Objective To identify and isolate the variant gene associated with gastric adenocarcinoma and clone the fragment of variant gene.Methods By arbitrarily primer polymerase chain reaction (AP-PCR), DNA samples from 5 matched gastric adenocarcinoma and non-tumor gastric tissues were analysed. Results The produced AP-PCR profiles were different in each matched gastric adenocarcinoma and non-tumor gastric tissue. One differentiated amplified DNA fragments PW2.2 from a matched gastric adenocarcinoma were cloned. The result of Southern blot hybridization with PW2.2 as a probe showing that this fragment was also found in some other gastric adenocarcinoma samples. Conclusion AP-PCR fingerprinting assay can be used to identify and clone the variant genes associated with gastric adenocarcinoma.
Objective To discuss the changes of c-kit/scf mRNA and protein in guinea pig gallbladder fed on high cholesterol diet. Methods Twenty guinea pigs were divided into two equal groups of 10 each：the control group and lithogenic group. Normal diet and high cholesterol diet was given to each group respectively. The period of stone permeation was six weeks. RT-PCR and Western blot were used to determin the expressions of c-kit and scf mRNA and protein. Results RT-PCR results showed that the expressions of c-kit mRNA（t=6.985，P＜0.01） and scf mRNA （t=6.028， P＜0.01）decreased significantly in lithogenic group compared with the control group. Western blot results showed that the expressions of c-kit protein （t=10.256， P＜0.01） and scf protein （t=9.586， P＜0.01）decreased significantly in lithogenic group compared with the control group. Conclusions The expressions of c-kit/scf mRNA and protein decrease during the formation of cholesterol gallstones in guinea pigs fed on high cholesterol diet. Inhibition of c-kit/scf pathway may play a role in the formation of cholesterol gallstones.
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 evaluate the potential of specific mRNA marker keratin 19(K19) to detect micrometastasis by reverse transcriptase polymerase chain reaction (RT-PCR) .Methods One hundred and ninty four regional lymph nodes harvested from 6 cases of benign diseases, 4 cases of breast carcinoma, 5 cases of gastric carcinoma and 12 cases of colorectal carcinoma patients were examined by conventional pathology and amplifying tissue specific K19 mRNA by RT-PCR separately, then the two methods were compared with each other. Results None of the 34 lymph nodes which were pathological metastasis-negative from benign diseases expressed K19 mRNA by RT-PCR, all of the 28 regional lymph nodes which were pathological metastasis-positive from malignant cases showed trains of K19 mRNA by RT-PCR. Of the 132 lymph nodes which were pathological metastasis-negative from malignant cases, 11 lymph nodes were detected with micrometastasis by genetic diagnosis.Conclusion Genetic diagnosis of lymph node micrometastasis is more sensitive than conventional pathology and has diagnostic value and merits further study.
Objective To investigate whether mutations exist in codon 58 and codon 347 of the rhodopsin gene in patients with autosomal dominant retinitis pigmentosa(ADRP)． Methods Point mutations at codons 58 and 347 were detected by restriction endonuclease digestion of exons 1 and 5 amplified by polymerase chain reaction(PCR)．This method was applied to screen genomic DNAs from 57 patients of 38 families with ADRP and 60 normal controls． Results Four patients from one family of ADRP were confirmed to have a point mutation at the second nucleotide of codon 58,and 6 patients from two families of ADRP were found to have a mutation at codon 347．None of these mutations were found in 60 normal subjects． Conclusion It is suggested that molecular genetic heterogeneity exists within ADRP and some subtypes of ADRP are caused by points mutations of the rhodopsin gene． (Chin J Ocul Fundus Dis,1998,14:108-110)
Objective To verify whetheriris pigment epithelial cells(IPECs)possess the similar potential of specific phagocytosis to retinal outer segments(ROS) with retinal pigment epithelialcells(RPECs). Methods IPESc were isolated from neonatal bovines with Hu's method,and were cultured.The cultured cells were identified by immunohistochemical methods with antibodies to cytokeratin and s-100.Total RNA of IPECs was extracted by Trizol.The specific primers for mannose-receptor andbeta;-actin were designed according to their sequence from Genbank.The mRNA expression of these proteins in the IPECs was analyzed by reverse transcription polymerase chain-reaction (RT-PCT).Results The Cultured IPECs have no contamination of other cells .The extracted RNA was ideal and had no degradation.RT-PCR analysis showed that mannose-receptor's mRNA was expressed in cultured IPECs in vitro.ConclusionCultured IPECs may express the mannose-receptor,and may have similar potential of phagocytosis to ROS with REPCs.
Objective To investigate the polymorphism of the vitamin D receptor gene (VDR)TaqⅠin relation to diabetic retinopathy. Method Fragment length discrepant allele specific PCR(FLDAS-PCR) were used to determine VDR genetypes in 158 patients with diabetic retinopathy and in 198 normal subjects. Results The frequency distribution of VDR genotypes in diabetic retinopathy patients was 106 (67.1%) in TT, 33(20.9%) in Tt, 19(12.0%) in tt; and in normal persons was 165 (83.3%) in TT, 23(11.6%) in Tt, 10 (5.1%) in tt. There was a significant difference between diabetic retinopathy patients and normal persons in distribution of VDR gene TaqⅠgenotypes(Plt;0.05). Conclusions There is some distribution alterations of VDR gene polymorphism in diabetic retinopathy patients. (Chin J Ocul Fundus Dis, 2006, 22: 94-96)
Objective To explore the effect of connective tissue growth factor on the pathogenesis of hypertrophic scar and keloid tissue. Methods The content of hydroxyproline was determined and the expression of connective tissue growth factor gene was detected by the reverse transcription-polymerase chain reaction and image analysis technique in 5 normal skins, 15 hypertrophic scars and 7 keloid tissues. Results The contents of hydroxyproline in the hypertrophic scar(84.10±1.76) and keloid tissue (92.38±2.04) were significantly higher than that of normal skin tissue (26.52 ± 4.10) (P lt; 0.01). The index of connective tissue growth factor mRNA in the hypertrophic scar (0.78 ± 0.63) and keloid tissue (0.84 ± 0.04) were higher than that of normal skin tissue ( 0.09 ± 0.25) (P lt; 0.01). Conclusion Connective tissue growth factor may play an important role in promoting the fibrotic process of hypertrophic scar and keloid tissue.