[Clinical observations of sequential interferon therapy following complete response to telbivudine treatment in chronic hepatitis B patients].

OBJECTIVE: To investigate the therapeutic benefit of sequential interferon alpha-1b (IFNa-1b) in patients with hepatitis B e antigen (HBeAg)-positive chronic hepatitis B (CHB) who showed early complete response to telbivudine (LdT) treatment, and to explore the clinical value of serum hepatitis B surface antigen (HBsAg) for predicting sustained virological response (SVR). METHODS: Twenty HBeAg+ CHB patients who had shown a complete response to LdT therapy before treatment week 52 were divided into two treatment groups: one continued on the LdT treatment for an additional 6 months, and the other switched to IFNa-1b for 6 months. Each patient presented for follow-up examinations at 1, 2, 3, 6, 9, 12, 18, 24, 30 and 36 months after treatment cessation. Serum levels of alanine aminotransferase (ALT) and creatinine were detected by an automated biochemical analyzer. HBV DNA load was determined by real-time PCR. HBsAg and HBeAg levels were assessed by chemiluminescence. RESULTS: The relapse rate was lower in the group treated with sequential IFN than in the group who continued LdT treatment (30% vs. 40%, P more than 0.05). The area under the receiver operating characteristic curve at week 24 (0.689) was significantly higher than at week 12 and week 48 (0.652 vs. 0.545, P less than 0.05). Decline in serum HBsAg levels at week 24 were predictive of SVR after treatment cessation. Patients showing a decrease more than 1000 IU/ml in serum HBsAg levels at week 24 had a significantly higher SVR rate than the patients who showed a decrease less than 1000 IU/ ml (90.9%(10/11) vs. 33.3%(3/9), P less than 0.05). At the end of treatment, patients showing a decrease less than 200 IU/ml of serum HBsAg levels had a significantly higher SVR rate than those showing more than 200 IU/ml (100% vs. 53.3%, P less than 0.05). CONCLUSION: Sequential IFNa-1b consolidation therapy does not reduce the rate of relapse after treatment cessation. However, patients with a decrease in serum HBsAg levels of more than 1000 IU/ml at treatment week 24 are more likely to achieve SVR.

Ammonium 2-(2,4-dichloro-phen-oxy)acetate hemihydrate.

The title compound, NH(4) (+)·C(8)H(7)Cl(2)O(6) (-)·0.5H(2)O, was prepared by the reaction of 2-(2,4-dichloro-phen-oxy)-acetic acid and ammonia in water at 367 K. The mol-ecular structure and packing are stabilized by N-H⋯O and O-H⋯O inter-molecular hydrogen-bond inter-actions.

[Dynamic changes of Th1/Th2 type cytokines in peripheral blood of patients with hepatitis B e antigen-positive chronic hepatitis B treated with telbivudine].

OBJECTIVE: To investigate the dynamic changes of Th1/Th2 type cytokines in peripheral blood of patients with hepatitis B e antigen-positive chronic hepatitis B treated with telbivudine (LDT). METHODS: The levels of IL-2, IL-4, IL-6, IL-10, TNF alpha, IFN gamma in the blood cells of HBeAg positive chronic hepatitis B patients were measured at 0, 4, 8, 12, 24, 48 weeks after LDT treatment by fluorescence activated cell sorting (FACS), the levels and dynamic changes of Th1/Th2 type cytokines in groups of complete response, partial response, non-response, virologic breakthrough were compared. RESULTS: The levels of Th1 type cytokines in complete response group were higher than those in groups of partial response, non-response and virologic breakthrough, however, the levels of Th2 type cytokines showed an opposite trend compared with Th1 type cytokines. There were no significant differences between each group. In complete response group, the levels of IL-2, TNF alpha and IFN gamma were higher than baseline 12 weeks after LDT treatment (P < 0.05). In partial response group the level of IFN gamma was higher than baseline 24 weeks after LDT treatment (P < 0.05). In non-response group, the levels of IL-6 and IL-10 were higher than baseline at 48 weeks after LDT treatment (P < 0.05). In virologic breakthrough group, the level of IL-4 was higher than baseline 24 weeks after LDT treatment (P < 0.05), while the level of IL-6 was higher than baseline 12 weeks after LDT treatment (P < 0.05). CONCLUSIONS: The balance of Th1/Th2 type cytokines plays an important role in the outcome of patients with hepatitis B e antigen-positive chronic hepatitis B treated with LDT. The immune response of patients with chronic hepatitis B is improved to some extent after LDT therapy.

[A clinical study of N-acetylcysteine treatment in chronic hepatitis B patients].

OBJECTIVE: To evaluate the efficacy and safety of N-acetylcysteine (NAC) and glutathione (GSH) in treating chronic hepatitis B patients. METHODS: Seventy-five patients with chronic hepatitis B were treated daily with an injection containing the same basic therapeutic drugs and randomly divided into a NAC group (50 patients) and a GSH group (25 patients). A daily dose of 8 grams of NAC and 1.2 grams of GSH was added to the injections of the two groups respectively. The trial lasted 28 days. Hepatic function and other biochemistry parameters (TBil, PTA, ALB et al) were tested on experimental day 0 and on days 7, 14, 21 and 28. The evaluation on the total effective rates of the NAC and GSH groups was based on the decreases of serum TBil and the increases of PTA. RESULTS: Both NAC and GSH have therapeutic effects. The total effective rate was 84% in the NAC group and 72% in the GSH group. The rate of side effects was 13% in the NAC group. CONCLUSION: NAC and GSH can decrease the level of serum TBil and increase PTA, but NAC was more effective in decreasing TBil than GSH. Serious adverse effects of NAC were not observed during the period of our treatment.

[Short-term results of telbivudine versus entecavir treatments in HBeAg-positive chronic hepatitis B patients in China].

OBJECTIVE: To evaluate the efficacy and safety of telbivudine (LDT) versus entecavir treatments in hepatitis B e antigen (HBeAg)-positive chronic hepatitis B (CHB) patients. METHODS: Eighty HBeAg-positive compensated CHB patients with HBV DNA more than 6 log10 copies/ml and serum ALT 2 x ULN were divided into two groups: a telbivudine treatment group, and a entecavir treatment group. HBV DNA, ALT and HBeAg were surveyed at baseline and at 12 and 24 weeks. The efficacy and safety of the two nucleoside analogues were assessed at 12 and 24 weeks. RESULTS: Undetectable serum HBV DNA levels of the telbivudine group (50% and 80%) were similar to those of the entecavir B group (50% and 70%) according to the polymerase-chain-reaction assay at week 12 and 24. There were no significant differences in the normalization of alanine aminotransferase levels between the two groups at week 12 and 24 (52.5% vs 60.0%, 77.5% vs 75.0%). The mean reductions in serum HBV DNA from the baseline levels at week 12 and 24 were similar between the two groups [5.27 vs.5.36, 6.49 vs.6.18 log (on a base-10 scale) copies per milliliter]. More patients in the telbivudine group had HBeAg seroconversion at week 12 than those in the entecavir group (20.0% vs 5.0%, P = 0.043); however, there was no significant difference between the two groups at week 24 (27.5% vs 17.5%). No adverse reactions were found in either group. CONCLUSION: There was no significant difference in HBV DNA undetectable rates and the ALT normalization rates between the two groups in a short-term therapy (24 weeks), but the telbivudine group had a higher rate in HBeAg seroconversion than that in the entecavir group at week 12.

Activation of extracellular signal-related kinases 1 and 2 in Sertoli cells in experimentally cryptorchid rhesus monkeys.

AIM: To assess the spatiotemporal changes in the expression of extracellular signal-regulated kinases 1 and 2 (ERK1/2), c-Jun N-terminal kinases (JNK) and p38 mitogen-activated protein kinases (MAPK) in response to heat stress in the cryptorchid testis, and to investigate a possible relation to Sertoli cell dedifferentiation. METHODS: Immunohistochemistry and western blot were used to examine the expression and activation of ERK1/2, p38 and JNK in the cryptorchid testis at various stages after experimental cryptorchidism. RESULTS: The abdominal temperature did not obviously change the total ERK1/2 expression but significantly activated phospho-ERK1/2 in the Sertoli cells of the cryptorchid testis. Heat stress increased total JNK expression in the Sertoli cells of the cryptorchid testis but did not activate phospho-JNK. Neither total p38 nor phospho-p38 was induced by heat stress in the Sertoli cells of the cryptorchid testis. Changes in the spatiotemporal expression of cytokeratin 18 (CK18), a marker of immature or undifferentiated Sertoli cells, were induced in the cryptorchid testis in a pattern similar to the activation of ERK1/2. CONCLUSION: The activation of ERK1/2 in the testis may be related to dedifferentiation of Sertoli cells under heat stress induced by experimental cryptorchidism.

Stem cell factor modulates the expression of steroidogenesis related proteins and FSHR during ovarian follicular development.

Stem cell factor (SCF) is essential for the development of primordial follicles. By using cultured ovaries from neonatal rats, the effect of SCF on early follicular development was investigated. Steroidogenesis is a hallmark of follicular development. Expressions of three key protein factors in steroidogenesis, SF-1, StAR, and P450arom, were investigated using immunohistochemistry and in situ hybridization. SF-1 and StAR proteins were expressed in all ovarian cells. P450arom mRNA was localized exclusively in oocytes implying that estrogen might be synthesized by oocytes at this stage. SCF up-regulated the mRNA and protein expression of these proteins, suggesting SCF might promote the production of estrogen during this period of time. To study the differentiation status of follicular cells, the expression of FSHR and its response to SCF treatment was examined by using semi-quantitative RT-PCR. The results showed that SCF inhibited the expression of FSHR mRNA. It was also observed that SCF stimulated the expression of basic fibroblast growth factor (bFGF) in oocytes. Inactivation of bFGF by its neutralizing antibody resulted in a reversal of the inhibitory effect of SCF on the expression of FSHR. Therefore, the FSHR inhibitory effect of SCF could be mediated by bFGF. In summary, it seems that, at the early stages of follicular development, SCF might stimulate oocytes to produce estrogen while it inhibits the differentiation of granulosa cells that are the major sources of estrogen at the late stages of follicular development.

Expression of HSP105 and HSP60 during germ cell apoptosis in the heat-treated testes of adult cynomolgus monkeys (macaca fascicularis).

To confirm that transient increase in temperature of the testis (43C for 30 minutes once daily for 2 consecutive days) could induce apoptosis of germ cells in non-human primates and to investigate the possible roles of Hsp105 and Hsp60 in regulation of germ cell loss, we conducted the study on eight cynomolgus monkeys. The sperm concentration on day 28 after heat shock decreased to 8.4% of pretreatment levels and recovered to baseline on day 144. Using the TUNEL assay, increased numbers of apoptotic spermatocytes and round spermatids were detected on days 3, 8, and 30 post heat treatment. Hsp105 and Hsp60 mRNA and protein levels were analyzed using in situ hybridization, RT-PCR, immunohistochemical and Western blot methods. Hsp105 was confined to nuclei of spermatids before treatment, decreased dramatically with the loss of spermatids on days 3, 8, and 30, before returning to baseline levels on days 84 and 144. The expression of Hsp60 was high on days 3, 8, 30 and was only detected in Sertoli cells and spermatogonia. These results suggested that exposure of the testis to heat resulted in selective, but reversible damage to the seminiferous epithelium via increased germ cell apoptosis. Temporal changes in the expression pattern of Hsp105 and Hsp60 in relation to germ cell death suggests they may be involved in key processes in regulation of germ cell apoptosis.

Therapeutic effect of autologous dendritic cell vaccine on patients with chronic hepatitis B: a clinical study.

AIM: To investigate the therapeutic effect of autologous HBsAg-loaded dendritic cells (DCs) on patients with chronic hepatitis B. METHODS: Monocytes were isolated from fresh peripheral blood of 19 chronic HBV-infected patients by Ficoll-Hypaque density gradient centrifugation and cultured by plastic-adherence methods. DCs were induced and proliferated in the culture medium with recombinant human granulocyte-macrophage-colony-stimulating factor (rhGM-CSF) and human interleukin-4 (rhIL-4). DCs pulsed with HBsAg for twelve hours were injected into patients subcutaneously twice at intervals of two weeks. Two patients received 100 mg oral lamivudine daily for 12 mo at the same time. HBV-DNA and viral markers in sera of patients were tested every two months. RESULTS: By the end of 2003, 11 of 19 (57.9%) patients had a clinical response to DC-treatment. HBeAg of 10 (52.6%) patients became negative, and the copies of HBV-DNA decreased 10(1.77+/-2.39) on average (t = 3.13, P<0.01). Two cases co-treated with DCs and lamivudine had a complete clinical response. There were no significant differences in the efficient rate between the cases with ALT level lower than 2XULN and those with ALT level higher than 2XULN before treatment (chi(2) = 0.0026). CONCLUSION: Autologous DC-vaccine induced in vitro can effectively suppress HBV replication, reduce the virus load in sera, eliminate HBeAg and promote HBeAg/anti-HBe transformation. Not only the patients with high serum ALT levels but also those with normal ALT levels can respond to DC vaccine treatment, and the treatment combining DCs with lamivudine can eliminate viruses more effectively.

Replication of hepatitis B virus in primary duck hepatocytes transfected with linear viral DNA.

AIM: To explore the expression and replication of hepatitis B virus (HBV) DNA in primary duck hepatocytes (PDHs). METHODS: Complete HBV genome was transfected into PDHs by electroporation (transfected group, 1.19 x 10(12) copies of linear HBV DNA/1 x 10(7) PDHs). After 1-5 d of transfection, HBsAg and HBeAg in the supernatant and lysate of PDHs were measured with the IMX System. Meanwhile, replicative intermediates of HBV DNA were analyzed by Southern blotting and Dot blotting. PDHs electroporated were used as control group. RESULTS: HBsAg in the hepatocyte lysates of transfected group was 15.24 (1 d), 14.55 (3 d) and 5.13 (5 d; P/N values, positive > or =2.1) respectively. HBeAg was negative (<2.1). Both HBsAg and HBeAg were negative in the supernatant of transfected group. Dot blotting revealed that HBV DNA was strongly positive in the transfected group and negative in the control group. Southern blot analysis of intracellular total DNA indicated that there were relaxed circular (rc DNA), covalently closed circular (ccc DNA), and single-stranded (ss DNA) HBV DNA replicative intermediates in the transfected group, there was no integrated HBV DNA in the cellular genome. These parameters were negative in control group. CONCLUSION: Expression and replication of HBV genes can occur in hepatocytes from non-mammalian species. HBV replication has no critical species-specificity, and yet hepatic-specific regulating factors in hepatocytes may be essential for viral replication.

[Effects of mitomycin C on the expression and transport of ice-nuclei proteins of Erwinia herbicola].

Abstract: In this paper, Mitomycin C (MMC) was added to different kinds of medium to study the effects of different cultural conditions on the Erwinia herbicola 10025A. For the first time it was confirmed that the expressed activity of the ice-nuclei active protein was different from its transportable manner from the ice nucleation active bacteria (Erwinia herbicola 10025A). The findings indicated that MMC could stimulate the SOS response,and induce the synthesis of some enzymes and proteins, which take part in repairing the damaged DNA. The effects of the MMC on the E. herbicola under different media were different. It could increase the ice nucleation activity of the E. herbicola, forming new small vesicles, which are secreted to the outside of membrane. The importance of this research for study the living mechanism of cells ander poor condition was discussed.

[A single factor analysis of the prognosis of 301 hepatitis failure cases and a study of a scoring system on their prognostic assessment].

OBJECTIVES: To analyze the factors related to prognosis of hepatitis failure and to determine the factors which significantly affect it, and to build a scoring system for assessment of the prognosis of hepatitis failure and also to examine its efficacy for clinical use. METHODS: Clinical data from 301 patients were analyzed retrospectively. The correlated degree between those single factors and prognosis of hepatitis failure was explored by logistic regression analysis. Independent risk factors of prognosis and those correlated coefficients, which were from logistic regression analysis, were used to build a scoring system. This system was used in analyzing the clinical data of 275 patients to examine its efficacy of the prognostic assessment. RESULTS: The factors that significantly affected the prognosis of hepatitis failure included age, clinical typing, hepatic coma, total bilirubin, and others (P < 0.01). Some factors, including PTA, blood urea, sodium and hepatic coma, were independent risk factors of prognosis. The scoring system built gave different scores between the effective treatment group and ineffective treatment group with statistical significance (P < 0.01). When the score was less than 40, the probability of a recovery was 76.9%; when the score was 40 to 80, the probability of a recovery was only 12.5%. When the score was more than 80, the probability of a recovery dropped to 0%. CONCLUSIONS: The factors, including PTA, blood urea and sodium and hepatic coma, are important in building a scoring system to assess the prognosis of hepatitis failure. The scoring system we built is very effective in evaluating the prognosis of hepatitis failure.

[Clinical investigation of famciclovir in chronic hepatitis B patients irresponsive to alpha interferon treatment].

OBJECTIVES: 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. METHODS: 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. RESULTS: 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). CONCLUSION: 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.

[A multi-center clinical study of N-acetylcysteine on chronic hepatitis B].

OBJECTIVES: To evaluate the effectiveness and safety of N-acetylcysteine (NAC) in treating chronic hepatitis B patients. METHODS: 144 patients with chronic hepatitis B (total bilirubin, TBil>170 mmol/L) from several centers were chosen for a randomized and double blind clinical trial. The patients were divided into a NAC group and a placebo group and all of them were treated with an injection containing the same standardized therapeutic drugs. A daily dose of 8 microgram NAC was added to the injection of the NAC group. The trial lasted 45 days. Hepatic function and other biochemistry parameters were checked at the experimental day 0 and days 15, 30, 45. RESULTS: Each group consisted of 72 patients of similar demology and disease characteristics. During the trial, 28 cases of the 144 patients dropped out. In the NAC group, at day 0 and day 30, the TBil were 401.7 vs. 149.2 and 160.1+/-160.6. In the placebo group, the TBil on the corresponding days were 384.1+/-134.0 and 216.3+/-199.9. Its decrease in the NAC group was 62% and 42% in the placebo group. At day 0 and day 45 of treatment, the effective PTa increase rate was 72% in the NAC group and 54% in the placebo group. The total effective rate (TBil + PTa) was 90% in the NAC group and 69% in the placebo group. The parameters of the two groups showed a remarkable difference. The rate of side effects was 14% in the NAC and 5% in the placebo groups. CONCLUSION: NAC can decrease the level of serum TBil, increase the PTa and reduce the time of hospitalization. NAC showed no serious adverse effects during the period of our treatment. We find that NCA is effective and secure in treating chronic hepatitis B patients.

[Effects of different salt and alkali stresses on ion distribution in Red globe/Beta grapevines].

The potted Red globe/Beta grapevines were selected to irrigated with NaCl, Na2SO4, NaHCO3, NH4Cl, (NH4)2SO4. Hence, the ions which induced leaf etiolation were screened and the impacts of different salt and alkali on ion distribution in different organs of grapevines were investigated. It was found that NaHCO3 exerted the greatest effects on grapevines, leaf etiolation at 14 days after treatment. By contrast, NaCl and NH4Cl treatments induced leaf etiolation at 28 days after treatment. The Na+ content in all the detected organs were significantly increased under NaHCO3 and NaCl treatment, and Na+ content in root under NaHCO3 treatment was 6.4 times as that in control root. NaHCO3 and NaCl treatments significantly decreased K+ content in the organs with the exception of leaf. NaHCO3 treatment significantly decreased K/Na in different organs, which declined to 0.1 in root. By contrast, NaCl treatment significantly decreased K/Na in the detected organs with exception of stem. Besides, the transport of Ca2+, Mg2+, Fe2+ to aboveground organs was significantly decreased by NaHCO3 and NaCl treatments. K/Na ratio in the detected organs were decreased under NH4Cl, (NH4) 2SO4 and Na2SO4 treatments, especially under NH4 Cl treatment. Taken together, NaHCO3 was the primary factor resulting in leaf etiolation, followed by NaCl and NH4Cl, while (NH4) 2SO4 and Na2SO4 produced impacts.

Establishment and assessment of two methods for quantitative detection of serum duck hepatitis B virus DNA.

AIM: To establish and assess the methods for quantitative detection of serum duck hepatitis B virus (DHBV) DNA by quantitative membrane hybridization using DHBV DNA probe labeled directly with alkaline phosphatase and fluorescence quantitative PCR (qPCR). METHODS: Probes of DHBV DNA labeled directly with alkaline phosphatase and chemiluminescent substrate CDP-star were used in this assay. DHBV DNA was detected by autoradiography, and then scanned by DNA dot-blot. In addition, three primers derived from DHBV DNA S gene were designed. Semi-nested primer was labeled by AmpliSensor. Standard curve of the positive standards of DHBV DNA was established after asymmetric preamplification, semi-nested amplification and on-line detection. Results from 100 samples detected separately by alkaline phosphatase direct-labeled DHBV DNA probe with dot-blot hybridization and digoxigenin-labeled DHBV DNA probe hybridization. Seventy samples of duck serum were tested by fluorescent qPCR and digoxigenin-labeled DHBV DNA probe in dot-blot hybridization assay and the correlation of results was analysed. RESULTS: Sensitivity of alkaline phosphatase direct-labeled DHBV DNA probe was 10 pg. The coincidence was 100% compared with digoxigenin-labeled DHBV DNA probe assay. After 30 cycles, amplification products showed two bands of about 180 bp and 70 bp by 20 g/L agarose gel electrophoresis. Concentration of amplification products was in direct proportion to the initial concentration of positive standards. The detection index was in direct proportion to the quantity of amplification products accumulated in the current cycle. The initial concentration of positive standards was in inverse proportion to the number of cycles needed for enough quantities of amplification products. Correlation coefficient of the results was (0.97, P<0.01) between fluorescent qPCR and dot-blot hybridization. CONCLUSION: Alkaline phosphatase direct-labeled DHBV DNA probe in dot-blot hybridization and fluorescent qPCR can be used as valuable means to quantify DHBV DNA in serum.

Effects of electroporation on primary rat hepatocytes in vitro.

AIM: To investigate the effects of electroporation on primary rat hepatocyte and to optimize the electroporation conditions introducing foreign genes into primary hepatocytes. METHODS: A single-pulse procedure was performed at low voltage (220-400 V) but with high capacitance (500-950 microF). Hepatocytes were divided into 4 groups according to the electroporation conditions: group I, 220 V and 500 microF; group II, 220 V and 950 microF; group III, 400 V and 950 microF,and group IV. The control group was freshly isolated hepatocytes and directly cultured under the same conditions as those of electroporation groups. The effects of electroporation on primary rat hepatocytes were detected by trypan blue exclusion (TBE) and MTT analysis. Besides, albumin (Alb), alanine transaminase (ALT) and lactate dehydrogenase (LDH) in the supernatants of cultured hepatocytes were measured by biochemical assay. RESULTS: Between day 1 and day 15 after incubation, primary rat hepatocytes of each electroporation group appeared normal, being the same with those of control group. TBE staining showed that slight hepatocyte damage and high survival rate were found in the electroporation groups and the control group. Cultured for 3, 7, 11 and 15 days, hepatocyte viability was approximately 92.6+/-2.5 %, 89.5+/-3.3 %, 82.0+/-3.5 % and 74.3+/-1.2 %, respectively. MTT analysis indicated that the viabilities of hepatocytes had no significant difference between each electroporation group, and those were similar to that of control group. At the 36th hour after electroporation, Alb, ALT and LDH in the supernatants of control group were 5.3+/-0.1 g x L(-1), 183.7+/-8.4 nkat x L(-1) and 896.8+/-58.5 nkat x L(-1); those of group II were 5.7+/-0.1 g x L(-1), 215.4+/-16.7 nkat x L(-1) and 1063.8+/-51.8 nkat x L(-1); and those of group III were 5.8+/-0.2 g x L(-1), 217.1+/-8.4 nkat x L(-1) and 1063.8+/-10.0 nkat x L(-1). Statistically, the proteins of group II and group III were significantly higher than those of control group (P<0.05), whereas the protein production of group I, Alb, ALT and LDH were 5.3+/-0.2 g x L(-1), 205.4+/-3.3 nkat x L(-1) and 1035.4+/-116.9 nkat x L(-1), were similar to those of control group. At the same time, TBE and MTT analysis indicated that there was no significant cell viability difference between electroporation groups and control group. CONCLUSION: This single-pulse electroporation procedure performed at low voltage (220-400 V) but with high capacitance (950 microF) is one of the optimal choices to introduce foreign genes into primary rat hepatocyte.

The natural history of chronic hepatitis B: a retrospective study.

OBJECTIVE: To clarify the natural history of chronic hepatitis B so as to evaluate its long-term therapeutic outcome of the patients and the efficacy of antiviral drugs. METHODS: A cohort of 183 biopsy-proven chronic hepatitis B patients (mean age of 31.75+/-8.03 years, male/female ratio: 152:31) and 247 controls were followed up retrospectively for 11.81+/-4.08 years. This study was focused on long-term clinical outcome including the rates of liver cirrhosis, hepatocellular carcinoma and death, apart from the long-term effect of antiviral drugs and prognostic factors. RESULTS: In the 183 chronic hepatitis B patients, 22 (12.02%) developed liver cirrhosis, 12 (6.56%) developed hepatocellular carcinoma, and 20 (10.93%) died. The 5-, 10- and 15-year survival rates were 97.27%, 91.62%, and 84.47%, respectively. The 5-, 10- and 15-year incidence rates of HCC were 0, 3.19%, and 11.56%, respectively. In the 247 controls, 6 (2.43%) died; none of them developed cirrhosis or HCC. The rates of death, liver cirrhosis, and HCC in the hepatitis B patients were markedly different (P<0.005) compared with the controls. The overall mortality of hepatitis B patients was 4.5-fold higher than the general population. Cox multiple regression analysis showed that old age, severe histological injury, and positive HBeAg were closely related to liver cirrhosis; old age, severe histological injury, and male were major factors leading to death. The independent variable of predicted HCC was not found. CONCLUSION: The long-term outcome of hepatitis B patients is poor and the efficacy of antiviral drugs needs further study.

[One year and half treatment with lamivudine in active cirrhosis resulting from chronic hepatitis B].

OBJECTIVE: To study the therapy effect of long term lamivudine treatment on active cirrhosis following chronic hepatitis B, and explore the methods for abnormalities resulting from lamivudine withdrawing. METHODS: 58 patients received lamivudine 100 mg orally everyday for 18 months. The changes were observed and wrote down, including clinical symptoms and signs, aminotransferase, virology indexes, and the abnormalities after lamivudine withdrawing, then further to find out plans for the latter. RESULTS: (1) After lamivudine treatment, there were 35 patients whose situation stabilized, life quality improved, child-pugh score declined, and liver function turned better. (2) The level of HBV DNA decreased at least 10(3) copies/ml. HBeAg of 33.3% patients (13/39) became negative. (3) Among the 10 patients who stopped lamivudine of their own accord, and came again after 3 - 6 months because of hepatitis B recurring, two were treated with interferon for one month, then turning to liver-protecting methods for deteriorating, the other eight only received liver-protecting and immune-regulating treatment, whose liver function improved. CONCLUSIONS: Long term treatment with lamivudine for active cirrhosis following chronic hepatitis B can improve liver function and life quality, prevent exacerbation. And it is not advisable to use interferon for hepatitis B relapsing after lamivudine withdrawing.

[Clinical research on the treatment effect of autologous dendritic cell vaccine on the patients with chronic hepatitis B].

OBJECTIVE: To investigate the treatment effect of autologous HBsAg-loaded dendritic cells (DCs) on patients with chronic hepatitis B (CHB). METHODS: Monocytes were isolated from fresh peripheral blood of 19 CHB patients by Ficoll-Hypaque density gradient centrifugating and cultured with plastic -adherence method. DCs were induced and proliferated from the monocytes with granulocyte-macrophage clony stimulating factor (GM-CSF) and interleukin-4 (IL-4) for seven days. After being incubated with HBsAg for two hours, DCs were injected to patients subcutaneously twice at the interval of two weeks. HBV DNA level, alanine aminotransferase (ALT) level, and HBV markers in the serum of patients were tested every two months. RESULTS: 11 of the 19 (57.9%) patients responded to DC-treatment clinically. The rates of HBeAg clearance and HBeAg/anti-HBe seroconversion were 52.6% (10/19) and 26.3% (5/19) respectively, and the copies of HBV DNA decreased by 10(1.77 2.39) (t = 3.13, P < 0.01). Two patients who were treated in combination with lamivudine had complete clinical response. There was no difference in the trial effect between the DC treatment and the other two antiviral methods, and in the efficient rate between the patients whose ALT levels were high before treatment and those whose ALT levels were normal. CONCLUSION: The autologous HBsAg-loaded DCs can effectively suppress HBV replication, reduce virus load in serum, eliminate HBeAg and promote HBeAg/ anti-HBe seroconversion. The patients whose ALT levels are high or normal can response clinically to DCs treatment. DCs in combination with lamivudine can eliminate virus more effectively.