Hao Yu-qing, Zhou Xue-dong, Xiao Xiao-rong, Lu Jun-jun, Zhang Fu-chun, Hu Tao, Wu Hong-kun, Chen Xin-mei
Key Laboratory for Oral Biomedical Engineering, Ministry of Education, West China of Stomatology College, Sichuan University, Chengdu 610041, China.
Chin Med J (Engl). 2005 Jan 20;118(2):155-60.
Cecropin-XJ belongs to cecropin-B, which is the most potent antibacterial peptide found naturally. The aim of this study was to investigate the effects of cecropin-XJ on growth and adherence of oral cariogenic bacteria.
Four oral cariogenic bacteria (Streptococcus mutans, Lactobacillus acidophilus, Actinomyces viscosus and Actinomyces naeslundii) were chosen for this experiment. The minimal inhibitory concentrations (MICs) and reductive percent of bacterial growth were used to assay the antibacterial activity of cecropin-XJ. Mammalian cytotoxicity of cecropin-XJ was tested with human periodontal membrane fibroblasts by tetrazolium (MTT) colorimetric assay. The bacterial morphological changes induced by cecropin-XJ were examined on scanning electron microscope (SEM). The influence of cecropin-XJ on bacterial adhesion to saliva-coated hydroxyapatite (S-HA) was measured by scintillation counting.
The MICs of cecropin-XJ for inhibition of the growth of four bacteria ranged from 4.0 to 42.8 micromol/L with the highest susceptible to A. naeslundii and the lowest susceptible to L. acidophilus. At pH 6.8, 5.5 and 8.2, 1/2 MIC of cecropin-XJ reduced the number of viable bacteria by 40.9%, 67.8% and 32.8% for S. mutans and by 28.1%, 57.2% and 37.9% for L. acidophilus. The activities against S. mutans and L. acidophilus increased at pH 5.5 compared with pH 6.8 (P < 0.01, respectively). In present of 50% saliva, 1/2 MIC of the peptide decreased the direct count of viable cells by 29.2% and 14.4% for S. mutans and L. acidophilus, respectively (P < 0.01 and P > 0.05, respectively), whereas almost no reduction counts were detected in the presence of 20% serum for both bacteria (P > 0.05, respectively). Mammalian cytotoxicity of cecropin-XJ from 1.0 to 100 micromol/L exhibited no cytotoxicity against human periodontal membrane fibroblasts (P > 0.05). Bacterial morphological changes induced by MIC of cecropin-XJ examined on SEM showed cell surface disruption. Furthermore, the ability of A. naeslundii adhesion to S-HA decreased significantly with MIC of cecropin-XJ for 10 and 20 minutes (P = 0.001 and 0.000, respectively), and S. mutans, A. viscosus to S-HA decreased significantly with MIC of cecropin-XJ for 20 minutes (P = 0.000, respectively).
Cecropin-XJ exhibited bactericidal action against cariogenic pathogens, and the antibacterial activity enhanced in the acid environment. The results also demonstrate that cecropin-XJ prevents S. mutans and actinomyces adsorption to S-HA. These findings suggest that Cecropin-XJ may have potential to prevent caries.
天蚕素-XJ属于天蚕素-B，是天然发现的最有效的抗菌肽。本研究的目的是探讨天蚕素-XJ对口腔致龋菌生长和黏附的影响。
本实验选用4种口腔致龋菌（变形链球菌、嗜酸乳杆菌、黏性放线菌和内氏放线菌）。采用最低抑菌浓度（MIC）和细菌生长还原率测定天蚕素-XJ的抗菌活性。采用四氮唑盐（MTT）比色法检测天蚕素-XJ对人牙周膜成纤维细胞的细胞毒性。通过扫描电子显微镜（SEM）观察天蚕素-XJ诱导的细菌形态变化。采用闪烁计数法测定天蚕素-XJ对细菌黏附于唾液包被羟基磷灰石（S-HA）的影响。
天蚕素-XJ对4种细菌生长抑制的MIC范围为4.0至42.8微摩尔/升，对内氏放线菌最敏感，对嗜酸乳杆菌最不敏感。在pH 6.8、5.5和8.2时，1/2 MIC的天蚕素-XJ使变形链球菌的活菌数分别减少40.9%、67.8%和32.8%，使嗜酸乳杆菌的活菌数分别减少28.1%、57.2%和37.9%。与pH 6.8相比，在pH 5.5时，天蚕素-XJ对变形链球菌和嗜酸乳杆菌的活性增加（P分别<0.01）。在50%唾液存在下，1/2 MIC的该肽使变形链球菌和嗜酸乳杆菌的活菌直接计数分别减少29.2%和14.4%（P分别<0.01和P>0.05），而在20%血清存在下，两种细菌的活菌计数几乎没有减少（P均>0.05）。1.0至100微摩尔/升的天蚕素-XJ对人牙周膜成纤维细胞无细胞毒性（P>0.05）。在SEM下观察到，天蚕素-XJ的MIC诱导的细菌形态变化显示细胞表面破坏。此外，内氏放线菌对S-HA的黏附能力在天蚕素-XJ作用10分钟和20分钟时显著降低（P分别为0.001和0.000），变形链球菌、黏性放线菌对S-HA的黏附能力在天蚕素-XJ作用20分钟时显著降低（P均为0.000）。
天蚕素-XJ对致龋病原体具有杀菌作用，且在酸性环境中抗菌活性增强。结果还表明，天蚕素-XJ可阻止变形链球菌和放线菌吸附于S-HA。这些发现提示天蚕素-XJ可能具有预防龋齿的潜力。
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