The effect of rapamycin on T cell development in mice

Rapamycin (RAPA) is a strong immunosuppressant with a chemical structure similar to that of FK506, although it acts by a mechanism different from both FK506 and cyclosporin A. The effect of RAPA on T cell development in mice was investigated in this study. RAPA caused significant thymic atrophy. The major histological change in the RAPA-treated thymus was thinning of the cortex. No other apparent damage in the cortex or medulla was observed. Consistent with these histological findings, in vivo thymocyte cycling was blocked by RAPA before the S phase, and the ex vivo and in vitro proliferation of the thymocytes was also strongly repressed by the drug. According to electron microscopy and DNA fragmentation assays, RAPA did not induce apoptosis. These results indicate that the repressed thymocyte proliferation is a major mechanism causing RAPA-induced thymic atrophy. Further, RAPA had no effect on thymocyte apoptosis induced by anti-CD3 or ionomycin, and the drug did not interfere with deletion of CD4⁺8⁺ thymocytes or peripheral Vβ6⁺ T cells induced by anti-CD3 or Mls-1^a, respectively. These data suggest that RAPA does not hamper the negative selection. There was a relative increase in the CD3^? fraction of the de novo developing CD4 and CD8 double-positive (DP) thymocytes in the RAPA-treated mice. Moreover, there were relative increases of the CD3^? fractions of the CD4 or CD8 single-positive (SP) cells in both the thymi and periphery. The generation of the CD3^? SP under the influence of RAPA could be used as a useful model for further study of the function and signal transduction of these CD3-defective SP cells.     

BTLA对调节性T细胞发育及功能的影响

目的 研究B和T淋巴细胞弱化因子(B and T lymphocyte attenuator,BTLA)对调节性T细胞(regulatory T cell,Treg)发育和功能的影响.方法 构建在Treg中特异性敲除BTLA基因的小鼠模型,使用流式细胞术检测该模型小鼠中枢及外周各淋巴器官中T细胞外周环境稳态、T细胞的活...     

T cell development and function in CrkL-deficient mice

The adapter protein CrkL has been implicated in multiple signal transduction pathways in hematopoietic cells. In T lymphocytes, the recruitment of CrkL-C3G complexes has been correlated with hyporesponsiveness, implicating CrkL as a potential negative regulator. To test this hypothesis we examined T cell activation in CrkL-deficient mice. The CrkL(-/-) genotype was partially embryonic lethal. In viable CrkL(-/-) mice, peripheral blood counts were normal. The thymus from CrkL(-/-) mice had 40% fewer cells compared to littermates, but the proportion of thymocyte subsets was comparable. There was no discernable alteration in T cell function as reflected by T cell numbers, expression of memory markers, IL-2 production, proliferation, and differentiation into Th1/Th2 phenotypes. Immunization induced comparable levels of IgG2a and IgG1 antibodies. Chimeric mice, generated by transfer of CrkL(-/-) fetal liver cells into irradiated RAG2(-/-) recipients, also showed normal T cell function, arguing against selection via partial embryonic lethality. Our results indicate that CrkL is not absolutely required for T cell development or function, and argue against it being an essential component of a negative regulatory pathway in TCR signaling.     

Vav1蛋白与T细胞功能

Vav1表达于人体所有血细胞相对分子质量为95000的蛋白,其在T细胞抗原受体（TCR）激活后发生迅速的酪氨酸磷酸化后活化,作为鸟苷酸交换因子（GEF）发挥重要作用。Vav1主要参与T细胞信号传导途径,传导信号激活钙流（[Ca^2＋]流）、细胞外信号调节激酶（ERK）以及转录因子NF—κB。Vav1蛋白在T细胞生长发育、成熟T细胞激活、细胞因子合成及细胞骨架活动中发挥重要作用。     

Notch信号通路在T淋巴细胞发育和功能中的作用

T淋巴细胞的发育是由严格的遗传程序调控的精细过程,而Notch信号通路是这一高度复杂遗传程序中最为关键的一环.Notch还与T细胞的激活和功能密切相关.此外,大量的临床和实验室研究证实,Notch信号途径的关键分子是多种人类遗传病的致病基因.Notch1受体基因的染色体易位或点突变是大多数成人T淋巴母细胞白血病的原因.该文将结合此研究室的工作,综述这一领域的相关进展.     

The effect of nitric oxide on sperm cell function and embryo development

PROBLEM: Nitric oxide (NO) has been known to have multifunctional roles both in the male and female reproductive systems. We investigated the effects of sodium nitroprusside (SNP)-dependent NO release on sperm cell function and embryo development to elucidate the mechanisms of action of NO. METHOD OF STUDY: Semen samples from 20 healthy men were processed by the swim-up method. Sperm motility, hyperactivation, and acrosome reaction were examined following incubation with various concentrations of SNP. The concentration of 10 nM to 1 mM was used for sperm motility and hyperactivation measurement and 1 microM to 1 mM for examining the effect on acrosome reaction. Embryo development to blastocyst stage was assessed using 100 nM to 1 mM of SNP added before transferring the mouse embryos into the culture medium. Finally, to understand the mechanism of action of NO, changes in embryo development were examined after zygotes were treated with various concentrations ranging up to 1 mM of 8-bromo-cGMP, an analog of cGMP. RESULTS: Both sperm motility and hyperactivation were significantly reduced at 100 microM and 1 mM concentrations of SNP after 6 hr of incubation. After 24 hr of incubation, they were greatly decreased with all, except the 10 nM concentration of SNP. The percentage of acrosomal-reacted spermatozoa was increased with the increasing concentration of SNP following incubation with 10 microM and 1 mM of SNP. Embryo development was arrested since the two-cell embryonic stage with all except the 100 nM concentration of SNP, and inhibited by 200 microM of SNP regardless of SNP treatment stage. However, embryo development was not influenced by 8-bromo-cGMP. CONCLUSIONS: We concluded that SNP-inhibited sperm cell function and embryo development in a dose- and time-dependent manner, and the inhibitory effect on embryo development, may not be a stage-specific treatment mediated via a cGMP-independent pathway. This result suggests that NO may be enough to affect the fecundity potential in vivo.     

HIV-induced deletion of antigen-specific T cell function is MHC restricted.

When antigen-specific T cells are pulsed by antigen-presenting cells (APC) in the presence of HIV they are functionally deleted following subsequent exposure to syngeneic APC in the absence of HIV. Recombinant soluble HIV envelope (gp120) is able to induce a similar effect which, unlike that induced by HIV, is reversible. Neither HIV nor gp120 affect the ability to respond to IL-2. Thus it is only antigen-specific responses involving the T cell receptor pathways and CD4/MHC class II interaction that appear to be inhibited by HIV-1 and gp120. Furthermore, the functional impairment caused by HIV-1 is specific to the T cells that respond to the antigen in co-culture with HIV, as there is no apparent effect on 'bystander'-activated T cells specific for another antigen. Antigen-specific T cell lines may be deleted by a signalling mechanism which involves molecules other than gp120/CD4 but still requires MHC class II restriction.     

Coronin-1 expression in T lymphocytes: insights into protein function during T cell development and activation

Coronin has been described as an actin-binding protein of Dictyostelium discoideum, and it has been demonstrated to play a role in cell migration, cytokinesis and phagocytosis. Coronin-related proteins are found in many eukaryotic species, including Coronin-1 in mammals whose expression is enriched in the hematopoietic tissues. Here, we characterize Coronin-1 gene and protein expression in mouse embryonic and adult T lymphocytes. Coronin-1 is expressed throughout T cell ontogeny and in peripheral alphabeta T cells. Expression varies along thymic cell development, with maximum levels observed in embryonic early thymocytes and, in the adults, the selected TCRalphabeta(+) single-positive thymocytes. Subcellular localization analysis indicates that Coronin-1 is in equilibrium between the cytosol and the cell cortex, where it accumulates in F-actin-rich membrane protrusions induced by polarized activation of TCR-CD3-stimulated T cells. These data are consistent with a role of Coronin-1 in T cell differentiation/activation events involving membrane dynamisms and the cortical actin cytoskeleton.     

Instant conditional transgenesis in the mouse hematopoietic compartment

Adoptive transfer of retrovirally transduced stem cells has recently been described for instant transgenesis in the hematopoietic compartment of mice. This method circumvents the need to manipulate the germline. However, cell type specific gene expression in this ‘retrogenic’ mouse model has remained tedious. Here we report a single retroviral vector-based method to rapidly generate conditional retrogenic mice. For this purpose, mutated loxP-flanked DNA segments are transduced into hematopoietic stem cells isolated from Cre recombinase transgenic mice, which are subsequently transferred into immunodeficient mice. In this way gene expression can be restricted to hematopoietic cell lineages of choice in the acquired immune system.     

CTLA4—Ig影响T细胞功能及机理的初步研究

目的：研究CTLA4-Ig融合蛋白对外周血T淋巴细胞活化的抑制作用及机理。方法：分离新鲜人外周血单个核细胞,观察CTLA4-Ig对淋巴细胞转化和混合淋巴细胞培养的作用,用流式细胞术观察CTLA4-Ig对T细胞表面CD25表达的影响,最近通过EMSA方法检测激活T细胞中核蛋白的变化。结果：CTLA4-Ig能够抑制淋巴细胞转化和混合淋巴细胞反应,减低T细胞表面CD25的表达,抑制核内与IL-2表达相关的RE/AP结合因子的活化。结论：CTLA4-Ig通过多种信号途径对T细胞的活化起作用,其机制之一是抑制核内结合于IL-2增强子RE/AP位点的核因子的活化。     

Heme oxygenase-1 is not required for mouse regulatory T cell development and function

CD4 regulatory T cells (Treg) ensure peripheral tolerance to self-antigens and limit the deleterious effects associated with inflammatory and immune responses by mechanisms that remain to be fully understood. The enzyme heme oxygenase-1 (HO-1), through its known anti-inflammatory activity, is a candidate for a functional role in Treg activity. We compared wild-type and heme oxygenase-1-deficient (hmox-1(-/-)) mice in order to assess the role of HO-1 in mouse Treg development and function under physiologic conditions. The frequency of CD25+ and Foxp3+ Treg was similar in hmox-1(-/-) and hmox-1(+/+) mice. More importantly, CD4+ CD25+ Treg purified from either hmox-1(-/-) or hmox-1(+/+) mice were equally efficient in controlling the proliferation in vitro and the expansion in vivo of CD4+ CD25- T cells, whether or not these responder cells expressed HO-1. In addition, induction of expression of HO-1 in vivo did not affect Treg suppressor function. As shown before, expression of HO-1 was higher in Treg than in naive T cells; however, naturally activated Foxp3- T cells displayed equal amount of HO-1 mRNA as Treg. Finally, we conclude that under physiological conditions in mice, Treg development, maintenance and function are independent of HO-1 activity.     

T cell deletion follows chronic antigen specific T cell activation in vivo

Exposure of mice transgenic for a TCR (F5) to cognate peptideantigen results in thymic depletion of CD4⁺ CD8⁺ cells and expansionand activation of peripheral CD8⁺ TCR(tg)⁺ T cells.in the thymusapoptotic DNA ladder is evident as early as 3 h after peptideinjection. Long exposure of intact or thymectomized F5 TCR transgenicmice to peptide antigen leads to depletion of most of the peripheralCD8⁺ T cells bearing the F5 receptor, with the remaining cellshaving lower levels of transgenic TCR compared with non-treatedanimals. In the thymus of intact F5 TCR transgenic mice suchcontinuous exposure to antigen results in the reappearance ofCD4⁺CD8⁺ with lower levels of the transgenic receptor.