Jiyao Wang

Inhibition by curcumin of multiple sites of the transforming growth factor-beta1 signalling pathway ameliorates the progression of liver fibrosis induced by carbon tetrachloride in rats

BackgroundAt present there is no effective and accepted therapy for hepatic fibrosis. Transforming growth factor (TGF)-β1 signaling pathway contributes greatly to hepatic fibrosis. Reducing TGF-β synthesis or inhibiting components of its complex signaling pathway represent important therapeutic targets. The aim of the study was to investigate the effect of curcumin on liver fibrosis and whether curcumin attenuates the TGF-β1 signaling pathway.MethodsSprague–Dawley rat was induced liver fibrosis by carbon tetrachloride (CCl4) for six weeks together with or without curcumin, and hepatic histopathology and collagen content were employed to quantify liver necro-inflammation and fibrosis. Moreover, the mRNA and protein expression levels of TGF-β1, Smad2, phosphorylated Smad2, Smad3, Smad7 and connective tissue growth factor (CTGF) were determined by quantitative real time-PCR, Western blot, or immunohistochemistry.ResultsRats treated with curcumin improved liver necro-inflammation, and reduced liver fibrosis in association with decreased α-smooth muscle actin expression, and decreased collagen deposition. Furthermore, curcumin significantly attenuated expressions of TGFβ1, Smad2, phosphorylated Smad2, Smad3, and CTGF and induced expression of the Smad7.ConclusionsCurcumin significantly attenuated the severity of CCl4-induced liver inflammation and fibrosis through inhibition of TGF-β1/Smad signalling pathway and CTGF expression. These data suggest that curcumin might be an effective antifibrotic drug in the prevention of liver disease progression.

Impact of nucleos(t)ide analogues on the estimated glomerular filtration rate in patients with chronic hepatitis B: a prospective cohort study in China.

Chronic hepatitis B therapy with nucleos(t)ide analogues, particularly tenofovir or adefovir, may affect renal function. To date, there has not been a head‐to‐head controlled study to assess estimated glomerular filtration rate (eGFR) fluctuations in nucleos(t)ide‐treated CHB patients. We aimed to evaluate the long‐term effects of nucleos(t)ide on eGFR in Chinese patients with chronic hepatitis B. This prospective cohort study included 275 patients. Patient subgroups included those treated with lamivudine (n = 50), adefovir (n = 60), telbivudine (n = 68) and entecavir (n = 61); untreated patients (n = 36) served as control. After an average follow‐up duration of 23 months, eGFR calculated by Cockcroft–Gault and Modification of Diet in Renal Disease formulas increased by 18.35 mL/min and 19.34 mL/min (P < 0.0001) in the telbivudine group, respectively, and decreased by 10.95 mL/min and 12.17 mL/min (P = 0.0001) in the adefovir group, respectively. Even if renal function was normal or mildly impaired at baseline, eGFR increased significantly more in the telbivudine group than in the other groups (P < 0.001). More patients in the adefovir group (23%) had a ≥20% decrease in eGFR than the other groups (P < 0.0001). More patients in the telbivudine group (31%) had a ≥20% increase in eGFR than the other groups (P < 0.0001). In conclusion, prolonged telbivudine therapy resulted in improved eGFR, while adefovir therapy was associated with decreased eGFR. Lamivudine and entecavir therapy did not significantly influence eGFR.

Peroxiredoxin 2: a potential biomarker for early diagnosis of Hepatitis B Virus related liver fibrosis identified by proteomic analysis of the plasma

BackgroundLiver fibrosis is a middle stage in the course of chronic Hepatitis B virus (HBV) infection, which will develop into cirrhosis and eventually hepatocellular carcinoma (HCC) if not treated at the early stage. Considering the limitations and patients reluctance to undergo liver biopsy, a reliable, noninvasive diagnostic system to predict and assess treatment and prognosis of liver fibrosis is needed. The aim of this study was to identify biomarkers for early diagnosis of HBV related liver fibrosis.MethodPlasma samples from 7 healthy volunteers and 27 HBV infected patients with different stages of fibrosis were selected for 2-DIGE proteomic screening. One-way ANOVA analysis was used to assess differences in protein expression among all groups. The alteration was further confirmed by western blotting. Plasma levels of 25 serological variables in 42 healthy volunteers and 68 patients were measured to establish a decision tree for the detection of various stages fibrosis.ResultThe up-regulated proteins along with fibrosis progress included fibrinogen, collagen, macroglobulin, hemopexin, antitrypsin, prealbumin and thioredoxin peroxidase. The down-regulated proteins included haptoglobin, serotransferrin, CD5 antigen like protein, clusterin, apolipoprotein and leucine-rich alpha-2-glycoprotein. For the discrimination of milder stage fibrosis, the area under curve for Prx II was the highest. Four variables (PT, Pre, HA and Prx II) were selected from the 25 variables to construct the decision tree. In a training group, the correct prediction percentage for normal control, milder fibrosis, significant fibrosis and early cirrhosis was 100%, 88.9%, 95.2% and 100%, respectively, with an overall correct percent of 95.9%.ConclusionThis study showed that 2-D DIGE-based proteomic analysis of the plasma was helpful in screening for new plasma biomarkers for liver disease. The significant up-expression of Prx II could be used in the early diagnosis of HBV related liver fibrosis.

Glycyrrhizin regulates CD4 + T cell response during liver fibrogenesis via JNK, ERK and PI3K/AKT pathway☆

The aims of this study were to elucidate the immunomodulatory effects of glycyrrhizin (GL) on CD4(+)T cell responses during liver fibrogenesis. To obtain in vivo evidence about the effects of GL on CD4(+)T cells in livers and spleens of concanavalin A (ConA)-induced mouse model, mice were administrated with ConA together with or without GL for 8 weeks. Mice treated with GL dramatically prevented liver inflammation and fibrosis. Besides, GL inhibited the infiltration of T helper (Th) cell type 1, Th2, Th17 and regulatory T cells (Treg) in livers and spleens of mouse fibrosis models, and regulated the Th1/Th2 and Treg/Th17 balances respectively to a relative dominance of Th1 and Treg lineages in livers. Moreover, GL dramatically enhanced the antifibrotic cytokine interferon (IFN)-γ and interleukin (IL)-10. GL at a concentration of 10 or 100 μg/mL was respectively incubated with ConA-stimulated splenic CD4(+)T cells in vitro, and JNK inhibitor (SP600125), ERK inhibitor (U0126), p38 inhibitor (SB203580) or PI3K/AKT inhibitor (LY29400225) was added during the incubation. Notably, GL not only inhibited ConA-induced proliferation of splenic CD4(+)T cells but also enhanced the mRNAs of IFN-γ and IL-10 in these cells. Be similar to the effects of GL, SP600125, U0126 and LY29400225, however not SB203580, also inhibited ConA-induced CD4(+)T cell proliferation, indicating the involvement of JNK, ERK and PI3K/AKT in this process. Moreover, GL significantly inhibited ConA-induced phosphorylation of JNK, ERK and PI3K/AKT in vitro. Collectively, GL might alleviate liver injury and fibrosis progression via regulation of CD4(+)T cell response in JNK, ERK and PI3K/AKT-dependent pathways.

Enhanced high-mobility group box 1 (HMGB1) modulates regulatory T cells (Treg)/T helper 17 (Th17) balance via toll-like receptor (TLR)-4-interleukin (IL)-6 pathway in patients with chronic hepatitis B.

High‐mobility group box 1 (HMGB1) proteins are substantially up‐regulated in acute and chronic hepatitis. However, the immunopathogenic role of HMGB1 in patients with chronic hepatitis B (CHB) has not been elucidated. In this study, using a cohort of 36 CHB patients, we demonstrated a crucial role for HMGB1 to modulate balance between regulatory T (Treg) and T helper 17 (Th17) cells via the toll‐like receptor (TLR)‐4‐interleukin (IL)‐6 pathway. Serum HMGB1 levels were dramatically higher in CHB patients and increased along with liver injury, inflammation and fibrosis. Notably, HMGB1 increased along with decreased Treg/Th17 cells ratios in the periphery or intrahepatic microenvironment, which provides a clue for HMGB1 to favour Th17 responses whereas inhibit Treg responses. For in vitro studies, serum pools were constructed with serum from CHB patients at an advanced stage, whereas peripheral blood mononuclear cells (PBMC) pools were constructed with cells from those at an early stage. CHB‐serum significantly enhanced retinoic acid‐related orphan receptor‐γt (RORγt), whereas they inhibited forkhead box P3 (Foxp3) expression in CHB‐PBMC, which could be reversed by blocking of HMGB1, TLR4, or IL‐6. Besides, recombinant HMGB1 (rHMGB1) dose‐dependently up‐regulated RORγt whereas down‐regulated Foxp3 expression in CHB‐PBMC, and meanwhile, rHMGB1 enhanced TLR4 and IL‐6 expression in CHB‐PBMC. Moreover, the axis of HMGB1–TLR4‐IL‐6–Treg/Th17 required noncontact interactions between CD4 and non‐CD4 cells. In addition, rHMGB1 down‐regulated anti‐inflammatory proteins on CD4+CD25+ cells whereas up‐regulated pro‐inflammatory cytokines in CD4+CD25− cells. In summary, enriched HMGB1 in CHB patients shifts Treg/Th17 balance to Th17 dominance via the TLR4‐IL‐6 pathway, which exacerbates liver injury and inflammation.

Early diagnosis and therapeutic choice of Klebsiella pneumoniae liver abscess

Nowadays, pyogenic liver abscess (PLA) is still a common and severe intra-abdominal infection, and Klebsiella pneumoniae had emerged as the most common pathogenic bacteria worldwide in the past ten years. Our study aims to achieve an early pathogenic diagnosis and rational therapy modality for Klebsiella pneumoniae liver abscess (KLA) through clinical data analysis. A total of 197 inpatients in Zhongshan Hospital, Shanghai, diagnosed as having liver abscess between March 2001 and September 2009 were enrolled. Patients with monomicrobial infection were divided into two groups: patients with K. pneumoniae liver abscess (KLA group, n = 106) and those with non-Klebsiella pneumoniae liver abscess (NKLA group, n = 56). A retrospective analysis was made between these two groups on the aspects of underlying diseases, clinical characteristics, laboratory data, culture results, and imaging findings. To evaluate the effects of different medical interventions, monomicrobial KLA patients were further divided into four subgroups (percutaneous liver aspiration, aspiration plus antibiotics flushing, aspiration plus retained catheter, and aspiration plus antibiotics flushing and retained catheter), and corresponding therapeutic effects were analyzed. KLA was more likely to occur in patients with coexisting diseases such as diabetes mellitus (53.77% vs 25.00%, P = 0.001) and hepatic adipose infiltration (16.04% vs 5.36%, P = 0.029). Compared to NKLA group, clinical characteristics including abdominal pain (40.57% vs 57.14%, P = 0.044), hypodynamia (19.81% vs 46.43%, P = 0.001), and hepatomegaly (4.72% vs 14.29%, P = 0.033) were much milder, but with a higher fasting blood glucose level (7.84 ± 0.36 vs 5.76 ± 0.30, P = 0.001) on admission in KLA group. In addition, KLA abscess often appeared singly in the right lobe of the liver with gas forming nature (32.88% vs 13.51%, P = 0.039), unsmooth rim (71.23% vs 40.54%, P = 0.002), and dynamic septum enhancement (41.10% vs 16.22%, P = 0.009). Compared to mono aspiration subgroup, additional antibiotic flushing could not further improve clinical outcomes of KLA patients (P > 0.05); however, the retained catheter showed obvious advantage in reducing abscess diameter (34.38 ± 3.25 mm vs 22.67 ± 2.37 mm, P = 0.017). It can be concluded that the strong association with diabetes, milder clinical symptoms, and gas-forming nature in CT images makes early pathogenic diagnosis of KLA possible. Comparatively, ultrasonography-guided percutaneous liver aspiration with retained catheter may be the most rational intervention modality of KLA.

Serum microRNA-124 is a novel biomarker for liver necroinflammation in patients with chronic hepatitis B virus infection

Patients with chronic hepatitis B virus (HBV) infection and normal or mildly increased transaminases may have sustained significant liver damage, as verified by liver biopsy. However, no suitable noninvasive method exists for identifying liver necroinflammation in such patients. We aimed to investigate the power of microRNA‐124 as a novel biomarker for liver necroinflammation. A total of 131 recruited patients with chronic HBV infection underwent liver biopsy for grading of necroinflammation (G) and staging of fibrosis (S). Thirty healthy individuals were included as controls (HCs). Serum microRNA‐124 and microRNA‐122 levels were measured using qRT‐PCR. Forty‐five patients from the study population receiving entecavir therapy were monitored for changes in serum microRNA‐124 levels in association with improved liver histology. The capacity of serum microRNA‐124 levels in discriminating the grade of liver necroinflammation was compared with alanine aminotransferase (ALT) with liver biopsy validation. Serum microRNA‐124 levels were significantly higher in patients with chronic HBV infection than in HCs (P < 0.0001). Patients with considerable liver necroinflammation (G ≥ 2) had significantly higher serum miRNA‐124 levels than those without or with mild necroinflammation (P < 0.0001). After 48 weeks of antiviral therapy, serum microRNA‐124 levels considerably declined in 45 patients (P < 0.0001), which were associated with histological improvement. In patients with normal ALT and a serum HBV DNA load >104 copies/mL, receiver operating characteristic (ROC) curve of serum microRNA‐124 levels yielded an area under ROC curve (AUC) of 0.840, with 58.3% sensitivity and 91.7% specificity in discriminating between moderate‐to‐severe liver necroinflammation (G ≥ 2).

Expression, localization and possible functions of aquaporins 3 and 8 in rat digestive system.

Although aquaporins (AQPs) play important roles in transcellular water movement, their precise quantification and localization remains controversial. We investigated expression levels and localizations of AQP3 and AQP8 and their possible functions in the rat digestive system using real-time polymerase chain reactions, western blot analysis and immunohistochemistry. We investigated the expression levels and localizations of AQP3 and AQP8 in esophagus, forestomach, glandular stomach, duodenum, jejunum, ileum, proximal and distal colon, and liver. AQP3 was expressed in the basolateral membranes of stratified epithelia (esophagus and forestomach) and simple columnar epithelia (glandular stomach, ileum, and proximal and distal colon). Expression was particularly abundant in the esophagus, and proximal and distal colon. AQP8 was found in the subapical compartment of columnar epithelial cells of the jejunum, ileum, proximal colon and liver; the most intense staining occurred in the jejunum. Our results suggest that AQP3 and AQP8 play significant roles in intestinal function and/or fluid homeostasis and may be an important subject for future investigation of disorders that involve disruption of intestinal fluid homeostasis, such as inflammatory bowel disease and irritable bowel syndrome.

Clinical analysis of 275 cases of acute drug-induced liver disease

In order to analyze the causative drugs, clinical manifestation and pathological characteristics of the patients with acute drug-induced liver disease, from January 2000 to December 2005, 275 cases diagnosed as acute drug-induced liver diseases according to Maria Criterion and hospitalized in Zhongshan Hospital of Fudan University were retrospectively reviewed. Each was determined by drug history, clinical symptoms and signs, laboratory tests and therapeutic effects. In 41 cases, the diagnosis was confirmed by liver biopsy. The proportion of acute drug-induced liver disease among all of the acute liver injuries was annually increased. The most common drugs which induced acute liver injuries were traditional Chinese herb medicine (23.3%, 64/275 cases), antineoplastic (15.3%, 42/275), hormones and other immunosuppressant agents (13.8%, 38/275), antihypertensive drugs and other cardiovascular drugs (10.2%, 28/275), NSAIDs (8.7%, 24/275) respectively. Hepatocellular injury was the predominant type in these cases (132 cases, 48%). The principal clinical manifestation included nausea (54.8%), fatigue (50.2%), jaundice (35.6%). 27.9% patients were asymptomatic. Most patients were cured with good prognosis. The total effective rate was 94.2% after treatment. The clinicians should pay attention to the prevention, diagnosis and therapy of drug-induced liver disease.

Natural history of serum HBV-RNA in chronic HBV infection

Virus‐like particles encapsulating HBV‐RNA represent a serum biomarker for assessing viral replication activity in clinical practice. However, baseline levels of serum HBV‐RNA and their associations with viral replicative intermediates and liver disease in phases of chronic hepatitis B remain unknown. In this cross‐sectional study, 102 patients were categorized into immune‐tolerant (IT), HBeAg‐positive immune active (HBeAg+IA), inactive carrier (IC) and HBeAg‐negative immune active (HBeAg‐IA) phases. HBV‐RNA in serum samples and in 66 paired liver biopsies were quantified and correlated with serum ALT levels, histopathological scores and the levels of other viral replicative intermediates. Mean levels of serum HBV‐RNA differed among phases, with the highest levels among IT (6.78 ± 0.83 log10 copies mL−1) patients, followed by HBeAg+IA (5.73 ± 1.16 log10 copies mL−1), HBeAg‐IA (4.52 ± 1.25 log10 copies mL−1) and IC (2.96 ± 0.40 log10 copies mL−1) patients. Serum HBV‐RNA levels correlated with HBV DNA in all phases, although correlations with other viral replicative intermediates weakened or disappeared when cases were stratified into phases. Distinct compositions of viral products were found among phases: the ratio of HBsAg to serum HBV‐RNA was highest in IC patients, while the ratio of serum HBV‐RNA to intrahepatic HBV‐RNA and the ratio of intrahepatic HBV‐DNA to intrahepatic HBV‐RNA were significantly higher in IT patients. In conclusion, baseline levels of HBV‐RNA and the composition of viral replicative intermediates differ significantly across the natural course of chronic HBV infection. These findings shed light on the nature of viral replication and pathogenesis of disease among different phases of chronic HBV infection.