Reversal of Sp1 transactivation and TGFß1/SMAD1 signaling by H2S prevent nickel-induced fibroblast activation.

Nickel as a heavy metal is known to bring threat to human health, and nickel exposure is associated with changes in fibroblast activation which may contribute to its fibrotic properties. H2S has recently emerged as an important gasotransmitter involved in numerous cellular signal transduction and pathophysiological responses. Interaction of nickel and H2S on fibroblast cell activation has not been studied so far. Here, we showed that a lower dose of nickel (200 µM) induced the activation of human fibroblast cells, as evidenced by increased cell growth, migration and higher expressions of α-smooth muscle actin (αSMA) and fibronectin, while high dose of nickel (1 mM) inhibited cell viability. Nickel reduced intracellular thiol contents and stimulated oxidative stress. Nickel also repressed the mRNA and protein expression of cystathionine gamma-lyase (CSE, a H2S-generating gene) and blocked the endogenous production of H2S. Exogenously applied NaHS (a H2S donor) had no effect on nickel-induced cell viability but significantly attenuated nickel-stimulated cell migration and the expression of αSMA and fibronectin. In contrast, CSE deficiency worsened nickel-induced αSMA expression. Moreover, H2S incubation reversed nickel-stimulated TGFß1/SMAD1 signal and blocked TGFß1-initiated expressions of αSMA and fibronectin. Nickel inhibited the interaction of Sp1 with CSE promoter but strengthened the binding of Sp1 with TGFß1 promoter, which was reversed by exogenously applied NaHS. These data reveal that H2S protects from nickel-stimulated fibroblast activation and CSE/H2S system can be a potential target for the treatment of tissue fibrosis induced by nickel.

Periplaneta Americana Extract May Attenuate Renal Fibrosis through Inhibiting Janus Tyrosine Kinase 2/Signal Transducer and Activator of Transcription 3 Pathway.

BACKGROUND: Periplaneta americana is one of the ancient insect groups with the strongest vitality. Periplaneta americana extract (PAE) has been explored as an alternative remedy for many diseases. Although much progress has been made in the study about PAE, the role of the drug in renal disease is rarely reported, especially in renal fibrosis. This study was designed to evaluate the renoprotective effect of PAE treatment to renal fibrosis. METHOD: An in vivo, unilateral ureteral obstruction (UUO) mouse model was built. Then the mice were treated with PAE (100 mg/kg body weight) once daily by oral gavage, again starting on the day of UUO and continued for 1 week. At the end of 1 week, the mice were sacrificed; kidney samples were collected for further analysis. In vitro, Boston University mouse proximal tubular cells were plated in 35-mm dishes at a density of 0.3 * 106 cells/dish. Then the cells were treated with 5-ng/mL TGF-ß1 in serum-free DMEM medium for an indicated length of time. The experimental groups were pretreated with the indicated concentrations of PAE (0.3125 mg/mL). The cells were further cultured for 24 h, and then cells were monitored morphologically or collected for biochemical analyses. RESULTS: Both in vivo and vitro PAE inhibits the expression of FN and alpha-smooth muscle actin and suppresses renal fibrosis. Importantly, PAE protects against renal fibrosis by inhibiting Janus tyrosine kinase 2 (JAK)/signal transducer and activator of transcription 3 (STAT) tyrosine phosphorylation. CONCLUSION: PAE attenuates renal fibrosis through the suppression of the JAK2/STAT3 pathway.

Inhibition of extracellular matrix production and remodeling by doxycycline in smooth muscle cells.

Alterations in the extracellular matrix (ECM) production and remodeling of smooth muscle cells (SMCs) have been implicated in processes related to the differentiation in atherosclerosis. Due to the anti-atherosclerotic properties of the tetracyclines, we aimed to investigate whether cholesterol supplementation changes the effect of doxycycline over the ECM proteins synthesis and whether isoprenylated proteins and Rho A protein activation are affected. SMC primary culture isolated from chicks exposed to atherogenic factors in vivo (a cholesterol-rich diet, SMC-Ch), comparing it with control cultures isolated after a standard diet (SMC-C). After treatment with 20 nM doxycycline, [H3]-proline and [H3]-mevalonate incorporation were used to measure the synthesis of collagen and isoprenylated proteins, respectively. Real-time PCR was assessed to determine col1a2, col2a1, col3a1, fibronectin, and mmp2 gene expression and the pull-down technique was applied to determine the Rho A activation state. A higher synthesis of collagens and isoprenylated proteins in SMC-Ch than in SMC-C was determined showing that doxycycline inhibits ECM production and remodeling in both SMC types of cultures. Moreover, preliminary results about the effect of doxycycline on protein isoprenylation and Rho A protein activation led us to discuss the possibility that membrane G-protein activation pathways could mediate the molecular mechanism.

Quercetin promotes cell apoptosis and inhibits the expression of MMP-9 and fibronectin via the AKT and ERK signalling pathways in human glioma cells.

Gliomas are the most common and malignant primary brain tumours and are associated with a poor prognosis despite the availability of multiple therapeutic options. Quercetin, a traditional Chinese medicinal herb, is an important flavonoid and has anti-cancer activity. Here, we evaluated whether quercetin could inhibit glioma cell viability and migration and promote apoptosis. The treatment of U87-MG glioblastoma and U251 and SHG44 glioma cell lines with different concentrations of quercetin inhibited cell viability in a dose-dependent manner. Wound healing assays indicated that quercetin significantly decreased glioma cell migration. ß-galactosidase staining, DNA staining and Annexin V-EGF/PI double staining assays demonstrated that quercetin promoted cell senescence and apoptosis. In addition, the protein levels of p-AKT, p-ERK, Bcl-2, matrix metallopeptidase 9 (MMP-9) and fibronectin (FN) were significantly reduced following quercetin treatment. Therefore, we conclude that quercetin might inhibit the viability and migration and promote the senescence and apoptosis of glioma cells by suppressing the Ras/MAPK/ERK and PI3K/AKT signalling pathways. Quercetin might be a potential candidate for the clinical treatment of glioma.

Diabetic nephropathy is resistant to oral L-arginine or L-citrulline supplementation.

Our recent publication showed that pharmacological blockade of arginases confers kidney protection in diabetic nephropathy via a nitric oxide (NO) synthase (NOS)3-dependent mechanism. Arginase competes with endothelial NOS (eNOS) for the common substrate L-arginine. Lack of L-arginine results in reduced NO production and eNOS uncoupling, which lead to endothelial dysfunction. Therefore, we hypothesized that L-arginine or L-citrulline supplementation would ameliorate diabetic nephropathy. DBA mice injected with multiple low doses of vehicle or streptozotocin (50 mg/kg ip for 5 days) were provided drinking water with or without L-arginine (1.5%, 6.05 g·kg(-1)·day(-1)) or L-citrulline (1.66%, 5.73 g·kg(-1)·day(-1)) for 9 wk. Nonsupplemented diabetic mice showed significant increases in albuminuria, blood urea nitrogen, glomerular histopathological changes, kidney macrophage recruitment, kidney TNF-α and fibronectin mRNA expression, kidney arginase activity, kidney arginase-2 protein expression, and urinary oxidative stress along with a significant reduction of nephrin and eNOS protein expression and kidney nitrite + nitrate compared with normal mice after 9 wk of diabetes. Surprisingly, L-arginine or L-citrulline supplementation in diabetic mice did not affect any of these parameters despite greatly increasing kidney and plasma arginine levels. These findings demonstrate that chronic L-arginine or L-citrulline supplementation does not prevent or reduce renal injury in a model of type 1 diabetes.

Selenium suppresses lipopolysaccharide-induced fibrosis in peritoneal mesothelial cells through inhibition of epithelial-to-mesenchymal transition.

Peritoneal fibrosis resulting from long-term clinical peritoneal dialysis has been the main reason of dropout from peritoneal dialysis. Peritonitis as a common complication of peritoneal dialysis treatment may lead to the occurrences of peritoneal fibrosis. We cultured peritoneal mesothelial cells with lipopolysaccharides (LPS) in order to stimulate the environment of peritonitis and investigate whether lipopolysaccharides could induce epithelial-to-mesenchymal transition (EMT). Oxidative stress could stimulate fibrogenesis while selenium has antioxidant properties. So, this study also explored whether selenium supplementation affects lipopolysaccharide-induced EMT and fibrosis. We found that lipopolysaccharides could activate EMT changes such as the loss of E-cadherin and the increase of α-smooth muscle actin (α-SMA), collagen I, vimentin, and fibronectin (FN), while selenium inhibits EMT by modulating reactive oxygen species (ROS) generation and ROS/MMP-9 signaling pathways in peritoneal mesothelial cells. Moreover, it was revealed that selenium decreased the EMT events of peritoneal mesothelial cells via inhibition of PI3k/AKT pathways. In conclusion, these findings enable a better understanding of the mechanism of peritoneal fibrosis and explore a new idea for the prevention and treatment.

[Effect of Moutan Cortex on AGEs-induced mesangial cell proliferation and basement membrane thickening].

OBJECTIVE: To investigate the effect of Moutan Cortex on mesangial proliferation and basement membrane thickening induced by advanced glycation end products (AGEs). METHOD: The glomerular mesangial cells (MC) injury model was established by inducing by AGEs. The cell were divided into 6 groups: the blank group ( BSA, 200 mg L-1) , the model group (AGEs, 200 mg L-1), the positive control group (AG, 10 mmol L L-1), and drug administration groups, namely the Moutan Cortex-treated high-dose group (2 x 10(-4) g mL(- 1)), the Moutan Cortex-treated medium-dose group (1 x 10(-4) g mL-1 ), and the Moutan Cortex-treated low-dose group (0. 5 x 10(-4) g . mL(-1)). The MTT method was performed to observe the effect of Moutan Cortex on the proliferation of MC. The content of fibronectin (FN) and collagen secretion 1V (Col IV) in cell supernatant were detected by ELISA kits. The western blot analysis was carried out to observe the FN expression. The Real-time PCR analysis was applied to examine the Col IV mRNA expression. RESULT: AGEs significantly increased AGEs-induced MC proliferation and FN and Col 1V secretion. The western blot analysis showed that MC could down-regulate the FN expression of MC secretion. According to the results of the real-time PCR assay, MC could down-regulate AGEs-induced MC secretion Col IV mRNA expression. CONCLUSION: MC had a certain protective effect on MC cultured under AGEs conditions. MC could remarkably inhibit the composition and secretion of Col IV and FN in matrix and the basement membrane thickening, and provide an experimental basis for the treatment of diabetic nephropathy.

The effect of Centella asiatica, vitamins, glycolic acid and their mixtures preparations in stimulating collagen and fibronectin synthesis in cultured human skin fibroblast.

Centella asiatica (Linn.) Urban is well known in promoting wound healing and provides significant benefits in skin care and therapeutic products formulation. Glycolic acid and vitamins also play a role in the enhancement of collagen and fibronectin synthesis. Here, we evaluate the specific effect of Centella asiatica (CA), vitamins, glycolic acid and their mixture preparations to stimulate collagen and fibronectin synthesis in cultured human fibroblast cells. The fibroblast cells are incubated with CA, glycolic acid, vitamins and their mixture preparations for 48 h. The cell lysates were analyzed for protein content and collagen synthesis by direct binding enzyme immunoassay. The fibronectin of the cultured supernatant was measured by sandwich enzyme immunoassay. The results showed that CA, glycolic acid, vitamins A, E and C significantly stimulate collagen and fibronectin synthesis in the fibroblast. Addition of glycolic acid and vitamins to CA further increased the levels of collagen and fibronectin synthesis to 8.55 and 23.75 µg/100 µg, respectively. CA, glycolic acid, vitamins A, E, and C, and their mixtures demonstrated stimulatory effect on both extra-cellular matrix synthesis of collagen and fibronectin in in vitro studies on human foreskin fibroblasts, which is beneficial to skin care and therapeutic products formulation.

Pentacyclic triterpenes from Manilkara bidentata resin. Isolation, identification and biological properties.

Three pentacyclic triterpenes were isolated for the first time from resinous plant Manilkara bidentata. Ultrasound-assisted extraction with ethanol was chosen after a comparison of various extraction methods. Analysis of the extract was performed by HPLC with evaporative light scattering detection and semi-preparative HPLC has enabled us to isolate two urs-12-enes (3ß-O-acetyl-α-amyrin and 3ß-O-trans cinnamyl-α-amyrin) and a lupane-type derivative (3ß-O-trans cinnamyl lupeol). Structures were elucidated on the basis of HRESIMS, atmospheric pressure photoionization MS, and homo- and heteronuclear correlation NMR experiments. Antioxidant and anti-inflammatory activities were determined on Manilkara extract and isolated fractions. We have also investigated their action on collagen and fibronectin synthesis, two very important proteins of the extracellular matrix. Thus, Manilkara extract was able to decrease IL-1ß and IL-8 pro-inflammatory cytokines. These activities exhibit the potential use of Manilkara extract as an anti-inflammatory and anti-aging ingredient for pharmaceutical and cosmetic industries.

A new multistep induction protocol for the transdifferentiation of bone marrow stromal stem cells into GABAergic neuron-like cells.

BACKGROUND: Bone marrow stromal stem cells (BMSC) are appropriate source of multipotent stem cells that are ideally suited for use in various cell-based therapies. It can be differentiated into neuronal-like cells under appropriate conditions. This study examined the effectiveness of co-stimulation of creatine and retinoic acid in increasing the differentiation of BMSC into GABAergic neuron-like cells (GNLC). METHODS: BMSC isolated from the femurs and tibias of adult rats were cultured in DMEM/F12 medium supplemented with 10% FBS, pre-induced using ß-mercaptoethanol ß-ME) and induced using retinoic acid (RA) and creatine. Immunostaining of neurofilament 200 kDa, neurofilament 160 kDa, nestin, fibronectin, Gamma-amino butyric acid (GABA) and glutamic acid decarboxylase (GAD) 65/67 were used to evaluate the transdifferentiation of BMSC into GNLC and to evaluate the effectiveness of pre-induction and induction assays. The expression of genes that encode fibronectin, octamer-binding transcription factor 4 (Oct-4), GAD 65/67 and the vesicular GABA transporter was examined in BMSC and GNLC by using RT-PCR assays during transdifferentiation of BMSC into GLNC. RESULTS: Co-stimulation with RA and creatine during the induction stage doubled the rates of GABAergic differentiation compared with induction using creatine alone, resulting in a 71.6% yield for GLNC. RT-PCR showed no expression of Oct-4 and fibronectin after the induction stage. CONCLUSION: The results of this study showed that the application of ß-ME, RA, and creatine induced the transdifferentiation of BMSC into GLNC.

Hydrogen peroxide-mediated oxidative stress and collagen synthesis in cardiac fibroblasts: blockade by tanshinone IIA.

ETHNOPHARMACOLOGICAL RELEVANCE: We have recently reported that tanshinone IIA attenuated cardiac fibrosis in two-kidney, two-clip renovascular hypertensive rats via inhibiting NAD(P)H oxidase. However, little is known about the cellular and molecular mechanisms of tanshinone IIA mediated anti-fibrotic effects in cardiac fibroblasts after H(2)O(2) stimulation. The present study was performed to investigate whether H(2)O(2) may increase collagen synthesis in cardiac fibroblasts by affecting the expression and activity of NAD(P)H oxidase and whether the effects of H(2)O(2) on cardiac fibroblasts can be blocked by treatment of tanshinone IIA. MATERIALS AND METHODS: Cardiac fibroblasts were treated with H(2)O(2) (100 µmol/L) in the presence or absence of tanshinone IIA (1 µmol/L), NAD(P)H oxidase inhibitors diphenyleneiodonium (10 µmol/L), siRNA-p47phox, siRNA-Nox2 and siRNA-Nox4. Collagen synthesis was measured by [(3)H]proline incorporation, O(2)(-) production were determined by flow cytometry and DHE fluorescence microscopy. NAD(P)H oxidase activity was measured by lucigenin-enhanced chemiluminescence. RESULTS: H(2)O(2) induced the activity of NAD(P)H oxidase, O(2)(-) production, collagen synthesis and fibronectin expression in cardiac fibroblasts, and DPI abolished this induction. Exposure of adult rat cardiac fibroblasts to H(2)O(2) had time-dependent increase in the expression of p47phox, Nox2 and Nox4 oxidases. In addition, tanshinone IIA significantly inhibited H(2)O(2)-induced collagen synthesis via attenuation of O(2)(-) generation and NAD(P)H oxidase activity. Moreover, siRNA-mediated knockdown of p47phox, Nox2 and Nox4 inhibited H(2)O(2)-induced NADPH oxidase activity. H(2)O(2)-induced collagen synthesis and fibronectin expression were also inhibited by p47phox, Nox2 and Nox4 knock down. CONCLUSIONS: Our data show that NAD(P)H oxidase plays a significant role in regulating collagen synthesis in H(2)O(2)-stimulated cardiac fibroblasts. Inhibition of NAD(P)H oxidase with tanshinone IIA completely blocked the H(2)O(2)-stimulated collagen production, which will raise the experimental basis for using tanshinone IIA to cardiac fibrosis in clinic.

EGCG inhibits CTGF expression via blocking NF-κB activation in cardiac fibroblast.

Connective tissue growth factor (CTGF) has been reported to play an important role in tissue fibrosis and presents a promising therapeutic target for fibrotic diseases. In heart, inappropriate increase in level of CTGF promotes fibroblast proliferation and extracellular matrix (ECM) accumulation, thereby exacerbating cardiac hypertrophy and subsequent failure. Epigallocatechin-3-gallate (EGCG), the major polyphenol found in green tea, possesses multiple protective effects on the cardiovascular system including cardiac fibrosis. However, the molecular mechanism by which EGCG exerts its anti-fibrotic effects has not been well investigated. In this study, we found that EGCG could significantly reduce collagen synthesis, fibronectin (FN) expression and cell proliferation in rat cardiac fibroblasts stimulated with angiotensinII (AngII). It also ameliorated cardiac fibrosis in rats submitted to abdominal aortic constriction (AAC). Moreover, EGCG attenuated the excessive expression of CTGF induced by AAC or AngII, and reduced the nuclear translocation of NF-κB p65 subunit and degradation of IκB-α. Subsequently, we demonstrated that in cardiac fibroblasts NF-κB inhibition could suppress AngII-induced CTGF expression. Taken together, these findings provide the first evidence that the effect of EGCG against cardiac fibrosis may be attributed to its inhibition on NF-κB activation and subsequent CTGF overexpression, suggesting the therapeutic potential of EGCG on the prevention of cardiac remodeling in patients with pressure overload hypertrophy.

[Dracorhodin perchlorate inhibit high glucose induce serum and glucocorticoid induced protein kinase 1 and fibronectin expression in human mesangial cells].

OBJECTIVE: To investigate the effect of dracorhodin perchlorate (DP) on inhibiting high glucose-induced serum and glucocorticoid induced protein kinase 1 (SGK1) and fibronectin (FN) expression in human mesangial cells (HMC), and its mechanism of prevention and treatment on renal fibrosis in diabetic nephropathy (DN) . METHOD: The HMC were divided into normal glucose group (NG group, 5.5 mmol x L(-1) D-glucose), normal glucose +low DP group (NG + LDP group, 5.5 mmol x L(-1) D-glucose +7.5 micromol x L(-1) DP), normal glucose +high DP group (NG + HDP group, 5.5 mmol x L(-1) D-glucose + 15 micromol x L(-1) DP), high glucose group (HG group,25 mmol x L(-1) D-glucose), high glucose +low DP group (HG + LDP group, 25 mmol x L(-1) D-glucose + 7.5 micromol x L(-1) DP)and high glucose +high DP group (HG +HDP group, 25 mmol x L(-1) D-glucose + 15 micromol x L(-1) DP). Each group was examined at 24 hours. The levels of SGK1 and FN mRNA was detected by real-time fluorescence quantitative PCR,and the expression of SGK1 and FN protein was detected by Western blot or indirect immunofluorescence. RESULT: A basal level of SGK1 and FN in HMC were detected in NG group, and the level of SGK1 and FN mRNA and protein were not evidently different compared to that of NG group adding 7.5 micromol x L(-1) DP for 24 hours. On the other hand, the levels of SGK1 and FN mRNA and protein were obviously decreased by adding 15 micromol x L(-1) DP for 24 hours. Compared to NG group, the levels of SGK1 and FN mRNA and protein were increased in HG group after stimulating for 24 hours (P < 0.01). Compared to HG group, the level of SGK1 and FN mRNA and protein were evidently reduced in HG + LDP and HG + HDP groups by adding 7.5 micromol x L(-1) DP and 15 micromol x L(-1) DP for 24 hours (P < 0.01). CONCLUSION: Dracorhodin perchlorate can inhibit high glucose-induced serum and glucocorticoid induced protein kinase 1 (SGK1) and fibronectin(FN) expression in human mesangial cells, and this may be part of the mechanism of preventing and treating renal fibrosis of DN.

Fibronectin synthesis by high glucose level mediated proliferation of mouse embryonic stem cells: Involvement of ANG II and TGF-beta1.

The role of individual supplements necessary for the long-term self-renewal of embryonic stem (ES) cells is poorly characterized in feeder/serum-free culture systems. This study sought to characterize the relationship between the effects of glucose on ES cell proliferation and fibronectin (FN) synthesis, and to assess the mechanisms responsible for these cellular effects of glucose. Treatment of the two ES cells (ES-E14TG2a and ES-R1) with 25 mM glucose (high glucose) increased the expression levels of FN mRNA and protein. In addition, high glucose and ANG II synergistically increased FN expression level, which coincident with data showing that high glucose increased the mRNA expression of angiotensin II (ANG II) type 1 receptor (AT(1)R), angiotensinogen, and FN, but not ANG II type 2 receptor. High glucose also increased the intracellular calcium (Ca(2+)) concentration and pan-protein kinase C (PKC) phosphorylation. Inhibition of the Ca(2+)/PKC pathway blocked high glucose-induced FN expression. High glucose or ANG II also synergistically increased transforming growth factor-beta1 (TGF-beta(1)) expression, while pretreatment with losartan abolished the high glucose-induced increase in TGF-beta(1) production. Moreover, TGF-beta(1)-specific small interfering RNA inhibited high glucose-induced FN expression and c-Jun N-terminal kinase (JNK) activation. The JNK inhibitor SP600125 blocked high glucose-induced FN expression and inhibited cell cycle regulatory protein expression induced by high glucose or TGF-beta(1). In this study, inhibition of AT(1)R, Ca(2+)/PKC, TGF-beta(1), JNK, FN receptor blocked the high glucose-induced DNA synthesis, increased the cell population in S phase, and the number of cells. It is concluded that high glucose increases FN synthesis through the ANG II or TGF-beta1 pathways, which in part mediates proliferation of mouse ES cells.

[Mechanism of the effect of Tongluo Recipe against glomerular sclerosis in rats].

OBJECTIVE: To investigate the effects of Tongluo Recipe on the expression of collagen IV (Col IV), fibronectin (FN), laminin (LN), transforming growth factor-beta1 (TGF-beta1) in rat renal tissues and explore the mechanism underlying these effects in rats with glomerular sclerosis. METHODS: The pathological changes in the renal tissues of rats with glomerular sclerosis were observed microscopically, and the expressions of Col IV, FN, LN, and TGF-beta1 were detected using immunohistochemical staining and image analysis system. RESULTS: Tongluo Recipe significantly decreased the expressions of Col IV, FN, LN and TGF-beta1 in the renal tissue of rats with glomerular sclerosis (P<0.05 or P<0.01) and obviously alleviated the renal pathologies (P<0.01). CONCLUSION: The therapeutic effects of Tongluo Recipe are probably mediated by lowered expressions of Col IV, FN, LN and TGF-beta1.

[Effects of yishen huoxue decoction on proliferating cell nuclear antigen expression and extracellular matrix in 5/6 nephrectomized rats].

OBJECTIVE: To investigate the effects and mechanisms of Yishen Huoxue decoction (YHD) on chronic renal failure (CRF) rats induced by 5/6 nephrectomy. METHODS: The glomerulosclerosis model was established by 5/6 nephrectomy in rats. Experimental animals were allocated into the normal group, the model group, the YHD group and the benazepril group. Urine protein of 24 h (UP) at the 6th and 12th weekend after operation, blood urea nitrogen (BUN) and creatinine (SCr), albumin (Alb) and haemoglobin (HB) at the 12th weekend were measured, renal pathology changes were examined with light microscope, the expressions of proliferating cell nuclear antigen (PCNA) and fibronectin (Fn) were examined by immunohistochemistry and mRNA expressions of connective tissue growth factor (CTGF) and plasminogen activator inhibitor-1 (PAI-1) by RT-PCR at the 12th weekend. RESULTS: Compared with those in the normal group, the levels of UP, BUN and SCr, the area of glomerular mesangial matrix, the FN deposition, PCNA expression in glomeruli and tubular interstitium and mRNA expressions of CTGF and PAI-1 were all significantly higher in the model group (P < 0.05). All the above-mentioned indexes were lower in the YHD group than those in the model group (P < 0.05). PCNA positively expressed cells in glomeruli of the normal, model group, YHD group and benazepril group was 7.00 +/- 2.24,34.78 +/- 6.96,15.75 +/- 2.61 and 15.50 +/- 2.57 respectively, positively correlated to the expression of CTGF, PAI-1, FN and SCr level. CONCLUSION: YHD could delay the progression of CRF in 5/6 nephrectomized rats, and the mechanisms were mainly related to the inhibition on renal cell proliferation and it induced over-expression of cytokines, and accumulation of extracellular matrix.

[Renal protective effect of Shenkang pill on diabetic rats].

OBJECTIVE: To investigate the effects of Shenkang pill on renal function and extracellular matrix secretion on the diabetic rats. METHOD: The diabetic rat models were induced by intraperitoneal injection of streptozotocin (STZ) and randomly divided into 3 groups' model control group; Capoten group and Shenkangwan group. Some normal other rats were used as normal control group. All rats were treated with corresponding drugs for 8 weeks. During and after the treatment, the general state, blood and urine glucose levels, excretion rate of the 24 hour urine protein and albumin, serum creatinine and blood urea nitrogen contents, kidney weight and relative kidney weight were measured. The mRNA of fibronectin(FN) in the kidney also detected by semi-quantitative reverse transcription polymerase chain reaction(RT-PCR). RESULT: Diabetes mellitus and renal lesions occurred in the three model groups. The expression of FN mRNA of the kidney in diabetic rats increased obviously. Shenkang pill could improve the general state and renal function of the diabetic rats, decrease the blood glucose levels and the excretion rate of the 24 hour urine protein and albumin, reduce the expression of FN mRNA in kidney. CONCLUSION: Shenkang pill has a certain protective effect on the diabetic kidney.

[Astragalus and angelica mixture inhibits the renal tubular epithelial cell injury induced by aristolochic-acid I].

OBJECTIVE: To investigate whether Chinese herb astragalus and angelica mixture (A&A) have influence on the renal tubular epithelial cell injury induced by aristolochic-acid I. METHODS: Human proximal tubular epithelial cell line HK-2 was pre-treated with AA-I (2.5 mg/L) for 4 hours. Cells were then treated with or without A&A for additional 44 hours. Cell apoptosis was evaluated by using FACS. Secreted fibronectin (FN) and TGF-beta1 levels were assayed by ELISA. The changes of AA-I-induced FN, TGF-beta1 level and the rate of apoptosis were compared before and after A&A treatment. RESULTS: There was basement secretion of TGF-beta1 by HK-2 cells (7.05+/-1.98 microg/L). Both normal serum (N-S) and A&A could not induce the cells to secret TGF-beta1(6.35+/-1.99 microg/L and 6.57+/-2.19 microg/L, vs. control, P>0.05). AA-I could induce the TGF-beta1 secretion by HK-2 cells (18.26+/-5.98 microg/L, vs. control, P<0.05). A&A could block AA-I-induced TGF-beta1 secretion by 63.5% (6.66+/-0.70 microg/L, vs. AA-I, P<0.05). It also suppressed AA-I-induced cell apoptosis by 93.7% (3.32%+/-0.41% vs. 19.19%+/-6.32% respectively, P<0.001) and FN secretion by 44% (1.64+/-1.11 folds vs. 2.93+/-0.87 folds respectively, P<0.05). CONCLUSION: A&A inhibits AA-I induced injury in human renal proximal tubule epithelial cells, whose mechanism may be partially through blocking TGF-beta1 secretion.

[Experimental study on effect of matrine in alleviating renal tubulointerstitial fibrosis in unilateral ureteral obstruction model].

OBJECTIVE: To observe the effects of matrine on renal tubulointerstitial fibrosis in unilateral ureteral obstruction (UUO) rat model and to explore its mechanisms. METHODS: Model rats of UUO were established and randomly divided into 6 groups: the normal group, the sham group, the UUO model group, the fusinopril treated group (F group), the high dose and low dose matrine treated groups (HM and LM). The expressions of matrix metalloproteinase-3 ( MMP-3), tissue inhibitor of metalloproteinase-1 (TIMP-1), fibronectin (FN), connective tissue growth factor (CTGF) and alpha-smooth muscle actin (alpha-SMA) were determined semi-quantitatively by immunohistochemistry on the 14th day. RESULTS: The expression of MMP-3 in the UUO model rats was significantly lowed on the 14th day, compared with that in rats not modeled in the normal and the sham groups (P < 0.05), but it was higher in the three treated groups than that in the UUO model group (P < 0.05); the expressions of TIMP-1, FN, CTGF and alpha-SMA were lower in the normal group and sham UUO group than those in all the UUO model groups, while they were lower obviously in the three treated groups than those in the UUO model group (P < 0.05); there was no significant difference in the above-mentioned indexes between HM group and F group (P < 0.05). CONCLUSION: Matrine could decrease the expressions of TIMP-1, FN, CTGF and alpha-SMA in the tubulointerstitium, and partly restore the expression of MMP-3, so as to delay the progression of renal tubulointerstitial fibrosis.

[Effect of shenkangwan on TGF-beta1 and FN in rat mesangial cells cultured by high glucose].

OBJECTIVE: To investigate the effect of Shenkangwan on secretion of transforming growth factor-beta1 (TGF-beta1) and Fibronectin (FN) and their mRNA expression in rat mesangial cells (MC)cultured by high glucose and explore its molecule mechanism on treating diabetic nephropathy (DN). METHODS: Rat MC were cultured in vivo and divided into six groups: low glucose group, high glucose group, normal rat serum group, Capoten tablets group, Shenkangwan high dose group and low dose group. Seventy two hours later, the secretion of TGF-beta1 and FN in MC were detected by enzyme linked immunosorbent assay (ELISA). The expression of TGF-beta1, and FN mRNA were determined by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: High gluscose could induce secretion of TGF-beta1 and FN in MC and increase expression of TGF-beta1 and FN mRNA. While Shenkangwan could repress these effects. CONCLUSION: Shenkangwan can suppress the secretion of the extracellular matrix in MC via TGF-beta1 signal way.