Esposito Giuseppe, Nobile Nicola, Gigli Stefano, Seguella Luisa, Pesce Marcella, d'Alessandro Alessandra, Bruzzese Eugenia, Capoccia Elena, Steardo Luca, Cuomo Rosario, Sarnelli Giovanni
Department of Physiology and Pharmacology "Vittorio Erspamer", Sapienza University of Rome Rome, Italy.
Department of Clinical Medicine and Surgery, University of Naples Federico II Naples, Italy.
Front Pharmacol. 2016 May 9;7:120. doi: 10.3389/fphar.2016.00120. eCollection 2016.
Clostridium difficile infections (CDIs) caused by Clostridium difficile toxin A (TcdA) lead to severe ulceration, inflammation and bleeding of the colon, and are difficult to treat.
The study aimed to evaluate the effect of rifaximin on TcdA-induced apoptosis in intestinal epithelial cells and investigate the role of PXR in its mechanism of action.
Caco-2 cells were incubated with TcdA and treated with rifaximin (0.1-10 μM) with or without ketoconazole (10 μM). The transepithelial electrical resistance (TEER) and viability of the treated cells was determined. Also, the expression of zona occludens-1 (ZO-1), toll-like receptor 4 (TLR4), Bcl-2-associated X protein (Bax), transforming growth factor-β-activated kinase-1 (TAK1), myeloid differentiation factor 88 (MyD88), and nuclear factor-kappaB (NF-κB) was determined.
Rifaximin treatment (0.1, 1.0, and 10 μM) caused a significant and concentration-dependent increase in the TEER of Caco-2 cells (360, 480, and 680% vs. TcdA treatment) 24 h after the treatment and improved their viability (61, 79, and 105%). Treatment also concentration-dependently decreased the expression of Bax protein (-29, -65, and -77%) and increased the expression of ZO-1 (25, 54, and 87%) and occludin (71, 114, and 262%) versus TcdA treatment. The expression of TLR4 (-33, -50, and -75%), MyD88 (-29, -60, and -81%) and TAK1 (-37, -63, and -79%) were also reduced with rifaximin versus TcdA treatment. Ketoconazole treatment inhibited these effects.
Rifaximin improved TcdA-induced toxicity in Caco-2 cells by the PXR-dependent TLR4/MyD88/NF-κB pathway mechanism, and may be useful in the treatment of CDIs.
艰难梭菌毒素A（TcdA）引起的艰难梭菌感染（CDIs）会导致结肠严重溃疡、炎症和出血，且难以治疗。
本研究旨在评估利福昔明对TcdA诱导的肠上皮细胞凋亡的影响，并探讨孕烷X受体（PXR）在其作用机制中的作用。
将Caco-2细胞与TcdA孵育，并在有或无酮康唑（10 μM）的情况下用利福昔明（0.1 - 10 μM）处理。测定处理后细胞的跨上皮电阻（TEER）和活力。此外，还测定了紧密连接蛋白1（ZO-1）、Toll样受体4（TLR4）、Bcl-2相关X蛋白（Bax）、转化生长因子-β激活激酶1（TAK1）、髓样分化因子88（MyD88）和核因子-κB（NF-κB）的表达。
利福昔明处理（0.1、1.0和10 μM）在处理后24小时使Caco-2细胞的TEER显著且呈浓度依赖性增加（分别比TcdA处理增加360%、480%和680%），并改善了细胞活力（分别为61%、79%和105%）。与TcdA处理相比，处理还呈浓度依赖性降低了Bax蛋白的表达（分别降低29%、65%和77%），增加了ZO-1（分别增加25%、54%和87%）和闭合蛋白（分别增加71%、114%和262%）的表达。与TcdA处理相比，利福昔明处理还降低了TLR4（分别降低33%、50%和75%）、MyD88（分别降低29%、60%和81%）和TAK1（分别降低37%、63%和79%）的表达。酮康唑处理抑制了这些作用。
利福昔明通过PXR依赖性TLR4/MyD88/NF-κB途径机制改善了TcdA诱导的Caco-2细胞毒性，可能对CDIs的治疗有用。
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