外周血CD4^(+)PD-1^(+)Tcells及CD4^(+)T淋巴细胞ATP含量与复发性卵巢癌疗效的相关性分析

目的 探讨外周血CD4^(+)程序性细胞死亡受体-1(PD-1)^(+)T cells及CD4^(+)T淋巴细胞三磷酸腺苷(ATP)含量与复发性卵巢癌疗效的相关性。方法 选取30例复发性卵巢癌患者为复发组,30例未复发卵巢癌患者为非复发组;另选取30名同期体检健康者作为对照组。评估外周血CD4^(+)PD-1^(+)T cells及CD4^(+)T淋巴细胞ATP含量与复发性卵巢癌疗效的相关性。结果 复发组和非复发组的CD4^(+)PD-1^(+)T cells较对照组明显升高(P<0.05)。复发组和非复发组的CD4^(+)T淋巴细胞ATP含量较对照组明显降低(P<0.05)。复发组治疗后CD4^(+)PD-1^(+)Tcells显著低于治疗前(P<0.05),治疗后CD4^(+)T淋巴细胞ATP含量显著高于治疗前(P<0.05)。CD4^(+)PD-1^(+)T cells与复发性卵巢癌疗效成负相关(r=-0.393,P=0.039),CD4^(+)T淋巴细胞ATP含量与复发性卵巢癌疗效成正相关(r=0.449,P=0.031)。结论 复发性卵巢癌患者外周血CD4^(+)PD-1^(+)T cells及CD4^(+)T淋巴细胞ATP含量与疗效密切相关。

Peripheral CD4^(+)CD8^(+) double positive T cells:A potential marker to evaluate renal impairment susceptibility during systemic lupus erythematosus

Lupus nephritis(LN) has a high incidence in systemic lupus erythematosus(SLE) patients, but there is a lack of sensitive predictive markers. The purpose of the study was to investigate the association between the CD4^(+)CD8^(+)double positive T(DPT) lymphocytes and LN. The study included patients with SLE without renal impairment(SLE-NRI), LN, nephritic syndrome(NS), or nephritis. Peripheral blood lymphocyte subsets were analyzed by flow cytometry. Biochemical measurements were performed with peripheral blood in accordance with the recommendations proposed by the National Center for Clinical Laboratories. The proportions of DPT cells in the LN group were significantly higher than that in the SLE-NRI group(t=4.012, P<0.001), NS group(t=3.240,P=0.001), and nephritis group(t=2.57, P=0.011). In the LN group, the risk of renal impairment increased significantly in a DPT cells proportion-dependent manner. The risk of LN was 5.136 times(95% confidence interval, 2.115–12.473) higher in cases with a high proportion of DPT cells than those whose proportion of DPT cells within the normal range. These findings indicated that the proportion of DPT cells could be a potential marker to evaluate LN susceptibility, and the interference of NS and nephritis could be effectively excluded when assessing the risk of renal impairment during SLE with DPT cell proportion.

Revolutionizing tumor immunotherapy:unleashing the power of progenitor exhausted T cells

In exploring persistent infections and malignancies, a distinctive subgroup of CD8^(+) T cells, progenitor exhausted CD8^(+) T(Tpex) cells, has been identified. These Tpex cells are notable for their remarkable self-renewal and rapid proliferation abilities. Recent strides in immunotherapy have demonstrated that Tpex cells expand and differentiate into responsive exhausted CD8^(+) T cells, thus underscoring their critical role in the immunotherapeutic retort. Clinical examinations have further clarified a robust positive correlation between the proportional abundance of Tpex cells and enhanced clinical prognosis. Tpex cells have found noteworthy applications in the formulation of inventive immunotherapeutic approaches against tumors. This review describes the functions of Tpex cells in the tumor milieu, particularly their potential utility in tumor immunotherapy. Precisely directing Tpex cells may be essential to achieving successful outcomes in immunotherapy against tumors.

Analysis of CD4^+CD25^+ Regulatory T Cells and Foxp3 mRNA in the Peripheral Blood of Patients with Asthma

The changes of CD4^＋CD25^＋ regulatory T cells （CD4^＋CD25^＋ Treg） and Foxp3 mRNA in peripheral blood mononuclear cells （PBMCs） from patients with asthma were investigated in order to elucidate the possible roles of CD4^＋CD25^＋ Treg in the development of asthma. The peripheral blood samples were collected from 29 healthy controls （normal control group） and 78 patients with asthma which included 30 patients in exacerbation group, 25 patients in persistent group, and 23 patients in remission group. By using flow cytometry and RT-PCR, the CD4^＋CD25^＋ Treg ratio and Foxp3 mRNA in PBMCs were detected. The CD4^＋CD25^＋ Treg ratio and Foxp3 mRNA in PBMCs of exacerbation and persistent groups were lower than that of remission and normal control groups （P〈0.05）. Although the CD4^＋CD25^＋ Treg ratio and Foxp3 mRNA of remission group were also lower than that of normal control group, there was no significant difference between them （P〉0.05）. As compared with persistent group, exacerbation group had lower CD4^＋CD25^＋ Treg ratio and Foxp3 mRNA （P〈0.05）. It was indicated that the decrease of CD4^＋CD25^＋ Treg ratio and its function in PBMCs may be responsible for pathogenesis of asthma.

An Association between Immunosenescence and CD4^+CD25^+ Regulatory T Cells: A Systematic Review

Objective Age-related increment of the prevalence of CD4^＋CD25^＋ regulatory T （Treg） cells were described controversially, and whether such changes explain immune dysfunction in the elderly is still unclear. The aim of this systematic review is to evaluate the role of the Tregs in immunosenescence. Methods Medline and manual searches were performed to identify all published epidemiological and animal studies investigating the efficacy of the association between immunosenescence and Treg cells. Results It was founded that the frequency, phenotypic characteristics, and number/function of Tregs were altered significantly with aging. Medical conditions in individuals with advanced ageas well as apoptosis intensity of Treg cells had an impact on the accumulation of Tregs which in turn could deteriorate cytotoxic activity of CD8＋ T and NK cells and production of IL-2. The range of immune cells that could be suppressed by Treg cells was quite wide and covered CD4^＋CD25^＋ T cells, NK cells, dendritic cells and even monocytes. These changes were observed both in humans and experimental animals. Besides, it was believed that frequency of Tregs increased with age and was accompanied by intensified suppressive activity for Tregs in patients, for example, with Alzheimer disease （AD） and Parkinson disease （PD）. The impaired condition of CD4＋ T cells, so-called immunosenescence, rendered transplant recipients less responsive to an allogeneic kidney graft, an effect that was limited to transplant recipients who were aged over 60 years. Conclusions Treg cells are associated with immunosenescence. All these changes contribute to the aging-related decline of immune responses and lead to the higher risk of immune-mediated diseases, cancer or infections in aged individuals.

Influence of Danshen Injection on airway inflammation and CD4^+ CD25^+ regulatory T cells of asthmatic rats

Objective： To investigate the influence of Danshen Injection on airway inflammation and CD4^＋CD25^＋ regulatory T cells（CD4^＋CD25^＋ Tr） of asthmatic rats, and elucidate the possible mechanism of Danshen Injection in treatment of asthma. Methods： 30 Wister rats were randomly divided into control group, asthma group and Danshen Injection treated group. Bronchoalveolar lavage fluids （BALF） were collected, and cytology studies were conducted. Lung tissues were obtained and pathologic analyses were done with hematoxylin and eosin stain （HE）. Flow cytometry was used to detect the CD4^＋CD25^＋ Tr ratio in peripheral blood mononuclear cells （PBMCs）. Results： Total cell, the percentage of lymphocytes, neutrophils and eosinophils （Eos） in BALF of Danshen Injection-treated group were lower than that in asthma group （P〈0.05, P〈0.01）. Compared with asthma group, less infiltration of inflammatory cells in lung tissues was observed in Danshen Injection-treated group. CD4^＋CD25^＋ Tr of asthma group was lower than that of control and Danshen Injection treated group （P〈0.05）. Conclusion： Danshen Injection can suppress airway inflammation of asthmatic rats, probably by increasing the number of CD4^＋CD25^＋ Tr.

Role of LAP^+CD4^+ T cells in the tumor microenvironment of colorectal cancer

AIM To investigate the abundance and potential functions of LAP^+CD4^+ T cells in colorectal cancer(CRC). METHODS Proportions of LAP^+CD4^+ T cells were examined in peripheral blood and tumor/paratumor tissues of CRC patients and healthy controls using flow cytometry. Expression of phenotypic markers such as forkhead box(Fox)p3, cytotoxic T-lymphocyte-associated protein(CTLA)-4, chemokine CC receptor (CCR)4 and CCR5 was measured using flow cytometry. LAP^-CD4^+ and LAP^+CD4^+ T cells were isolated using a magnetic cellsorting system and cell purity was analyzed by flow cytometry. Real-time quantitative polymerase chain reaction was used to measure expression of cytokines interleukin (IL)-10 and transforming growth factor(TGF)-β.RESULTS The proportion of LAP^+CD4^+ T cells was significantly higher in peripheral blood from patients (9.44% ± 3.18%) than healthy controls (1.49% ± 1.00%, P < 0.001). Among patients, the proportion of LAP^+CD4^+ T cells was significantly higher in tumor tissues(11.76% ± 3.74%) compared with paratumor tissues (3.87% ± 1.64%, P < 0.001). We also observed positive correlations between the proportion of LAP^+CD4^+ T cells and TNM stage(P < 0.001), distant metastasis(P < 0.001) and serum level of carcinoembryonic antigen(P < 0.05). Magnetic-activated cell sorting gave an overall enrichment of LAP^+CD4^+ T cells (95.02% ± 2.87%), which was similar for LAP^-CD4^+ T cells(94.75% ± 2.76%). In contrast to LAP^-CD4^+ T cells, LAP^+CD4^+ T cells showed lower Foxp3 expression but significantly higher levels of CTLA-4, CCR4 and CCR5(P < 0.01). LAP^+CD4^+ T cells expressed significantly larger amounts of IL-10 and TGF-β but lower levels of IL-2, IL-4, IL-17 and interferon-γ, compared with LAPCD4+ T cells.CONCLUSION LAP^+CD4^+ T cells accumulated in the tumor microenvironment of CRC patients and were involved in immune evasion mediated by IL-10 and TGF-β.

Glutamine deprivation impairs function of infiltrating CD8^(+)T cells in hepatocellular carcinoma by inducing mitochondrial damage and apoptosis

BACKGROUND The functions of infiltrating CD8^(+)T cells are often impaired due to tumor cells causing nutrient deprivation in the tumor microenvironment.Thus,the mechanisms of CD8^(+)T cell dysfunction have become a hot research topic,and there is increased interest on how changes in metabolomics correlate with CD8^(+)T cell dysfunction.AIM To investigate whether and how glutamine metabolism affects the function of infiltrating CD8^(+)T cells in hepatocellular carcinoma.METHODS Immunohistochemical staining and immunofluorescence were performed on surgically resected liver tissues from patients.Differentially expressed genes in infiltrating CD8^(+)T cells in hepatocellular carcinoma were detected using RNA sequencing.Activated CD8^(+)T cells were co-cultured with Huh-7 cells for 3 d.The function and mitochondrial status of CD8^(+)T cells were analyzed by flow cytometry,quantitative real-time polymerase chain reaction,and transmission electron microscopy.Next,CD8^(+)T cells were treated with the mitochondrial protective and damaging agents.Functional alterations in CD8^(+)T cells were detected by flow cytometry.Then,complete medium without glutamine was used to culture cells and their functional changes and mitochondrial status were detected.RESULTS There were a large number of infiltrating PD-1+CD8^(+)T cells in liver cancer tissues.Next,we cocultured CD8^(+)T cells and Huh-7 cells to explore the regulatory effect of hepatoma cells on CD8^(+)T cells.Flow cytometry results revealed increased PD-1 expression and decreased secretion of perforin(PRF1)and granzyme B(GZMB)by CD8^(+)T cells in the co-culture group.Meanwhile,JC-1 staining was decreased and the levels of reactive oxygen species and apoptosis were increased in CD8^(+)T cells of the co-culture group;additionally,the mitochondria of these cells were swollen.When CD8^(+)T cells were treated with the mitochondrial protective and damaging agents,their function was restored and inhibited,respectively,through the mitochondrial damage and apoptotic pathways.Subsequently,complete medium without glutamine was used to culture cells.As expected,CD8^(+)T cells showed functional downregulation,mitochondrial damage,and apoptosis.CONCLUSION Glutamine deprivation impairs the function of infiltrating CD8^(+)T cells in hepatocellular carcinoma through the mitochondrial damage and apoptotic pathways.

Isolation and identification of CD4^+CD25^+ regulatory T cells in rat

Objective： To establish a stable and high efficient method for collection of CD4^＋CD25^＋ regulatory T cells from rats in vitro. Methods： CD4^＋CD25^＋ regulatory T cells were isolated from the rat splenic cells through two steps by magic cell sorting （MACS） system. The first step was negative selection of CD4^＋T cells by cocktail antibodies and anti-IgG magic microbeads, and the second step was positive selection of CD25^＋T cells by anti-CD25 PE and anti-PE magic microbeads. The purity and viability of separated cells were measured by flow cytometry （FACS） and Trypan blue staining. The suppressive ability of seperated cells on the proliferation of CD4^＋CD25^- T cells was assessed by cell proliferation assay. Results： The purity of negatively enriched CD4^＋ T cells was 79%-87% （83.6%±2.5% ） , and the purity of positively enriched CD4^＋CD25^＋ T cells was 86%- 93% （ 90.2±1.8% ） with the viability of 92%～95% （92.8% ± 3.4% ）. The enriched cells significantly suppressed the proliferation of CD4^＋CD25^- T cells in mixed lymphocyte culture （P 〈 0.05）. Conclusion： An effective method can be established for enrichment of CD4^＋CD25^＋ regulatory T cells in two steps by MACS, with satisfied cell purity, viability and function.

Effect of Homoharringtonine on Bone Morrow CD34^+CD7^+ Cells in Chronic Granulocytic Leukemia

Objective： To explore the effect of homoharringtonine （HHT） on bone morrow CD34^＋CD7^＋ cells in chronic granulocytic leukemia （CGL）. Methods： The changes of bone morrow CD34^＋CD7^＋ cells were observed after the treatment of HHT in 23 cases with CGL. The proliferation and apoptosis of CD34^＋CD7^＋ cells treated with HHT in vitro were studied. Results： The proportion of CD34^＋CD7^＋ cells in CGL （0.145±0.021） was higher than that of normal control （0.052±0.013）. The proportion of CD34^＋CD7^＋ cells in patients who got cytogenetic responses to HHT （0.072±0.020） decreased remarkably, but not in those patients who did not got cytogenetic responses to HHT, （0.137±0.023）. the proliferation of CD34^＋ cells was inhibited and the proportion of CD34^＋CD7^＋ cells decreased after cultured with HHT （0.134 in 24 h, 0.126 in 48 h and 0.102 in 72）. The apoptosis rate of CD34^＋CD7^＋ cells was higher than that in CD34^＋CDT cells （35.39%±4.39% versus 24.57%±4.01%, P〈0.05） 72 h after culture with HHT. Conclusion： The proportion of CD34^＋CD7^＋ cells in CGL was higher than that of normal control and HHT may inhibit the proliferation and induce apoptosis of bone marrow CD34^＋CD7^＋ cells.

CD4^+CD25^(high) Regulatory Cells in Peripheral Blood of NSCLC Patients

The proportion and changes of CD4^＋CD25^high regulatory T cells （Trs） in peripheral blood of non-small cell lung cancer （NSCLC） patients were analyzed and their clinical significance explored. The peripheral blood was collected from 61 patients with NSCLC and 15 healthy controls. By using monoclonal antibodies, the blood samples were evaluated with the flow cytometry for lymphocyte subsets （CD3^＋, CD4^＋ and CD8^＋） and CD4^＋CD25^high Tr cells. The results showed that the proportion of CD4^＋CD25^high Tr cells in NSCLC group was significantly higher than in control group [（4.36 ±2.07） % vs （2.04±1.03） %, P〈0.01]. The proportion of CD4^＋CD25^ high Tr cells in late stage was higher than that in early stage [stages Ⅰ ＋Ⅱ （2.264±0.6） %; stage Ⅲ（3.284± 1.38） %; stage IV （6.06 4±4.08） %] （P〈0.05）. Kaplan-Meier survival analysis revealed that the prognosis of the patients who had higher proportion of CD4^＋CD25^high Tr cells in peripheral blood was worse （P=0.0026）. In conclusion, the relative increase in CD4^＋CD25^high Tr cells in peripheral blood may be related to im- munosuppression and tumor progression in patients with NSCLC. This finding suggests that CD4^＋CD25^high Tr cells in peripheral blood of NSCLC may be positive for prognosis analysis. The use of depletion of the CD4^＋CD25^high Tr cell therapy to treat NSCLC patients may be an effective strategy.

Effect of Mg^(2+)level on the functions of CD8^(+)T lymphocytes and NK cells in patients with COVID-19

Objective:To investigate the changes of Mg^(2+) levels in serum and peripheral blood mononuclear cells(PBMCs)of patients with COVID-19 and its effects on the functions of CD8^(+)T lymphocytes and NK cells.Methods:A total of 165 COVID-19 patients hospitalized in Ezhou Central Hospital from January 20 to February 20,2020 were divided into mild/common group(98 cases)and severe/critical group(67 cases).At the same time,34 healthy persons were selected as the control group.Peripheral blood was collected and PBMCs were isolated,the level of Mg^(2+) in serum and PBMCs was detected.The subsets of CD8^(+)T lymphocytes and NK cell and the expression levels of their surface inhibitory molecular PD-1 and activator molecular NKG2D were detected by flow cytometry.The correlation between Mg^(2+) concentration and the expression levels of PD-1 and NKG2D was also analyzed.Results:Compared with the control group,the concentration of Mg^(2+) in serum and PBMCs,the counts of CD8^(+)T lymphocytes and NK cell in patients with mild/common and severe/critical groups were significantly reduced(P<0.05),while the expression level of surface inhibitory molecular PD-1 were significantly increased(P<0.05),while the expression level of the activation molecule NKG2D were significantly decreased(P<0.05).However,the changes of the above indicators in patients with severe/critical group were greater than those in the mild/common group(P<0.05).In addition,the Mg^(2+) concentration in COVID-19 patients was negatively correlated with the expression level of PD-1 on CD8^(+)T lymphocytes and NK cells(P<0.05),and positively correlated with the expression levels of NKG2D(P<0.05).Conclusion:The concentration of Mg^(2+) in the serum and PBMCs of COVID-19 patients is significantly reduced,which may cause the function of CD8^(+)T lymphocytes and NK cells to be inhibited.

CD68^+、CD163^+巨噬细胞对食管癌浸润及预后的影响

目的探讨CD68+、CD163+巨噬细胞对食管癌浸润及预后的影响。方法收集手术切除的食管癌组织标本84例、食管癌旁组织标本49例,采用免疫组化法检测组织标本中CD68+、CD163+巨噬细胞浸润密度,并对患者预后进行分析。结果食管癌组织中CD68+、CD163+巨噬细胞浸润密度每视野(35.69±14.50)个和(29.78±12.36)个,中位数分别为32.8个和27.1个;癌旁正常组织中CD68+、CD163+巨噬细胞浸润密度每视野(4.26±2.30)个和(2.33±1.71)个,中位数分别为3.4个和2.2个。食管癌组织中CD68+、CD163+巨噬细胞浸润密度明显高于癌旁正常组织(P均<0.05)。TNMⅢ期者、有淋巴结转移者、低分化程度者CD68+、CD163+巨噬细胞浸润密度明显高于TNMⅠ~Ⅱ期者、无淋巴结转移者、高中分化程度者(P均<0.05);患者术后1年、3年、5年生存率分别为83%,67%,46%,中位生存时间为44.21个月。Spearman相关分析显示,CD68+、CD163+巨噬细胞浸润密度与患者生存时间呈现负相关性(P<0.05);多因素Cox回归分析TNM分期、淋巴结转移、分化程度、CD163+巨噬细胞浸润密度是影响患者预后的独立危险因素(P均<0.05)。结论食管癌患者局部病灶组织中CD68+、CD163+巨噬细胞浸润密度明显增高,CD163+巨噬细胞浸润密度增高是影响患者预后的独立因素。

人外周血CD68^+单核细胞体外的分选及诱导分化为破骨细胞

目的建立高纯度人活性破骨细胞方法,用于对破骨细胞生物化学和分子生物学的研究。方法用免疫磁珠法在人类外周血单核细胞中分选出CD68+细胞,用流式细胞仪分析分选效能,体外在10-8 mol/L地塞米松及25 μg/L巨噬细胞集落刺激因子的培养条件下用16 μg/L可溶性核因子κB受体活化子配体诱导(s-RANKL)分化,用降钙素受体抗体免疫组化染色及抗酒石酸磷酸酶染色鉴定,扫描电镜观察骨片贴壁细胞及骨吸收陷窝。结果分选获得CD68+单核细胞纯度为(93.06±0.61)%(n=4),经核因子κB受体活化子配体诱导后降钙素受体抗体免疫组化染色阳性,抗酒石酸磷酸酶染色,扫描电镜观察有骨吸收陷窝形成。结论人外周血单核细胞中CD68+细胞是破骨前体细胞,在RANKL在诱导下分化为成熟破骨细胞。

CD68^+巨噬细胞表达与肺腺癌侵袭性及临床病理分期相关性分析

目的通过肺腺癌肿瘤实质中CD68^+巨噬细胞浸润情况的分析,及肿瘤实质中Ki67指数表达情况,探讨肿瘤相关巨噬细胞(TAMs)与肺腺癌临床病理分期及肿瘤侵袭能力相关性。方法选取南京市胸科医院2017年间130例肺腺癌组织切片,应用免疫组织化学技术观察并计数CD68^+巨噬细胞在肺腺癌肿瘤实质及间质浸润情况及Ki67指数表达情况分析其相关性。结果(1)肺腺癌肿瘤实质内TAMs表达低于肿瘤癌旁间质内TAMs表达(P<0.05)。(2)肺腺癌肿瘤实质及间质内TAMs表达与Ki67表达有相关性(P=0.0416)。(3)Ⅲ~Ⅳ期NSCLC肿瘤实质组织中CD68^+阳性TAMs表达显著高于Ⅰ~Ⅱ期,差异有统计学意义(P<0.05)。结论结果显示CD68+巨噬细胞浸润情况与肿瘤TNM分期、及Ki67表达有一定相关性。肿瘤相关巨噬细胞(TAMs)浸润肿瘤实质及间质导致肺内炎症微环境,与肿瘤侵袭能力正相关。

肝癌组织中CD68^+肿瘤相关巨噬细胞数量与Ki-67蛋白表达及原发性肝癌预后的关系

目的探讨原发性肝癌(HCC)患者肝癌组织中CD68+肿瘤相关巨噬细胞(CD68+TAM)的数量与增殖指标Ki-67及肝癌预后的关系,寻找评估肝癌预后更可靠的指标。方法应用免疫组织化学SABC法检测73例HCC组织中CD68^+TAM的分布情况及增殖指标Ki-67蛋白表达,运用化学发光测定法检测血清AFP水平。结果 CD68^+TAM高密度组Ki-67阳性率明显高于低密度组(P=0.0191)。CD68^+TAM数量与HCC患者的年龄、性别无关(P>0.05),而与HCC病理分级有关(P=2.83E-04)。血清AFP水平与HCC患者的年龄、性别、病理分级无关(P>0.05)。CD68^+TAM高密度组的总生存时间显著短于低密度组(P=0.0004)。而AFP高水平组与低水平组比较,总生存时间、Ki-67阳性率差异均无统计学意义(P>0.05)。结论 HCC患者肝癌组织中CD68^+TAM数量与肝癌增殖活性密切相关,是HCC预后的独立危险因素。与AFP相比,CD68+TAM有望成为评估肝癌预后更可靠的指标。

CD45RO^+T细胞和CD68^+细胞在外阴上皮内非瘤样病变和外阴癌中的侵润

目的:探讨CD45RO+T细胞和CD68+细胞在外阴上皮内非瘤样病变和外阴癌发生、发展过程中的侵润及作用。方法:免疫组化S-P法分别检测CD45RO+T细胞、CD68+细胞在各20例外阴鳞状上皮细胞增生、外阴硬化性苔藓、外阴鳞状上皮癌病变中的侵润,取10例外阴正常皮肤作对照。结果:CD45RO+T细胞和CD68+细胞在外阴正常皮肤中侵润水平极低,明显低于外阴鳞状上皮细胞增生、外阴硬化性苔藓和外阴鳞状细胞癌病变(P<0.05)。它们在外阴硬化性苔藓晚期病变中侵润明显高于早期病变(P<0.05);在中高分化外阴鳞状细胞癌组织中侵润明显高于低分化外阴鳞状细胞癌组织(P<0.05)。CD45RO+T细胞在中高分化外阴鳞状细胞癌组织中侵润最高,其次是晚期硬化性苔藓(P<0.05);CD68+细胞在中高分化外阴鳞状细胞癌和晚期硬化性苔藓病变中侵润最高,但二者之间没有统计学差异(P>0.05);它们均明显高于外阴鳞状细胞增生、低分化外阴鳞状细胞癌和早期硬化性苔藓(P<0.05),而在后三者之间侵润没有明显差异(P>0.05)。CD68+细胞和CD45RO+T细胞在外阴不同病变中侵润具有正相关性,相关系数为0.742(P<0.001),而且CD68+细胞多出现在CD45RO+T细胞的周围,且多在病变浅层。结论:CD45RO+T细胞和CD68+细胞参与皮肤的免疫反应,它们相互协同作用,在外阴上皮内非瘤样病变和外阴癌发生发展过程中发挥免疫调节作用。

Effects of estrogen on CD4^+ CD25^+ regulatory T cell in peripheral blood during pregnancy

Objective To investigate the effects of estrogen(E_2)level on regulatory T cells(Treg)in peripheral blood during pregnancy.Methods:A total of 30 healthy non-pregnant women were selected as control group,90 pregnant women of early,middle and late pregnancy and 30 postpartum women at 1 month after parturition were selected as experimental groups including early pregnancy group,middle pregnancy group and late pregnancy group;the proportions of CD4^+CD25^+Treg and CD4^+CD25^+CD127^-Treg among CD4 T cells were detected by flow cytometry;the serum estrogen content in peripheral blood was detected by electrochemical immune luminescence method.Results:E_2 level was coincident with the change of Tregs number during pregnancy.The estrogen content in peripheral blood increased gradually from early pregnancy to late pregnancy,then decreased significantly after parturition,and the level at 1 month after parturition down to the level in non-pregnancy group(P>0.05);the level of E_2 in pregnancy groups were significantly higher than those in non-pregnancy group(P<0.01);and there were significant differences among early pregnancy group,middle pregnancy group and late pregnancy group(P<0.05).The proportions of CD4^+CD25^+Treg and CD4^+CD25^+CD127^-Treg in pregnancy groups were significantly higher than those in non-pregnancy group(P<0.05),but decreased significantly after parturition,and there was no significant difference between non-pregnancy group and postpartum women group(P>0.05):the proportions in middle and late pregnancy groups were significantly higher than those in early pregnancy group(P<0.05).but decreased slightly in late pregnancy group,there was no significant difference between late pregnancy group and middle pregnancy group(P>0.05).There was correlation between Tregs number with estrogen level during pregnancy.The proportion of CD4^+CD25^+Treg and CD4^+CD25^+CD 127^-Treg were positively correlated with estrogen level.Conclusions:High proportion of CD4^+CD25^+Trcg and CD4^+CD25^+CD127^-Treg is closely related to the high level of E,during pregnancy.It suggested that high level of estrogen may induce an increase of CD4^+CD25^+Treg in peripheral blood.and then influence the immune function of pregnant women.The results of this experiment might play an important role of estrogen in immune-modulation during pregnancy.

肿瘤相关巨噬细胞及CD8^(+)/CD68^(+)细胞比值是影响肺腺癌患者预后的独立危险因素

目的:探讨CD68^(+)肿瘤相关巨噬细胞(TAM)、CD8^(+)T细胞、Foxp3^(+)Treg细胞等在肺腺癌(LUAD)组织中浸润分布及其与患者预后的关系。方法:收集2004年9月至2009年4月间在苏州大学附属第三医院手术切除的93例LUAD组织及78例癌旁组织,采用组织芯片(TMA)及多重免疫荧光(mIF)技术检测其中的免疫细胞浸润与分布,Wilcoxon秩和检验比较癌与癌旁组织、癌巢与间质中浸润水平的差异,χ^(2)检验分析其浸润水平及CD8^(+)/CD68^(+)细胞比值与临床病理特征的关系,Kaplan-Meier法和COX模型分析影响患者OS的潜在危险因素。结果:与癌旁组织比较,癌组织中CD68^(+)TAM、CD8^(+)T细胞、Foxp3^(+)Treg细胞浸润水平均显著增加(均P<0.01),间质CD68^(+)TAM、CD8^(+)T细胞的浸润水平均显著高于癌巢(均P<0.01)。总CD68^(+)TAM、癌巢及间质CD68^(+)TAM浸润水平与淋巴结转移呈正向关联(均P<0.05),癌巢CD68^(+)TAM浸润水平与T分期呈正向关联(P<0.05),间质CD68^(+)TAM浸润水平与病理分级呈正向关联(P<0.05);癌组织中CD8^(+)/CD68^(+)细胞比值与病理分级、淋巴结转移均呈负向关联(均P<0.05)。Kaplan-Meier生存分析显示,LUAD组织中总CD68^(+)TAM、癌巢及间质CD68^(+)TAM高浸润患者OS均短于低浸润患者(P<0.05或P<0.01)、癌组织中CD8^(+)/CD68^(+)细胞比值高患者OS显著长于低比值患者(P<0.05)。多因素COX模型分析示,LUAD患者年龄、TNM分期及癌组织中CD8^(+)/CD68^(+)细胞比值是影响患者预后的独立风险因素(P<0.05或P<0.01)。结论:高度浸润的CD68^(+)TAM与LUAD的进展、侵袭、转移和不良预后显著关联,而高CD8^(+)/CD68^(+)细胞比值是影响LUAD患者OS的独立保护因素。

Specific CD8^+ T cell response immunotherapy for hepatocellular carcinoma and viral hepatitis

Hepatocellular carcinoma(HCC), chronic hepatitis B(CHB) and chronic hepatitis C(CHC) are characterized by exhaustion of the specific CD8^+ T cell response. This process involves enhancement of negative costimulatory molecules, such as programmed cell death protein-1(PD-1), cytotoxic T-lymphocyte antigen-4(CTLA-4), 2B4, Tim-3, CD160 and LAG-3, which is linked to intrahepatic overexpression of some of the cognate ligands, such as PD-L1, on antigen presenting cells and thereby favouring a tolerogenic environment. Therapies that disrupt these negative signalling mechanisms represent promising therapeutic tools with the potential to restore reactivity of the specific CD8^+ T cell response. In this review we discuss the impressive in vitro and in vivo results that have been recently achieved in HCC, CHB and CHC by blocking these negative receptors with monoclonal antibodies against these immune checkpoint modulators. The article mainly focuses on the role of CTLA-4 and PD-1 blocking monoclonal antibodies, the first ones to have reached clinical practice. The humanized monoclonal antibodies against CTLA-4(tremelimumab and ipilimumab) and PD-1(nivolumab and pembrolizumab) have yielded good results in testing of HCC and chronic viral hepatitis patients. Trelimumab, in particular, has shown a significant increase in the time to progression in HCC, while nivolumab has shown a remarkable effect on hepatitis C viral load reduction. The research on the role of ipilimumab, nivolumab and pembrolizumab on HCC is currently underway.