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首页> 外文期刊>Journal of Huazhong University of Science and Technology >Paired Immunoglobin-like Receptors A and B Are New Targets for Inducing Dendritic Cells Tolerance in Mice
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Paired Immunoglobin-like Receptors A and B Are New Targets for Inducing Dendritic Cells Tolerance in Mice

机译：配对的免疫球蛋白样受体A和B是诱导小鼠树突细胞耐受性的新目标

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摘要

The expression of paired immunoglobin-like receptors A (PIR-A) and B (PIR-B) and their relationship with tolerogenic dendritic cells (T-DC) in mice were investigated. The mouse DCs line, DC2.4 cells were cultured with the recombinant murine interleukin-10 (IL-10) and recombinant human transforming growth factor β1 (TGF-β1) respectively to develop the T-DC and stimulated with lipopolysaccharide (LPS) for 48 h to induce the mature dendritic cells (LPS-DC). Special small interfering RNAs (siRNA) molecule for PIR-B was chemically synthesized and transfected into DC2.4 cells (Si-DC) by lip2000. The expression of PIRs on DC2.4 cells were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR), flow cytometry (FCM) and Western blot. Realtime reverse transcriptase-polymerase chain reaction (Realtime-PCR) was applied for measurement of PIR-A and CD80, CD86, MHC-Ⅱ mRNA expression. The allogeneic stimulating capacity of DCs was measured by mixed lymphocyte reaction (MLR) using ~3H-thymidine incorporation test. The concentration of IFN-y in supernatants of MLR from distinct groups was analyzed by ELISA. The results showed that PIR positive rate was (28.65±8.12)% examined by FCM on DC2.4 cells. PIR positive rate was increased dramatically to (54.21±6.34)%, (58.78±4.70)%, (48.24±6.75)% respectively for IL-10, TGF-β1 and LPS induction (P< 0.01), but there was no significantly different among the three groups (P> 0.01). The semi-quantitative RT-PCR and Western blot revealed that IL-10 and TGF-β1 induced the higher PIR-B level and lower PIR-A level. On the contrary, the LPS down-regulated the PIR-B expression and up-regulated the PIR-A expression. Realtime PCR examination demonstrated that PIR-A and co-stimulating molecules such as CD80, CD86 and MHC-Ⅱ were increased significantly after stimulation with LPS. Compared with the DC2.4 cells and the LPS-DC, the T-DCs inhibited alloactived T cell proliferation and down-regulated the IFN-γ secretion in MLR supernatant. Si-DC promoted the T cell proliferation (P< 0.01) and enhanced the IFN-y secretion (P< 0.01). It was concluded that up-regulating the PIR-B and down-regulating the PIR-A expression were the general feature of phenotype and constructed the new targets for dendritic cells to acquire immune tolerance in mice. Overexpression of PIR-B can inhibit the up-regulation of the PIR-A, CD80, CD86 and MHC-Ⅱ expression, which might be the molecular mechanism for the T-DC.
机译：研究了成对的免疫球蛋白样受体A（PIR-A）和B（PIR-B）的表达及其与耐受性树突状细胞（T-DC）的关系。小鼠DCs系DC2.4细胞分别用重组鼠白介素10（IL-10）和重组人转化生长因子β1（TGF-β1）培养以形成T-DC，并用脂多糖（LPS）刺激48小时诱导成熟的树突状细胞（LPS-DC）。化学合成用于PIR-B的特殊小干扰RNA（siRNA）分子，并通过lip2000将其转染到DC2.4细胞（Si-DC）中。通过半定量逆转录聚合酶链反应（RT-PCR），流式细胞术（FCM）和Western blot检测PIR在DC2.4细胞上的表达。应用实时逆转录-聚合酶链反应（Realtime-PCR）检测PIR-A和CD80，CD86，MHC-Ⅱ的mRNA表达。通过〜3H-胸苷掺入试验，通过混合淋巴细胞反应（MLR）测量DC的同种异体刺激能力。通过ELISA分析不同组的MLR上清液中IFN-γ的浓度。结果表明，FCM检测DC2.4细胞的PIR阳性率为（28.65±8.12）％。 IL-10，TGF-β1和LPS诱导的PIR阳性率分别显着增加至（54.21±6.34）％，（58.78±4.70）％，（48.24±6.75）％（P <0.01），但无显着性三组之间的差异（P> 0.01）。半定量RT-PCR和蛋白质印迹显示IL-10和TGF-β1诱导较高的PIR-B水平和较低的PIR-A水平。相反，LPS下调PIR-B表达而上调PIR-A表达。实时PCR检测显示，LPS刺激后，PIR-A和共刺激分子如CD80，CD86和MHC-Ⅱ显着增加。与DC2.4细胞和LPS-DC相比，T-DCs抑制了同种异体T细胞增殖，并下调了MLR上清液中IFN-γ的分泌。 Si-DC促进T细胞增殖（P <0.01）并增强IFN-y分泌（P <0.01）。结论是，上调PIR-B和下调PIR-A表达是表型的普遍特征，并为树突状细胞建立了新的靶标，以使其具有免疫耐受性。 PIR-B的过表达可抑制PIR-A，CD80，CD86和MHC-Ⅱ表达的上调，这可能是T-DC的分子机制。

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