Search

Background: The biochemical response is a crucial indicator of prognosis in chronic hepatitis B (CHB) patients treated with nucleotide/nucleoside analogues (NAs). The impact of hepatitis D virus (HDV) infection on alanine aminotransferase normalization is elusive. Methods: The longitudinal study recruited 1185 CHB patients who received NAs. These patients were tested for anti-HDV antibody and HDV RNA at the initiation of anti-hepatitis B virus (HBV) therapy and annually for patients who were HDV-seropositive. ALT levels were examined at the first and second year of anti-HBV therapy. ALT abnormality was defined as ALT levels above 40 IU/mL in both male and female, and the risk factors associated with ALT abnormality were analysed. Results: Rates of seropositivity for anti-HDV and HDV RNA were 2.0% and 0.8% among 1185 NA-treated CHB patients, respectively. The strongest factor associated with ALT abnormality (>40 IU/mL) after first year treatment with NAs was HDV RNA seropositivity at year 1 (odds ratio [OR]/95% confidence interval [CI]: 31.44/3.49–283.56, P = 0.002), followed by liver cirrhosis (2.18/1.51–3.15, P

Background/Aims Hepatitis C Virus (HCV) infection is diagnosed by the presence of antibody to HCV and/or HCV RNA. This study aimed to evaluate the accuracy of anti-HCV titer (S/CO ratio) in predicting HCV viremia in patients with or without hepatitis B virus (HBV) dual infection. Methods Anti-HCV seropositive patients who were treatment-naïve consecutively enrolled. Anti-HCV antibodies were detected using a commercially chemiluminescent microparticle immunoassay. HCV RNA was detected by real-time PCR method. Results A total of 1321 including1196 mono-infected and 125 HBV dually infected patients were analyzed. The best cut-off value of anti-HCV titer in predicting HCV viremia was 9.95 (AUROC 0.99, P Conclusions Anti-HCV titer strongly predicted HCV viremia. This excellent performance could be generalized to either HCV mono-infected or HBV dually infected patients.

Background Mitochondrial dysfunction is a newly established risk factor for the development of renal fibrosis. Cell survival and injury repair is facilitated by mitochondrial biogenesis. Nuclear respiratory factor 1 (NRF-1) is a transcriptional regulation factor that plays a central role in the regulation of mitochondrial biogenesis. However, the transcription factor of this process in renal fibrosis is unknown. Thus, we hereby discussed the correlations of NRF-1 and renal interstitial fibrosis. Materials and methods In vitro fibrosis model was established by treatment with transforming growth factor-β1 (TGF-β1) in NRK-49F (Normal Rat kidney fibroblast). We investigated the ROS production, mitochondrial biogenesis and fibrogenic marker (e.q. fibronectin) during the progression of renal fibrosis by kit and Western blotting assay. Here, we used that two distinct mechanisms regulate NRF-1 activation and degradation of NRF-1. NRF-1 was transfect by pcDNA-NRF-1 overexpression gene to evaluate the NRF-1 activity of the therapeutic effect in renal fibrosis. In addition, NRF-1 was silenced by shRNA-NRF-1 to evaluate the significance of NRF-1. ELISA was used to evaluate the secreted fibronectin. Immunofluorescence staining was used to assay the in situ expression of proteins (e.g. fibronectin, NRF-1). Results Under renal fibrosis conditions, TGF-β1 (5 ng/ml) increased ROS. Simultaneously, TGF-β1-induced extracellular fibronectin by ELISA assay. In addition, TGF-β1 decreased expression of mitochondrial biogenesis. This is the first time to demonstrate that expression of NRF-1 is significantly decreased in renal fibrosis. However, NRK49F was a transfection with pcDNA-NRF-1 (2 μg/ml) expression vector dramatically reverse TGF-β1-induced cellular fibrosis concomitantly with the suppression of fibronectin (both intracellular and extracellular fibronectin). More importantly, transfection with shRNA-NRF-1 (2 μg/ml) significantly increased the expression of fibronectin of both intercellular and extracellular origins in NRK-49F cells. Discussion These finding suggest that NRF-1 plays a pivotal role on renal cellular fibrosis. Moreover, NRF-1 might act as a novel renal fibrosis antagonist by down-regulating fibrosis signaling in renal fibroblast cells.

Peroxisome proliferator-activated receptor coactivator-1 alpha (PGC-1α) is a transcriptional coactivator that regulates energy metabolism and mitochondrial biogenesis. Recently, mitochondrial dysfunction has been indicated as an established risk factor for the development of renal fibrosis. However, whether PGC-1α is involved in the pathogenesis of renal fibrosis is unknown. In this study, we treated NRK-49F (normal rat kidney fibroblast) cells with transforming growth factor-beta 1 (TGF-β1) for 24 h to establish an in vitro fibrosis model. TGF-β1 induced the upregulation of type I collagen, fibronectin, TGF-β receptor I (TGFβ-RI), TGFβ-RII, Smad4, and pSmad2/3, as well as PGC-1α. NRK-49F cells transfected with pcDNA-PGC-1α showed significantly increased expression of fibronectin and type I collagen, as revealed by western blot assay. Interestingly, transfection with PGC-1α-siRNA caused a stark reversal of TGF-β1-induced cellular fibrosis, with concomitant suppression of fibronectin and type I collagen, as revealed by western blot and immunofluorescence assays. Moreover, SB431542 (TGFβ-RI), LY294002 (PI3K/Akt), and SB203580 (p38 MAPK), inhibitors of TGF-β-associated pathways, markedly suppressed TGF-β1-induced PGC-1α upregulation. These results implicate a role of PGC-1α in renal interstitial fibrosis mediated via the TGFβ-RI, PI3K/Akt, and p38 MAPK pathways. Our findings that PGC-1α-siRNA downregulates fibronectin and type I collagen suggest that it can be used as a novel molecular treatment for renal fibrosis.

Fibrosis is the important pathway for end-stage renal failure. Glucose has been demonstrated to be the most important fibrogenesis-inducing agent according to previous studies. Despite diosgenin has been demonstrated to be anti-inflammatory, the possible role in fibrosis regulation of diosgenin remain to be investigated. In this study, renal proximal tubular epithelial cells (designated as HK-2) were treated with high concentration of glucose (HG, 27.5mM) to determine whether diosgenin (0.1, 1 and 10 μM) has the effects to regulate renal cellular fibrosis. We found that 10 μM of diosgenin exert optimal inhibitory effects on high glucose-induced fibronectin expression in HK-2 cells. In addition, diosgenin markedly inhibited HG-induced increase in α-smooth muscle actin (α-SMA) and HG-induced decrease in E-cadherin. In addition, diosgenin antagonizes high glucose-induced epithelial-to-mesenchymal transition (EMT) signals partly by enhancing the catabolism of Snail in renal cells. Collectively, these data suggest that diosgenin has the potential to inhibit high glucose-induced renal tubular fibrosis possibly through EMT pathway.

Thymic stromal lymphopoietin (TSLP) has previously been linked to allergic inflammatory diseases, and tissue fibrosis and organ dysfunction may also arise from such inflammation. It remains unclear, however, whether TSLP plays any role in the occurrence of renal fibrosis, so this study investigated that possibility. An in vitro fibrosis model was established by treating normal rat kidney fibroblast (NRK-49F) cells with transforming growth factor-β1 (TGF-β1), after which the levels of various fibrogenic markers (e.g., fibronectin) and downstream fibrogenic signal proteins (e.g., smad 7) were investigated. Also, TSLP shRNA was used to silence the effects of TSLP, while an ELISA was conducted to evaluate the fibronectin secretions. The level of fibronectin in the NRK-49F cells was dose- and time-dependently increased by the administration of exogenous TSLP (P

Renal fibrosis progresses to end stage of diabetes kidney disease, which causes irreversible progressive proximal tubular injury. In a previous study, 20-hydroxyecdysterone (20-HE), a phytoecdysteroid, attenuated renal injury in diabetes models. However, the fibrosis regulatory role remains to be investigated. Methods: The proximal tubular epithelial cells (designated as HK-2) were treated for 48 h with TGF-β1 (5ng/ml) in different concentrations of 20-HE (0 to 500 nM/ml) in the last 24 h of culture. The extracellular fibronectin was measured by ELISA assay. Western blot and immunofluorescence were used to evaluate the expression of TGF-β1/Smads transducer (including Smad2/3, 4, and 7), epithelial and mesenchymal markers (e.g. E-cadherin and α-smooth muscle actin) and Snail (transcriptional regulators for EMT). Results: 20-HE reverses TGF-β1-induced increase in fibronectin (both intracellular and extracellular fibronectin). Simultaneously, 20-HE reverses TGF-β1-induced down-regulation of Smad7. In addition, 20-HE significantly attenuates TGF-β1-induced upregulation of Smad2/3 and pSmad2/3, and downregulation of E-Cadherin. Moreover, 20-HE dramatically suppresses TGF-β1-induced increases in the expression of Snail. Conclusion: We propose that 20-HE is a potential fibrosis antagonist for renal proximal tubule cells. 20-HE might act through suppressing post-receptor signaling of TGF-β1 and restoring tubule epithelial character by blocking the expression of Snail.

Objective Oleanolic acid (OA) and its glycosides have been reported to prevent bone loss by inhibiting the formation of osteoclasts. However, because bone formation and resorption are balanced processes in bone metabolism, no studies have described the effect of OA on osteogenesis. The aim of the present study was to evaluate the osteoprotective effect of OA in rats with ovariectomy (OVX)-induced osteoporosis and to search for the molecular targets of OA in bone mesenchymal stem cells (bMSCs). Methods Two-month-old female mice that underwent OVX were treated with OA (20 mg/kg a day). After 2 weeks and after 3 months, bone mass was evaluated by micro-CT, morphometry, and immunohistochemical detection. In addition, the expression of 256 genes was measured via microarray and confirmed by real-time reverse transcription-polymerase chain reaction. The effects of OA on the activities of bMSCs were also observed in vitro using alkaline phosphatase and cell proliferation assays. Results Micro-CT displayed only a tendency for bone loss at 2 weeks but a decrease in bone mass at 3 months after OVX. OA treatment increased osteoblast number, increasing osteocalcin and runt-related protein 2 protein levels in vivo and facilitating the osteoblastic differentiation of bMSCs in vitro at doses of 10(-6) and 10(-5) M. Gene expression profile analysis revealed that OVX caused a marked dysregulation of gene expression, especially at 2 weeks, some of which was rescued by OA. Few of these genes overlapped, but their functions were involved in the Notch signaling pathway between two phases of the osteoporotic process. Conclusions OA exerts an osteoprotective effect in OVX-induced osteoporotic rats and stimulates the osteoblastic differentiation of bMSCs in vitro. The molecular mechanism of this effect might be related to the Notch signaling pathway and requires further investigation.

Background Hyperglycemia is the most important risk factor in the progression of renal fibrosis in diabetic kidney. Based on previous studies, interleukin-7 (IL-7) may exert antifibrotic activities in pulmonary fibrosis model. However, the role of IL-7 in the pathogenesis of renal tubulointerstitial fibrosis remains unclear. Thus, we hereby elucidate the effects of IL-7 in cultured renal proximal tubular epithelial cells (designated as HK-2) treated under hyperglycemic condition. Methods Cells were cultured in high glucose (27.5 mM) for 2 days. Different concentration of IL-7 (10, 50, 100 or 200 ng/ml) was added in the last 24 h of culture. ELISA was used to evaluate the secreted protein such as fibronectin and TGF-β 1 . Western blot was used to examine the EMT marker (including α-smooth muscle actin (α-SMA) and E-cadherin), signal transducer (including Smad Smad2/3 and Smad7) and EMT initiator (e.g. Snail). Immunofluorescence staining was used to assay the in situ expression of proteins (e.g. fibronectin and Snail). Results We found that IL-7 significantly attenuated high glucose-inhibited cellular growth and high glucose-induced fibrosis. More importantly, high glucose-induced up-regulation of fibronectin, TGF-β, TGF-β RII and pSmad2/3 was markedly inhibited by IL-7. On the contrary, high glucose-induced down-regulation of Smad7 was significantly reversed by IL-7 instead. IL-7 markedly inhibited high glucose-induced increase in α-SMA and Snail and decrease in E-cadherin. Conclusion We demonstrate that IL-7 has the potential to inhibit high glucose-induced renal proximal tubular fibrosis partly by modulating Smads and EMT pathway.

Introduction Multiple myeloma is a malignant immunoproliferative disorder with lipoprotein abnormalities. We report a case of falsely low concentrations of LDL-cholesterol (LDL-C) and artifactural undetectable HDL-cholesterol (HDL-C) as measured with direct methods in a patient of multiple myeloma with IgG κ monoclonal gammapathy and significant hyperlipidemia. Case report The patient had HDL-C and LDL-C concentrations in the 0.63–0.71 mmol/l and 2.22–2.36 mmol/l ranges, respectively, as measured by a traditional semi-quantitative electrophoresis method. The observation of falsely low concentrations of LDL-C and artifactural undetectable HDL-C might result in the mismanagement of patients of multiple myeloma with monoclonal gammapathy, because the LDL-C and HDL-C concentrations are positive and negative risk factors of cardiovascular diseases. Conclusions Care must be taken when using the homogenous method for direct measurement of LDL-cholesterol and HDL-cholesterol in patients of multiple myeloma with monoclonal paraprotein.

Diabetic nephropathy is characterized by early hypertrophy in both glomerular and tubuloepithelial elements. However, no studies to date have established a direct causal link between hyperglycaemia and renal hypertrophy. Our previous studies have found that high glucose does not induce cellular hypertrophy or expression of TGF-beta1 (transforming growth factor-beta1) in distal renal tubule cells [Yang, Guh, Yang, Lai, Tsai, Hung, Chang and Chuang (1998) J. Am. Soc. Nephrol. 9, 182-193]. In the present study, we used AGEs (advanced glycation end-products) to mimic long-term hyperglycaemia. Similar to glucose, AGEs did not induce TGF-beta1 mRNA in distal renal tubule cells [MDCK (Madin-Darby canine kidney) cells]; however, TGF-beta1 bioactivity was increased significantly. This result indicated post-translational regulation. Since TSP-1 (thrombospondin-1) has been demonstrated to activate latent TGF-beta1 in a variety of systems, the following experiments were performed. We found that AGEs dose-dependently increased both intracellular and extracellular levels of TSP-1. Purified TSP-1, like AGEs, increased the cellular protein content. Furthermore, anti-TSP-1 neutralizing antibodies attenuated the AGE-induced increase in TGF-beta1 bioactivity and hypertrophy. Thus TSP-1 might mediate AGE-induced distal renal tubule hypertrophy. In addition, we observed several putative transcription factor binding sites in the TSP-1 promoter, including those for AP-1 (activator protein-1), CREB (cAMP response element binding protein), NF-kappaB (nuclear factor-kappaB), SRF (serum response factor) and HSF (heat-shock factor), by sequence mapping. We used an enhancer assay to screen possible transcription factors involved. We showed that AP-1 and CREB were specifically induced by AGEs; furthermore, TFD (transcription factor decoy) for AP-1 could attenuate the AGE-induced increases in TSP-1 levels and cellular hypertrophy. Thus regulation of TSP-1 might be critical for hyperglycaemic distal tubule hypertrophy. Furthermore, TSP-1 TFD might be a potential approach to ameliorate diabetic renal hypertrophy.

Mitochondrial dysfunction is a newly established risk factor for the development of renal fibrosis. Cell survival and injury repair is facilitated by mitochondrial biogenesis. NRF-1 is a key transcription factor regulating the function of mitochondria.However, the role of this process in renal fibrosis is unknown. Thus, we hereby discuss the correlations of NRF-1 and renal interstitial fibrosis. Materials and Methods: NRK-49F (Normal Rat kidney fibroblast) cells were used. In vitro fibrosis model was established by treatment with transforming growth factor-b1 (TGF-b1). The fibrogenic marker (e.q. fibronectin) and fibrogenic signal proteins (e.q. pSmad2/3,Smad2/3,Smad7) were investigated. Here, we used that two distinct mechanisms regulate NRF1 activition and degradation of NRF1. NRF-1 was transfect by NRF-1 overexpression gene to evaluate NRF-1 activity of the therapeutic effect in renal fibrosis. In addition, NRF-1 was silenced by NRF-1 shRNA to evaluate the significance of NRF-1. ELISA was used to evaluate the secreted fibronectin. Western blot was used to examine the intracellular fibrogenic signal proteins (e.q. pSmad2/3,Smad2/3,Smad7). Immunofluorescence staining was used to assay the in situ expression of proteins (e.g. fibronectin). Results: Our study demonstrated that expression of NRF-1 is significantly decreased in renal fibrosis. However, transfection with pcDNA-NRF-1 (2mg/mL) expression vector did dramatically reverse TGF-b1-induced cellular fibrosis concomitantly with the suppression of fibronectin (both intracellular and extracellular fibronectin) and fibrogenic signal protein. Intriguingly, NRF-1 shRNA dramatically increased TGF-b1-induced cellular fibrosis. Collectively, these finding suggest that NRF-1 plays a pivotal role on renal cellular fibrosis. Moreover, NRF-1 might act as a novel renal fibrosis antagonist by down-regulating fibrosis signaling in renal fibroblast cells.

Renal interstitial fibrosis is characterized by increased extracellular matrix (ECM) synthesis. Epithelial-mesenchymal transition (EMT) in kidneys is driven by regulated expression of fibrogenic cytokines such as transforming growth factor-beta (TGF-β). Yam, or Dioscorea alata (DA) is an important herb in Chinese medicine widely used for the treatment of clinical diabetes mellitus. However, the fibrosis regulatory effect of DA is unclear. Thus, we examined TGF-β signaling mechanisms against EMT in rat fibroblast cells (NRK-49F). The characterization of DA water-extracts used various methods; after inducing cellular fibrosis in NRK-49F cells by treatment with β-hydroxybutyrate (β-HB) (10 mM), we used Western blotting to examine the protein expression in the TGF-β-related signal protein type I and type II TGF-β receptors, Smads2 and Smad3 (Smad2/3), pSmad2 and Smad3 (pSmad2/3), Smads4, Smads7, and EMT markers. These markers included E-cadherin, alpha-smooth muscle actin (α-SMA), and matrix metalloproteinase-2 (MMP-2). Bioactive TGF-β and fibronectin levels in the culture media were determined using ELISA. Expressions of fibronectin and Snail transcription factor, an EMT-regulatory transcription factor, were assessed by immunofluorescence staining. DA extract dose-dependently (50–200 µg/mL) suppressed β-HB-induced expression of fibronectin in NRK-49F cells concomitantly with the inhibition of Smad2/3, pSmad2/3, and Smad4. By contrast, Smad7 expression was significantly increased. DA extract caused a decrease in α-SMA (α-smooth muscle actin) and MMP-2 levels, and an increase in E-cadherin expression. We propose that DA extract might act as a novel fibrosis antagonist, which acts partly by down regulating the TGF-β/smad signaling pathway and modulating EMT expression.

Dysregulation of epithelial-to-mesenchymal transition (EMT) may contribute to renal fibrogenesis. Our previous study indicated that bone morphogenetic protein-2 (BMP-2) significantly reversed transforming growth factor (TGF)-β1-induced renal interstitial fibrosis. In this study, we examined the underlying mechanism and elucidate the regulation of EMT process under BMP-2 treatment. Cultured renal interstitial fibroblast (NRK-49F) was treated with TGF-β1 (10 ng/ml) with or without BMP-2 (10-250 ng/ml) for 24 h. The expression of α-smooth muscle actin (α-SMA), E-cadherin, fibronectin, or Snail transcriptional factors was analyzed by immunofluorescence staining or Western blotting. Cell migration was analyzed by wound-healing assay. NRK-49F treated with TGF-β1 induced significant EMT including upregulatioin of α-SMA, fibronectin, and snail proteins and down-regulation of E-cadherin. Interestingly, co-treatment with BMP-2 dose-dependently reversed TGF-β1-induced cellular fibrosis, cell migration, and above EMT change. The above effect was closely correlated with Snail since BMP-2 dose- and time-course dependently induced a significant decrease in the level of Snail. Moreover, Snail siRNA significantly reversed TGF-β1-induced increases in the level of α-SMA and fibronectin (intracellular and extracellular). We suppose that BMP-2 have the potential to attenuate TGF-β1-induced renal interstitial fibrosis by attenuating Snail expression and reversing EMT process.

Icariin (ICA) is an active component of Herba Epimedium effective in preventing osteoporosis. Bone mesenchymal stem cells (bMSCs) are an important target by which ICA promotes osteogenesis. However, its molecular mechanisms are poorly defined. In the present study, we induced osteoporosis in rats by corticosterone (CORT) and ovariectomy (OVX), treated both with ICA for 2 weeks or 3 months. As results, both models displayed bone loss tendency within 2 weeks and a significant bone loss after 3 months. ICA promoted bMSCs diffenentiation from CORT rat, and increased the secretion of osteocalcin, collagen I, runt-related transcription factor 2 in OVX model. Gene profile revealed a marked shift of gene expression by ICA, with much more significance in CORT rats. These potential molecular targets were involved in cell communication, adhesion, cycle and cytokines secretion. But very few genes overlapped in these two models, suggesting the effects and molecular mechanisms of ICA on osteoporosis might be pathogenesis-dependent. However, the Notch signaling pathway was common in both models, and should be paid close attention to for further study.

We evaluated the performance of a hepatitis C virus (HCV) antigen/antibody combination test [Murex HCV Antigen/Antibody Combination Test (Murex Ag/Ab test)] by comparing it with the current third-generation HCV antibody enzyme immunoassay (anti-HCV). A total of 403 serum samples were consecutively collected from four patient groups: healthy controls ( n =100); HCV-infected patients (HCV group, n =102); Human immunodeficiency virus (HIV)/HCV-infected patients (HIV/HCV group, n =100); and patients with uremia (uremia group, n =101). Performances were evaluated for the Murex Ag/Ab, anti-HCV, and HCV RNA in the HIV/HCV and uremia patient groups. In the HCV group, all 102 samples showed concordant positive and negative results for anti-HCV, Murex Ag/Ab, and HCV RNA tests. In the HIV/HCV group, all 100 samples were positive for both anti-HCV and Murex Ag/Ab tests, whereas 88 patients (88%) were HCV RNA positive. In the uremia group, 14 (69.0%) of the 23 anti-HCV-positive patients were HCV RNA positive, whereas 14 (77.8%) of the 18 Murex Ag/Ab–positive patients were HCV RNA positive. None of anti-HCV-negative or Murex Ag/Ab–negative patients were HCV RNA positive. Based on the HCV RNA assay, the sensitivities for both anti-HCV and Murex Ag/Ab assays were 100%, whereas the specificities of these two assays were 89.7% and 95.4%, respectively. With good sensitivity and specificity, the Murex Ag/Ab assay could be a useful alternative diagnostic tool, especially in immunocompromised populations, such as patients with uremia or those infected with HIV.