粘附分子对CD3介导的胸腺细胞[Ca^2＋]i应答的影响

ＬＦＡ－１和ＩＣＡＭ－１广泛表达于各胸腺细胞亚群，但ＩＣＡＭ－１在ＰＮＡ￣＋细胞的表达下调。本文报道：用抗ＬＦＡ－１／ＩＣＡＭ－１和抗ＣＤ３单抗，分析了粘附分子ＬＦＡ－１／ＩＣＡＭ－１对抗ＣＤ３诱导的胸腺细胞［Ｃａ￣（２＋）］ｉ应答的影响。结果显示，可溶性抗ＬＦＡ－１／ＩＣＡＭ－１可抑制ＣｏｎＡ刺激的胸腺细胞增殖，且以抗ＬＦＡ－１抗体的作用更为显著，在ＣｏｎＡ或抗ＣＤ３诱导的胸腺细胞［Ｃａ￣（２＋）］ｉ应答中，抗ＬＦＡ－１单抗可明显抑制［Ｃａ￣（２＋）ｉ升高。但如果用二抗交联ＣＤ３和ＬＦＡ－１，胸腺细胞［Ｃａ￣（２＋）ｉ则显著高于单独交联ＣＤ３时的水平（Ｐ＜０．０１），而ＣＤ３与ＩＣＡＭ－ｌ交联却无此效应，此外，仅交联ＬＦＡ－１或ＩＣＡＭ－１也无诱导［Ｃａ￣（２＋）］ｉ应答的作用。提示在ＬＦＡ－ｌ与ＩＣＡＭ－１介导的胸腺细胞与胸腺基质细胞相互作用中，ＬＦＡ－１可为ＴＣＲ／ＣＤ３途径介导的胸腺细胞活化提供复合刺激信号。     

枸杞多糖2对辐射损伤小鼠免疫功能恢复的影响

从枸杞子中提取的枸杞多糖（ｌｙｃｉｕｍｂａｒｂａｒｕｍｐｏｌｙｓａｃｃｈａｒｉｄｅ，ＬＢＰ）有较好的免疫增强作用，可促进Ｔ、Ｂ淋巴细胞的功能，增强机体免疫监视功能以及降低抗肿瘤化疗药物引起的免疫抑制等作用。本文研究了ＬＢＰ２对辐射损伤小鼠免疫功能恢复的影响，ＬＢＰ２能明显促进辐射损伤小鼠免疫功能的恢复，照射后３０ｄ，胸腺指数、脾细胞对ＣｏｎＡ、ＬＰＳ的增殖反应、ＭＬＲ、ＤＴＨ及ＰＦＣ均较照射对照组明显增强。     

内皮素对成年鼠离体心室肌细胞内游离钙浓度的影响及机制

目的和方法：采用分离的Ｓｐｒａｇｕｅ－Ｄａｗｌｅｙ大鼠心室肌细胞,以Ｆｕｒａ－２／ＡＭ荧光指示剂负载,检测心肌细胞内游离钙浓度（［Ｃａ２＋］ｉ）变化,探讨内皮素－１（ＥＴ－１）对［Ｃａ２＋］ｉ的作用及其机制。结果：ＥＴ－１（１×１０－７ｍｏｌ／Ｌ）引起［Ｃａ２＋］ｉ升高分两个时相：快速相和持续相,可被ＥＴＡ的特异性受体阻断剂ＢＱ１２３（２×１０－６ｍｏｌ／Ｌ）所阻断。移去细胞外液钙以及用百日咳毒素（２００ｎｇ／ｍＬ）处理１０ｈ后,ＥＴ－１仍引起快速相,但持续相消失。Ｒｙａｎｏｄｉｎｅ（４μｍｏｌ／Ｌ）和异搏定（２×１０－５ｍｏｌ／Ｌ）对ＥＴ－１诱导［Ｃａ２＋］ｉ升高的作用无显著影响。结论：ＥＴ－１升高［Ｃａ２＋］ｉ是通过ＥＴＡ受体介导;快速相［Ｃａ２＋］ｉ升高主要由胞内Ｒｙａｎｏｄｉｎｅ不敏感的钙池释放造成,与百日咳毒素敏感的Ｇ蛋白无关;持续相［Ｃａ２＋］ｉ升高主要由胞外Ｃａ２＋内流引起,不是通过电压依赖性Ｌ－型钙通道介导,与百日咳毒素敏感的Ｇ蛋白有关     

长春新碱诱导的自噬性凋亡与细胞内游离Ca^2＋浓度的相关性研究

目的 研究长春新碱（ＶＣＲ）诱导的Ｌ－０２细胞自噬性凋亡细胞内游离钙离子浓度（［Ｃａ＾２＋］ｉ）的变化,以及自噬特异性抑制剂３－ｍｅｔｈｙｌａｄｅｎｉｎｅ（３ＭＡ）对此自噬性凋亡和［Ｃａ＾２＋］ｉ的影响。方法 应用已建立的ＶＣＲ诱导的Ｌ－０２细胞自噬性凋亡模型,使用电镜、流式细胞术检测细胞;用Ｆｌｕｏ－３／ＡＭ荧光探针经流式细胞仪测定Ｌ－０２细胞平均［Ｃａ＾２＋］ｉ。结果 电镜及流式细胞术检测证实     

尼可地尔降低血管平滑肌细胞内ATP诱导的游离钙浓度

目的：探讨尼可地尔对血管平滑肌细胞内游离钙（［Ｃａ２＋］ｉ）的影响及机理。方法：培养的兔主动脉平滑肌细胞加入Ｆｕｒａ－２ＡＭ２５μｍｏｌ／Ｌ,在３７℃下孵育５０ｍｉｎ,［Ｃａ２＋］ｉ用荧光分光光度计检测。结果：ＡＴＰ（０１ｍｍｏｌ／Ｌ）诱导的［Ｃａ２＋］ｉ峰相和持续相增加可被尼可地尔抑制,且呈剂量依赖性,尼可地尔（１０μｍｏｌ／Ｌ）的抑制作用可被优降糖（１０μｍｏｌ／Ｌ）完全阻断（峰相：５３０±３１ｖｓ５４４±４１ｎｍｏｌ／Ｌ,持续相：３７０±１９ｖｓ３８１±１１ｎｍｏｌ／Ｌ,Ｐ＞００５）;在无钙溶液中,先给尼可地尔能显著抑制ＡＴＰ诱导的［Ｃａ２＋］ｉ峰相增加。结论：尼可地尔抑制ＡＴＰ诱导的［Ｃａ２＋］ｉ增加,可能与减少细胞外钙内流及细胞内钙释放有关。     

α-MSH对EGTA发热的作用

目的：观察α－ＭＳＨ对ＥＧＴＡ发热反应的作用及其可能机制。方法：建立ＥＧＴＡ发热模型；在离体条件下，应用Ｆｕｒａ－２荧光指示剂测定细胞内Ｃａ２＋浓度（［Ｃａ２＋］ｉ）；体外培养下丘脑神经细胞。结果：α－ＭＳＨ能明显抑制ＥＧＴＡ性发热反应（Ｐ＜０．０１）；ＥＧＴＡ可以降低下丘脑神经细胞［Ｃａ２＋］ｉ水平（Ｐ＜０．０１），但α－ＭＳＨ不影响正常下丘脑神经细胞［Ｃａ２＋］ｉ及ＥＧＴＡ对下丘脑神经细胞［Ｃａ２＋］ｉ的作用（Ｐ＞０．０５）；ＥＧＴＡ可刺激体外培养的下丘脑神经细胞释放ＣＲＨ（Ｐ＜０．０５），而α－ＭＳＨ能抑制ＥＧＴＡ的这种作用（Ｐ＜０．０５）。结论：α－ＭＳＨ抑制中枢发热介质ＣＲＨ的产生可能是降低ＥＧＴＡ发热反应的主要机制之一；中枢ＣＲＨ的产生和释放增加可能是ＥＧＴＡ性发热的一个重要因素。     

乙酰胆碱对大鼠腹腔巨噬细胞内钙离子浓度和表面Ia抗原表达的影响

用Ｆｕｒａ－２作为荧光探针测定大鼠腹腔巨噬细胞（Ｍ）内钙离子浓度（［Ｃａ￣（２＋）］ｉ），ＡＰＡＡＰ桥联酶标法检测Ｍ表面Ｉａ抗原的表达。结果表明：５×１０￣（－６）ｍｏｌ／Ｌ的乙酞胆碱（Ａｃｈ）可使Ｍ［Ｃａ￣（２＋）］ｉ；明显上升，可促进Ｍ表面Ｉ－Ａ和Ｉ－Ｅ抗原的表达，而阿托品（１０￣（－５）ｍｏｌ／Ｌ）可阻断Ａｃｈ升高［Ｃａ￣（２＋）］ｉ的作用。阿托品、三氟啦嚎（ＴＦＰ，５０μｍｏｌ／Ｌ）、ＥＧＴＡ（６ｍｍｏｌ／Ｌ）均可阻断Ｍ促进ＭＩａ抗原表达的作用，ｃＡＭＰ依赖性蛋白激酶抑制剂（ＰＫＩ，２５μｇ／ｍｌ）对Ａｃｈ促进ＭＩａ抗原表达的作用无影响。     

SRBC膜提取物对猪PBMNC第二信使的影响

胰酶水解绵羊红细胞（ＳＲＢＣ）释放膜表面活性蛋白组分Ⅲ（ＴＲＦ－Ⅲ），单独作用可使猪外周血单个核细胞（ＰＢＭＮＣ）胞内ｃＡＭＰ水平升高，以１００μｇ／ｍｌ浓度刺激达最高峰，由对照组０．９４±０．１４ｐｍｏｌ／Ｌ升高到２．７５±０．２５ｐｍｏｌ／Ｌ（Ｐ＜０．０１）。如果ＰＢＭＮＣ事先与腺苷酸环化酶（ＡＣａｓｅ）抑制剂ＬｉC１孵育后再以ＴＲＦ－Ⅲ或ＰＡＨ刺激。ｃＡＭＰ增高受到抑制（Ｐ＜０．０１）；而ＥＤＴＡ－２Ｎａ（一种磷酸二酯酶ＰＤＥ抑制剂）对此ｃＡＭＰ升高无影响。结果提示，此ｃＡＭＰ升高主要是通过活化ＡＣａｓｅ水解ＡＴＰ生成ｃＡＭＰ，而不像是抑制ＰＤＥ减少ｃＡＭＰ降解引起的。ＴＲＦ－Ⅲ诱导猪ＰＢＭＮＣ胞内Ｃａ~(２＋)浓度升高，以１００μｇ／ｍｌ刺激２分钟升高最多，由对照组的２４２±７ｎｍｏｌ／Ｌ升高到３２３±１５ｎｍｏｌ／Ｌ（Ｐ＜０．０１）。以ＥＧ－ＴＡ除去胞外Ｃａ~(２＋)再以ＴＲＦ－Ⅲ或ＰＡＨ刺激，仅观察到小范围［Ｃａ~(２＋)］ｉ升高。看来这一过程包括了刺激胞内Ｃａ~(２＋)释放和胞外Ｃａ~(２＋)内流两种方式。以上结果证明，ＴＲＦ－Ⅲ对淋巴细胞功能影响与细胞内第二信使有关。     

缺氧对培养的猪肺内皮细胞胞浆游离钙的影响

本文应用荧光探针Ｆｕｒａ－１／ＡＭ测定胞浆游离钙浓度（［Ｃａ＾２＋］ｉ）技术，观察培养的猪肺动脉内皮细胞［Ｃａ＾２＋］ｉ在缺氧时变化。实验发现：向细胞悬液中充氮气造成缺氧时，肺动脉内皮细胞［Ｃａ＾２＋］ｉ升高８１±２１％（Ｐ＜０．０５，ｎ＝８）。结果提示肺动脉内皮细胞钙信使系统可能参与缺氧所致血管反应。     

多抗甲素与IL—2协同促进骨髓移植小鼠免疫功能重建

目的：为了探讨多抗甲素ＰＡＡ与ＩＬ－２对骨髓移植（ＢＭＴ）小鼠免疫功能重建的影响。方法：用致死量照射的Ｂａｂｌ／ｃ小鼠移植同系小鼠骨髓细胞后,分别给予ＰＡＡ、ＩＬ－２或ＰＡＡ＋ＩＬ－２,３０ｄ后检查３组ＢＭＴ小鼠免疫功能重建的情况。结果：ＢＭＴ后３０ｄ,自然恢复小鼠免疫功能十分低下,但ＰＡＡ及ＩＬ－２用药鼠脾细胞对丝裂原反应,对ＳＲＢＣ特异的ＰＦＣ数,对同种异型小鼠脾细胞ＤＴＨ及ＭＬＲ均明显高于移     

驱虫斑鸠菊注射液对小鼠免疫功能的影响

目的:探讨驱虫斑鸠菊对小鼠免疫功能的影响。方法:采用[3H]-TdR掺入法和脾细胞介导羊红细胞定量溶血分光光度法以及迟发型超敏反应试验等观察了驱虫斑鸠菊对小鼠兔疫功能的作用。结果:驱虫斑鸠菊在体内外均可以明显抑制ConA刺激的小鼠T淋巴细胞的增殖反应和LPS刺激的小鼠B淋巴细胞的增殖反应（P<0.01）;对小鼠胸腺指数和脾脏指数、绵羊红细胞（SRBC）诱导的正常小鼠脾抗体细胞生成反应以及小鼠皮肤迟发型超敏反应都显示出明显的抑制作用,而且上述抑制作用与药物浓度有一定的剂量效应关系。结论:驱虫斑鸠菊对机体体液免疫、细胞免疫都有明显的抑制作用。     

转录因子GATA-3 mRNA在哮喘模型小鼠体内的表达

目的:探讨转录因子GATA-3 mRNA在哮喘模型小鼠体内的表达。 方法:建立卵白蛋白（OVA）致小鼠哮喘模型，计数支气管肺泡灌洗液（BALF）中炎症细胞总数和分类，评价肺组织炎症细胞浸润；ELISA检测BALF和脾细胞培养上清液中IL-4和IFN-γ浓度；RT-PCR检测脾细胞和肺组织GATA-3 mRNA表达水平。 结果:哮喘组BALF中炎症细胞总数及嗜酸粒细胞百分比明显高于对照组，支气管出现明显炎症细胞浸润、黏液分泌和支气管收缩，BALF和脾细胞培养上清液中IL-4明显高于对照组，肺组织和脾细胞GATA-3 mRNA表达水平均明显高于对照组。 结论:哮喘模型小鼠肺组织和脾细胞GATA-3 mRNA表达增加，可能在促进Th2细胞因子合成和介导气道炎症细胞浸润中具有重要作用。     

B cell stimulating activity of metallothionein in vitro

The effect of metallothionein (MT) on lymphocytes was studied in vitro. Rabbit MT induced the proliferative responses of mouse splenocytes at concentrations of 1-25 microg/ml. MT synergistically enhanced Con A- or LPS-induced proliferative response of splenocytes. A similar effect was also observed for rabbit splenocytes at a similar concentration. Free heavy metals such as Cd and Zn only weakly stimulated splenocytes. 2-mercaptoethanol (2-Me) abrogated the effect of MT, suggesting that thiols in MT play an important role for splenocyte response. MT stimulated splenocytes from MT-knockout mice as well as those from normal control mouse. The responder cells for MT stimulation were B cells and MT also induced B cell differentiation to produce Ig. MT induced the calcium influx of B cells, but not T cells. These results indicate that MT has a potent immunomodulating activity, especially on B cells.     

胚芽乳杆菌胞壁肽聚糖对细胞免疫的作用

用体外刺激、~8H-TdR掺入测定淋巴细胞增殖的方法观察到胚芽乳杆菌的胞壁肽聚糖(PG)可以直接诱导小鼠脾淋巴细胞的增殖反应,在所采用剂量范围内细胞增殖的倍数与PG剂量呈线性关系;但对胸腺细胞和分纯的脾T淋巴细胞则无直接作用。还观察到PG有协同ConA诱导小鼠脾细胞和胸腺细胞增殖作用。可见PG具有B细胞的丝裂原性,并有增强T细胞丝裂原的作用。     

酶联免疫斑点方法改进及应用

建立了ELISPOT方法并通过改进 ,应用到抗人CD8杂交瘤细胞及LACA小鼠体外脾细胞分泌IgG量的检测 ,同时观察六味地黄多糖CA4 3在体外对LACA小鼠脾细胞分泌IgG的影响。结果表明 10 μg/mlLPS作用下可明显提高小鼠脾细胞体外产生抗体的量和提高分泌抗体的细胞数。 5 0 μg/mlCA4 3在体外与LPS具有相似的作用 ,可以刺激小鼠脾细胞分泌IgG的量和分泌抗体的细胞数量增多。     

Changes of natural killer cell activity in different mouse lines by acute and asymptomatic flavivirus infections

Effect of certain flaviviruses on the activity of mouse natural killer (NK) cells was investigated using the classical mouse splenocyte system and YAC-1 cells for demonstration of NK cell cytotoxicity. Infection of mice with Langat and West Nile (WN) viruses was accompanied by temporary activation of NK cells. In mice infected with tick-borne encephalitis (TBE) virus the stimulation phase of NK cell cytotoxicity on days 2-4 post-infection (p.i.) was followed by suppression of their activity. As to the surface markers (sensitivity to antitheta and antiimmunoglobulin serum, respectively), the flavivirus-activated NK cells did not differ from the endogenous NK cells of intact mice. The stimulatory effect of flaviviruses on cytotoxicity of NK cells varied in different mouse lines. An increased NK cell activity at early stages of TBE virus infection was observed in mouse lines characterized by low (C57B1/6) and medium (BALB/c)--but not by high (CBA)-activity of their non-stimulated NK cells. Suppression of NK cell activity at later stages of TBE virus infection was not associated with virus multiplication in mouse splenocytes.     

Interleukin-12 is critical for induction of nitric oxide-mediated immunosuppression following vaccination of mice with attenuated Salmonella typhimurium.

Studies from our laboratory have shown that infection of mice with an attenuated strain of Salmonella typhimurium causes a marked suppression in the capacity of splenocytes to generate an in vitro plaque-forming cell (PFC) response to sheep erythrocytes. The suppression has been shown to be mediated by mature, adherent macrophages (Mphis) and nonadherent, precursor Mphis. Nitric oxide has been identified as the suppressor factor. The present study investigated the role of interleukin-12 (IL-12) in the generation of nitric oxide-mediated immunosuppression in this model. Salmonella inoculation resulted in marked suppression of PFC responses and high levels of nitrite production. When mice were treated with anti-IL-12 prior to inoculation, nitrite levels in splenocyte cultures were reduced by 75% and the suppression of PFC responses was prevented. The nonadherent splenocyte fraction from Salmonella-inoculated mice, which contains precursor Mphis and is weakly immunosuppressive, was treated with IL-12 in vitro. IL-12 augmented the capacity of this fraction to suppress PFC responses by normal splenocytes in a coculture system. Additionally, IL-12 induced nitrite and gamma interferon (IFN-gamma) production in a dose-dependent manner. Treatment with anti-IFN-gamma blocked nitrite production and suppression, indicating that IFN-gamma is an important intermediary in the pathway of IL-12-induced immunosuppression. These results indicate that IL-12 is critical for the induction of nitric oxide-mediated immunosuppression following S. typhimurium inoculation and, through its ability to stimulate IFN-gamma production, can induce nitric oxide-producing suppressor Mphis.     

IgM production of lymphocytes from C57BL/6N mice was stimulated by estrogen treated splenic adherent cells

Estrogens have diverse effects on cell growth, differentiation and homeostatic functions, and have been shown to play an important role in regulating immune system. In this study, we examined the effect of 17beta-estradiol (E2) on antibody production by splenocytes isolated from C57BL/6N mice. Our results suggest that the activation of immunoglobulin (Ig) M production by E2 requires direct cell-cell interaction between adherent and non-adherent cells in mouse splenocyte population, and the primary target of E2 is adherent cell population. In addition, we indicated that ER antagonist ICI 182780 suppressed this enhancing effect of E2. Both ERalpha agonist and ERalpha agonist enhanced IgM production of mouse splenocytes. ERs are expressed on plasma membrane as well as in nucleus. However, a plasma membrane-associated ER specific ligand has no stimulation effect on IgM production. In conclusion, our results indicate that adherent cells stimulated by E2 up-regulate IgM production of lymphocytes through the direct cell-cell interactions, and the enhancing effect of E2 is arouse through ERalpha and ERbeta on these cells.     

Ly-49D transfected NK cells show reduced interleukin-10 production in response to H-2d and increased lytic activity: implication by interaction using class I MHC alloantigen+ target cells in pregnancy

PROBLEM: Mating of CBA/J (H-2k) with DBA/2 (H-2d) males leads to a high rate of spontaneous resorption (about 40%), which is not seen in other mating combinations, such as CBA/J X BALB/c. The activation of natural killer cells (NK cells) seems to be a key mechanism for the maternal-fetal intolerance in allogeneic pregnancy, and recurrent spontaneous abortion. The effect of expression of the NK cell activating receptor Ly49D recognizing BALB/c or DBA/2 class I MHC was investigated. METHOD OF STUDY: Intracellular interleukin (IL)-100 production was detected and target cell survival rates were calculated after 22 hr coincubation of rat NK cells transfected, or not. with a murine Ly49D receptor, with either male BALB/c or male DBA/2 splenocytes, by using flow cytometry. RESULTS: Ly49D negative rat NK cells produced 13.7% more IL-10 than Ly49D positive rat NK cells, and more splenocytes were killed by Ly49D transfected rat NK cells (survival rate 2.45%) than by Ly49D negative rat NK cells (survival rate 4.36%). CONCLUSION: After physiological stimulation with BALB/c or DBA/2 splenocytes, rat NK cells are able to synthesize IL-10. Recognition of mouse splenocyte major histocompatibility complex (MHC) by Ly49D mice receptor decreased IL-10 production. The observed increase in killing activity might be a result of this phenomenon. NK cell activation via the Ly49D receptor might play an important role in pregnancy failure, but cannot explain why CBA/J X DBA/2 matings are abortion prone, and CBA/J X BALB/c matings are abortion resistant.