La、hVAP-33、eIF2B γ和HCV IRES特异性小干扰RNA抑制HCV的复制和表达

目的 证实La自身抗原、33kD人类囊相关膜蛋白(hVAP-33)和真核细胞翻译起始因子第3亚单位(eIF2B γ)是HCV在细胞内的协同感染因子,通过抑制Huh7细胞内这些因子的表达可抑制HCV复制和表达. 方法 分别设计合成3条HCV内部核糖体进入位点(IRES)的小干扰RNA(siRNAs),转染Huh7-HCV细胞后筛选出其中沉默效率最高的1条.以HCV假病毒感染Huh7细胞后48 h,分别以上述HCV IRES siRNA和之前试验中已经筛选出的La、hVAP-33和eIF2B γ特异性siRNAs单独或者不同组合转染Huh7-HCV细胞,利用荧光定量PCR方法检测HCV核心基因并计算△△CT值(相对定量法),比较不同siRNAs及组合对目的基因沉默的效率;同时以Western blot观察HCV核心蛋白表达量的差异.结果 La自身抗原与IRES特异性siRNA共转染对HCV表达的抑制效率最高,使HCV核心蛋白基因相对表达量下降了约41％;4种基因特异性siRNAs 对HCV在Huh7细胞中的复制和表达均有不同程度的抑制作用,La、hVAP-33和eIF2Bγ特异性siRNAs分别与IRES siRNA联合均较其单独转染对目的基因的抑制效率高.结论 可以认为La自身抗原、hVAP-33和eIF2Bγ是HCV在宿主细胞内的协同感染因子,通过对Huh7细胞内协同感染因子及HCV IRES基因沉默后可以显著减少HCV的表达.     

丙型肝炎病毒抑制DNA损伤修复相关基因生长阻滞和DNA损伤诱生蛋白45α的表达

目的 探讨HCV感染对DNA损伤修复相关基因生长阻滞和DNA损伤诱生蛋白45 α(GADD45 α)表达的影响. 方法 建立全基因HCV JFH1感染的Huh7,5.1细胞模型.用相对荧光定量PCR和Western blot分别检测HCV JFH1感染和未感染的Huh7.5.1细胞中GADD45 α mRNA和蛋白质的表达水平.组间数据比较用单因素方差分析.结果 HCV JFH1感染的Huh7.5.1细胞内有HCV RNA高水平复制及HCV NS5A蛋白质和核心蛋白的表达.与未感染Huh7.5.1细胞相比,HCV JFH1感染72h的细胞内GADD45αmRNA和蛋白质相对表达量均明显降低,分别为0.57±0.09比1.00±0.11和0.28±0.03比1.00±0.07,差异均有统计学意义(F值分别为75.407和560.04,P值均＜0.01). 结论 HCV下调GADD45 α的转录和蛋白质表达,影响DNA损伤修复,这可能是HCV感染致肝癌发生的机制之一.     

丙型肝炎病毒F蛋白抑制肝癌细胞增殖功能的研究

目的 研究HCV F蛋白的生物学功能.方法 扩增目的基因片段,通过酶切连接方法将编码HCV F蛋白的基因片段克隆至载体pSEB-3Flag中,用构建好的重组质粒pSEB-3Flag-F转染Huh7、SMMC7721细胞,经Blasticidine抗性筛选,拟建立HCV F蛋白稳定表达的细胞株,用RT-PCR方法检测HCV-F基因的表达.通过MTS、细胞计数、结晶紫以及流式细胞检测实验观察F蛋白对转染细胞增殖和细胞周期的影响.计量资料分析采用t检验.结果 成功建立了HCV F基因稳定表达的细胞株Huh7-F和SMMC7721-F,MTS、细胞计数实验均证实Huh7-F和SMMC7721-F细胞较对照细胞增殖速度减慢,Huh7-F稳定细胞株结晶紫定量A值为1.072±0.070,对照组为1.387±0.005(t=-6.347,P＜0.05);SMMC7721-F稳定细胞株A值为1.594±0.007,对照组为1.921±0.090(t=-4.81,P＜0.05).流式细胞仪检测Huh7-F稳定细胞株48hS期细胞百分比分别为47.12％±0.04％,对照组S期细胞数为55.32％000±1.45％(t=-7.99,P＜0.05);SMMC7721-F稳定细胞株S期细胞百分比分别为30.75％±0.09％,对照组S期细胞数为33.23％±0.28％(t=-5.91,P＜0.05).结论 稳定转染HCV F基因可以明显抑制Huh7细胞、SMMC7721细胞的增殖.     

丙型肝炎病毒非结构蛋白5A影响p53抑制肝癌细胞甲胎蛋白表达的分子机制

目的了解丙型肝炎病毒(HCV)非结构蛋白5A(NS5A)对p53抑制甲胎蛋白(AFP)基因表达的影响及分子机制。方法采用质粒转染技术及微粒体酶免疫法观察p53蛋白对Huh7肝癌细胞AFP表达的抑制作用及HCV NS5A对其抑制作用的影响;蛋白印迹实验观察HCV NS5A对p53蛋白表达的影响;谷胱甘肽转移酶沉淀实验鉴定HCV NS5A与p53蛋白能否相互作用形成复合物。结果转染pRc/CMV空质粒的Huh7细胞上清液AFP浓度为(14 322±2412)ng/ml,转染pCNS5A质粒的Huh7细胞上清液的AFP 浓度为(13 843±3218)ng/ml,两组问t=1.42,P>0.05;转染pC53-NS3质粒的Huh7细胞上清液的AFP 浓度为(10 241±1326)ng/ml,与上述两组比较,t值分别为2.41及2.38,P值均<0.05;pCNS5A和pC53- NS3共转染者AFP浓度为(14 582±1238)ng/ml,与pC53-NS3单独转染组比较,t=3.12,P<0.01; pCNS5A和pC53-NS3共转染者和pC53-NS3单独转染者Huh7细胞p53蛋白表达无变化。在谷胱甘肽转移酶沉淀实验中,加入谷胱甘肽-p53融合蛋白后出现HCV NS5A蛋白条带,而仅加入谷光甘肽者则未出现此条带。结论p53蛋白能抑制Huh7细胞AFP的表达,HCV NS5A能减轻p53蛋白对AFP表达的抑制作用。HCV NS5A不影响p53蛋白的表达但能与p53蛋白结合形成复合物是使p53功能失活的分子机制。     

基于2a基因型丙型肝炎病毒自身启动子的单顺反子复制子的构建和功能测定

目的 构建丙型肝炎病毒(HCV)单顺反子复制子,研究其在Huh7.5和Huh7.1细胞中的复制功能,为研究HCV复制规律和抗病毒药物的筛选建立模型. 方法 用Quick change点突变方法删除pJ6JFH 1B1aRL质粒上的Core-E1-E2-p7-NS2片段(约3090 bp),得到△pJ6JFH1B1aRL,然后测序,选择序列正确的克隆,用AgeⅠ和AvrⅡ酶切回收目的片段约2280bp,同时用AgeⅠ和AvrⅡ酶切pSGRJFH1和其突变体质粒,回收载体片段.将目的片段和载体片段连接,构建由HCV-IRES启动的单顺反子复制子pSGRm-JFH IblaRL以及其变异突变体pSGRm-JFH 1b1aRLGND,用其RNA转染Huh7.5和Huh7.1细胞,研究复制子在细胞中的复制情况.结果 经过Quick change和多步克隆方法成功构建HCV-IRES启动的单顺反子复制子,复制子RNA转染Huh7.5和Huh7.1细胞72h后复制达高峰,96h复制开始下降,而对照的突变体从24h至96h均没有复制.结论 成功构建了HCV-IRES单顺反子复制子,转染Huh7.5和Huh7.1细胞后在不同时间均有复制.     

含EGFP报告基因单顺反子HCV亚基因组复制子载体的构建和表达

目的构建含增强型绿色荧光蛋白(EGFP)报告基因的HCV复制子表达载体,并实现其在细胞中的复制表达。方法用分子生物学基因克隆技术对HCV 2a型复制子的基因进行改造,用EGFP基因替代HCV基因组中的包膜基因(E1和E2)体外构建重组单顺反子HCV亚基因组复制子真核表达质粒pcDNA-JFH1-EGFP,经限制性内切酶酶切分析和测序鉴定;脂质体介导转染人肝癌细胞系Huh-7细胞,用荧光显微镜观察EGFP表达,采用半定量RT-PCR方法检测重组复制子的HCV RNA负链,采用Western blot检测HCV NS3蛋白的复制表达,并观察IFN-α对重组质粒表达的HCV RNA复制的抑制作用。结果构建的4个重组质粒酶切分析与预期相符,HCV亚基因复制子表达载体中未发生EGFP和HCV编码区读码框架改变,转染重组载体Huh-7细胞检测到HCV负链及EGFP和HCV NS3蛋白表达。转染后48h,1 000IU/ml和2 000IU/ml IFN-α处理的细胞HCV RNA表达水平分别为未处理组的20.0%和7.6%。结论含EGFP报告基因的单顺反子HCV亚基因组复制子表达载体pcDNA-JFH1-EGFP构建成功,在Huh-7细胞中能有效复制表达,为进一步研究HCV提供了实验平台。     

小泛素样修饰物特异性蛋白酶3通过调节脂滴水平体外促进HepG2细胞丙型肝炎病毒复制

目的 探究丙型肝炎病毒(HCV)感染细胞小泛素样修饰物特异性蛋白酶3(SENP3)对脂滴水平的调节及其对HCV复制的影响。方法 以感染性HCV病毒颗粒感染人HepG2细胞,采用Western blot法检测感染前后SENP3蛋白表达水平。采用脂质体法转染SENP3-siRNA至HepG2细胞,在感染HCV后采用Western blot法检测HCV核心蛋白表达量,采用qRT-PCR法检测HCV RNA水平,采用油红O染色观察细胞脂滴水平。以软脂酸处理敲低SENP3后的HepG2细胞,采用qRT-PCR法检测感染HCV后HCV RNA水平。结果 在HCV感染HepG2细胞后1 d、3 d和6 d,HCV 核心蛋白相对表达量分别为0.01±0.00、0.17±0.02和0.43±0.04,SENP3蛋白相对表达量分别为0.23±0.04、0.42±0.03和0.46±0.04,差异显著(P<0.05);转染SENP3-siRNA可有效降低SENP3表达并降低HCV感染后细胞HCV核心蛋白表达水平;SENP3敲低细胞HCV RNA相对水平为54.2±11.4%,较转染非特异性siRNA细胞显著降低(P<0.01);敲低SENP3蛋白表达后细胞脂滴数量显著减少;敲低SENP3的HepG2细胞感染HCV后HCV RNA相对水平为58.2±5.2%,较转染非特异性siRNA细胞显著降低(P<0.01),而以软脂酸处理敲低SENP3的HepG2细胞,在感染HCV后HCV RNA相对水平为74.6±6.4%,较未经软脂酸处理细胞显著升高(P<0.01)。结论 在HCV感染肝细胞时SENP3可通过调节脂滴代谢促进HCV复制。     

携带IFN—β基因的丙型肝炎病毒微型基因组构建及抑制病毒复制的效应

目的构建丙型肝炎病毒（HCV）反应性干扰素（IFN）-β的微型基因组及其抑制效应的初步评价。方法提取poly I：C刺激的淋巴细胞总RNA,RT-PCR扩增IFN-β基因,克隆入前期构建的HCV微型基因组pT7-5U△131-315rI3Urz;PCR并鉴定插入方向,将反向插入的质粒命名为pT7-5U△131-315rIFNI3Urz。将微型基因组5U△131-315rIFNI3URz克隆入pcDNA3.1载体,获得pCMV-5U△131-315rIFNI3URz。将体外转录的5U△131-315rIFNI3URz RNA转染Huh7.5BB7细胞,48 h后检测IFN-β的表达。将pCMV-5U△131-315rIFNI3URz转染Huh7.5 JFH-1细胞系,荧光实时定量PCR法测定细胞中HCV RNA。结果成功建立了Huh7.5JFH-1细胞系;含有IFN-β的微型基因组在复制子细胞和HCV感染细胞中可以特异表达IFN-β;携带IFN-β的HCV微型基因组可以降低细胞中病毒的RNA拷贝水平,具有一定的剂量依赖效应。结论反向插入IFN-β基因的HCV微型基因组进入HCV感染细胞内表达IFN-β并靶向抑制病毒的复制,为抗HCV感染研究提供了一个新的方法和思路。     

醛酮还原酶1B10基因沉默对MHCC97H细胞生长和基因表达的影响

目的 探讨醛酮还原酶1B10(AKR1B10)基因在肝癌发生和发展过程中的作用及其机制.方法 化学合成AKR1B10序列特异性小干扰RNA,用脂质体LipofectaminrTM2000介导转染人肝癌细胞系MHCC97H,设实验组转染AKR1B10-小干扰RNA,设阴性对照组转染非编码序列双链小RNA,设空白对照组不作转染.用实时定量PCR、Western blot和酶活性检测法检测AKR1B10 mRNA和蛋白的表达情况,用细胞计数试剂盒CCK-8检测转染细胞的生长增殖变化,用流式细胞仪检测膜联蛋白V/碘化丙啶双标记的凋亡细胞变化,并用实时定量PCR检测c myc、c-fos、N-ras、ki-67、caspas 3、bax肿瘤相关基因的表达变化.用析因设计的方差分析和完全随机方差分析进行组间比较,LSD法进行多重比较.结果 转染24、48 h和72 h后,AKR1B10 mRNA的相对表达量在实验组分别为0.382±0.048、0.098±0.008和0.257±0.032;阴性对照组分别为0.797±0.092、0.742±0.086和0.794+0.105;空白对照组分别为0.808±0.118、0.716±0.083和0.706±0.067.不同转染时间和不同实验组中AKR1B10 mRNA的表达比较,F值分别为11.650和566.971,P值均＜0.01,差异均有统计学意义.转染24、48 h和72 h后,实验组的AKR1B10mRNA表达较空白对照组被明显抑制,抑制率分别为52.7%±5.6%、86.3%±4.4%和63.7%±6.1%,以转染48 h的抑制率最高(t=80.68,P＜0.01);阴性对照组的表达水平接近空白对照组(P＞0.05).Western blot结果显示,转染24、48 h和72 h后,实验组的AKR1B10蛋白表达水平均有下降,其中以转染72 h的降低幅度最大;阴性对照组的表达水平接近空白对照组.转染24、48、72h和96h后,实验组450nm处吸光度值分别为1.465±0.040、1.724±0.106、1.934±0.069和2.377±0.163;阴性对照组分别为1.528±0.044、2.019±0.091、2.711±0.204、3.151±0.159;空白对照组分别为1.567+0.057、2.102±0.099、2.642±0.198、3.069±0.180;不同转染时间和不同实验组间比较,F值分别为128.092、36.535,P值均＜0.01,差异均有统计学意义.转染了AKR1B10-siRNA的MHCC97H细胞生长受到抑制.与空白对照组相比,实验组的细胞生长抑制率在转染48、72 h和96 h分别为18.0%±1.6%、26.1%±3.2%和22.5%±1.1%,t值分别为19.197、13.093和23.553,P值均＜0.01,差异均有统计学意义;阴性对照组的细胞生长未受到明显抑制(P＞0.05).实验组、阴性对照组和空白对照组各基因mRNA的相对表达量:c-myc基因分别为1.047±0.156、1.737±0.193和1.631±0.128;c-fos基因分别为0.041+0.003、0.082±0.006和0.081±0.004; N-ras基因分别为0.082±0.009、0.156±0.013和0.133±0.015;ki-67基因分别为0.032±0.002、0.070±0.008和0.069±0.005; caspasc-3基因分别为0.148±0.018、0.110±0.009和0.108+0.012;bax基因分别为0.780±0.092、0.629±0.058和0.617±0.073,各基因的mRNA在不同组别表达差异均有统计学意义(F值分别为104.384、400.915、211.903、427.041、67.750和37.272,P值均＜0.01).结论 AKR1B10可能通过调节肿瘤相关基因的表达水平来促进细胞增殖、抑制细胞凋亡.     

干扰固醇调节元件结合蛋白-1c表达对细胞炎症趋化因子配体2和成纤维细胞生长因子21的影响

目的 研究L02肝细胞脂肪变模型中内质网应激状态下,干扰固醇调节元件结合蛋白(SREBP) -1c表达后肝细胞脂质代谢及肝细胞系统性炎症的变化情况及其意义。 方法 用油酸诱导L02肝细胞脂肪变,选用pSilencer 1.0-U6-4476质粒干扰SREBP-1c的表达,分为空白对照组、空质粒转染模型组及干扰质粒转染模型组。分别于0、24、48、72 h检测细胞炎症趋化因子配体(CCL)2和成纤维细胞生长因子(FGF) 21的mRNA和蛋白质的表达。组间比较用t检验,多个组间正态资料的比较用单因素方差分析。 结果 干扰SREBP-1c后,干扰质粒转染模型组较空质粒转染模型组细胞内脂肪滴数量明显减少,提示肝细胞脂肪变显著减轻。干扰质粒转染模型组CCL2基因在0、24、48、72h mRNA相对表达量分别为1.03±0.11、1.11±0.21、0.88±0.16、1.05±0.15,与空白对照组比较,P值均＞0.05,差异均无统计学意义。组内各时相点比较,P值均＞0.05,差异均无统计学意义。而对应的蛋白质表达量0、24、48、72h分别为1.19±0.15、1.07±0.18、0.48±0.14、0.05±0.24,低于空白对照组的4.15±0.18、4.09±0.21、3.91±0.13、4.06±0.12,两组比较,t值分别为2.78、3.67、4.10、4.89,P值均＜0.01,差异均有统计学意义。干扰质粒转染模型组组内各时相点比较,t值分别为1.11、2.25、3.17、1.01、3.27、3.51,P值均＜0.05,差异均有统计学意义。干扰质粒转染模型组FGF21基因mRNA相对表达量在0、24、48、72 h分别为1.01±0.08、0.91±0.22、0.98±0.20、1.02±0.12,与空白对照组比较,P值均＞0.05,差异均无统计学意义。干扰质粒转染模型组组内各时相点之间比较,t值分别为0.28、0.28、0.17、0.19、0.24、0.16,P值均＞0.05,差异均无统计学意义。而对应的蛋白质表达量下调趋势非常明显,0、24、48、72 h分别为0.81±0.05、0.66±0.12、0.58±0.08、0.19±0.13,与空白对照组的3.95±0.07、4.01±0.14、3.89±0.19、4.08±0.15比较,t值分别为3.21、3.78、4.54、5.17,P值均＜0.01,差异均有统计学意义。干扰质粒转染模型组组内各时相点比较,t值分别为1.08、1.76、3.54、1.24、4.21、3.79,P值均＜0.05,差异均有统计学意义。结论 SREBP-1c下调表达后能减轻内质网应激的后续效应,改善肝细胞脂质代谢,还可能通过CCL2通路减轻肝细胞内质网应激所致的炎症效应。     

The immunoneuroendocrine role of melatonin

Abstract: A tight, physiological link between the pineal gland and the immune system is emerging from a series of experimental studies. This link might reflect the evolutionary connection between self-recognition and reproduction. Pinealectomy or other experimental methods which inhibit melatonin synthesis and secretion induce a state of immunodepression which is counteracted by melatonin. In general, melatonin seems to have an immunoenhancing effect that is particularly apparent in immunodepressive states. The negative effect of acute stress or immunosuppressive pharmacological treatments on various immune parameters are counteracted by melatonin. It seems important to note that one of the main targets of melatonin is the thymus, i.e., the central organ of the immune system. The clinical use of melatonin as an immunotherapeutic agent seems promising in primary and secondary immunodeficiencies as well as in cancer immunotherapy. The immunoenhancing action of melatonin seems to be mediated by T-helper cell-derived opioid peptides as well as by lymphokines and, perhaps, by pituitary hormones. Melatonin-induced-immuno-opioids (MHO) and lymphokines imply the presence of specific binding sites or melatonin receptors on cells of the immune system. On the other hand, lymphokines such as -γ-interferon and interleukin-2 as well as thymic hormones can modulate the synthesis of melatonin in the pineal gland. The pineal gland might thus be viewed as the crux of a sophisticated immunoneuroendocrine network which functions as an unconscious, diffuse sensory organ.     

Response of rat pineal melatonin to calcium, magnesium, and lithium is circadian stage dependent

Abstract: Herein we documented the response of pineal melatonin production to electrolytes known to be effective on pineal function in view of a possible circadian stage dependence. We studied the release of melatonin by perifused rat pineal glands at 2 different circadian stages corresponding to the middle of the light and dark periods, i.e., respectively, 7 and 19 HALO (Hours After Light Onset, L:D = 12:12). The initial efflux rates were, as expected, much higher in the perifusates of glands removed from rats sacrificed during the dark phase than of those removed during the light phase. After 3 hr of perifusion, melatonin release reached similar levels which were found constant up to the 8th hr of perifusion, whatever the circadian stage. Perifusion of the glands with physiological concentrations for the rat of calcium (5.2 mmol/1) and magnesium (1.34 mmol/1) resulted in a stimulatory effect on the pineal glands removed from rats sacrificed in the middle of the dark period (19 HALO), whereas no effects were observed on the pineal glands removed from rats sacrificed during the light (7 HALO). Lithium (0.28 and 0.55 mmol/1) was ineffective on melatonin release in pineal glands removed 7 and 19 HALO. Our results show differences in the initial efflux rates of melatonin and in the response of perifused pineal glands to calcium and magnesium according to the circadian stage.     

Changes in connexin43, the gap junction protein of astrocytes, during development of the rat pineal gland

Abstract: The abundance of gap junctions between rat pineal astrocytes formed by connexin43 (Cx43) was studied during development. Levels and distribution of Cx43 were measured by immunoblotting and indirect immunofluorescence, respectively. The amount of Cx43 in cells located within the gland was low until about the 7th postnatal day and increased to adult values between the 14th and 21st days postpartum. Although astrocytes, recognized by their vimentin immunoreactivity, were scarce before birth, they were abundant by the 7th postnatal day suggesting that the low levels of Cx43 found at this age corresponded to a low expression of this protein. Localization of the immunoreactivity to Cx43 and vimentin showed a close correlation, indicating that mature or immature pineal astrocytes form gap junctions made of Cx43. Since Cx43 levels attained their adult values at about the time the innervation and the functional state of the gland reached maturity (2–3 weeks after birth), it is proposed that astrocyte gap junctions are involved in the function of the adult rat pineal gland.     

Conservative management of perforated duodenal diverticulum： A case report and review of the literature

Duodenal diverticula are a relatively common condition. They are asymptomatic, unless they become complicated, with perforation being the rarest but most severe complication. Surgical treatment is the most frequently performed approach. We report the case of a patient with a perforated duodenal diverticulum, which was diagnosed early and treated conservatively with antibiotics and percutaneous drainage of secondary retroperitoneal abscesses. We suggest this method could be an acceptable option for the management of similar cases, provided that the patient is in good general condition and without septic signs.     

Presence of immunoreactive growth hormone and prolactin in the ovine pineal gland

Abstract: The use of antisera raised against bovine growth hormone (GH) and ovine prolactin (PRL) enabled the detection of related immunoreactive (ir) sequences of proteins in ovine pineal tissue. The isolation of PRL-like ir-material was accomplished using a 0.25 M ammonium sulphate (pH 5.5) extraction followed by ethanol precipitation, whereas the resulting 2.0 M ammonium sulphate (pH 7.0) precipitate contained a GH-like immunoreactivity. Gel chromatography of the GH-like immunoreactivity (Sephadex G-100) indicated the presence of several GH-like fragments ranging in the M_r range of 7,000 to 55,000. Analyses of the PRL-like ir-material found in pineal tissue on HPLC using a TSK 545-DEAE column led to the resolution into a single peak of immunoreactivity. A single peak of activity was also observed following chromatofocusing and hydrophobic interaction chromatography of the ir-peak from the TSK 545-DEAE column. The PRL-like ir-material inhibited the binding of [¹²⁵I]ovine PRL-S14 to anti-ovine PRL antibodies without showing an affinity for binding to anti-rat PRL or anti-bovine GH antibodies. Scatchard analysis of the binding of pineal PRL-like ir-material and pituitary ovine PRL-S14 to liver membranes from day-20 pregnant rats revealed similar affinity constants (K_a of 4.7 ± 0.2 × 10⁹ M^-1). In addition, the replication of Nb 2 Node rat lymphoma cells was stimulated by pineal PRL-like ir-material, an effect known to be specific for lactogenic hormones. The pineal PRL-like immunoreactivity appeared on sodium dodecyl sulfate polyacrylamide gels as a single major band of M_r 24,000. The functional status of PRL-and GH-like ir-material in the ovine pineal remains to be determined, but evidence is presented that the overall protein synthesis rate of the rat pineal responded to circulating concentrations of PRL.     

Microbiology of human immunodeficiency virus anorectal disease

PURPOSE: Individuals who are seropositive for the human immunodeficiency virus are at high risk for opportunistic infection and anorectal disorders. Little prospective information is available regarding anorectal pathogens in these patients. METHODS: One hundred sixty-three HIV-seropositive patients presented to the colorectal clinic between 1989 and 1992. Forty-seven (29 percent) patients were thought to have an infectious process and were prospectively studied using a standardized multiculture protocol. RESULTS: Mean age was 33 (range, 19–59) years. All were male; high-risk behavior accounted for 87 percent of HIV transmissions. Presenting complaints included anorectal pain (79 percent), pus per anum (28 percent), and blood per anum (26 percent). Examination revealed perianal tenderness (60 percent), condyloma (38 percent), perianal ulcers (38 percent), and anal fissures (34 percent). Sixty-six sets of cultures were performed; 28 patients had one set, 15 had two sets, and 4 had three sets. Thirty-two of these 47 patients (68 percent) had positive cultures including herpes (50 percent), cytomegalovirus (25 percent),Neisseria gonorrhoeae (16 percent), chlamydia (16 percent), acidfast bacilli (2 percent), and others (9 percent). Six of 32 patients with positive cultures had more than one organism cultured. Sixteen (50 percent) patients with positive cultures were treated medically, 8 (25 percent) were treated surgically and 8 (25 percent) were treated with both modalities. Sixty-one procedures were performed on 17 patients for condylomata. Eighteen patients had 20 procedures for abscesses, 50 percent of whom had positive cultures for other than common bowel flora; all improved. Fourteen patients underwent 33 procedures for perianal fistulas.Mycobacterium fortuitum was cultured from one patient who required 13 procedures for abscesses and fistulas. Forty-five (96 percent) patients were followed for an average of 12.5 months ±2.9 SEM (range, 1–94 months). Symptoms were improved or resolved in 22 of 32 (69 percent) patients with positive cultures and in 11 of 13 (84 percent) with negative cultures. CONCLUSIONS: Specific pathogens may often be identified in human immunodeficiency virus-seropositive patients with anorectal disorders if aggressively sought. Although patients without specific pathogens identified may be expected to improve with planned empiric treatment, positive identification allows more directed therapy.