Detection Of HLA-A/DRB1 Gene Polymorphism In The Congenital Absence Of Vagina Patients And The Follow-up For The Patients With Sigmoid Neovagina

The congenital absence of vagina (MRKHS) is the second most common cause of primary amenorrhea. The etiology of the disease is still unknown. It was often suggested that it was a disease of multifatorial/polygenic inheritance. MRKHS involves congenital absence of the vagina, uterus and cervix. These patients were infertility and be failure to achieve intercourse. The disease decreased the quality of life of the patients. Sigmoid vaginal construction is the main method to treat MRKHS. To date , most of the Chinese literatures about the disease was reported about the surgical technique of the vaginal reconstruction. And there have been few reports of the etiology or the follow-up for the disease.The current study includes the following three parts as bellowsPartⅠThe HLA-A/DRB1 Gene Polymorphism Detected in Patients of the Congenital Absence of VaginaObjective: To analyze the association of the polymorphism of HLA-A/DRB1 alleles with MRKHS, and to offer the clues searching for susceptive gene of the disease. Methods: The objects were selected in the MRKHS patients who were follow-up or consulted in the Department of Obstetrics & Gynecology of the second hospital of Hebei Medical University from October 2005 to January 2007. MRKHS group : Include 30 Han-nationality patients with MRKHS in Hebei Province. All of the patients had normal female 46 XX karyotype. Control group : To select randomly 300 cases who were Han-nationality, healthy and no-relationship female in Hebei Province. The data were provided by Bone Marrow Bank of Hebei Blood Center. Gene loci of HLA-A,DRB1 alleles were detected by flow polymerase chain reaction sequence specific oligonucleotide probe (PCR-SSOP): To take 2 mL venous blood sample in the cases of MRKHS group→to anticoagulate the sample by EDTAK2→to extract the DNA by centrifugal collumn method→to amplificate and hybridize the DNA→to analyze the sample by the luminex100 equipment for hybridization. The statistical software SASv6.12 was used to analyze the data. The data were contrasted withχ2 test or Fisher Exact test. P values less than 0.05 considered significant. Calculate the odds ratio (OR) of the gene when the difference was significant.Results1 The genotype of the HLA-A alleles: 11 alleles were detected. The gene frequency of HLA-A*33 in MRKHS patients was significantly higher than that in controls (0.1167 vs 0.0300, P<0.05, OR>1). The gene frequency of HLA-A*24 in MRKHS patients was significantly lower than that in controls (0.0667 vs 0.1717, P<0.05, OR<1).2 The genotype of the HLA-DRB1 alleles: 12 DRB1 alleles were detected. The gene frequency of HLA-DRB1*13 in MRKHS patients was significantly higher than that in controls (0.1333 vs 0.0217, P<0.05, OR>1).Conclusions: In Han-nationality people in Hebei Province of China, maybe there have some relation between the allele of HLA-A*33 /HLA-DRB1*13 with MRKHS. It offer the clues searching for susceptive gene of MRKHS.PartⅡInvestigated Study on Quality of Life and Mental Status in Patients with Congenital Absence of VaginaObjective: To understand the quality of life (QOL) and the mental status in patients with congenital absence of vagina(MRKHS).Methods: 43 MRKHS patients and 43 control subjects were measured with the generic quality of life inventory-74 (GQOLI-74), self anxiety scale (SAS) and self depression scale (SDS). The statistical software SASv6.12 was used to analyze the data. The data were contrasted with t test (t'test when variances were unequal),χ2 test and analysis of correlation. P values less than 0.05 considered significant.Results: The scores of subfactors such as mental function and social function were lower in the MRKHS group (59.15±13.12 vs 67.57±13.51, 54.40±11.41 vs 62.90±12.40, P<0.01). The total score of GQOLI-74 in the MRKHS group was significantly lower than that in the control group (57.48±8.34 vs 62.82±9.79,P<0.01). The score of SAS in the MRKHS patients was significantly higher than that in the control group (45.98±10.32 vs 39.09±9.42). There was negative correlation between the total score of GQOLI-74 and the SAS (r =-0.64,P =0.0001). The mean index of SDS in the MRKHS patients was significantly higher than its in the control group (0.50±0.13 vs 0.41±0.11). There was negative correlation between the total score of GQOLI-74 and the mean index of SDS (r =-0.65,P =0.0001).Conclusions: It is suggested that the MRKHS patients have poor life quality. The patients more tented to have the anxiety and depression symptom.PartⅢMorphologic Change and Expression of Neuronal Nitric Oxide Synthase of the Neovaginal Mucosa in Patients Undergoing Vaginal Construction by Sigmoid ColonObjective: The aim of the study was to evaluate the changes in the histological morphology and the expression of neuronal nitric oxide synthase (nNOS) in the vaginal colonic mucosa of patients undergoing vaginal construction by sigmoid colon and to explore the putative reason for the vaginal discharge after artificial vaginal construction.Methods: The objects were selected in the MRKHS patients who were follow-up from October 2005 to January 2007. After knowing the fact 10 patients agreed to take part in present study. Biopsy of the neovaginal mucosal were taken in the upper one-third and the lower one-third (3 cm distal to the hymen) of the sigmoid neovagina. Biopsy of the normal mucosal were taken during the operation of sigmoid vaginal construction from 2005 to 2006. To investigate the morphologic change of the sigmoid neovaginal mucosa by the microscope after the samples had been dyed with the HE dyeing method. To select randomly 3 samples from the samples of the upper one-third of the sigmoid neovagina to investigate by the electron microscope. The specimen immunochemically stained for nNOS and were analyzed semiquantitatively. The statistical software SASv6.12 was used to analyze the data. The data were contrasted with Fisher Exact test. P values less than 0.05 considered significant.Results1 The general situations: Every case of MRKHS group had a normal female 46, XX karyotype. The average time was 27.6 months between operation and biopsy. The average time was 7.5 months when the discharge of the neovagina became to decrease after the operation. The mean time of the sexual intercourse was 18.3 months. The average PH value of the neovagina was 7.9.2 Pathological observations: The upper one-third of the sigmoid neovagina: There had no significant histological change in mucosal epithelial layer. The muscularis mucosa was hyperplasy and pachynsis. The density of the submucosal nerve plexus was significantly higher than that in controls (3.6±1.5 vs 1.7±0.8, P<0.05). The lower one-third of the sigmoid neovagina: There was squamous metaplasia in the mucosa in 7 cases. The number of the glands was decreased. Some of the glands were atrophy. The mesenchym of the mucosa was increased. The muscularis mucosa and the submucosa were disappeared.3 Observation with scanning electron microscope: There were some ultrastructural changes in the mucosa of the sigmoid neovagina. The surface of the mucosa was not smooth. The number of secretory vacuole from the goblet cells was decrease. Observation with transmission electron microscope: Some of the cristae in mitochondria were fusion or disappeared. Some rough endoplasmic reticulum was degranulation.4 The expression of the nNOS: There were nNOS expression in the samples of MRHKS group and control group. Compared to control group, the expression of nNOS in the epithelium and the gland of colonic mucosa were decreased significantly(P<0.05).Conclusions1 It was possible that there had some relation between the morphologic change of the neovaginal mucosa and the decrease of the sigmoid neovaginal discharge. Maybe the morphologic change induced by the functional alteration of the colon and the microenviroment change of the neovagina.2 The significance of the decrease of the nNOS expression hasn't been clear.These results suggested that there maybe have some relation between the decrease of the nNOS expression in the sigmoid neovaginal mucosa and the decrease of the artificial vaginal discharge.