CAJ | 학술논문

背景:目前用于组织工程韧带的支架材料胶原蛋白、聚乳酸、聚羟基乙酸、小肠黏膜下层、聚糖及纳米材料等均有降解速度快的特点,应用于宿主后有一定炎症反应.目的:观察蚕丝支架的机械强度和体外降解性,以及与巨噬细胞的反应.设计:对照观察.单位:实验于2004-09/2005-01在华中科技大学附属协和医院骨科完成.材料:市售白色家蚕蚕丝(20/22D),每30根蚕丝平行排列为一束支架.置入煮沸的5g/LNa2CO3中进行脱胶,Na2CO3溶液体积(mL)与生丝的质量(g)之比为1000.方法:①体外降解实验:8cm长蚕丝支架干燥后测初质量,然后分别浸泡于磷酸盐缓冲液和用磷酸盐缓冲液配制的1.0g/L胶原酶中,浸泡12周后测试蚕丝支架残质量,计算失质量率.同时进行拉伸试验,测量样品的极限抗拉强度.②单核细胞株RAW264.7的培养:将丝状支架组、对照组、脂多糖组均加入2×108 L-1巨噬细胞混悬液1mL中培养,培养第1,7天收集各组细胞培养上清,采用ELISA方法测定TNF-α含量.主要观察指标:①丝状支架在磷酸盐缓冲液和胶原酶中失重率和极限抗拉强度变化.②培养第1,7天各组细胞培养上清中TNF-α含量.结果:①丝状支架在胶原酶中8周后质量减少已超过50%,在磷酸盐缓冲液中未见明显改变.②胶原酶消化8周后丝状支架的极限抗拉强度下降超过原来的50%,在磷酸盐缓冲液中未见明显变化.③培养第1,7天丝状支架组巨噬细胞分泌少量TNF-α,脂多糖组上清中的TNF-α水平明显高于丝状支架组(P<0.01),丝状支架组与对照组TNF-α水平无差异(P>0.05).结论:丝状支架经12周降解后仍保持良好机械特性,培养第1,7天与巨噬细胞混合培养中,巨噬细胞对该材料具有免疫惰性.
배경:목전용우조직공정인대적지가재료효원단백、취유산、취간기을산、소장점막하층、취당급납미재료등균유강해속도쾌적특점,응용우숙주후유일정염증반응.목적:관찰잠사지가적궤계강도화체외강해성,이급여거서세포적반응.설계:대조관찰.단위:실험우2004-09/2005-01재화중과기대학부속협화의원골과완성.재료:시수백색가잠잠사(20/22D),매30근잠사평행배렬위일속지가.치입자비적5g/LNa2CO3중진행탈효,Na2CO3용액체적(mL)여생사적질량(g)지비위1000.방법:①체외강해실험:8cm장잠사지가간조후측초질량,연후분별침포우린산염완충액화용린산염완충액배제적1.0g/L효원매중,침포12주후측시잠사지가잔질량,계산실질량솔.동시진행랍신시험,측량양품적겁한항랍강도.②단핵세포주RAW264.7적배양:장사상지가조、대조조、지다당조균가입2×108 L-1거서세포혼현액1mL중배양,배양제1,7천수집각조세포배양상청,채용ELISA방법측정TNF-α함량.주요관찰지표:①사상지가재린산염완충액화효원매중실중솔화겁한항랍강도변화.②배양제1,7천각조세포배양상청중TNF-α함량.결과:①사상지가재효원매중8주후질량감소이초과50%,재린산염완충액중미견명현개변.②효원매소화8주후사상지가적겁한항랍강도하강초과원래적50%,재린산염완충액중미견명현변화.③배양제1,7천사상지가조거서세포분비소량TNF-α,지다당조상청중적TNF-α수평명현고우사상지가조(P<0.01),사상지가조여대조조TNF-α수평무차이(P>0.05).결론:사상지가경12주강해후잉보지량호궤계특성,배양제1,7천여거서세포혼합배양중,거서세포대해재료구유면역타성.
BACKGROUND: Presently, the biomaterial used in ligament tissue engineering such as collagen protein, polylactic acid, polyglycolic acid, small intestinal submucosa, glycan and nanomaterial are characterized by rapid degradation, resulting in inflammatory reaction after applying in host.OBJECTIVE: To investigate mechanical strength and in vitro degradation of silk scaffold and explore the reaction to macrophages.DESIGN: Controlled experiment.SETTING: Experiments were performed at the Department of Orthopaedics, Union Hospital, Huazhong University of Science and Technology from September 2004 to January 2005.MATERIALS: White raw Bombyx mori silkworm fibers of size 20/22 (according to the manufacturer) were obtained from the market. Bundles of 30 parallel fibers were prepared for a bundle of scaffold, which was put into fervens 5g/L Na2CO3for degumming. Ratio of Na2CO3 solution (Ml) to raw silk (g) was 1000.METHODS: In vitro degradation: 8cm long silk scaffold was weighed after drying. Subsequently, the silk scaffold was separately dipped into phosphate buffer saline (PBS) and 1.0g/L collagenase prepared with PBS. Twelve weeks later, silk scaffold was weighed to calculate weight loss rate. Simultaneously, tensile test was performed to detect the ultimate tensile strength (UTS) of samples. Culture of monocyte strain RAW264.7:2×108L-1 macrophage suspension (1mL) were separately added in a silk scaffold group, a control group and a lipopolysaccharide (LPS) group. At days 1 and 7, cell supernatant was collected from each group. Tumor necrosis factor-α(TNF-α) levels were measured by enzyme linked immunosorbent assay (ELISA).MAIN OUTCOME MEASURES: ① Changes in weight loss rate and UTS of the silk matrices after incubated with collagenase and the PBS. ②TNF-αlevels in the supernatant of each groups at days 1 and 7.RESULTS: Mass of silk matrices reduced by over 50% after incubated with collagenase for 8 weeks, but no change was found in PBS. UTS decreased by over 50% 8 weeks after incubated with collagenase, but no change was detected in PBS. At days 1 and 7, TNF-α levels in the supernatant was less in the silk scaffold group; TNF-α levels in the supernatant was significantly higher in the LPS group than in the silk scaffold group (P<0.01), but no significant difference in TNF-α levels was measured between the silk scaffold group and the control group (P>0.05).CONCLUSION: After 12-weeks degradation, silk scaffold still has good mechanical properties. Macrophages possess immunological inertia at days 1 and 7 after inoculated with macrophages.