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[1]許勇,薛彬,孟祥超,等.伸筋活血湯對(duì)低氧誘導(dǎo)因子-1α缺失小鼠椎間盤(pán)退變的影響…[J].中醫(yī)正骨,2018,30(01):12-17.
　XU Yong,XUE Bin,MENG Xiangchao,et al.Effect of Shenjin Huoxue Tang(伸筋活血湯)on intervertebral disc degeneration in mice with hypoxia-inducible factor-1α deficiency[J].The Journal of Traditional Chinese Orthopedics and Traumatology,2018,30(01):12-17.
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伸筋活血湯對(duì)低氧誘導(dǎo)因子-1α缺失小鼠椎間盤(pán)退變的影響…()
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《中醫(yī)正骨》[ISSN:1001-6015/CN:41-1162/R]

卷:
第30卷
期數(shù):
2018年01期
頁(yè)碼:
12-17
欄目:
基礎(chǔ)研究
出版日期:
2018-01-20

文章信息/Info

Title:
Effect of Shenjin Huoxue Tang(伸筋活血湯)on intervertebral disc degeneration in mice with hypoxia-inducible factor-1α deficiency
作者:
許勇; 薛彬; 孟祥超; 李中偉; 奚小冰; 李飛躍
上海交通大學(xué)醫(yī)學(xué)院附屬瑞金醫(yī)院,上海 200025
Author(s):
XU Yong; XUE Bin; MENG Xiangchao; LI Zhongwei; XI Xiaobing; LI Feiyue
Ruijin Hospital affiliated to Shanghai Jiaotong University medical college,Shanghai 200025,China
關(guān)鍵詞:
椎間盤(pán)退行性變;  缺氧誘導(dǎo)因子1;  伸筋活血湯;  小鼠;  動(dòng)物實(shí)驗(yàn)
Keywords:
Keywords intervertebral disc degeneration;  hypoxia-inducible factor-1;  Shenjin Huoxue Tang;  mice;  animal experimentation
摘要:
目的:觀(guān)察伸筋活血湯對(duì)低氧誘導(dǎo)因子(hypoxia-inducible factor,HIF)-1α缺失小鼠椎間盤(pán)退變的影響。方法:通過(guò)雜交繁殖獲得同窩HIF-1α缺失小鼠和正常小鼠。取2、4、8周齡HIF-1α缺失小鼠及正常小鼠,每個(gè)時(shí)間點(diǎn)2種小鼠各取5只,處死后取L3～L5節(jié)段腰椎組織制備切片,分別進(jìn)行HE染色和番紅-固綠染色。取16只4周齡HIF-1α缺失小鼠,隨機(jī)分為2組,每組8只,分別以伸筋活血湯和生理鹽水灌胃。灌胃每次0.12 mL,每天1次,共30 d。處死小鼠取L3～L5節(jié)段腰椎組織制備切片,分別進(jìn)行HE染色、番紅-固綠染色及免疫組化染色后觀(guān)察。結(jié)果:HE染色顯示,與正常小鼠相比,2周齡HIF-1α缺失小鼠髓核占整個(gè)椎間盤(pán)組織的比例減少,髓核中的細(xì)胞仍以含有大液泡的脊索細(xì)胞為主,但已有大量髓核細(xì)胞變小,髓核內(nèi)細(xì)胞結(jié)構(gòu)排列稍紊亂; 4周齡HIF-1α缺失小鼠椎間盤(pán)內(nèi)的細(xì)胞種類(lèi)和數(shù)量迅速下降,髓核細(xì)胞排列紊亂; 8周齡HIF-1α缺失小鼠髓核細(xì)胞數(shù)量減少十分明顯,纖維組織填充于髓核中,細(xì)胞外基質(zhì)增加。番紅-固綠染色顯示,與正常小鼠相比,2周齡HIF-1α缺失小鼠椎間盤(pán)中聚集蛋白聚糖減少,4周齡HIF-1α缺失小鼠椎間盤(pán)中聚集蛋白聚糖減少更加明顯,8周齡HIF-1α缺失小鼠髓核出現(xiàn)裂痕及鈣化,髓核骨化明顯。4周齡HIF-1α缺失小鼠經(jīng)藥物干預(yù)30 d后,HE染色結(jié)果顯示,伸筋活血湯組小鼠椎間盤(pán)終板無(wú)明顯破損和骨化,髓核細(xì)胞數(shù)量較多,排列較整齊; 生理鹽水組小鼠椎間盤(pán)髓核內(nèi)細(xì)胞呈梭形,體積變小,數(shù)量明顯減少,排列嚴(yán)重紊亂,細(xì)胞外基質(zhì)增加,髓核中出現(xiàn)裂痕,纖維組織填充于髓核中,甚至出現(xiàn)終板破壞。番紅-固綠染色結(jié)果顯示,伸筋活血湯組小鼠骨和軟骨終板染色明顯,椎間盤(pán)髓核內(nèi)細(xì)胞數(shù)量較多,椎間盤(pán)骨化現(xiàn)象不明顯; 生理鹽水組骨和軟骨終板染色較淺,出現(xiàn)裂痕及鈣化,細(xì)胞數(shù)量極少。免疫組化染色結(jié)果顯示,伸筋活血湯組聚集蛋白聚糖陽(yáng)性染色的細(xì)胞數(shù)量明顯較多,陽(yáng)性信號(hào)多分布在細(xì)胞內(nèi); 生理鹽水組聚集蛋白聚糖陽(yáng)性染色的細(xì)胞數(shù)量較少,陽(yáng)性信號(hào)多分布在細(xì)胞外。生理鹽水組聚集蛋白聚糖陽(yáng)性染色細(xì)胞的灰度值小于伸筋活血湯組(129.889±9.839,154.333±9.622,t=0.200,P=0.000)。結(jié)論:HIF-1α缺失可導(dǎo)致小鼠椎間盤(pán)退變,伸筋活血湯能夠延緩HIF-1α缺失小鼠的椎間盤(pán)退變。
Abstract:
ABSTRACT Objective:To observe the effect of Shenjin Huoxue Tang(伸筋活血湯,SJHXT)on intervertebral disc degeneration in mice with hypoxia-inducible factor(HIF)-1α deficiency.Methods:The littermate HIF-1α deficiency mice and normal mice were obtained through crossbreeding.Two-,4- and 8-week-old HIF-1α deficiency mice and normal mice were fetched,and 5 cases were randomly selected from each kind of mice at each time point and were executed,and then their vertebral tissues of L3-L5 were fetched out and were sectioned for HE staining and safranin O-fast green staining.Sixteen 4-week-old HIF-1α deficiency mice were fetched and were randomly divided into 2 groups,8 cases in each group,and they were intragastric administrated with SJHXT(SJHXT group)and normal saline(NS group)respectively,0.12 mL at a time,once a day for consecutive 30 days.Then the mice were executed,and their vertebral tissues of L3-L5 were fetched out and were sectioned for HE staining,safranin O-fast green staining and immunohistochemical staining.Results:The HE staining results showed that,compared to normal mice,(1)the proportion of nucleus pulposus to whole intervertebral disc of 2-week-old HIF-1α deficiency mice decreased,and the cells in nucleus pulposus were still dominated by notochord cells containing large vacuoles,and a large number of nucleus pulposus cells shrinked and their intracellular structure was slightly disorganized;(2)the cell type and number decreased rapidly in intervertebral disc and nucleus pulposus cells were disorganized in 4-week-old HIF-1α deficiency mice;(3)the nucleus pulposus cells decreased obviously and the extracellular matrix increased and the nucleus pulposus was filled with fibrous tissues in 8-week-old HIF-1α deficiency mice.The safranin O-fast green staining results showed that,compared to normal mice,the aggrecan in intervertebral disc decreased in 2-week-old HIF-1α deficiency mice and decreased more obviously in 4-week-old HIF-1α deficiency mice; and cracks,calcifications and obvious ossification appeared in nucleus pulposus in 8-week-old HIF-1α deficiency mice.After 30-day drug intervention on 4-week-old HIF-1α deficiency mice,the HE staining results showed that(1)no obvious damage and ossification were found in intervertebral disc endplate and a large number of nucleus pulposus cells arranged in order in mice of SJHXT group;(2)the cells in intervertebral disc nucleus pulposus of mice in NS group decreased in volume and number and presented with fusiform cells and arranged in great disorder,moreover,the extracellular matrix increased and cracks and a large amount of fibrous tissues were found in nucleus pulposus and the destroyed endplate was also found.The safranin O-fast green staining results showed that(1)the stains in bony and cartilaginous endplates were obvious in mice of SJHXT group,moreover,there were abundant cells in intervertebral disc nucleus pulposus,and the intervertebral disc ossification was not so obvious;(2)the stains presented with light colour in bony and cartilaginous endplates of mice in NS group,moreover,cracks,calcifications and few cells appeared in bony and cartilaginous endplates.The immunohistochemical staining results showed that many cells with positive aggrecan staining were found in SJHXT group and few cells with positive aggrecan staining were found in NS group; and the positive signals mainly distributed inside the cells in SJHXT group and outside the cells in NS group.The gray values of cells with positive aggrecan staining were less in NS group compared to SJHXT group(129.889+/-9.839,154.333+/-9.622,t=0.200,P=0.000).Conclusion:HIF-1α deficiency can lead to intervertebral disc degeneration in mice,and SJHXT can delay intervertebral disc degeneration in mice with HIF-1α deficiency.

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備注/Memo

備注/Memo:
基金項(xiàng)目:上海市中醫(yī)藥傳承和創(chuàng)新項(xiàng)目(ZY3-CCCX-1-1014); 上海市名老中醫(yī)學(xué)術(shù)經(jīng)驗(yàn)研究工作室建設(shè)項(xiàng)目(SHGZS-2017010); 2017年全國(guó)名老中醫(yī)藥專(zhuān)家傳承工作室建設(shè)項(xiàng)目(MLZJGZS-2017001) 通訊作者:李飛躍 E-mail:[email protected]
更新日期/Last Update: 2018-06-02