中华口腔医学杂志
中華口腔醫學雜誌
중화구강의학잡지
Chinese Journal of Stomatology
2014年
1期
15-20
,共6页
王宏岩%谭丽思%刘俊超%李倩%潘亚萍%钟鸣
王宏巖%譚麗思%劉俊超%李倩%潘亞萍%鐘鳴
왕굉암%담려사%류준초%리천%반아평%종명
吸烟%紫单胞菌,龈%KB细胞%基质金属蛋白酶类
吸煙%紫單胞菌,齦%KB細胞%基質金屬蛋白酶類
흡연%자단포균,간%KB세포%기질금속단백매류
Smoking%Porphyromonas gingivalis%KB cells%Matrix metalloproteinases
目的 探讨慢性牙周炎患者中吸烟个体血清在牙龈卟啉单胞菌(Porphyromonas gingivalis,Pg)内化KB细胞及在内化过程中对KB细胞分泌基质金属蛋白酶(matrix metalloproteinase,MMP)1、MMP-9及金属蛋白酶组织抑制剂1(tissue inhibitor of metalloproteinase-1,TIMP-1)的作用.方法 抽取20例(吸烟个体10例,非吸烟个体10例)就诊于中国医科大学口腔医学院牙周科的慢性牙周炎患者前臂静脉血5 ml,离心提取血清.在试验组(吸烟组)和对照组(非吸烟组)中分别加入吸烟个体和非吸烟个体血清200、400及800μl至Pg感染KB细胞模型,作用12 h,脑心浸液琼脂培养基培养细菌5～7d,计数细菌菌落.采用人酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)试剂盒检测两组上清液中MMP-1、MMP-9及TIMP-1的浓度差异.采用单因素方差分析比较两组中分别加入不同体积(200、400、800μl)血清对Pg内化KB细胞的影响及KB细胞分泌MMP-1、MMP-9、TIMP-1的差异;采用t检验比较吸烟组与非吸烟组加入相同血清量Pg内化KB细胞及KB细胞分泌MMP-1、MMP-9、TIMP-1的浓度差异.结果 加入200、400、800μl血清时,吸烟组血清与非吸烟组血清引起Pg内化KB细胞的菌落数分别为(11.2±1.1) ×104、(12.6±1.2) ×104、(44.7±1.3)×104 CFU/ml和(33.6±1.4)×104、(38.9±1.1)×104、(11.2±1.2)×104 CFU/ml (P＜ 0.05).在加入200、400、800μl吸烟个体血清时,KB细胞分泌MMP-1的质量浓度分别为(107.2±21.5)、(165.9 ±20.2)及(434.4 ±48.0) μg/L,分泌MMP-9的质量浓度分别为(3.99±0.29)、(4.21±0.61)及(5.62±0.47)μg/L,分泌TIMP-1的质量浓度分别为(401.3 ±12.7)、(418.3 ±28.5)及(637.3 ±37.3) μg/L;加入200、400、800μl非吸烟个体血清时,KB细胞分泌MMP-1的质量浓度分别为(77.6±10.8)、(84.7±10.2)及(98.2±9.7)μg/L,分泌MMP-9的质量浓度分别为(3.84±0.52)、(4.02±0.68)及(4.25±0.37) μg/L,分泌TIMP-1的质量浓度分别为(67.3±26.9)、(89.4±22.7)及(78.2±16.5) μg/L.随着吸烟组血清剂量的增加,KB细胞表达MMP-1、MMP-9、TIMP-1的质量浓度增加,不同体积血清引起MMP-1、MMP-9及TIMP-1表达的差异具有统计学意义(P＜0.05);加入800μl吸烟组血清与加入800μl非吸烟组血清相比,KB细胞分泌MMP-1、MMP-9及TIMP-1的质量浓度均显著升高(P＜0.05).结论 吸烟个体的血清可能在Pg内化KB细胞的过程中发挥作用,并促进KB细胞产生MMP-1、MMP-9及TIMP-1;吸烟个体的口腔局部微环境可能有利于慢性牙周炎的进展及复发.
目的 探討慢性牙週炎患者中吸煙箇體血清在牙齦卟啉單胞菌(Porphyromonas gingivalis,Pg)內化KB細胞及在內化過程中對KB細胞分泌基質金屬蛋白酶(matrix metalloproteinase,MMP)1、MMP-9及金屬蛋白酶組織抑製劑1(tissue inhibitor of metalloproteinase-1,TIMP-1)的作用.方法 抽取20例(吸煙箇體10例,非吸煙箇體10例)就診于中國醫科大學口腔醫學院牙週科的慢性牙週炎患者前臂靜脈血5 ml,離心提取血清.在試驗組(吸煙組)和對照組(非吸煙組)中分彆加入吸煙箇體和非吸煙箇體血清200、400及800μl至Pg感染KB細胞模型,作用12 h,腦心浸液瓊脂培養基培養細菌5～7d,計數細菌菌落.採用人酶聯免疫吸附測定(enzyme-linked immunosorbent assay,ELISA)試劑盒檢測兩組上清液中MMP-1、MMP-9及TIMP-1的濃度差異.採用單因素方差分析比較兩組中分彆加入不同體積(200、400、800μl)血清對Pg內化KB細胞的影響及KB細胞分泌MMP-1、MMP-9、TIMP-1的差異;採用t檢驗比較吸煙組與非吸煙組加入相同血清量Pg內化KB細胞及KB細胞分泌MMP-1、MMP-9、TIMP-1的濃度差異.結果 加入200、400、800μl血清時,吸煙組血清與非吸煙組血清引起Pg內化KB細胞的菌落數分彆為(11.2±1.1) ×104、(12.6±1.2) ×104、(44.7±1.3)×104 CFU/ml和(33.6±1.4)×104、(38.9±1.1)×104、(11.2±1.2)×104 CFU/ml (P＜ 0.05).在加入200、400、800μl吸煙箇體血清時,KB細胞分泌MMP-1的質量濃度分彆為(107.2±21.5)、(165.9 ±20.2)及(434.4 ±48.0) μg/L,分泌MMP-9的質量濃度分彆為(3.99±0.29)、(4.21±0.61)及(5.62±0.47)μg/L,分泌TIMP-1的質量濃度分彆為(401.3 ±12.7)、(418.3 ±28.5)及(637.3 ±37.3) μg/L;加入200、400、800μl非吸煙箇體血清時,KB細胞分泌MMP-1的質量濃度分彆為(77.6±10.8)、(84.7±10.2)及(98.2±9.7)μg/L,分泌MMP-9的質量濃度分彆為(3.84±0.52)、(4.02±0.68)及(4.25±0.37) μg/L,分泌TIMP-1的質量濃度分彆為(67.3±26.9)、(89.4±22.7)及(78.2±16.5) μg/L.隨著吸煙組血清劑量的增加,KB細胞錶達MMP-1、MMP-9、TIMP-1的質量濃度增加,不同體積血清引起MMP-1、MMP-9及TIMP-1錶達的差異具有統計學意義(P＜0.05);加入800μl吸煙組血清與加入800μl非吸煙組血清相比,KB細胞分泌MMP-1、MMP-9及TIMP-1的質量濃度均顯著升高(P＜0.05).結論 吸煙箇體的血清可能在Pg內化KB細胞的過程中髮揮作用,併促進KB細胞產生MMP-1、MMP-9及TIMP-1;吸煙箇體的口腔跼部微環境可能有利于慢性牙週炎的進展及複髮.
목적 탐토만성아주염환자중흡연개체혈청재아간계람단포균(Porphyromonas gingivalis,Pg)내화KB세포급재내화과정중대KB세포분비기질금속단백매(matrix metalloproteinase,MMP)1、MMP-9급금속단백매조직억제제1(tissue inhibitor of metalloproteinase-1,TIMP-1)적작용.방법 추취20례(흡연개체10례,비흡연개체10례)취진우중국의과대학구강의학원아주과적만성아주염환자전비정맥혈5 ml,리심제취혈청.재시험조(흡연조)화대조조(비흡연조)중분별가입흡연개체화비흡연개체혈청200、400급800μl지Pg감염KB세포모형,작용12 h,뇌심침액경지배양기배양세균5～7d,계수세균균락.채용인매련면역흡부측정(enzyme-linked immunosorbent assay,ELISA)시제합검측량조상청액중MMP-1、MMP-9급TIMP-1적농도차이.채용단인소방차분석비교량조중분별가입불동체적(200、400、800μl)혈청대Pg내화KB세포적영향급KB세포분비MMP-1、MMP-9、TIMP-1적차이;채용t검험비교흡연조여비흡연조가입상동혈청량Pg내화KB세포급KB세포분비MMP-1、MMP-9、TIMP-1적농도차이.결과 가입200、400、800μl혈청시,흡연조혈청여비흡연조혈청인기Pg내화KB세포적균락수분별위(11.2±1.1) ×104、(12.6±1.2) ×104、(44.7±1.3)×104 CFU/ml화(33.6±1.4)×104、(38.9±1.1)×104、(11.2±1.2)×104 CFU/ml (P＜ 0.05).재가입200、400、800μl흡연개체혈청시,KB세포분비MMP-1적질량농도분별위(107.2±21.5)、(165.9 ±20.2)급(434.4 ±48.0) μg/L,분비MMP-9적질량농도분별위(3.99±0.29)、(4.21±0.61)급(5.62±0.47)μg/L,분비TIMP-1적질량농도분별위(401.3 ±12.7)、(418.3 ±28.5)급(637.3 ±37.3) μg/L;가입200、400、800μl비흡연개체혈청시,KB세포분비MMP-1적질량농도분별위(77.6±10.8)、(84.7±10.2)급(98.2±9.7)μg/L,분비MMP-9적질량농도분별위(3.84±0.52)、(4.02±0.68)급(4.25±0.37) μg/L,분비TIMP-1적질량농도분별위(67.3±26.9)、(89.4±22.7)급(78.2±16.5) μg/L.수착흡연조혈청제량적증가,KB세포표체MMP-1、MMP-9、TIMP-1적질량농도증가,불동체적혈청인기MMP-1、MMP-9급TIMP-1표체적차이구유통계학의의(P＜0.05);가입800μl흡연조혈청여가입800μl비흡연조혈청상비,KB세포분비MMP-1、MMP-9급TIMP-1적질량농도균현저승고(P＜0.05).결론 흡연개체적혈청가능재Pg내화KB세포적과정중발휘작용,병촉진KB세포산생MMP-1、MMP-9급TIMP-1;흡연개체적구강국부미배경가능유리우만성아주염적진전급복발.
Objective To investigate the effects of serum from smoking individuals or non-smoking individuals with periodontitis on Porphyromonas gingivalis(Pg) internalizing KB cells,and the expression of matrix metalloproteinase(MMP)-1,MMP-9,tissue inhibitor of metalloproteinase-1 (TIMP-1) in the culture supernatant of KB cells.Methods The venous blood of 20 periodontitis patients'(10 smoking and 10 non-smoking) was extracted under the informed consent and centrifuged for serum.The smoking-individual serum(Y group) and non-smoking-individual(N group)serum were added to the model of Pg internalizing KB cells for 12 hours,plated on brain-heart infusion(BHI) and incubated anaerobically at 37 ℃ for 5 days.The colony forming units(CFU) of cell-invasive bacteria were estimated by colony counting.M MP-1,MMP-9 and TIMP-1 protein levels in culture supernatant were determined by enzyme-linked immunosorbent assay (ELISA) in the two groups following co-culture of Pg with KB cells for 12 hours.Results The CFU were (11.2 ± 1.1) × 104,(12.6 ± 1.2) × 104,(44.7 ± 1.3) × 104 CFU/ml when adding 200,400,800 μl Y-group serum to the model of Pg co-culture with KB cells and when the serum was extracted from N group,the CFU were (33.6 ± 1.4) × 104,(38.9 ± 1.1) × 104,(11.2 ± 1.2) × 104 CFU/ml respectively.When 200,400,800 μl Y group-serum was added to co-culture fluid of Pg internalizing KB cells,the concentrations of MMP-1 secreted from KB cells were (107.2 ± 21.5),(165.9 ± 20.2),(434.4 ± 48.0) μg/L respectively,the concentrations of M MP-9 were (3.99 ±0.29),(4.21 ± 0.61),(5.62 ± 0.47) μg/L respectively,the concentrations of TIMP-1 were (401.3 ± 12.7),(418.3 ± 28.5),(637.3 ± 37.3) μg/L.When the serum (200,400,800 μl) extracted from N group,the concentration of MMP-1 and MMP-9 secreted by KB cell were (77.6±10.8),(84.7 ± 10.2) and (98.2 ±9.7) μg/L and (3.84 ±0.52),(4.02 ±0.68),(4.25 ±0.37) μg/L,respectively.The concentration of TIMP-1 were (67.3 ±26.9),(89.4 ± 22.7) and (78.2 ± 16.5) μg/L secreted by KB cells in the course of Pg internalized KB cell.With the increasing of Y group-serum,the more MMP-1,MMP-9 and TIMP-1 were secreted by KB cells (P ＜ 0.05).When 800 μl Y group-serum was added compared with N group-serum to the Pg co-culture with KB model,the more MMP-1,MMP-9 and TIMP-1 were secreted by KB cells (P＜0.05),when 400 μl Y group-serum was added compared with N group-serum to the Pg co-culture with KB model,the more MMP-1 and TIMP-1 were secreted by KB cells (P ＜ 0.05).Conclusions The smoking-serum might enhance Pg internalizing KB cells and enhance the expression of MMP-1,MMP-9 and TIMP-1 secreted from KB cells.The local microenvironment of smoking individual may contribute to the recurrence and progression of chronic periodontitis.
