中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2014年
11期
2515-2517
,共3页
孟繁钢%何爱珊%张志奇%张紫机%康焱%杨子波%龙毅%林子洪%吴刚
孟繁鋼%何愛珊%張誌奇%張紫機%康焱%楊子波%龍毅%林子洪%吳剛
맹번강%하애산%장지기%장자궤%강염%양자파%룡의%림자홍%오강
透明质酸%骨髓间充质干细胞%组织工程软骨
透明質痠%骨髓間充質榦細胞%組織工程軟骨
투명질산%골수간충질간세포%조직공정연골
Hyaluronic acid%Bone marrow mesenchymal stem cells%Tissue-engineering cartilage
目的 观察透明质酸对入骨髓间充质干细胞(hBMSCs)复合磷酸三钙-胶原材料体外构建组织工程软骨的影响.方法 将提取扩增的第3代hBMSCs体外与组织工程支架磷酸三钙-胶原复合,分两组进行三维培养:对照组和实验组(对照组培养液基础上加入100 mg/L透明质酸).培养2周后进行苏木素-伊红(HE)染色、阿尔新蓝染色、Ⅱ型胶原(COL2)免疫组织化学染色,实时定量逆转录聚合酶链反应(RT-PCR)和糖胺多糖(GAG)定量检测,比较体外成软骨效果.结果 HE染色结果显示骨髓间充质干细胞能够均匀分布于材料中,实验组细胞基质的分泌大于对照组.实验组阿尔新蓝染色,COL2免疫组织化学染色,COL2、软骨特异性基SOX9、聚集蛋白聚糖(Aggrecan)的基因表达分别为8.954±2.612/1.062±0.459、3.512 ±0.638/1.111 ±0.595、4.588±1.964/1.042±0.335,糖胺多糖(GAG)定量检测结果(21.741±2.633/16.717±0.595)也都显著高于对照组,表明100 mg/L透明质酸能够有效促进hBMSCs复合磷酸三钙-胶原体外成软骨.在COL10和COL1的基因表达上实验组也大于对照组(21.741 ±2.633/16.717 ±0.595),说明透明质酸除了能体外诱导hBMSCs成软骨分化外,还能进一步诱导其成熟发展.结论 100 mg/L透明质酸能够在三维组织工程支架中促进hBMSCs体外成软骨分化,并且能够进一步诱导其成熟发展.
目的 觀察透明質痠對入骨髓間充質榦細胞(hBMSCs)複閤燐痠三鈣-膠原材料體外構建組織工程軟骨的影響.方法 將提取擴增的第3代hBMSCs體外與組織工程支架燐痠三鈣-膠原複閤,分兩組進行三維培養:對照組和實驗組(對照組培養液基礎上加入100 mg/L透明質痠).培養2週後進行囌木素-伊紅(HE)染色、阿爾新藍染色、Ⅱ型膠原(COL2)免疫組織化學染色,實時定量逆轉錄聚閤酶鏈反應(RT-PCR)和糖胺多糖(GAG)定量檢測,比較體外成軟骨效果.結果 HE染色結果顯示骨髓間充質榦細胞能夠均勻分佈于材料中,實驗組細胞基質的分泌大于對照組.實驗組阿爾新藍染色,COL2免疫組織化學染色,COL2、軟骨特異性基SOX9、聚集蛋白聚糖(Aggrecan)的基因錶達分彆為8.954±2.612/1.062±0.459、3.512 ±0.638/1.111 ±0.595、4.588±1.964/1.042±0.335,糖胺多糖(GAG)定量檢測結果(21.741±2.633/16.717±0.595)也都顯著高于對照組,錶明100 mg/L透明質痠能夠有效促進hBMSCs複閤燐痠三鈣-膠原體外成軟骨.在COL10和COL1的基因錶達上實驗組也大于對照組(21.741 ±2.633/16.717 ±0.595),說明透明質痠除瞭能體外誘導hBMSCs成軟骨分化外,還能進一步誘導其成熟髮展.結論 100 mg/L透明質痠能夠在三維組織工程支架中促進hBMSCs體外成軟骨分化,併且能夠進一步誘導其成熟髮展.
목적 관찰투명질산대입골수간충질간세포(hBMSCs)복합린산삼개-효원재료체외구건조직공정연골적영향.방법 장제취확증적제3대hBMSCs체외여조직공정지가린산삼개-효원복합,분량조진행삼유배양:대조조화실험조(대조조배양액기출상가입100 mg/L투명질산).배양2주후진행소목소-이홍(HE)염색、아이신람염색、Ⅱ형효원(COL2)면역조직화학염색,실시정량역전록취합매련반응(RT-PCR)화당알다당(GAG)정량검측,비교체외성연골효과.결과 HE염색결과현시골수간충질간세포능구균균분포우재료중,실험조세포기질적분비대우대조조.실험조아이신람염색,COL2면역조직화학염색,COL2、연골특이성기SOX9、취집단백취당(Aggrecan)적기인표체분별위8.954±2.612/1.062±0.459、3.512 ±0.638/1.111 ±0.595、4.588±1.964/1.042±0.335,당알다당(GAG)정량검측결과(21.741±2.633/16.717±0.595)야도현저고우대조조,표명100 mg/L투명질산능구유효촉진hBMSCs복합린산삼개-효원체외성연골.재COL10화COL1적기인표체상실험조야대우대조조(21.741 ±2.633/16.717 ±0.595),설명투명질산제료능체외유도hBMSCs성연골분화외,환능진일보유도기성숙발전.결론 100 mg/L투명질산능구재삼유조직공정지가중촉진hBMSCs체외성연골분화,병차능구진일보유도기성숙발전.
Objective To investigate the effect of hyaluronan (HA) on engineered cartilage formation.Methods Human mesenchymal stem cell (hBMSCs) were isolated by density gradient centrifugation and were seeded onto tricalcium phosphate-collagen (TCP-COL) scaffolds.hBMSCs were cultured on the scaffold for 2 weeks in two groups:(1) control group ; (2) HA group (control group culture medium with 100 mg/L HA).The effect of hyaluronan on chondrogenesis was assessed by hematoxylin-eosin (HE) staining,alcian blue staining,anti type Ⅱ collagen immunohistochemical staining,Real-time polymerase chain reaction (RT-PCR) and glycosaminoglycans (GAG) quantification assay.Results The data of hematoxylin-eosin (HE) staining,alcian blue staining,anti type Ⅱ collagen immunohistochemical staining,RT-PCR [Collagen Ⅱ (COL2) (P ＜ 0.01),SRY-related high mobility group-box gene9 (SOX9) (P ＜ 0.01),Aggrecan (P ＜ 0.05)] (8.954 ± 2.612/1.062 ± 0.459,3.512 ± 0.638/1.111 ± 0.595 and 4.588 ± 1.964/1.042 ± 0.335,respectively) and GAG quantification assay (21.741 ± 2.633/16.717 ± 0.595) (P ＜ 0.05) shows HA (100 g/L) could significantly induce the chondrogenic differentiation of hBMSCs on the TCP-COL scaffold.In addition,in the HA group,a significant up-regulation of COL1 (P＜0.05) (3.258 ±1.038/1.127 ±0.678) andCOL10 (P＜0.05) (5.961 ±2.598/1.080 ±0.458) gene were detected compared with the control group,indicating that HA may also promote robust maturation of tissue-engineered cartilage.Conclusion HA not only could significantly induce the chondrogenic differentiation of hBMSCs on the TCP-COL scaffolds,but also could promote maturation of the tissue-engineered cartilage.
