CAJ | 학술논문

目的探讨肺炎衣原体(C.pn)感染对胰岛β细胞增殖和功能的影响.方法将C.pn在人喉癌细胞系(HEp-2细胞)中增殖培养后,以感染复数为0.08、0.20,干预时间为6,24 h的C.pn感染体外培养的胰岛β细胞系(INS-1细胞),通过光镜下观察细胞形态变化及免疫荧光单克隆抗体染色方法确认感染模型的建立.在不同感染复数(0,0.04,0.08,0.10,0.12,0.14,0.16,0.20)、不同干预时间(2,4,6,8,16,24,36 h)的C.pn作用下,利用四甲基偶氮唑蓝法、台盼蓝活细胞计数及流式细胞仪观察C.pn感染对INS-1细胞增殖和凋亡的影响；并通过放射免疫法测定C.pn感染后经高糖(25 mmol/L)和低糖(5.6 mmol/L)刺激的INS-1细胞胰岛素分泌量.结果与正常对照组相比,小剂量(0.08)加短时间(6 h)C.pn感染组INS-1细胞密度增加,细胞间隙变窄,大剂量(0.20)加长时间(24 h)C.pn感染组INS-1细胞骤缩、变圆、仍聚团生长.免疫荧光单克隆抗体染色显示特异性亮绿色荧光,证实C.pn感染INS-1细胞的体外模型成功建立.与正常对照组相比,小剂量(＜0.20)加短时间(＜24 h)的C.pn感染促进INS-1细胞增殖(t=-8.907,P ＜0.05)及INS-1细胞分泌胰岛素(t=-9.186,P＜0.05),而大剂量(＞0.20)加长时间(＞24 h)的C.pn感染则促进INS-1细胞凋亡(t=-37.306,P＜0.05)且抑制INS-1细胞分泌胰岛素(t=9.592,P＜0.05).结论 C.pn感染对胰岛β细胞增殖和功能具有双向作用,可为2型糖尿病早期防治提供新思路.
목적 탐토폐염의원체(C.pn)감염대이도β세포증식화공능적영향.방법 장C.pn재인후암세포계(HEp-2세포)중증식배양후,이감염복수위0.08、0.20,간예시간위6,24 h적C.pn감염체외배양적이도β세포계(INS-1세포),통과광경하관찰세포형태변화급면역형광단극륭항체염색방법학인감염모형적건립.재불동감염복수(0,0.04,0.08,0.10,0.12,0.14,0.16,0.20)、불동간예시간(2,4,6,8,16,24,36 h)적C.pn작용하,이용사갑기우담서람법、태반람활세포계수급류식세포의관찰C.pn감염대INS-1세포증식화조망적영향；병통과방사면역법측정C.pn감염후경고당(25 mmol/L)화저당(5.6 mmol/L)자격적INS-1세포이도소분비량.결과 여정상대조조상비,소제량(0.08)가단시간(6 h)C.pn감염조INS-1세포밀도증가,세포간극변착,대제량(0.20)가장시간(24 h)C.pn감염조INS-1세포취축、변원、잉취단생장.면역형광단극륭항체염색현시특이성량록색형광,증실C.pn감염INS-1세포적체외모형성공건립.여정상대조조상비,소제량(＜0.20)가단시간(＜24 h)적C.pn감염촉진INS-1세포증식(t=-8.907,P ＜0.05)급INS-1세포분비이도소(t=-9.186,P＜0.05),이대제량(＞0.20)가장시간(＞24 h)적C.pn감염칙촉진INS-1세포조망(t=-37.306,P＜0.05)차억제INS-1세포분비이도소(t=9.592,P＜0.05).결론 C.pn감염대이도β세포증식화공능구유쌍향작용,가위2형당뇨병조기방치제공신사로.
Objective To explore the effects of infection of Chlamydia pneumonia (C.pn) on the proliferation and function of pancreatic β cells.Methods C.pn had been cultured in the human laryngeal epidermoid carcinoma cell line(HEp-2 cells).Then the pancreatic β cells (INS-1 cells) were infected by C.pn,the multiplicity of infection were 0.08 and 0.20,the time for intervention were 6 h and 24 h.The changes of cell morphology under the light microscopy and the immunofluorescence monoclonal antibody staining method were used to confirm the establishment of infection model.Under the infection of C.pn with different multiplicity of infection (0,0.04,0.08,0.10,0.12,0.14,0.16,0.20) and different time (2,4,6,8,16,24,36 h),the methyl thiazolyl tetrazolium assay,the trypan blue and the flow cytometry were used to evaluate the effects of C.pn on the proliferation and function of INS-1 cells.Moreover,the low glucose (5.6 mmol/L) and high glucose (25 mmol/L)-stimulated insulin secretion of INS-1 cells,infected with C.pn,were measured through radioimmunoassay.Results Compared with normal control group,the cell density increased and the cell gap became more narrow in C.pn interventional group with a small dose (0.08) plus a short time (6 h).However,the cells became shrink and round,still agglomerated growing,in C.pn intervent-ional group with a large dose (0.20) plus a long time (24 h).The immunofluorescent monoclonal antibody staining showed a specificity bright green fluorescent,confirming the C.pn inflectional cell model in vitro had been successfully established.Compared with normal control group,the infection of C.pn with a small dose (＜ 0.20) plus a short time (＜ 24 h) could induce the proliferation of INS-1 cells (t =-8.907,P ＜ 0.05)and increase the glucose-stimulated insulin secretion of INS-1 cells (t =-9.186,P ＜ 0.05),but a large dose (＞ 0.20) plus a long time (＞ 24 h) could induce the apoptosis of INS-1 cells (t =-37.306,P ＜0.05) and decrease the glucose-stimulated insulin secretion of INS-1 cells (t =9.592,P ＜ 0.05).Conclusion The infection of C.pn has a two-way effect on the proliferation and function of pancreatic β cells,providing new ideas for early prevention of type 2 diabetes mellitus.