Enhancing Immunotherapeutic Value of Lymphocytes by Controlling Apoptosis

本研究的主要目的是确定被细胞因子激活的NK细胞和T细胞在遇到靶细胞上的激活配体时发生凋亡的机制。我们观察到，在体外用细胞因子IL2和IL15刺激的人原代NK细胞和T细胞，当它们的激活受体交联时，显示出广泛的凋亡潜力。为了进一步研究这种激活诱导NK细胞死亡，我们从早期的微阵列研究中得到了启发。我们的微阵列数据显示，在il - 2培养的细胞中，抗凋亡分子TOSO被下调了5倍。TOSO也被称为Fas凋亡抑制分子3 (FAIM3)，是免疫球蛋白基因超家族的成员，它抑制Fas介导的细胞凋亡。TOSO主要在某些种类的免疫细胞中表达。然而，TOSO在NK细胞活化的生理背景下的作用及其功能尚不清楚。这促使我们研究TOSO在活化诱导NK细胞死亡中的作用。我们还证实了TOSO水平下调的事实，并使用实时PCR测定了TOSO表达的相对水平。此外，我们发现受il - 2刺激的人NK细胞表达高水平的死亡受体Fas。为了研究TOSO在这一现象中的作用，我们正在克隆人类TOSO基因并在原代人NK细胞中过表达。我们还计划建立一个稳定表达TOSO的细胞系模型。

The main objective of this study is to determine the mechanism by which NK and T cells activated with cytokines undergo apoptosis when they encounter activating ligands on target cells. We observed that human primary NK and T cells that were stimulated in vitro with the cytokines IL2 and IL15 showed extensive potential to undergo apoptosis when their activation receptors were cross linked. In order to study this activation induced cell death in NK cells further, we drew incite from our earlier microarray study. Our microarray data on genes that are modulated upon stimulation of human NK cells in culture with IL2 showed that the antiapoptotic molecule, TOSO was down regulated in cells cultured with IL2 by 5 fold. TOSO, also called as Fas Apoptotic Inhibitory Molecule 3 (FAIM3) is a member of immunoglobulin gene superfamily and it inhibits Fas mediated apoptosis. TOSO is expressed mainly in certain classes of immune cells. However, the role of TOSO in the physiological context of NK cell activation and their function is not well known. This prompted us to look into the role of TOSO in activation induced cell death in NK cells. We also confirmed the fact that TOSO levels were down regulated and quantitated the relative levels of TOSO expression using real time PCR assays. Further, we found that human NK cells when stimulated with IL2 expressed high levels of the death receptor Fas. In order to study the involvement of TOSO in this phenomenon, we are in the process of cloning the human TOSO gene and over expressing it in primary human NK cells. We are also planning to establish a cell line model for stable expression of TOSO.