摘要目的:探讨外周血单个核细胞（peripheral blood mononuclear cells，PBMCs）源性外泌体经miR-1306改善子宫内膜容受性的机制。方法:运用皮下注射米非司酮制备着床障碍小鼠模型并分为着床障碍组、PBMCs干预组，同时设置正常组，每组12只。PBMCs干预组给予宫腔注射PBMCs源性外泌体干预，正常组和着床障碍组宫腔注射等体积缓冲液，比较各组小鼠的妊娠率和着床点数，RT-PCR法检测子宫内膜组织miR-1306表达，酶联免疫吸附试验检测子宫内膜组织活性氧（reactive oxygen species，ROS）、白细胞介素-6（interleukin-6，IL-6）、单核细胞趋化因子-1（monocyte chemcattractant protein-1，MCP-1）、超氧化物歧化酶（superoxide dismutase，SOD）水平，Western blotting法检测小鼠子宫内膜组织中妊娠相关蛋白表达。将子宫内膜上皮细胞分为对照组、实验组、阴性对照组、miR-1306 inhibitor组，除对照组外余下各组细胞均采取Transwell共培养PBMCs源性外泌体，使用MTT法、Edu法、Annexin-V/PI流式法分别检测细胞活性、增殖和凋亡情况，试剂盒检测细胞ROS水平，Western blotting检测相关蛋白表达。结果:着床障碍组小鼠妊娠率［8.33%（1/12）］、着床点数［0（0，0）个］和子宫内膜组织miR-1306表达（0.24±0.05）、SOD水平［（5.66±0.72）U/mL］均显著低于正常组［100%（12/12）、16.50（14.00，19.00）个、1.03±0.05、（8.69±1.21）U/mL，均 P<0.05］；子宫内膜组织ROS水平［（4.87±0.39）U/mL］、IL-6水平［（116.51±5.78）ng/L］、MCP-1水平［（36.84±3.56）μg/L］和KIR2DL4（0.87±0.06）、核因子红细胞2相关因子2（nuclear factor erythroid-2-related factor 2，Nrf2，0.76±0.06）、Kelch样环氧氯丙烷相关蛋白1（Kelch-like ECH-associated protein 1，Keap1，0.79±0.05）、受体相互作用蛋白酶1（receptor interacting protein 1，RIP1，0.94±0.04）和RIP3蛋白（0.86±0.05）表达水平均显著高于正常组［（2.41±0.19）U/mL、（83.79±6.68）ng/L、（12.32±2.09）μg/L、0.27±0.03、0.31±0.05、0.23±0.04、0.34±0.03、0.31±0.05，均 P<0.05］。PBMCs干预组小鼠妊娠率［75.00%（9/12）］、着床点数［13.00（13.00，14.75）个］和子宫内膜组织miR-1306表达（0.82±0.05）、SOD水平［（7.24±0.84）U/mL］均显著高于着床障碍组（均 P<0.05）；子宫内膜组织ROS［（3.43±0.30）U/mL］、IL-6［（94.69±3.99）ng/L］、MCP-1水平［（27.03±3.48）μg/L］和KIR2DL4（0.54±0.08）、Nrf2（0.48±0.05）、Keap1（0.43±0.05）、RIP1（0.56±0.05）、RIP3蛋白表达（0.49±0.03）均显著低于着床障碍组（均 P<0.05）。实验组细胞活性［（126.63±1.25）%］、增殖率［（53.54±2.82）%］和ROS水平［（3.12±0.31）U/mL］均显著高于对照组［100%、（23.18±3.07）%、（2.51±0.28）U/mL，均 P<0.05］，凋亡率［（5.69±0.47）%］、KIR2DL4（0.36±0.06）、Nrf2（0.30±0.06）、Keap1（0.26±0.04）、RIP1（0.27±0.05）和RIP3蛋白表达（0.26±0.06）均低于对照组［（27.13±2.97）%、0.84±0.06、0.75±0.05、0.68±0.05、0.80±0.06、0.80±0.07，均 P<0.05］。miR-1306 inhibitor组细胞活性［（83.48±5.34）%］、增殖率［（38.42±4.28）%］均显著低于阴性对照组［（127.12±4.08）%、（53.57±2.09）%，均 P<0.05］，细胞凋亡率［（13.63±1.77）%］、ROS水平［（6.49±0.62）U/mL］、KIR2DL4（0.67±0.07）、Nrf2（0.57±0.05）、Keap1（0.50±0.05）、RIP1（0.64±0.06）和RIP3蛋白表达（0.61±0.08）均显著高于阴性对照组［（5.71±0.78）%、（3.23±0.31）U/mL、0.36±0.07、0.30±0.07、0.27±0.06、0.28±0.07、0.28±0.06，均 P<0.05］。 结论:PBMCs源性外泌体可以提高着床障碍小鼠妊娠率，可能是通过促进miR-1306表达、抑制KIR2DL4表达以改善炎症反应，还与缓解子宫内膜过度氧化应激和细胞凋亡有关。

abstractsObjective:To explore the mechanism of peripheral blood mononuclear cells (PBMCs)-derived exosomes on improving endometrial receptivity by miR-1306.Methods:The models of mice with implantation disorders were prepared by subcutaneous injection of mifepristone. And they were divided into implantation disorder group and PBMCs intervention group, 12 mice in each group. The normal group was set up. PBMCs intervention group was given intrauterine injection of PBMCs-derived exosomes, while normal group and implantation disorder group were given the same volume of buffer solution. The pregnancy rates and number of embryos implantation in different groups were compared. The expression of miR-1306 in endometrial tissues was detected by RT-PCR. The levels of reactive oxygen species (ROS), interleukin-6 (IL-6), monocyte chemcattractant protein-1 (MCP-1) and superoxide dismutase (SOD) in endometrial tissues were detected by enzyme-linked immunosorbent assay. The expressions of pregnancy-related proteins in endometrial tissues were detected by Western blotting. The endometrial epithelial cells were divided into control group, experimental group, negative control group and miR-1306 inhibitor group. Except control group, cells were co-cultured with PBMCs-derived exosomes by Transwell in the other groups. The activity, proliferation and apoptosis of cells were detected by MTT, Edu and Annexin-V/PI flow, respectively. The level of ROS in cells was detected by kits. And expressions of the related proteins were detected by Western blotting.Results:The pregnancy rate [8.33% (1/12), number of embryos implantation [0(0, 0)], expression of miR-1306 in endometrial tissue (0.24±0.05) and SOD level [(5.66±0.72) U/mL] in implantation disorder group were significantly lower than those in normal group [100% (12/12), 16.50 (14.00, 19.00), 1.03±0.05, (8.69±1.21) U/mL, P<0.05], while levels of ROS [(4.87±0.39) U/mL], IL-6 [(116.51±5.78) ng/L] and MCP-1 in endometrial tissues [(36.84±3.56) μg/L], and expressions of KIR2DL4 (0.87±0.06), nuclear factor erythroid-2-related factor 2 (Nrf2, 0.76±0.06), Kelch-like ECH-associated protein1 (Keap1, 0.79±0.05), receptor interacting protein 1 (RIP1, 0.94±0.04) and RIP3 (0.86±0.05) were significantly higher than those in normal group [(2.41±0.19) U/mL, (83.79±6.68) ng/L, (12.32±2.09) ng/mL, 0.27±0.03, 0.31±0.05, 0.23±0.04, 0.34±0.03, 0.31±0.05, all P<0.05]. The pregnancy rate [75.00% (9/12)], number of embryos implantation [13.00 (13.00, 14.75)], expression of miR-1306 in endometrial tissues (0.82±0.05) and SOD level [(7.24±0.84) U/mL] in PBMCs intervention group were significantly higher than those in implantation disorder group (all P<0.05), while levels of ROS [(3.43±0.30) U/mL], IL-6 [(94.69±3.99) ng/L] and MCP-1 in endometrial tissues [(27.03±3.48) μg/L], and expressions of KIR2DL4 (0.54±0.08), Nrf2 (0.48±0.05), Keap1 (0.43±0.05), RIP1 (0.56±0.05) and RIP3 (0.49±0.03) were significantly lower than those in implantation disorder group (all P<0.05). The cells activity [(126.63±1.25)%], proliferation rate [(53.54±2.82)%] and ROS level [(3.12±0.31) U/mL] in experimental group were significantly higher than those in control group [100%, (23.18±3.07)%, (2.51±0.28) U/mL, all P<0.05], while apoptosis rate [(5.69±0.47)%], expressions of KIR2DL4 (0.36±0.06), Nrf2 (0.30±0.06), Keap1 (0.26±0.04), RIP1 (0.27±0.05) and RIP3 (0.26±0.06) were lower than those in control group [(27.13±2.97)%, 0.84±0.06, 0.75±0.05, 0.68±0.05, 0.80±0.06, 0.80±0.07, all P<0.05]. The cells activity [(83.48±5.34)%] and proliferation rate [(38.42±4.28)%] in miR-1306 inhibitor group were significantly lower than those in negative control group [(127.12±4.08)%, (53.57±2.09)%, all P<0.05], while apoptosis rate [(13.63±1.77)%], ROS level [(6.49±0.62) U/mL], expressions of KIR2DL4 (0.67±0.07), Nrf2 (0.57±0.05), Keap1 (0.50±0.05), RIP1 (0.64±0.06) and RIP3 (0.61±0.08) were significantly higher than those in negative control group [(5.71±0.78)%, (3.23±0.31) U/mL, 0.36±0.07, 0.30±0.07, 0.27±0.06, 0.28±0.07, 0.28±0.06, all P<0.05]. Conclusion:PBMCs-derived exosomes can improve pregnancy rate in mice with implantation disorders, which may improve inflammatory response by promoting miR-1306 expression and inhibiting KIR2DL4 expression, and is also related to relieving excessive oxidative stress and apoptosis of endometrium.