CAJ | 학술논문

目的探讨纤维蛋白支架对神经干细胞和星形胶质细胞分化及增殖的影响.方法分别培养胚胎大鼠脊髓来源的神经干细胞和新生鼠脊髓神经胶质细胞,接种于纤维蛋白支架上,同时用多聚赖氨酸修饰的玻片作为对照.于体外培养不同时间后,用神经丝蛋白(NF200)对神经细胞进行免疫荧光染色,测量各复孔(n=4)内NF阳性细胞的突起长度,计算其平均值;用胶质纤维酸性蛋白(GFAP)对胶质细胞进行染色,各复孔(n=4)内统计5个不同视野的胶质细胞总数和GFAP阳性细胞数,计算GFAP阳性细胞相对数量的平均值.比较在纤维蛋白支架和玻片上神经干细胞分化、神经纤维延伸及神经胶质细胞增殖的差异.同时用免疫印迹技术对荧光染色结果进行验证.上述实验各重复3次.结果纤维蛋白支架组的NF阳性纤维明显长于对照组,GFAP阳性星形胶质细胞相对数量明显少于对照组,GFAP的表达水平明显低于对照组.结论纤维蛋白支架可促进神经干细胞向神经细胞分化,并有利于神经纤维的延伸而抑制星形胶质细胞的增殖和成熟.
목적 탐토섬유단백지가대신경간세포화성형효질세포분화급증식적영향.방법 분별배양배태대서척수래원적신경간세포화신생서척수신경효질세포,접충우섬유단백지가상,동시용다취뢰안산수식적파편작위대조.우체외배양불동시간후,용신경사단백(NF200)대신경세포진행면역형광염색,측량각복공(n=4)내NF양성세포적돌기장도,계산기평균치;용효질섬유산성단백(GFAP)대효질세포진행염색,각복공(n=4)내통계5개불동시야적효질세포총수화GFAP양성세포수,계산GFAP양성세포상대수량적평균치.비교재섬유단백지가화파편상신경간세포분화、신경섬유연신급신경효질세포증식적차이.동시용면역인적기술대형광염색결과진행험증.상술실험각중복3차.결과 섬유단백지가조적NF양성섬유명현장우대조조,GFAP양성성형효질세포상대수량명현소우대조조,GFAP적표체수평명현저우대조조.결론 섬유단백지가가촉진신경간세포향신경세포분화,병유리우신경섬유적연신이억제성형효질세포적증식화성숙.
Objective To investigate the effects of the fibrin scaffold on the differentiation and the proliferation of neural stem cells and astrocytes. Methods Neural stem cells and the gliocytes derived from spinal cord were cultured in vitro respectively. The purified neural stem cells or gliocytes were seeded separately onto the fibrin scaffolds as experimental group and the glass slides modified with poly-L-lysine(PLL)as control group. At different time in culture the neural stem cells were immunofluorescence stained with antibodies against the marker of neurons I.e. Neurofilament(NF).The length of NF-positive neuritis was masured and the average value was calculated in the culture well (n=4). The gliocytes were immunofluorescence stained with antibodies against the marker of astrocytes I.e. Glial fibrillary acidic protein (GFAP ). The total number of the cells and the GFAP-positive cells were counted from 5 different fields of vision in the culture well (n=4), then the average ratio of GFAP-positive cells was calculated. The differentiation of neural stem cells, the extension of neurites and the proliferation of astrocytes on the fibrin scaffolds were compared with those on the slides. The protein of GFAP was detected by Western blotting to analyse the mature degree of astrocytes. All above experiments were repeated 3 times respectively. Results Immunofluorescence staining showed that the NF-positive neurites in the fibrin scaffold group were longer than those in the control group, whereas GFAP-positive cells were fewer than those in the control group. The expression of GFAP in the cells on the scaffold was lower than that in the control group.Conclusion The fibrin scaffold could promote differentiation of the neural stem cells to neurons and extension of the neurites. Meanwhile, the scaffold could inhibit proliferation and mature of the astrocytes.