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
4期
1615-1619
,共5页
程百祥%张旻%杜静%陈慧%李轶杰%陈永进
程百祥%張旻%杜靜%陳慧%李軼傑%陳永進
정백상%장민%두정%진혜%리질걸%진영진
间质干细胞%P38MAPK%细胞膜片
間質榦細胞%P38MAPK%細胞膜片
간질간세포%P38MAPK%세포막편
Mesenchymal stem cells%P38MAPK%Cell sheets
目的研究P38丝裂原激活的蛋白激酶（P38MAPK）信号通路在压力调控骨髓间充质干细胞（BMSCs）膜片成软骨响应中作用。方法将含有抗坏血酸培养基构建的兔BMSCs细胞膜片分为空白对照组和加压组（静态液压加载装置给予细胞膜片120 kPa、1 h/d、连续4 d的压力刺激），采用Western blot及real time-PCR检测P38MAPK信号通路的表达，real time-PCR技术检测成软骨基因的表达。结果兔BMSCs传代后细胞生长状态稳定，呈梭形；成骨诱导后，茜素红染色可观察到矿化结节，碱性磷酸酶活性染色呈阳性，成脂诱导后油红O染色细胞内出现红色脂滴；Western blot结果显示，与对照组（0.165±0.035）比较，压力刺激下兔BMSCs细胞膜片中P38MAPK蛋白表达量（0.820±0.108）显著升高（P＜0.05），同时real time-PCR结果表明：加压组兔BMSCs 细胞膜片中 P38MAPK 的 mRNA 表达量（0.651±0.105）高于对照组（0.166±0.046）（P＜0.05），且成软骨基因（Sox-9、Aggrecan及Col-Ⅱ）的mRNA表达量（3.323±0.729、0.901±0.162、1.151±0.105）也明显高于对照组（0.541±0.121、0.335±0.094、0.466±0.158）（ P ＜0.05）。结论 P38MAPK信号通路在压力调控BMSCs膜片成软骨响应中起正调节作用。
目的研究P38絲裂原激活的蛋白激酶（P38MAPK）信號通路在壓力調控骨髓間充質榦細胞（BMSCs）膜片成軟骨響應中作用。方法將含有抗壞血痠培養基構建的兔BMSCs細胞膜片分為空白對照組和加壓組（靜態液壓加載裝置給予細胞膜片120 kPa、1 h/d、連續4 d的壓力刺激），採用Western blot及real time-PCR檢測P38MAPK信號通路的錶達，real time-PCR技術檢測成軟骨基因的錶達。結果兔BMSCs傳代後細胞生長狀態穩定，呈梭形；成骨誘導後，茜素紅染色可觀察到礦化結節，堿性燐痠酶活性染色呈暘性，成脂誘導後油紅O染色細胞內齣現紅色脂滴；Western blot結果顯示，與對照組（0.165±0.035）比較，壓力刺激下兔BMSCs細胞膜片中P38MAPK蛋白錶達量（0.820±0.108）顯著升高（P＜0.05），同時real time-PCR結果錶明：加壓組兔BMSCs 細胞膜片中 P38MAPK 的 mRNA 錶達量（0.651±0.105）高于對照組（0.166±0.046）（P＜0.05），且成軟骨基因（Sox-9、Aggrecan及Col-Ⅱ）的mRNA錶達量（3.323±0.729、0.901±0.162、1.151±0.105）也明顯高于對照組（0.541±0.121、0.335±0.094、0.466±0.158）（ P ＜0.05）。結論 P38MAPK信號通路在壓力調控BMSCs膜片成軟骨響應中起正調節作用。
목적연구P38사렬원격활적단백격매（P38MAPK）신호통로재압력조공골수간충질간세포（BMSCs）막편성연골향응중작용。방법장함유항배혈산배양기구건적토BMSCs세포막편분위공백대조조화가압조（정태액압가재장치급여세포막편120 kPa、1 h/d、련속4 d적압력자격），채용Western blot급real time-PCR검측P38MAPK신호통로적표체，real time-PCR기술검측성연골기인적표체。결과토BMSCs전대후세포생장상태은정，정사형；성골유도후，천소홍염색가관찰도광화결절，감성린산매활성염색정양성，성지유도후유홍O염색세포내출현홍색지적；Western blot결과현시，여대조조（0.165±0.035）비교，압력자격하토BMSCs세포막편중P38MAPK단백표체량（0.820±0.108）현저승고（P＜0.05），동시real time-PCR결과표명：가압조토BMSCs 세포막편중 P38MAPK 적 mRNA 표체량（0.651±0.105）고우대조조（0.166±0.046）（P＜0.05），차성연골기인（Sox-9、Aggrecan급Col-Ⅱ）적mRNA표체량（3.323±0.729、0.901±0.162、1.151±0.105）야명현고우대조조（0.541±0.121、0.335±0.094、0.466±0.158）（ P ＜0.05）。결론 P38MAPK신호통로재압력조공BMSCs막편성연골향응중기정조절작용。
Objective To investigate the role of the P38 mitogen activated protein kinases ( P38MAPK) signal pathway in the mechanical pressure regulated chondrogenesis of bone marrow mesenchymal stem cell ( BMSC) sheets.Methods The rabbit BMSCs sheets obtained after being cultured by standard medium with Vitamin C were divided into control group and pressure group .The rabbits BMSCs sheets in the latter group were treated with 120 KPa hydrostatic pressure for 1 h/day with 4 consecutive days ,then the P38 MAPK protein and gene expressions were examined by Western blot and real time-PCR,respectively .Real time-PCR was also used to measure the chondrogenic gene expressions in BMSCs sheets .Results The subcultured rabbit BMSCs grew well with typical shuttle shape .The mineralized nodules were observed with alizarin Bordeaux staining and the cells were positive to alkaline phosphates staining while the red lipid droplet existed in adipose cells with oil red O staining after adipose induction .In addtition,Western blot confirmed the higher protein expression of P 38MAPK in rabbit BMSCs sheets in the pressure group(0.820 ±0.108)than that of the control group(0.165 ±0.035)(P<0.05).Furthermore,the Real Time-PCR showed us that the pressure increased the mRNA expression of P 38MAPK in rabbit BMSCs sheets (0.651 ±0.105 ) when comparing withthe control ones (0.166 ±0.046),and the mRNA levels of chondrogenic genes (Sox-9,Aggrecan and Col-Ⅱ) in rabbit BMSCs sheets in the pressure group ( 3.323 ±0.729 ,0.901 ±0.162 ,1.151 ±0.105 ) were significantly higher than those of the control group (0.541 ±0.121,0.335 ±0.094,0.466 ±0.158)(P<0.05). Conclusions The P38MAPK signal pathway plays a positive role in the mechanical pressure regulated chondrogenesis of BMSCs sheets .
