人类疱疹病毒7型感染脐血单个核细胞产生TNF—α

用生物活性法,检测人类疱疹病毒7型Glasgow株和南京地方株YY5及HHV-6GS株感染单个核细胞培养上清中肿瘤坏死因子－α（TNF-α）的水平。结果发现,HHV-7对也能较强地诱生TNF-α,但达到峰值时间（3～4d）迟于HHV-6GS株（2d）;在感染24h上请中,GS株产生TNF-α量明显多于YY5株、Glasgow株产生量（P＜0．05）,三者与未感染单个核细胞比较都存在显著差异（P＜0．05）,但Glasgow珠和YY5株之间无差异（P＞0．05）。结果表明,HHV-6、HHV-7都能通过刺激单个核细胞产生TNF-α而发挥免疫调节功能。     

不同镇痛方法对患者IL-6、IL-8、IL-10的影响

目的评价不同镇痛方法对盆腔手术患者血浆中细胞因子变化的影响,并与传统术后镇痛法进行比较,探讨术后充分镇痛对免疫功能的影响。方法根据镇痛方法不同,将60例行子宫切除患者随机分为3组：第1组患者为术后根据临床需要,临时给予哌替啶50mg肌肉注射（Ⅰ组,n=20）;第Ⅱ组患者为罗哌卡因复合芬太尼硬膜外镇痛组（Ⅱ组,n=20）;第Ⅲ组患者为芬太尼静脉镇痛组（Ⅲ组,n=20）;观察麻醉前30min、手术后30min、2h、24h、48h和72h六个时点患者血清中白细胞介素-6（IL-6）、白细胞介素-8（IL-8）、白细胞介素-10（IL-10）水平的变化。结果3组患者术后血清IL-6、IL-8、IL-10水平与麻醉前值比较均升高（P〈0．01）,一般在术后24h达峰值。比较血清IL-6、IL-8、IL-10浓度变化,Ⅱ和Ⅲ组抑制这3种细胞因子释放的能力明显强于Ⅰ组（P〈0．05）,与Ⅲ组比较,Ⅱ组更为明显（P〈0．05）。结论硬膜外局麻药复合阿片受体激动药镇痛模式可更有效地降低术后炎性直激反应。     

人IL—6及其受体拮抗剂的研究进展

人IL－6及其受体拮抗剂的研究主要集中于两个方面：单克隆抗体和突变体,针对人IL－6和人IL－6R的活性区域构建的单克隆抗体对于临床治疗多发性骨髓瘤显示出了很好的短期疗效,根据与生长激素（GH）及其受体复合物GH／（GHbp)2的结构对比,推测人IL－6和IL－6Ra的活性位点,结合定点突变技术,设计IL－6突变体,IL－6R突变体和IL－6突变体－IL－6Ra融合蛋白,它们对天然hIL-6的生物活性显示出了明显的拮抗作用。     

细菌内毒素对纤维细胞合成IL-6、IL-8的诱导作用

探讨细菌内毒素（LPS）对成纤维细胞合成IL－6、IL—8的诱导作用。以不同浓度的LPS刺激NIH3Th细胞,用EI。ISA法检测受刺激后的NIH3Th细胞培养肝清波中的IL－6、IL－8浓度,受LPS刺激后,NIH3Th细胞培养上清波中的11。－6含复为3．7fi．31n玖rnl（8）,7．3I2．2吟rTil（8）;11。－8含展为3．8ilO．94吟rnl（SN,8．39if．52略加（48h）,与对照组有显著性差别,对见lPS叶诱导NIH3Th细胞合成分泌IL－6、IL—8的水平增高,从而参与抗炎反应和炎症过程。细菌内毒素对纤维细胞合成IL-6、IL-8的诱导作用@闻平$镇江医学院…     

人类疱疹病毒6型可诱生TNF－α

用生物活性法和双抗体夹心桥联酶免疫吸附（ＥＬＩＳＡ）法检测了人类疱疹病素６型（ＨＨＶ－６）ＧＳ株和南京地方株ＣＮ５,８,１０感染的淋巴细胞培养上清中的肿瘤坏死因子（ＴＮＦ）的水平,发现培养２４ｈ即可检出高水平的ＴＮＦ,４８～７２ｈ述到峰值,此后逐渐下降,与未感染耐照组比较有及其显著的差异（Ｐ＜０．００１）。ＧＳ株与地方株同诱生ＴＮＦ水平无儿著性差异（Ｐ＞０．１）,三株地方株诱生ＴＮＦ也无显著性差异（Ｐ＞０．０５）。ＴＮＦ－α单抗可以完全中和培养上清中ＴＮＦ的活性,证实上清中有ＴＮＦ－α。与ＬＰＳ比较,ＨＨＶ－６诱生ＴＮＦ－α的能力要强得多。     

猪白细胞介素_6在巴斯德毕氏酵母（Pichia pastoris）中的分泌表达

白细胞介素6（Interleukin_6, IL_6）是一种具有多种生物学效应的细胞因子,在疾病诊断与疫苗佐剂领域有广阔的应用前景。在本试验中,猪白细胞介素_6（pIL_6）的cDNA序列被克隆入甲醇酵母（Pichia pastoris）分泌表达载体pPIC9K中,并转化入P. pastoris GS115菌株。其重组菌株GS115/pPIC9K_IL6经1% 甲醇诱导后,能分泌表达分子量约为24.5KD的重组蛋白,Western blot确证为pIL_6。该酵母表达产物无N端糖基化修饰。用依赖IL6生长的B9细胞株检测提纯后的pIL_6,其生物学活性可达8×104IU/mg。     

Balb／c小鼠左右两侧大脑皮层IL—1β，IL—6含量的不对称性

为探讨左右侧大脑皮层白细胞介素1β（interdeukine-1β,IL-1β）和白细胞介素6（interleukine-6,IL-6)的含量状况及与左右侧大脑皮层免疫调控异质性的关系,。分别取正常及细胞菌脂多糖（lipopolysaccharuide,LPS)刺激2h后的Balb/c小鼠的左右侧大脑皮层,制备匀浆液,用ELISA法检测匀浆液中IL－1β,IL-6的含量,结果显示,正常小鼠大脑皮层IL－1β,IL-6含量均为右侧显著高于左侧;LPS刺激后,左侧大脑皮层IL－6含量明显升高,右侧大脑皮层IL－6含量显著高于左侧大脑皮层,而左右侧大脑皮层IL－1β含量无明显差异,该实验结果表明,Balb/c小鼠在正常生理状态下两大脑皮层IL－1β,IL－6含量存在不对称性,左右侧大脑皮层免疫调控异质性可能与细胞因子的不对称性有关。     

Ghrelin抑制H202诱导的肺泡上皮A549细胞中IL-8的产生

目的：通过细胞培养的方法,初步研究了生长激素释放肽Ghrelin在氧化应激相关的肺泡上皮细胞炎症反应中的作用。方法：首先是用双氧水（H202）刺激A549细胞建立肺泡上皮细胞炎症反应模型,分别加入不同浓度的Ghrelin及普通培养基培养A549细胞。用酶联免疫吸附技术（ELISA）从蛋白水平检测上清液中IL-8的含量,采用逆转录聚合酶链反应技术（RT—PCR）检测炎性细胞因子IL．8mRNA的表达。结果：H2O2可以使A549细胞IL-8蛋白的释放及IL-8mRNA的表达明显升高（P〈0．05）,而用Gllrelin干预后IL．8的释放及IL-8mRNA的表达被抑制,明显低于单纯H2O2刺激的模型组（P〈0．05）,且随着浓度的增加Ghrelin的这种抑制作用逐渐增强。结论：分别从蛋白水平和基因水平证明了Ghrelin能够抑制H202诱导的肺泡上皮A549细胞中IL-8的产生,由此推测Ghrelin可能能够抑制以COPD、支气管哮喘为代表的氧化应激相关的肺部炎症反应。     

HHV—6体外感染对外周血单个核细胞IL—6,IL—8的诱生和NK活性的影响

采用生物活性法和／或酶联免疫吸附法和以及乳酸脱氢酶释放法,研究了人类疱疹病毒６型南京地方株ＣＮ８对人体周血单个核细胞的ＩＬ－６,ＩＬ－８的诱生和ＮＫ活性的影响,并与国外的ＧＳ侏作比较。结果发现,ＨＨＶ－６ＣＮ８,ＧＳ两种病毒感染均苛诱导ＰＢＭＣｓ产生ＩＬ－８,４８ｈ达到峰值。     

人胎脾细胞IL-2和IL-6的产生及其与NK细胞功能发育的关系

人类胚胎发育时期,脾细胞IL-2和IL-6的产生及其与NK细胞功能发育关系的研究结果表明,胚胎20周龄前IL-2的活性和NK活性细胞基本缺乏,但可分泌低水平的IL-6;随个体发育,IL-2、IL-6的产生和NK细胞活性均逐渐增强,三者间呈直线正相关关系(r>0.86);出生前,IL-6的产生和NK细胞活性显著低于成人组(p<0.01),而IL-2的产生巳达成人水平(p>0.05)。最后,对在胚胎发育过程中IL-2和IL-6的产生,及其与NK细胞功能发育间的关系进行了讨论。     

人类疱疹病毒6型感染细胞免疫学特性研究

采用间接免疫荧光法、ＡＰＡＡＰ法及ＭＴＴ法,研究了人类疱疹病毒６型（ＨＨＶ６）中国南京地方株ＣＮ５感染细胞病毒抗原表达的形态学和动力学特征、ＣＤ抗原表达阳性细胞百分率的变化及ＰＨＡ诱导的细胞增殖反应的改变。结果显示,ＣＮ５感染脐血单个核细胞（ＣＢＭＣｓ）后８～１２ｈ即可在细胞内检出病毒抗原,至接种后４８ｈ,病毒抗原阳性细胞可达３６％;ＣＮ５感染ＣＢＭＣｓ和成人外周血单个核细胞（ＰＢＭＣｓ）后可引起两者ＣＤ３阳性细胞减少、ＣＤ４阳性细胞增多,而对ＣＤ２、ＣＤ８、ＣＤ４５ＲＡ阳性细胞百分率未见明显影响;ＣＮ５感染细胞裂解液对ＰＨＡ诱导的ＰＢＭＣｓ增殖反应具有抑制作用,这种抑制作用与该裂解液的蛋白浓度之间呈一剂量依赖关系,且可被ＨＨＶ６抗血清所逆转。     

Human herpesvirus-6-specific interleukin 10-producing CD4+ T cells suppress the CD4+ T-cell response in infected individuals

Human herpesvirus‐6 (HHV‐6) infection normally persists for the lifetime of the host and may reactivate with immunosuppression. The mechanism behind HHV‐6 latent infection is still not fully understood. In this study, we observed that decreased proliferation of CD4⁺ T cells and PBMCs but not CD8⁺ T cells from HHV‐6‐infected individuals was stimulated with HHV‐6‐infected cell lysates. Moreover, HHV‐6‐stimulated CD4⁺ T cells from HHV‐6‐infected individuals have suppressive activity on naïve CD4⁺ T and CD8⁺ T cells from HHV‐6‐uninfected individuals. However, no increased proportion of CD4⁺ CD25⁺ Treg cells from HHV‐6‐infected individuals contributed to the suppressive activity of the HHV‐6‐stimulated CD4⁺ T cells from HHV‐6‐infected individuals. Transwell experiments, ELISA and anti‐IL‐10 antibody blocking experiment demonstrated that IL‐10 may be the suppressive cytokine required for suppressive activity of CD4⁺ T cells from HHV‐6‐infected individuals. Results of intracellular interleukin (IL)‐10 and IL‐4 further implicated the HHV‐6‐speciflc IL‐10‐producing CD4⁺ T cells in the suppressive activity of CD4⁺ T cells from HHV‐6‐infected individuals. Results of intracellular interferon (IFN)‐γ demonstrated a decreased frequency of HHV‐6‐speciflc IFN‐γ‐producing CD4⁺ T, but not CD8⁺ T cells in HHV‐6‐infected individuals, indicating that it was the CD4⁺ Th1 responses in HHV‐6‐infected individuals that were selectively impaired. Our findings indicated that HHV‐6‐specific IL‐10‐producing CD4⁺ T cells from HHV‐6‐infected individuals possess T regulatory type 1 cell activity: immunosuppression, high levels of IL‐10 production, with a few cells expressing IFN‐γ, but none expressing IL‐4. These cells may play an important role in latent HHV‐6 infection.     

Cytokine and cellular responses to human herpesvirus‐6B in patients with B‐acute lymphoblastic leukemia

Primary infection with human herpesvirus‐6 (HHV‐6), is followed by its lifelong persistence in the host. Most T‐cell responses to HHV‐6 have been characterized using peripheral blood from healthy adults; however, the role of HHV‐6 infection in immune modulation has not been elucidated for some diseases. Therefore, in this study the immune response to HHV‐6 infection in patients with B‐acute lymphoblastic leukemia (B‐ALL) was analyzed. HHV‐6 load was quantified in blood samples taken at the time of diagnosis of leukemia and on remission. The same concentrations of anti‐ and pro‐inflammatory cytokines (IL‐4, IL‐1, IL‐6, IL‐8, IL‐12p70, IL‐17a, TNF‐α and IFN‐γ) were detected in plasma samples from 20 patients with and 20 without detectable HHV‐6 virus loads in blood. Characterization of T‐cell responses to HHV‐6 showed low specific T‐cells frequencies of 2.08% and 1.46% in patients with and without detectable viral loads, respectively. IFN‐γ‐producing T cells were detected in 0.03%–0.23% and in 0%–0.2% of CD4+T cells, respectively. Strong production of IL‐6 was detected in medium supernatants of challenged T‐cells whatever the HHV‐6 status of the patients (973.51 ± 210.06 versus 825.70 ± 210.81 pg/mL). However, concentrations of TNF‐α and IFN‐γ were low. Thus, no association between plasma concentrations of cytokines and detection of HHV‐6 in blood was identified, suggesting that HHV‐6 is not strongly associated with development of B‐ALL. The low viral loads detected may correspond with latently infected cells. Alternatively, HHV‐6B specific immune responses may be below the detection threshold of the assays used.     

双歧杆菌对裸鼠腹腔巨噬细胞产生IL—1及IL—6的影响

给裸小鼠腹腔注射活的青春型双歧杆菌,并以小鼠胸腺细胞增殖法及ＥＬＩＳＡ法分别检测了裸鼠腹腔巨噬细胞分泌的ＩＬ１活性及ＩＬ６含量。结果表明：实验组裸鼠腹腔巨噬细胞分泌的ＩＬ１活性以及ＩＬ６含量均显著高于对照组,两者均具有统计学意义（ｐ＜００１）。这提示青春型双歧杆菌可激活巨噬细胞产生ＩＬ１以及ＩＬ６,它们在该菌调节机体免疫反应中可能起一定作用。     

Constitutive release of IL6 by human papillomavirus type 16 (HPV16)-harboring keratinocytes: a mechanism augmenting the NK-cell-mediated lysis of HPV-bearing neoplastic cells.

In the present study we demonstrate that the cultured human keratinocyte cell line (SK-v cells) harboring and expressing integrated human papillomavirus type 16 (HPV16) DNA sequences constitutively releases IL6, which is known as a pleiotropic immunoregulatory cytokine of potential antitumor properties. The presence of IL6 activity in SK-v cell-conditioned media (SK-v CM) was demonstrated by tritiated thymidine incorporation into IL6-dependent B9 murine plasmacytoma cells. The effect on B9 cells was specific since it could be inhibited by anti-IL6 neutralizing antibodies but not by a normal control serum. IL6 did not affect SK-v cell growth; however, it significantly augmented NK cell activity of human peripheral blood lymphocytes against both K562 erytholeukemic and SK-v cells as assessed by 51Cr release assay. SK-v CM displayed NK cell-augmenting activity that copurified with IL6 activity in both size exclusion and anion-exchange HPLC. Furthermore, SK-v cell-derived NK cell stimulatory activity could be neutralized with anti-IL6 antibodies. These results suggest that HPV-harboring neoplastic cells can release IL6 which may indirectly mediate tumor death by augmentation of NK cell activity.     

A组轮状病毒NSP6蛋白表达及免疫学性质研究

轮状病毒(RV）NSP6与NSP5由同一基因片段编码,至今对NSP6 性质了解很少。用基因重组表达和免疫学方法,重组表达了A组人RV NSP6蛋白,进行了NSP6的动物免疫及其抗原反应性、免疫原性研究以及RV感染细胞中NSP6的合成及亚细胞分布研究。研究结果表明,NSP6可在原核系统中高效表达,表达蛋白占菌体总蛋白的34.2%;NSP6免疫豚鼠血清抗体可特异性识别菌体细胞中表达的NSP6和SA11及Wa病毒感染的MA104细胞中合成的NSP6蛋白;病毒感染细胞中合成的NSP6在感染后3h就可检测到,12h表达量达到最高;NSP6在病毒感染细胞质中呈弥散状分布,并主要积聚在细胞核的周围,未观察到毒质体样结构。研究结果对深入了解RV NSP6的结构与功能具有重要的意义,具有重要的潜在应用价值。     

Identification of antigenic proteins encoded by human herpesvirus 8 and seroprevalence in the general population and among patients with and without Kaposi's sarcoma

To establish a sensitive and specific antibody assay, potent antigenic proteins encoded by human herpesvirus 8 (HHV8) were studied. Fifteen recombinant HHV8-encoded proteins were produced as glutathione S-transferase fusion proteins. The sera from AIDS-associated Kaposi's sarcoma (KS) patients reacted with four proteins encoded by open reading frames (ORFs) K8.1, 59, 65, and 73 in a Western blot assay. An enzyme-linked immunosorbent assay (ELISA) using these four proteins as antigens (mixed-antigen ELISA) revealed that all 26 sera derived from KS patients (24 with and 2 without human immunodeficiency virus infection) became positive for anti-HHV8 antibodies. The presence of HHV8 was demonstrated in 14 (1. 4%) of 1,004 sera from the Japanese general population and 10 (1.9%) of 527 sera from patients without HHV8-associated diseases. The presence of immunoglobulin G (IgG) and IgM antibodies against HHV8 examined further by the mixed-antigen ELISA and Western blotting revealed IgG antibody in all ELISA-positive sera, while IgM antibody against ORF K8.1 was absent. These data suggest that the ORF 73 and 65 proteins are potent antigens for a sensitive serological assay.     

Natural killer cell-dependent mycobacteriostatic and mycobactericidal activity in human macrophages

Host defense mechanisms against Mycobacterium avium complex (MAC) are poorly understood. Recent evidence suggests the role of NK cells in the host defense against some intracellular pathogens. We investigated whether NK cells play a role in MAC infection. IL-2-activated human NK cells were incubated with human monocyte-derived macrophages either before or after infection with MAC. Macrophages were lysed 3 and 5 days after infection for quantitation of viable intracellular organisms. Although no killing was observed by nonstimulated macrophages, exposure to IL-2-treated NK cells for 24 h before infection induced macrophage to kill 70 +/- 8% of intracellular MAC by 3 days, and 81% +/- 4% in 5 days (p less than 0.01 for both compared with control). Killing was not blocked by incubation with anti-TNF antibody (Ab) or anti-IFN-gamma Ab. Similarly, incubation of macrophages for 24 h with supernatant obtained from IL-2 activated NK cells was associated with 74 +/- 4% killing of intracellular MAC in 3 days and 81 +/- 6% in 5 days (p less than 0.01 for both compared with control). However, the supernatant-mediated activation was partially blocked by anti-TNF Ab (46 +/- 6%; p less than 0.05) but not by anti-IFN gamma Ab. When infected macrophages were incubated with NK cells 24 h after infection for 48 h, they killed 54 +/- 3% of intracellular M. avium in 3 days and 73 +/- 5% in 5 days (p less than 0.02 for both compared with control). This effect was also not blocked by either anti-TNF or anti-IFN gamma Ab. These results suggest that activated NK cells may have an important role in the intracellular killing of MAC and that the NK-mediated activation of macrophages is in part mediated by TNF.     

Viral IL-6 blocks neutrophil infiltration during acute inflammation

Pathologies arising as a consequence of human herpesvirus-8 (HHV8) infections are closely associated with the autocrine activity of a HHV8 encoded IL-6 (vIL-6), which promotes proliferation of infected cells and their resistance to apoptosis. In this present report, studies show that vIL-6 may also be important in influencing the host's immunological response to secondary infections. Using peritoneal inflammation as a model of acute bacterial infection, vIL-6 was found to specifically block neutrophil recruitment in vivo through regulation of inflammatory chemokine expression. This response was substantiated in vitro where activation of STAT3 in human peritoneal mesothelial cells by vIL-6 was associated with enhanced CCL2 release. Although vIL-6 did not effect CXCL8 production, IL-1beta-induced secretion of this neutrophil-activating chemokine was significantly suppressed by vIL-6. These data suggest that vIL-6 has the capacity to suppress innate immune responses and thereby influence the outcome of opportunistic infections in HHV8-associated disease.     

Effects of natural and recombinant IL 2 on regulation of IFN gamma production and natural killer activity: lack of involvement of the Tac antigen for these immunoregulatory effects

Highly enriched populations of human large granular lymphocytes (LGL), natural killer (NK) cells, and T cells were obtained from low and high density fractions, respectively, of discontinuous Percoll gradients. The NK cells were composed of 75 to 90% LGL, with the majority of the contaminating cells being monocytes. The T cells were greater than 95% OKT3+. The proliferative and cytotoxic progenitors in both fractions were examined by using a limiting dilution assay with interleukin 2 (IL 2) from four sources: 1) crude supernatant of a gibbon lymphoma (MLA-144), 2) purified (150,000-fold) MLA-144 IL 2, 3) partially purified human IL 2, and 4) purified recombinant human IL 2. The proliferative capacity was measured at day 7 by [3H]thymidine incorporation, whereas the progenitors of cells with NK-like activity were evaluated by assessing cytotoxic activity against K562 cells at day 8 in a 4-hr 51Cr-release assay. The frequency of proliferative progenitors among T cells was approximately 1/5 and was approximately 1/60 with LGL. Titration of the highly purified IL 2 preparation demonstrated that LGL proliferated with as little as 2 U of IL 2. The frequency of detectable cytotoxic progenitors in the LGL population, however, fell sharply when less than 40 U of IL 2 were employed. The T cells failed to demonstrate cytotoxic activity against the NK-susceptible target cells at any concentration of IL 2 tested. The IL 2 preparations also were examined for their ability to directly and rapidly enhance the cytotoxic activity of highly purified NK cells. All four preparations of IL 2 enhanced the cytotoxic activity of LGL without any detectable accessory requirement after incubation for as little as 6 hr, even though the MLA-144 IL 2 preparations were devoid of detectable interferons (IFN). These data indicate that IL 2 has dual effects on NK cells, regulating their activity was well as promoting their proliferation. Collectively, these results demonstrate that highly purified IL 2, devoid of other detectable lymphokines, is capable of supporting the growth of human NK cells and augmenting their in vitro activity. In parallel experiments, these same IL 2 preparations were quite active in causing the proliferation of T lymphocytes, clearly demonstrating a role of IL 2 in promoting the proliferation of NK cells as well as T cells. The mechanism of IL 2 boosting appears to be a direct interaction with LGL, resulting in the production of IFN gamma.(ABSTRACT TRUNCATED AT 400 WORDS)