中华烧伤杂志
中華燒傷雜誌
중화소상잡지
16
2014年
2期
148-152
,共5页
侯玉森%柴家科%刘玲英%段红杰%郁永辉%胡泉%褚万立%王一贺%罗红敏
侯玉森%柴傢科%劉玲英%段紅傑%鬱永輝%鬍泉%褚萬立%王一賀%囉紅敏
후옥삼%시가과%류령영%단홍걸%욱영휘%호천%저만립%왕일하%라홍민
间质干细胞%脂多糖类%细胞增殖%细胞凋亡
間質榦細胞%脂多糖類%細胞增殖%細胞凋亡
간질간세포%지다당류%세포증식%세포조망
Mesenchymal stem cells%Lipopolysaccharides%Cell proliferation%Apoptosis
目的 体外观察不同浓度LPS对人脐带间充质干细胞(hUCMSC)增殖以及凋亡的影响,并探讨可能机制. 方法 取足月剖宫产健康胎儿的脐带组织,采用组织块法体外分离培养hUCMSC,实验选用第3代细胞.按随机数字表法将细胞分为对照组及0.1、1.0、10.0、100.0 μg/mLLPS干预组,各LPS干预组细胞培养液中添加对应浓度的LPS,对照组细胞培养液中不添加LPS.对LPS干预组细胞,刺激12、24、48 h采用噻唑蓝法检测细胞增殖活性(每组每时相点样本数为5),数据以吸光度值表示;刺激24 h采用吖啶橙-溴化乙啶(AO-EB)染色观察细胞凋亡情况(每组样本数为4),并采用流式细胞仪检测细胞凋亡率(每组样本数为5).对照组细胞于相同时相点行相应检测.对数据行单因素方差分析、LSD-t检验. 结果 (1)刺激12h,各组细胞增殖活性无明显差异(t值为-1.67 ～1.33,P值均大于0.05).与对照组比较,0.1、1.0、10.0μg/mL LPS干预组刺激24、48 h细胞增殖活性均明显增高(t值为-13.42 ～17.34,P＜0.05或P＜0.01),其中以1.0μg/mL LPS干预组最明显;100.0 μg/mL LPS干预组刺激24、48 h细胞增殖活性明显降低(t值分别为8.64、17.34,P值均小于0.01).(2)AO-EB染色显示,对照组和0.1、1.0、10.0μg/mL LPS干预组未见明显细胞凋亡,100.0 μg/mL LPS干预组可见明显细胞凋亡.(3)流式细胞仪显示,对照组及0.1、1.0、10.0、100.0 μg/mL LPS干预组细胞凋亡率分别为(3.1±0.6)％、(2.6±0.7)％、(2.9±0.8)％、(3.1±0.4)％、(25.1±2.7)％,组间比较差异有统计学意义(F=272.19,P＜0.01).0.1、1.0、10.0 μg/mL LPS干预组细胞凋亡率与对照组相近(t值分别为1.22、0.57、-0.14,P值均大于0.05),100.0 μg/mL LPS干预组细胞凋亡率明显高于对照组(t=-17.63,P＜0.01). 结论 低浓度水平LPS促进hUCMSC增殖,但随着LPS浓度逐渐增大,hUCMSC增殖活性下降甚至凋亡,这可能与不同浓度LPS激活的主要分子信号通路各异有关.
目的 體外觀察不同濃度LPS對人臍帶間充質榦細胞(hUCMSC)增殖以及凋亡的影響,併探討可能機製. 方法 取足月剖宮產健康胎兒的臍帶組織,採用組織塊法體外分離培養hUCMSC,實驗選用第3代細胞.按隨機數字錶法將細胞分為對照組及0.1、1.0、10.0、100.0 μg/mLLPS榦預組,各LPS榦預組細胞培養液中添加對應濃度的LPS,對照組細胞培養液中不添加LPS.對LPS榦預組細胞,刺激12、24、48 h採用噻唑藍法檢測細胞增殖活性(每組每時相點樣本數為5),數據以吸光度值錶示;刺激24 h採用吖啶橙-溴化乙啶(AO-EB)染色觀察細胞凋亡情況(每組樣本數為4),併採用流式細胞儀檢測細胞凋亡率(每組樣本數為5).對照組細胞于相同時相點行相應檢測.對數據行單因素方差分析、LSD-t檢驗. 結果 (1)刺激12h,各組細胞增殖活性無明顯差異(t值為-1.67 ～1.33,P值均大于0.05).與對照組比較,0.1、1.0、10.0μg/mL LPS榦預組刺激24、48 h細胞增殖活性均明顯增高(t值為-13.42 ～17.34,P＜0.05或P＜0.01),其中以1.0μg/mL LPS榦預組最明顯;100.0 μg/mL LPS榦預組刺激24、48 h細胞增殖活性明顯降低(t值分彆為8.64、17.34,P值均小于0.01).(2)AO-EB染色顯示,對照組和0.1、1.0、10.0μg/mL LPS榦預組未見明顯細胞凋亡,100.0 μg/mL LPS榦預組可見明顯細胞凋亡.(3)流式細胞儀顯示,對照組及0.1、1.0、10.0、100.0 μg/mL LPS榦預組細胞凋亡率分彆為(3.1±0.6)％、(2.6±0.7)％、(2.9±0.8)％、(3.1±0.4)％、(25.1±2.7)％,組間比較差異有統計學意義(F=272.19,P＜0.01).0.1、1.0、10.0 μg/mL LPS榦預組細胞凋亡率與對照組相近(t值分彆為1.22、0.57、-0.14,P值均大于0.05),100.0 μg/mL LPS榦預組細胞凋亡率明顯高于對照組(t=-17.63,P＜0.01). 結論 低濃度水平LPS促進hUCMSC增殖,但隨著LPS濃度逐漸增大,hUCMSC增殖活性下降甚至凋亡,這可能與不同濃度LPS激活的主要分子信號通路各異有關.
목적 체외관찰불동농도LPS대인제대간충질간세포(hUCMSC)증식이급조망적영향,병탐토가능궤제. 방법 취족월부궁산건강태인적제대조직,채용조직괴법체외분리배양hUCMSC,실험선용제3대세포.안수궤수자표법장세포분위대조조급0.1、1.0、10.0、100.0 μg/mLLPS간예조,각LPS간예조세포배양액중첨가대응농도적LPS,대조조세포배양액중불첨가LPS.대LPS간예조세포,자격12、24、48 h채용새서람법검측세포증식활성(매조매시상점양본수위5),수거이흡광도치표시;자격24 h채용아정등-추화을정(AO-EB)염색관찰세포조망정황(매조양본수위4),병채용류식세포의검측세포조망솔(매조양본수위5).대조조세포우상동시상점행상응검측.대수거행단인소방차분석、LSD-t검험. 결과 (1)자격12h,각조세포증식활성무명현차이(t치위-1.67 ～1.33,P치균대우0.05).여대조조비교,0.1、1.0、10.0μg/mL LPS간예조자격24、48 h세포증식활성균명현증고(t치위-13.42 ～17.34,P＜0.05혹P＜0.01),기중이1.0μg/mL LPS간예조최명현;100.0 μg/mL LPS간예조자격24、48 h세포증식활성명현강저(t치분별위8.64、17.34,P치균소우0.01).(2)AO-EB염색현시,대조조화0.1、1.0、10.0μg/mL LPS간예조미견명현세포조망,100.0 μg/mL LPS간예조가견명현세포조망.(3)류식세포의현시,대조조급0.1、1.0、10.0、100.0 μg/mL LPS간예조세포조망솔분별위(3.1±0.6)％、(2.6±0.7)％、(2.9±0.8)％、(3.1±0.4)％、(25.1±2.7)％,조간비교차이유통계학의의(F=272.19,P＜0.01).0.1、1.0、10.0 μg/mL LPS간예조세포조망솔여대조조상근(t치분별위1.22、0.57、-0.14,P치균대우0.05),100.0 μg/mL LPS간예조세포조망솔명현고우대조조(t=-17.63,P＜0.01). 결론 저농도수평LPS촉진hUCMSC증식,단수착LPS농도축점증대,hUCMSC증식활성하강심지조망,저가능여불동농도LPS격활적주요분자신호통로각이유관.
Objective To investigate the effects of different concentrations of lipopolysaccharide (LPS) on proliferation and apoptosis of human umbilical cord mesenchymal stem cells (hUCMSCs) in vitro,and to explore their possible mechanism.Methods hUCMSCs from umbilical cord tissue of full-term healthy fetus delivered by caesarean section were isolated and cultured in vitro using tissue attachment method.The 3rd passage hUCMSCs were used in the study.Cells were divided into groups A,B,C,D,and E,which were treated with DMEM/F12 medium containing 0,0.1,1.0,10.0,and 100.0 μg/mL of LPS respectively.In groups B,C,D,and E,methyl-thiazole-tetrazolium assay was used to detect proliferative activity of hUCMSCs at post treatment hour (PTH) 12,24,and 48 (denoted as absorption value),with 5 samples in each group at each time point; apoptosis of hUCMSCs at PBH 24 was identified with acridine orange-ethidium bromide (AO-EB) staining,with 4 samples in each group; apoptotic rate of hUCMSCs was determined by flow cytometer,with 5 samples in each group.Above-mentioned indexes were determined in group A at the same time points.Data were processed with analysis of variance and LSD-t test.Results (1) There was no statistically significant difference in proliferative activity of hUCMSCs at PTH 12 among groups A,B,C,D,and E (with t values from-1.67 to 1.33,P values above 0.05).Compared with that of group A,proliferative activity of hUCMSCs was increased in groups B,C,and D at PTH 24 and 48 (with t values from-13.42 to 17.34,P ＜0.05 or P ＜ 0.01),especially so in group C.Proliferative activity of hUCMSCs was lower in group E at PTH 24 and 48 than in group A (with t values respectively 8.64 and 17.34,P values below 0.01).(2) Obvious apoptosis of hUCMSCs was observed in group E but not in the other 4 groups with AO-EB staining.(3) Apoptosis rates of hUCMSCs in groups A,B,C,D,and E were respectively (3.1 ±0.6)％,(2.6±0.7)％,(2.9±0.8)％,(3.1 ±0.4)％,(25.1 ±2.7)％ (F =272.19,P ＜0.01).Apoptotic rate of hUCMSCs in group B,C,or D was respectively close to that in group A (with t values respectively 1.22,0.57,-0.14,P values above 0.05),but it was higher in group E than in group A (t =-17.63,P ＜ 0.01).Conclusions hUCMSCs proliferation may be promoted by low concentration of LPS.hUCMSCs proliferation is inhibited or induced to apoptosis along with the increase in concentration of LPS,and it may be related to activation of different major molecular signaling pathways by different concentrations of LPS.
