南方医科大学学报
南方醫科大學學報
남방의과대학학보
JOURNAL OF SOUTHERN MEDICAL UNIVERSITY
2014年
2期
174-179
,共6页
屈娅荣%何肖龙%王勤%张立科%龙敏%罗军%张文炳%曹虹
屈婭榮%何肖龍%王勤%張立科%龍敏%囉軍%張文炳%曹虹
굴아영%하초룡%왕근%장립과%룡민%라군%장문병%조홍
尿路致病性大肠杆菌%外膜蛋白T%基因敲除%黏附
尿路緻病性大腸桿菌%外膜蛋白T%基因敲除%黏附
뇨로치병성대장간균%외막단백T%기인고제%점부
uropathogenic Escherichia Coli%outer membrane protein T%gene knockout%adhesion
目的：探讨大肠杆菌外膜蛋白T（OmpT）参与尿路致病性大肠杆菌致病的机制。方法通过建立人膀胱癌移行上皮细胞模型，检测野生株CFT073、ompT基因敲除株COTD、回补株COTD（pST）的体外黏附情况；比较野生株、敲除株和回补株对细胞外基质的体外黏附能力；通过荧光定量PCR方法，比较ompT基因敲除以后，iha和iroN基因的mRNA表达水平；建立小鼠动物模型，比较有无OmpT菌株所致小鼠膀胱和肾脏的细菌载量、血液菌浓度及炎症因子IL-6和IL-8的蛋白表达水平，验证OmpT的致炎作用。结果野生株细胞黏附率为（8.81±1.13）%，敲除株为（4.62±0.39）%，敲除株的黏附率明显低于野生株（P<0.05）；野生株的细胞外基质黏附率为（8.85±0.79）%，敲除株为（4.95±0.59）%，敲除株的黏附率明显低于野生株（P<0.05）；与敲除株比较，iha和iroN基因在野生株中的表达水平分别是敲除株的2.1和3.8倍；野生株感染的小鼠膀胱组织中细菌载量均数为6.36±0.06，敲除株为6.01±0.07，回补株为6.29±0.06，野生株感染的小鼠肾脏组织中细菌载量为6.25±0.05，敲除株为5.87±0.06，差异均有统计学意义（P<0.05）；野生株和回补株所致小鼠膀胱和肾脏组织炎症中IL-6检出的阳性率分别为60%和50%，而ompT基因敲除株则为12.5%。IL-8的表达情况同IL-6。结论初步证实OmpT通过发挥毒力因子的作用，从而影响尿路致病性大肠杆菌对宿主细胞的黏附，其可能机制是影响细菌对细胞外基质的黏附和参与iha和iroN基因的表达并在致炎过程中发挥重要作用。
目的：探討大腸桿菌外膜蛋白T（OmpT）參與尿路緻病性大腸桿菌緻病的機製。方法通過建立人膀胱癌移行上皮細胞模型，檢測野生株CFT073、ompT基因敲除株COTD、迴補株COTD（pST）的體外黏附情況；比較野生株、敲除株和迴補株對細胞外基質的體外黏附能力；通過熒光定量PCR方法，比較ompT基因敲除以後，iha和iroN基因的mRNA錶達水平；建立小鼠動物模型，比較有無OmpT菌株所緻小鼠膀胱和腎髒的細菌載量、血液菌濃度及炎癥因子IL-6和IL-8的蛋白錶達水平，驗證OmpT的緻炎作用。結果野生株細胞黏附率為（8.81±1.13）%，敲除株為（4.62±0.39）%，敲除株的黏附率明顯低于野生株（P<0.05）；野生株的細胞外基質黏附率為（8.85±0.79）%，敲除株為（4.95±0.59）%，敲除株的黏附率明顯低于野生株（P<0.05）；與敲除株比較，iha和iroN基因在野生株中的錶達水平分彆是敲除株的2.1和3.8倍；野生株感染的小鼠膀胱組織中細菌載量均數為6.36±0.06，敲除株為6.01±0.07，迴補株為6.29±0.06，野生株感染的小鼠腎髒組織中細菌載量為6.25±0.05，敲除株為5.87±0.06，差異均有統計學意義（P<0.05）；野生株和迴補株所緻小鼠膀胱和腎髒組織炎癥中IL-6檢齣的暘性率分彆為60%和50%，而ompT基因敲除株則為12.5%。IL-8的錶達情況同IL-6。結論初步證實OmpT通過髮揮毒力因子的作用，從而影響尿路緻病性大腸桿菌對宿主細胞的黏附，其可能機製是影響細菌對細胞外基質的黏附和參與iha和iroN基因的錶達併在緻炎過程中髮揮重要作用。
목적：탐토대장간균외막단백T（OmpT）삼여뇨로치병성대장간균치병적궤제。방법통과건립인방광암이행상피세포모형，검측야생주CFT073、ompT기인고제주COTD、회보주COTD（pST）적체외점부정황；비교야생주、고제주화회보주대세포외기질적체외점부능력；통과형광정량PCR방법，비교ompT기인고제이후，iha화iroN기인적mRNA표체수평；건립소서동물모형，비교유무OmpT균주소치소서방광화신장적세균재량、혈액균농도급염증인자IL-6화IL-8적단백표체수평，험증OmpT적치염작용。결과야생주세포점부솔위（8.81±1.13）%，고제주위（4.62±0.39）%，고제주적점부솔명현저우야생주（P<0.05）；야생주적세포외기질점부솔위（8.85±0.79）%，고제주위（4.95±0.59）%，고제주적점부솔명현저우야생주（P<0.05）；여고제주비교，iha화iroN기인재야생주중적표체수평분별시고제주적2.1화3.8배；야생주감염적소서방광조직중세균재량균수위6.36±0.06，고제주위6.01±0.07，회보주위6.29±0.06，야생주감염적소서신장조직중세균재량위6.25±0.05，고제주위5.87±0.06，차이균유통계학의의（P<0.05）；야생주화회보주소치소서방광화신장조직염증중IL-6검출적양성솔분별위60%화50%，이ompT기인고제주칙위12.5%。IL-8적표체정황동IL-6。결론초보증실OmpT통과발휘독력인자적작용，종이영향뇨로치병성대장간균대숙주세포적점부，기가능궤제시영향세균대세포외기질적점부화삼여iha화iroN기인적표체병재치염과정중발휘중요작용。
Objective To study the role of outer membrane protein T (OmpT) in the pathogenesis of uropathogenic Escherichia. coli. Methods In cultured human bladder epithelial cell line 5637, we examined the adhesion ability of wild-type (CFT073), ompT gene knockout (COTD), and revertant (pST) strains of E.coli to the cells and the extracellular matrix (ECM). The expressions of the adhesion gene iha and virulence gene iroN were detected by real-time PCR. Murine models of urinary tract infection with the 3 strains were established to evaluate the bacterial burden of the bladder and kidney tissue and bacterial counts in blood. We also detected the expressions of interleukin-6 (IL-6) and IL-8 in the bladder and kidney tissues of the mice. Results The COTD strain showed a significantly lower cell adhesion rate than CFT073 strain [(4.62±0.39)%vs (8.81±1.13)%, P<0.05] with also a lower ECM-adhesion rate [(4.95 ± 0.59)%vs (8.85 ± 0.79)%, P<0.05]. The mRNA expressions of iha and iroN in CFT073 strain were 2.1 and 3.8 times that of COTD strain. In the mouse model, the mean bacterial load of CFT073 strain in the bladder tissue was 6.36±0.06, significantly greater than that of COTD (6.01±0.07) and revertant (6.29±0.06) strains (P<0.05);the bacterial load of CFT073 strain in the kidney tissue was also significantly higher than that of COTD strain (6.25 ± 0.05 vs 5.87 ± 0.06, P<0.05). In mice infected with the wild-type, knockout, and revertant strains, the detection rates of IL-6, which were identical to those of IL-8, in the inflammatory bladder and kidney tissues were 60%, 12.5%, and 50%, respectively. Conclusions OmpT may regulate the expression of the adhesion gene iha and the transferrin gene iroN to affect the adhesion of uropathogenic E.coli to host cells.
