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目的探讨c-Met受体阻断剂卡博替尼(Cabozantinib,XL-184)能否抑制产单核细胞李斯特菌(Listeria monocytogenes,LM)进入Caco-2细胞并降低对细胞的损伤。方法通过侵袭实验研究XL-184在抑制LM进入Caco-2细胞中的作用,并研究剂量及时间与侵袭率的关系;Caco-2细胞种植于Transwell上室,LM感染单层Caco-2细胞,然后在上室中加入辣根过氧化物酶(HRP),通过测定跨上皮细胞电阻(TEER)值及计数从上室渗透到下室的LM数量和HRP浓度,预测X-184对单层细胞通透性的保护作用;检测乳酸脱氢酶(LDH)释放水平推测细胞膜的渗透性;荧光染色鉴定细胞活性。结果相比对照组,侵袭实验显示,XL-184处理组的LM侵袭率明显降低(P=0.000),且侵袭率随XL-184浓度和作用时间的增加而显著降低,联合用药效果优于单独用药(P<0.05)。细胞通透性实验表明,XL-184可明显抑制LM诱导的跨上皮细胞电阻(TEER)的降低(P<0.05),并可降低辣根过氧化物酶和细菌介导的细胞通透性(P=0.000);细胞活性实验表明,XL-184可提高细胞存活率(P<0.01);同时,可显著降低LM诱导的LDH释放(P<0.05)。结论 XL-184可能在抑制LM侵袭Caco-2及降低LM诱导的Caco-2损伤中起重要作用。因此,XL-184有望成为潜在的治疗和预防李斯特菌感染的有效药物。
Objective To investigate whether Cabozantinib (XL-184), a c-Met receptor antagonist, can inhibit the entry of Listeria monocytogenes (LM) into Caco-2 cells and reduce the damage to cells. Methods The effect of XL-184 on the inhibition of LM entry into Caco-2 cells was studied by invasion assay and the relationship between dose and time and invasion rate was studied. Caco-2 cells were seeded on the upper chamber of Transwell and monolayers of Caco-2 cells were infected by LM. Horseradish peroxidase (HRP) was then added to the upper chamber and X-184 was evaluated for monolayer cells by measuring the trans-epithelial cell resistance (TEER) value and counting the number of LM permeating into the lower chamber from the upper chamber and the HRP concentration Permeability of the protective effect; detection of lactate dehydrogenase (LDH) release level speculate membrane permeability; fluorescent staining to identify cell activity. Results Compared with the control group, the invasion assay showed that the LM invasion rate of XL-184 treatment group was significantly lower (P = 0.000), and the invasion rate was significantly decreased with the increase of XL-184 concentration and action time, Medication (P <0.05). Cell permeability experiments showed that XL-184 significantly inhibited the LM-induced decrease of TEER (P <0.05) and decreased the activity of horseradish peroxidase and bacterial-mediated cell permeability P = 0.000). The cell viability assay showed that XL-184 increased the cell viability (P <0.01) and at the same time decreased the LM-induced LDH release (P <0.05). Conclusion XL-184 may play an important role in inhibiting the invasion of Caco-2 by LM and reducing the damage of LM-induced Caco-2. Therefore, XL-184 is expected to become a potential drug for the treatment and prevention of listeriosis.