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Objective:To explore the role and mechanism of the B7-H1 expressed by auto-keratinocytes in the intermingled skin grafting model in vitro(MELC). Methods:The intermingled skin grafting model(MELC) was established in vitro. Flow cytometry was performed to detect the expressions of B7-H1 in keratinocytes. The expressions of PD-1 in lymphocytes were measured at the same time. The levels of IL-10,Foxp3 and GATA-3 mRNA in lymphocytes were detected by Real-time quantitative PCR. Results: Through flow eytometry,in the MELC with auto-keratinocytes,the expression of B7-H1 in auto-keratinocytes and the PD-1 in lymphocytes were rising trend and the rising rate was in time-dependent manners(P<0. 01). RT-PCR assay indicated that the relative levels of IL-10, Foxp3,GATA-3mRNA expression were significant raised and the rising rate was in time-dependent manners (P<0. 01). Conclusion:In the intermingled skin grafting model,the auto-keratinocytes could express B7-H1 to enhance the expression of PD-1 in T cells. When B7-H1 combined with PD-1,the Th2 cells and Foxp3+Tregs were induced and suppressed the immune response in the intermingled skin grafting model.
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Objective:To investigate the possible role of NKT cells in unexplained recurrent spontaneous abortion by measuring the NKT cell numbers,maturity and cytokine secretion of the placenta of mice with unexplained recurrent spontaneous abortion.Methods:Normal pregnancy model in hybrid by feeding CBA / J and BABL/C in a cage,and the model of unexplained recurrent spontaneous abortion was established by feeding CBA / J and DBA2/J in a cage.The number of NKT and CD3~+T cells was determined by flow-cytometry;Th1/Th2-relative cytokines were assayed by ELISA and T-bet expression was determinded by fluorescence quantitative PCR.Results:There was not significant change of CD3~+ T cells when compared between normal pregnancy and unexplained recurrent spontaneous abortion group (P＞0.05).In the course of normal pregnancy,the IFN-γ secreted by placenta lymphocytes decreased gradually,accompanied by the decline of NKT cell number and the proportion of mature cells;whereas in the course of unexplained recurrent spontaneous abortion,it was on the opposite.There was significant difference of T-bet mRNA expression between the two groups.T-bet mRNA expression was related to the proportion of mature NKT cells or placenta IFN-γ secretion by lymphocytes.Conclusion:Unexplained recurrent spontaneous abortion may be related to NKT cells disorders,NKT cells are of low-mature proportion and inadequate secretion of IFN-γ during early pregnancy,whereas are shown high-mature proportion and excessive secretion of IFN-γ during latter pregnancy;the anomaly of T-bet mRNA expression may be one of the factors leading to NKT cells disorder.
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<p><b>OBJECTIVE</b>To observe the immune reaction in the mixed culture of host lymphocytes with allogenic and host endothelial cells.</p><p><b>METHODS</b>The host epithelial cells and lymphocytes from burn patients and allogenic epithelial cells were mix-cultured in different ratios, so as to simulate the local immune micro-environment of host skin island in intermingled skin grafting. In addition, the cells from normal human subjects were also mix-cultured as control. The lymphocyte cpm values were detected by (3)H-TdR and HLA molecules and T cell subgroup were determined by immunohistological technique.</p><p><b>RESULTS</b>(1) The lymphocyte proliferation reaction could be effectively inhibited by the epithelial cells from burn patients but not from normal control. (2) The inhibition of host lymphocyte proliferation could not be mediated by the HLA-DQ molecules of epithelium from burn patients. (3) The positive expression rate of HLA-DR of epithelia from burn patients was evidently higher that that from normal control (P < 0.05), (4) The CD8 expression of lymphocyte in burn patients was significantly higher than that in normal control (P < 0.01), while the CD4 expression in burn patients was lower than that in normal control (P < 0.01). But there was no obvious difference of the CD3 expression between patients and normal subjects (P > 0.05).</p><p><b>CONCLUSION</b>The lymphocyte proliferation reaction could be obviously inhibited by the host epithelium, which might be related to the specific immune state of the host lymphocytes and epithelium of burn patients.</p>
Subject(s)
Humans , Cell Communication , Allergy and Immunology , Physiology , Cell Culture Techniques , Cell Division , Epithelial Cells , Allergy and Immunology , Physiology , Lymphocytes , Allergy and Immunology , Physiology , Skin Transplantation , Allergy and ImmunologyABSTRACT
Aim To explore the mechanism of the immune tolerance induced by the auto-islets in the intermingled skin grafting. Methods The mixed epidermal cell and lymphocyte culture (MELC) was established, in which auto-keratinocytes were added to mimic the effect exerted by the auto-islets in vivo. Results Auto-keratinocytes could inhibit the lympho-proliferation induced by alloantigen. Conclusion The immune tolerance induced by auto-islets may be related to the function of auto-keratinocytes as antigen presenting cells.
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Objective:As the function of ??T cells and NK in the immunological therapy is showing more and more important,the characteristics of ?? T cells,NK and LAK were analyzed comparatively after that they were richened by isolating and incubating in vitro.Methods:The cells were collected using MACS after the cells were panned respectively with special monoclonal antibodies.Then the characteristics including proliferation,phenotype,cytotoxin and the down regulation blocked by specific antibodies were analyzed.Results:The ?? T cells can expand 600～800 times after culturing 2 weeks and the percent of CD3,CD8 and ?? expressed on the collected cells were 72.29%,58.02% and 65.98% respectively ?? T cells showed the high cytotoxin to K562(NK sensitive cell line),Raji and XG 7 cell lines(NK non sensitive cell lines).The percentage of cytotoxin reached 35.98%,52.27% and 69.08% respectively compared with ??T respectively.No obviously change of percent of ??T cytotxic ability to these target cells were observed using special MHC class I monoclonal antibody to block the ??T cell before coculturing the target cells with ?? T cells.Conclusion:All of ?? T cells,NK and LAK showed the non specific cytotoxin to tumor cells.The cytotoxic capability of ?? T cells did not be effected after blocking with MHC class I monoclonal antibody.These results demonstrated that ?? T cell could kill more kinds of tumor cells than NK and LAK.
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Objective: Full expression of class I HLA molecules on tumor cells is pivotal in priming the tumor antigen-specific CTLs for effective iramunotherapies. Tumor cells display abnonnal expression of HLA class I molecules in human ovarian carcinoma. Methods: In this study, the expression of class I molecules in human ovarian cancer cells was determined by using Western bloting, immunohistochemistry testing and flow cytometry. The mRNA of TAP (transporter associated with antigen processing) and LMP(low molecular weight polypeptide) were exannined at the same time by RT-PCR. Results: It has been shown that class I molecules were unable to be, or only weakly, expressed in five of eight ovarian cancer cell lines. On these cell lines abnormal in class I expression, no or only little mRNA were detected for TAP1, TAP2, LMP2 and(or) LMP7 genes. Class I expression, however, could be partially recovered by incubating the tumor cells at 25℃ when compared with those at 37℃ . Conclusion: The results suggested that the dysfunction of TAP and IMP genes, following by a defective expression of class I molecules, might be one of the mechanisms enable ovarian cancer cells to escape from immune surveillance.
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When 56 unrelated subjects in Shanghai were typed for HLA-A, B, C, D, DR and DQ with the homozygous typing cells (HTCs) and sera of 3 rd Asia-Oceania Histocompatib(?)l(?)tv Workshop Conference, only five of the eleven Dw 3 positive subjects were identified to be associated with the correspondingly designated DR 3. For the other six subjects, Dw 3 were co-expressed with Dw 7 c (Dw7+Dw17) and DR7 in the same halotype, resulting in a Dw "triplet" when the third D specificity was detected in the other halotypes. The Dw 3 was thus divided into two forms characterized by the two different haplotypes in which it was involved: HLA-B17-DR 3-DRw52-Dw3 and HLA-B13-DR 7-DR w53-DQ w2-Dw7 c-Dw3. With circumstancial evidences, the co-expression of Dw 3 and Dw 7 c was attributable to a tentative DQw 2 split in Chinese, which, as a part of the antigen epitopes, might take part In the expression of Dw3 and Dw7 c specificities and could be recognized by the HTCs.
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Objective:To lay background for studying rejection mechanisms in xenotransplantation, class II DQA genes of swine leukocyte antigens (SLA) of three Chinese pig strains GXP, GZP and YNP were cloned and sequenced.Methods:RT-PCR was performed to proliferate SLA-DQA genes (GXPDQA, GZPDQA and YNPDQA),which were cloned into pGEM-T Easy vector for sequencing and analysis.Results:All three allelic sequences examined, are not identical to those reported, which allows the sequences receiving their accession numbers from GenBank as AY102473, AY102474 and AY102475, which encompass an open reading frame of 765, 768 and 768 nucleotides respectively.Conclusion:Three new alleles of SLA-DQA genes were obtained. Analysis of the amino acid sequences of swine, human and mouse DQA (or equivalent) also indicated that GXPDQA, GZPDQA and YNPDQA were more similar to human DQA than mouse H-2-A?, which might provide an understanding of how the immune system evolved.
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Trichosanthin (Tk), a crystal plant protein isolated and purified from root tubes of a Chinese medicinal herb Trichosanthes kiritowii Maxim (Cucurbitaceae), was demonstrated to be capable of inducing immune suppression in man, and the suppression could be eased up by depleting the culture of CD 8~+ cells with monoclonal anti bodies as reported in our previous work (1). This paper desc ribes that the diminition of suppression,or the restoration of low- responsiveness, however, was restricted in certain subjects we studied. For the others, the suppression kept unchanged or even got more vigorous with the same treatment. For this reason, healthy subjects were categoried as CD 8 -mediators (M~+) and non-mediators (M ~- ). In addition, there were some interesting points revealed as follows. 1 ) M~+ and M~- were determined as a phenotipically stable and reproduceable traits; 2) The difference between two categories can only be observed in the MLR- Tk testing system but not in the non-specific PHA- Tk system in which PHA initiated lymphoproliferation was depressed by T K; 3 ) Instead of stimu lator, only responding PBMC in one-way MLC was affectad by depleting CD8 cells, which resulted in the distinguishable phenotypes of M~+ and M~-; 4) The supernatants collected from Tk sensitized. cultured PBMC could show suppressive activity if the cell donors were M~+, but not M~-.