【作者】 赵振刚；

【导师】 韩涛； 高英堂；

【作者基本信息】 天津医科大学 ， 内科学， 2010， 硕士

【摘要】 目的：研究乙肝病毒变异与乙肝相关性慢加急性肝衰竭之间的关系。方法：通过收集20例乙肝相关性慢加急性肝衰竭(ACLF)患者及19例慢性乙型肝炎(CHB)患者血清及临床资料,建立多个基因区巢氏PCR方法,对所有患者HBV DNA的前S区、S区、BCP区、C区、前C区、P区进行扩增、测序,应用BioEdit(7.0.9.0)生物学分析软件进行比对分析,使用SPSS17.0软件应用Fisher确切概率法及t检验进行统计学分析,进一步了解乙肝病毒变异与乙肝相关性慢加急性肝衰竭之间的关系。结果：一、入组患者临床资料：乙肝相关性慢加急性肝衰竭组(简称ACLF组)男14例,女6例,CHB组男14例,女5例,两组性别组成无差异；所有入组患者年龄24-71岁,ACLF与CHB组年龄分别为49.70±14.33 vs 40.37±10.91岁,ACLF组年龄高于CHB组(P值小于0.05)；ALB、ALT、TBIL在ACLF和CHB组的情分别为：27.31±3.39 vs43.16±4.35(g/L)、303.20±408.92 vs 146.79±307.79(U/L)、287.37±133.02 vs 19.75±19.99(μmol/L), ACLF组ALB水平低于CHB组,而TBIL明显高于CHB组(P值小于0.05)；ACLF和CHB组HBV DNA分别为：5.82±1.52 vs 7.67±0.85(copies/ml, log10), ACLF组较CHB组低(P值小于0.05)；ACLF组中HBeAg(+)者8例,HBeAg(-)12例,CHB组中HBeAg(+)者14例,HBeAg(-)5例,ACLF组HBeAg(-)患者所占比例高于CHB组(P值小于0.05)；ACLF组中HBeAg(+) HBeAg(-)患者的HBV DNA水平分别为：6.21±1.51vs 5.70±1.60 (copies/ml,log10),二者无差异(P值大于0.05)；ACLF组HBeAg(+)、HBeAg(-)患者的好转率分别为62.50%(5/8)vs 66.67%(8/12)，二者无差异(P值大于0.05)。二、测序结果：1.HBV基因型：20例ACLF患者中16例成功完成S区测序,其基因型均为C型；19例CHB患者中17例完成S区测序,其中仅两例为B型,余均为C型。2.前S区：缺失变异在两组中均有出现,前S2区缺失变异在ACLF组发生率为35.00%,而在CHB组则较低为5.26%，ACLF组更易发生前S2区缺失变异(P值小于0.05)。3.BCP区+前C区+C区：G1896A、G1899A、G1896A+G1899A在ACLF组发生率明显偏高于CHB组(56.25% vs 12.50%,37.50% vs 6.25%,31.25% vs 0.00%, P值均小于0.05)；而A1762T、G1764A、A1762T+G1764A变异在两组中均频发(81.25% vs75.00%,87.50% vs 75.00%,81.25% vs 75.00%),但两组间比较无明显差异(P值大与0.05)；T1753C、A1853C、T1754C、C1766T和T1768A变异在两组中发生率均较低(18.75%vs 6.25%, 0.00% vs 0.00%,0.00% vs 0.00%, 6.25% vs 0.00%, 6.25% vs 0.00%),且在两组间比较差异均无统计学意义(P值大于0.05)。4.P+S区：G587A变异在两组均未发生；ACLF组C766G发生率高于CHB组(25.00% vs 0.00%,P值小于0.05)。所有患者核苷类似物的耐药变异位点检测仅发现1例ACLF组患者出现了YMDD变异。5.1例ACLF患者HBV DNA基因序贯分析：该患者基因型为C型,发病初期出现了前S2区缺失变异、G1896A+G1899A+A1762T+G1764A、T1961C及C766G变异,经过治疗病情好转后前S2区缺失变异、G1896A+G1899A及T1961C变异均消失,但A1762T+G1764A及C766G变异仍持续存在。结论：前S2区缺失变异、G1896A、G1899A和G1896A+G1899A变异与乙肝相关性慢加急性肝衰竭的发生相关；A1762T、G1764A、A1762T+G1764A变异在慢性乙型肝炎及乙肝相关性慢加急性肝衰竭中普遍存在。更多还原

【Abstract】 AIM:To investigate the relationship between Hepatitis B Virus(HBV) mutations and HBV-related acute-on-chronic liver failure(ACLF).METHODS:Serum were collected from 20 HBV-related ACLF and 19 chronic hepatitis B(CHB) patients.A sensitive and reliable nested PCR assay was performed to analyze the target HBV gene fragments,which were further sequenced and analyzed by BioEdit(7.0.9.0).Fisher’s exact probability or t test were calculated by SPSS 17.0 to analyze the relationship between HBV mutations and HBV-related ACLF.RESULTS:Cilinical data:Between ACLF and CHB group,there were significant difference of age(49.70±14.33 vs 40.37±10.91years, P<0.05),ALB (27.31±3.39 vs 43.16±4.35g/L,P<0.05),TBIL(287.37±133.02 vs 19.75±19.99μmol/L, P< 0.05), HBeAg(+) (40.00% vs 73.68%, P<0.05),and HBV DNA (5.82±1.52 vs 7.67±0.85copies/ml,log10, P<0.05),whereas there was no significant difference in gender and ALT. No difference of HBV DNA and clinical improvement rate was found between HBeAg(+) and HBeAg(-) ACLF patients [6.21±1.51vs5.70±1.60 (copies/ml,log10),62.50% vs 66.67%, P> 0.05].Squence analysis:1. Genotype: 16 of 20 ACLF cases were sequenced successfully,all of 16 ACLF patients were genotype C.In CHB group,17 of 19 cases were sequenced completely,genotype B was only found in 2 cases,and other 15 were genotype C.2. We detected PreS1 and PreS2 deletion, PreS2 were found in 8 cases,7 patients(35.00%) were in the ACLF group and 1 patient(5.26%) was in the CHB group.There were significant correlations between PreS2 deletion and HBV-related acute-on-chronic liver failure (P<0.05).3. G1896A,G1899A and G1896A+G1899A mutations occurrence were significantly high in ACLF patients than in CHB patients (56.25% vs 12.50%, 37.50% vs 6.25%,31.25% vs 0.00%,P<0.05).Although A1762T,G1764A, A1762T+G1764A mutations were observed in two groups (81.25% vs 75.00%, 87.50% vs 75.00%,81.25% vs 75.00%),no difference was found between two groups(P>0.05).4. No G587A mutation was found in two groups,4 of 16 patients presented C766G mutation (25.00% vs 0.00%,P<0.05)and only 1 patient presented YMDD mutation.5. A patient with ACLF was monitored more than 2 months,HBV mutations analysis showed that PreS2 deletion,G1896A+G1899A+A1762T+ G1764A,C766G and T1961C mutations occurred at early stage of ACLF,PreS2 deletion,G1896A+G1899A and T1961C muataions disappeared with clinical condition improved,but A1762T+G1764A and C766G mutations still existed.CONCLUSIONS:PreS2 deletion,G1896A,G1899A and G1896A+G1899A mutations were associated with HBV-related ACLF.Although A1762T,G1764A and A1762T+G1764A mutations often existed in both HBV-related ACLF and CHB patients,there was no relationship bewteen A1762T,G1764A,A1762T+G1764A mutations and HBV-related ACLF.更多还原