Purpose To investigate the relationship between mitochondrial DNA 11778 mutation and clinical characteristics of patients with Laber is hereditary optic neuropathy(LHON). Methods PCR RFLPs (MaeⅢ) and mutation specific primer PCR(MSP-PCR) were used simultaneously to detect mitochondrial DNA 11778 mutation. Results Among 10 subjects who habored 11778 mutation,one was a carrier and nine were patients with LHON.Of the nine patients,six were males and three were females.The age of onset ranged from 12 to 25 years old and the onset interval of the two eyed varied between 0 to 6 months. The visual acuity was CF/10cm-0.1 except one who lost her vision after delivery but recovered gradually.The results of visual field,VEP and color vision were abnormal but ERG and systemic status were all normal. Conclusion Molecular biological detection of the ten subjects showed that they all habored mtDNA 11778 mutation.The existence of carrier and visual recovery imlied that mtDNA mutation was a primary cause of LHON,but other factors such as endocrine disorder might influence the pathogenesis of LHON. (Chin J Ocul Fundus Dis,1998,14:156-158)
Objective To construct specifically expressed vascular endothelial growth factor (VEGF)165 gene in retina. Methods Rho promoter, specifically expressed in retina, was amplified by polymerase chain reaction (PCR) from the genomic DNA of a BLAB/C rat, then it was cut with restriction enzymes and cloned into the plasmid pcDNA3.1+-VEGF165 to form recombinant plasmid pcDNA3.1+-rho-VEGF165. The correct recombinant plasmid pcDNA3.1+-rho-VEGF165 was identified by restriction enzymes and PCR, and was transferred by jetPEI into cultured human navel vein endothelial cells and human retinal pigment epithelial (RPE) cells. The expression of VEGF protein in human navel vein endothelial and RPE cells was detected by immunocytochemical staining and protraction of the growth curve of the cells. Results In human RPE cells, the expression of VEGF protein was more in recombinant plasmidpcDNA3.1+-rho-VEGF165 than that in plasmidpcDNA3.1+-rho-VEGF165 ; in human navel vein endothelial cells, no obvious difference of the expression of VEGF protein between recombinant plasmid pcDNA3.1+-rho-VEGF165 and plasmid pcDNA3.1+-rho-VEGF165 was found. Conclusions The construction of pcDNA3.1+-rho-VEGF165 carrier may provide the basic material for the study of the nosogenesis of VEGF in retinal neovascularization, and establish the foundation to set up the model of transgenic mice with VEGF specific expressing in retina. (Chin J Ocul Fundus Dis, 2005,21:106-108)
Objective To investigate the expression of T cell receptor (TCR) Vβ8.3 gene on CD4+ T lymphocytes in the rats with experimental autoimmune uveoretinitis (EAU). Methods Eighteen Lewis rats were divided into EAU, complete Freund′s adjuvant, and the control group. Inter photoreceptor retinoid-binding protein (IRBP) R16 peptide was synthesized using Fmoc procedure for induction of EAU. Magnetic absorption cell sorting (MACS) me thod was used to isolate the CD4+T lymphocytes from the spleen of the rats. Flow cytometry was used to monitor the efficiency of isolation. The expression of TCR Vβ8.3 gene segment on CD4+T lymphocytes was determined by fluorescent quantitative polymerase chain reaction. Results EAU was successfully induced in the Lewis rats immunized with IRBP R16 peptide. The proportion of CD4+T lymphocytes isolated by means of MACS was statistically higher than that before isolation (P＜0.001). The expression of TCR Vβ8.3 gene segment on CD4+ T lymphocytes in EAU rats was significantly higher than that in the control (P＜0.05). Conclusions There is a predominant usage of antigen-specific TCR Vβ 8.3 gene in EAU rats induced by IR BP R16 peptide, which may serve as a target for immunotherapy of EAU. (Chin J Ocul Fundus Dis，2004，20：165-167)
Objective To detect and analyse the mutations in rhodopsin gene of members in a family affected by autosomal dominant retinitis pigmentosa (ADRP). Methods Using the polymerase chain reaction (PCR), we amplified exon 1-5 of rhodopsin gene in patients with ADRP,and analyzed it with direct sequence measuement. Results The Gly-182-Asp mutation in the rhodopsin gene was detected in most of affected members of this ADRP family, but no mutation was detected in two affected members and the control ones. Conclusion We cannot regard the Gly-182-Asp mutation in the rhodopsin gene as the pathagenic factor of the ADRP family. It is likely there is a new gene next to the rhodopsin gene. (Chin J Ocul Fundus Dis, 2002, 18: 256-258)
ObjectiveTo systematically review the diagnostic value of PCR-single-strand conformational polymorphism (PCR-SSCP) method for detecting rpoB gene mutation of rifampin-resistant mycobacterium tuberculosis. MethodsSuch databases as PubMed, Web of Science, The Cochrane Library (Issue 2, 2014), CBM, VIP and WanFang Data were electronically and comprehensively searched for relevant studies on the diagnostic value of PCR-SSCP method for detecting rpoB gene mutation of rifampin-resistant mycobacterium tuberculosis from inception to January 1st, 2014. Literature screening according to the inclusion and exclusion criteria, data extraction and methodological quality assessment were completed by two reviewers independently. Meta-analysis was then conducted using Meta-Disc 1.4 and Stata 12.0. ResultsA total of 10 studies were included involving 1 299 cases. The results of meta-analysis showed SEN=0.92 (95%CI 0.90 to 0.94, P=0.019 3), SPE=0.97 (95%CI 0.95 to 0.98, P < 0.000 1), +LR=23.68 (95%CI 8.71 to 64.37, P < 0.000 1), -LR=0.10 (95%CI 0.06 to 0.15, P=0.023 1), DOR=257.16 (95%CI 96.82 to 683.02, P=0.020 0), and SROC area under the curve (AUC) was 0.971 5, and Q* was 0.922 3. The results of sensitivity analysis (after removing studies with sample size less than 100, Chinese studies and QUADAS more than 10 studies) showed that, the results were stable with reliable conclusion. ConclusionPCR-SSCP method has a fairly high value in the diagnosis of rpoB gene mutation of rifampinresistant mycobacterium tuberculosis.
ObjectiveTo explore the variation of the structure of the intestinal flora between healthy people and patients with obstructive jaundice perioperatively. MethodsFrom February 2013 to August 2014, 20 patients with obstructive jaundice and 10 healthy persons (normal control group) in our hospitol were selected as the research object. The first stool specimens of the research object after admission were obtained and the total fecal bacteria DNA were extracted. After polymerase chain reaction amplification, the changes in the structure of bacterial flora were dynamic observed by using denaturing gradient gel electrophoresis (DGGE), and the gel bands were analyzed by using Quantity One software. The similarity and diversity of flora structure, and principal component analysis (PCA) were analyzed. ResultsSignificant differences of colonic microflora were found between patients with obstructive jaundice and healthy people; advantage intestinal flora in obstructive jaundice patients was significant lower than the normal control group. With the extension of time and degree of obstruction aggravated, a descending trend was found in number, abundance, and diversity of the intestinal microflora (P < 0.05). ConclusionThere is significant differences in the structure of colon bacteria in patients with obstructive jaundice and healthy persons.
Objective To investigate the mutations of the gene in Chinese patients with X linked juvenile retinoschisis (XLRS), and to provide the genetic diagnosis and consultation of heredity for the patients and their families. Methods Genomic DNA was isolated from leukocytes of 29 male patients with XLRS, 38 female carriers and 100 normal controls (the patients and the carriers were from 12 families). All 6 exons of XLRS1 gene were amplified by polhism (SSCP) assay. The positions and types of XLRS1 gene mutations were determined by direct sequencing. Results Eleven different XLRS1 mutations were identified in these 12 families, including one frameshift mutation due to base loss of the first exon: c.22delT（L9CfsX20）, one nonsense mutation due to base loss of the first exon (Trp163X), one splice donor site mutation(c.52+2 Trarr;C; IVS1+2T to C), and eight missense mutation due to base replacement（Ser73Pro, Arg102Gln, Asp145His, Arg156Gly, Arg200Cys, Arg209His, Arg213Gln, and Cys223Arg). No gene mutation was detected in the control group. Four new mutations included frmaeshift mutation（L9CfsX20）and mutations of Asp145His, Arg156Gly, and Trp163X at the fifth exon. A newly discovered non-disease-related polymorphism (NSP) was the c.576C to T (Pro192Pro) change at the sixth exon. Conclusion Eleven different XLRS1 mutations were detected, which is the cause of XLRS in Chinese people. The detection of gene mutations may provide the guidance of genetic diagnosis and the consultation of family heredity for the patients and their families. (Chin J Ocul Fundus Dis, 2006, 22: 77-81)
Objective To analyze the association of human leucocyte antigen (HLA)-DRB and -DQB alleles with Ealesprime; disease, and to investigate the potential immunogenetics mechanism of Ealesprime; disease. Methods Gene loci of HLA-DRB and -DQB1 alleles were detected by polymerase chain reaction-sequence specific primer (PCR-SSP) in 27 Han-nationality patients with Ealesprime; disease in Northern China and 30 age and sex-matched normal persons as control, then statistics package for social science (SPSS) for Windows ver 13.0 software was used to analyze the distribution features of frequency of HLA-DRB and -DQB1 alleles in the two groups. Results Compared with the control group, the frequency of HLA-DRB104 allele increased obviously in the patients with Ealesprime; disease［odds ratio (OR)=3.20 ，OR 95% confidence interval（CI）=1.00-10.21, and P=0.047］. Simultaneously, no statistically significant difference of the distribution of any other DRB or DQB1 allele between the two groups was found (Pgt;0.05). Conclusions In hannationality people in Northern China, DRB104 is positively associated with Ealesprime; disease, suggesting that DRB104 may confer a major influence on Ealesprime; disease. Turbulence of immune function begotten by infect-agents attack may occur in the individuals with Ealesprime; disease due to the specific hereditary diathesis of HLA, which may cause the occurrence and development of Eales disease. (Chin J Ocul Fundus Dis, 2006, 22: 90-93)
The aim of the this study was to search for bacterial DNA sequences in cholesterol gallstones with negative bacterial culture by NP-PCR technique. Bacterial gene fragments were amplified in vitro from DNA which were extracted from cholesterol gallstones in gallbladder for identifying the existence of bacteria. The gallbladder gallstones of 30 patients were analysed. Bacterial DNA was found in the stones of 26 patients, indicating that most cholesterol gallstones harbor bacterial DNA.
ObjectiveTo investigate the expressions and clinical significance of human telomerase reverse transcriptase (hTERT) mRNA and γglutamyl transpeptidase mRNA-H (GGT mRNA-H) in the peripheral blood of small hepatocellular carcinoma (HCC) patients. MethodsThe expressions of hTERT mRNA and GGT mRNA-H were detected in the peripheral blood of thirty patients with small HCC by RT-PCR, eighteen patients with benign liver diseases, and twelve normal volunteers. ResultsThe positive rate of hTERT mRNA and GGT mRNA-H expression in patients with small HCC were 80.0% (24/30) and 46.7%(14/30), respectively. In patients with hepatitic cirrhosis the positive rate of hTERT mRNA expression was 33.3% (6/18), while the expression of GGT mRNA was not detected. Both the expressions of hTERT mRNA and GGT mRNA-H were negative in all normal volunteers. The combination analysis of hTERT mRNA and GGT mRNA-H expression achieved positive rate of 86.7% in the diagnosis of small HCC, which was significantly higher than the positive rate of AFP (26.7%)， Plt;0.05. ConclusionThe hTERT mRNA and GGT mRNA-H are significantly expressed in small HCC patients, the combination analysis of hTERT mRNA and GGT mRNA-H seems to be useful in the early diagnosis of small HCC.