缺血性糖尿病足部溃疡皮肤Vδ1T细胞胰岛素样生长因子1与诱生型一氧化氮合酶的表达

背景:皮肤Vδ1T细胞分泌的一系列细胞因子能促进伤口愈合,但与糖尿病足部皮肤组织溃疡难以愈合的关系尚未见报道。目的:观察缺血性糖尿病患者足部溃疡皮肤Vδ1T细胞中胰岛素样生长因子1与诱生型一氧化氮合酶的表达。方法:选取缺血性糖尿病足部溃疡患者及正常对照者足部全层皮肤,各10例。双酶标免疫荧光标记测量Vδ1T细胞及胰岛素样生长因子1、诱生型一氧化氮合酶蛋白表达。结果与结论:缺血性糖尿病足部溃疡患者Vδ1T细胞中胰岛素样生长因子1蛋白表达较对照者明显减少(P<0.01);诱生型一氧化氮合酶蛋白表达明显增加(P<0.01)。提示Vδ1T细胞中胰岛素样生长因子1减少可能与缺血性糖尿病足部溃疡难以愈合有关,而胰岛素样生长因子1减少与溃疡皮肤Vδ1T细胞诱生型一氧化氮合酶表达增加,产生氧化应激有关。

Vδ1 T细胞和TINs细胞表达情况与宫颈癌患者临床分期及预后的关系

目的探讨Vδ1 T细胞和肿瘤浸润性中性粒细胞(tumor infiltrating neutrophil,TINs)细胞表达情况及其与宫颈癌临床分期及预后关系。方法选取我院2012年1月至2017年1月期间经组织病理学确诊的60例宫颈癌患者为研究组,根据临床分期分为Ⅰ~Ⅳ期,患者数22例、18例、12例、8例,并选择同期100例良性病变者作为对照组。采用流式细胞术检测所有患者组织标本中Vδ1 T细胞表达水平,检测CD66b+TINs水平,IL^(-1)1、IL-8、IL^(-1)8炎性细胞因子,CCL2、CCL5、CXCL10趋化因子mRNA水平,并评价上述细胞因子表达水平与宫颈癌临床分期之间的相关性;采用ROC曲线评估Vδ1 T细胞和TINs细胞联合检测对宫颈癌患者预后预测价值。结果不同临床分期宫颈癌患者炎性细胞因子(IL^(-1)1、IL-8、IL^(-1)8)水平以及趋化因子(CCL2、CCL5、CXCL10)水平相比对照组均明显升高,且Ⅲ期和Ⅳ期患者高于Ⅰ期组患者(P<0.001);多元Logistic回归分析结果显示γδT细胞比例、CD66b+TINs百分比以及IL^(-1)1、IL-8、IL^(-1)8以及CCL2、CCL5均属于影响宫颈癌患者临床分期的危险因素(P<0.05);随访3年,生存率为68.33%(41/60),ROC曲线分析结果显示联合诊断曲线下面积为0.895(95%CI:0.795~0.965),灵敏度为84.2%,特异度73.2%,约登指数为0.574,具有最佳诊断效能。结论宫颈癌患者Vδ1 T细胞和TINs细胞随着临床分期进展而升高,联合检测对于宫颈癌患者预后具有一定的预测价值。

慢性心力衰竭患者外周血Vδ1T细胞数量及功能变化

目的观察慢性心力衰竭(CHF)患者外周血Vδ1 T细胞数量和功能的改变,探讨Vδ1 T细胞在CHF发病中的作用。方法选取2012年3月至2013年12月在该院入住的CHF患者50例及同期入院进行健康体检者50例,抽取CHF患者及健康对照(HC)者外周静脉血10 ml。采用流式细胞术测定其外周血Vδ1 T细胞比例及Vδ1 T细胞表面Foxp3表达情况。采用CFSE染色方法检测Vδ1 T细胞对CD4 T细胞增殖能力的影响。采用流式细胞术测定Vδ1 T细胞与CD4 T细胞共孵育后对CD4 T细胞分泌IFN-γ及TNF-α能力的影响。结果 CHF患者外周血Vδ1 T细胞数量〔(1.59±0.77)%〕与HC〔(1.41±0.344)%〕相比无统计学差异(P>0.05)。与HC外周血Vδ1 T细胞表面Foxp3表达水平〔(19.82±2.46)%〕相比,CHF组Foxp3表达水平〔(1.23±0.86)%〕显著降低(P<0.05)。HC外周血Vδ1 T细胞可显著抑制CD4 T细胞增殖和IFN-γ及TNF-α分泌能力,而CHF患者外周血Vδ1 T细胞不具有此功能。结论 CHF患者外周血Vδ1 T细胞功能丧失,致使外周血CD4 T细胞增殖和分泌IFN-γ及TNF-α能力增强,CHF患者免疫功能的这种改变可能与CHF的发生相关。

MICA反应性Vδ1γδT细胞对上皮肿瘤细胞的细胞毒作用

目的研究人类MHCⅠ类链相关基因A(MICA)是否可以在体外诱导Vδ1γδT细胞的扩增,并且对MICA反应性Vδ1γδT细胞的细胞毒作用进行初步研究。方法采用分子克隆技术原核表达并纯化MICA蛋白,使用重组MICA蛋白在体外诱导肿瘤组织中的Vδ1γδ肿瘤浸润T淋巴细胞(TILs),3-(4,5-二甲基噻唑-乙-YL)-2,5-二甲基-四唑溴盐(MTT)法检测MICA反应性Vδ1γδTILs的细胞毒活性。结果原核表达系统pET30表达并纯化了MICA;固相化的MICA蛋白可以诱导肿瘤组织中的Vδ1γδTILs在体外扩增,同时Vδ1γδTILs对MICA+的肿瘤细胞具有明显杀伤活性。结论MICA反应性Vδ1γδT细胞可能是肿瘤过继免疫治疗的新的候选效应细胞。

荧光定量RT-PCR技术检测健康人外周血γδT细胞细胞受体分子Vδ1～8亚群分布

目的：检测和分析健康成人外周血中γδT细胞的T细胞受体（ TCR）分子Vδ1~8亚群分布情况。方法：采集16名健康成人外周血，荧光定量反转录聚合酶链式反应检测其γδT细胞TCR分子Vδ1~8亚群的基因相对表达量，计算各亚群所占百分比。结果：健康成人外周血γδT细胞TCR分子Vδ1~8亚群的比例分别为：Vδ1（7.19±0.33）%、Vδ2（54.27±7.9）%、Vδ3（20.23±0.03）%、Vδ4（3.80±0.02）%、Vδ5（2.08±0.02）%、Vδ6（0.81±0.25）%、Vδ7（7.21±0.02）%、Vδ8（4.41±0.04）%。 Vδ2亚群均明显高于其他亚群（P〈0.01）。结论：我国健康成年人外周血γδT细胞TCR分子Vδ1~8亚群分布由多到少依次为Vδ2〉Vδ3〉Vδ1〉Vδ8〉Vδ7〉Vδ4〉Vδ5〉Vδ6。

缺血性糖尿病足部溃疡皮肤VδT1细胞中IGF-1表达的研究

目的探讨缺血性糖尿病足部溃疡皮肤Vδ1T细胞中胰岛素样生长因子-1(IGF-1)的表达及其临床意义。方法 2008年9月￣2010年8月年10月在南方医院住院和门诊的20例患者自愿捐献的标本,分为两组,每组10人,缺血性糖尿病足部溃疡(IDF)组:男4例,女6例;正常对照(NC)组:男5例,女5例;双酶标免疫荧光标记Vδ1T细胞及IGF-1蛋白的表达,以Olympus FluoView FV10i激光共聚焦显微镜观察和摄像,并用IMAGE-PROPLUS6.0分析IGF-1的表达。结果 IDF组及NC组中Vδ1阳性细胞的数量无明显差异(P>0.05),IDF组Vδ1T细胞中IGF-1的表达较NC组减少,差异有统计学意义(P<0.001)。结论缺血性糖尿病足部溃疡皮肤Vδ1T细胞中IGF-1的表达减少,提示Vδ1T细胞中IGF-1的减少可能与缺血性糖尿病足部溃疡难以愈合有关。

Wilfoside C3N Promotes Tumor Cell Death by Activating Gammadelta T Cells-Mediated Anti-tumor Immunity

Vδ1^+γδ T lymphocytes are known to play important roles in anti-tumor immunity.We recently reported an anti-tumor activity of wilfoside C3 N,an active component extracted from Chinese medicinal herbs.In the current study,we evaluated the role of Vδ1^+γδ T cells in C3 N anti-tumor activity using an in vitro cell co-culture model.We found that C3 N induced the ECA109 tumor cells to undergo apoptosis in the presence of Vδ1^+γδ T cells.The level of ECA109 apoptosis maximized when both C3 N and Vδ1 + γδ T cells were present,which correlated with the increased expression of Fas on ECA109 and Fas ligand on Vδ1^+γδ T cells induced by C3 N.In addition,C3 N also enhanced secretion of cytokines,perforin and granzymes by Vδ1^+γδ T cells.These observations suggest that activation of Vδ1^+ γδ T cells may play a critical role in C3N-mediated anti-tumor activity.

γδT cells:Major advances in basic and clinical research in tumor immunotherapy

γδT cells are a kind of innate immune T cell.They have not attracted sufficient attention because they account for only a small proportion of all immune cells,and many basic factors related to these cells remain unclear.However,in recent years,with the rapid development of tumor immunotherapy,γδT cells have attracted increasing attention because of their ability to exert cytotoxic effects on most tumor cells without major histocompatibility complex(MHC)restriction.An increasing number of basic studies have focused on the development,antigen recognition,activation,and antitumor immune response ofγδT cells.Additionally,γδT cell-based immunotherapeutic strategies are being developed,and the number of clinical trials investigating such strategies is increasing.This review mainly summarizes the progress of basic research and the clinical application ofγδT cells in tumor immunotherapy to provide a theoretical basis for further the development ofγδT cell-based strategies in the future.

Analysis of tumor-infiltrating gamma delta T cells in rectal cancer

AIM: To investigate the regulatory effect of Vδ1 T cells and the antitumor activity of Vδ2 T cells in rectal cancer.METHODS: Peripheral blood, tumor tissues and paracarcinoma tissues from 20 rectal cancer patients were collected. Na?ve CD4 T cells from the peripheral blood of rectal cancer patients were purified by negative selection using a Naive CD4+ T Cell Isolation Kit Ⅱ(Miltenyi Biotec). Tumor tissues and para-carcinoma tissues were minced into small pieces and digested in a triple enzyme mixture containing collagenase type Ⅳ, hyaluronidase, and deoxyribonuclease for 2 h at room temperature. After digestion, the cells were washed twice in RPMI1640 and cultured in RPMI1640 containing 10% human serum supplemented with L-glutamine and 2-mercaptoethanol and 1000 U/m L of IL-2 for the generation of T cells. Vδ1 T cells and Vδ2 T cells from tumor tissues and para-carcinoma tissues were expanded by anti-TCR gδ antibodies. The inhibitory effects of Vδ1 T cells on na?ve CD4 T cells were analyzed using the CFSE method. The cytotoxicity of Vδ2 T cells on rectal cancer lines was determined by the LDH method.RESULTS: The percentage of Vδ1 T cells in rectal tumortissues from rectal cancer patients was significantly increased, and positively correlated with the T stage. The percentage of Vδ2 T cells in rectal tumor tissues from rectal cancer patients was significantly decreased, and negatively correlated with the T stage. After culture for 14 d with 1 mg/m L anti-TCR gδ antibodies, the percentage of Vδ1 T cells from para-carcinoma tissues was 21.45% ± 4.64%, and the percentage of Vδ2 T cells was 38.64% ± 8.05%. After culture for 14 d, the percentage of Vδ1 T cells from rectal cancer tissues was 67.45% ± 11.75% and the percentage of Vδ2 T cells was 8.94% ± 2.85%. Tumor-infiltrating Vδ1 T cells had strong inhibitory effects, and tumor-infiltrating Vδ2 T cells showed strong cytolytic activity. The inhibitory effects of Vδ1 T cells from para-carcinoma tissues and from rectal cancer tissue were not significantly different. In addition, the cytolytic activities of Vδ2 T cells from para-carcinoma tissues and from rectal cancer tissues were not significantly different.CONCLUSION: A percentage imbalance in Vδ1 and Vδ2 T cells in rectal cancer patients may contribute to the development of rectal cancer.

Distinct Pattern of Human Vδ/51 γδ/5 T Cells Recognizing MICA

γδ T cells represent one unique recognition pattern, the limited recognition, which distinguishes from the specific recognition for αβ T cells and pattern recognition for macrophages. Vδ1 γδT cell is the major subset of human γδT cells, which predominates in mucosal tissue including the intestinal epithelia. Presently, a few antigens that human Vδ1TCR can recognize have been identified. Among them, MHC class I chain-related molecules A （MICA） have been studied most intensively. Besides Vδ1TCR, MICA is also the ligand of NKG2D, a C-type lectin-like activating immunoreceptor. In human, only Vδ1 cells can simultaneously express both types of receptors of MICA while NK cells, αβ T cells and other subsets of γδT cells likewise express NKG2D. Although the precise mechanisms are still enigmatic, this distinct pattern of Vδ1 cells recognizing MICA predicts unique biological significance of Vδ1 cells in immune defense. Recent years, some progresses have been made in this issue. In this review we summarize the related reports and put forward some novel views based on our group＇s studies.

肺癌患者外周血及组织中γδT细胞的分布特征及意义

目的探讨肺癌患者外周血和组织中γδT细胞及其亚群的分布特征及临床意义。方法流式细胞术检测10例肺癌患者外周血、20例健康对照者外周血、15例肺癌患者癌组织及其癌旁正常组织中γδT细胞、Vδ1 T细胞和Vδ2 T细胞的表达水平。结果肺癌患者外周血中Vδ2 T细胞的表达水平显著低于健康对照者,差异有统计学意义(P<0.001);肺癌患者癌组织中浸润的γδT细胞(P<0.001)和Vδ1 T细胞(P<0.001)表达水平显著高于癌旁正常组织;肺癌患者癌组织中浸润的Vδ1 T细胞表达水平与淋巴结转移相关(P<0.05)。结论肺癌患者外周血及癌组织中γδT细胞及其亚群具有截然不同的分布特征,癌组织浸润的Vδ1 T细胞表达水平与淋巴结转移有关,提示γδT细胞及其亚群可能参与肺癌病程的进展。

γδ T淋巴细胞的抗肿瘤作用

粘膜免疫和固有免疫在对肿瘤的防御中起着重要作用。γδ型T细胞大量的分布于粘膜相关淋巴组织,其作用介于固有免疫和适应性免疫之间并且是MHC非限制性的。因此其在抗肿瘤免疫中的作用也是不可忽视的。以下将对其两个主要的功能亚群VγVδ2 T细胞和Vδ1 T细胞向肿瘤组织内浸润、对肿瘤细胞的识别及杀伤机制进行简要综述。