中华临床医师杂志（电子版）
中華臨床醫師雜誌（電子版）
중화림상의사잡지（전자판）
CHINESE JOURNAL OF CLINICIANS(ELECTRONIC VERSION)
2013年
24期
11559-11562
,共4页
合湫%苏恒%李超%沈涛%薛元明%严新民
閤湫%囌恆%李超%瀋濤%薛元明%嚴新民
합추%소항%리초%침도%설원명%엄신민
胰岛%小鼠%糖尿病%葡萄糖刺激下的胰岛素分泌
胰島%小鼠%糖尿病%葡萄糖刺激下的胰島素分泌
이도%소서%당뇨병%포도당자격하적이도소분비
Islets of langerhans%Mice%Diabetes mellitus%Glucose stimulation of the insulin secretion
目的：探讨炎性细胞因子IL-1β、IFN-γ对小鼠原代胰岛葡萄糖刺激下胰岛素分泌功能的影响。方法（1）经胰腺表面多点注射灌注胶原酶消化，不连续密度梯度离心法分离、纯化小鼠胰岛。分离后胰岛培养48 h后换用无糖KRB缓冲液培养30 min，分别于含0、2.8、22.2 mmol/L葡萄糖的KRB缓冲液中孵育60 min，放射免疫法检测上清液中胰岛素浓度（GSIS）；（2）小鼠胰岛分别加入不同浓度的IL-1β（0.25 ng/ml、2.5 ng/ml）和（或）IFN-γ（100 U/ml、1000 U/ml），24 h换无糖KRB缓冲液培养30 min，于含22.2 mmol/L葡萄糖的KRB缓冲液中孵育1 h，放射免疫法检测细胞上清液中胰岛素浓度；（3）采用SPSS 11.5对数据进行分析，多组间比较采用方差分析，两组间比较采用t检验，以P＜0.05为有统计学差异。结果（1）与无葡萄糖刺激组相比，小鼠胰岛在22.2 mmol/L葡萄糖刺激时胰岛素分泌达高峰[（142.48±1.22）μIU/ml vs.（40.61±0.74）μIU/ml，t＝-123.669，P＜0.001]；2.8 mmol/L葡萄糖刺激时胰岛素分泌较22.2 mmol/L葡萄糖刺激时下降；（2）与单纯22.2 mmol/L葡萄糖刺激组相比，IL-1β（0.25 ng/ml）组[（113.83±5.65）μIU/ml vs.（142.48±1.22）μIU/ml，t＝8.759，P＜0.05]、IL-1β（2.5 ng/ml）组[（91.01±2.62）μIU/ml vs.（142.48±1.22）μIU/ml，t＝30.874，P＜0.05]葡萄糖刺激下胰岛素分泌显著下降，下降程度随IL-1β的剂量增加而下降更明显；与单纯葡萄糖刺激组相比，IFN-γ（100 U/ml）组[（116.09±6.11）μIU/ml vs.（142.48±1.22）μIU/ml，t＝7.336，P＜0.05]、IFN-γ（1000 U/ml）组[（71.78±3.51）μIU/ml vs.（142.48±1.22）μIU/ml，t＝13.687，P＜0.001]；（3）与IL-1β（2.5 ng/ml）组及IFN-γ（1000 U/ml）组相比，IL-1β联合IFN-γ组葡萄糖刺激下胰岛素分泌显著下降（F＝329.098，P＜0.001）。结论 IL-1β和IFN-γ对原代小鼠原代胰岛的胰岛素分泌功能有抑制作用，且两者间有协同效应。
目的：探討炎性細胞因子IL-1β、IFN-γ對小鼠原代胰島葡萄糖刺激下胰島素分泌功能的影響。方法（1）經胰腺錶麵多點註射灌註膠原酶消化，不連續密度梯度離心法分離、純化小鼠胰島。分離後胰島培養48 h後換用無糖KRB緩遲液培養30 min，分彆于含0、2.8、22.2 mmol/L葡萄糖的KRB緩遲液中孵育60 min，放射免疫法檢測上清液中胰島素濃度（GSIS）；（2）小鼠胰島分彆加入不同濃度的IL-1β（0.25 ng/ml、2.5 ng/ml）和（或）IFN-γ（100 U/ml、1000 U/ml），24 h換無糖KRB緩遲液培養30 min，于含22.2 mmol/L葡萄糖的KRB緩遲液中孵育1 h，放射免疫法檢測細胞上清液中胰島素濃度；（3）採用SPSS 11.5對數據進行分析，多組間比較採用方差分析，兩組間比較採用t檢驗，以P＜0.05為有統計學差異。結果（1）與無葡萄糖刺激組相比，小鼠胰島在22.2 mmol/L葡萄糖刺激時胰島素分泌達高峰[（142.48±1.22）μIU/ml vs.（40.61±0.74）μIU/ml，t＝-123.669，P＜0.001]；2.8 mmol/L葡萄糖刺激時胰島素分泌較22.2 mmol/L葡萄糖刺激時下降；（2）與單純22.2 mmol/L葡萄糖刺激組相比，IL-1β（0.25 ng/ml）組[（113.83±5.65）μIU/ml vs.（142.48±1.22）μIU/ml，t＝8.759，P＜0.05]、IL-1β（2.5 ng/ml）組[（91.01±2.62）μIU/ml vs.（142.48±1.22）μIU/ml，t＝30.874，P＜0.05]葡萄糖刺激下胰島素分泌顯著下降，下降程度隨IL-1β的劑量增加而下降更明顯；與單純葡萄糖刺激組相比，IFN-γ（100 U/ml）組[（116.09±6.11）μIU/ml vs.（142.48±1.22）μIU/ml，t＝7.336，P＜0.05]、IFN-γ（1000 U/ml）組[（71.78±3.51）μIU/ml vs.（142.48±1.22）μIU/ml，t＝13.687，P＜0.001]；（3）與IL-1β（2.5 ng/ml）組及IFN-γ（1000 U/ml）組相比，IL-1β聯閤IFN-γ組葡萄糖刺激下胰島素分泌顯著下降（F＝329.098，P＜0.001）。結論 IL-1β和IFN-γ對原代小鼠原代胰島的胰島素分泌功能有抑製作用，且兩者間有協同效應。
목적：탐토염성세포인자IL-1β、IFN-γ대소서원대이도포도당자격하이도소분비공능적영향。방법（1）경이선표면다점주사관주효원매소화，불련속밀도제도리심법분리、순화소서이도。분리후이도배양48 h후환용무당KRB완충액배양30 min，분별우함0、2.8、22.2 mmol/L포도당적KRB완충액중부육60 min，방사면역법검측상청액중이도소농도（GSIS）；（2）소서이도분별가입불동농도적IL-1β（0.25 ng/ml、2.5 ng/ml）화（혹）IFN-γ（100 U/ml、1000 U/ml），24 h환무당KRB완충액배양30 min，우함22.2 mmol/L포도당적KRB완충액중부육1 h，방사면역법검측세포상청액중이도소농도；（3）채용SPSS 11.5대수거진행분석，다조간비교채용방차분석，량조간비교채용t검험，이P＜0.05위유통계학차이。결과（1）여무포도당자격조상비，소서이도재22.2 mmol/L포도당자격시이도소분비체고봉[（142.48±1.22）μIU/ml vs.（40.61±0.74）μIU/ml，t＝-123.669，P＜0.001]；2.8 mmol/L포도당자격시이도소분비교22.2 mmol/L포도당자격시하강；（2）여단순22.2 mmol/L포도당자격조상비，IL-1β（0.25 ng/ml）조[（113.83±5.65）μIU/ml vs.（142.48±1.22）μIU/ml，t＝8.759，P＜0.05]、IL-1β（2.5 ng/ml）조[（91.01±2.62）μIU/ml vs.（142.48±1.22）μIU/ml，t＝30.874，P＜0.05]포도당자격하이도소분비현저하강，하강정도수IL-1β적제량증가이하강경명현；여단순포도당자격조상비，IFN-γ（100 U/ml）조[（116.09±6.11）μIU/ml vs.（142.48±1.22）μIU/ml，t＝7.336，P＜0.05]、IFN-γ（1000 U/ml）조[（71.78±3.51）μIU/ml vs.（142.48±1.22）μIU/ml，t＝13.687，P＜0.001]；（3）여IL-1β（2.5 ng/ml）조급IFN-γ（1000 U/ml）조상비，IL-1β연합IFN-γ조포도당자격하이도소분비현저하강（F＝329.098，P＜0.001）。결론 IL-1β화IFN-γ대원대소서원대이도적이도소분비공능유억제작용，차량자간유협동효응。
Objective To investigate the effect of inflammatory cytokines IL-1βand IFN-γon the insulin secretion in mouse islets. Methods (1)The isolation of mouse pancreatic islets was carried out by stationary digestion after multi-point injection of collagenase solution. A discontinuous histopaque solution (1119, 1110, 1080 and 1060) was applied to the purification of the islets. Isolated mouse islets were cultured in DMEM for 48 hours then incubated for 30 minutes in Krebs-Ringer bicarbonate (KRB) buffer without glucose. KRB was then discarded and replaced with fresh buffer containing 0 mmol/L, 2.8 mmol/L or 22.2 mmol/L glucose for 1 hour. Supernatants were collected for insulin assays by an insulin RIA kit (GSIS). (2)IL-1β(0.25 ng/ml, 2.5 ng/ml) and/or IFN-γ(100 U/ml, 1000 U/ml) were added in islets for 24 hours then GSIS was measured as indicated with 22.2 mmol/L glucose stimulation.(3) Data were analyzed by ANOVA with correction for multiple comparisons or by LSD's t-test with SPSS 11.5. Significance was assumed at P<0.05, Extremely significance at P<0.001. Results (1) The insulin level reaches a peak in response to 22.2 mmol/L compared with 0 mmol/L glucose stimulation [(142.48±1.22) μIU/ml vs. (40.61±0.74)μIU/ml, t=-123.669, P<0.001]. The insulin level in response to 2.8 mmol/L glucose stimulation was lower than 22.2 mmol/L glucose stimulation. (2) Compared with 22.2 mmol/L glucose stimulation alone group, the insulin level was significantly lower in IL-1β(0.25 ng/ml) group [(113.83±5.65)μIU/ml vs. (142.48±1.22)μIU/ml, t = 8.759, P < 0.05] and IL-1β(2.5 ng/ml) group [(91.01±2.62)μIU/ml vs. (142.48±1.22)μIU/ml, t=30.874, P<0.05]. Compared with 22.2 mmol/L glucose stimulation alone group, the insulin level was significantly lower in IFN-γ(100 U/ml) group [(116.09±6.11)μIU/ml vs. (142.48±1.22)μIU/ml, t=7.336, P< 0.05] and IFN-γ(1000 U/ml) group [(71.78±3.51)μIU/ml vs. (142.48±1.22)μIU/ml, t=13.687, P<0.001]. (3) Compared with IL-1β(2.5 ng/ml) and IFN-γ(1000 U/ml) groups, the insulin level in response to 22.2 mmol/L glucose stimulation was significantly lower in IL-1βplus IFN-γgroup (F=329.098, P<0.001). Conclusion Glucose-induced insulin secretion was inhibited by inflammatory cytokines IL-1β, IFN-γor in cooperation in mouse islets.
