Myd88 knockdown with RNA interference induces in vitro immune hyporesponsiveness in dendritic cells from rhesus monkeys
Myd88 knockdown with RNA interference induces in vitro immune hyporesponsiveness in dendritic cells from rhesus monkeys
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RNA 干扰敲低 Myd88 会导致恒河猴树突状细胞体外免疫反应低下
DOI：
10.1007/s00251-022-01260-x
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发表时间：
2022-03-18
期刊：
影响因子：
3.2
通讯作者：
Hu, Mingdao
中科院分区：
文献类型：
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作者：
Wang, Qiuhong;Cun, Dongyun;Hu, Mingdao
未成熟树突状细胞（imDC）被激活并成熟以启动适应性免疫应答，导致同种异体移植排斥和移植失败。髓样分化因子88（Myd 88）是Toll样受体（TLR）信号通路中的关键因子。在这里，我们研究了Myd 88沉默对DC功能和免疫应答的影响。通过免疫磁珠法从恒河猴骨髓中分离CD 34+细胞，然后用表达Myd 88特异性短发夹RNA（sh-Myd 88）的腺病毒感染。将sh-NC（非靶向阴性对照）或sh-Myd 88感染的DC用脂多糖（LPS）再处理48小时以诱导DCS成熟。通过MHCII、CD 80和CD 86的免疫荧光染色鉴定DC的成熟。Annexin V/PI染色检测DC凋亡。通过ELISA评估DC相关细胞因子水平（IFN-γ和IL-12）。采用混合淋巴细胞反应（MLR）检测Myd 88沉默的DC对体外T淋巴细胞的影响。结果显示，与对照组和sh-NC感染组相比，Myd 88沉默组的DC具有较低的MHCII、CD 80、CD 86和DC相关细胞因子（IFN-γ和IL-12）水平。Myd 88对DCs的凋亡无影响。MLR表明Myd 88沉默可以有效地阻断体外LPS激活的T细胞增殖。这些数据与致耐受性DC的特征一致。综上所述，Myd 88沉默可抑制imDCs的成熟，减轻免疫排斥反应，为临床肝移植免疫耐受提供参考。
Immature dendritic cells (imDCs) are activated and mature to initiate an adaptive immune response, resulting in allograft rejection and transplantation failure. Myeloid differentiation factor 88 (Myd88) is a key factor in the Toll-like receptor (TLR) signaling pathway. Here, we investigated the effect of Myd88 silencing on DC function and immune response. CD34 + cells were isolated from the bone marrow of rhesus monkeys by the immunomagnetic bead method and then infected with an adenovirus expressing Myd88-specific short hairpin RNA (sh-Myd88). sh-NC (nontargeting negative control)- or sh-Myd88-infected DCs were treated with lipopolysaccharide (LPS) for another 48 h to induce DCS maturation. The maturation of DCs was identified by immunofluorescence staining for MHCII, CD80, and CD86. DC apoptosis was examined using Annexin V/PI staining. DC-related cytokine levels (IFN-γ and IL-12) were assessed by ELISA. A mixed lymphocyte reaction (MLR) was performed to test the effect of Myd88-silenced DCs on T lymphocytes in vitro. The results showed that compared with control or sh-NC-infected DCs, Myd88-silenced DCs had lower MHCII, CD80, CD86, and DC-related cytokine (IFN-γ and IL-12) levels. Myd88 did not affect the apoptosis of DCs. MLR demonstrated that Myd88 silencing could effectively block LPS-activated T cell proliferation in vitro. These data were consistent with the characteristics of tolerogenic DCs. In conclusion, our data indicated that Myd88 silencing could inhibit the maturation of imDCs and alleviate immune rejection, which provides a reference for immune tolerance in clinical liver transplantation.