Hepatitis C virus (HCV), one of the major pathogens of viral hepatitis, causes significant hazards in humans. Interferon treatment in combination with ribavirin is used as the first line clinical treatment for HCV infection. However, good response to this treatment has only been observed in few patients and repeated recurrence has also been reported frequently. Therefore, new antiviral agents and therapies are in urgent demand. Here, we report a newly constructed Escherichia coli RNase P based M1GS ribozyme that can specifically and efficiently target the core gene of HCV. The guide sequence (GS) of this M1IGS was designed according to the sequence of the core coding region of HCV genome. The GS was then covalently linked to the 3' terminus of M1 RNA, the catalytic subunit of RNase P from Escherichia coli. The specification of this sequence-specific ribozyme, M1GS, was then examined using an in vitro cleavage assay. The cytotoxicity and its activity in inhibition of HCV gene expression and viral proliferation were further studied in vivo. Our results show that the reconstructed M1GS ribozyme displayed obvious catalytic activity in cleaving target mRNAs fragment in vitro. Notable reduction in the expression of HCV core protein and a 1 000-fold reduction in viral growth were also observed in cultured HCV infected Huh7.5.1 cells expressing the functional M1GS ribozyme. This study demonstrated a direct evidence for the antiviral activity of the customized M1GS-HCV/C141 ribozyme, and thus provided a promising new strategy for clinical treatment of HCV infection.
Antiviral Agents ; pharmacology ; Escherichia coli ; genetics ; Genetic Engineering ; Hepacivirus ; genetics ; physiology ; RNA, Catalytic ; genetics ; pharmacology ; RNA, Guide ; genetics ; Ribonuclease P ; genetics ; Viral Core Proteins ; genetics

OBJECTIVE To study the molecular cytogenetic alterations of transitional cell carcinoma of the urinary bladder with exfoliated cells by fluorescence in situ hybridization(FISH) analysis of chromosome-specific probes.METHODS FISH was performed using 3,7,9 and 17 of chromosome-specific probes to examine chromosome aberration of exfoliated cells in 50 urine samples from patients with transitional cell urinary bladder carcinoma.RESULTS The frequency of numerical aberration of chromosomes 3,7,9 and 17 was 28%,32%,56% and 38% in urinary exfoliated cells,respectively.Loss of chromosome 9 was the most common finding,but it was not correlated with pathological grade of cancer and stage of the disease.Abnormality of chromosomes 3,7 and 17 was associated with the clinical stage.CONCLUSIONS A number of chromosome aberrations are detected in transitional cell carcinoma of the urinary bladder by FISH technique which provides a basis for further understanding of its molecular pathogenesis.It is a rapid,accurate and very sensitive method and can be used in clinical diagnosis.

Objective To study the protective effects on ovarian function by caloric restriction (CR) and its mechanism.Methods Thirty female C57BL/6 mice of 8 weeks old were randomly divided into two groups,including ad libitum (AL) group and caloric restriction (CR) group.The general situation and ovarian function of those mice were compared and evaluated.Ovarian follicles were counted by hematoxylin-eosin staining.Anti-Miillerian Hormone(AMH) mRNA expression of the ovary were detected by using real-time PCR.The concentrations of serum estradiol,progesterone of the mice were measured by ELISA.And the fertility of mice by mating trials were evaluated,SIRT3,Hypoxia inducible factor 1α (HIF-1α) and catalase (CAT) mRNA expression of the mice ovaries were detected by Real-Time PCR.Results The total follicles were 546 in CR mice and 286 in AL mice.The proportion of primordial follicles were 38.6％ (211/546)in ovaries of CR mice and 29.4％ (84/286)in ovaries of AL mice,which reached statistical difference.The proportion of atretic follicles 5.3％ (29/546) in ovaries of CR mice,compared with 16.8％ (48/286) in AL mice,was significantly decreased (P ＜ 0.05).The AMH mRNA expression in CR mice ovaries was 3.37 times of that of AL mice (P ＜ 0.05).The serum concentration of estradiol in CR mice was up to (5.3 ± 1.6) pmol/L,which was much higher than (3.6 ± 1.6) pmol/L in AL mice.While,the progesterone concentration of (0.4 ±0.3) nmol/L in CR mice was lower than (1.4 ± 0.8) nmol/L in AL mice (P ＜ 0.05).Fertility and survival of offsprings were both improved in CR mice.The expression level of SIRT3 mRNA in CR mice ovary was 1.39 times,CAT was 1.55 times and HIF-1 α was 0.31 times of those in AL mice (P ＜ 0.05).Conclusions Caloric restriction can delay the ovary aging process through reduce follicle depletion by suppressing follicle recruitment and ovulation.The function of ovarian reserve and reproductive endocrine was effectively protected.Caloric restriction can reduce the incidence of follicular atresia,its mechanism might be associated with anti-oxidative stress.

Objective To explore the clinical characteristics and diagnostic methods of cerebral schistosomiasis.Methods The data of CT,MRI and the cerebral spine fluid(CSF) immunoassay in 42 patients with cerebral schistosomiasis were analyzed retrospectively.Results Forty cerebral schistosomiasis patients were diagnosed by the iconography and immunoassay,1 by the postoperative pathological examination and 1 by diagnostic treatment with praziquantel.Conclusions CT and MRI are of great value to determination of the position and nature of the cerebral schistosomiasis.The sensitivity and accuracy of MRI are better for indicating the main pathological changes of the disease than those of CT.The CSF immunoassay plays an important role in the differential diagnosis of cerebral schistosomiasis.

Lymph metastasis has great impact on the prognosis of pancreatic cancer patients, which can relfect the biological and invasive potential of pancreatic cancer. However, currently, there is no standard in the clinical management of the lymph metastasis in pancreatic cancer. In this report, we will discuss and summarize the followings:lymph metastatic rate and its impact on prognosis, the rule of lymph metastasis, sentinel lymph node, intra-operative lymph nodes mapping, TNM staging, regional lymph nodes resection, number of lymph nodes examined, lymph node ratio, guiding adjuvant treatments, lymphatic targeted therapy.

Objective To investigate the changes of CD 4+CD25+CD127 low regulatory T ( Treg ) cells in peripheral blood samples from patients with lung cancer and their correlation with clinicopathologic features of lung cancer .Methods Flow cytometry was used to measure the percentages of CD 4, CD8, nat-ural killer ( NK) and Treg cells in peripheral blood samples collected from 160 patients with lung cancer and 60 healthy subjects .The correlations between the levels of Treg cells and clinicopathologic features of lung cancer were analyzed .The percentages of Treg cells in 60 patients with lung cancer before and after surgery were compared .Results The percentages of CD 4+and NK cells and the ratios of CD 4+/CD8+cells in pa-tients with lung cancer were significantly lower than those in healthy control , while the percentages of Treg cells in patients with lung cancer were decreased as compared with those in healthy subjects .The percenta-ges of Treg cells in patients with advanced cancer were significantly higher than those in patients at early stage (P<0.05), which dropped significantly after surgery (P<0.01).Conclusion The results of this study indicated that Treg cells in patients with lung cancer might inhibit antitumor immune responses and correlate with the progression of cancer .It would be worthwhile to check Treg cells in patients with lung cancer for monitoring the prognosis and treatment effects .

Background and purpose:Ultrasound is a regular screening method of solid pseudopapillary tumor of the pancreas (SPTP). This study was to summarize the diagnostic value of ultrasound to SPTP.Methods:Clinical and ultrasound data of 62 SPTP cases in Fudan University Shanghai Cancer Center were retrospectively collected and analyzed.Results:Five cases of SPTP were undetected by ultrasound in the group. The features of ultrasound including: large mass located at the body and tail of the pancreas, clear boundary and regular shape, low ultrasound with uneven signal, or low signal mixed with no signal. A few cases have calciifcation and blood signal. Most of the cases presented no dilation of main pancreatic duct and bile duct and regional lymph nodes enlargement. Conclusion:Ultrasound can be used to detect SPTP which has special ultrasound signal features.

OBJECTIVE:To establish the method for simultaneous determination of the content of berberine hydrochloride, baicalin and osthole in Jinchan zhiyang capsules. METHODS:HPLC method was adopted. The determination was performed in Hypersil BDS C18 column with mobile phase consisted of methanol-acetonitrile-0.1%phosphoric acid-three triethylamine(50∶30∶19∶1, V/V/V/V) at the flow rate of 1.0 mL/min. The detection wavelengths were set at 265 nm (berberine hydrochloride),280 nm (baicalin)and 322 nm(osthole). The column temperature was set at 30 ℃,and sample size was 10 μL. RESULTS:The linear range of berberine hydrochloride,baicalin and osthole were 80-800 ng(r＝0.999 8),60-600 ng(r＝0.999 9),60-600 ng(r＝0.999 6),respectively. RSDs of precision,stability and reproducibility tests were all lower than 2.0%. The limits of quantitation were 80,60,60 ng,respectively,and the limits of detection were 24,20,20 ng,respectively. The recovery rates were 97.4%-98.3%(RSD＝0.33%,n＝6),98.4%-99.6%(RSD＝0.42%,n＝6)and 96.9%-99.0%(RSD＝0.92%,n＝6),respectively. RSDs of durability tests were all lower than 1.2%. CONCLUSIONS:The method is simple, accurate, precise, stable, reproducible and durable. It can be used for simultaneous determination of berberine hydrochloride,baicalin and osthole in Jinchan zhiyang capsules.

Objective To observe the effect and safety of the aversion therapy with furazolidone on patients with alcohol dependence.Methods 90 patients with alcohol dependence were randomly divided into the aversion therapy group and the control group with 45 cases in each group. The cases of the aversion therapy group were treated by aversion therapy with furazolidone and those of the control group were treated with routine therapy. The changes of the blood pressure, pulse and respiratory rate before and after drinking were observed and the rate of successful abstinence in one year was investigated.Results The effect of the aversion therapy group treated with furazolidone was significantly better than that of the control group ( P<0.05). The aversion therapy was safe.Conclusion The aversion therapy with furazolidone is more effective and safe.

OBJECTIVE:To establish the method for simultaneous determination of berberine hydrochloride and baicalin in Jianpi zhixiening granules. METHODS:HPLC switching walvelength method was adopted. The determination was performed on Hypersil BDS C18 column with mobile phase consisted of methanol-0.45％ phosphoric acid solution-triethylamine(50:49:1,V/V/V) at the flow rate of 1.0 mL/min. The detection wavelength was set at 265 nm(berberine hydrochloride)and 280 nm(baicalin). The column temperature was set at 30 ℃,and sample size was 10 μL. RESULTS:The linear range of berberine hydrochloride and baicalin were 60.3-312.8 ng(r=0.9997)and 81.5-368.9 ng(r=0.9999). The limits of quantitation were 0.6668,0.7740 ng,andthe limits of detection were 0.2226,0.2580 ng,respectively. RSDs of intermediate precision,stability and repeatability tests were all lower than 1.0％. The recoveries were 96.48％-99.30％(RSD=1.06％,n=6) and 95.20％-99.39％(RSD=1.66％,n=6), respectively. RSDs of durability test were all lower than 2.0％. CONCLUSIONS:The method is simple, precise, stable, reproducible,accurate and durable. It can be used for simultaneous determination of berberine hydrochloride and baicalin in Jianpi zhixiening granules.