滴虫病是最常见的非病毒性性传播感染，增加了宫颈癌的风险。阴道滴虫（T. vaginalis）可以调节宿主细胞中的促炎细胞因子产生。 Toll样受体（TLRs）是哺乳动物细胞中的一类模式识别受体（PRRs），在各种宿主细胞中表达，并在识别病原体和促炎反应中起重要作用。本研究的目的是研究TLR5在暴露于T. vaginalis的宫颈癌细胞（HeLa）和人阴道上皮细胞（HVECs）中的作用。首先，细胞和寄生虫分别在RPMI和酪蛋白酵母浸出物麦芽糖（TYM）中培养。细胞和寄生虫在RPMI-TYM培养基中培养（9：1体积比共培养）后，HVECs和HeLa细胞被T. vaginalis滴虫刺激（37℃，5% CO2孵育24小时）。在RNA提取和cDNA合成后，使用实时聚合酶链反应评估TLR5，IRAK1和NF-κB的基因表达水平。此外，使用蛋白印迹法测量蛋白水平。所有的测试和对照都使用β-actin作为内部控制进行归一化。实时聚合酶链反应结果显示与对照组相比，T. vaginalis暴露的HVECs和HeLa细胞中TLR5，IRAK1和NF-κB的基因表达显著增加（p < 0.05）。此外，蛋白印迹分析显示在寄生虫暴露后，两组中TLR5和NF-κB蛋白水平显著增加（p < 0.05）。这些发现揭示了宿主-寄生虫相互作用，并表明T. vaginalis能够刺激TLR5并激活相关途径。版权所有（c）2023 Soraya Mohammadi Esfanjani，Leili Aghebati Maleki，Sanam Nami，Mina Ebrahimi，Amir Baghbanzadeh，Gregorio Perez-Cordon，Sonia M Rodrigues Oliveira，Maria de Lourdes Pereira，Aleksandra Barać，Ehsan Ahmadpour，Jelena Micić。

Trichomoniasis is the most common non-viral sexually transmitted infection that increases the risk of cervical cancer. Trichomonas vaginalis (T. vaginalis) can regulate the pro-inflammatory cytokine production in the host cells. Toll-like receptors (TLRs) are a family of the pattern recognition receptors (PRRs) of mammalian cells, expressed in various host cells and have an important role in recognizing pathogens, and pro-inflammatory responses. The aim of the present study is to investigate the role of TLR5 in cervical cancer cells (HeLa) and human vaginal epithelial cells (HVECs) exposed to T. vaginalis.First, the cells and parasites were cultured in RPMI and trypticase yeast extract maltose (TYM), respectively. After adaption of parasite and epithelial cells by RPMI-TYM medium co-culture (9:1 vol/vol), HVECs and HeLa cells were stimulated with T. vaginalis trophozoites (24-hour incubation at 37 °C, 5% CO2). Following RNA extraction and cDNA synthesis, the gene expression levels of TLR5, IRAK1, and NF-κB were assessed using real-time PCR. Besides, the protein levels were measured using western blotting. All tests and controls were normalized using β-actin as a housekeeping control.Real-time PCR results showed an increased gene expression of TLR5, IRAK1, and NF-κB in T. vaginalis exposed HVECs and HeLa cells compared to the control group (p < 0.05). Additionally, western blot analysis showed a statistically significant increase in TLR5, and NF-κB proteins in both groups after exposure to the parasite (p < 0.05).These findings provide insight into the host-parasite interaction, and the results indicated that T. vaginalis could stimulate TLR5 and activate related pathways.Copyright (c) 2023 Soraya Mohammadi Esfanjani, Leili Aghebati Maleki, Sanam Nami, Mina Ebrahimi, Amir Baghbanzadeh, Gregorio Perez-Cordon, Sonia M Rodrigues Oliveira, Maria de Lourdes Pereira, Aleksandra Barać, Ehsan Ahmadpour, Jelena Micić.