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目的:研究硫酸酯化天麻多糖(GEPS)对皮质酮(CORT)诱导损伤的保护作用。方法:采用氨基磺酸-甲酰胺法经透析冷冻干燥等处理制得GEPS。在GEPS各剂量(0,250,500,1 000μg·ml~(-1))与PC12细胞孵育30 min之后,换用CORT作用48 h。采用CCK-8法判断细胞损伤程度;采用乳酸脱氢酶(LDH)释放法检测细胞凋亡率;采用DAPI染色法观察细胞凋亡形态。结果:GEPS各剂量组的PC12细胞存活率较模型组显著上升(P<0.05),且呈浓度依赖性。经GEPS作用后能有效缓解LDH的释放,与CORT损伤模型组相比,差异有统计学意义(P<0.05),且随着GEPS浓度的上升,LDH的释放率逐渐下降。倒置显微镜下可见CORT损伤模型组PC12细胞折光度下降,突触收缩,细胞聚集,贴壁不良,有许多呈悬浮状,但经过GEPS预处理的PC12细胞随着GEPS浓度的上升,生存状态明显好转。与CORT致损伤组相比,GEPS各浓度组凋亡的细胞核形态学改变有所改善,凋亡细胞明显减少。结论:GEPS对CORT诱导PC12细胞损伤具有一定的保护作用,但与天麻多糖(GEP)相比提高不大,其他方面的生物学活性有待进一步研究。
Objective: To study the protective effect of sulfated Gastrodia Polysaccharide (GEPS) on cortisone (CORT) -induced injury. METHODS: GEPS was prepared by sulfamic acid-formamide method after dialysis and freeze-drying. The cells were incubated with PC12 cells for 30 min at each dose of GEPS (0, 250, 500 and 1000 μg · ml -1) and switched to CORT for 48 h. The degree of cell damage was determined by CCK-8 method. The apoptosis rate was detected by lactate dehydrogenase (LDH) release assay. The apoptosis morphology was observed by DAPI staining. Results: The survival rate of PC12 cells in each dose group of GEPS was significantly higher than that of model group (P <0.05), and in a dose-dependent manner. The effect of GEPS on LDH release was relieved. Compared with CORT model, the difference was statistically significant (P <0.05). The release rate of LDH decreased with the increase of GEPS concentration. Under inverted microscope, PC12 cells in CORT injury group showed decreased refraction, synaptic contraction, cell aggregation and poor adherentity, and many were suspended. However, the survival rate of PC12 cells pretreated with GEPS increased significantly with the increase of GEPS concentration . Compared with CORT-induced injury group, the morphological changes of apoptotic nuclei in GEPS groups were improved, and apoptotic cells were significantly reduced. CONCLUSION: GEPS can protect PC12 cells against CORT-induced injury in some degree, but not significantly up-regulated compared with GEP. The biological activity of GEPS remains to be further studied.