Department of Endodontics, School and Hospital of Stomatology, Wenzhou Medical University, Wenzhou 325027, China.
School and Hospital of Stomatology, Wenzhou Medical University, Wenzhou 325027, China.
Biomed Res Int. 2021 Mar 23;2021:6692328. doi: 10.1155/2021/6692328. eCollection 2021.
To investigate the effects of ficin on biofilm formation of conditionally cariogenic (). Biomass and metabolic activity of biofilm were assessed using crystal violet assay, colony-forming unit (CFU) counting, and MTT assay. Extracellular polysaccharide (EPS) synthesis was displayed by SEM imaging, bacteria/EPS staining, and anthrone method while acid production was revealed by lactic acid assay. Growth curve and live/dead bacterial staining were conducted to monitor bacterial growth state in both planktonic and biofilm form. Total protein and extracellular proteins of biofilm were analyzed by protein/bacterial staining and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), severally. qRT-PCR was conducted to detect acid production, acid tolerance, and biofilm formation associated genes. Crystal violet assay, CFU counting, and MTT assay showed that the suppression effect of ficin on biofilm formation was concentration dependent. 4 mg/mL ficin had significant inhibitory effect on biofilm formation including biomass, metabolic activity, EPS synthesis, and lactic acid production ( < 0.05). The growth curves from 0 mg/mL to 4 mg/mL ficin were aligned with each other. There was no significant difference among different ficin groups in terms of live/dead bacterial staining result ( > 0.05). Protein/bacterial staining outcome indicated that ficin inhibit both total protein and biofilm formation during the biofilm development. There were more relatively small molecular weight protein bands in extracellular proteins of 4 mg/mL ficin group when compared with the control. Generally, ficin could inhibit biofilm formation and reduce cariogenic virulence of effectively in vitro; thus, it could be a potential anticaries agent.
为了研究ficin 对条件致龋菌 () 生物膜形成的影响。采用结晶紫法、菌落形成单位 (CFU) 计数和 MTT 法评估生物膜的生物量和代谢活性。通过扫描电镜成像、细菌/ EPS 染色和蒽酮法显示胞外多糖 (EPS) 的合成，通过乳酸测定法显示产酸情况。通过生长曲线和活菌/死菌染色来监测浮游和生物膜形式的细菌生长状态。通过蛋白/细菌染色和十二烷基硫酸钠聚丙烯酰胺凝胶电泳 (SDS-PAGE) 分别分析生物膜的总蛋白和胞外蛋白。通过 qRT-PCR 检测产酸、耐酸和生物膜形成相关基因。结晶紫法、CFU 计数和 MTT 法显示 ficin 对 生物膜形成的抑制作用呈浓度依赖性。4mg/mL ficin 对生物膜形成具有显著的抑制作用，包括生物量、代谢活性、EPS 合成和乳酸产生（<0.05）。从 0mg/mL 到 4mg/mL ficin 的生长曲线彼此一致。不同 ficin 组之间活菌/死菌染色结果无显著差异（>0.05）。蛋白/细菌染色结果表明 ficin 在生物膜发育过程中抑制总蛋白和生物膜形成。与对照组相比，4mg/mL ficin 组的胞外蛋白中相对分子量较小的蛋白带更多。总的来说，ficin 可有效抑制体外生物膜形成并降低 的致龋毒力；因此，它可能是一种潜在的抗龋剂。
