放线菌酮诱导大鼠脾细胞凋亡的电镜观察

利用透射电镜观察了放线菌酮体内诱导大鼠脾细胞凋亡的形态学变化，结果显示，腹腔注射放线菌酮４小时后，大鼠脾细胞发生凋亡，凋亡脾细胞核和胞质发生一系列形态学变化。主要表现为胞核染色质浓缩，重排，呈不同形态，多数凋亡脾细胞的粗面内质网增殖，凋亡小体形成，其线粒体也大量增加，或单个分布或团聚，肿胀，空泡化。结果提示，凋亡脾细胞的主要细胞器主动参与凋亡过程。     

放线菌酮诱导大鼠胸腺细胞凋亡的电镜观察

利用透射电镜详细观察了放线菌酮体内诱导大鼠腺细胞凋亡的形态学变化。观察显示，腹腔注射放线菌酮４小时后，大鼠胸腺细胞发生凋亡，凋亡胸腺细胞胞核和胞质发生一系列形态学变化，产生凋亡小体。主要表现为染色质断裂、浓缩、边集、大部分细胞核变成花瓣状，其它细胞核变成半月状、黑洞样和空泡状；粗面内质网大量增殖，并包裹细胞成分形成自噬体；线粒体增多，嵴紊乱并空泡化。凋亡细胞及其形成的凋亡小体被其它细胞吞噬清除。结     

放线菌酮诱导大鼠脾细胞凋亡的电镜观察

利用透射电镜观察了放线菌酮体内诱导大鼠脾细胞凋亡的形态学变化,结果显示,腹腔注射放线菌酮4小时后,大鼠脾细胞发生凋亡,凋亡脾细胞核和胞质发生一系列形态学变化.主要表现为胞核染色质浓缩、重排、呈不同形态;多数凋亡脾细胞的粗面内质网增殖,凋亡小体形成,其线粒体也大量增加,或单个分布或团聚,肿胀、空泡化.结果提示,凋亡脾细胞的主要细胞器主动参与凋亡过程.     

肝细胞癌凋亡病变的原位末端记显示

原位末端标记（ＩＳＥＬ）是显示凋亡的新技术，按Ｇａｖｒｉｅｌｉ法对４６例未接受化疗与放疗的人肝细胞癌（ＨＣＣ）作回顾性（ＩＳＥＬ）研究，结果显示的对照纱列中中阴性与阳性均与理论相符，４６例ＨＣＣＩＳＥＬ（＋）可分为：（１?单个细胞（＋），（２）点状（＋）；（３）小片状（＋）；（４）大片（＋）。Ｉ级ＨＣＣ以单细胞（＋）较多；Ⅱ，Ⅲ级惟小片状（＋）较多（Ｐ〈０．０５）。单个细胞（＋）凋亡经典概念，而片     

生后大鼠发育不同时期各级卵泡凋亡的观察

目的：观察研究生后不同阶段大鼠卵巢卵泡的生长发育与卵泡凋亡的关系、凋亡过程及规律，探索性激素血液浓度变化与卵泡凋亡的内在联系。方法：用DNA缺口原位末端标记检测法，对生后20-140天不同阶段大鼠卵巢卵泡的生长发育及卵泡凋亡进行了光镜观察，同时对生后各阶段鼠血液孕激素(progesterone P）,卵泡刺激素（follicle stimulating hormone FSH)、黄体生成素(luteinizing lormone LH)、雌二醇(estradiol E2)浓度进行放免检测。结果：生后20天鼠卵泡即可见到凋亡发生，卵泡凋亡始于卵母细胞，继则是与其相邻的卵泡细胞，卵泡细胞的凋亡从内向外依次发生。卵泡的凋亡数目随着卵巢的生长发育而逐渐增多，生后40天可见到黄体细胞凋亡发生，至50-60天可见到卵泡膜内膜细胞凋亡及白体形成，出生80天后卵巢发育已完全成熟，此时卵泡的凋亡数也达到了高峰；放免结果经方差分析显示孕激素P＞0．05，差异显著，推测其浓度变化与卵泡凋亡有内在联系。     

前列腺癌雄激素受体表达与细胞凋亡的关系

目的：探讨前列腺癌雄激素受体与细胞凋亡的关系及其生物学特性,方法：56例前列腺癌和20例良性前列腺增生症,应用免疫组化方法检测石蜡蜡包埋的组织切片中雄激素受体（AR）表达和应用原位末端标记技术（TUNEL）检测细胞凋亡指数（AI）。结果：AR在良恶性前列腺组织中的表达差异无显著性（P＞0．05）。AR和AI与前列腺癌Gleason分级无关。AI在前列腺癌组织中的表达明显高于良性前列腺增生症（P＜0．05）,AR阳性的前列腺癌AI亦明显高于AR阴性的前列腺癌。结论：前列腺癌组织中AR的表达可诱导其细胞凋亡,并对前列腺癌的功能性分类和内分泌治疗的疗效判断具有实用的临床意义。     

双标记方法检测骨肉瘤细胞增生和细胞凋亡

本研究应用ＡＰＡＡＰ和原位末端标记（ｉｎｓｉｔｕｅｎｄｌａｂｅｌｉｎｇ，ＩＳＥＬ）双标记方法，检测了８０例骨肉瘤细胞的增生和细胞凋亡，旨在建立一种同时显示细胞增生和细胞凋亡的方法，便于病理医生更好地观察肿瘤细胞增生和细胞凋亡的关系。     

双标忆方法检测骨肉瘤细胞增生和细胞凋亡

本研究应用ＡＰＡＡＰ和原位末端标记（ＩＳＥＬ）双标记方法，检测了８０例骨肉瘤细胞的增生和细胞凋亡，旨在建立一种同时显示细胞增生和细胞凋亡的方法，便于病理医生更好地观察肿瘤细胞增生和细胞凋亡的关系。     

金黄色葡萄球菌及其L型诱导血管内皮细胞凋亡的研究

金黄色葡萄球菌 (简称金葡菌 )是临床上常见的主要致病菌之一 ,金葡菌感染可以诱发宿主细胞凋亡。我们用原位末端标记法 (TUNEL)、流式细胞术检测金葡菌及其L型诱导体外培养的人脐静脉内皮细胞 (HUVECs)凋亡情况。将处于指数生长期的HUVECs中陈旧培养液弃去 ,加入 1.8ml营养液和 0 .2ml 10 8CFU ml作为诱导因素的金葡菌(标准菌株 :CMCC2 60 75株 )及其L型进行实验 ,同时设立生理盐水对照 ,各组于诱导因素加入后 2、4、6、8及 10h分别收集 1次细胞进行检测。TUNEL染色法示金葡菌及其L型均能诱导HUVECs发生凋亡 ;流式细胞仪 (F…     

早期凋亡的T淋巴细胞诱导生成耐受性树突状细胞的实验研究

目的： 探讨早期凋亡T淋巴细胞的抑制性免疫调节性能。方法： 采用定时紫外线照射诱导T淋巴细胞早期凋亡，深低温反复冻融获得坏死T淋巴细胞。体外诱导、纯化并培养骨髓源性不成熟树突状细胞（imDCs）,imDCs分别和早期凋亡或坏死T淋巴细胞共培养。用流式细胞仪、双夹心ELISA、［³H］掺入混合淋巴细胞反应等方法分析imDCs吞噬早期凋亡或坏死T淋巴细胞后，在不同处理条件下，MHC-Ⅱ、CD40、CD80、CD86的表达水平、分泌IL-12 p70以及刺激T淋巴细胞增殖能力的差异。结果： imDCs和坏死细胞碎片共培养后明显趋于成熟，其MHCⅡ和CD40、CD80、CD86的表达水平显著上调；分泌较高水平的IL-12 p70；和同种异体处女T淋巴细胞混合培养后显著刺激处女T淋巴细胞增殖。 imDCs和早期凋亡的T淋巴细胞共培养后,其MHC-Ⅱ、CD40、CD80和CD86的表达维持较低水平；仅分泌较低水平IL-12 p70；和同种异体处女T淋巴细胞混合培养后不能刺激淋巴细胞增殖。此外，早期凋亡的T淋巴细胞孵育上清显著抑制了吞噬坏死细胞碎片后的imDCs表达共刺激分子CD40、CD80、CD86。当TGFβ₁中和抗体和早期凋亡T淋巴细胞同加入imDCs，在表达MHC-Ⅱ、CD40、CD80、CD86，分泌IL-12 p70，刺激处女T淋巴细胞增殖等方面和吞噬坏死T淋巴细的DCs相比无显著差异。结论： 早期凋亡T淋巴细胞通过释放免疫抑制性细胞因子TGFβ₁，诱导imDCs呈现出耐受性DCs（TolDCs）的免疫表型及生物学特征，从而发挥抑制性免疫调节作用。     

肝细胞癌凋亡病变的原位末端标记显示

原位末端标记（ＩＳＥＬ）是显示凋亡的新技术。按Ｇａｖｒｉｅｌｉ法对４６例未接受过化疗与放疗的人肝细胞癌（ＨＣＣ）作回顾性（ＩＳＥＬ）研究，结果显示在对照系列中阴性与阳性均与理论相符，４６例ＨＣＣＩＳＥＬ（＋）可分为：①单个细胞（＋）；②点状（＋）；③小片状（＋）；④大片（＋）．Ⅰ级ＨＣＣ以单细胞（＋）较多；Ⅱ、Ⅲ级以小片状（＋）较多（Ｐ＜０．０５）．单个细胞（＋）为凋亡经典概念，而片状（＋）报道还不多。本文小片状（＋）很普遍，常位于癌巢中央，其ＨＥ形态以核浓缩为主，此结果表明，通常ＨＣＣ所见小片状＂变性坏死样改变＂可能为凋亡而非坏死。     

组织细胞性坏死性淋巴结炎的电镜和原位末端标记观察

目的 从分子生物学角度,探讨组织性坏死性淋巴结炎的组织发生,提高认识,避免误诊。方法 对34例组织细胞性坏死性淋巴结炎标本行HE和原位末端标记TUNEL染色,光镜观察,其中4例做电镜检查。结果 显示多样的组织细胞主要为T淋巴细胞,以T淋巴细胞大量增生和凋亡为其特征。结论 必须抓住组织细胞性坏死性淋巴结炎的T细胞大量增生和凋亡的形态特征,才能确诊。     

Apoptosis in the human liver during allograft rejection and end-stage liver disease

The contribution of apoptosis (programmed cell death) to cellular damage in human liver disease is unknown. Using the in situ DNA end labelling method (ISEL), evidence was sought of programmed cell death (PCD) in liver tissue from patients with various liver diseases. In particular, the study aimed to determine whether PCD is involved in either the loss of interlobular bile ducts (vanishing bile duct syndrome—VBDS) or the perivenular hepatocyte drop-out, both of which are characteristic of irreversible graft rejection. Large numbers of apoptotic hepatocytes were found in pervenular areas in tissues taken from patients with chronic graft rejection. Significant hepatocyte apoptosis, was not seen in long-term stable allografts, primary biliary cirrhosis, cholestasis, paracetamol-induced fulminant hepatic failure, or fulminant hepatic failure of indeterminate origin (non-A, non-B, non-C hepatitis). Bile ducts rarely stained positively, but mononuclear cells present in the post-transplant tissues were frequently positive, showing nuclear or cytoplasmic staining. The presence of cytoplasmic staining suggested that some mononuclear cells had ingested apoptotic DNA from other cellular sources. PCD may thus contribute to the perivenular hepatocyte loss in chronic rejection. The absence of ductular epithelial cell staining suggests that PCD is not involved significantly in the bile duct loss of VBDS. Furthermore, apoptosis of monomuclear cells implies that PCD may be involved in regulating the inflammatory cell infiltration of graft rejection.     

狼疮性BXSB小鼠脾脏淋巴细胞增殖与凋亡的初步分析

为了比较全面准确地了解系统性红斑狼疮 (SLE )BXSB小鼠的发病过程中 ,淋巴细胞增殖与凋亡的动力学变化及其机制。采用细胞双色荧光染色的标记技术 ,检测了脾脏淋巴细胞中的增殖细胞和凋亡细胞的百分率 ,并且测定了巨噬细胞吞噬凋亡细胞的能力。结果发现 ,发病的雄性BXSB小鼠和雌性BXSB小鼠脾脏中增殖的CD4 + T淋巴细胞和B淋巴细胞百分率显著高于对照C5 7小鼠 ,而凋亡的B淋巴细胞的百分率显著低于对照C5 7小鼠 ;但是 ,雌雄BXSB小鼠和对照C5 7小鼠巨噬细胞吞噬凋亡细胞的吞噬指数相同。本研究结果表明 ,在BXSB小鼠的SLE发病过程中 ,淋巴细胞的增殖速度异常升高、而凋亡速度下降 ,可能与其脾脏肿大有关 ;而且淋巴细胞的增殖与凋亡的失衡与巨噬细胞的功能无关 ,可能与淋巴细胞内在的异常有关。     

震动伤引起的家犬外周血淋巴细胞凋亡及其机制

应用免疫细胞化学和酶细胞化学技术，观察了震动伤引起的家犬外周血淋巴细胞凋亡，Ｔ淋巴细胞数的变化及其发生机制。结果表明，实验所用２００、１００和８０三种垂直加速度值（Ｇ）在致伤后不同时间，均可引起淋巴细胞的凋亡，其凋亡率随Ｇ值增加而增大；观察伤后不同时间细胞凋亡的变化，发现在伤后３ｄ其凋亡率达到峰值，约为伤前的５～８倍；而外周血Ｔ淋巴细胞数在伤后明显降低，与凋亡呈现出相反的变化趋势；还发现伤后３ｄ与凋亡相关的ｐ５３和Ｂａｘ蛋白阳性率分别为伤前值的２．３和１．８倍，二者在诱发淋巴细胞凋亡中可能起着重要作用。     

Apoptosis in histiocytic necrotizing lymphadenitis

Cell death can now be divided into necrosis and apoptosis, which are different in their morphology, biochemistry and biological significance. The present study was designed to investigate cell death in histiocytic necrotizing lymphadenitis (HNL). The features of cell death in 10 cases of HNL were analyzed using histiomorphology, ultrastructure and in situ apoptosis detection (ApopTag) methods. Two patterns of cell death were discerned. One was apoptosis of individual cells and the other was necrosis. The first pattern could be observed in all cases and the morphological features of the dead cells were consistent with those of apop tosis, which included distinctive cell volume shrinking and chromatin condensation. The apoptotic cells and bodies could frequently be found to be phagocytosed by the histiocytes. ApopTag was positively stained in most of the morphologically apoptotic cells. By double staining, most ApopTag positive cells were found to be T lymphocytes. A previous report showed that the majority of the proliferative cells were T lymphocytes. Based on those results, if was speculated that the main pathological characteristics of HNL therefore consisted of apoptosis and the proliferation of T lymphocytes.     

Lymphocyte subpopulations in pyogranulomas of caseous lymphadenitis.

Pyogranulomas of ovine caseous lymphadenitis (CLA) are encapsulated lesions resulting from infections with Corynebacterium pseudotuberculosis, a bacterial pathogen able to grow within macrophages. Immunohistology of CLA lesions showed a band of lymphocytes lining the inside of the collagen capsule in intimate contact with necrotic tissue, the intracapsular lymphocytes being organized into three layers. The innermost layer, immediately adjacent to the central necrotic tissue consisted of a narrow band of MHC class II staining macrophages. Cells staining for CD4, CD8 and gamma delta T cell markers were unevenly distributed throughout the lymphoid layer, tending to be more numerous immediately external to the macrophage layer. The intracapsular lymphoid tissue contained a high proportion of CD8+ lymphocytes (CD4:CD8, 1.5:1) and of gamma delta lymphocytes (CD4:CD8:gamma delta, 1:0.7:0.8). External to the T cell-rich zone and adjacent to the surrounding collagen capsule was a dense band of cells, a proportion of which stained atypically for CD45R and were tentatively identified as B cells. CD8+ and gamma delta+ T cells showed similar distributions and their relative abundance, compared with CD4+ T cells, was a distinguishing feature of the CLA lesion. Staining for factor VIII-related antigen clearly showed endothelial venules throughout the intracapsular lymphoid tissue. The presence of endothelial venules and the organized architecture of the lymphoid tissue teleologically argues that lymphocytes are continually recruited into chronic CLA lesions and play an important role in the ongoing disease process.     

Molecular Methods for the Identification of Apoptosis in Tissues

Abstract

There has been an upsurge in the interest paid to the process of apoptosis in recent years as a consequence of the realization that apoptosis is important in the regulation of cell number in normal tissues and in a wide variety of diseases. However, the methods used to identify apoptosis in tissues have not progressed. In this review, the development of novel methods for the identification of apoptosis in histological material is discussed. These methods rely on the degradation of DNA that is seen in apoptotic cells and that can be labeled by the incorporation of a biotinylated nucleotide, followed by an enzyme histochemical demonstration method. The problems inherent in the use of these techniques are discussed, particularly relating to situations other than apoptosis where degraded DNA will occur. These methods will prove to be a useful adjunct to morphological examination for the identification and quantification of apoptosis. (The J Histotechnol 17:261, 1994)