Clinical characteristics of 638 patients infected with hepatitis C virus by post blood transfusion,in Shanghai / 中华流行病学杂志

Objective To investigate me clinical-epidemiologic characteristics of patients with hepatitis C virus(HCV)infection by post blood transfusion.Methods Polymerase chain reaction (PCR)and enzynle linked immunosorbent assay(ELISA)were used to detect HCV RNA and anti.HCV,respectively.Analysis was performed on patients'age distribution,cause of primary diseases,years ofexposure,ingredient and amount of transfusion,incubation period,disorder on liver function and changes on abdominal ultrasound image,etc.Results HCV RNA levels were higher than 3.0log10 copy/ml in 90.8%infected patients、with a median as 6.10 log10 copy/ml.19.2%of the patients showed viral load 3.0 to 4.0 iog10 copy/ml,and 66.1%of them showed 5.0 to 6.0 log10 copy/ml.Only 14.7%of the infected persons had HCV RNA levels higher than 7.0 log10 copy/ml.Eighty-one point five percent(44/54)of the infected persons were confirmed as HCV RNA positive by HCV RNA qualitative analysis with HCV genotype as primarily type 1.99.8%(636/637)of the pmients were detected as anti-HCV positive by serological test.The sensitivity of serological test was higher than both quantitative and qualitative HCV RNA assays(P=0.000,P=0.000,respectively).HCV infection post blood transfusion was more seen in common people at 40 to 60 years old Most cases(85.7%)had their first exposure during 1990 to 1994.More than 10% of the cases had primary diseases aS obstetrics,orthopedics or gastrointestinal tract hemorrhage.79.9%of the patients received whole blood product transfusion.The mean interval between transfusion and clinical diagnosis was 8.5±5.5 years.90.1%of the infected patients had liver function damage,while most of them showed elevated alanine aminotransferase(ALT)no more than 5 upper limits of normal(ULN).wheteas Serum total bilirubin(TBIL).ALT and aspartate aminotransferase(AST)≥5×ULN level were showing more clinicaI manifestations(P=0.000.P=0.001,P=0.009,respectively).Abdominal ultrasound among 8.9%of the infected persons showed changes in cirrhosis,and most of them werc older than 50years of age.Conclusion Most of the post transfusion HCV infected cases happened in adulthood,and were mainly exposed during 1990 to 1994.Infected pmients usually had their liver function damaged with elevated ALT no more than 5×ULN and with medium HCV RNA levels.HCV genotype was mainly for type 1.Patients who weTe ofolder age showed higher incidence ofcirrhosis.If a patients'infection period Was longer than 5 years,he/she would show higher incidence of cirrhosis.

Results of 3 years of continuous entecavir treatment in nucleos(t)ide-naive chronic hepatitis B patients / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To evaluate the virological, serological and biochemical outcomes of 3 years of entecavir (ETV) treatment in nucleoside-naive chronic hepatitis B patients.</p><p><b>METHODS</b>This study was divided into two stages: Patients receiving either ETV 0.5 mg/d (n = 258) or lamivudine (LAM) 100 mg/d (n = 261) entered the initial 96-week randomized, double blind, controlled efficacy study. Patients not achieving a consolidated response (HBV DNA less than 0.7 MEq/ml, ALT less than 1.25 times*ULN, and if HBeAg-positive at baseline, loss of HBeAg for >or= 24 weeks), or those experienced viral breakthrough or relapse, entered a 48-week entecavir rollover study.</p><p><b>RESULTS</b>96 weeks after the treatment, 79% of ETV treated and 46% of LAM treated patients had HBV DNA less than 300 copies/ml (P < 0.0001), 96% of ETV treated and 92% of LAM treated patients had normalized ALT (P = 0.06). 21% of ETV treated and 23% of LAM treated patients achieved HBeAg seroconversion. Among the 160 patients received continuous ETV for 144 weeks, 89% had undetectable serum HBV DNA, 86% showed ALT normalization, and 27% achieved HBeAg seroconversion. ETV resistance was rare: only 3 patients showed ETV resistance 96 weeks after the treatment, and additional 2 patients developed ETV resistance during the following 48 weeks, genotyping indicated the ETV resistance was caused by gene mutation. Adverse event rates in ETV-treated patients were similar to those in LAM-treated patients, but fewer ALT flares were observed in ETV-treated patients.</p><p><b>CONCLUSIONS</b>This study demonstrates that ETV treatment results in long-term HBV suppression and ALT normalization in Chinese CHB patients, and is associated with low rate of drug resistance.</p>

Effects of CpG-ODN combined with HBsAg on the phenotype, function and the activity of NF-kappa B and AP-1 of monocyte-derived dendritic cells in chronic hepatitis B patients / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effect of synthetic non-methylated CpG-ODN combined with recombined HBsAg on the phenotype, function and the expression level of nucleic transcription factor NF-kappa B (NF-kB) and AP-1 of monocyte-derived dendritic cells (DC) in patients with chronic hepatitis B (CHB).</p><p><b>METHODS</b>Purified monocytes were isolated from the peripheral blood of CHB patients and healthy volunteers and cultured with human granulocyte-monocyte colony stimulating factor added together with interleukin-4. On the fifth day of culture, CpG-ODN, HBsAg and other reagents, such as TNF alpha and PBS, were added to the medium, and 18 hours later cells were collected for the detection of surface molecules (HLA-DR/CD86/CD1a). IL-12p70 levels in the supernatant and stimulating capacity to allogenic T lymphocytes were detected. The nucleic proteins of NF-kB and AP-1 in DC were extracted and purified for the gel shift assay.</p><p><b>RESULTS</b>Compared with those of the PBS group, the expression rates of HLA-DR of DC treated with CpG-ODN and/or HBsAg were obviously increased. Both the IL-12p70 level and the stimulating capacity of DC to allogenic T lymphocytes in mixed lymphocyte reaction increased significantly in the CpG-ODN group and in the CpG-ODN/HBsAg combination group (P less than 0.01 and 0.05, respectively). The expression rates of CD1a were raised only in the CpG-ODN/HBsAg combination group. Three kinds of immunological adjuvants, TNF alpha, CpG-ODN and CpG-ODN/HBsAg enhanced the expression of nucleic NF-kB and inhibited the expression of AP-1 in DC.</p><p><b>CONCLUSION</b>CpG-ODN, like TNF alpha, has remarkable stimulatory effect on the impaired phenotype and function of monocyte-derived DC in patients with CHB; CpG-ODN and HBsAg have a synergetic effect in increasing the antigen presenting function. The regulating ability of CpG-ODN and TNF alpha on the expression levels of NF-kB and AP-1 might be one of the mechanisms of their immunostimulatory effects on DC of the CHB patients.</p>

The effect of N-acetyl-L-cysteine on endoplasmic reticulum stress mediated apoptosis of HepG2 cells / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To analyze the mechanisms of NAC on endoplasmic reticulum (ER) stress mediated cells apoptosis of HepG2 cells and to evaluate the potential role of NAC in the treatment of liver injury.</p><p><b>METHODS</b>HepG2 cells were treated with H2O2 to make a model of oxidative ER stress mediated apoptosis. To evaluate the apoptosis, various methods such as MTT, DNA ladder, Western blot and flow cytometry were used. Then the optimal dosage and incubation time of NAC intervention in apoptosis were ascertained, and the differences between induction and intervention of apoptosis, including the percentage of apoptosis, the expression of apoptotic protein (GRP78, Caspase-12, PARP) and the production of reactive oxygen species (ROS) were compared.</p><p><b>RESULTS</b>The activity of the cells decreased by H2O2 (0, 1, 3, 5 mmol/L) treatments in a dose-dependent manner. The ratio of apoptotic cells increased (0.7%+/-0.5%, 26.4%+/-1.8%, 29.7%+/-1.2% and 51.2%+/-9.4%, respectively) as did the production of ROS (14.0%+/-0.5%, 95.2%+/-0.1%, 97.5%+/-0.2% and 98.3%+/-0.2%, respectively). The HepG2 cells showed typical morphologic change of ER stress 6 hr after they were treated with 3 mmol/L H2O2. ER stress mediated-apoptosis was confirmed by Western blot. NAC (10 mmol/L and 20 mmol/L) protected cells from apoptosis. Typical features of ER stress apoptosis were seen accompanied by diminishing the ratio of apoptotic cells from 29.7%+/-1.2% to 23.3%+/-4.7% and 14.3%+/-1.2%. The production of ROS also decreased from 97.5%+/-0.2% to 52.2%+/-0.8% and 51.2%+/-2.9%. The effect was related to the concentration: 20 mmol/L NAC was more effective than 10 mmol/L.</p><p><b>CONCLUSIONS</b>As an oxidizing agent, H2O2 may induce ROS in cells and induce oxidative stress, causing ER stress and apoptosis. NAC can inhibit the procession of ROS directly and prevent injuries to the hepatocytes.</p>

Toll like receptor 9 expression and functions of plasmacytoid dendritic cells in the peripheral blood of patients with chronic hepatitis B virus infection / 中华传染病杂志

Objective To investigate the expression of Toll-like receptor(TLR) 9 of circulating plasmacytoid dendritic cell (pDC) and analyze the frequency and interferon (IFN)-?production of circulating pDC during hepatitis B virus(HBV) infection.Methods Peripheral blood was collected from 69 HBV-infected patients,including 14 cases of asymptomatie HBV infection,30 cases of chro- nic hepatitis B(CHB),25 cases of HBV-related liver cirrhosis,and 21 healthy blood donors as con- trols.Flow cytometry was used to analyze the frequency of circulating pDC and the mean fluorescence intensity(MFI) of TLR9.Fresh peripheral blood mononuclear cells(PBMC) were stimulated by CpG ODN 2216 for 24 h in vitro.IFN-?in the supernatant was measured using enzyme-linked immunosorbent assay(ELISA) to analyze the frequency and IFN-?production of pDC during HBV in- fection.Data were analyzed with SPSS 13.0 for windows.Results Compared with healthy controls (62.6?10.7),the MFI of TLR9 of patients with asymptomatic HBV infection,those of CHB and HBV-related cirrhosis were significantly reduced (P

The degree of HBV suppression with 24 week telbivudine- or lamivudine-treatment in hepatitis B patients predicts the efficacy of the treatment at week 52 / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To investigate the possibilities of an association between the degrees of HBV suppression with nucleoside treatments at week 24 and week 52 in hepatitis B patients and to find a useful predictor for treatment efficacy.</p><p><b>METHODS</b>In this phase III, double-blind, multicenter trial, we compared the efficacy of telbivudine treatment with lamivudine treatment in 332 Chinese compensated chronic hepatitis B patients. The patients were randomly assigned to a daily 600 mg telbivudine treatment group or daily 100 mg lamivudine group for 24 weeks. They were then categorized into 4 groups according to their serum HBV DNA levels (copies/ml) at week 24: a PCR-undetectable group (< 300 copies/ml); a QL- < 10(3) copies/ml group; a 10(3)-<10(4) copies/ml group; and a > or = 10(4) copies/ml group. The treatments were continued as they previously had been for another 28 weeks and the patients serum HBV DNA levels were examined again.</p><p><b>RESULTS</b>At week 52, mean reductions of serum HBV DNA were significantly greater in the telbivudine-treated patients than in the lamivudine-treated group (6.2 log10 vs 5.4 log10, t = 3.6, P < 0.01). Viral resistance was twice as common in lamivudine-treated patients compared to those receiving telbivudine. Telbivudine was well-tolerated with an adverse event profile similar to that of lamivudine. The lower the HBV DNA level achieved at week 24, the higher HBV DNA non-detectable by PCR. ALT normalization and HBeAg seroconversion achieved at week 52, and viral resistance at week 48 decreased parallel to the degree of HBV DNA inhibition.</p><p><b>CONCLUSION</b>HBV DNA PCR-undetectable at week 24 in nucleoside-treated hepatitis B patients suggests a better efficacy at week 52 and lower viral resistance at week 48. The degree of suppression of HBV at week 24 may be used as a predictor of 1-year outcome.</p>

Correlation between liver stem cell activation and histopathologic changes in liver with hepatic failure / 中华病理学杂志

<p><b>OBJECTIVE</b>To investigate the correlation between liver stem cell activation and histopathologic changes in liver transplant recipients with hepatic failure.</p><p><b>METHODS</b>A total of 33 cases of hepatic failure were enrolled into the study. Donor liver tissues were used as normal controls. Histopathologic changes, presence of hepatotropic virus antigens, history of artificial liver therapy and number of c-kit positive cells were analyzed.</p><p><b>RESULTS</b>There were a total of 25 males and 8 females. The age of patients ranged from 21 to 64 years. Among the 33 patients studied, 6 suffered from acute liver failure, 5 from subacute liver failure and the remaining from acute-on-chronic liver failure (associated with cirrhosis). Thirteen patients had a history of artificial liver therapy. Activated liver stem cells expressed c-kit monoclonal antibody but were negative for toluidine blue stain. The number of c-kit-positive cells in acute liver failure, subacute liver failure and acute-on-chronic liver failure were 3.50 +/- 2.66 (0 to 8) per mm(2), 11.47 +/- 8.85 (3 to 30) per mm(2) and 15.50 +/- 10.95 (5 to 45) per mm(2), respectively (P < 0.05). The number of c-kit-positive cells in cases with or without artificial liver therapy showed no statistically significant difference.</p><p><b>CONCLUSIONS</b>The poor prognosis of acute liver failure is mainly due to massive liver necrosis and insufficient stem cell activation. Liver stem cell level is increased whenever there is progression into subacute liver failure and chronic liver failure. Actively treating acute liver failure with stimulation of the self-regeneration system in liver is thus useful.</p>

Histological changes in the livers of chronic hepatitis B patients with persistently normal serum alanine transaminase levels / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the histological changes in livers of chronic hepatitis B (CHB) patients with persistently normal serum ALT levels (PNAL).</p><p><b>METHODS</b>274 CHB patients who had percutaneous liver biopsies and had a detectable viral load (lower limit of detection is 10(3) copies/ml) in our department between October 2003 and February 2007 were included in this study. Among these patients, 139 had PNAL, group A, (with at least 3 normal serum ALT levels, with intervals of more than two months over a period of 12 or more months before the biopsy). The other 135 patients, group B, had abnormal serum ALT levels during the same period. The histological changes in the livers of the two groups of patients were compared.</p><p><b>RESULTS</b>Sixty-six (47.5%) patients with PNAL had normal liver histology, but significant pathohistological changes such as significant necroinflammation, fibrosis and/or cirrhosis were found in 33 (23.7%) patients. Thirteen (9.4%) had established cirrhosis. When compared to patients within (0-0.75)x upper limit of normal (ULN) ALT, patients within (0.76-1.00)x ULN ALT had higher scores of histological changes (43.5% vs. 19.8%, P < 0.05). In the PNAL group, scores of histological changes increased sharply in parallel with an age increase of older than 40 yrs. However neither viral loads nor HBeAg statuses of the PNAL patients had any predictive meaning to the scores of the histological findings.</p><p><b>CONCLUSIONS</b>23.7% of our CHB patients with PNAL, regardless of what their HBeAg statuses or viral load levels were, had significant liver pathohistological changes. Liver biopsies should be considered in CHB patients with PNAL, especially those older than 40 yrs and with a higher ALT within (0.76-1) x ULN.</p>

Foscarnet sodium for treatment in patients with severe chronic hepatitis B / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effectiveness of foscarnet sodium in the treatment of severe chronic hepatitis B.</p><p><b>METHODS</b>Two hundred and eight patients were enrolled in a multicenter, double-blind, controlled study. The patients received foscarnet sodium (foscarnet group) or saline (control group) injections for 4 weeks, and were then followed for 24 weeks.</p><p><b>RESULTS</b>HBV DNA negative rate was 12.8% in the foscarnet group and 7.1% in the control group at the end of treatment; and it was 5.5% and 3.0% at the end of the follow-up period respectively (P > 0.05). The rate of HBV DNA decrease of more than 2 log copies/ml was 53.2% in the foscarnet group and 16.2% in the control group at the end of treatment, and 23.9% and 8.1% (P < 0.01) respectively at the end of the follow-up period. The rate of HBV DNA < 10(5) copies/ml was 64.2% and 30.3% at week 4 in the two groups respectively, and 40.4% and 22.2% (P < 0.01) at the end of the follow-up period. HBeAg negative rate was 17.3% and 5.8% at the end of the treatment, and 22% and 5.4% at the end of the follow-up period (P < 0.01). The rate of HBeAg seroconversion was 12.7% and 3.7% at week 4, and 16.7% and 1.5% at the end of the follow-up period. Response rate was 60.6% and 21.2% at the end of week 4 (P < 0.05).</p><p><b>CONCLUSION</b>Foscarnet sodium injection has a good effect on severe chronic hepatitis B patients and it is safe to use on them.</p>

Preparation and identification of hammerhead ribozyme in vitro against caspase-12 mRNA fragments / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To design and synthesize ribozymes targeting 138 and 218 sites of the mRNA nucleotide of mouse caspase-12, a key intermedium of ER stress mediated apoptosis, and to identify their activities through in vitro transcription and cleavage.</p><p><b>METHODS</b>The mouse caspase-12 gene fragment was obtained by RT-PCR and cloned into the PGEM-T vector under the control of T7 RNA polymerase promoter. The transcription product of the target was labeled with a-32P UTP, while ribozymes were not labeled. Ribozyme and target RNA were incubated for 90 min at 37 degree C in a reaction buffer to perform the cleavage reaction.</p><p><b>RESULTS</b>It was found that under a condition of 37 degree C, pH 7.5 and with Mg2+ in a concentration of 10 mmol/L, Rz138 and Rz218 both cleaved targets at predicted sites, and the cleavage efficiency of Rz138 was 100%.</p><p><b>CONCLUSION</b>Rz138 and Rz218 prepared in vitro possess the perfect specific catalytic cleavage activity. Rz138 has excellent cleavage efficiency. It may be a promising tool to prevent ER stress induced apoptosis through catalytic cleavage of caspase-12 mRNA in vivo. It also can be used to verify whether caspase-12 is necessary in ER stress induced apoptosis.</p>

A study of the dosage and efficacy of entecavir for treating hepatitis B virus / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To evaluate the antiviral activity and safety of entecavir in patients with chronic HBV infection as a preliminarily step in selecting 0.1 mg or 0.5 mg as a better dosage for a further large scale clinical trial.</p><p><b>METHODS</b>This was a randomized, double-blinded, placebo-controlled and dose-ranging trial of entecavir usage in 212 patients with chronic HBV infection. The patients were randomly assigned to 3 groups: 0.1 mg entecavir (69), 0.5 mg entecavir (72) and, placebo (71) groups and treated for 28 days. The patients were then followed for 56 days without treatment.</p><p><b>RESULTS</b>The proportion of subjects who achieved the primary endpoint at day 28, with their HBV DNA level decreased >2 log or undetectable, was significantly greater in the entecavir 0.1 mg and 0.5 mg dose groups compared with the placebo group (P < 0.01 for both comparisons). The mean change from baseline in HBV DNA levels at day 28 was greater for entecavir 0.1mg and 0.5 mg groups compared with the placebo group (both P < 0.01). The mean change from baseline in HBV DNA levels at day 28 for entecavir 0.5 mg group was greater than that of the entecavir 0.1 mg group (P < 0.01). During the 56-day post-dosing follow-up phase, the entecavir 0.5 mg group was associated with greater and more sustained suppression of viral replication than the entecavir 0.1 mg group (P < 0.01). There were no clinically meaningful differences in the incidence of any adverse events between the entecavir dosing and the placebo groups.</p><p><b>CONCLUSION</b>Entecavir at both 0.1 mg and 0.5 mg doses demonstrated superior antiviral activity compared with a placebo. Since the entecavir 0.5 mg dose appears to have greater antiviral activity than the 0.1 mg dose and with a comparable safety and tolerability profile, the 0.5 mg entecavir dose could be used in further trials.</p>

Clinical investigation of famciclovir in chronic hepatitis B patients irresponsive to alpha interferon treatment / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To evaluate the efficacy and safety of famciclovir on the decreasing levels of serum HBV-DNA and ALT and HBeAg/antiHBe seroconversion in chronic hepatitis B patients irresponsive to 3 months treatment with alpha interferon.</p><p><b>METHODS</b>Two hundred and nineteen patients with chronic HBV infection, defined as positive HBsAg, HBeAg and HBV DNA, were enrolled and randomly half-and- half put into famciclovir and placebo groups. The two groups received either famciclovir 500 mg tid or a placebo treatment for 24 weeks, and then were followed-up for another 24 weeks with no treatment.</p><p><b>RESULTS</b>At the end of 24 weeks, the log value of HBV DNA dropped from 6.54+/-1.26 to 5.70+/-2.03 in the famciclovirt group and were elevated from 6.30+/-1.32 to 6.51+/-1.65 in the placebo group (P < 0.01). The rate of cases with persistence HBV DNA dropped 2 log of quantity in the famciclovir group and was 28.28% (28/99); it was 9.47% (9/95) in the placebo group (P < 0.01). Those with persistence negative HBV DNA was 28.28% (28/99) in the flamciclovir treated group and 14.74% (14/95) in the placebo group (P < 0.05). Those persistently being HBeAg negative were 7.69% (7/91) in the famciclovir treated group and 3.33% (3/90) in the placebo group (P > 0.05). The HBeAg/antiHBe seroconversion was 4.40% (4/91) in the famciclovir group and 2.22% (2/90) in the placebo group (P > 0.05). The percentage of cases with normal of ALT level was 15.15% in the famciclovir group and 6.35% in the placebo group (P < 0.05).</p><p><b>CONCLUSION</b>Famciclovir is effective in inhibiting HBV DNA replication and in decreasing serum ALT levels. The rate of HBeAg/antiHBe seroconversion in the famciclovir treated group was similar to that of the placebo group. Famciclovir was well tolerated without severe adverse effects during our treatment.</p>

Caspase-12 expression and activation in the pathogenesis of acute hepatic failure induced by lipopolysaccharide and D-galactosamine / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the role of caspase-12 expression on hepatocyte apoptosis in an experimental model of acute hepatic failure (AHF).</p><p><b>METHODS</b>A mouse experimental model of AHF was developed by intraperitoneal injection of lipopolysaccharide (LPS) and D-galactosamine (D-Gal). Hepatocyte apoptosis was examined by DNA agarose gel and liver pathology. Caspase-12 mRNA expression in liver was detected by reverse transcriptase PCR (RT-PCR) method. The expression of caspase-12, GRP78 proteins in livers was determined by Western blot.</p><p><b>RESULTS</b>Caspase-12 mRNA expression in the livers increased significantly from 5 to 7 hours after administration of LPS and D-Gal. Typical manifestation of hepatocyte apoptosis appeared at 5 hours after the drug administration. After 5 hours the level of serum ALT and AST were remarkably increased, and they reached the peak at 7 hours. The expression of procaspase-12 protein decreased obviously at 7 hours. Seven hours after the drug administration, hepatocyte apoptosis and necrosis both started. The marker of endoplasmic reticulum (ER) stress, Bip/GRP78 was activated during the development of hepatocyte apoptosis. The level of Bip/GRP78 protein was gradually increased at 5 hours after the drug induction.</p><p><b>CONCLUSION</b>Hepatocyte apoptosis plays an important role in the pathogenesis of AHF. Caspase-12 induced ER stress involves in hepatocyte apoptosis. It suggests that inhibition of caspase-12 activation might be a potential strategy in the treatment of AHF in the future.</p>

Inhibition of mouse hepatocyte apoptosis by anti-caspase-12 small interfering RNA / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To study the inhibition of primary mouse hepatocyte apoptosis by small interfering RNA (siRNAs) against caspase-12.</p><p><b>METHODS</b>The Balb/c mouse primary hepatocytes were isolated in situ with two-step liver perfusion with 0.5 g/L collagenase type IV, and apoptosis were induced with 4 micromol/L thapsigargin (TG). The three kingds of siRNAs targeting different gene sites (130, 214, 521) were synthetized chemically. The single-stranded RNAs were annealed to produce double-stranded siRNAs, then the mouse primary hepatocytes were transfected by oligofectamine package. The inhibition of caspase-12 was analyzed with RT-PCR and Western-blot. The viable hepatocytes following the induction of apoptosis were evaluated with MTT.</p><p><b>RESULTS</b>All the three kinds of siRNAs could obviously inhibit normal mouse hepatocyte caspase-12 mRNA. The siRNA (214) were more effective than the other two when the concentration was 100 nmol/L. The caspase-12 mRNA expression was inhibited by 52.08%, while that of siRNA (521) was 30.73% (t=4.30, P <0.05). However when the concentration was 200 nmol/L, the inhibitions were similar (88.07%, 86.22% and 89.41% respectively). siRNA (214) could downregulate the expression of apoptotic hepatocytes procaspase-12 by 51.43% ( t=4.30, P <0.01). Contrasted with apoptotic hepatocytes, the cell activity, which was analyzed with MTT, increased by 48.76% (t=2.23, P <0.01).</p><p><b>CONCLUSION</b>siRNAs could effectively downregulate the expression of caspase-12 at mRNA and protein levels and prevent mouse primary hepatocytes from apoptosis.</p>

Clone, expression and cleavage activity of anti-caspase-7 hammerhead ribozyme in vitro / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To design hammerhead ribozymes against mouse caspase-7 and to study their expression and cleavage activity in vitro.</p><p><b>METHODS</b>The secondary structures of ribozyme and caspase-7 genes were analyzed and simulated by computer. Ribozymes DNA sequences were synthesized by automatic synthetic apparatus. Caspase-7 DNA sequence was acquired by reverse transcription PCR. Ribozymes and caspase-7 DNA sequences were separately cloned into pBSKneo U6 and pGEM-T vectors. Ribozymes and caspase-7 mRNA were obtained by transcription in vitro, and ribozymes cleavage activity was identified by cleavage experiment in vitro.</p><p><b>RESULTS</b>Two ribozymes named Rz333 and Rz394 targeting 333 and 394 sites in caspase-7 mRNA were designed by computer software, and their DNA sequences were synthesized. The expression vector of caspase-7 and plasmids containing Rz333 and Rz394 were reconstructed successfully. Ribozymes and caspase-7 mRNA were expressed by in vitro transcription. In vitro cleavage experiments showed that Rz333 cleaved caspase-7 mRNA and produced 243nt and 744nt segments. The cleavage efficiency is 67.98%, while Rz394 cannot cleave caspase-7 mRNA.</p><p><b>CONCLUSIONS</b>Rz333 can site-specifically cleave caspase-7 mRNA.</p>

The experimental study on the expression of toll-like receptor 2 in fulminant hepatic failure / 中华肝脏病杂志

<p><b>OBJECTIVE</b>In order to explore the role of toll-like receptors 2 (TLR2) in initiating inflammatory response, the expression of TLR2 of the liver and IL-18, TNF-alpha and IFN-gamma of plasma in fulminant hepatic failure was analysed.</p><p><b>METHODS</b>D-galactosamine (D-Gal, 900 mg/kg) and lipopolysaccharide (LPS, 10 microg/kg) were administered intraperitoneally into the BALB/C mice. To evaluate the hepatic injury, serum transaminase (ALT and AST) and plasma IL-18, TNF-alpha and IFN-gamma were determined and the mortality was observed at various time points following the intraperitoneal injection. The level of TLR2 mRNA was measured by semiquantitative RT-PCR. The protein expression of TLR2 in the liver was detected by immunohistochemistry. The data was analyzed by SAS software.</p><p><b>RESULTS</b>After 4 hours of intraperitoneal injection of D-Gal/LPS, the serum transaminase and plasma IL-18, TNF-alpha and IFN-gamma levels were elevated. The treated mice began to die at 7 hours. The mortality reached up to 80% at 10 h. TLR2 mRNA was expressed at a low level in liver tissues of normal mice, while it was significantly increased and maintained at a higher level following intraperitoneal injection with D-Gal/LPS. The expression of TLR2 protein was similar to that of the TLR2 mRNA, and the expression of TLR2 mRNA was positively correlated with the concentration of plasma IL-18, TNF-alpha and IFN-gamma (r=0.36, P=0.02; r = 0.48, P 0.003; r = 0.72, P<0.001) at different time points.</p><p><b>CONCLUSIONS</b>Our results showed that TLR2 was involved in initiating and inducing the expression of proinflammation cytokines in this model of fulminant hepatic failure. The results suggest that adjusting the expression of TLR2 might be a new strategy in preventing the development of infectious diseases</p>

Effects of growth factors and extracellular matrix on proliferation and differentiation of fetal liver progenitor cell in vitro / 中华肝脏病杂志

<p><b>OBJECTIVE</b>Liver development needs a number of growth factors and components of the extracellular matrix. The study is to explore how growth factors and extracellular matrix regulate proliferation and differentiation of fetal liver progenitor cell.</p><p><b>METHODS</b>We demonstrate isolation of hepatic progenitor/stem cells from ED 14.5 SD rat liver, which contains a large number of hepatoblasts. Proliferation assay-3H thymidine incorporation was used to detect the effect of growth factors on proliferation of hepatic progenitor cell. Growth factor and extracellular matrix were added and stem cell clone formation was counted. Mark of bile duct and hepatocyte were detected with double-marker immunocytochemistry.</p><p><b>RESULTS</b>Progenitor liver cells displayed clonogenic capacity, expressed markers of hepatocytes and bile duct cells and G-6-P. HGF, EGF can accelerate DNA synthesis and stem cell clone formation of hepatic progenitor cell. Extracellular matrix collagen I, collagen IV or laminin were essential for formation of stem cell clone. Single cell culture needed HGF, EGF, extracellular matrix and supernatant of mix cell (which contained fetal parenchymal cells, mesenchymal cells and hematopoietic cells) culture.</p><p><b>CONCLUSION</b>Growth factors especially HGF and EGF play crucial role in proliferation and differentiation of liver progenitor cell. Some factors secreted from mesenchymal cell and hematopoietic cells may be involved.</p>

Effect of Tauroursodeoxycholic acid on cytochrome C-mediated apoptosis in HepG2 cells / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effect of Tauroursodeoxycholic acid (TUDCA) on Taurodeoxycholic acid (TDCA)-induced HepG2 cell apoptosis and to clarify the molecular mechanism of its anti-apoptosis effect of TUDCA.</p><p><b>METHODS</b>Morphologic evaluation of apoptotic cells was performed by Hoechst 33258 staining and electron microscope. DNA fragment was detected by electrophoresis on 1.5% agarose gels. Apoptosis rate was measured by flow cytometry using PI dye. Following incubation of HepG2 cells either with TDCA alone, or coincubation with TUDCA and TDCA, the releasing level of cytochrome c from mitochondria into cytosol was determined by western blot, also the activity of caspase-3, 8, 9.</p><p><b>RESULTS</b>Incubating the cells with 400 micromol/L TDCA for 12 h induced the cells apoptosis significantly. The apoptotic rate decreased from 50.35% +/- 2.20% to 13.78% +/- 0.84% after coincubation with TUDCA, and this anti-apoptotic effect of TUDCA was confirmed by morphological and DNA ladder detection. TUDCA significantly inhibited the release of cytochrome C from mitochondria into cytosol, and the activity of caspase-9, 3 (t > or = 13.00, P < 0.01), especially at 12 h, caspase-3 activity decreased by 54.9% (t = 16.88, P < 0.01) and 52.5%, however it had no obvious effect on the activity of caspase-8 (t = 1.94, P > 0.05).</p><p><b>CONCLUSIONS</b>TUDCA prevents HepG2 cells apoptosis induced by TDCA through modulating mitochondrial membrane stability, inhibiting the release of cytochrome c and the activation of procaspase-9 and 3. Anti-apoptotic mechanism of TUDCA may be considered to be one of the most important reasons that TUDCA exerts significant efficacy in the treatment of cholestatic liver diseases.</p>

A clinical research on oxymatrine for the treatment of chronic hepatitis B / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the clinical effect of oxymatrine on chronic viral hepatitis B and to look for new methods for treating hepatitis B.</p><p><b>METHODS</b>Multi-center, controlled study was used. In this study, 196 patients were allocated to oxymatrine, oxymatrine with Ara-AMP, IFN-a1b, and glucose groups to observe ALT, AST and viral marker changes.</p><p><b>RESULTS</b>At the end of treatment, the rate of normal ALT, the negative rate of HBV DNA and HBeAg, and the positive rate of HBeAb were similar in oxymatrine, oxymatrine with Ara-AMP, and IFN-a1b groups. It was higher than that of glucose group. After 12 months follow up, the total effective rate is 40.8%, 60.8% and 43.1% in oxymatrine, oxymatrine with Ara-AMP, and IFN-a1b groups, respectively.</p><p><b>CONCLUSIONS</b>Oxymatrine, oxymatrine with Ara-AMP, and IFN-a1b are effective to treat hepatitis B with a good negative rate of HBV DNA and HBeAg and positive rate of HBeAb.</p>

Immunostimulatory role of CpG-containing oligodeoxynucleotide on the monocyte-derived dendritic cell in patients with chronic hepatitis B / 中华传染病杂志

0.05).However,neither CpG-ODN nor hTNF-? failed in improving the expression rates of CD1a.Conclusions CpG-ODN,like hTNF-?,has remarkable im- munostimulatory effect on the differentiation and maturation of monocyte-derived DC from patients with chronic hepatitis B.