本文關鍵詞：HBsAg陽性孕婦胎盤組織HBV cccDNA檢測及臨床意義研究　出處：《桂林醫(yī)學院》2015年碩士論文　論文類型：學位論文

  更多相關文章： 乙型肝炎病毒 胎盤 宮內感染 原位PCR 共價閉合環(huán)狀DNA

【摘要】：目的:乙型肝炎病毒(Hepatitis B Virus,HBV)主要通過母嬰垂直傳播引起,其中宮內感染是最主要的傳播途徑,是造成免疫預防失敗的主要原因。本實驗將通過一種新型的原位檢測方法,檢測HBsAg陽性孕婦胎盤組織中HBVcccDNA存在情況,進一步探討其意義及乙型肝炎病毒母嬰傳播機制,研究其與宮內感染發(fā)生的關系。方法:以52例北京302醫(yī)院婦產中心進行產檢并分娩的HBsAg陽性的孕婦產婦胎盤組織作為實驗組,5例乙型肝炎病毒五項標志物均為陰性的健康孕婦的胎盤組織作為陰性對照組,同時以HBsAg與HBcAg均陽性的肝癌肝穿樣本作為陽性對照組。收集孕婦及其新生兒臨床資料,胎兒娩出后30min內于臍帶中央切取2.0cm×2.0cm×1.5cm大小的胎盤組織,立即置于10%中性福爾馬林中固定,經梯度酒精脫水、石蠟包埋、4.5μm連續(xù)切片后使用。Envision免疫組織化學染色方法檢測石蠟包埋胎盤組織各類細胞中HBsAg、HBcAg表達情況;原位聚合酶鏈反應方法(in situ polymerase chain reaction,IS-PCR)檢測胎盤組織中HBV cccDNA表達情況。每次實驗均設立陰性對照組與陽性對照組。以新生兒出生后24h內股靜脈血中HBsAg、HBeAg或HBV DNA任一項為陽性作為新生兒發(fā)生宮內感染的標志。結果:1.52例HBsAg陽性孕婦中,共有21例新生兒發(fā)生宮內感染,總的感染率為40.38%(21/52)。大三陽組孕婦新生兒HBV感染率為89.47%(17/19),高于小三陽組孕婦12.12%(4/33),P0.05,差異有統(tǒng)計學意義。HBV DNA高載量組(104~109IU/ml)與低載量組(102~104 IU/ml)新生兒HBV感染率分別為77.78%(14/18)、25.00%(3/12),P=0.014,差異有統(tǒng)計學意義。2.免疫組織化學染色實驗組胎盤組織HBs Ag與HBcAg沒有陽性信號,陽性對照組HBsAg與HBcAg均為陽性,陰性對照未檢出陽性信號。3.原位聚合酶鏈反應在HBs Ag陽性孕婦胎盤組織中未檢出HBV cccDNA陽性信號,陽性對照組HBV cccDNA檢測有陽性信號,陰性對照組未檢測到HBV ccc DNA陽性信號。結論:胎盤組織有可能不是發(fā)生HBV宮內感染的主要途徑,可能通過其他途徑發(fā)生感染。宮內感染的高危因素有待于大樣本量的實驗證實。
[Abstract]:Objective: hepatitis B virus (Hepatitis B, Virus, HBV) mainly through vertical transmission caused by intrauterine infection, which is the main route of transmission, is a major cause of immunization failure. This experiment will be through a novel in situ detection methods, HBVcccDNA detection of HBsAg positive placenta in the presence of further study the significance of maternal transmission of hepatitis B virus and study its mechanism, and the relationship between the occurrence of intrauterine infection. Methods: 52 cases of No.302 Hospital of Beijing Maternity Center for production inspection and delivery of the placenta HBsAg positive pregnant women as the experimental group, 5 cases of hepatitis B virus markers were five placentas of healthy pregnant women as negative the negative control group, while HBsAg and HBcAg were positive in liver samples as positive control group. Clinical data of pregnant women and their newborns, 30min after delivery of fetus The placenta is 2.0cm * 2.0cm * 1.5cm size in the central cord cut, immediately placed in 10% neutral formalin, after gradient alcohol dehydration, paraffin embedding, 4.5 m serial sections after using.Envision immunohistochemical staining method to detect the paraffin embedded placental tissue of various cell HBsAg, HBcAg expression; in situ polymerase chain (in situ polymerase chain reaction method, reaction, IS-PCR) expression of HBV cccDNA detection in placenta of each experiment were set up. The negative control group and positive control group. The newborn within 24h after femoral venous blood HBsAg, HBeAg or HBV DNA for any positive signs as neonates of intrauterine infection. Results: 1.52 cases HBsAg positive pregnant women, a total of 21 cases of neonatal intrauterine infection, the total infection rate was 40.38% (21/52). 3 group of neonatal HBV infection rate was 89.47% (17/ 19), higher than that of small Sanyang Group 12.12% (4/33), P0.05, there was a significant difference between the.HBV DNA group (104~109IU/ml) with high load and low load group (102~104 IU/ml) neonatal HBV infection rate was 77.78% (14/18), 25% (3/12), P=0.014, the difference has statistical significance of immunohistochemical.2. staining in experimental group placenta HBs Ag and HBcAg have no positive signal, positive control group HBsAg and HBcAg were all positive and negative controls were detected.3. positive signals in situ polymerase chain reaction cccDNA HBV positive signals were not detected in HBs Ag positive placenta, positive control group HBV cccDNA detected positive signal, the negative control group, positive signals were not detected by HBV CCC DNA. Conclusion: the placenta may not be the main route of HBV intrauterine infection, may be infected by other means. The risk factors of intrauterine infection remains to be confirmed by large sample experiments.

【學位授予單位】：桂林醫(yī)學院
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R714.251

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