1.湖南中医药大学第一附属医院,湖南省长沙市雨花区韶山中路95号,410007
2.湖南中医药大学
pengqinghua@hnucm.edu.cn
收稿:2024-03-11,
修回:2024-05-21,
纸质出版:2024-09-02
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欧晨,宋厚盼,谢薇等.滋阴明目方对视网膜色素变性模型小鼠视网膜组织凋亡及内质网应激的影响[J].中医杂志,2024,65(17):1808-1815.
OU Chen,SONG Houpan,XIE Wei,et al.The Effect of Ziyin Mingmu Formula (滋阴明目方) on Retinal Tissue Apoptosis and Endoplasmic Reticulum Stress in Retinitis Pigmentosa Model Mice[J].Journal of Traditional Chinese Medicine,2024,65(17):1808-1815.
欧晨,宋厚盼,谢薇等.滋阴明目方对视网膜色素变性模型小鼠视网膜组织凋亡及内质网应激的影响[J].中医杂志,2024,65(17):1808-1815. DOI: 10.13288/j.11-2166/r.2024.17.011.
OU Chen,SONG Houpan,XIE Wei,et al.The Effect of Ziyin Mingmu Formula (滋阴明目方) on Retinal Tissue Apoptosis and Endoplasmic Reticulum Stress in Retinitis Pigmentosa Model Mice[J].Journal of Traditional Chinese Medicine,2024,65(17):1808-1815. DOI: 10.13288/j.11-2166/r.2024.17.011.
目的
2
探讨滋阴明目方治疗视网膜色素变性的可能机制。
方法
2
将60只rd10小鼠采用随机数字表法分为模型组、维生素A组和滋阴明目方低、中、高剂量组,每组12只;另将12只C57BL/6小鼠作为空白组。滋阴明目方低、中、高剂量组小鼠分别给予滋阴明目方13.5、27.0、54.0 g/(kg·d)灌胃,维生素A组小鼠给予维生素A软胶囊750 IU/(kg·d)灌胃,空白组和模型组小鼠给予生理盐水13.5 ml/(kg·d)灌胃。各组均每天灌胃1次,共28天。运用眼底照相、光学相干断层扫描检测小鼠眼底、视网膜形态,TUNEL法检测小鼠视网膜组织细胞凋亡情况,微滴式数字PCR和免疫荧光双染检测小鼠视网膜组织葡萄糖调节蛋白78(GRP78)、1型内质网转膜蛋白激酶(IRE1)、活化转录因子6(ATF6)、C/EBP同源蛋白(CHOP) mRNA和蛋白表达水平。
结果
2
模型组小鼠视盘色蜡黄,眼底萎缩样改变,可见大片色素沉着,视网膜血管变细,可见大量血管白鞘,视网膜细胞核排列紊乱,可见大量凋亡细胞;滋阴明目方各剂量组和维生素A组小鼠眼底状况较模型组均有不同程度改善,凋亡细胞减少,其中以滋阴明目方中、高剂量组改善较显著。与空白组比较,模型组小鼠视网膜厚度减少,GRP78、IRE1、ATF6、CHOP mRNA及蛋白表达均升高(
P
<0.01)。与模型组比较,维生素A组和滋阴明目方中、高剂量组视网膜厚度均增加,GRP78、IRE1、ATF6 mRNA和蛋白表达均降低;滋阴明目方低、中、高剂量组CHOP mRNA和蛋白表达降低(
P
<0.05或
P
<0.01)。与维生素A组和滋阴明目方低剂量组比较,滋阴明目方高剂量组视网膜厚度增加(
P
<0.01)。与维生素A组比较,滋阴明目方中、高剂量组GRP78、ATF6、IRE1、CHOP mRNA表达降低,GRP78、CHOP蛋白表达降低(
P
<0.05或
P
<0.01)。与滋阴明目方低剂量组比较,滋阴明目方中、高剂量组CHOP蛋白表达降低(
P
<0.01)。
结论
2
滋阴明目方可以通过抑制内质网应激反应以减少视网膜组织的细胞凋亡,进而延缓视网膜病变进展,保护视功能,且具有剂量依赖性。
Objective
2
To study the effect of
Ziyin Mingmu Formula
(滋阴明目方) on retinitis pigmentosa mice and its possible mechanism.
Methods
2
Sixty rd10 mice were randomly divided into model group, vitamin A group, and
Ziyin Mingmu Formula
low-, medium-, and high-dose groups, 12 mice in each group; twelve C57BL/6 mice as the blank group. The mice in
Ziyin Mingmu Formula
low-, medium-, and high-dose groups were given 13.5, 27.0 and 54.0 g/(kg·d) of
Ziyin Mingmu Formula
by gavage, respectively; mice in the Vitamin A group were given 750 IU/(kg·d) of vitamin A soft capsule by gavage; mice in the blank group and model group were given 13.5 ml/(kg·d) of physiological saline by gavage. Each group was gavaged once a day for 28 days. Fundus photography and optical coherence tomography were used to detect the morphology of the fundus and retina, TUNEL method was used to detect apoptosis, microtitre digital PCR and immunofluorescence double staining were used to detect
glucose-regulated protein 78 (GRP78), inositol-requiring enzyme 1 (IRE1), activated transcription factor 6 (ATF6), and C/EBP homologous protein (CHOP) mRNA and protein expression levels.
Results
2
In the model group, optic discs were yellowish, fundus was atrophic with large pigmentation. The retinal blood vessels were thinned, a large number of vascular white sheaths were seen, the retinal cell nuclei disorganised, and a large number of apoptotic cells were seen; the fundus conditions of the mice in all dosages of
Ziyin Mingmu Formula
and the vitamin A group were improved to varying degrees compared with those of the model group, the apoptotic cells reduced, with
Ziyin Mingmu Formula
middle- and high-dose groups showing more significant improvement. Compared with the blank group, the retinal thickness of mice in the model group reduced, and the expression of GRP78, IRE1, ATF6, CHOP mRNA and protein elevated (
P
<0.01). Compared with the model group, retinal thickness increased and GRP78, IRE1, ATF6 mRNA and protein expression decreased in the vitamin A group and
Ziyin Mingmu Formula
medium- and high-dose groups; CHOP mRNA and protein expression decreased in low-, medium-, and high-dose groups of
Ziyin Mingmu Formula
(
P
<0.05 or
P
<0.01). Compared with vitamin A group and the low-dose group of
Ziyin Mingmu Formula
, retinal thickness increased in the high-dose group of
Ziyin Mingmu Formula
(
P
<0.01). Compared with vitamin A group, GRP78, ATF6, IRE1, and CHOP mRNA expression decreased in the medium- and high-dose groups of
Ziyin Mingmu Formula
, and GRP78 and CHOP protein expression decreased (
P
<0.05 or
P
<0.01). Compared with the low-dose group of
Ziyin Mingmu Formula
, CHOP protein expression reduced in the medium- and high-dose groups of
Ziyin Mingmu Formula
(
P
<0.01).
Conclusion
2
Ziyin Mingmu Formula
can reduce apoptosis of retinal tissues by inhibiting the endoplasmic reticulum stress response, thus slowing down the progression of retinopathy and protecting visual function in a dose-dependent manner.
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