Immunobiology of Hybrid Resistance

描述(申请人提供)：受辐射的H2杂合F1杂合小鼠对H2纯合子亲本株骨髓移植的杂交抗性违反了移植遗传学定律，并由自然杀伤(NK)细胞而不是T淋巴细胞介导。NK细胞表达Ly49受体，接受MHC I类抗原的负信号，如Ly49C或I-H2-KB、Ly49A或G2-H2-DD。也有一些在细胞质区域缺乏抑制基序的Ly49受体与含有激活基序的DAP12蛋白共表达，如Ly49D、H和U。NK细胞的宿主防御功能在很大程度上受这些受体的调节，这些受体表达在NK细胞的不同部分。Ly49受体非常多态，决定了小鼠排斥骨髓移植、杀死肿瘤细胞以及抵抗病毒和其他感染的遗传能力。根据我们最近的研究，我们有三个具体目标。 目的：验证一种假设，即表达Ly49A/G2受体的NK细胞不仅不能杀伤DD+靶细胞，而且还作用于缺乏Ly49A/G2受体的Ly49D+NK细胞以抑制其功能或活性。该方法将激活B6Ly49A转基因小鼠的NK细胞，并测试它们照射B6宿主排斥H_2d骨髓移植的能力。 目的2：验证某些不能排斥H_2d骨髓移植的小鼠品系缺乏Ly49D样受体或在其所有NK细胞上表达Ly49G2或A的假设。该方法将通过同时表达Ly49D和绿色荧光蛋白(GFP)的构建体来转导FVB小鼠的干细胞。这些细胞将被转移到照射FVB小鼠以产生骨髓细胞嵌合体，通过流式细胞仪检测GFP。这些小鼠将接受h2d骨髓移植的挑战，以确定它们现在是否可以排斥。将给予F(ab‘)2个抗Ly49G2和A的单抗片段来阻断阴性信号。目的：验证激活受体(如Ly49H)介导小鼠对Friend白血病病毒(FV)和小鼠巨细胞病毒(MCMV)感染的遗传抵抗力，而抑制性受体对感染抵抗力较差的假设。方法包括使用转基因抑制受体的小鼠，如B6，Ly49C或A，以测试病毒抵抗力，并通过骨髓细胞转移的方式将Ly49H基因转移到易感小鼠，如目标2。这些研究将扩大NK细胞免疫生物学的知识。

DESCRIPTION (provided by applicant): Hybrid resistance in irradiated H2 heterozygous F1 hybrid mice to grafts of H2 homozygous parental strain marrow grafts defies the laws of transplantation genetics and is mediated by natural killer (NK) cells rather than T lymphocytes. NK cells express Ly49 receptors that receive negative signals from MHC class I antigens, e.g. Ly49C or I - H2-Kb, Ly49A or G2 - H2-Dd. There are also Ly49 receptors lacking an inhibitory motif in the cytoplasmic domain which are co-expressed with DAP12 proteins containing an activating motif, e.g. Ly49D, H, and U. Host defense functions of NK cells are largely regulated by these receptors that are expressed on various fractions of NK cells. The Ly49 receptors are very polymorphic and determine the genetic ability of mice to reject marrow grafts, to kill tumor cells and to resist virus and other infections. Based on our recent studies, we have three specific aims. Aim 1: Test the hypothesis that NK cells that express Ly49A/G2 receptors not only fail to kill Dd+ target cells, but also act on Ly49D+ NK cells lacking Ly49A/G2 to inhibit their function or viability. The approach will be to activate NK cells of B6.Ly49A transgenic mice and test their ability of irradiate B6 hosts to reject H2d marrow grafts. Aim 2: Test the hypothesis that certain strains of mice that fail to reject H2d marrow grafts lack Ly49D-like receptors or express Ly49G2 or A on all of their NK cells. The approach will be to transduce stem cells of FVB mice with a construct expressing both Ly49D and green fluorescent protein (GFP). These cells will be transferred to irradiate FVB mice to generate marrow cell chimeras, detected by flow cytometry for GFP. These mice will be challenged with H2d marrow grafts to determine if they can now reject. F(ab')2 monoclonal antibody fragments to Ly49G2 and A will be given to block negative signals. Aim 3: Test the hypothesis that activating receptors, e.g. Ly49H, mediate the genetic resistance of mice to Friend leukemia virus (FV) and murine cytomegalovirus (MCMV) infections, while inhibitory receptors are responsible for poor resistance to infections. Approaches include using mice transgenic for inhibitory receptors, e.g. B6.Ly49C or A, for testing viral resistance, and transferring Ly49H genes to susceptible mice by means of marrow cell transfers, as in Aim 2. These studies will amplify knowledge of NK cell immunobiology.