Calicecotomy Combined with Trans-renal Parenchyma Pneumatic Lithotripsy for Complicated Staghorn Renal Calculi / 中国微创外科杂志

Objective To evaluate the efficacy of calicecotomy combined with trans-renal parenchyma pneumatic lithotripsy for complicated staghorn renal calculi.Methods The severe hydrocalycosis was incised using electrocautery,then pneumatic lithotripsy was performed and the broken stones were taken out.For those patients with stenotic entrance to renal calyces without hydrocalycosis,we stabbed into the stones with the lithotriptic pole(1 mm in diameter) through renal parenchyma and took the broken stones out of the entrance.Results The renal pedicle were not blocked in 19 cases.The operation time was 90-150 minutes,with a mean of 120 minutes.There was no blood transfusion with the blood loss ranging form 100 to 250 ml.The procedures were successful in 17 cases without residual stones after operation;intraoperative residual sand-like calculi were found in 1 case and removed by irrigation and drainage through nephrostomy tube;intraoperative missing calyceal calculi occurred in 1 case and were cleared by extracorporeal shock wave lithotripsy(ESWL).A follow-up for 10-60 months(mean,18 months)in 15 patients showed recurrence in 2 ones,and the stones were removed by ESWL.Conclusions Calicecotomy combined with trans-renal parenchyma pneumatic lithotripsy for complicated staghorn renal calculi has the advantages of less blood loss and definite efficacy.

Study on the clinic,neuro-electrophysiology and molecular biology of Machado-Joseph disease / 临床神经病学杂志

Objective　To study the clinic, neuro-electrophysiology and molecular biology of Machado-Joseph disease (MJD).Methods　Family visiting, physical examination and the blood samples were analysed on molecular biology in 44 members of a family with MJD.The cases of inpatients were examined on cerebrospinal fluid and neuro-electrophysiology.Results　10 patients of the family attacked,which were consisted with autosomal dominant inheritance type. Age of the onset was 8～38 years old. The clinical characteristic was progressive severe spinocerebellar of ataxia,faciolingual myokymia,bulging eyes.Change of denervated muscle was revealed by neuro-etectrophysiological examination. Light atrophy was observed in cerebellar,brain stem, spinal cord.The genetic defect of MJD was located the long arm of chromosome 14 between D14S280 and D14S81, their distance was 3.0 cm.All tested patients had their CAG repeated expansion from 72 to 84 in the MJD gene.Conclusion　MJD is a neuro-degenerative disorder of autosomal dominant inheritance. The disease was clinically characterized by progressive severe spinocerebellar ataxia, no obvious changes of cerebrospinal fluid,neuro-electrophysiology, CT and MRI.The genetic defect of MJD was located the long arm of chromosome 14.The number of CAG repeated expansion mutation was associated with the age of the onset.

The immune responses to hepatitis B gene vaccine in mice and the immune adjuvant effect of cytokines / 医学研究生学报

Objectives：To observe the effect of eukaryotic expression vectors coding IL-2 and IL-12 on immune responses induced by DNA immunization of HBV surface antigen（pCR3.1-S）in BABL/c(H-2d) and the protection against P815 mastocytoma cells stable expressing HBV surface antigen in mice after immunized with HBV gene vaccine.Methods：The immunization was performed by intramuscular injection,three weeks later,we directly inoculated P815-HBV-S into mice by subcutaneous injection .Tumor growth was measured every five days.Anti-HBs in serum was detected by ELISA and HBsAg specific cytotoxic T lymphocytes (CTLs) activity was measured by 51 Chromium release assay.Results：Eight weeks after immunization,the A value of mice serum in 450 nm and CTLs activity of mice codiog IL-2 and IL-12 eukaryotic expression vectors were significant higher(P<0.05) than that of mice intramuscular injected HBV-S DNA vaccine,these values are significant higher than that of mice injected pCR3.1(P<0.05).The spleen cells CTLs activity have decreased obviously after treated with anti-CD8+ monoclonal antibody and have no significant change after treated with anti-CD4+ monoclonal antibody.The HBV-S gene vaccine could evidently inhibit the tumor growth,prolong the survival period (>38.2 days) and improve the survival rate in mice.Conclusions：The DNA vaccine of HBV ( pCR3.1-S) had strong antigenicity in cellular and humoral immunity and had marked killing effect on HBV infected cells in vivo，which could be promoted by vector coding murine IL-2 or IL-12.CTLs activity was performed by CD8+ cells.

Tissue inhibitors of metalloproteinase-1 and -2 (TIMP-1 and TIMP-2) mRNA and antigens location in the liver of patients with cirrhosis / 中华传染病杂志

Objective To study the expression and distribution of TIMP 1 and TIMP 2 in liver tissue of cirrhosis patient and to investigate the roles and pathogenesis of TIMP 1 and TIMP 2 in liver cirrhosis. Methods TIMP 1 and TIMP 2 proteins and mRNA were detected with immunohistochemistry and in situ hybridization methods using monoclonal antibodies and cDNA probes. Results mRNA and proteins of TIMP 1 and TIMP 2 were detected in all the liver tissues from 40 liver cirrhosis patients, all in cytoplasm but not nucleus. TIMP 1 and TIMP 2 were found co exist in all samples, while TIMP 1 concentration was higher. Conclusions mRNA and protein of TIMP 1 and TIMP 2 are found in all the cirrhosis patient samples. Liver TIMP 1 and TIMP 2 concentrations increase with the progression of liver cirrhosis, decrease the degradation of extracellular matrix proteins, resulting in the initiation and the development of liver fibrosis and liver cirrhosis.

CONSTRUCTION,IDENTIFICATION AND EXPRESSION OF FRAMEWORK PLASMID pAd.HCV-CE1 OF ADENOVIRUS EXPRESSION VECTOR OF HCV-CE1 / 解放军医学杂志

The HCV core gene and E1 gene derived from the plasmid pBRTM/HCV1 3011 by using polymerase chain reaction(PCR) was inserted into the backward position of cytomegalovirus(CMV) immediate early promotor element of framework plasmid (pAd.CMV Link 1) of adenovirus expression vector, then the recombinant plasmid pAd.HCV CE1 was obtained. The inserted DNA of pAd.HCV CE1 was confirmed to be HCV core and E1 gene by endonuclease, PCR and sequencing. HCV core gene was expressed transiently with lipofectamine 2000 coated in human hepatoblastoma 7721 cells which was confirmed by immunofluorescence. The results indicate that the recombinant framework plasmid of adenovirus expression vector pAd.HCV CE1 can express HCV core and E1 gene. This should be useful to pack adenovirus expression vector which can express HCV core and E1 gene.

INTRACELLULAR INHIBITION OF VIRAL GENE EXPRESSION BY HAMMERHEAD RIBOZYMES AGAINST HEPATITIS C VIRUS 5′ NCR AND C REGIONS / 解放军医学杂志

To investigate the potential use of synthetic stabilized ribozymes for the treatment of chronic hepatitis C virus (HCV) infection,we designed and synthesized 2 hammerhead ribozymes (Rz213 and Rz498) targeting conserved sites in the 5′noncoding region (NCR) and C gene of HCV RNA.Constructed to the eukaryotic vector pcDNA3, the two ribozymes were respectively or simultaneously transfected with lipofectamine into WISHnc transgenic cells, which could express permanently HCV C luciferase protein under the control of HCV 5′NCR.The expression of C luciferase was measured by luminometer.The results showed that the luciferase activities were significantly down regulated in the WISHnc cells, and the inhibitory rates were 42.94%～67.81% within 7 days after ribozymes transfection. There was no significant differences between Rz213 or Rz498 and co transfection, but adding the target site of ribozymes might prevent host cells from the loss of ribozyme therapeutic effect due to viral gene mutation.

Quantitation of hepatitis C virus RNA in amniotic fluid of gravida infected by hepatitis C virus / 中华妇产科杂志

Objective To detect hepatitis C virus (HCV) RNA in amniotic fluid of gravida and investigate mother-to-infant transmission of HCV. Methods Thirty-four HCV seropositive gravida (experimental group) were engaged. Fluorescence quantitative polymerase chain reaction (PCR) based on amplisensor assay and reverse transcription -PCR (RT-nPCR) was used. Serum HCV RNA positive sera were genotyped by RFLP analysis of PCR products from 5′NC region. Sera and amniotic fluid samples of 40 normal gravida were set as the control group. Results In the experimental group, HCV RNA was detected in amniotic fluid (5.9%, 2/34) of 2 cases. HCV RNA titers were 10 5 and 10 6 copy/ml respectively. No HCV RNA was detected in the amniotic fluid and sera of the control (n=40). Conclusions HCV RNA was rarely detected in amniotic fluid. The amniotic fluid is not the main route of HCV mother-to-infant transmission.

Quantitive analysis of the inhibition of HCV IRES mediated HCV core protein expression in cells by inhibitor RNA / 中华传染病杂志

Objective To study the inhibition of HCV IRES mediated HCV core protein expression in cells by inhibitor RNA. Methods Plasmid pcRz-IRNA, a eukaryotic expression vector with IRNA and two self cleavage ribozyme overhang at both sides respectively, was constructed and co-transfected with pcHCVcluc (containing HCV NCR, core and Luc genome) into the HHCC cell line (Human Hepatocellular Carcinoma cell line). Immunoflurescence tests were applied to detect the co-transfected cells, which were thereafter analysed with confocal microscope quantitatively. Luciferase activity was valued using Luc Assay System (Promega). Results The cotransfected cells expressed HCV core protein, and the fluorecein in which was reduced significantly in comparison with control. Conclusions IRNA can inhibit the expression of HCV IRES mediated core protein in the cotransfected cells.

Specific cellular immune responses and effect against subcutaneously transplanting tumor induced by DNA vaccine of HBV surface gene in mice / 中华传染病杂志

Objective To observe the specific immune responses and the protection against P815 mastocytoma cells stably expressing HBV surface antigen in H-2 d mice after DNA immunization of HBV surface antigen gene (pCR3.1-S). Methods The immunization was performed by intramuscular injection of DNA vaccine (pCR3.1-S). P815-HBV-S was inoculated subcutaneously into mice three weeks after DNA immunization. The tumor growth was measured every five days. HBsAg specific cytotoxic T lymphocyte (CTL) activity was measured by 51 Chromiunm release assay. Results HBV DNA vaccine can evidently inhibit the tumor growth, prolong the survival period and improve the survival rate in mice. Meanwhile, HBsAg specific CTL activity was obviously increased after DNA immunization. Conclusions The results show that the DNA vaccine, pCR3.1-S, has strong antigenecity in cellular immunity and has marked killing effect on HBV infected cells in vivo. DNA vaccine against HBV may be useful for both prophylactic and therapeutic purposes.

Study on the clinic,neuro electrophysiology and molecular biology of Machado Joseph disease / 临床神经病学杂志

Objective To study the clinic, neuro electrophysiology and molecular biology of Machado Joseph disease (MJD).Methods Family visiting, physical examination and the blood samples were analysed on molecular biology in 44 members of a family with MJD.The cases of inpatients were examined on cerebrospinal fluid and neuro electrophysiology.Results 10 patients of the family attacked,which were consisted with autosomal dominant inheritance type. Age of the onset was 8~38 years old. The clinical characteristic was progressive severe spinocerebellar of ataxia,faciolingual myokymia,bulging eyes.Change of denervated muscle was revealed by neuro etectrophysiological examination. Light atrophy was observed in cerebellar,brain stem, spinal cord.The genetic defect of MJD was located the long arm of chromosome 14 between D 14 S 280 and D 14 S 81 , their distance was 3.0 cm.All tested patients had their CAG repeated expansion from 72 to 84 in the MJD gene.Conclusion MJD is a neuro degenerative disorder of autosomal dominant inheritance. The disease was clinically characterized by progressive severe spinocerebellar ataxia, no obvious changes of cerebrospinal fluid,neuro electrophysiology, CT and MRI.The genetic defect of MJD was located the long arm of chromosome 14.The number of CAG repeated expansion mutation was associated with the age of the onset.

INHIBITION OF HBV BY RIBOZYME IN ADENOVIRUS EXPRESSION SYSTEM / 解放军医学杂志

To investigate the inhibition of HBV expression by multi-target ribozyme-expressing adenovirus vector in vivo, the ribozyme and muta-ribozyme were cloned into adenovirus vector pAd-CMV. By using the adenovirus expression system, the two plasmids were co-transfected into 293 cell line with adenovirus gene recombinant plasmids pJM17. The recombinant adenovirus were detected by plaque assay. Then the recombinant adenovirus infected the 2.2.15 cells. Using ELISA, dot-bolt hybridization and image analysis system, the inhibition of HBV expression was detected. The ribozyme and muta-ribozyme were cloned into the adenovirus vector pAd-CMV. The recombinant adenovirus could infect 2.2.15 cells and express the ribozyme. The ribozyme could decrease the expression of HBeAg by 87.1%. All these results strongly indicate that ribozyme can inhibit HBV expression in the cell, which represents a potential gene therapy for HBV infection.