Biron C A, Natuk R J, Welsh R M
J Immunol. 1986 Mar 15;136(6):2280-6.
Cytolytic lymphocytes were isolated from the spleens of lymphocytic choriomeningitis virus (LCMV)-infected mice and were characterized in regards to function, cell size, antigen phenotype, and cell morphology. Only 2% of the Lyt-2+ cells from uninfected mice were large granular lymphocytes (LGL), whereas 21% of the Lyt-2+ cells isolated 7 days postinfection were LGL. The day 7 Lyt-2+ populations contained all of the LCMV-specific, class I histocompatibility antigen-restricted cytotoxic T lymphocyte (CTL) activity, but no natural killer (NK) cell activity. The NK cell activity was consistently recovered in Lyt-2- populations isolated from both control mice and mice on day 7 postinfection. The LGL isolated on day 7 postinfection were concluded to be predominantly T cells and not NK cells because 1) the proportions of LGL in fractionated cell populations 7 days postinfection correlated with levels of CTL-mediated lysis but not NK cell-mediated lysis, 2) they were recovered in the Lyt-2+ population, and 3) antibody to asialo GM1, known to eliminate NK cell-mediated lysis but not T cell-mediated lysis, dramatically reduced NK cell LGL numbers in vivo on day 3 postinfection but only marginally affected LGL numbers on day 7. Virus-induced inflammation elicited a 50-fold increase in LGL numbers in the peritoneum on day 7 postinfection. The peritoneal exudate LGL were also associated with CTL activity and were resistant to treatment with antibody to asialo GM1. These results indicate that in vivo-generated CTL have the morphology of LGL and that the appearance of cytoplasmic granules correlates with the ability of cells to mediate lysis. To focus on cells being stimulated during infections, activated blast cells were separated from small resting cells by centrifugal elutriation. Coincidental with the peak in overall spleen leukocyte cytotoxic activity, the peaks of blast NK cells and CTL were at days 3 and 7 postinfection respectively. More than 50% of the blast lymphocytes isolated on either day 3 or day 7 postinfection were LGL. The CTL activity in the blast populations on day 7 postinfection was mediated by Lyt-2+ cells, and 37 to 64% of these Lyt-2+ blast cells were LGL. Cytolytic NK cell and CTL LGL could not be distinguished by morphology or by cell densities, because they overlapped in low density Percoll gradient fractions. Since this technique has been used to enrich for LGL, these data indicate that heterogeneity in LGL populations may result from the presence of both CTL and NK cell LGL.
从感染淋巴细胞性脉络丛脑膜炎病毒（LCMV）的小鼠脾脏中分离出溶细胞性淋巴细胞，并对其功能、细胞大小、抗原表型和细胞形态进行了鉴定。未感染小鼠的Lyt-2⁺细胞中只有2%是大颗粒淋巴细胞（LGL），而感染后7天分离出的Lyt-2⁺细胞中有21%是LGL。感染后第7天的Lyt-2⁺细胞群体包含了所有LCMV特异性的、I类组织相容性抗原限制的细胞毒性T淋巴细胞（CTL）活性，但没有自然杀伤（NK）细胞活性。在从对照小鼠和感染后第7天的小鼠中分离出的Lyt-2⁻细胞群体中，始终能检测到NK细胞活性。得出感染后第7天分离出的LGL主要是T细胞而非NK细胞的结论，原因如下：1）感染后7天分级细胞群体中LGL的比例与CTL介导的裂解水平相关，但与NK细胞介导的裂解水平无关；2）它们在Lyt-2⁺细胞群体中被分离出来；3）抗唾液酸GM1抗体已知可消除NK细胞介导的裂解，但不影响T细胞介导的裂解，在感染后第3天能显著减少体内NK细胞LGL的数量，但对第7天的LGL数量影响很小。病毒诱导的炎症在感染后第7天使腹膜中的LGL数量增加了50倍。腹膜渗出液中的LGL也与CTL活性相关，并且对抗唾液酸GM1抗体的处理具有抗性。这些结果表明，体内产生的CTL具有LGL的形态，并且细胞质颗粒的出现与细胞介导裂解的能力相关。为了聚焦于感染期间被刺激的细胞，通过离心淘洗将活化的母细胞与小的静止细胞分离。与脾脏白细胞总体细胞毒性活性的峰值同时出现的是，母细胞NK细胞和CTL的峰值分别出现在感染后第3天和第7天。在感染后第3天或第7天分离出的母细胞淋巴细胞中，超过50%是LGL。感染后第7天母细胞群体中的CTL活性由Lyt-2⁺细胞介导，这些Lyt-2⁺母细胞中有37%至64%是LGL。溶细胞性NK细胞和CTL LGL无法通过形态或细胞密度来区分，因为它们在低密度Percoll梯度组分中重叠。由于该技术已被用于富集LGL，这些数据表明LGL群体的异质性可能是由于CTL和NK细胞LGL的共同存在。
