Federal University of Pernambuco (UFPE), Keizo Asami Institute (iLIKA), Recife, Pernambuco, Brazil.
Federal University of Pernambuco (UFPE), Biochemistry Department, Recife, Pernambuco, Brazil.
Microb Pathog. 2024 Nov;196:107007. doi: 10.1016/j.micpath.2024.107007. Epub 2024 Oct 10.
The present study aimed to evaluate the anti-staphylococcal, antibiofilm, cytotoxicity and trypanocidal activity, mechanisms of parasite death and immunomodulatory effect of CrataBL encapsulated into liposomes (CrataBL-Lipo). CrataBL-Lipo were prepared by the freeze-thaw technique and characterized. Anti-staphylococcal and antibiofilm activities of CrataBL and CrataBL-Lipo were evaluated against standard and clinical strains of Staphylococcus aureus susceptible and resistant. Thus, broth microdilution method was performed to determine the Minimum Inhibitory Concentration (MIC). Antibiofilm activity at subinhibitory concentrations was evaluated using the crystal violet staining method. Cytotoxicity of CrataBL-Lipo was verified in L929 fibroblasts and J774A.1 macrophages by determining the inhibitory concentration necessary to kill 50 % of cells (IC). Trypanocidal activities of CrataBL-Lipo was evaluated in Trypanosoma cruzi and the efficacy was expressed as the concentration necessary to kill 50 % of parasites (EC). The mechanisms of parasite death and immunomodulatory effect of CrataBL-Lipo were evaluated using flow cytometry analysis. CrataBL-Lipo presented Ø of 101.9 ± 1.3 nm (PDI = 0.245), ζ of +33.8 ± 1.3 mV and %EE = 80 ± 0.84 %. CrataBL-Lipo presented anti-staphylococcal activity (MIC = 0.56 mg/mL to 0.72 mg/mL). CrataBL-Lipo inhibited 45.4 %-75.6 % of biofilm formation. No cytotoxicity of CrataBL-Lipo was found (IC > 100 mg/L). CrataBL-Lipo presented EC of 1.1 mg/L, presenting autophagy, apoptosis and necrosis as death profile. In addition, CrataBL-Lipo reduced the production of IL-10 and TNF-α levels, causing an immunomodulatory effect. CrataBL-Lipo has a therapeutic potential for the treatment of staphylococcal infections and Chagas disease exhibiting a high degree of selectivity for the microorganism, and immunomodulatory properties.
本研究旨在评估 CrataBL 包封于脂质体(CrataBL-Lipo)后的抗葡萄球菌、抗生物膜、细胞毒性和杀锥虫活性、寄生虫死亡机制和免疫调节作用。采用冻融法制备 CrataBL-Lipo 并进行表征。采用肉汤微量稀释法测定最小抑菌浓度(MIC),评估 CrataBL 和 CrataBL-Lipo 对标准和临床敏感和耐药金黄色葡萄球菌株的抗葡萄球菌和抗生物膜活性。采用结晶紫染色法评估亚抑菌浓度下的抗生物膜活性。通过测定半数细胞杀伤所需的抑制浓度(IC),在 L929 成纤维细胞和 J774A.1 巨噬细胞中验证 CrataBL-Lipo 的细胞毒性。采用测定半数寄生虫杀伤所需的浓度(EC),评估 CrataBL-Lipo 在 Trypanosoma cruzi 中的杀锥虫活性。用流式细胞术分析评估 CrataBL-Lipo 的寄生虫死亡机制和免疫调节作用。CrataBL-Lipo 的平均粒径为 101.9 ± 1.3nm(PDI = 0.245),ζ电位为+33.8 ± 1.3mV,%EE = 80 ± 0.84%。CrataBL-Lipo 具有抗葡萄球菌活性(MIC 为 0.56 至 0.72mg/mL)。CrataBL-Lipo 抑制 45.4%-75.6%的生物膜形成。未发现 CrataBL-Lipo 的细胞毒性(IC>100mg/L)。CrataBL-Lipo 的 EC 为 1.1mg/L,表现出自噬、凋亡和坏死作为死亡模式。此外,CrataBL-Lipo 降低了 IL-10 和 TNF-α 水平的产生,具有免疫调节作用。CrataBL-Lipo 具有治疗金黄色葡萄球菌感染和恰加斯病的潜力,对微生物具有高度选择性,并具有免疫调节特性。