目的 探討脂肪間充質(zhì)干細(xì)胞外泌體（ADSCs-Exos）對子宮內(nèi)膜異位癥細(xì)胞（12Z）增殖以及纖維化相關(guān)因子表達(dá)的影響。方法 采用超速離心法提取ADSCs-Exos，并通過透射電鏡（TEM）、納米顆粒跟蹤分析（NTA）、Western blotting鑒定；采用PKH26標(biāo)記外泌體法觀察12Z細(xì)胞對外泌體的吞噬內(nèi)化情況；采用CCK-8法檢測ADSCs-Exos （12.5、25.0、50.0 μg·mL^-1）對12Z細(xì)胞增殖的影響；將12Z細(xì)胞分為：對照組、模型組、ADSCs-Exos（12.5、25.0、50.0 μg·mL^-1）組，除對照組外，其余各組采用TGF-β1（5 ng·mL^-1）誘導(dǎo)細(xì)胞纖維化，給藥24 h后采用實(shí)時熒光定量PCR（qRT-PCR）檢測纖維化因子基因mRNA的表達(dá)。結(jié)果 提取的ADSCs-Exos呈現(xiàn)圓形顆粒，清晰可見；總萃取物的平均粒徑為122.6 nm，濃度為1.8×10⁶顆粒·mL^-1；CD9、CD63和TSG101在ADSCs-Exos均有表達(dá)，且Calnexin蛋白未表達(dá)，符合外泌體的特征。12Z細(xì)胞可以成功將ADSCs-Exos吞噬內(nèi)化；CCK-8檢測結(jié)果顯示，與對照組相比，采用25 μg·mL^-1 ADSCs-Exos干預(yù)24 h后12Z細(xì)胞的相對存活率顯著降低（P＜0.01）。qRT-PCR結(jié)果顯示，與對照組相比，模型組細(xì)胞的CTGF、COL1A1、α-SMAmRNA表達(dá)水平顯著升高（P＜0.001）；與模型組比較，12.5、25.0 μg·mL^-1 ADSCs-Exos組CTGF、COL1A1、α-SMA mRNA表達(dá)水平降低（P＜0.05、0.01、0.001）；50.0 μg·mL^-1 ADSCs-Exos組α-SMA mRNA表達(dá)水平明顯降低（P＜0.001）。結(jié)論 ADSCs-Exos可被12Z細(xì)胞所吞噬，抑制12Z細(xì)胞的增殖，并且降低細(xì)胞纖維化相關(guān)因子基因的表達(dá)水平。

Objective To investigate the effects of adipose-derived stem cell-derived exosomes (ADSCs-Exos) on the proliferation and fibrosis-related factor expression of endometriosis cells (12Z). Methods The ADSCs-Exos were extracted by ultracentrifugation, identified by transmission electron microscopy, nanoparticle tracking analysis (NTA), and Western blotting. The uptake of exosomes by 12Z cells was observed by PKH26 labeling exosome method. The effects of ADSCs-Exos (12.5, 25.0, 50.0 μg·mL^-1) on the proliferation of 12Z cells were detected by CCK-8 assay. The 12Z cells were divided into four groups: control group, model group, and ADSCs-Exos (12.5, 25.0, 50.0 μg·mL^-1) group. Except for the control group, the other groups were induced to fibrosis by TGF-β1 (5 ng·mL^-1) for 24 h, and the expression of fibrosis-related gene mRNA was detected by real-time quantitative PCR (qRT-PCR) after drug treatment. Results The ADSCs-Exos extracted showed round particles that were clearly visible. The average particle size of the total extract was 122.6 nm, and the concentration was 1.8×10⁶ particles·mL^-1. CD9, CD63, and TSG101 were expressed in ADSCs-Exos, and Calnexin protein was not expressed, consistent with the characteristics of exosomes. 12Z cells successfully internalized and phagocytosed ADSCs-Exos. The CCK-8 assay results showed that compared with the control group, the relative survival rate of 12Z cells was significantly lower after 24 h of treatment with 25.0 μg·mL^-1 ADSCs-Exos (P < 0.01). The qRT-qPCR results showed that compared with the control group, the expression levels of CTGF, COL1A1, and α -SMA mRNA in the model group were significantly incresed (P < 0.001). Compared with the model group, the expression levels of CTGF, COL1A1, and α -SMA mRNA in the 12.5, and 25.0 μg·mL^-1 ADSCs-Exos groups were lower (P < 0.05, 0.01, 0.001), and the expression level of α-SMA mRNA in the 50.0 μg·mL^-1 ADSCs-Exos group was significantly lower (P < 0.001). Conclusion Adipose mesenchymal stem cell exosomes can be swallowed by endometriosis cells, which can further inhibit the proliferation of endometriosis cells and reduce the expression level of cell fibrosis-related factor genes, thus playing a role in the treatment of endometriosis.

R965

國家自然科學(xué)基金委員會項(xiàng)目（82104920）;山東省自然科學(xué)基金委員會項(xiàng)目（ZR2021QH129）;山東省中醫(yī)藥重點(diǎn)學(xué)科方劑學(xué)（魯衛(wèi)中醫(yī)藥科教字〔2022〕4號）