Comparison between HIV- and CMV-specific T cell responses in long-term HIV infected donors

The mechanisms underlying non-progression in HIV-1 infection are not well understood; however, this state has been associated previously with strong HIV-1-specific CD8+ T cell responses and the preservation of proliferative CD4+ T cell responses to HIV-1 antigens. Using a combination of interferon-gamma (IFN-gamma) ELISpot assays and tetramer staining, the HIV-1-specific CD8+ T cell populations were quantified and characterized in untreated long-term HIV-1-infected non-progressors and individuals with slowly progressive disease, both in relation to CD4+ T cell responses, and in comparison with responses to cytomegalovirus (CMV) antigens. High levels of CD8+ T cell responses specific for HIV-1 or CMV were observed, but neither their frequency nor their phenotype seemed to differ between the two patient groups. Moreover, while CMV-specific CD4+ T cell responses were preserved in these donors, IFN-gamma release by HIV-1-specific CD4+ T cells was generally low. These data raise questions with regard to the role played by CD8+ T cells in the establishment and maintenance of long-term non-progression.     

Apoptosis of T lymphocytes isolated from peripheral blood of patients with acute exacerbation of chronic obstructive pulmonary disease

Purpose

Chronic obstructive pulmonary disease (COPD) is characterized by chronic inflammation of the airways and progressive destruction of lung parenchyma. Apoptosis is critical for the maintenance of normal tissue homeostasis and is in equilibrium with proliferation and differentiation. This study was undertaken to investigate relationship between apoptosis of peripheral blood lymphocytes during exacerbation of COPD and inflammatory response that characterizes this condition.

Materials and Methods

Seventeen patients with COPD exacerbation, 21 stable COPD, and 12 control subjects were included. T lymphocytes were isolated from peripheral blood using MACS. Apoptosis of T lymphocytes was assessed with FACS using annexin V and 7-aminoactinomycin. Serum levels of interleukin (IL)-6, IL-8 and tumor necrosis factor (TNF)-α were determined by an immunoassay technique.

Results

There was significantly increased percentage of apoptotic lymphocytes, CD 4⁺, and CD 8⁺ T cells in the peripheral blood of patients with exacerbation of COPD compared with stable COPD. Serum levels of IL-6, IL-8, and TNF-α were significantly increased in patients with exacerbation of COPD compared with stable COPD. Only TNF-α presented a positive correlation with apoptotic lymphocytes in patients with exacerbation of COPD.

Conclusion

Increased apoptotic lymphocytes may be associated with upregulation of TNF-α in the peripheral blood of patients with acute exacerbation of COPD.     

Progressive loss of IL-2-expandable HIV-1-specific cytotoxic T lymphocytes during asymptomatic HIV infection

In HIV-1 infection, circulating HIV-1-specific cytotoxic T lymphocytes (CTL) exist in different states of activation, including activated cytotoxic cells and memory cells. We report that a subpopulation of HIV-1-specific CTL is capable of clonal expansion upon culture with IL-2 without exogenous antigen. The IL-2-expandable HIV-1-specific CTL precursor frequency was reduced in patients with advancing infection, although HIV-1-specific memory CTL could still be detected by stimulation in vitro with allele-specific HIV-1 peptide. Longitudinal analysis during advancing infection showed a progressive decline in the IL-2-expandable HIV-1-specific CTL precursor (CTLp) frequency without a decline in Epstein-Barr virus (EBV)-specific or allo-specific CTLp frequencies. To address mechanisms that may contribute to the decline in the IL-2-expandable HIV-specific CTL response, the requirements for in vitro generation of HIV-1-specific and EBV-specific effector CTL were examined. In the absence of exogenous IL-2 in limiting dilution, generation of EBV-specific CD8⁺ effector CTL was dependent upon help from CD4⁺ cells. CD4⁺ help for EBV-specific CD8⁺ CTL was observed in asymptomatic HIV infection but not in advanced infection. In the presence of exogenous IL-2, CD4⁺ cells could also provide help for the optimal generation of HIV-1 peptide-specific CD8⁺ CTL, because in vitro depletion of CD4⁺ cells prior to culture using stimulation with an MHC class I-restricted HIV-1 peptide reduced the peptide-specific CD8⁺ CTL response. We conclude that there is a decline in the IL-2-expandable HIV-1-specific CTL response during advancing infection. There are a number of possible mechanisms for this decline, including a reduction in CD4⁺ T cell help for in vivo antigen-activated CD8⁺ T cells.     

IL-18对系统性红斑狼疮患者淋巴细胞凋亡和P53蛋白表达的影响

目的　研究系统性红斑狼疮 (SLE)患者外周血淋巴细胞在体外IL 1 8刺激培养下细胞凋亡及P53蛋白表达情况。方法　AnnexinV联合PI染色定量法及免疫荧光染色法 ,分析了 44例SLE患者和 30例正常人外周淋巴细胞在体外IL 1 8刺激培养后凋亡发生率 ,凋亡相关基因P53蛋白的表达以及淋巴细胞凋亡发生与疾病活动性的相关性。结果　在IL 1 8刺激培养作用下 ,活动期SLE淋巴细胞凋亡发生率较正常人显著增高 (P <0 .0 1 ) ,而静止期则无明显变化 (P >0 .0 5)。P53蛋白表达在活动期SLE较正常人显著性下降 (P <0 .0 1 ) ,静止期无明显变化 (P >0 .0 5)。P53的表达与疾病活动指数SLEDAI之间有明显的相关性 (P <0 .0 1 )。结论　IL 1 8可引起SLE患者PBL凋亡率的增高 ,表明IL 1 8在体内凋亡或凋亡相关性免疫机制中起着一定的作用     

经血感染HIV/AIDS患者pDCs与艾滋病疾病进展关系的研究

目的 了解中国HIV/AIDS患者外周血中类浆细胞样树突状细胞（plasmacytoid dendritic cells，pDC）绝对值的变化，分析其与HIV感染及疾病进展的关系。方法采集56例未经治疗的H1VIMDS患者及34例健康人抗凝静脉血，采用流式细胞仪单平台检测技术，分析计算三色荧光抗体标记的全血中pDC（Lin1^＋／HLA-DR^＋／CD123^＋）绝对值。结果MDS组pDC绝对值为3．31个／μl（0．31～6．46个/μl），显著低于正常对照组、HIV慢性感染组及HIV感染疾病长期不进展（long term non-progressors，LTNP）组（P〈0．01）。HIV慢性感染组pDC绝对值为5．27个/μl（2．25～9．80个/μl），显著低于正常对照组及LTNP组（P〈0．01）。LTNP组pDC绝对值为9．95个／μl（6．16～20．88个／μl），显著高于正常对照组（7．84个/μl，3．45～13．97个／μl，P〈0．01）。56例HIV/AIDS患者pDC绝对值与CD4^＋T淋巴细胞绝对值及百分率呈显著正相关（r=0．422，P〈0．01；r=0．488，P〈0．01），与病毒载量呈显著负相关（r=-0．444，P〈0．05）。结论HIV／AIDS患者pDC数量减少，并且随疾病进展逐渐下降；HIV感染LTNP的血pDC数量显著高于HIV慢悱感染者及MDS患者，甚至高于正常对照。提示外周血pDC在控制HIV感染方面有重要作用，与艾滋病疾病进展明显相关。     

类风湿关节炎患者外周血淋巴细胞的凋亡及其意义

目的研究类风湿关节炎(RA)患者外周血淋巴细胞的凋亡及其意义。方法采用流式细胞术和激光共聚焦显微镜,检测35例RA患者和30例健康志愿者的外周血淋巴细胞抵抗凋亡的能力,包括分析新鲜分离和培养的淋巴细胞凋亡率及其与临床病情严重度的相关性。结果两组间新鲜分离的淋巴细胞凋亡率差异无统计学意义[(2.63±1.47)%比(3.53±2.04)%,P>0.05]。活化淋巴细胞检测的结果显示,RA组的凋亡率明显低于对照组[(12.08±6.06)%比(18.66±7.27)%,P<0.05],并与DAS28评分呈负相关(r=-0.617,P=0.008)。结论RA患者活化的外周血淋巴细胞出现凋亡缺陷,并与RA病情活动度相关。提示这些细胞募集到组织损害和自身免疫炎性反应进程异常的部位并对炎性反应进程的发展和维持起重要作用。     

Immunophenotypic analysis of peripheral blood leukocytes at different stages of HIV infection. An analysis of asymptomatic, ARC, and AIDS populations

Blood leukocytes from 51 patients with acquired immune deficiency syndrome (AIDS) or AIDS-related syndrome (ARC) were immunophenotyped with the use of monoclonal antibodies and flow cytometry. The patients were placed into four clinically defined groups: HIV-positive asymptomatic (HIV+/A, 8); persistent generalized adenopathy (14); Kaposi's sarcoma (12); and opportunistic infections (17). Immunophenotypes were compared between groups. Statistically significant differences were seen in absolute lymphocyte counts, total T-cells, helper/inducer T-cells, the helper inducer subset of CD4+ lymphocytes, the suppressor inducer subset of CD4+ lymphocytes, activated helper T-cells, and natural killer cells. CD8+ cells and subsets were not statistically different between groups, possibly obscured by large ranges, but median values suggested differences. Results indicate a pattern of increasing or decreasing numbers of certain subpopulations as HIV infection progresses.     

Phenotype of peripheral blood lymphocytes and serum immunoglobulin concentration in patients with early rheumatoid arthritis

We studied the phenotype of peripheral blood lymphocytes and serum immunoglobulin concentration in patients with early rheumatoid arthritis and rheumatoid arthritis of more than 12 months duration. Subpopulations of CDIgM⁺, CD25⁺, and HLA-DR⁺ lymphocytes and IgA concentration differed in these groups of patients with rheumatoid arthritis. The count of lymphocytes carrying CDIgM⁺ and HLA-DR⁺ receptors correlated with activity of rheumatoid arthritis. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 141, No. 1, pp. 92–94, January, 2006     

Evidence for autoantibody-induced CD4 depletion mediated by apoptotic and non-apoptotic mechanisms in HIV-positive long-term surviving haemophilia patients

It is believed that autoimmune phenomena and apoptosis contribute to CD4 depletion. We investigated 11 long‐term (>20 years) HIV‐infected haemophilia patients and 10 healthy controls. Using four‐colour‐fluorescence flow cytometry, we studied the proportions of CD3⁺CD4⁺ and CD3⁺CD4^– blood lymphocytes that were CD95⁺, CD95L⁺, immune complex⁺ (IC⁺, consisting of IgM, IgG, C3d and/or gp120), and were viable or non‐viable (propidium iodide⁺ = PI⁺). In addition, we studied viability of CD4⁺IgG⁺ patient lymphocytes using the apoptosis marker annexin and the permeability indicator 7‐amino actinomycin D (7‐AAD). HIV⁺ patients had a higher proportion of CD3⁺CD4⁺IgG⁺PI⁺ lymphocytes than healthy controls (median: 3·7%versus 0·3%; P = 0·00001). These non‐viable IgG‐coated lymphocytes might have been killed in vivo by ADCC or complement lysis; 9·1% of the circulating CD3⁺CD4⁺ blood lymphocytes were IgG⁺PI^– (controls: 2·5%; P = 0·001). These viable IgG‐coated lymphocytes might be targets for phagocytosis or anti‐CD95 autoantibody‐mediated apoptosis. Because HIV⁺ patients and healthy controls had similar proportions of PI⁺ or PI^– CD3⁺CD4⁺ lymphocytes that carried CD95L on the surface, and because CD3⁺CD4⁺CD95L⁺ cells that were IgG⁺, C3d⁺ and/or gp120^– were increased in HIV⁺ patients, the role of CD95L‐induced apoptosis in long‐term HIV‐infected haemophilia patients remains unclear. The findings that HIV⁺ patients had higher proportions of CD3⁺CD4⁺CD95⁺ (PI⁺: 6·5%versus 1·4%; P = 0·00002; PI^–: 55·8%versus 44·4%; P = 0·04) blood lymphocytes and that the proportion of CD4⁺IgG⁺Annexin⁺7‐AAD^– blood lymphocytes was associated inversely with peripheral CD4 counts (r = ?0·636; P < 0·05) suggest that attachment of IgG to CD4⁺ blood lymphocytes (anti‐CD95?) induces in some lymphocytes apoptosis with subsequent depletion of these IgG‐coated apoptotic CD4⁺ lymphocytes from the circulation. We found supporting evidence for the contention that autoantibody‐induced apoptotic and non‐apoptotic mechanisms contribute to CD4 depletion in long‐term HIV‐infected haemophilia patients.     

210例HIV/AIDS患者淋巴细胞计数与机会性感染相关分析

目的分析HIV/AIDS患者总淋巴细胞计数（TLC）与机会性感染的关系,为HIV/AIDS患者机会性感染的治疗及一级、二级预防提供参考依据。方法对2009年6月至2011年5月210例HIV/AIDS患者的淋巴细胞及出现的机会性感染进行分析。将患者分为TLC〉1300个/μl组（G1组）和TLC≤1300个/μl组（G2组）,比较两组患者机会性感染发生率的差异。结果 210例HIV/AIDS患者机会性感染的总感染率为86.7%,主要的机会性感染为口腔念珠菌感染（56.2%）、细菌性肺炎（46.7%）、肺结核（42.4%）、败血症（21.4%）、感染性腹泻（20.5%）。G2组患者机会性感染的发生率为93.6%,高于G1组患者（55.3%）,差异有统计学意义（P〈0.05）;随着TLC的下降,患者发生机会性感染的机率增高。结论 HIV/AIDS患者机会性感染的发生率高,TLC是HIV/AIDS患者发生机会性感染的独立危险因素。因此,应定期监测HIV/AIDS患者TLC,加强患者机会性感染的一级和二级预防。     

The influence of substance P on the proliferation of peripheral blood lymphocytes from normal individuals and birch pollen-allergic patients

We have studied the influence of substance P (SP) on the proliferative response of concanavalin A (ConA)-activated peripheral blood lymphocytes from 16 birch pollen-allergic patients, sampled before and during the pollen season, and from 15 normal individuals. The median response to ConA 3 micrograms/ml in the presence of SP 10(-11)-10(-6) M, was in most instances within +/- 10% of the control value for cells from both healthy and atopic individuals. However, the individual differences were considerable. Analysis of the proliferative responses to ConA of the cells from the allergic patients sampled before and during season, revealed higher responses in the presence of 10(-6) than of 10(-7) M SP. This was in contrast to the findings in the normal individuals: only half of their cells showed such increased responses. This difference in response frequency was statistically significant between allergic patients before season and normal individuals (P less than 0.05) and between allergic patients during season and normal individuals (P less than 0.01). The difference in proliferation rate in the SP concentration interval, 10(-6) to 10(-7) M, for the cells from allergic patients, sampled both before and during season, was significantly different from the cells from healthy individuals (P less than 0.03 and P less than 0.001 respectively). The cells sampled from four allergic subjects during the birch pollen season showed a more profoundly decreased response to ConA in the presence of SP 10(-8) M, compared with their cells sampled before season. Such responses were never seen with cells sampled before season and with cells from normal individuals. The results suggest an involvement of SP in the immunoregulation, particularly in patients exhibiting allergic reactions to birch pollen.     

Longitudinal study of plasma HIV-1 RNA concentrations during the asymptomatic stage of HIV infection measured using AMPLICOR HIV monitor and NASBA HIV-1 RNA QT tests

The predictive value of two methods for measuring HIV RNA concentration in plasma was assessed in relation to CD4 lymphocyte counts during the asymptomatic period of infection. The design was a retrospective longitudinal case-control study for a mean period of 60 months involving 20 asymptomatic patients included in the French National prospective survey. The CD4 counts in these patients during the last 36 months of the study were stable (non-progressors) or declined (progressors). Plasma RNA concentrations were determined in each subject annually using the AMPLICOR and NASBA techniques. Only AMPLICOR gave RNA titers above the cut-off value for all the patients. The techniques agreed satisfactorily, although there was a difference, median 0.4 log₁₀, between the AMPLICOR and NASBA values. The non-progressors had low and stable RNA concentrations. The concentration was higher in the progressors, according to the AMPLICOR technique, from their inclusion in the study, and according to the NASBA technique, from 1 year after inclusion. However, only four of ten individual progressors had stable plasma HIV RNA concentrations significantly above those of the non-progressors before the decline in their CD4 counts. These were all and only the patients with a decline in lymphocyte counts more than 100 CD4/mm³/year. In each of the other progressors, the RNA concentration was not significantly different from those of the non-progressors. Thus, when making decisions about therapy, plasma HIV RNA determinations cannot be used in place of CD4 counts and may provide valuable additional information. J. Med. Virol. 54:60–68, 1998. © 1998 Wiley-Liss, Inc.     

非结核分枝杆菌感染患者外周血免疫细胞改变及其临床意义

目的分析非结核分枝杆菌(nontuberculosis mycobacterium,NTM)感染患者外周血免疫细胞表达率的改变以及临床意义.方法选择2016年4月至2017年4月期间苏州市第五人民医院住院的69例NTM感染患者作为病例组,选取同期本院健康体检者66例作为对照组.采用流式细胞术对健康对照组和NTM感染患者(NTM组)外周血免疫细胞表达率进行检测,并比较各组之间的差异.进一步根据感染部位的差异将NTM组分为单侧感染组(29例)和双侧感染组(40例),比较两组之间免疫细胞表达水平的差异.结果与对照组相比较,NTM组的总的T淋巴细胞表达率没有显著改变,而B淋巴细胞与单核细胞表达率出现显著上调[(10.50±0.56)%比(12.26±0.57)%,(5.61±0.24)%比(6.72±0.32)%,t值分别为2.209和2.774,P值均<0.05],自然杀伤(natural killer,NK)细胞表达率出现显著下调(21.87±0.98)%比(18.63±0.98)%,t=2.341,P<0.05).对T细胞亚群的分析发现,NTM组的CD4+T细胞、CD8+T细胞、CD8+CD28-T细胞的表达率与对照组相比,差异无统计学意义(P>0.05),而CD4+CD25HT细胞的表达率较对照组显著上调[(8.05±0.35)%比(3.69±0.19)%,t=11.000,P<0.05],CD8+CD28+T细胞表达率较对照组显著下调[(23.76±0.90)%比(27.07±0.74)%,t=3.039,P<0.05].进一步依据患者感染部位比较分析发现,单侧感染组与双侧感染组的T淋巴细胞、B淋巴细胞、NK细胞、单核细胞在外周血中表达率的差异均无统计学意义(P>0.05).T细胞亚群的结果分析显示,CD4+T细胞、CD8+T细胞、CD4+CD25HT细胞在单侧感染组与双侧感染组之间的表达率差异无统计学意义(P>0.05),而双侧感染组的CD8+CD28+T细胞表达比例较单侧感染组显著下降[(21.99±1.02)%比(26.21±1.52)%,t=2.397,P<0.05]、CD8+CD28-T细胞表达比率较单侧感染组显著上调[(24.48±1.90)%比(18.19±1.60)%,t=2.404,P<0.05].结论NTM感染患者外周血中一些免疫细胞的表达率出现了显著改变,可能对疾病的发生与发展产生了重要影响.     

SLE患者外周血中T、B细胞表面Fas和bcl—2表达的研究

目的 探讨活动期SLE患者外周血中T、B细胞表面Fas和bcl-2的表达水平及其与T、B细胞凋亡的关系。方法 采用流式细胞术,测定活动期SLE患者外周血T、B细胞表面Fas和bcl-2的表达,并同时测定患者血中T、B细胞的凋亡。结果 ①活动期SLE患者外周血中B细胞及CD8T细胞表面bcl-2的表达明显高于正常人（分别为P＜0．05）和P＜0．01）;CD4^ 及CD8^ T细胞表面Fas的表达高于正常人（分别为P＜0．05和P＜0．01）;B细胞表面Fas的表达降低（P＜0．01）;CD4^ T细胞bcl-2的表达下降（P＜0．01）。②活动期SLE患者血中B细胞的凋亡率明显下降,CD8^ T细胞数校正常对照组增加,CD4^ T细胞低于正常人（分别为：P＜0．01、P＜0．05和P＜0．01）。结论 SLE患者体内淋巴细胞凋亡的异常与T、B细胞表面Fas和bcl-2基因的表达的异常有关。     

存放温度及时间对于HIV/AIDS患者外周血CD4+、CD8+细胞测定结果的影响

目的　研究HIV AIDS患者外周血CD4 、CD8 淋巴细胞数在不同条件下 (时间、温度和处理过程 )的变化。方法　选取HIV AIDS患者 34例 ,用流式细胞术检测在 4℃条件下放置不同时间(2、2 4、4 8、72h)的外周血CD4 、CD8 细胞数的变化 ,对经过处理的外周血 (处理过的血样 )CD4 、CD8 细胞数的变化进行比较 ;对室温条件下放置不同时间 (2、2 4、4 8、72h)处理过的血样CD4 、CD8 细胞数的变化进行比较。结果　在 4℃时 ,全血放置 2、2 4、4 8、72h的CD4 细胞计数差异无显著意义(P >0 0 5 ) ,而CD8 细胞数放置 72h时则差异有显著意义 (P <0 0 5 ) ;处理过的样品放置 72hCD4 细胞计数差异才有显著意义 (P <0 0 5 ) ,而CD8 细胞数在 2 4h时差异就有显著意义 (P <0 0 5 )。在室温时 ,处理过血样放置 2、2 4、4 8、72h的CD4 细胞计数差异无显著意义 (P >0 0 5 ) ,而CD8 细胞数在 4 8h则差异有显著意义 (P <0 0 5 )。结论　抗凝全血在 4℃放置 4 8h ,检测CD4 、CD8 淋巴细胞数 ,结果是可靠的。处理过血样在室温放置 2 4h ,检测CD4 、CD8 淋巴细胞数 ,结果是可靠的。 2 4～4 8h虽然CD4 淋巴细胞没有变化 ,但CD8 淋巴细胞却发生明显的变化 ,两者比例必然发生变化。     

Expression of CTLA-4 (CD152) in peripheral blood T cells of children with influenza virus infection including encephalopathy in comparison with respiratory syncytial virus infection

Influenza virus and respiratory syncytial virus (RSV) are the most common causes of acute severe respiratory infection in children during the winter. There have been few reports about peripheral blood T cell activation in vivo in influenza virus infection and conflicting results concerning peripheral blood T cells activation in RSV infection. Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4, CD152) is a receptor present on T cells that plays a critical role in the down-regulation of antigen-activated immune responses. To clarify the status of peripheral blood T cells, we investigated intracellular CTLA-4 expression in T cells in patients with influenza virus and RSV infection. We collected blood samples from 15 patients with influenza virus infection, including three with complications of influenza virus-associated encephalopathy and 18 patients with RSV infection, as well as 44 healthy children. We determined the intracellular expression of CTLA-4 in CD4+ and CD8+ T cells by flow cytometry. There were no significant differences in the percentages of intracellular CTLA-4-positive CD4+ T cells and CD8+ T cells by age. The percentages of intracellular CTLA-4-positive CD4+ T cells in the patients with influenza virus infection were significantly higher than those in healthy children (P < 0.01). In particular, the patients with influenza virus-associated encephalopathy had sevenfold higher percentages of CTLA-4-positive CD4+ T cells than influenza patients without encephalopathy (P < 0.05). The patients with influenza virus-associated encephalopathy had increased percentages of CTLA-4-positive CD8+ cells at the acute stage in comparison with the convalescent stage and in control subjects (P < 0.01, respectively). RSV patients showed no increase in CTLA-4-positive CD4+ T cells or CD8+ T cells. The immunological status of peripheral T cell activation is substantially different in influenza virus infection and RSV infection. The patients with RSV infection did not show any increase in CTLA-4-positive peripheral blood T cells. There was a remarkable increase in intracellular CTLA-4 in CD4+ and CD8+ T cells in influenza virus-associated encephalopathy. Down-regulation of antigen-activated peripheral blood T cell activation might play an important role in the pathogenesis of influenza virus-associated encephalopathy and host defence against influenza virus infection.     

Ribavirin inhibits DNA,RNA, and protein synthesis in PHA-stimulated human peripheral blood mononuclear cells: possible explanation for therapeutic efficacy in patients with chronic HCV infection

The treatment of choice for patients infected chronically with HCV is the combination of IFN-alpha and ribavirin. Monotherapy with ribavirin leads to a clinical and histological improvement, but its exact mechanism of action is unknown. Therefore, the effect of ribavirin on synthesis of inflammatory cytokines and on apoptosis in stimulated peripheral blood mononuclear cells (PBMCs) was investigated. PBMCs were isolated from the blood of HCV infected patients and from healthy volunteers. The effect of ribavirin on IFN-gamma and IL-1beta release in the supernatant of unstimulated and phytohemagglutinin (PHA) stimulated PBMCs was investigated by enzyme linked immunosorbent assay (ELISA). The effect on total DNA, RNA, and protein synthesis was analyzed by measurement of 3H-thymidine, 3H-uridine and 3H-leucine incorporation into cellular macromolecules. Ribavirin led to a dose-dependent decrease of the IFN-gamma but an increase of IL-1beta release into the supernatant of PHA-stimulated PBMCs. At the same time, a dose-dependent decrease of total DNA, RNA, and protein synthesis in cultures of PHA-stimulated PBMCs was demonstrated. These effects could be compensated by the addition of equimolar amounts of guanosine. The rate of apoptotic CD45+ and CD14+ cells in PBMCs cultures increased in a dose-dependent manner. Our data suggest that ribavirin administration to chronically HCV-infected patients could lead to a decrease of the synthesis of proinflammatory cytokines (e.g., IFN-gamma) by an inhibition of total DNA-, RNA-, and protein-synthesis and by induction of apoptosis in the cells of the inflammatory infiltrate. Furthermore, ribavirin could influence the synthesis of viral particles in the hepatocytes.     

Virological analysis and phenotypic characterization of peripheral blood lymphocytes of hepatitis C virus-infected patients with and without mixed cryoglobulinaemia

In clinical and pathological terms hepatitis C virus (HCV)-infected patients can be subdivided into two main groups with and without mixed cryoglobulinaemia (MC). Involvement of blood mononuclear cells by HCV has potentially important implications. To this end, HCV-RNA levels in peripheral blood lymphocytes (PBL) preparations of 20 chronically HCV-infected patients with MC were measured and compared with those found in a group of 20 patients without MC matched for age, serum HCV-RNA, infectious genotype, source and presumable duration of infection. Phenotypic abnormalities of PBL subsets in each group of patients were determined by cell surface marker expression and compared. Results showed a significant enrichment of HCV-RNA in PBL of MC patients compared with a non-MC group (P = 0.01). Different distribution of HCV-RNA was accompanied by evidence of an increased frequency of circulating B cells. These data indicate that MC patients are characterized distinctly by a higher quota of cell-associated viral load.