外源褪黑素对鹿茸软骨细胞生长发育的影响

doi:10.16431/j.cnki.1671-7236.2020.07.004

摘要/Abstract

摘要： 试验旨在探讨褪黑素对体外培养的鹿茸软骨细胞增殖、周期及凋亡的影响。利用甲苯胺蓝、茜素红S、阿利新蓝染色鉴定软骨细胞，采用外源添加褪黑素的方法，用不同浓度（0、400、800、1 200、1 600和2 000 pg/mL）、不同时间（24、48和72 h）处理鹿茸软骨细胞，CCK-8法检测细胞增殖。选取800 pg/mL褪黑素处理软骨细胞24 h，利用流式细胞仪检测细胞周期的分布及细胞凋亡情况，采用ELISA试剂盒检测细胞培养液中睾酮含量。结果显示，经不同浓度的褪黑素处理鹿茸软骨细胞不同时间后，800 pg/mL褪黑素处理鹿茸软骨细胞24 h细胞活力极显著增加（P<0.01）。软骨细胞经褪黑素处理后，与对照组相比，褪黑素处理组细胞G1期比例显著降低（P<0.05）；G2期比例极显著增加（P<0.01），细胞被阻滞在G2期；褪黑素处理组细胞早期凋亡率无显著变化（P>0.05），晚期凋亡率显著下降（P<0.05）；ELISA检测睾酮分泌水平均显著上升（P<0.05）。综上，外源添加800 pg/mL褪黑素可以促进鹿茸软骨细胞的增殖，抑制软骨细胞的晚期凋亡，促进睾酮的分泌；以上结果可为阐明褪黑素参与鹿茸生长发育机制提供参考。

关键词: 鹿茸; 褪黑素; 软骨细胞; 增殖

Abstract: This study was aimed to investigate the effects of melatonin on proliferation,cell cycle and apoptosis of antler cartilages cultured in vitro.The chondrocytes were identified by toluidine blue,alizarin red S and alexin blue staining,the exogenous addition of melatonin was used at different concentrations (0,400,800,1 200,1 600 and 2 000 pg/mL),velvet antler chondrocytes were treated at different times (24,48 and 72 h),and cell proliferation was detected by CCK-8 method.The chondrocyte was treated with 800 pg/mL melatonin for 24 h.The cell cycle distribution and apoptosis were detected by flow cytometry.The testosterone content in the cell culture medium was determined by ELISA kit.The results showed that after treatment with different concentrations of melatonin for different time,the cell viability of velvet cartilage cells treated with 800 pg/mL melatonin for 24 h was extremely significantly increased (P<0.01).After treatment with melatonin in chondrocytes,the proportion of G1 phase in melatonin-treated group was significantly lower than that in control group (P<0.05).The proportion of G2 phase was extremely significantly increased (P<0.01),and the cells were arrested in G2 phase.There was no significant change in the early apoptotic rate of melatonin treatment group (P>0.05),and the late apoptotic rate was significantly decreased (P<0.05).The testosterone secretion level was significantly increased by ELISA (P<0.05).In conclusion,the addition of 800 pg/mL melatonin could promote the proliferation of antler chondrocytes and inhibit the late apoptosis of chondrocytes to promote the secretion of testosterone,which provided a reference for elucidating the mechanism of melatonin involved in the growth and development of antler.

Key words: velvet antler; melatonin; chondrocytes; proliferation

中图分类号:

S825

李可可, 孙旭阳, 陈超, 荆焕松, 许学林, 游思, 赵靖波, 熊家军. 外源褪黑素对鹿茸软骨细胞生长发育的影响[J]. 中国畜牧兽医, 2020, 47(7): 1997-2004.

LI Keke, SUN Xuyang, CHEN Chao, JING Huansong, XU Xuelin, YOU Si, ZHAO Jingbo, XIONG Jiajun. Effect of Exogenous Melatonin on the Growth and Development of Antler Chondrocytes[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(7): 1997-2004.

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