外源性硫化氢通过上调SIRT1延缓心肌早衰抑制尿毒症大鼠心肌纤维化

doi:10.11958/20220662

天津医药 ›› 2022, Vol. 50 ›› Issue (12): 1276-1281.doi: 10.11958/20220662

外源性硫化氢通过上调SIRT1延缓心肌早衰抑制尿毒症大鼠心肌纤维化

刘达¹(), 肖婷², 梁彪³, 宋熊¹, 王森¹, 赵俊雄¹, 杨军¹^,^△()

1 南华大学附属第一医院心血管内科（邮编421000）
2 深圳市龙华区中心医院心血管内科
3 宁乡市人民医院心血管内科

收稿日期:2022-05-05 修回日期:2022-05-26 出版日期:2022-12-15 发布日期:2022-12-30
通讯作者: 杨军 E-mail:945268433@qq.com;yangjunketizu@163.com
作者简介:刘达（1996），男，硕士在读，主要从事心血管疾病方面研究。E-mail：945268433@qq.com
基金资助:
国家自然科学基金面上项目(81870230);湖南省卫生健康委员会科技计划重点项目(20201913);湖南省自然科学基金科卫联合基金项目(2019JJ80033);深圳市龙华区医疗卫生机构区级科研项目(2020035)

Exogenous H₂S delays premature myocardial aging and inhibits myocardial fibrosis in uremic rats by upregulating SIRT1

LIU Da¹(), XIAO Ting², LIANG Biao³, SONG Xiong¹, WANG Sen¹, ZHAO Junxiong¹, YANG Jun¹^,^△()

1 Department of Cardiology, the First Affiliated Hospital of University of South China, Hengyang 421000, China
2 Department of Cardiology, Longhua District Central Hospital
3 Department of Cardiology, People's Hospital of Ningxiang

Received:2022-05-05 Revised:2022-05-26 Published:2022-12-15 Online:2022-12-30
Contact: YANG Jun E-mail:945268433@qq.com;yangjunketizu@163.com

刘达, 肖婷, 梁彪, 宋熊, 王森, 赵俊雄, 杨军. 外源性硫化氢通过上调SIRT1延缓心肌早衰抑制尿毒症大鼠心肌纤维化[J]. 天津医药, 2022, 50(12): 1276-1281.

LIU Da, XIAO Ting, LIANG Biao, SONG Xiong, WANG Sen, ZHAO Junxiong, YANG Jun. Exogenous H₂S delays premature myocardial aging and inhibits myocardial fibrosis in uremic rats by upregulating SIRT1[J]. Tianjin Medical Journal, 2022, 50(12): 1276-1281.

摘要/Abstract

摘要：

目的探讨外源性硫化氢（H₂S）对尿毒症心肌病（UCM）大鼠心肌早衰和心肌纤维化的影响。方法雄性SD大鼠48只按随机数字表法分成假手术组（Sham组）、模型组（UCM组）、H₂S治疗组（UCM+H₂S组）以及H₂S对照组（H₂S组）。其中UCM组、UCM+H₂S组用经典5/6肾切除法造模，而Sham组及H₂S组仅游离双肾。建模后UCM+H₂S组和H₂S组大鼠每日注射硫氢化钠（NaHS，50 μmol/kg），Sham组和UCM组注射同剂量生理盐水。干预8周，心脏超声检测大鼠左心室的短轴缩短率（LVFS）；Masson染色法评估心肌纤维化情况，计算胶原容积分数（CVF）；免疫组化染色检测心肌Ⅲ型胶原表达；β-半乳糖苷酶染色检测衰老心肌细胞；蛋白免疫印迹法检测心肌中沉默信息调节因子-1（SIRT1）、NOD样受体家族3（NLRP3）、白细胞介素（IL）-1β、P21、P19、P53蛋白表达。结果与Sham组比较，UCM组LVFS明显下降，心肌CVF、Ⅲ型胶原的阳性表达率、β-半乳糖苷酶染色阳性率升高，NLRP3、IL-1β、P19、P21及P53表达增多，SIRT1蛋白表达减少（P<0.05）；而UCM+H₂S组较UCM组大鼠心肌CVF、Ⅲ型胶原、β-半乳糖苷酶染色阳性率降低，NLRP3、IL-1β、P19、P21及P53蛋白表达量减少，SIRT1蛋白表达增多（P<0.05）。结论外源性H₂S能改善尿毒症大鼠心肌纤维化，机制可能与其上调SIRT1抑制心肌早衰有关。

关键词: 硫化氢, 衰老, 尿毒症, NLR家族，含PYRIN结构域蛋白3, 沉默信息调节因子1, 心肌纤维化

Abstract:

Objective To investigate the effect of hydrogen sulfide (H₂S) on premature myocardial aging and myocardial fibrosis in uremic rats. Methods Forty-eight male SD rats were randomly divided into the sham operation group (Sham group), the model group (uremic cardiomyopathy, UCM group), the H₂S treatment group (UCM+H₂S group) and the H₂S group. Rats in the UCM group and the UCM+H₂S group were modeled by the classic 5/6 nephrectomy method. After the model was established, rats in the UCM+H₂S group and the H₂S group were injected with sodium hydrosulfide (NaHS, 50 μmol/kg) every day for 8 weeks, while the Sham group and the UCM group were injected with the same amount of normal saline. Left ventricular shortening rate (LVFS) was measured by cardiac ultrasound. Masson staining was used to evaluate myocardial fibrosis, and collagen volume fraction (CVF) was calculated. The expression of type Ⅲ collagen in myocardium was detected by immunohistochemistry. β -galactosidase staining was used to detect senescent myocardial cells. The expression levels of SIRT1, NLRP3, interleukin (IL) -1β, P21, P19 and P53 proteins in myocardium were measured by Western blot assay. Results Compared with the Sham group, LVFS was significantly decreased in the UCM group, the relative expression levels of CVF, type Ⅲ collagen and the positive rate of β -galactosidase staining were increased, the expressions of NLRP3, IL-1β, P19, P21 and P53 were increased, and the expression of SIRT1 protein was decreased (P<0.05). Compared with the UCM group, the relative expression levels of CVF, type Ⅲ collagen and positive staining rate of β-galactosidase were decreased in the UCM+H₂S group, and the protein expressions of NLRP3, IL-1β, P19, P21 and P53 were decreased, while the protein expression of SIRT1 was increased (P<0.05). Conclusion Exogenous H₂S can improve myocardial fibrosis in uremic rats, and the mechanism may be related to the upregulation of SIRT1 and the inhibition of premature myocardial senescence.

Key words: hydrogen sulfide, aging, uremia, NLR family, pyrin domain-containing 3 protein, silent information regulator-1, myocardial fibrosis

中图分类号:

R542.2+3

图/表 7

表1 各组大鼠血清BUN、Scr比较（$\bar{x}\pm s$）

Tab.1 Comparison of serum BUN and Scr between the four groups

组别	n	BUN（mmol/L）	Scr（μmol/L）
Sham组	12	11.11±2.43	31.75±6.02
UCM组	8	25.09±7.04^a	109.50±12.45^a
UCM+H₂S组	9	23.13±6.23^a	106.20±11.69^a
H₂S组	11	10.67±1.94	32.14±5.61
F		27.000^＊	158.100^＊

图1 各组大鼠心功能超声图像

Fig.1 Ultrasound image of rat heart function in each group

图2 各组大鼠心肌组织胶原纤维Masson染色（×400）

Fig.2 Masson staining of collagen fibers in myocardial tissue of rats in each group (×400)

图3 各组大鼠心肌组织Ⅲ型胶原表达情况（免疫组化染色，×400）

Fig.3 Expression of type Ⅲ collagen in myocardial tissue of rats in each group (immunohistochemical staining, ×400)

图4 各组大鼠左室心肌组织衰老情况（β-半乳糖苷酶染色，×400）

Fig.4 The aging level of myocardial tissue of rats in each group (β -galactosidase staining, ×400)

图5 各组大鼠心肌组织中SIRT1、P21、P19、P53及NLRP3、IL-1β蛋白表达

Fig.5 The protein expression levels of SIRT1, P21, P19，P53, NLRP3 and IL-1β in myocardial tissue of rats in each group

表3 各组大鼠心肌SIRT1、P21、P19、P53及NLRP3、IL-1β蛋白表达水平比较（n=7，$\bar{x}\pm s$）

Tab.3 Comparison of SIRT1, P21, P19, P53, NLRP3 and IL-1β protein expression levels in myocardial tissue of rats between the four groups

组别	SIRT1	P21	P19
Sham组	0.87±0.07	0.69±0.02	1.18±0.08
UCM组	0.69±0.05^a	0.93±0.04^a	1.46±0.03^a
UCM+H₂S组	0.83±0.04^b	0.68±0.02^b	1.25±0.03^b
H₂S组	0.83±0.01	0.69±0.02	1.29±0.04
F	6.381^＊	51.420^＊	14.150^＊
组别	P53	NLRP3	IL-1β
Sham组	0.57±0.01	0.68±0.03	0.79±0.07
UCM组	0.79±0.02^a	0.89±0.04^a	1.16±0.10^a
UCM+H₂S组	0.60±0.01^b	0.84±0.02^b	0.92±0.08^b
H₂S组	0.59±0.01	0.69±0.04	0.81±0.06
F	96.370^＊	23.890^＊	11.980^＊

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外源性硫化氢通过上调SIRT1延缓心肌早衰抑制尿毒症大鼠心肌纤维化

Exogenous H₂S delays premature myocardial aging and inhibits myocardial fibrosis in uremic rats by upregulating SIRT1

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外源性硫化氢通过上调SIRT1延缓心肌早衰抑制尿毒症大鼠心肌纤维化

Exogenous H2S delays premature myocardial aging and inhibits myocardial fibrosis in uremic rats by upregulating SIRT1

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Exogenous H₂S delays premature myocardial aging and inhibits myocardial fibrosis in uremic rats by upregulating SIRT1