Effects of cytochrome c on early steroid-induced femoral head necrosis

doi:10.3969/j.issn.2095-4344.2016.42.003

Abstract

Abstract:

BACKGROUND: Studies have shown that apoptosis is closely related to the pathogenesis of steroid-induced femoral head necrosis. The release of cytochrome c plays a very important role in the process of apoptosis.
OBJECTIVE: To observe the effects of cytochrome c on early steroid-induced femoral head necrosis.
METHODS: Twenty-four healthy adult male New Zealand rabbits at 5 months old were randomly divided into model group and control group (n=12 per group). Models of early steroid-induced femoral head necrosis were established by intragluteal injection of hormone combined with ear vein injection of horse serum. In the control group, rabbits were given ear vein injection of the same amount of physiological saline. At 2, 4 and 6 weeks after model establishment, histopathological changes of bilateral femoral head were observed by optical microscope, and the ratio of empty lacuna was calculated. Apoptosis of osteocytes was determined by TUNEL assay, and apoptotic index was calculated. Immunohistochemical method was used to determine cytochrome c and to calculate cytochrome c-positive expression rate.
RESULTS AND CONCLUSION: (1) The ratio of empty lacuna and apoptotic index: The model of early steroid-induced femoral head necrosis was successfully established in the experiment. Compared with the control group, ratio of empty lacuna, apoptotic index and expression rate of cytochrome c in osteocytes were significantly higher in the model group (P < 0.05 or P < 0.01). (2) Correlation analysis: Ratio of empty lacuna was significantly positively associated with apoptotic index at various time points in the model group (r=0.856, P < 0.01). Expression rate of cytochrome c was significantly positively associated with apoptotic index at various time points in the model group (r=0.824, P < 0.01). (3) These findings confirm that cytochrome c-involved apoptosis of osteocytes may play an important role in steroid-induced femoral head necrosis. Expression rate of cytochrome c in osteocytes is remarkably positively associated with the occurrence of steroid-induced femoral head necrosis in rabbits.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: Tissue Engineering, Apoptosis, Models, Animal

CLC Number:

R318

Ji Hao-yu1, Wang Xiao-long2 . Effects of cytochrome c on early steroid-induced femoral head necrosis[J]. Chinese Journal of Tissue Engineering Research, 2016, 20(42): 6252-6258.

Figures/Tables 1

2.1 实验动物数量分析兔24只均进入结果分析，实验过程无死亡等脱失现象的发生，全部进入结果分析。 2.2 早期激素性股骨头坏死模型兔骨组织病理学变化苏木精-伊红染色显示，模型组从建模后第2周后开始出现股骨头坏死表现，主要表现为软骨及软骨下松质出现空骨陷窝(图1A)，部分骨陷窝内骨细胞消失，骨细胞胞浆中充满脂滴，部分骨细胞胞核受挤压被挤向一边，髓腔内造血细胞减少，脂肪细胞增生肥大、数目增多(图1C)，骨小梁稍变细、甚至断裂。随着观察时间延长，实验组上述表现越明显。对照组骨小梁结构完整，排列规则，骨细胞清晰可见，中央可见较大、深染的细胞核，空骨陷窝少见，髓腔内造血细胞丰富，脂肪细胞较少，大小、形态正常(图1B)，股骨头软骨层完整、软骨细胞排列整齐(图1D)。模型组各时间点的股骨头空骨陷窝率均高于对照组，差异均有显著性意义(P < 0.05；表1)。 2.3 早期激素性股骨头坏死模型兔骨组织细胞凋亡变化 TUNEL检测显示，对照组在整个实验过程中凋亡骨细胞很少，凋亡细胞数相对稳定；而模型组从建模后第2周开始光学显微镜下可见较多胞浆染成棕黄色的TUNEL阳性细胞(即凋亡细胞)，随着造模时间的延长，凋亡细胞数逐渐增多，细胞凋亡指数逐渐增高。模型组各时间点的凋亡指数均明显高于与对照组，差异均有显著性意义(P < 0.01；表1)。 2.4 早期激素性股骨头坏死模型兔骨组织细胞色素c的阳性表达变化免疫组织化学染色显示，模型组从第2周后开始出现较多胞浆染成棕褐色的细胞色素c阳性骨细胞，随着造模时间的延长，细胞色素c阳性细胞数逐渐增多(图2)。模型组各时间点的骨细胞细胞色素c阳性表达率均明显高于对照组，差异均有显著性意义(P < 0.01；表1)。 2.5 相关性分析结果 Pearson相关性分析结果显示：模型组各时间点的细胞凋亡指数与空骨陷窝率呈明显正相关(r=0.856，P < 0.01)，且模型组的骨细胞细胞色素c阳性表达率与细胞凋亡指数同样呈明显正相关(r=0.824，P < 0.01)，差异均有显著性意义，表明在早期激素性股骨头坏死中，股骨头病变部位的细胞色素c与细胞凋亡指数具有显著相关性。"

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