by immunohistochemistry). After stratification by sex, location, tumor differentiation, RUNX3 status, KRAS/BRAF status, or p53 status, CIMP-high was persistently correlated with TGFBR2 mutation. In contrast, RUNX3, KRAS, or BRAF status was no longer correlated with TGFBR2 mutation after stratification by CIMP status. In conclusion, TGFBR2 mutation is associated with CIMP-high and indirectly with RUNX3 methylation. Our findings emphasize the importance of analyzing global epigenomic status (for which CIMP status is a surrogate marker) when correlating a single epigenetic event (eg, RUNX3 methylation) with any other molecular or clinicopathologic variables.

Lower expression levels of the transforming growth factor beta receptor type II protein are associated with a less aggressive tumor phenotype and improved survival among patients with clear cell renal cell carcinoma

Loss of expression of the transforming growth factor beta type II receptor (TbetaRII) has been implicated as an important event in renal carcinogenesis; however, its role as a potential prognostic factor remains poorly understood. Using archived tumor samples and long-term follow-up on a cohort of 280 clear cell renal cell carcinoma (ccRCC) patients treated with surgery from 1980 to 1998, we evaluated the association of TbetaRII expression and cancer-specific survival in both a univariate and multivariate setting. Low tumor expression of TbetaRII is associated with a less aggressive tumor phenotype at time of surgery. Moreover, those patients with lower levels of TbetaRII expression experience better cancer-specific survival than patients with higher levels of TbetaRII expression (log rank P = .034). Based on a Cox proportional hazard model adjusting for age, patients with tumors showing low (hazard ratio, 0.49; 95% confidence interval, 0.27-0.88) and moderate (hazard ratio, 0.7; 95% confidence interval, 0.40-1.23) TbetaRII expression are at decreased risk of RCC death compared with patients with tumors having high levels of TbetaRII expression. Adjustment for well-known pathologic predictors of RCC outcome attenuates the association of TbetaRII expression and ccRCC survival. Of interest, the association with TbetaRII expression appears more pronounced among those patients with tumors showing less aggressive phenotypes. Data from this investigation are the first to suggest that loss of TbetaRII expression is associated with improved ccRCC patient survival, especially among those patients with less aggressive disease profiles at time of surgery.

Familial thoracic aortic aneurysms and dissections: Three families with early‐onset ascending and descending aortic dissections in women

Ascending thoracic aortic aneurysms leading to type A dissections can be inherited in an autosomal dominant manner with variable age of onset and decreased penetrance, primarily in women. Three families are described with autosomal dominant inheritance of either ascending aortic aneurysms leading to type A dissections or type B dissections, and a young age of onset of aortic dissections in both men and women. Pedigree analysis suggests that a de novo mutation is responsible for the disease in one family. The discordant age of onset of aortic disease in a monozygotic twin pair in a different family indicates that environmental or stochastic factors may influence the variable expression of disease. Genetic analysis of one family excluded linkage to known loci for TAAD (TAAD1, TAAD2, FAA1, or FBN1) and sequence analysis failed to identify mutations in TGFBR2, the gene encoding transforming growth factor beta receptor type II. Thus, a novel unidentified loci may be responsible for the phenotype in these three families.

Mutational analysis of transforming growth factor-beta receptor type II and Smad3 tumor suppressor genes in prolactinomas

The presence of transforming growth factor-beta receptor type II and Smad3 gene abnormality was examined in human prolactinomas by single-strand conformation polymorphism screening for gene abnormality and band shift in tumor tissue and peripheral blood samples obtained from 14 patients treated for prolactinoma. Direct sequencing of exon 4 of the transforming growth factor-beta receptor type II gene, which showed a band shift by single-strand conformation polymorphism, disclosed a single nucleotide polymorphism (rs2228048) (alleles A/G; average allele frequency, G:0.817, A:0.183) and not an active mutation. Seven of the 14 cases showed base substitution (G --> A) of the single nucleotide polymorphism. Two of these 7 cases showed base substitution (G --> A) in the blood sample in accordance with neoplastic transformation. Direct sequencing of exon 2 of the Smad3 gene, which showed band shift by single-strand conformation polymorphism, disclosed single nucleotide polymorphism (rs1065080) (alleles C/T; average allele frequency, C:0.947, T:0.053) and no active mutation. Four of the 14 cases showed base substitution (C --> T) of the single nucleotide polymorphism. Blood samples were available for 2 of these 4 cases, and 1 of these 2 cases showed base substitution (C --> T) in accordance with neoplastic transformation. Estimation of the drug sensitivity of pituitary adenoma based on analysis of peripheral blood cells needs special care.

Contribution of activin receptor–like kinase 5 (transforming growth factor β receptor type I) signaling to the fibrotic phenotype of scleroderma fibroblasts

To use a specific transforming growth factor beta receptor type I (TGFbetaRI; activin receptor-like kinase 5 [ALK-5]) kinase inhibitor (SD208) to determine the role of activation of the TGFbetaRI kinase (ALK-5) in maintaining the profibrotic phenotype of dermal fibroblasts in systemic sclerosis (SSc). The effect of SD208 on the expression of key biochemical markers of the fibrotic phenotype was compared in fibroblasts cultured from clinically involved (lesional) and clinically uninvolved skin of patients with diffuse cutaneous SSc (dcSSc) and in fibroblasts from healthy controls matched for age, sex, and anatomic site. Protein expression was compared together with the ability of fibroblasts to adhere to the extracellular matrix and to remodel and contract a free-floating fibroblast-populated type I collagen lattice. Inhibiting TGFbetaRI kinase reduced the expression of a cohort of fibrotic markers by dermal fibroblasts from patients with dcSSc, including type I collagen and beta1 integrin. Moreover, inhibition also attenuated the elevated adhesive and contractile abilities of dcSSc fibroblasts. Our data suggest that some of the key profibrotic features of lesional SSc fibroblasts are dependent upon ALK-5 activity. Thus, TGFbetaRI kinase-mediated signaling may contribute to dermal fibrosis in dcSSc.

Matrix contraction by dermal fibroblasts requires syndecan 4: Insights into pathological scarring in chronic fibrotic disease

Chronic fibrotic diseases, such as diffuse systemic sclerosis (dSSc), are characterized by excessive scarring. The cell type that causes scarring is the myofibroblast. We show that fibroblasts isolated from scars of dSSc patients retain their myofibroblast phenotype in cell culture. We find that antagonizing transforming growth factor-beta receptor type I with the specific ALK5 inhibitor SB431542 alleviates the elevated matrix contraction by dSSc fibroblasts. dSSc fibroblasts show elevated ERK activation, and antagonizing MEK with U0126 suppresses the contractile abilities of dSSc dermal fibroblasts. TGFbeta-induced matrix contraction requires the proteglycan syndecan 4, as TGFbeta cannot induce a contractile phenotype in syndecan 4 −/ − dermal fibroblasts or induce phosphorylation of ERK. Lesional dSSc fibroblasts overexpress syndecan 4. Syndecan 4 siRNA reduces the activated phenotype of SSc fibroblasts. Thus antagonizing syndecan 4 would be expected to alleviate scarring in chronic fibrotic diseases, including dSSc.

Matrix Contraction by Dermal Fibroblasts Requires Transforming Growth Factor-β/Activin-Linked Kinase 5, Heparan Sulfate-Containing Proteoglycans, and MEK/ERK: Insights into Pathological Scarring in Chronic Fibrotic Disease

Scarring is characterized by excessive synthesis and contraction of extracellular matrix. Here, we show that fibroblasts from scarred (lesional) areas of patients with the chronic fibrotic disorder diffuse scleroderma [diffuse systemic sclerosis (dSSc)] show an enhanced ability to adhere to and contract extracellular matrix, relative to fibroblasts from unscarred (nonlesional) areas of dSSc patients and dermal fibroblasts from normal, healthy individuals. The contractile abilities of normal and dSSc dermal fibroblasts were suppressed by blocking heparin sulfate-containing proteoglycan biosynthesis or antagonizing transforming growth factor-beta receptor type I [activin-linked kinase (ALK5)] or ras/mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK). Compared with both normal and nonlesional fibroblasts, lesional dSSc fibroblasts overexpressed the heparin sulfate-containing proteoglycan syndecan 4. We also found that the procontractile signals from transforming growth factor (TGF)-beta were integrated through syndecan 4 and MEK/ERK because the ability of TGFbeta to induce contraction of dermal fibroblasts was prevented by MEK antagonism. TGFbeta could not induce a contractile phenotype or phosphorylate ERK in syndecan 4(-/-) dermal fibroblasts. These results suggest that integrating TGFbeta and ERK signals via syndecan 4 is essential for the contractile ability of dermal fibroblasts. We conclude that antagonizing MEK/ERK, TGFbeta1/ALK5, or syndecan 4 may alleviate scarring in chronic fibrotic disease.

Effects of salvianolic acid B on expressions of TGF-β1 and its receptors in liver of rats with dimethylnitrosamine-induced hepatic fibrosis

To study the therapeutic mechanism of salvianolic acid B (SA-B) in treating hepatic fibrosis. Twenty-four Wistar rats were randomly divided into 3 groups: normal control group, untreated group and SA-B-treated group. Rats in the untreated and SA-B-treated groups were injected intraperitoneally with 0.5% dimethylnitrosamine (DMN) for 4 weeks, daily for 3 days each week at a dose of 10 microg/kg, to induce hepatic fibrosis. Then, rats in the SA-B-treated group were given SA-B orally for another 4 weeks. Hydroxyproline (Hyp) contents in liver tissue of the rats in 3 groups were determined with HCl hydrolysis, and collagen type I, transforming growth factor-beta 1 (TGF-beta1), transforming growth factor-beta receptor type I (TbetaRI) and transforming growth factor-beta receptor type II (TbetaRII) were detected by Western blotting. The Hyp content and the expressions of collagen type I, TGF-beta1, TbetaRI and TbetaRII in the liver tissue of rats in the untreated group increased significantly as compared with those in the normal control group, while those in SA-B-treated group decreased significantly as compared with those in the untreated group. The therapeutic mechanism of SA-B in treating hepatic fibrosis may be related to inhibiting the expressions of TGF-beta1 and its receptors.