State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China.
The First People's Hospital of Yunnan Province, Kunming, China.
Arch Oral Biol. 2017 Aug;80:41-50. doi: 10.1016/j.archoralbio.2017.03.017. Epub 2017 Mar 25.
Antimicrobial peptides (AMPs) have shown the ability to inhibit planktonic bacteria and biofilms. The objectives of this study were to de novo design and synthesize a series of cationic, amphipathic α-helical AMPs that would be shorter, less cytotoxic, and more potent than existing AMPs against cariogenic bacteria.
Three short AMPs (GH8, GLLWHLLH-NH; GH12, GLLWHLLHHLLH-NH; and GH16, GLLWHLLHHLLHLLHH-NH) were designed, synthesized and characterized structurally. Minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) against eight major cariogenic bacteria were tested to select the most promising peptide. Scanning electron microscopy (SEM) was used to observe the bacterial membrane after treatment with selected peptides. The bactericidal kinetics, effects on biofilm and cytotoxity were further investigated.
Of the three AMPs, GH12 had the most balanced structural parameters and a high content of α-helical structure. GH12 had a MIC of 4.0-8.0μg/mL and MBC of 8.0-32.0μg/mL. The corresponding values for the other two AMPs were 2- to 64- fold higher. In time-kill assays, GH12 killed all bacterial strains within 60min at 4- fold MBC. SEM observed lysis and pore formation of the cytomembrane after treatment with GH12. 8.0μg/mL GH12 inhibited Streptococcus mutans biofilm formation. Confocal laser scanning microscopy showed that GH12 effectively reduced the biomass of 1-day-old S. mutans biofilm. Cytotoxicity assays indicated that GH12 showed little toxic effect on the viability of human gingival fibroblasts.
These results indicate that GH12 shows antimicrobial activity against cariogenic bacteria and biofilms in vitro.
抗菌肽（AMPs）已显示出抑制浮游细菌和生物膜的能力。本研究的目的是从头设计和合成一系列阳离子、两亲性α-螺旋 AMPs，这些 AMPs 比现有的 AMPs 更短、细胞毒性更小、对致龋菌更有效。
设计、合成并结构表征了三种短 AMP（GH8，GLLWHLLH-NH；GH12，GLLWHLLHHLLH-NH；和 GH16，GLLWHLLHHLLHLLHH-NH）。测试了最小抑菌浓度（MIC）和最小杀菌浓度（MBC）对八种主要致龋菌，以选择最有前途的肽。扫描电子显微镜（SEM）用于观察经选定肽处理后的细菌膜。进一步研究了杀菌动力学、对生物膜的影响和细胞毒性。
在这三种 AMP 中，GH12 具有最平衡的结构参数和较高的α-螺旋结构含量。GH12 的 MIC 为 4.0-8.0μg/mL，MBC 为 8.0-32.0μg/mL。其他两种 AMP 的相应值高 2 至 64 倍。在时间杀伤试验中，GH12 在 4 倍 MBC 下 60min 内杀死所有细菌株。SEM 观察到 GH12 处理后细胞质膜的裂解和孔形成。8.0μg/mL 的 GH12 抑制变形链球菌生物膜的形成。共聚焦激光扫描显微镜显示 GH12 有效减少了 1 天龄变形链球菌生物膜的生物量。细胞毒性试验表明 GH12 对人牙龈成纤维细胞的活力几乎没有毒性作用。
这些结果表明 GH12 对致龋菌和生物膜具有体外抗菌活性。
