Pilose antler regulates the expression of ADAMTS-4/TIMP-3 gene in the extracellular matrix in a rabbit model of osteoarthritis

doi:10.3969/j.issn.2095-4344.0366

Abstract

Abstract:

BACKGROUND: Pilose antler has been shown to be effective for osteoarthritis, but its association with ADAMTS-4/TIMP-3 gene (main composition of extracellular matrix) is rarely reported.
OBJECTIVE: To investigate the effect of pilose antler on the expression of ADAMTS-4/TIMP-3 gene in rabbit models of osteoarthritis, and to further elucidate the mechanism of pilose antler in the prevention and treatment of osteoarthritis.
METHODS: Forty-two healthy female New Zealand white rabbits were selected and used for establishing the models of osteoarthritis by anterior cruciate ligament rupture. The rabbit models were then randomized into three groups: control group (normal saline), low-dose (0.21 g/kg) and high-dose (0.84 g/kg) pilose antler groups. After intervention for 9 weeks, the content of hydroxyproline in both knees was detected. The expression levels of mRNA of cartilage target genes in each group were detected by RT-PCR. Protein expression levels of aggrecan and collagen type II were determined by western blot assay.
RESULTS AND CONCLUSION: Compared with the control group, the content of hydroxyproline in cartilage in the low-dose and high-dose pilose antler groups was significantly increased, and the content in the high-dose pilose antler group was significantly higher than that in the low-dose pilose antler group (P < 0.05). The order of mRNA expression level of ADAMTS-4 was as follows: high-dose pilose antler group < low-dose pilose antler group < control group (P < 0.05). The order of mRNA expression levels of metallopeptidase inhibitor 3, aggrecan and collagen type II was as follows: high-dose pilose antler group > low-dose pilose antler group > control group (P < 0.05). The expression levels of aggrecan and collagen type II were the highest in the high-dose pilose antler group, followed by the low-dose pilose antler group, and the lowest in the control group (P < 0.05). Our findings indicate that pilose antler can repair the injured cartilage by inhibiting the secretion of ADAMTS-4, increasing the secretion of metallopeptidase inhibitor 3 and preventing the degradation of aggrecan and collagen type II.

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: Drugs, Chinese Herbal, Osteoarthritis, Extracellular Matrix, Cartilage, Tissue Engineering

CLC Number:

R318

Sun Zhi-tao1, Feng Hua-long2, He Sheng-hua1, Lai Ju-yi1, Huang Fei-qiang2, Wang Ye-guang1. Pilose antler regulates the expression of ADAMTS-4/TIMP-3 gene in the extracellular matrix in a rabbit model of osteoarthritis[J]. Chinese Journal of Tissue Engineering Research, 2018, 22(32): 5133-5138.

Figures/Tables 3

2.1 实验动物数量分析 42只新西兰大白兔全部进入结果分析，无脱落。 2.2 造模后兔软骨形态变化及病理改变鹿茸干预2疗程后高剂量鹿茸组可见关节面不平整，平台边缘可见较浅的软骨溃瘍，软骨表面有毛糙；低剂量鹿茸组软骨显示关节面有较深廉烂，软骨下骨隐约可见，关节边缘软组织增生明显；对照组关节面软骨全层剥脱，软骨下骨显露，关节边缘软组织增生明显(图1)。高剂量鹿茸组光镜下可见表层软骨细胞肿胀变圆，扁平的软骨细胞减少，软骨细胞簇集；低剂量鹿茸组表层及中层细胞收缩，细胞数量弥漫性增多，出现大量簇集细胞团；对照组大部分软骨的表层细胞消失，软骨表面出现溃烂缺损，软骨细胞数量减少，可见坏死崩解细胞，潮线紊乱甚至消失(图2)。 2.3 各组羟脯氨酸含量的比较高剂量鹿茸组湿质量和干质量组羟脯氨酸的含量显著高于低剂量鹿茸组(t=3.41，P=0.00；t=2.73，P=0.02)。高剂量鹿茸组湿质量和干质量组羟脯氨酸的含量显著高于对照组 (t=6.56，P=0.00；t=6.97，P=0.00)。低剂量鹿茸组湿质量和干质量组的羟脯氨酸含量显著高于对照组 (t=3.15，P=0.01；t=4.24，P=0.00)，表1。 2.4 各组软骨中靶基因mRNA的表达高剂量鹿茸组ADAMTS-4 mRNA的表达显著低于低剂量鹿茸组和对照组(t=3.05，P=0.01；t=7.24，P=0.00)；低剂量鹿茸组ADAMTS-4 mRNA的表达显著低于对照组(t=4.19，P=0.00)。而高剂量鹿茸组TIMP-3、蛋白聚糖、Ⅱ型胶原蛋白mRNA的表达显著高于低剂量鹿茸组和对照组(t=2.45，P=0.03；t=2.57，P=0.02；t=3.76，P=0.00；t=6.02，P=0.00；t=6.36，P=0.00；t=6.74，P=0.00)；低剂量鹿茸组TIMP-3、蛋白聚糖、Ⅱ型胶原蛋白mRNA的表达显著高于对照组(t=3.57，P=0.00；t=3.79，P=0.00；t=2.97，P=0.01)，见表2。 2.5 各组软骨中蛋白聚糖和Ⅱ型胶原蛋白的表达高剂量鹿茸组蛋白聚糖的表达显著高于低剂量鹿茸组和对照组(t=2.15，P=0.048；t=7.28，P=0.00)；低剂量鹿茸组蛋白聚糖的表达显著高于对照组(t=5.13，P=0.00)。高剂量鹿茸组Ⅱ型胶原蛋白的表达显著高于低剂量鹿茸组和对照组(t=6.29，P=0.00；t=8.72，P=0.00)；低剂量鹿茸组Ⅱ型胶原蛋白的表达显著高于对照组(t=2.43，P=0.03)，见表3，图3。"

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