The Notch γ-secretase inhibitor ameliorates kidney fibrosis via inhibition of TGF-β/Smad2/3 signaling pathway activation

Kidney fibrosis is a common feature of chronic kidney disease (CKD). A recent study suggests that abnormal Notch signaling activation contributes to the development of renal fibrosis. However, the molecular mechanism that regulates this process remains unexplored. Unilateral ureteral obstruction (UUO) or sham-operated C57BL6 mice (aged 10 weeks) were randomly assigned to receive dibenzazepine (DBZ, 250 μg/100 g/d) or vehicle for 7 days. Histologic examinations were performed on the kidneys using Masson's trichrome staining and immunohistochemistry. Real-time PCR and western blot analysis were used for detection of mRNA expression and protein phosphorylation. The expression of Notch 1, 3, and 4, Notch intracellular domain (NICD), and its target genes Hes1 and HeyL were upregulated in UUO mice, while the increase in NICD protein was significantly attenuated by DBZ. After 7 days, the severity of renal fibrosis and expression of fibrotic markers, including collagen 1α1/3α1, fibronectin, and α-smooth muscle actin, were markedly increased in UUO compared with sham mice. In contrast, administration of DBZ markedly attenuated these effects. Furthermore, DBZ significantly inhibited UUO-induced expression of transforming growth factor (TGF)-β, phosphorylated Smad 2, and Smad 3. Mechanistically, Notch signaling activation in tubular epithelial cells enhanced fibroblast proliferation and activation in a coculture experiment. Our study provides evidence that Notch signaling is implicated in renal fibrogenesis. The Notch inhibitor DBZ can ameliorate this process via inhibition of the TGF-β/Smad2/3 signaling pathway, and might be a novel drug for preventing chronic kidney disease.     

Pirfenidone modulates macrophage polarization and ameliorates radiation-induced lung fibrosis by inhibiting the TGF-β1/Smad3 pathway

Radiation-induced lung injury (RILI) mainly contributes to the complications of thoracic radiotherapy. RILI can be divided into radiation pneumonia (RP) and radiation-induced lung fibrosis (RILF). Once RILF occurs, patients will eventually develop irreversible respiratory failure; thus, a new treatment strategy to prevent RILI is urgently needed. This study explored the therapeutic effect of pirfenidone (PFD), a Food and Drug Administration (FDA)-approved drug for (IPF) treatment, and its mechanism in the treatment of RILF. In vivo, C57BL/6 mice received a 50 Gy dose of X-ray radiation to the whole thorax with or without the administration of PFD. Collagen deposition and fibrosis in the lung were reversed by PFD treatment, which was associated with reduced M2 macrophage infiltration and inhibition of the transforming growth factor-β1 (TGF-β1)/Drosophila mothers against the decapentaplegic 3 (Smad3) signalling pathway. Moreover, PFD treatment decreased the radiation-induced expression of TGF-β1 and phosphorylation of Smad3 in alveolar epithelial cells (AECs) and vascular endothelial cells (VECs). Furthermore, IL-4–induced M2 macrophage polarization and IL-13–induced M2 macrophage polarization were suppressed by PFD treatment in vitro, resulting in reductions in the release of arginase-1 (ARG-1), chitinase 3-like 3 (YM-1) and TGF-β1. Notably, the PFD-induced inhibitory effects on M2 macrophage polarization were associated with downregulation of nuclear factor kappa-B (NF-κB) p50 activity. Additionally, PFD could significantly inhibit ionizing radiation-induced chemokine secretion in MLE-12 cells and consequently impair the migration of RAW264.7 cells. PFD could also eliminate TGF-β1 from M2 macrophages by attenuating the activation of TGF-β1/Smad3. In conclusion, PFD is a potential therapeutic agent to ameliorate fibrosis in RILF by reducing M2 macrophage infiltration and inhibiting the activation of TGF-β1/Smad3.     

Dynamics of TGF-β/Smad signaling

The physiological responses to TGF-β stimulation are diverse and vary amongst different cell types and environmental conditions. Even though the principal molecular components of the canonical and the non-canonical TGF-β signaling pathways have been largely identified, the mechanism that underlies the well-established context dependent physiological responses remains a mystery. Understanding how the components of TGF-β signaling function as a system and how this system functions in the context of the global cellular regulatory network requires a more quantitative and systematic approach. Here, we review the recent progress in understanding TGF-β biology using integration of mathematical modeling and quantitative experimental analysis. These studies reveal many interesting dynamics of TGF-β signaling and how cells quantitatively decode variable doses of TGF-β stimulation.     

MFAP2 promotes HSCs activation through FBN1/TGF-β/Smad3 pathway

Liver fibrosis is a chronic inflammatory process characterized by the accumulation of extracellular matrix (ECM), which contributes to cirrhosis and hepatocellular carcinoma. Increasing evidence suggests that the activation of hepatic stellate cells (HSCs) under an inflammatory state leads to the secretion of collagens, which can cause cirrhosis. In this study, we analysed data from the Gene Expression Omnibus (GEO) databases to identify differentially expressed genes (DEGs) between quiescent and fibrotic HSCs. We found that Microfibril Associated Protein 2 (MFAP2) was elevated in carbon tetrachloride (CCl4)-induced liver fibrosis and Transforming Growth Factor-Beta 1 (TGF-β1)-activated HSCs. Knockdown of MFAP2 inhibited HSC proliferation and partially attenuated TGF-β-stimulated fibrogenesis markers. Bioinformatics analysis revealed that Fibrillin-1 (FBN1) was correlated with MFAP2, and the expression of FBN1 was significantly upregulated after MFAP2 overexpression. Silencing MFAP2 partially attenuated the activation of HSCs by inhibiting HSC proliferation and decreasing collagen deposits. In vitro results showed that the inhibition of MFAP2 alleviated hepatic fibrosis by inhibiting the activation and inducing the apoptosis of active HSCs in a CCl4-induced mouse model. In conclusion, our results suggest that MFAP2 is a potential target for the clinical treatment of liver fibrosis.     

Acetylshikonin from Zicao ameliorates renal dysfunction and fibrosis in diabetic mice by inhibiting TGF-β1/Smad pathway

Diabetic nephropathy (DN) is the major cause of end-stage renal disease in diabetic patients. Zicao, a well-known Chinese traditional medicine, has attracted much attention due to its beneficial effects in various medical fields. In this study, we attempted to investigate the effects and mechanisms of action of acetylshikonin, the main ingredient of Zicao, on renal dysfunction in DN. Our results showed that administration with acetylshikonin not only decreased blood urea nitrogen, urine creatinine and the mean kidney-to-body weight ratio in streptozotocin-induced diabetic mice, but also restored the loss of body weight, whereas the blood glucose was not changed. Masson’s trichrome staining showed that acetylshikonin treatment resulted in a marked decrease in kidney fibrosis from diabetic mice. The increased expression of fibrosis proteins, such as plasminogen activator inhibitor type 1 (PAI-1), connective tissue growth factor, and collagen III and IV, were reduced after acetylshikonin administration. In addition, the expressions of interleukin-1β, interleukin-6, monocyte chemoattractant protein-1, intercellular adhesion molecule 1 and infiltration of macrophages in kidney tissues were decreased in acetylshikonin-treated diabetic mice. Acetylshikonin led to a reduction of transforming growth factor-β1 (TGF-β1) expression and Smad-2/3 phosphorylation, as accompanied by increased Smad7 expression. Furthermore, in vitro treatment with acetylshikonin markedly attenuated TGF-β1-induced the PAI-1, collagen III and IV, and Smad-2/3 phosphorylation in HK2 immortalized human proximal tubule epithelial cells. Acetylshikonin also prevented epithelial-to-mesenchymal transition induced by TGF-β1. Collectively, our study provides evidences that acetylshikonin attenuates renal fibrosis though inhibiting TGF-β1/Smad signaling pathway, suggesting that acetylshikonin may be a novel therapeutic agent for the treatment of DN.     

Fluorofenidone ameliorates cholestasis and fibrosis by inhibiting hepatic Erk/-Egr-1 signaling and Tgfβ1/Smad pathway in mice

Cholestasis is characterized by intrahepatic accumulation of bile acids (BAs), resulting in liver injury, fibrosis, and liver failure. To date, only ursodeoxycholic acid and obeticholic acid have been approved for the treatment of cholestasis. As fluorofenidone (AKF-PD) was previously reported to play significant anti-fibrotic and anti-inflammatory roles in various diseases, we investigated whether AKF-PD ameliorates cholestasis. A mouse model of cholestasis was constructed by administering a 0.1 % 3,5-diethoxycarbonyl-1,4-dihydroxychollidine (DDC) diet for 14 days. Male C57BL/6 J mice were treated with either AKF-PD or pirfenidone (PD) orally in addition to the DDC diet. Serum and liver tissues were subsequently collected and analyzed. We found that AKF-PD significantly reduced the levels of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and total bile salts (TBA), as well as hepatic bile acids (BAs) levels. Hepatic histological analyses demonstrated that AKF-PD markedly attenuated hepatic inflammation and fibrosis. Further mechanistic analyses revealed that AKF-PD markedly inhibited expression of Cyp7a1, an enzyme key to BAs synthesis, by increasing Fxr nuclear translocation, and decreased hepatic inflammation by attenuating Erk/-Egr-1-mediated expression of inflammatory cytokines and chemokines Tnfα, Il-1β, Il-6, Ccl2, Ccl5 and Cxcl10. Moreover, AKF-PD was found to substantially reduce liver fibrosis via inhibition of Tgfβ1/Smad pathway in our mouse model. Here, we found that AKF-PD effectively attenuates cholestasis and hepatic fibrosis in the mouse model of DDC-induced cholestasis. As such, AKF-PD warrants further investigation as a candidate drug for treatment of cholestasis.     

Periodontal ligament-associated protein-1 knockout mice regulate the differentiation of osteoclasts and osteoblasts through TGF-β1/Smad signaling pathway

It has been known that periodontal ligament-associated protein-1 (PLAP-1/Asporin) not only inhibits cartilage formation in osteoarthritis, but it also influences the healing of skull defect. However, the effect and mechanism of PLAP-1/Asporin on the mutual regulation of osteoclasts and osteoblasts in periodontitis are not clear. In this study, we utilized a PLAP-1/Asporin gene knockout (KO) mouse model to research this unknown issue. We cultured mouse bone marrow mesenchymal stem cells with Porphyromonas gingivalis lipopolysaccharide (P.g. LPS) for osteogenic induction in vitro. The molecular mechanism of PLAP-1/Asporin in the regulation of osteoblasts was detected by immunoprecipitation, immunofluorescence, and inhibitors of signaling pathways. The results showed that the KO of PLAP-1/Asporin promoted osteogenic differentiation through transforming growth factor beta 1 (TGF-β1)/Smad3 in inflammatory environments. We further found the KO of PLAP-1/Asporin inhibited osteoclast differentiation and promoted osteogenic differentiation through the TGF-β1/Smad signaling pathway in an inflammatory coculture system. The experimental periodontitis model was established by silk ligation and the alveolar bone formation in PLAP-1/Asporin KO mice was promoted through TGF-β1/Smad3 signaling pathway. The subcutaneous osteogenesis model in nude mice also confirmed that the KO of PLAP-1/Asporin promoted bone formation by the histochemical staining. In conclusion, PLAP-1/Asporin regulated the differentiation of osteoclasts and osteoblasts through TGF-β1/Smad signaling pathway. The results of this study lay a theoretical foundation for the further study of the pathological mechanism underlying alveolar bone resorption, and the prevention and treatment of periodontitis.     

PI3K/Akt pathway mediates high glucose-induced lipogenesis and extracellular matrix accumulation in HKC cells through regulation of SREBP-1 and TGF-β1

Previous studies have shown that high glucose stimulates renal SREBP-1 gene expression and increases renal tubular cells lipid metabolism, however, the mechanisms remain elusive. In the present study we demonstrated that PI3K/Akt pathway was activated in human renal proximal tubular cell line (HKC) exposed to high glucose accompanied with up-regulation of SREBP-1, TGF-β1, lipid droplets deposits and extracellular matrix production. Inhibition of PI3K/Akt pathway by chemical LY294002 or specific short hairpin RNA (shRNA) vector prevented SREBP-1 and TGF-β1 up-regulation, as well as ameliorated HKC cells lipogenesis and extracellular matrix accumulation. These findings indicate that PI3K/Akt pathway potentially mediates high glucose-induced lipogenesis and extracellular matrix accumulation in HKC cells.     

PCBP2 promotes the development of glioma by regulating FHL3/TGF-β/Smad signaling pathway

The purpose of this study was to investigate the role of Poly (C)-binding protein 2 (PCBP2) and the related signaling pathway in glioma progression. Quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC) were performed to measure PCBP2 messenger RNA and protein expression in glioma tissues or cells. Cell transfection was completed using Lipofectamine 2000. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, Transwell assay and flow cytometry assay were used to explore the effects of PCBP2 expression on biological behaviors of glioma cells. Western blot assay was used for the detection of pathway related proteins. Expression of PCBP2 in glioma tissues and cells were higher than that in paracancerous tissues and normal cells (both p < .01). Moreover, the elevated expression of PCBP2 was significantly correlated with tumor size (p = .001) and WHO stage (p = .010). Knockdown of PCBP2 could suppress proliferation, migration and invasion of glioma cells and promote apoptosis. Besides, the expression of transforming growth factor-β (TGF-β) pathway related proteins TGF-β1, p-Smad2 and p-Smad7 were decreased following the downregulation of PCBP2. PCBP2 also inhibited FHL3 expression by binding to FHL3-3′UTR. The inhibition of FHL3 could reverse the antitumor action caused by PCBP2 silencing. In vivo assay, PCBP2 was also found to inhibit the tumor growth of glioma. PCBP2 activates TGF-β/Smad signaling pathway by inhibiting FHL3 expression, thus promoting the development and progression of glioma.     

Induction of galectin-1 by TGF-β1 accelerates fibrosis through enhancing nuclear retention of Smad2

Fibrosis is one of the most serious side effects in cancer patients undergoing radio-/ chemo-therapy, especially of the lung, pancreas or kidney. Based on our previous finding that galectin-1 (Gal-1) was significantly increased during radiation-induced lung fibrosis in areas of pulmonary fibrosis, we herein clarified the roles and action mechanisms of Gal-1 during fibrosis. Our results revealed that treatment with TGF-β1 induced the differentiation of fibroblast cell lines (NIH3T3 and IMR-90) to myofibroblasts, as evidenced by increased expression of the fibrotic markers smooth muscle actin-alpha (α-SMA), fibronectin, and collagen (Col-1). We also observed marked and time-dependent increases in the expression level and nuclear accumulation of Gal-1. The TGF-β1-induced increases in Gal-1, α-SMA and Col-1 were decreased by inhibitors of PI3-kinase and p38 MAPK, but not ERK. Gal-1 knockdown using shRNA decreased the phosphorylation and nuclear retention of Smad2, preventing the differentiation of fibroblasts. Gal-1 interacted with Smad2 and phosphorylated Smad2, which may accelerate fibrotic processes. In addition, up-regulation of Gal-1 expression was demonstrated in a bleomycin (BLM)-induced mouse model of lung fibrosis in vivo. Together, our results indicate that Gal-1 may promote the TGF-β1-induced differentiation of fibroblasts by sustaining nuclear localization of Smad2, and could be a potential target for the treatment of pulmonary fibrotic diseases.     

Protective effect of TRPV1 against renal fibrosis via inhibition of TGF-β/Smad signaling in DOCA-salt hypertension

To investigate the effects of the transient receptor potential vanilloid type 1 (TRPV1) channel on renal extracellular matrix (ECM) protein expression including collagen deposition and the transforming growth factor β (TGF-β)/Smad signaling pathway during salt-dependent hypertension, wild-type (WT) and TRPV1-null (TRPV1?/?) mutant mice were uninephrectomized and given deoxycorticosterone acetate (DOCA)-salt for 4 wks. TRPV1 gene ablation exaggerated DOCA-salt-induced impairment of renal function as evidenced by increased albumin excretion (μg/24 h) compared with WT mice (83.7 ± 7.1 versus 28.3 ± 4.8, P < 0.05), but had no apparent effect on mean arterial pressure (mmHg) as determined by radiotelemetry (141 ± 4 versus 138 ± 3, P > 0.05). Morphological analysis showed that DOCA-salt-induced glomerulosclerosis, tubular injury and macrophage infiltration (cells/mm2) were increased in TRPV1?/? compared with WT mice (0.74 ± 0.08 versus 0.34 ± 0.04; 3.14 ± 0.26 versus 2.00 ± 0.31; 68 ± 5 versus 40 ± 4, P < 0.05). Immunostaining studies showed that DOCA-salt treatment decreased nephrin but increased collagen type I and IV as well as phosphorylated Smad2/3 staining in kidneys of TRPV1?/? compared with WT mice. Hydroxyproline assay and Western blot showed that DOCA-salt treatment increased collagen content (μg/mg dry tissue) and fibronectin protein expression (%β-actin arbitrary units) in the kidney of TRPV1?/? compared with WT mice (26.7 ± 2.7 versus 17.4 ± 1.8; 0.93 ± 0.07 versus 0.65 ± 0.08, P < 0.05). Acceleration of renal ECM protein deposition in DOCA-salt-treated TRPV1?/? mice was accompanied by increased TGF-β1, as well as phosphorylation of Smad2/3 protein expression (%β-actin arbitrary units) compared with DOCA-salt-treated WT mice (0.61 ± 0.07 versus 0.32 ± 0.05; 0.57 ± 0.07 versus 0.25 ± 0.05; 0.71 ± 0.08 versus 0.40 ± 0.06, P < 0.05). These results show that exaggerated renal functional and structural injuries are accompanied by increased production of ECM protein and activation of the TGF-β/Smad2/3 signaling pathway. These data suggest that activation of TRPV1 attenuates the progression of renal fibrosis possibly via suppression of the TGF-β and its downstream regulatory signaling pathway.     

Metformin attenuates post-epidural fibrosis by inhibiting the TGF-β1/Smad3 and HMGB1/TLR4 signaling pathways

Excessive post-epidural fibrosis is a common cause of recurrent back pain after spinal surgery. Though various treatment methods have been conducted, the safe and effective drug for alleviating post-epidural fibrosis remains largely unknown. Metformin, a medicine used in the treatment of type 2 diabetes, has been noted to relieve fibrosis in various organs. In the present study, we aimed to explore the roles and mechanisms of metformin in scar formation in a mouse model of laminectomy. Post-epidural fibrosis developed in a mouse model of laminectomy by spinous process and the T12-L2 vertebral plate with a rongeur. With the administration of metformin, post-epidural fibrosis was reduced, accompanied with decreased collagen and fibronectin in the scar tissues. Mechanistically, metformin decreased fibronectin and collagen deposition in fibroblast cells, and this effect was dependent on the HMGB1/TLR4 and TGF-β1/Smad3 signalling pathways. In addition, metformin influenced the metabolomics of the fibroblast cells. Taken together, our study suggests that metformin may be a potential option to mitigate epidural fibrosis after laminectomy.     

17β-Estradiol mediates TGFBR3/Smad2/3 signaling to attenuate the fibrosis of TGF-β1-induced bovine endometrial epithelial cells via GPER

Abnormal function and fibrosis of endometrium caused by cows' endometritis pose difficult implantation of embryos and uterine cavity adhesions. 17β-Estradiol (E2) serves as the most effective aromatized estrogen, and its synthetase and receptors have been detected in the endometrium. Studies have demonstrated the positive role of estrogen in combating pathological fibrosis in diverse diseases. However, it is still unknown whether E2 regulates endometrium fibrosis in bovine endometritis. Herein, we evaluated the expression patterns of transforming growth factor-β1 (TGF-β1), epithelial-mesenchymal transformation (EMT)-related proteins (α-SMA, vimentin N-cadherin and E-cadherin), cytochrome P450 19A1 (CYP19A1), and G protein-coupled estrogen receptor (GPER) in bovine healthy endometrium and Inflammatory endometrium. Our data showed that the inflamed endometrium presented low CYP19A1 and GPER expression, and significantly higher EMT process versus the normal tissue. Moreover, we established a TGF-β1-induced fibrosis model in BEND cells, and found that E2 inhibited the EMT process of BEND cells in a dose-dependent manner. The anti-fibrotic effect of E2 was blocked by the GPER inhibitor G15, but not the estrogen nuclear receptors (ERs) inhibitor ICI182780. Moreover, the GPER agonist G1 inhibited fibrosis and Smad2/3 phosphorylation but increased the expression of TGFBR3 in BEND cells. Transfection with TGFBR3 small interfering RNA blocked the effect of G1 on fibrosis of BEND cells and upregulated the expression of P-Smad2/3. Our in vivo data also showed that E2 and G1 affected uterus fibrosis in mice endometritis model caused by LPS, which was associated with the inhibition of TGFBR3/Smad2/3 signaling. In conclusion, our data implied that E2 alleviates the fibrosis of TGF-β1-induced BEND cells, which is associated with the GPER mediation of TGFBR3/Smad2/3 signaling.     

Endoglin promotes TGF-β/Smad1 signaling in scleroderma fibroblasts

TGF-β is the primary inducer of extracellular matrix proteins in scleroderma (systemic sclerosis, SSc). Previous studies indicate that in a subset of SSc fibroblasts TGF-β signaling is activated via elevated levels of activin receptor-like kinase (ALK) 1 and phosphorylated Smad1 (pSmad1). The goal of this study was to determine the role of endoglin/ALK1 in TGF-β/Smad1 signaling in SSc fibroblasts. In SSc fibroblasts, increased levels of endoglin correlated with high levels of pSmad1, collagen, and connective tissue growth factor (CCN2). Endoglin depletion via siRNA in SSc fibroblasts inhibited pSmad1 but did not affect pSmad2/3. Following endoglin depletion mRNA and protein levels of collagen and CCN2 were significantly decreased in SSc fibroblasts but remained unchanged in normal fibroblasts. ALK1 was expressed at similar levels in SSc and normal fibroblasts. Depletion of ALK1 resulted in inhibition of pSmad1 and a moderate but significant reduction of mRNA and protein levels of collagen and CCN2 in SSc fibroblasts. Furthermore, constitutively high levels of endoglin were found in complexes with ALK1 in SSc fibroblasts. Overexpression of constitutively active ALK1 (caALK1) in normal and SSc fibroblasts led to a moderate increase of collagen and CCN2. However, caALK1 potently induced endothelin 1 (ET-1) mRNA and protein levels in SSc fibroblasts. Additional experiments demonstrated that endoglin and ALK1 mediate TGF-β induction of ET-1 in SSc and normal fibroblasts. In conclusion, this study has revealed an important profibrotic role of endoglin in SSc fibroblasts. The endoglin/ALK1/Smad1 pathway could be a therapeutic target in patients with SSc if appropriately blocked.     

Calreticulin regulates TGF-β1-induced epithelial mesenchymal transition through modulating Smad signaling and calcium signaling

As a Ca²⁺ binding protein, calreticulin (CRT) has many functions and plays an important role in a variety of tumors. The role of CRT in TGF-β1-induced EMT is unknown. In this study, we demonstrated in vitro that TGF-β1-induced EMT elevated the expression of CRT in A549 lung cancer cells. Subsequently, we confirmed that overexpression CRT had no capacity to induce A549 cells EMT alone, but successfully enhanced TGF-β1-induced-EMT. Furthermore, knockdown of CRT in A549 cells significantly suppressed changes of EMT marks expression induced by TGF-β1. On treatment with TGF-β1, overexpression of CRT could enhance the phosphorylation of both Smad2 and Smad3. Consistently, the knockdown of CRT by siRNA-CRT could inhibit Smad signaling pathway activated by TGF-β1. These results indicated that CRT regulates EMT induced by TGF-β1 through Smad signaling pathway. Finally, TGF-β1-induced-EMT enhanced store-operated Ca²⁺ influx in A549 cells. CRT knockdown was able to abolish the effect of TGF-β1 on thapsigargin (TG) −induced Ca²⁺ release, but had failed to reduce store-operated Ca²⁺ influx. The alteration of intracellular Ca²⁺ concentration by TG or BAPTA-AM was able to regulate EMT induced by TGF-β1 through Smad signaling pathway. Together, these data identify that CRT regulates TGF-β1-induced-EMT through modulating Smad signaling. Furthermore, TGF-β1-induced-EMT is highly calcium-dependent, CRT was partly involved in it.     

Protection from obesity and diabetes by blockade of TGF-β/Smad3 signaling

Imbalances in glucose and energy homeostasis are at the core of the worldwide epidemic of obesity and diabetes. Here, we illustrate an important role of the TGF-β/Smad3 signaling pathway in regulating glucose and energy homeostasis. Smad3-deficient mice are protected from diet-induced obesity and diabetes. Interestingly, the metabolic protection is accompanied by Smad3(-)(/-) white adipose tissue acquiring the bioenergetic and gene expression profile of brown fat/skeletal muscle. Smad3(-/-) adipocytes demonstrate a marked increase in mitochondrial biogenesis, with a corresponding increase in basal respiration, and Smad3 acts as a repressor of PGC-1α expression. We observe significant correlation between TGF-β1 levels and adiposity in rodents and humans. Further, systemic blockade of TGF-β signaling protects mice from obesity, diabetes, and hepatic steatosis. Together, these results demonstrate that TGF-β signaling regulates glucose tolerance and energy homeostasis and suggest that modulation of TGF-β activity might be an effective treatment strategy for obesity and diabetes.     

Crosstalk between TGF-β1 and CXCR3 signaling during urethral fibrosis

Urethral fibrosis is an important pathological feature of urethral stricture. TGF-β1 and CXC chemokine receptor 3 (CXCR3) signaling have been reported as the critical pathways involved in the pathology of fibrosis. Here, we collected the urine samples from the patients with recurring urethral stricture, recurring stricture treated by cystostomy, and age- and gender-matched healthy people. ELISA detection revealed that TGF-β1 level was significantly up-regulated for the urethral stricture patients. By contrast, flow cytometry, real-time PCR detection, and immunofluoresecent staining showed that urethral stricture resulted in decreased expression of CXCR3. TGF-β1 treatment could increase cell proliferation and migration ability of urethra fibroblasts, whereas IP-10/CXCR3 signaling showed the opposite effect. Further, we found a crosstalk between TGF-β1 and CXCR3 signaling in the regulation of urethral fibrosis. Thus, pharmacological intervention of TGF-β1 or CXCR3 signaling has a potential as the therapeutic target for the prevention of urethral fibrosis.