T cell activation correlates with an increased proportion of antigen among the materials acquired from target cells

We have investigated the density of peptides required to elicit different biological responses in cytotoxic T lymphocytes (CTL), including trogocytosis (i.e., the phenomenon whereby the lymphocytes actively capture fragments of plasma membrane from those cells with which they establish an immune synapse). We have used two separate mouse models of CTL recognising defined peptides presented by MHC class I molecules. In both systems, triggering of cytotoxicity and capture of membrane components reached saturation with low densities of ligand. On the other hand, down-modulation of cell-surface levels of TCR, induction of IFN-gamma production and detection of peptide captured required much higher ligand densities. Interestingly, fratricide (i.e., killing between CTL sharing the same specificity), a mechanism proposed to account for CTL exhaustion, was detected only at antigen concentrations still well above that second threshold leading to full blown activation. Taken together, our results show that the different thresholds that govern the elicitation of different CTL functions correlate with different proportions of antigen among the target cell components being captured via trogocytosis.     

Ex vivo stimulation of cord blood mononuclear cells by dexamethasone and interleukin-7 results in the maturation of interferon-gamma-secreting effector memory T cells

The effects of dexamethasone phosphate and interleukin‐7 upon the proliferation of T‐cells and the production of interferon‐γ in the newborn's cord blood mononuclear cell cultures were studied. The capability of dexamethasone to enhance T‐cell proliferation induced by anti‐CD3 with interleukin‐7 in some newborn cord blood mononuclear cell cultures was identified. Dexamethasone suppressed production of interferon‐γ in 68‐h cell cultures stimulated with anti‐CD3 both in the presence of interleukin‐7 and without it. However, a 68‐h cultivation of newborn blood cells with dexamethasone, anti‐CD3 and interleukin‐7 resulted in the accumulation of T‐lymphocytes capable of producing interferon‐γ after restimulation. As a result of it the amount of interferon‐γ producing CD7⁺ T‐cells and the concentration of interferon‐γ in cultural supernatants were maximal in the cell cultures incubated with anti‐CD3, interleukin‐7 and dexamethasone during the first 68 h and subsequently restimulated with phorbol 12‐myristate 13‐acetate and ionomycin. The stimulation of neonatal or adult blood cells by dexamethasone, anti‐CD3 and interleukine‐7 also causes a decrease in the number of naïve T‐cells and central memory cells and an increase in the number of effector memory CD7⁺CD45RA⁺CD62L^– cells in cultures. It is possible that these effects are caused by the influence of dexamethasone on IL‐7 receptor expression: it is known that IL‐7 receptor alpha‐chain gene is a glucocorticoid‐inducible gene.     

Depletion of mitochondrial DNA in HIV-1-infected patients and its amelioration by antiretroviral therapy

Mitochondrial DNA (mtDNA) of peripheral blood mononuclear cells (PBMCs) collected from Human immunodeficiency virus 1 (HIV-1)-infected patients and healthy controls were measured longitudinally using real-time polymerase chain reaction to evaluate the effects of antiretroviral agents on mtDNA synthesis in vivo and to assess the value of monitoring mtDNA in PBMCs to predict adverse events amongst these patients. MtDNA levels in PBMCs were significantly decreased in treatment-naive HIV-1-infected patients compared with healthy people. MtDNA levels were not only significantly correlated with CD4(+) T-cell count, but also inversely correlated with HIV-1 viral load. MtDNA levels in untreated patients and healthy controls were stable during the period of observation. On the other hand, amongst patients treated with regimens containing AZT/3TC or d4T/3TC, mtDNA increased during treatment and recovered to levels comparable to healthy controls. In contrast, mtDNA decreased immediately after the initiation of an AZT/ddC-containing regimen. We did not find a correlation between mtDNA levels and changes in clinical parameters. There was no significant difference in mtDNA levels between patients with and those without lipoatrophy. Furthermore, there was no obvious difference in mtDNA levels amongst those patients exhibiting signs and symptoms of peripheral neuropathy. In conclusion, the decrease in mtDNA levels in PBMCs amongst HIV-1-infected patients and its amelioration by antiretroviral therapy may suggest the influence of direct effects on mitochondria or mtDNA by HIV-1 infection. Further investigations are needed to elucidate the mechanisms contributing to decreased mtDNA and the value of mtDNA measurement in the care of HIV-1-infected individuals.     

神经营养素受体在人外周血T细胞中表达的研究

目的 研究人外周血CD4^ /CD8^ T细胞4种神经营养素受体基因的转录。方法 应用尼龙手法分离出T细胞，磁式细胞分离法（MACS）分离CD4^ /CD8^ T细胞亚群，再以RT－PCR法研究4种神经营养素受体在两种T细胞亚群上的表达。结果 未经刺激的CD4^ /CD8^ T细胞亚群不表达任何神经营养素受体。经PHA或PPD刺激后，CD4^ /CD8^ T细胞亚群表达trkA,CD8^ T细胞亚群表达trkC；而在各种状态下的T细胞上均未见表达trkB及p75^NGFR。结论神经营养素受体在两种T细胞亚群中有不同的表达格局，提示不同T细胞亚群受神经营养素调节的模式可能各不相同。     

巨噬细胞激活因子的分离纯化

由Ｔ细胞株（Ｍｏｌｔ ４）产生的巨噬细胞激活因子（ＭＡＦ）用阴离子交换和凝胶过滤色谱纯化。纯化的ＭＡＦ进行ＰＡＧＥ出现一条带，进行ＳＤＳ－ＰＡＧＥ证实其为单亚基，分子量为６００００；ＩＥＰ表明其等电点在ｐＨ３．５～４．５之间。     

CTLA-4-FasL融合分子的构建及其生物学特性的鉴定

文章通过特异的引物分别扩增出CTLA 4和FasL胞外区的cDNA ,将它们拼接后 ,克隆入真核表达载体pcDNA3 1( + )中 ,进行表达、纯化 ,获得CTLA 4 FasL融合蛋白。Westernblot分析显示了该融合蛋白具有CTLA4 胞外区和FasL胞外区的抗原性。体外试验表明 ,该融合蛋白可以结合Jurkat细胞表面的Fas受体和Raji细胞表面的B7分子 ,表明了该分子双特异性的特点。该融合蛋白能够直接诱导Jurkat细胞发生凋亡 ,且此凋亡效应伴随Raji细胞的参与而增强 ,初步证实了该分子的免疫抑制效应 ,从而为进一步研究该融合蛋白特性及应用奠定了基础。     

Viral hemagglutinin augments peptide-specific cytotoxic T cell responses

In attempt to increase the induction of peptide-specific cytolytic T cells (CTL) we investigated the effect of the Newcastle disease virus (NDV) hemagglutinin-neuraminidase (HN) gene product on the activation of peptide-specific CTL. Spleen cells of CH3 mice immunized against the influenza nucleoprotein peptide 50–63 (NP 50–63) were restimulated in vitro (i) with peptide-pulsed syngeneic fibroblast cells (Ltk^?) as antigen-presenting cells, which were in addition (ii) infected with NDV or (iii) stably transfected with the HN cDN A of NDV. A greater than sixfold increase in peptide-specific CTL responses was observed in cultures restimulated with peptide-pulsed Ltk^? cells which co-expressed viral hemagglutinin due to either infection or transfection. A similar augmentation was seen in CTL responses against other types of antigen (major histocompatibility complex alloantigens, minor histocompatibility antigens or tumor antigens) when suboptimal cultures were stimulated with the respective antigen-presenting cells modified by NDV infection. These findings suggest that NDV or viral HN expressed on antigen-presenting cells or tumor cells can exert a T cell co-stimulatory function.     

Rapid tyrosine phosphorylation of Grb2 and Shc in T cells exposed to anti-CD3, anti-CD4, and anti-CD45 stimuli: differential effects of aging

Two adapter proteins, Grb2 and Shc, have recently been implicated in the transmission of activation signals from the stimulated T cell receptor to Ras. We show here that in vitro stimulation of mouse splenic T cells with crosslinked anti-CD3 antibody leads within 30 s to phosphorylation of both Grb2 and Shc. Treatment with crosslinked anti-CD45 antibody leads to phosphorylation of Grb2 and also to a slight retardation in the mobility of this protein in an SDS polyacrylamide gel; both changes are seen within 30 s of crosslinking. Crosslinked anti-CD4 antibody leads to phosphorylation of Shc and to the phosphorylation of a 30-kDa protein that cross-reacts with anti-Grb2 antibodies. Aging leads to a decline in CD3-stimulated phosphorylation of Shc (but not Grb2), and to an increase in CD4-stimulated phosphorylation of Grb2, Shc, and the 30-kDa Grb2-like protein. Increased tyrosinephosphorylation of Grb2 after exposure to either anti-CD3 or anti-CD45 suggests that Grb2 may be a common substrate for both CD3-linked kinases and the CD45 phosphatase. The differences between T cells from young and old mice suggest that aging may lead to a set of alterations in kinase/substrate coupling that contribute to immune dysfunction in the elderly, and that activation of the Ras pathway might be impaired by aging in T lymphocytes.     

Neurological Manifestations of COVID-19 Feature T Cell Exhaustion and Dedifferentiated Monocytes in Cerebrospinal Fluid

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过继转输胚胎抗原耐受T细胞诱导自然流产孕鼠母胎免疫耐受

探讨过继转输胚胎抗原耐受T细胞对小鼠自然流产模型妊娠预后及宿主孕鼠免疫细胞对父系抗原免疫耐受状态的影响。以CBA/J×BALB/c为正常妊娠模型 ,CBA/J×DBA/ 2为自然流产模型 ,将自然流产模型CBA/J孕鼠于孕 4d (着床期 )分别腹腔注射大鼠抗小鼠CD80和CD86mAb或大鼠同型IgG。于孕 9d ,应用免疫磁珠阴性分选三组孕鼠的脾脏T细胞 ,并将三组T细胞分别转输至孕 4d的CBA/J×DBA/ 2孕鼠。于宿主孕鼠孕第 9天 ,采用单向混合淋巴细胞反应分析宿主孕鼠脾脏免疫细胞对父系抗原的增殖能力 ,并用流式细胞术分析经父系抗原刺激的宿主孕鼠脾脏T细胞内IL 2表达水平。于孕 1 4d分别观察宿主孕鼠的胚胎吸收率。结果显示 ,过继转输胚胎抗原耐受T细胞和转输正常妊娠模型孕鼠的T细胞均可诱导宿主孕鼠脾脏免疫细胞对父系抗原的增殖能力及IL 2的表达显著下降 (P <0 0 5 ) ,孕 1 4d胚胎吸收率也显著下降 (P <0 0 5 )。这些结果表明 ,于孕早期过继转输胚胎抗原耐受T细胞和转输正常妊娠模型孕鼠的T细胞能诱导宿主孕鼠母 胎免疫耐受 ,防止母体对胚胎的免疫排斥 ,从而使自然流产模型的妊娠预后达到正常妊娠水平。