胞苷环3′,5′一磷酸对T原淋巴细胞株JM的抑制作用

在从S至G_2/M期和从G_1至S期的过程中,3′,5′-cCMP能抑制用胸腺嘧啶同步化的JM-T原淋巴细胞株细胞的增殖。而5-CMP,5-dCMP和2′,3′-cCMP却能在S期促进细胞生长。对于非同步化生长的JM细胞,3′,5′-cCMP也显示出抑制作用。     

IL-2产生能力和敏感性的测定

本文仅就IL-2的产生能力和敏感性的测定方法介绍如下:一、人的IL-2的制备可用扁桃体或外周血淋巴细胞.常用Ficoll比重离心法,从几名正常人外周血中分得单个核细胞,再用尼龙绵吸附法分得非粘附性细胞,在含有1%FCS,5×10~(-5)M的2ME,10mM的Hepes,0.2%PHA的RPM1640培养液中培养24-48小时,采集上清液即为IL-2的粗制品.如在培养液中加入淋巴母细胞传代株作为同种异型抗原刺激,或加入5-10μg/ml的PMA(乙酸肉豆寇佛波醇)、5×10~(-6)M消炎痛(indo methacin)都可增加IL-2的产生能力.最     

小儿急性肾炎患者IL-2、IL-6、TNF水平的变化

目的:探讨了小儿急性肾炎患者血清白介素-2(IL-2)、白介素-6(IL-6)、肿瘤坏死因子(TNF)在急性肾炎发生、发展过程中的变化及相互关系。方法:用放射免疫分析检测了31例急性肾炎患儿血清中IL-2、IL-6、TNF水平,并以30例正常健康儿童作比较。结果:急性肾炎患儿血清中IL-6、TNF水平显著高于正常人组(P<0.01),而IL-2水平显著低于正常人组(P<0.01)。直线相关分析显示,IL-6、TNF水平与BUN呈正相关(P<0.01)。结论:血清IL-2、IL-6、TNF在小儿急性肾炎的发生、发展过程中相互作用,观察其浓度的变化对探讨其发病机理及指导用药均有重要临床价值。     

谷氨酰胺、地塞米松对内毒素血症大鼠血清IL-8,IL-10的影响

目的通过观察谷氨酰胺、地塞米松对内毒素血症大鼠血清IL-8,IL-10的影响,探讨地塞米松、谷氨酰胺对内毒素血症大鼠的保护途径.方法选健康18日龄Wistar大鼠120只,随机取8只腹腔注射生理盐水作对照组(0h),余大鼠按腹腔注射药物不同分成内毒素(L)、谷氨酰胺预防(Gln)地塞米松治疗(D1)组,每组40只,各组于加LPS后2、4、6、24及72h分别断头取血,用放免法测定IL-8,IL-10.结果(1)L组IL-8在2、24h出现两次升高,前者升高程度更明显,差异显著,P<0.01;D1和C1n组各时间点IL-8均低于L组,尤其以2h抑制作用更强,差异显著,P<0.01;Gln组第一次高峰在6h.(2)L组IL-10在2、72h出现两次升高,后者升高程度更明显,差异显著,P<0.01;D1和Gln组第二次升高均提前至24h,增幅最高的为D1组,而其它各时间点IL-10均以Gln组升高明显,差异显著,P<0.01.结论细胞因子IL-8/IL-10在内毒素血症中起主要的损害与抗损害作用,地塞米松治疗对机体能抑制损伤/促进修复;谷氨酰胺不仅有与地塞米松相似的作用,而且可使损伤延迟,二者都能使修复作用提前.     

HBV感染患者2′,5′寡腺苷酸合成酶、IL-2和IL-12水平检测及意义

目的:选用2′,5′寡腺苷酸合成酶(2-5OAS)作为干扰素观察指标,IL-2、IL-12作为Th1应答观察指标,了解内源性干扰素系统和Th1应答在HBV感染发病机制中作用。方法:用放射同位素法测定单核细胞2-5OAS活性;ELISA法测定血清IL-2、IL-12。结果:无症状HBsAg携带组2-5OAS、IL-2、IL-12含量与正常对照组无显著差异(P>0.05),急性肝炎组均显著高于正常对照组(P<0.01),慢性乙型肝炎轻、中、重度组、慢性重型肝炎组、肝硬化组、肝癌组均显著低于正常对照组(P<0.05),并且随慢性肝炎病情加重以及肝硬化、肝癌发生而递减,其中肝硬化、肝癌组处于最低水平(与慢性肝炎各组比较P<0.05)。结论:在HBV感染发病过程的不同阶段和不同临床分型患者中,其内源性干扰素系统和Th1应答反应都是有显著差异的,细胞免疫对病毒感染的痊愈起主导作用。     

尖锐湿疣患者治疗前后血清IL-2、IL-6和TNF-α水平检测的临床意义

为探讨尖锐湿疣患者治疗前后的血清IL-2、IL-6、TNFα-水平及其临床意义,分别应用RIA法和ELISA法对42例尖锐湿疣患者测定血清IL-2、IL-6和TNFα-水平,并与30名健康人(正常组)作比较。结果表明,尖锐湿疣患者在治疗前血清TNF-α水平显著高于正常组(P<0.01),而IL-2、IL-6则显著低于正常组(P<0.01),经6个月治疗后,与正常组比较仍有显著性差异(P<0.05)。结论:尖锐湿疣患者的免疫功能存在明显缺陷。     

Bach2对Th2细胞内IL-33生物学功能的影响

目的：探讨Bach2与Th2细胞内IL-33生物学功能的关系。方法：构建Bach2基因的条件敲除小鼠(Bach2-CKO小鼠),将Bach2-CKO小鼠与正常小鼠分为两组,IL-33经鼻诱导Th2型免疫应答,对比分析两组小鼠肺部炎症情况;从小鼠脾脏中分离na?ve CD4~+T细胞并诱导分化为Th2细胞;下调(或上调)细胞内Bach2表达后给予IL-33刺激,检测Th2型细胞因子表达。结果：与正常小鼠相比,IL-33显著增加Bach2-CKO小鼠肺组织内炎症细胞浸润、黏液腺分泌以及IL-4、IL-5和IL-13mRNA表达;与正常Th2细胞相比,IL-33更加显著地促进Bach2-/-Th2细胞IL-5和IL-13表达(P<0.01);上调细胞内Bach2表达显著抑制IL-33相关IL-5和IL-13表达(P<0.01)。结论：Bach2参与Th2细胞内IL-33/ST2信号通路调控,可有效抑制该细胞内IL-33的生物学功能。     

IL-6诱导鼻咽癌CNE-2细胞分泌IL-10的机制研究

目的探讨IL-6诱导鼻咽癌CNE-2细胞分泌IL-10的分子机制。方法 IL-6体外刺激鼻咽癌细胞系CNE-2细胞,或加入抗IL-6受体的抗体和信号通路抑制剂预孵育1 h,IL-6再进行刺激,RT-PCR检测IL-10的mRNA表达水平,酶联免疫吸附试验检测细胞培养上清中IL-10的分泌水平。结果与未刺激组相比,IL-6刺激的CNE-2细胞显著上调IL-10的表达和分泌,差异具有极显著性统计学意义(P0.01),并以剂量和时间依赖的方式增加;加入抗IL-6受体的抗体或NF-κB抑制剂预孵育后,IL-6刺激CNE-2细胞表达和分泌IL-10的水平显著下降,差异具有极显著性统计学意义(P0.01)。而PI3K抑制剂、p38/MAPK抑制剂、JNK抑制剂、STAT3抑制剂和MEK1/2抑制剂对IL-6诱导的IL-10表达和分泌水平则无明显的影响。结论 IL-6经IL-6R/NF-κB信号诱导CNE-2细胞分泌IL-10,抑制IL-6信号有助于鼻咽癌的临床免疫治疗。     

尿毒症血液透析患者血清GH/IGF-1、IL-2、IL-4水平的变化及相关性研究

目的 观察尿毒症血液透析(HD)患者血清GH/IGF-1、IL-2、IL-4水平的变化及其相互关系,以探讨其在免疫-内分泌平衡中的作用.方法 选择终末期尿毒症行HD患者35例及健康对照者20例,于清晨空腹采血,采用双抗体夹心ABC-ELISA法测定血清IGF-1、IL-2、IL-4的含量,化学发光法测定GH水平,多元相关分析研究它们之间的关系.结果 HD患者血清细胞因子IGF-1、IL-2水平与对照组比较明显升高,差异均有显著性(P<0.01);GH、IL-4与对照组比较降低,但无显著性差异(P0.05).多元相关分析表明HD患者血清GH与IGF-1(r=-0.38,P<0.05),GH与IL-4(r=-0.42,P<0.05)均呈显著负相关,而与IL-2无明显相关性;IGF-1与IL-2(r=0.45,P<0.01),IL-2与IL-4(r=0.53,P<0.01),IL-4与IGF-1(r=0.57,P<0.01)均呈显著正相关.结论 HD患者血清IL-2、IL-4水平变化与GH/IGF-1改变相关联,这种变化及其相互影响是免疫-内分泌平衡失调的因素之一.     

5-氮-2’-脱氧胞苷对人卵巢癌细胞SKOV3凋亡及HMLH1和HMSH2表达的影响

目的探讨5-氮-2’-脱氧胞苷(5-Aza-2’-deoxycytidine,5-Aza-CdR)对人卵巢癌细胞系SKOV3增殖、凋亡及错配修复基因HMLH1和HMSH2表达的影响。方法用0.5、5和50μmol/L特异性甲基转移酶抑制剂5-Aza-CdR处理SKOV3细胞3d,继续常规培养7d后,采用四唑盐(MTT)比色法检测生长活性,流式细胞术分析细胞的凋亡,半定量RT-PCR检测细胞HMLH1和HMSH2 mRNA的表达水平。结果人卵巢癌细胞系SKOV3经0.5、5和50μmol/L5-Aza-CdR处理后,均能明显抑制肿瘤细胞生长。各组细胞的凋亡率分别为10.59%±1.57%、17.52%±1.72%、34.10%±1.45%,显著高于对照组的5.35%±0.86%(P<0.01);且凋亡率与5-Aza-CdR剂量成正相关(F=227.6,P<0.01)。在SKOV3中HMLH1和HMSH2呈弱表达,经5-Aza-CdR处理后mRNA表达量有不同程度的增加,且与药物存在剂量依赖性。结论5-Aza-CdR可逆转卵巢癌细胞系中存在的HMLH1和HMSH2甲基化,DNA错配修复基因甲基化与卵巢癌的发生、发展有关。     

Human Leucocyte Migration Inhibitory Factor (LIF)

The cyclic nucleotides guanosine 3',5'-cyclic monophosphoric acid (3',5'-cGMP) and cytidine 2',3'-cyclic monophosphoric acid (2',3'-cCMP) but not cyclic phosphodiesters derived from the bases adenine and uracil preserved LIF activity against the blocking effect of the serine protease inhibitor phenylmethylsulphonyl fluoride (PMSF). Phosphomonoesters derived from guanosine and cytidine as well as 2',3'-cGMP and 3',5'-cCMP were all inactive, indicating specificity for phosphodiester bonds and their respective positions in the two active nucleotides. The protection afforded by 3',5'-cGMP and 2',3'-cCMP was dose dependent. Thus, using 10(-3) M PMSF, 3',5'-cGMP was active at concentrations higher than 10(-5) to 10(-4) M, and 2',3'-cCMP at concentrations higher than 3 X 10(-4) to 10(-3) M. The more pronounced LIF-inhibitory effect obtained by increased concentrations of PMSF could be overcome by raising the levels of the nucleotides, indicating that the interactions between PMSF and the nucleotides with LIF were mutally exclusive. The possibility that 3',5'-cGMP and perhaps 2',3'-cCMP function as modulators of LIF is discussed, and models for the function of this lymphokine are proposed.     

Modulation of histamine type II receptors on CD8+ T cells by interleukin-2 and cimetidine.

CD8+ T cells are known to play a major role in regulating immune functions under normal and disease conditions. In this study a radioligand binding assay was used to quantitate histamine type II (H2) receptors on activated T cells. The objective was to examine the expression of H2 receptors on T cells during activation with interleukin-2 (IL-2) and treatment with cimetidine. Activated suppressor T cells induced by concanavalin A+IL-2 showed a significant (p less than 0.01) increase in H2 receptors compared to the control nonactivated T cells. The T cells expressing the H2 receptors were identified as CD8+ cells; those among them that had an enhanced level of H2 receptors were identified as CD25+. Treatment of activated suppressor cells with the H2 receptor antagonist cimetidine at a concentration of 10(-5) M significantly reduced the number of H2 receptors. Suppressor cells induced by Con A+IL-2 were able to suppress both IgG and IgM production that was reversible with cimetidine. Incubation of lymphocytes with 50 U/ml IL-2 alone in 3-day culture significantly (p less than 0.005) enhanced H2 receptor expression. These studies demonstrate that activated suppressor T cells that are CD8+CD25+ have enhanced levels of H2 receptors and can be modulated by cimetidine.     

Growth of certain myeloid leukemic cells can be stimulated by interleukin-2.

The effect of recombinant interleukin-2 (IL-2) on the proliferation of T-cell depleted leukemic blasts was evaluated in 23 patients with acute myelogenous leukemia (AML). For this purpose, the effect of IL-2 on cell growth, [3H]-thymidine incorporation into the blasts and the expression of IL-2 receptors on cell surface using T-cell depleted blasts were studied. The results showed that IL-2 stimulated [3H]-thymidine incorporation significantly in blasts of 8 out of 23 cases of AML. An IL-2 induced increase in cell number was directly demonstrated in seven out of eight IL-2 responsive patients studied. IL-2 stimulated the proliferation of blasts in monocytic lineage (M4 and M5), but not all M4/M5 leukemics responded to rIL-2. Stimulation of the growth of leukemic cells was not correlated with the expression of Tac antigen on the cell surface, but it was significantly correlated with the expression of IL-2 receptor (IL-2R) beta chain on the cell surface. These results indicate that IL-2 is an active growth factor in certain myeloid leukemia cells, especially of monocytic type.     

Triptolide suppresses T-lymphocyte proliferation by inhibiting interleukin-2 receptor expression,but spares interleukin-2 production and mRNA expression

The purpose of this study was to elucidate the mechanism of action of triptolide on the T-lymphocyte-mediated immune response. Lymphocytes were incubated with a suboptimal dose oof Con A or PHA in the presence or absence of varying doses of triptolide to assess the effect of triptolide on lymphocyte proliferation, interleukin-2(IL-2) production and IL-2 receptor expression. Then, Con A or PHA induced T-blast cells were cultured with a sufficient dose of recombinant human IL-2 in the presence or absence of triptolide to evaluate the effect of triptolide on the interaction of IL-2 and IL-2 receptors. The effect of triptolide on the immune response in vivo was also investigated. The results of these studies clearly demonstrated that triptolide selectively inhibited the T-lymphocyte proliferative response to Con A and PHA, but had less effect on LPS-induced B-lymphocyte proliferation. Triptolide also suppressed the expression of IL-2 receptors on PHA induced T-blast cells, but did not alter the production of IL-2 by mouse splenic cells and human tonsil lymphocytes. Furthermore, the results also showed that triptolide at higher concentration had a slight inhibitory effect on the interaction of IL-2 and IL-2 receptors, and addition of exogenous IL-2 did not reverse the inhibiting action of triptolide on T-cell proliferation. Taken together, these results suggest that triptolide inhibits T-lymphocyte proliferation mainly by affecting IL-2 receptor expression rather than IL-2 production.     

Regulation of interleukin-2 receptor expression and receptor release

The generation and cell surface expression of IL-2 receptors was monitored by: (i) an ELISA that permits quantitative determination of detergent-solubilized or soluble IL-2 receptors; and (ii) detection of the binding of 125I-labelled recombinant IL-2 and of anti-IL-2 receptor antibodies to receptor bearing cells. Upon lectin stimulation both high and low affinity IL-2 receptors became expressed in parallel at the cell surface. Both high and low affinity receptors were upregulated by IL-2. Upon lectin activation the amount of cell-associated receptors increased and on day 2 of the culture period IL-2 receptors were detectable in the culture supernatant. IL-2 upregulated both high and low affinity IL-2R expression on T-lymphoblasts. IL-2R bearing leukemic cells and T lymphoblasts released IL-2R when cultured in vitro. IL-2R release by T lymphoblasts was enhanced dramatically by IL-2. On the other hand, IL-2-receptor positive leukemic cells released receptors in an IL-2 independent manner. Release of receptors could also be detected in serum-free medium. At least a part of the released receptors could be specifically bound to immobilized pure recombinant IL-2 and to monoclonal anti-IL-2-receptor antibodies. Small but significant amounts of soluble IL-2 receptors were detectable in the sera of normal mice. In sera of mice inoculated with IL-2-receptor positive syngeneic leukemic cells, elevated levels of IL-2 receptors were detectable. Release of IL-2 receptors seems to represent one of the major routes by which the receptors are cleared from the cell surface.     

Down-regulation of interleukin-2 receptor expression on natural killer cells/large granular lymphocytes by interleukin-4.

It has recently been demonstrated that IL-4 inhibits IL-2 receptor expression on T cells. Studies have also shown that IL-4 can inhibit IL-2-induced natural killer cell (NK) cytotoxicity, and that IL-4 in combination with IL-2 and large granular lymphocyte (NK/LGL) cells suppresses Ig synthesis. Therefore, we examine whether IL-2 receptor expression on NK/LGL cells is affected with or without IL-4, using fluorescent receptor analysis. Our results demonstrate that IL-4 inhibits/down-regulates the expression of IL-2 receptors on either phytohemagglutinin or IL-2-stimulated NK/LGL cells.     

Analysis of dynamics and functions of high-affinity interleukin-2 receptors

Activated T cells express at least two affinity classes of interleukin-2 (IL-2) receptors. The number of low-affinity receptors is normally 10–30 times greater than that of the high-affinity receptors. In this report, normal human T cells are used in a cellular system in which the number of low-affinity receptors can be manipulated. The resulting receptor composition, which has been characterized in a previous report, contain such decreased levels of low-affinity IL-2 receptors that almost half of the surface pool of anti-IL-2 receptor antibody (anti-Tac) binding sites is associated with high-affinity receptors. By using such cells the dynamics and functions of high-affinity IL-2 receptors were studied and compared with receptors on a cell population expressing the normal 10–30-fold excess of anti-Tac binding sites over high-affinity IL-2 receptors. The results reveal that the rapid turnover of high-affinity IL-2 receptors is independent of the quantitative level of Tac antigen expression. The rapid kinetic of IL-2 internalization results in a 80–90% reduction of the steady-state levels of high-affinity receptors in the presence of IL-2. Most importantly, by using a cell population that expresses very low levels of Tac antigens, it became evident that IL-2 internalization is associated with an immediate substantial decrease of the surface level of anti-Tac binding sites. The Tac antigen thus appeared to be internalized together with the high-affinity IL-2 receptor complex but nevertheless the normal 10–30-fold excess of Tac antigens, over high-affinity IL-2 receptors, seems not to influence the process of internalization.     

Investigations on the interaction of tartaric acid derivative/human serum albumin tissue adhesive with J774A.1 mouse macrophage cells through SEM, IL-6 cytokine and gene expression techniques

We developed a novel tissue adhesive consisting of human serum albumin (HSA) and tartaric acid derivative (TAD). Four different concentrations of TAD namely, 0.05 mM, 0.1 mM, 0.2 mM and 0.3 mM were mixed with 40%, 42% and 44% HSA individually and were made in the form of disks. J774A.1 mouse macrophage cells were seeded on top of these disks. The disks were pre-treated with sterile water and Eagle's medium before every seeding. All the seeding was incubated from 1 day to 3 days before making any investigations on it. SEM images were recorded and it was observed that these cells adhered to these materials very well. Mouse IL-6 cytokine expressions were studied using ELISA. It was seen from the cytokine expression results that the release of IL-6 was minimum at 0.3 mM TAD concentrations with 44% HSA disks. No significant difference was observed in the cytokine expressions of IL-6 at 42% and 44% HSA at all concentrations of TAD studied in this work. mRNA gene expressions of IL-6 were investigated using RT-PCR technique. In 40% HSA, the gene expression level of IL-6 gene did not change during 3-day-culture in the range of TAD concentration of 0.05 mmol to 0.2 mmol. However, 0.3 mM TAD suppressed the gene expression at all concentration of HSA. In 42% HSA, although 0.05 mM and 0.1 mM TAD did not affect the gene expression, 0.2 mM and 0.3 mM TAD induced the expression level with incubation time. In 44% HSA, all the concentration of TAD increased the expression level even though the cytokine expression levels were quite low. Hence it could be thought that the expression at the cytokine level is quite insignificant where as it is to be considered at the gene expression level. On the whole, 0.3 mM TAD with 44% HSA could be considered as a challenging material as a tissue adhesive material for use in the field of tissue engineering.