Fuzheng Huayu Formula () prevents rat renal interstitial fibrosis induced by HgCl2 via antioxidative stress and down-regulation of nuclear factor-kappa B activity.

OBJECTIVE: To investigate the mechanism of action of Fuzheng Huayu Formula (, FZHY) against renal interstitial fibrosis (RIF) relating to oxidative injury and nuclear factor-kappa B (NF-κB) activity. METHODS: Thirty-two Sprague-Dawley rats were randomly divided into 3 groups: normal group, model group and FZHY treatment group. The RIF model was induced by oral administration of HgCl2 at a dose of 8 mg/kg body weight once a day for 9 weeks. Meanwhile, rats in FZHY treatment group orally took FZHY at a dose of 4.0 g/kg rat weight for 9 weeks. The content of hydroxyproline (Hyp) and collagen deposition in kidney were observed. The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), the content of glutathione (GSH) and malondialdehyde (MDA) of kidney were tested. The expressions of inhibitor-κappa B (IκB), phospho-IκB (p-IκB), tumor necrosis factor-α (TNF-α), matrix metalloproteinase-2 (MMP-2) and α-smooth muscle actin (α-SMA) were analyzed by Western blot. α-SMA expression was also observed by immunofluorescent staining. MMP-2 activity was measured by gelatin zymography. NF-κB activation was determined by electrophoretic mobility shift assay. RESULTS: Renal interstitial fibrosis was induced by HgCl2, demonstrated by remarkably increased Hyp contents and excessive collagen deposition in kidney (P<0.01). FZHY significantly inhibited renal interstitial collagen deposition and reduced Hyp content of the HgCl2-treated rats (P<0.01). GSH content decreased obviously, and MDA content increased signifificantly in HgCl2-treated rats compared with that of normal rats (P<0.01). FZHY significantly increased GSH content and decreased MDA content in the model rats (P<0.01). The expression α-SMA was increased in model rats compared with that of normal rats, FZHY signifificantly decreased its expression (P<0.01). The expressions of p-IκB and TNF-α and MMP-2, MMP-2 activity, and NF-κB activation were increased in model group compared with that in normal group (P<0.01), FZHY signifificantly decreased NF-κB activation, MMP-2 activity and p-IκB and TNF-α expressions (P<0.01). CONCLUSIONS: FZHY could protect kidney from oxidative injury intoxicated by HgCl2, and antagonized oxidative stress-stimulated NF-κB activity through inhibition of IκB phosphorylation in the interstitial fibrotic kidney, these effects importantly contributed to FZHY action mechanism against renal interstitial fifibrosis.

[Performance of FibroScan in evaluating the curative effects of traditional Chinese medicine on liver fibrosis].

OBJECTIVE: To assess the performance of FibroScan in evaluating the curative effects of traditional Chinese medicine (TCM) on liver fibrosis, and to analyze factors influencing the diagnostic accuracy. METHODS: Data of FibroScan values, types of disease, use of drug, liver function indexes, prothrombin time (PT) and international normalized ratio (INR) were collected at both pre- (1 month prior) and post-FibroScan for 102 patients who underwent at least two FibroScan procedures. Patients were subgrouped according to presence of fibrosis, presence of cirrhosis, and TCM formulation and statistically analyzed. RESULTS: The pre- and post-FibroScan mean liver stiffness measurements (LSMs) were significantly different when the variation of LSM was more than or equal to2 kPa for the non-fibrotic group (vs. the fibrotic group), or when the variation wasmore than or equal to4 kPa for the cirrhotic group (vs. the non-cirrhotic group). In addition, the three TCM formulation groups showed significant differences, with the most robust difference exhibited between the FuZheng HuaYu formulation group and the other treatment groups (P = 0.010). No significant differences were observed for the liver function indexes, PT, or INR. However, the post-FibroScan levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and gamma-glutamyltransferase (GGT) was significantly reduced in patients with reduced LSM. CONCLUSION: FibroScan may be a useful non-invasive clinical tool for evaluating the comprehensive curative effect of treatments for chronic liver diseases, and its performance is not obviously impacted by ALT, AST, GGT, PT, and INR. The criteria for efficacy established by FibroScan are 2 kPa for the patients without liver fibrosis and 4 kPa for patients with liver cirrhosis.

[Biochemical characteristics of traditional Chinese medicine syndromes and their elements in patients with hepatitis B cirrhosis].

OBJECTIVE: To investigate the characteristics of traditional Chinese medicine (TCM) syndromes and their elements in patients with post hepatitic cirrhosis by analyzing the relationships between signs and symptoms and biochemical parameters. METHODS: A total of 440 patients with hepatitis B cirrhosis treated in Shanghai Public Health Center and Shuguang Hospital, Longhua Hospital and Central Hospital of Putuo District Affiliated to Shanghai University of Traditional Chinese Medicine during January 2002 to January 2006 were enrolled in this study. Signs and symptoms and biochemical information of patients were collected by using a self-designed questionnaire regarding the four examinations of TCM. Signs and symptoms were firstly analyzed to find the frequency of occurrence. Then, the patients were divided into two groups according to non-existent or existent sign and symptom and the correlations between the signs and symptoms which occurred most frequently and their biochemical parameters were analyzed. RESULTS: Sixteen symptoms which occurred most frequently were fatigue, colored urine, liver palms, opaque complexion, string-like pulse, weakness at waist and knees, dry month and bitter taste in the mouth, profuse dreaminess and poor sleepiness, heaviness of limbs, abdominal distention, yellow eyes, fine pulse, impetuosity and susceptibility to rage, splenomegaly, poor appetite, and distension and fullness in the chest and hypochondrium. A previous study on syndrome differentiation of 900 patients with post hepatitic cirrhosis showed 4 syndrome patterns: internal accumulation of dampness-heat, liver-kidney yin deficiency, internal accumulation of blood stasis-heat, and liver depression and spleen deficiency. Further analysis showed that internal accumulation of dampness-heat syndrome was characterized by obvious hepatic inflammation, poor synthesis function and more ascites. Liver-kidney yin deficiency syndrome was characterized by low-level hepatic inflammation, poor synthesis function and more ascites. Internal accumulation of blood stasis-heat syndrome was characterized by low-level hepatic inflammation, poorer synthesis function, ascites and splenomegalia, and liver depression and spleen deficiency syndrome was characterized by slight hepatic inflammation, synthesis function injury, decreased internal portal vein diameter and less ascites. CONCLUSION: Different syndrome patterns have different pathological features, showing the complexity and polymorphism of syndrome construction.

Effects of vitamin E on mercuric chloride-induced renal interstitial fibrosis in rats and the antioxidative mechanism.

OBJECTIVE: To observe the effects of vitamin E (Vit E) on mercuric chloride (HgCl2)-induced renal interstitial fibrosis (RIF) in rats and discuss its antioxidative mechanism. METHODS: A total of 32 Sprague-Dawley rats were randomly assigned to three groups: normal group, model group and Vit E group. RIF was induced by oral administration of HgCl(2) at a dose of 8 mg/kg body weight once a day for 9 weeks. Rats in Vit E group were administered with Vit E capsule at 100 mg/kg body weight, and rats in normal and model groups were treated with normal saline. At the end of the 9th week, rats were sacrificed and renal hydroxyproline (Hyp)'s trichrome and periodic acid-silver methenamine (PASM) staining. The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and contents of glutathione (GSH) and malondialdehyde (MDA) in kidney tissue were tested with commercial kits. The expressions of nuclear factor-κB (NF-κB), inhibitor-κB (IκB), phospho-IκB (p-IκB) and tumor necrosis factor-α (TNF-α) were determined by Western blot. The expression of α-smooth muscle actin (α-SMA) was assayed by Western blot and immunofluorescent staining. RESULTS: Renal Hyp content, HE, Masson's trichrome and PASM staining results and α-SMA expression confirmed development of HgCl2-induced RIF in rats. Oxidative stress markers GSH, GSH-Px and MDA confirmed oxidative stress in RIF rats. Compared with model rats, rats in Vit E group had lower kidney Hyp content (P<0.01). GSH and MDA contents decreased significantly in Vit E group compared with model group (P<0.01). The expressions of NF-κB and IκB had no significant difference among all groups (P>0.05). In Vit E group, the expressions of p-IκB and TNF-α decreased significantly compared with model group (P<0.01). The expression of α-SMA in Vit E group was also decreased significantly compared with model group (P<0.01). CONCLUSION: Vit E has a protective effect on experimental RIF induced by HgCl(2) in rats and it is related to inhibition of lipid peroxidation, which involves blocking of NF-κB signaling pathway and the activation of cells producing extracellular matrix.

Salvianolic acid B prevents epithelial-to-mesenchymal transition through the TGF-beta1 signal transduction pathway in vivo and in vitro.

BACKGROUND: Salvianolic Acid B (Sal B) is a water-soluble component from Danshen (a traditional Chinese herb widely used for chronic renal diseases) with anti-oxidative and cell protective properties. Sal B also has potential protective effects on renal diseases. Tubular epithelial cells can undergo epithelial-to-mesenchymal transition (EMT), which plays an important role in the pathogenesis of renal interstitial fibrosis (RIF) and is mainly regulated by TGF-beta1/Smads pathway. The aims of the study are to investigate the effect of Sal B on tubular EMT in vivo and in vitro, and to elucidate its underlying mechanism against EMT related to TGF-beta1/Smads pathway. RESULTS: For in vivo experiments, RIF was induced in rats by oral administration of HgCl2 and prophylaxised with Sal B and vitamin E. The protein expression of E-cadherin was down-regulated, while the expression of alpha-SMA, TGF-beta1, TbetaR-I, p-Smad2/3 and the activity of matrix metalloproteinase-2 (MMP-2) were up-regulated in kidneys of model rats when compared with those of normal rats. In contrast, Sal B and vitamin E significantly attenuated the expression of alpha-SMA, TGF-beta1, TbetaR-I, p-Smad2/3, and MMP-2 activity, but increased E-cadherin expression. For in vitro experiments, HK-2 cells were incubated with TGF-beta1 to induce EMT, and the cells were co-cultured with 1 and 10 microM Sal B or SB-431542 (a specific inhibitor of TbetaR-I kinase). TGF-beta1 induced a typical EMT in HK-2 cells, while it was blocked by Sal B and SB-431542, as evidenced by blocking morphologic transformation, restoring E-cadherin and CK-18 expression, inhibiting alpha-SMA expression and F-actin reorganization, and down-regulating MMP-2/9 activities in TGF-beta1 mediated HK-2 cells. Furthermore, Sal B and SB-431542 profoundly down-regulated the expressions of TbetaR-I and p-Smad2/3 but prevented the decreased expression of Smad7 in TGF-beta1 stimulated HK-2 cells. CONCLUSIONS: Sal B can prevent tubular EMT in the fibrotic kidney induced by HgCl2 as well as HK-2 cells triggered by TGF-beta1, the mechanism of Sal B is closely related to the regulation of TGF-beta1/Smads pathway, manifested as the inhibition of TGF-beta1 expression, suppression of TbetaR-I expression and function, down-regulation of Smad2/3 phosphorylation, and restoration of the down-regulation of Smad7, as well as inhibition of MMP-2 activity.

[Effects of fuzheng huayu recipe in antagonizing rat's renal interstitial fibrosis].

OBJECTIVE: To observe the effects of Fuzheng Huayu Recipe (FHR) on rat's renal interstitial fibrosis induced by mercuric chloride (HgCl2), and to explore preliminarily its mechanism of action. METHODS: Rats were randomly divided into four groups: the normal group, the model group, the FHR group and the vitamin E group, the latter two were treated respectively by FHR 4.6 g/kg and vitamin E 100 mg/kg. Rats model was established by oral administration of 8 mg/kg HgCl2 for 9 weeks. Serum creatinine (Cr) and urea nitrogen (BUN) content were tested with corresponding test kits; hydroxyproline (Hyp) content in kidney was assayed with hydrochloric acid hydrolysis; renal histologic change was observed with HE, Masson and methenamine silver (PASM) staining; and collagen type I (Col I), as well as protein expressions of fibronectin (FN) and alpha-smooth muscle actin (alpha-SMA) was determined with Western blot. RESULTS: Compared with the model group, the kidney/body weight ratio, serum levels of Cr and BUN, kidney Hyp content, and severity of renal interstitial fibrosis in the two treated groups were significantly lower (P<0.05 or P<0.01), and the improvements were more significant in the FHR group than those in the vitamin E group; Col I and FN protein expression was also weaker in the two treated group (Col, P<0.05; FN, P<0.01); while the expression of alpha-SMA was lower in the FHR group (P<0.01), but it wasn't in the vitamin E group (P>0.05). CONCLUSION: FHR could improve the HgCl2-induced renal function injury in rats, decrease extracellular matrix deposition and restrain renal interstitial fibrosis, the mechanism of action might be related with its inhibitory effect on myofibroblast activation.

Fuzheng Huayu recipe and vitamin E reverse renal interstitial fibrosis through counteracting TGF-beta1-induced epithelial-to-mesenchymal transition.

AIM: To investigate the mechanism of action of Fuzheng Huayu recipe (FZHY) and vitamin E (Vit E) against renal interstitial fibrosis related to transforming growth factor-beta1 (TGF-beta1) mediated tubular epithelial-to-mesenchymal transition. MATERIALS AND METHODS: Renal interstitial fibrosis was induced by administration of HgCl(2) at a dose of 8 mg/kg body weight once a day for 9 weeks. Rats were randomly divided into four groups: normal, model, FZHY, and Vit E group. Rats in the latter two groups were treated with the FZHY recipe and Vit E respectively. HK-2 cells were treated with TGF-beta1 for 24h, followed by incubation with either SB-431542 (a potent and specific inhibitor of TbetaR-I kinase) or FZHY drug-containing serum for another 24h. Hyp content in rat kidney tissue was assayed with Jamall's method and collagen deposition in kidney was visualized using Masson stain. Protein expression of TGF-beta1, TbetaR-I, Smad2, p-Smad2, Smad3, and p-Smad3 was analyzed by Western blotting. Protein expression and the location of Smad3 in kidney was assayed by immunohistochemistry, E-cadherin, cytokeratin 18 (CK-18), alpha-SMA and TGF-beta1 by immunofluorescent stain. RESULTS: FZHY and Vit E inhibited renal collagen deposition and reduced Hyp content significantly. They upregulated E-cadherin protein expression and down-regulated the protein expression of alpha-SMA, TGF-beta1, p-Smad2, p-Smad3, and TbetaR-I. Lastly, they inhibited the nuclear translocation of Smad3 in fibrotic kidney tissue. FZHY drug-containing serum significantly upregulated the expression of CK-18 and down-regulated the expression of alpha-SMA, TbetaR-I, p-Smad2/3 in TGF-beta1 stimulated HK-2 cells. CONCLUSION: The mechanism of action of FZHY and Vit E against renal interstitial fibrosis is related to the reversal of tubular EMT induced by TGF-beta1.

[Effects of Fuzheng Huayu recipe on renal toxic interstitial fibrosis in rats].

OBJECTIVE: To investigate the effects of Fuzheng Huayu Recipe on renal interstitial fibrosis in rats induced by the toxic substances. METHODS: Forty Wistar rats were divided into 3 groups, including the normal group, model control and Fuzheng Huayu Recipe treated group. The renal fibrotic model was induced with intraperitoneal injection of dimethylnitrosamine (DMN) for 4 weeks and oral administration of food containing heavy metals such as Hg, etc for 8 weeks. From the 5th week, the rats were orally administered Fuzheng Huayu Recipe for 4 weeks. The renal tissue was stained with HE and periodic acid-silver metheramine, respectively. The serum creatinine and urea nitrogen were assayed with the kits, and the renal hydroxyproline (Hyp) contents in the homogenate of kidney tissue were measured with HCl hydrolysis. RESULTS: In the model group, the renal tubule was swollen, and the tubular epithelial cells were degenerated, necrosed and detached, with the infiltration of inflammatory cells; the tubular interstitium was expanded, characterized with the deposition of large collagens; the tubular basement membrane became thicker and wrinkled; and the renal Hyp content and serum creatinine and urea nitrogen levels increased remarkably as compared to the normal group. After the treatment with Fuzheng Huayu Recipe, the renal tubular inflammation and interstitial collagen deposition were alleviated, and the serum creatinine and urea nitrogen levels decreased significantly as compared to the model group. CONCLUSION: Administration of DMN and heavy metals can induce the renal interstitial fibrosis and impair the renal function. Fuzheng Huayu Recipe can improve the impaired renal functions and reverse the renal interstitial fibrosis.