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Effects of Chinese Medicine Tong xinluo on Diabetic Nephropathy via Inhibiting TGF- β 1-Induced Epithelial-to-Mesenchymal Transition.

Zhang N, Gao Y, Zou D, Wang J, Li J, Zhou S, Zhu Z, Zhao X, Xu L, Zhang H - Evid Based Complement Alternat Med (2014)

Bottom Line: As a result, the expressions of collagen IV (Col IV) and fibronectin (FN) were significantly decreased in TXL group.In vivo, 24 h-UAER (24-hour urine albumin excretion ratio) and BUN (blood urea nitrogen) were decreased and Ccr (creatinine clearance ratio) was increased in TXL group compared with DN group.In summary, the present study demonstrates that TXL successfully inhibits TGF- β 1-induced epithelial-to-mesenchymal transition in DN, which may account for the therapeutic efficacy in TXL-mediated renoprotection.

View Article: PubMed Central - PubMed

Affiliation: School of Traditional Chinese Medicine, Capital Medical University, No. 10, Youanmenwai, Xitoutiao, Fengtai District, Beijing 100069, China.

ABSTRACT
Diabetic nephropathy (DN) is a major cause of chronic kidney failure and characterized by interstitial and glomeruli fibrosis. Epithelial-to-mesenchymal transition (EMT) plays an important role in the pathogenesis of DN. Tong xinluo (TXL), a Chinese herbal compound, has been used in China with established therapeutic efficacy in patients with DN. To investigate the molecular mechanism of TXL improving DN, KK-Ay mice were selected as models for the evaluation of pathogenesis and treatment in DN. In vitro, TGF- β 1 was used to induce EMT. Western blot (WB), immunofluorescence staining, and real-time polymerase chain reaction (RT-PCR) were applied to detect the changes of EMT markers in vivo and in vitro, respectively. Results showed the expressions of TGF- β 1 and its downstream proteins smad3/p-smad3 were greatly reduced in TXL group; meantime, TXL restored the expression of smad7. As a result, the expressions of collagen IV (Col IV) and fibronectin (FN) were significantly decreased in TXL group. In vivo, 24 h-UAER (24-hour urine albumin excretion ratio) and BUN (blood urea nitrogen) were decreased and Ccr (creatinine clearance ratio) was increased in TXL group compared with DN group. In summary, the present study demonstrates that TXL successfully inhibits TGF- β 1-induced epithelial-to-mesenchymal transition in DN, which may account for the therapeutic efficacy in TXL-mediated renoprotection.

No MeSH data available.


Related in: MedlinePlus

Effects of TXL on the expressions of TGF-β1, E-CA, and α-SMA in renal tissues of KK-Ay mice. (a) mRNA expression of TGF-β1 was determined by RT-PCR with β-actin as an internal control. (b) mRNA expressions of E-CA and α-SMA were determined by RT-PCR with β-actin as an internal control. (c) Representative bands of TGF-β1, E-CA, and α-SMA detected by western blot. (d) Representative immunofluorescence staining photographs of E-CA and α-SMA, visualized by confocal microscope. Images are shown at 20x. (e) Mean fluorescence activity of E-CA and α-SMA analyzed by image-pro plus 6.0 software. (f) Densitometry analysis of TGF-β1, E-CA, and α-SMA bands from (c), normalized to β-actin.
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fig1: Effects of TXL on the expressions of TGF-β1, E-CA, and α-SMA in renal tissues of KK-Ay mice. (a) mRNA expression of TGF-β1 was determined by RT-PCR with β-actin as an internal control. (b) mRNA expressions of E-CA and α-SMA were determined by RT-PCR with β-actin as an internal control. (c) Representative bands of TGF-β1, E-CA, and α-SMA detected by western blot. (d) Representative immunofluorescence staining photographs of E-CA and α-SMA, visualized by confocal microscope. Images are shown at 20x. (e) Mean fluorescence activity of E-CA and α-SMA analyzed by image-pro plus 6.0 software. (f) Densitometry analysis of TGF-β1, E-CA, and α-SMA bands from (c), normalized to β-actin.

Mentions: Overwhelming evidences implicate that TGF-β1 acts as the key mediator of tubular EMT [30]. To observe whether TXL affected TGF-β1 expression and markers of EMT in vivo, RT-PCR was first performed to examine the expression of TGF-β1 at 24 weeks of age. RT-PCR results showed that TGF-β1 was significantly enhanced in DN group compared with normal group (Figure 1(a), *P < 0.05). After being treated with TXL for 12 weeks, expression of TGF-β1 was significantly decreased compared with DN group (Figure 1(a), ▲P < 0.05). Additionally, Western blot results were consistent with RT-PCR results (Figures 1(c) and 1(f)). TXL suppressed TGF-β1 expression both at mRNA and protein levels. Next, to further verify the effect of TXL on tubular EMT, E-CA and α-SMA were detected by RT-PCR, western blot, and immunofluorescence staining, respectively. RT-PCR and western blot results showed that α-SMA was enhanced and E-CA was decreased in DN group compared with normal group (Figures 1(b), 1(c), and 1(f), *P < 0.05). More importantly, TXL significantly decreased α-SMA expression and increased E-CA expression compared with DN group both at mRNA and protein levels (Figures 1(b), 1(c), and 1(f), ▲P < 0.05). Immunofluorescence staining showed the similar results that epithelial marker E-CA was significantly decreased, while α-SMA was increased in DN group compared with normal group (Figures 1(d) and 1(e), *P < 0.05). More importantly, TXL treatment significantly restored E-CA and α-SMA expressions (Figures 1(d) and 1(e), ▲P < 0.05). These results demonstrated that TXL can inhibit TGF-β1 expression and EMT in DN.


Effects of Chinese Medicine Tong xinluo on Diabetic Nephropathy via Inhibiting TGF- β 1-Induced Epithelial-to-Mesenchymal Transition.

Zhang N, Gao Y, Zou D, Wang J, Li J, Zhou S, Zhu Z, Zhao X, Xu L, Zhang H - Evid Based Complement Alternat Med (2014)

Effects of TXL on the expressions of TGF-β1, E-CA, and α-SMA in renal tissues of KK-Ay mice. (a) mRNA expression of TGF-β1 was determined by RT-PCR with β-actin as an internal control. (b) mRNA expressions of E-CA and α-SMA were determined by RT-PCR with β-actin as an internal control. (c) Representative bands of TGF-β1, E-CA, and α-SMA detected by western blot. (d) Representative immunofluorescence staining photographs of E-CA and α-SMA, visualized by confocal microscope. Images are shown at 20x. (e) Mean fluorescence activity of E-CA and α-SMA analyzed by image-pro plus 6.0 software. (f) Densitometry analysis of TGF-β1, E-CA, and α-SMA bands from (c), normalized to β-actin.
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Related In: Results  -  Collection

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fig1: Effects of TXL on the expressions of TGF-β1, E-CA, and α-SMA in renal tissues of KK-Ay mice. (a) mRNA expression of TGF-β1 was determined by RT-PCR with β-actin as an internal control. (b) mRNA expressions of E-CA and α-SMA were determined by RT-PCR with β-actin as an internal control. (c) Representative bands of TGF-β1, E-CA, and α-SMA detected by western blot. (d) Representative immunofluorescence staining photographs of E-CA and α-SMA, visualized by confocal microscope. Images are shown at 20x. (e) Mean fluorescence activity of E-CA and α-SMA analyzed by image-pro plus 6.0 software. (f) Densitometry analysis of TGF-β1, E-CA, and α-SMA bands from (c), normalized to β-actin.
Mentions: Overwhelming evidences implicate that TGF-β1 acts as the key mediator of tubular EMT [30]. To observe whether TXL affected TGF-β1 expression and markers of EMT in vivo, RT-PCR was first performed to examine the expression of TGF-β1 at 24 weeks of age. RT-PCR results showed that TGF-β1 was significantly enhanced in DN group compared with normal group (Figure 1(a), *P < 0.05). After being treated with TXL for 12 weeks, expression of TGF-β1 was significantly decreased compared with DN group (Figure 1(a), ▲P < 0.05). Additionally, Western blot results were consistent with RT-PCR results (Figures 1(c) and 1(f)). TXL suppressed TGF-β1 expression both at mRNA and protein levels. Next, to further verify the effect of TXL on tubular EMT, E-CA and α-SMA were detected by RT-PCR, western blot, and immunofluorescence staining, respectively. RT-PCR and western blot results showed that α-SMA was enhanced and E-CA was decreased in DN group compared with normal group (Figures 1(b), 1(c), and 1(f), *P < 0.05). More importantly, TXL significantly decreased α-SMA expression and increased E-CA expression compared with DN group both at mRNA and protein levels (Figures 1(b), 1(c), and 1(f), ▲P < 0.05). Immunofluorescence staining showed the similar results that epithelial marker E-CA was significantly decreased, while α-SMA was increased in DN group compared with normal group (Figures 1(d) and 1(e), *P < 0.05). More importantly, TXL treatment significantly restored E-CA and α-SMA expressions (Figures 1(d) and 1(e), ▲P < 0.05). These results demonstrated that TXL can inhibit TGF-β1 expression and EMT in DN.

Bottom Line: As a result, the expressions of collagen IV (Col IV) and fibronectin (FN) were significantly decreased in TXL group.In vivo, 24 h-UAER (24-hour urine albumin excretion ratio) and BUN (blood urea nitrogen) were decreased and Ccr (creatinine clearance ratio) was increased in TXL group compared with DN group.In summary, the present study demonstrates that TXL successfully inhibits TGF- β 1-induced epithelial-to-mesenchymal transition in DN, which may account for the therapeutic efficacy in TXL-mediated renoprotection.

View Article: PubMed Central - PubMed

Affiliation: School of Traditional Chinese Medicine, Capital Medical University, No. 10, Youanmenwai, Xitoutiao, Fengtai District, Beijing 100069, China.

ABSTRACT
Diabetic nephropathy (DN) is a major cause of chronic kidney failure and characterized by interstitial and glomeruli fibrosis. Epithelial-to-mesenchymal transition (EMT) plays an important role in the pathogenesis of DN. Tong xinluo (TXL), a Chinese herbal compound, has been used in China with established therapeutic efficacy in patients with DN. To investigate the molecular mechanism of TXL improving DN, KK-Ay mice were selected as models for the evaluation of pathogenesis and treatment in DN. In vitro, TGF- β 1 was used to induce EMT. Western blot (WB), immunofluorescence staining, and real-time polymerase chain reaction (RT-PCR) were applied to detect the changes of EMT markers in vivo and in vitro, respectively. Results showed the expressions of TGF- β 1 and its downstream proteins smad3/p-smad3 were greatly reduced in TXL group; meantime, TXL restored the expression of smad7. As a result, the expressions of collagen IV (Col IV) and fibronectin (FN) were significantly decreased in TXL group. In vivo, 24 h-UAER (24-hour urine albumin excretion ratio) and BUN (blood urea nitrogen) were decreased and Ccr (creatinine clearance ratio) was increased in TXL group compared with DN group. In summary, the present study demonstrates that TXL successfully inhibits TGF- β 1-induced epithelial-to-mesenchymal transition in DN, which may account for the therapeutic efficacy in TXL-mediated renoprotection.

No MeSH data available.


Related in: MedlinePlus