目的 通过观察卵巢早衰 POF 患者外周血CD4+CD25+调节性T细胞 Treg 及干扰素-γ（IFN-γ）、转化生长因子-β(TGF-β)的变化，探讨POF的免疫学发病机制。 方法 收集2011年12月－2012年9月就诊的POF患者17例，卵巢储备功能减退 DOR 患者11例，以及生殖中心健康育龄女性16例，流式细胞仪定量检测外周血Treg数量，Elisa方法检测血清IFN-γ、TGF-β的水平，并以FSH/LH评价卵巢储备功能，进行相关性分析。 结果 与对照组相比，POF组和DOR组IFN-γ水平增高 P＜0.01 、TGF-β水平降低 P＜0.01 ，POF患者及DOR患者Treg比例降低 P＜0.01 ，IFN-γ的增高与卵巢储备功能的下降呈显著正相关 r=0.70，P＜0.01 。 结论 Treg 和IFN-γ、TGF-β水平与卵巢早衰密切相关，IFN-γ对评估卵巢储备功能、预测卵巢早衰具有参考价值。
Objective To investigate the expression of Human leucocyte antigen(HLA)-DP, -DQ, -DR and CD40 in human retinal pigment epithelial (RPE) cells, to determine their molecule expression in immune response process, and their abilities to stimulate T lymphocyte activation. Methods Human RPE cells were cultured with or without (IFN respectively. Expression of HLA-DP, -DQ, -DR and CD40 was measured by immunohistochemical staining. Meanwhile, peripheral blood mononuclear cells (PBMC) were cocultured with RPE cells in vitro, and then the expression of activated lymphocytes CD69 was measured by fluorescence activated cell sorter(FACS). Results Expression of HLA-DP, -DQ, -DR and CD40 antigen were enhanced by gamma;-interferon inducement. Increasing amount of CD69 positive lymphocytes were found in the co-culture system of RPE cells and PBMC. Conclusion T-lymphocytes in the peripheral blood were activated by human RPE cells which is antigen presenting cells with immunological characteristics potential.
ObjectiveTo summarize the clinical, radiological and pathological characteristics of acquired immune deficiency syndrome (AIDS) combined with Pneumocystis carinii pneumonia (PCP), so as to improve the clinicians' understanding of the disease. MethodsThe clinical data of 50 AIDS patients combined with PCP admitted between February 2006 and May 2015 were retrospectively analyzed, including medical history, physical signs, laboratory examination, chest high resolution CT (HRCT), pathological characteristics, treatment and prognosis, etc. ResultsThe clinical features of AIDS patients combined with PCP included cough, dyspnea and fever, without obvious positive signs in the lung.The patients were divided as a mild group, a moderate group and a severe group according to the levels of PaO2.There was significant difference among three groups in serum albumin level [(23±3) g/L vs. (30±5) g/L and (28±6) g/L, P < 0.01].There were no significant differences among three groups in CD4+ T lymphocyte and lactate dehydrogenase (LDH) (P > 0.05).The typical chest radiograph feature of HRCT was ground-glass shadows in both lungs, and may be associated with reticular shadows or "gravel sign" and cyst.Of 50 patients, 16 patients were diagnosed via pathology of transbronchial lung biopsy(TBLB) and only 5 patients were diagnosed via silver staining of the bronchoalveolar lavage fluid (BALF).The other patients were clinically diagnosed.100% of the patients were treated with sulfamethoxazole (SMZco), 64%with caspofungin, and 72% with glucocorticoid.All the patients relieved with no death in hospital. ConclusionWhen a patient got cough, dyspnea and fever, especially ground glass on HRCT in both lungs, AIDS combined with PCP should be highly considered, and diagnostic treatment with SMZco and CD4+ T lymphocyte measurement should be conducted as soon as possible, so as to reduce misdiagnosis and mortality.
Objective To observe the turbulence of regional intra-artery implantation chemotherapy and peripheral venous chemotherapy on immunologic function of patients. Methods Two weeks after radical operation of gastric carcinoma, chemotherapy was performed. Eighty-three patients were divided into two groups, one (42 patients) received peripheral venous chemotherapy (PVC) and the other (41 patients) received regional intra-artery implantation pump chemotherapy (RAIPC). The serum T-lymphocyte subsets and immunoglobulin level before and 1-4 days after the chemotherapy were measured. Results After PVC, proportion of CD3 and CD4, CD4/CD8 and IgG, IgA, IgM concentration in PVC group were significantly decreased compared with those before PVC （P＜0.05， P＜0.01）, and it is the same when compared with postRAIPC patients except for CD4 and IgM （P＜0.05）. In RAIPC group, there were no significant changes in proportion of CD3, CD4 and CD8, CD4/CD8 and IgG, IgA concentration between pre- and post-RAIPC patients. Conclusion After radical operation of gastric carcinoma, RAIPC affects the immunologic function more moderate than PVC.
Objective To observe the therapeutic effect of mensenchymal stem cells (MSCs) for experimental autoimmune uveitis (EAU). Methods MSCs were obtained from Wistar rats and selected by plastic adherence. Lewis rats were divided into treatment group and control group, six rats in each group. EAU models were induced by immunization with an emulsion (0.2 ml) containing 30 mu;g interphotoreceptor retinoid-binding protein derived peptide R16 and complete Freundprime;s adjuvant. The clinical manifestations of two groups were observed. Nine to 11 day after modeling, 1 ml MSCs suspension, which contained 5times;106 MSCs, were injected into the rats in treatment group via tail vein, and the rats in control group were given equal volume of phosphate buffer solution. Fifteen day after modeling, the eyes were collected to test the proportion of interferon gamma;, interleukin-17 and Foxp3 positive cells by flow cytometry. The clinical scores were analyzed by mixed linear model and statistical analysis of variance of repeated measurement data. The results of flow cytometry were analyzed using independent-sample t test. Results Six days after immunization, mild dilatation and congestion of iris vascular was observed. Nine days after immunization, mild muddy anterior chamber, myosis and absent pupillary reaction to light were observed. Twelve days after immunization, muddy anterior chamber, occlusion of pupil and dimmed or disappeared red reflex were observed, and then inflammation was slowly reduced. From 11 to 15 days after immunization, the clinical score of treatment group was lower than that in control group, the difference was statistically significant (t=2.42, 2.21, 4.16, 5.24, 4.03; P＜0.05). The results of flow cytometry showed that MSCs treatment could decrease the proportion of CD4＋T cells, Th1 cells and Th17 cells, increase the proportion of Treg cells. Conclusion MSCs treatment can ameliorate EAU, up-regulate the expression of Treg cells and down-regulate the expression of CD4＋T cells, Th1 cells and Th17 cells.
ObjectiveTo evaluate the effects and mechanism of indoleamine 2, 3-dioxygenase (IDO) modified rat bone marrow mesenchymal stem cells (BMSCs) in composite tissue allograft rejection. MethodsBMSCs isolated from Brown Norway (BN) rats (aged, 4-6 weeks) were infected by IDO[green fluorescent protein (GFP)]-lentivirus. The high expression target gene and biological activity cell line (IDO-BMSCs) were screened. IDO mRNA and protein expressions were detected by RT-PCR and Western blot. The biological activity of IDO in supernatant was detected by measuring the amount of kynurenine generation. In mixed lymphocyte reaction system, different numbers of IDO-BMSCs mixed with responding cells (peripheral blood mononuclear cell isolated from 4-6-week-old LEWIS rats, as recipient) and stimulating cells (peripheral blood mononuclear cell isolated from BN rats, as donor), with the cells ratios of 1:5:5, 1:10:10, 1:50:50, and 1:100:100 (as experimental groups 1, 2, 3, and 4, respectively). Each reaction system was blocked by 1 mmol/L 1-methyl-tryptophan (1-MT) (IDO specific inhibitor). IDO-BMSCs mixed with responding cells (1:5) as the negative control group, responding cells mixed with stimulating cells (1:1) as positive control group; and IDO-BMSCs were cultured in RPMI 1640 medium alone as blank control group. MTT assay was used to detect the T lymphocytes proliferation at 5 days. Furthermore, GFP-BMSCs (group A), IDO-BMSCs (group B), and normal saline (group C) were infused via the tail vein of allogeneic limb transplantation rats, and graft survival time and rejection were observed in each group. ResultsThe IDO expression of BMSCs after genetic modification was higher than that before genetic modification. IDO-BMSCs could significantly improved kynurenine concentration in culture medium supernatant when compared with GFP-BMSCs (P<0.05). Before adding 1-MT, with the ratio of IDO-BMSCs to responding cells decreased, T lymphocytes proliferation rate increased in experimental groups 1, 2, and 3, showing significant differences between groups (P<0.05); there was no significant difference between experimental group 4 and the positive control group (P>0.05). After adding 1-MT, T lymphocytes proliferation rate was significantly higher than that before adding 1-MT in the other experimental groups (P<0.05) except experimental group 4 (P>0.05). In vivo, IDO-BMSCs could promote colonization in allograft, inhibit transplantation rejection, and prolong survival time of composite tissue allograft; the survival time of composite tissue allograft was (11.5±0.6) days in group A, (14.5±0.8) days in group B, and (9.0±0.3) days in group C, and it was significantly longer in group B than in groups A and C, and in group A than in group C (P<0.05). ConclusionIDO-BMSCs can promote the survival of allogeneic composite tissue grafts in rats, and its mechanism may involve in inhibition of T lymphocytes proliferation and promotion their own colonization in allograft.
Objective To observe the proportion changes of CD4+CD25+FOXP3+ T cells in peripheral blood of patients with VogtKoyanagiHarada disease (VKH) before and after one month of treatment. Methods he peripheral blood samples from 15 patients with VKH disease before and after one month of treatment by glucocorticoid, and from 15 healthy volunteers were collected，and lymphocytes were separated from them. CD4+CD25+ regulatory T cells were labeled by antibodies of cell surface marker CD4、CD25 and transcription factor FOXP3. The proportion of CD4+CD25+FOXP3+ T cells were detected by flow cytometry. Results Before the treatment, the percentage of CD4+CD25+FOXP3+ T cells in periphery blood was（0.30plusmn;0.19）% of CD4+ cell in VKH patients, and（1.41plusmn;0.52）% in control group, the difference was statistically significant（t=7.665,Plt;0.01）; after one month of treatment, the VKH patients group was（1.28plusmn;0.54）% which close to the control group. However there were two patients whose CD4+CD25+ T cells increased extraordinarily after one month of treatment. Conclusions The proportion of CD4+CD25+ FOCP3+ T cells in periphery blood in VKH patients were lower than control group obviously before treatment, but were close to control group after treatment. Those results indicated that VKH diseases may be associated with the decreased proportion of CD4+CD25+ regulatory T cells.
ObjectiveTo investigate the method for generating anchor chemric T lymphocytes that can target tumor associated glycoprotein-72 (TAG72) antigen and analyze their repressive effects on proliferation of TAG72 positive hepatocarcinoma cells. MethodsFirstly, peripheral blood mononuclear cells (PBMCs) from healthy volunteers were isolated. And then, CD8+ T cells were isolated from PBMCs via magnetic activated cell sorting (MACS). These lymphocytes were transfected with recombinant vector, anti-TAG72-scFv-CD28-pcDNA3, through Lipofectamine2000 to gernerate anchor chimeric TAG72-specific CD8+ T cells. SMMC7721 (TAG72 positive) hepatocarcinoma cells were co-cultured with chimeric T lymphocytes and their cell cycles were analyzed by flow cytometry (FCM). ResultsAnchor chmeric T lymphcytes targetting TAG72 recognized TAG72 positive SMM7721 cells and repressive effects on their proliferation were observed by flow cytometry. ConclusionAnchor chmeric T lymphcytes targetting TAG72 on tumor surface can specifically recognize TAG72 positive hepatocarcinoma cells and may exert repressive effect on their proliferation.
Objective To observe the effects of Thymosin α1 （Tα1） on acute rejection after liver transplantation and immune function of T cells. Methods Twenty recipients of liver transplantation due to primary hepatic carcinoma were divided into two groups: Tα1 group (n=10) and control group (n=10). Tα1 group received subcutaneous injection of Tα1 1.6 mg on the first day after liver transplantation and then twice a week for at least one month. Both Tα1 group and control group took same immunodepressants. Core biopsies were carried to compare the incidence rate of acute rejection between Tα1 group and control group. Peripheral T cellular immune function in these two groups was detected on 1 d before, 1 week, 2 weeks and 1 month after transplantation. Results There was not significant difference of incidence rate of acute rejection between Tα1 group and control group (Pgt;0.05). In the Tα1 group, CD4+, CD8+ lymphocyte cell counts and the CD4+/CD8+ ratio were significantly higher than those in the control group in 2 weeks and 1 month after transplantation (P＜0.05). Conclusion Use of Tα1 in recipients who also takes rountine immunosuppressants dose not increase the risk of occurring acute rejection after liver transplantation. Tα1 can significantly increase CD4+, CD8+ counts and CD4+/CD8+ ratio, which shows that Tα1 may improve recipients’ cellular immune function.