干扰素联合病毒唑对肝炎患者正,负链丙型肝炎病毒RNA的影响

为评价干扰素联合病毒唑的抗病毒效果，用套式反转录－聚合酶链反应技术观察其对慢性丙型肝炎患者正、负链丙型肝炎病毒ＲＮＡ（ＨＣＶＲＮＡ）的作用。结果显示，在对联合抗病毒治疗有反应的患者中，多数患者ＡＬＴ降至正常，血清正链阳性率由９２．３１％降至３８．４６％（Ｐ＜０．００５），周围血单核细胞（ＰＢＭＣ）中负链阳性率由７６．９２％降至３８．４６％（Ｐ＜０．０５），但对ＰＢＭＣ中正链ＨＣＶＲＮＡ影响不大。治疗停止后，半数以上患者病情复发，病情复发者均为ＰＢＭＣ中正链及负链ＨＣＶＲＮＡ持续阳性者。提示单纯检测血清正链ＨＣＶＲＮＡ作为疗效考核及病情转归的判断指标有一定的局限性。同时检测血清及ＰＢＭＣ中正、负链ＨＣＶＲＮＡ，具有重要的临床意义。本研究干扰素联合病毒唑的抗病毒效果与单用干扰素接近，似说明病毒唑并未增加干扰素的疗效     

周围血白细胞的复制型丙型肝炎病毒RNA的检测及临床意义

用简并引物作套式反转录。聚合酶链反应检测正、负链丙型肝炎病毒（ＨＣＶ）ＲＮＡ。显示３０例急、慢性丙型肝炎患者的血清及血浆中，７例无症状抗－ＨＣＶ阳性者的血清、血浆及周围血白细胞（ＰＢＬ／Ｃ）中，均未检出负链ＨＣＶＲＮＡ。慢性丙型肝炎者ＰＢＩ（中正、负链ＨＣＶＲＮＡ的检出率高于急性丙型肝炎及无症状抗－ＨＣＶ阳性者（Ｐ＜０．０５～０．００１）。１７例经肝组织学检查的患者中，急性肝炎（ＡＨ）者ＰＢＬ／Ｃ的正、负链ＨＣＶＲＮＡ检出率低于慢性活动性肝炎（ＣＡＨ）者（Ｐ＜０．０５）。１例ＡＨ及６例ＣＡＨ患者肝组织内正、负链ＨＣＶＲＮＡ全部阳性。证实丙型肝炎患者的ＰＢＬ／Ｃ确可被ＨＣＶ感染，病程越长，被ＨＣＶ感染的可能性越大；病情活动者，ＰＢＬ／Ｃ中负链ＨＣＶＲＮＡ的检出率越高。提示ＨＣＶ不仅可以感染ＰＢＬＣ，而且可在其中复制；负链ＨＣＶＲＮＡ的出现与病情活动有关。     

丙型肝炎患者血清和外周血单个核细胞中HCV RNA检测及意义

目的 了解丙肝患者血清和外周血单个核细胞（ＰＢＭＣ）中ＨＣＶＲＮＡ存在情况及有其临床意义，方法 应用套式ＰＣＲ检测４６例急性丙型肝炎（丙型肝炎以下简称丙肝）和４２例慢性丙型肝炎患者血清和ＰＢＭＣ中ＨＣＶＲＮＡ。结果 慢性丙型肝炎患者ＰＢＭＣ中ＨＣＶＲＮＡ检出率显著高于急性丙型患者（Ｐ〈０．００１），急，慢性丙型肝患者血清和慢性丙肝患者ＰＢＭＣ中ＨＣＶ ＲＮＡ检出率显著高于ＡＬＴ正常的抗－ＨＣＶ阳性     

原位杂交法检测外周血单个核细胞中HCV RNA

目的比较慢性丙型肝炎患者用干扰素治疗前以及治疗后３个月ＰＢＭＣ中ＨＣＶＲＮＡ。方法应用地高辛素标记ＨＣＶＲＮＡ正链及负链探针，建立原位杂交方法检测外周血单个核细胞（ＰＭＢＣ）中的ＨＣＶＲＮＡ。结果治疗前１９例患者正链ＨＣＶＲＮＡ阳性，８例负链ＨＣＶＲＮＡ阳性，用正链探针杂交在较多的细胞中出现杂交信号，负链探针杂交仅在少数细胞中出现杂交信号，ＨＣＶＲＮＡ在ＰＢＭＣ胞浆中呈均质性分布。用干扰素治疗结束后３个月２０例患者中９例ＨＣＶＲＮＡ转阴性，近期治愈率４５％。结论原位杂交技术的敏感性及特异性较高，且重复性较好，是研究ＨＣＶＲＮＡ在组织中定位分布和病毒复制场所一种切实可行的方法     

慢性丙型肝炎患者外周血单个核细胞中HCV RNA和NS5抗原的检测

目的 研究丙型肝炎病毒（ＨＣＶ）感染外周血个核细胞（ＰＢＭＣ）的情况及其意义。方法 运用非核素原位杂交法（ＮＩＳＨ）和抗生蛋白链菌素－生物素法（ＳＡＢＣ）分别检测２０例慢性丙型肝炎患者ＰＢＭＣ中的ＨＣＶ ＲＮＡ和ＮＳ５抗原。结果 ２０例患者中有８例（４０％）,ＰＢＭＣ中ＨＣＶ ＲＮＡ呈阳性,其中６例（３０％）ＰＢＭＣ中ＮＳ５抗原同时呈阳性。ＨＣＶＲＮＡ主要分布于胞浆中,而ＮＳ５抗原还可以出现在胞膜     

丙型病毒性肝炎患者外周血单核细胞中HCV—RNA的存在及其意义

作者用套式ＰＣＲ检测２５例丙型病毒性肝炎（丙肝）患者血清和外周血单核细胞（ＰＢＭＣ）中ＨＣＶ－ＲＮＡ，结果显示血清阳性２１例，ＰＢＭＣ阳性１８例，表现在大部分丙肝患者ＰＢＭＣ中有ＨＣＶ－ＲＮＡ的存在。且急性丙肝患者ＰＢＭＣ中ＨＣＶ－ＲＮＡ阳性率明显低于慢性丙肝患者，提示ＰＢＭＣ中ＨＣＶ－ＲＮＡ的存在可能与感染ＨＣＶ后的慢性化有关。     

慢性丙型肝炎患者外周血单个核细胞丙型肝炎病毒检测及其意义

目的 探讨外周血单个核细胞（ＰＢＭＣｓ）在丙型肝炎病毒（ＨＣＶ）的感染中的作用。方法 对２２例慢性丙型肝炎患者２１例抗－ＨＣＶ（＋）血液管析患者及１２例健康献血员的ＰＢＭＣｓ分别进行ＨＣＶＲＮＡ,ＨＣＶ抗原检测及电镜观察。结果 （１）２２生丙型肝炎肝炎患者ＰＢＭＣｓ中有７７．３％（１７／２２）ＨＣＶＲＮＡ阳性,（２）感染ＨＣＶ的ＰＢＭＣｓ中电镜下发现复制的ＨＣＶ颗粒;（３）ＨＣＶ颗粒阳笥者的血清和     

丙型肝炎患者血清中高变区1(HVR1)抗体的检测与分析

目的　了解丙型肝炎患者体内抗ＨＶＲ１ 的产生情况,并探讨其与病情发展及预后的关系．方法　用含ＨＶＲ１ 序列的合成肽对４５ 例急、慢性肝炎患者血清中抗ＨＶＲ１ 作了ＥＬＩＳＡ 检测,并对所有血清作ＨＣＶＲＮＡ 检测．结果　急性丙型肝炎患者血清中抗ＨＶＲ１ 阳性率为７０－６ ％ ,与慢性丙型肝炎患者早期血清（１２－５ ％ ） 相比差异显著性（ Ｐ＜ ０－０５） ,而且两者的平均抗体滴度相差亦非常显著（４－２ ±０－６ ｖｓ ２－８ ±０－７ ,Ｐ＜ ０－０１） ． 慢性丙型肝炎现症患者血清抗ＨＶＲ１ 的阳性率为７２－７ ％ ,其中抗ＨＶＲ１ 阳性血清中ＨＣＶＲＮＡ 的阳性率也很高（７５－０ ％ ） ．结论　ＨＣＶ 感染早期抗ＨＶＲ１ 的产生及其滴度的高低可能与疾病的转归有关;慢性丙型肝炎患者体内抗ＨＶＲ１ 的存在可能反映了ＨＶＲ１ 变异引起的免疫逃避．     

原位反转录PCR检测慢性丙型肝炎患者外周血单个核细胞中HCV

目的研究慢性丙型肝炎患者外周血单个核细胞（ｐｅｒｉｐｈｅｒａｌｂｌｏｏｄｍｏｎｏｎｕｃｌｅａｒｃｅｌ，ＰＢＭＣ）中丙型肝炎病毒的定位分布及复制。方法直接原位反转录－ＰＣＲ（标记引物）对２０例慢性丙型肝炎患者的ＰＢＭＣ内丙型肝炎病毒（ＨＣＶ）的分布、定位及存在形式进行研究。结果１６例患者外周血单个核细胞的胞浆内发现ＨＣＶＲＮＡ阳性信号，并有一例患者的胞浆内发现ＨＣＶＲＮＡ负链信号。结论ＨＣＶ可感染ＰＢＭＣ并在其中复制。原位ＲＴ－ＰＣＲ为进一步研究ＨＣＶ进入人体内的分布、潜伏、复制状况以及丙型肝炎治疗、药物研究奠定了基础。     

用支链DNA测定慢性丙型肝炎患者血清HCV RNA浓度来预测干扰素疗法的疗效

用支链ＤＮＡ测定慢性丙型肝炎患者血清ＨＣＶＲＮＡ浓度来预测干扰素疗法的疗效［英］／ＯｓｈｉｔａＭ…Ｇａｓｔｒｏｅｎｔｅｒｏｌｏｇｙ．－１９９５，１０８（４）．－Ａ１１３９已证实干扰素（ＩＦＮ）疗法对慢性丙型肝炎是有效的，而且治疗前血清ＨＣＶＲＮＡ浓度...     

HCV RNA在外周血单个核细胞中的复制与丙肝慢性化及复发的关系

通过对外周血单个核细胞（PBMC）中正链及负链HCV RNA的检测来探讨HCV在PBMC中的复制与丙肝慢性化及复发的关系。应用巢式RT—PCR对71例慢性丙型肝炎患者的血清和PBMC进行正负链HCV RNA的检测,其中15例干扰素治疗。并对10例正常人的血清和PBMC进行了HCV RNA检测。71例慢性丙型肝炎患者血清中正链HCV RNA阳性率73．2％,负链HCV RNA阳性率为0,PBMC中正链HCV RNA阳性率为60．5％,负链HCV RNA阳性率为38．0％。15例干扰素治疗患者在治疗后血清正链HCV RNA转阴率为73．3％,PBMC中正链HCV RNA转阴率为45．5％,负链转阴率为71．4％。正常对照组血清及PBMC中HCV RNA阳性率为0。这说明HCV RNA在PBMC中的复制与丙肝慢性化及复发存在一定的关系。干扰素治疗丙肝具有一定疗效,但不能彻底清除病毒。     

外周血单个核细胞内HCV RNA是慢性丙型肝炎干扰素治疗效果的预测因子

目的 探讨慢性丙型肝炎干扰素(IFN)治疗结束时，外周血单个核细胞(PBMC)中HCV RNA检测对持续性应答的预测作用。方法 对因输血而感染的慢性HCV肝炎患者进行IFN治疗24周，在治疗12周即取得完全应答的患者，24周治疗结束时检测PBMC中HCV RNA，并对患者进行长期的血清HCV RNA监测。结果 治疗结束时，有9例患者的PBMC HCV RNA为阳性，7例为阴性。在PBMC中HCV RNA阳性患者，停止治疗的6个月内9例中8例血清HCV RNA阳转，1年内所有患者的HCV RNA复发。而PBMC HCV RNA阴性者，随访至停药6月时，7例中仅1例HCV RNA阳转，1年内共2例HCV RNA复发，其余5例，随访至3．5年，血清中HCV RNA仍阴性。结论 IFN治疗结束时PBMC中HCV RNA检测能预测慢性丙型肝炎的治疗效果。     

Replication of hepatitis C virus in peripheral blood mononuclear cells. Effect of alpha-interferon therapy.

Hepatitis C virus (HCV) is a positive-stranded RNA virus which replicates through a negative-stranded RNA intermediate. Using a PCR procedure to detect positive and negative strands, we investigated the existence of HCV replication in lymphoid cells. Both positive and negative strands were found in the peripheral blood mononuclear cells (PBMC) of all patients (n = 10) with untreated chronic hepatitis C. No HCV sequences were detected in PBMC in any of the 8 healthy controls. Fifteen patients with chronic hepatitis C were studied at the end of a 12-month course of alpha-interferon therapy. The positive strand was detected in PBMC in all 9 non-responder patients, and the negative strand in 7. In contrast, in PBMC from responder patients (n = 6) the positive strand was found in 4 and the negative strand in only 2 cases. These results demonstrate that HCV can infect PBMC and replicate in these cells and that interferon seems to exert an inhibitory effect on this process. Persistence of HCV-RNA in PBMC may help explain disease relapse after successful interferon therapy.     

Testing for hepatitis C virus sequences in peripheral blood mononuclear cells of patients with chronic hepatitis C in the absence of serum hepatitis C virus RNA

Hepatitis C virus (HCV) is able to replicate in peripheral blood mononuclear cells (PBMC) of HCV-infected patients. Few data are available on PBMC testing for HCV RNA in serum HCV RNA negative patients, positive for anti-HCV and with histological evidence of chronic hepatitis. Twenty such patients were studied; of these, 11 were tested during interferon α (IFN) treatment, at the time of serum HCV RNA clearance and ALT normalisation: only one was found to be positive for HCV sequences in PBMC. Within 3 months of IFN withdrawal all 11 patients relapsed with high ALT and recurrence of serum HCV RNA. Of nine serum HCV RNA negative patients with chronic hepatitis C who were not receiving IFN when tested (four untreated patients and five patients who had already completed IFN schedule), PBMC HCV RNA was detected in four. Evidence of active HCV replication (presence of the minus strand genome) in PBMC was also observed in two cases. Thus, five of the 20 patients without detectable serum HCV RNA turned out to be carriers of HCV sequences in PBMC. These data indicate that: 1. PBMC are an extrahepatic replication site of HCV; this is true also in the absence of serum HCV RNA; 2. the role of PBMC as a “viral reservoir” after IFN-induced serum HCV RNA clearance is questioned; 3. the absence of both serum and PBMC HCV RNA in patients under IFN is not predictive of sustained viral loss; 4. testing for PBMC viral sequences might enhance the chances of detecting HCV infection.     

Effect of TT virus co‐infection on interferon response in chronic hepatitis C patients

Abstract: Aim: TT virus (TTV) is a single stranded DNA virus found in serum of patients with post‐transfusion non‐A to ‐G hepatitis. TTV‐DNA has been investigated in sera of patients with various liver diseases. This study aimed at finding whether co‐infection with TTV in HCV patients, may influence the effect of interferon (IFN) in complete elimination of HCV, and analysed the correlation between HCV and TTV by semi‐quantification of both HCV RNAs and TTV DNA. Methods: In 28 chronic hepatitis C (CH‐C) patients with TTV co‐infection, the presence of TTV DNA was checked in sera six months before and after the end of IFN therapy. Result: Five out of 28 patients became negative for both HCV‐RNA and TTV‐DNA following IFN therapy. But 10 out of 28 patients persistently remained positive for both. Among the remaining 13 patients, 5 tested negative for HCV‐RNA but positive for TTV‐DNA. Post IFN therapy changes in serum alanine aminotransferase (ALT) levels did not appear to be influenced by the presence of TTV co‐infection. HCV‐RNA was found to be the most important predictor of IFN response in CH‐C patients with TTV co‐infection. TTV DNA level in sera had no correlation with IFN response. In addition, there was no relationship between HCV RNA and TTV DNA. Conclusion: Based on these results, it can be concluded that the effectiveness of IFN in eliminating HCV does not seem to be influenced by co‐infection.     

HCV replication in PBMC and its influence on interferon therapy

AIM: To study hepatic virus C (HCV) RNA and HCV protein expression in peripheral blood mononuclear cells (PBMCs) of patients with HCV infection, and explore the relationship between the HCV RNA in the PBMCs and response to interferon (IFN) therapy. METHODS: Type-specific primers were designed and RT-nested PCR was used to detect the plus- and minus- strands of HCV RNA in PBMCs of 54 patients with HCV infection; Indirect immunofluorescence assay was applied to identify HCVNS5 protein expression in PBMCs; 6 month-, 3 MU-IFN regiment was administrated to observe the responses to IFN in 35 chronic hepatitis C patients with different HCV RNA status in PBMCs. RESULTS: HCV plus strand RNA was found in 10 of 19 (52.6 %) acute hepatitis C patients and 22 of 35 (62.9 %) chronic hepatitis C patients. HCV minus strand RNA was detected in 14 of 35 (40.0 %) chronic hepatitis C patients, but only one patient (5.3 %) with acute HCV infection was found to be minus HCV RNA positive. Though no HCV NS5 protein expression was found in the examined 10 cases of acute HCV infection, it was positive in 17 of 20 (85.0 %) chronic hepatitis C patients by indirect immunofluorescence assay. There are significant differences of positive rate of the minus-strand and HCVNS5 protein between acute and chronic hepatitis C groups (u=2.07, P<0.05 and u=4.43, P<0.01 respectively). The patients with minus-strand HCV RNA showed a significantly lower 6-month sustained response (SR-6) to IFN compared to those without minus-strand HCVRNA in PBMCs (biologically 14.3 % vs 42.8 %, chi(2)=4.12, P<0.05 and virologically 7.1 % vs 23.9 %, chi(2)=4.24, P<0.05). CONCLUSION: HCV is capable of infecting and replicating in PBMCs, and HCVNS5 protein was expressed in PBMCs. The patients with minus strand HCV RNA in PBMCs showed a significantly lower 6-month sustained response to IFN, suggesting that minus-strand HCV RNA in PBMCs may be one of the factors influencing response to IFN therapy.     

Effect of TT virus co-infection on interferon response in chronic hepatitis C patients

AIM: TT virus (TTV) is a single stranded DNA virus found in serum of patients with post-transfusion non-A to -G hepatitis. TTV-DNA has been investigated in sera of patients with various liver diseases. This study aimed at finding whether co-infection with TTV in HCV patients, may influence the effect of interferon (IFN) in complete elimination of HCV, and analysed the correlation between HCV and TTV by semi-quantification of both HCV RNAs and TTV DNA. METHODS: In 28 chronic hepatitis C (CH-C) patients with TTV co-infection, the presence of TTV DNA was checked in sera six months before and after the end of IFN therapy. RESULT: Five out of 28 patients became negative for both HCV-RNA and TTV-DNA following IFN therapy. But 10 out of 28 patients persistently remained positive for both. Among the remaining 13 patients, 5 tested negative for HCV-RNA but positive for TTV-DNA. Post IFN therapy changes in serum alanine aminotransferase (ALT) levels did not appear to be influenced by the presence of TTV co-infection. HCV-RNA was found to be the most important predictor of IFN response in CH-C patients with TTV co-infection. TTV DNA level in sera had no correlation with IFN response. In addition, there was no relationship between HCV RNA and TTV DNA. CONCLUSION: Based on these results, it can be concluded that the effectiveness of IFN in eliminating HCV does not seem to be influenced by co-infection.