首页|变异链球菌反义vicK RNA对口腔链球菌多菌种生物膜致龋性的抑制作用

变异链球菌反义vicK RNA对口腔链球菌多菌种生物膜致龋性的抑制作用

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目的 研究变异链球菌(Sm)反义vicKRNA(ASvicK)对3种常见口腔链球菌[Sm、血链球菌(Ss)和戈登链球菌(Sg)]构建的多菌种生物膜致龋性的调控作用.方法 通过重组质粒构建ASvicK过表达株,以Sm标准菌株UA159和ASvicK过表达株分别与Ss及Sg构建的多菌种生物膜(分别为UA159+Ss+Sg组、ASvicK+Ss+Sg组)为研究对象,扫描电镜观察生物膜结构;结晶紫染色法检测细菌生物膜总量的差异;通过乳酸试剂盒及蒽酮法评估生物膜产酸产糖能力;利用TaqMan荧光定量PCR及实时荧光定量PCR检测生物膜中3种菌的比例及Sm致龋相关基因的改变;进一步构建人牙釉质片生物膜脱矿模型,用显微硬度仪检测牙釉质表面硬度变化.结果ASvicK过表达后,Sm+Ss+Sg多菌种生物膜结构疏松.相较于UA159+Ss+Sg组,ASvicK+Ss+Sg组生物膜总量、乳酸产量分别显著降低78.93％及62.23％(均P＜0.001),而生物膜水不溶性和不溶性胞外多糖产量分别显著降低39.13％及68.00％(均P＜0.001).与UA159+Ss+Sg组相比,ASvicK+Ss+Sg多菌种生物膜中致龋菌Sm比例显著降低33.00％(P＜0.01),Sm致龋相关基因vicK/X、gtfB、gtfC、gtfD和ftf表达显著下调(P＜0.05),牙釉质片脱矿后显微硬度值显著上升至183.84％(P＜0.001).结论ASvicK过表达可降低口腔链球菌生物膜中致龋菌Sm的比例,并减弱口腔链球菌生物膜致龋毒力及致龋性,可能减缓龋病的进展.
Inhibitory effect of Streptococcus mutans antisense vicK RNA regulating the cariogenicity of oral streptococci multi-species biofilm
Objective To investigate the regulative effects of Streptococcus mutans(Sm)antisense vicK RNA(ASvicK)on the multi-species biofilm formed by three common oral streptococci(Sm,Streptococcus sanguinis,and Streptococcus gordonii)(Sm+Ss+Sg).Methods ASvicK over-expression strain was constructed by using a recombinant plasmid,and three-species biofilm UA159+Ss+Sg and ASvicK+Ss+Sg were cultured.The phenotypes of biofilms were detected by scanning electron microscopy(SEM).Crystal violet(CV)assay was used to detect biofilm biomass.Lactate kit and anthrone-sulfuric acid colorimetric assay were used to determine the abilities of lactic acid and exopolysaccharides production,respectively.The proportions of three-species and expression levels of the cariogenic-related genes in biofilms were detected by TaqMan fluorescence quantitative PCR and real-time fluorescence quantitative PCR.A biofilm demineralization model of human enamel slabs was further constructed,and the hardness of enamel surface was detected.Results Compared to UA159+Ss+Sg,over-expression of ASvicK could inhibit biofilm formation and lactic acid production in ASvicK+Ss+Sg biofilm significantly decreased by 78.93％(P＜0.001)and 62.23％(P＜0.001),respectively.With ASvicK over-expression,the amounts of water-insoluble and-soluble glucoses in ASvicK+Ss+Sg biofilm were reduced respectively by 39.13％(P＜0.001)and 68.00％(P＜0.001).Compared to the UA159+Ss+Sg Group,the proportion of Sm,the cariogenic bacteria,showed 33.00％reduction(P＜0.01)in Sm+Ss+Sg biofilm,and the gene expressions of cariogenic-relative genes vicK/X,gtfB/C/D,and ftf significantly decreased(P＜0.05).The micro-hardness value of enamel slabs after demineralization by ASvicK+Ss+Sg biofilm was significantly increased to 183.84％(P＜0.001).Conclusions ASvicK over-expression could reduce the Sm proportion and weaken the cariogenicity of oral Streptococcus biofilm,thereby possibly slowing down the progression of caries.

Dental cariesStreptococcus mutansAntisense vicK RNAStreptococcus sanguinisStreptococcus gordonii

陈虹、许萌萌、孙雨婷、余霜、杨德琴

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重庆医科大学附属口腔医院北部院区牙体牙髓科口腔疾病与生物医学重庆市重点实验室重庆市高校市级口腔生物医学工程重点实验室,重庆 401147

龋齿 链球菌,变异 反义vicK RNA 血链球菌 戈登链球菌

国家自然科学基金中国博士后科学基金重庆市自然科学基金博士后科学基金重庆市渝中区科技局基础研究与前沿探索项目重庆市博士后项目重庆市教委科学技术研究计划青年项目

821009912021M700627cstc2021jcyjbshX021120210124201001000599458382100991

2024

10.3760/cma.j.cn112144-20231031-00229

中华口腔医学杂志
中华医学会

中华口腔医学杂志

CSTPCD北大核心
影响因子：1.194
ISSN：1002-0098
年,卷(期)：2024.59(1)
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