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J Zhejiang Univ (Med Sci)  2021, Vol. 50 Issue (3): 352-360    DOI: 10.3724/zdxbyxb-2021-0164
    
Salt-inducible kinase 2 regulates energy metabolism in rats with cerebral ischemia-reperfusion
ZHANG Ran12(),LIU Yun12,ZHANG Cui1,MA Mengyao1,LI Shu12,*(),HONG Yun3,*()
1. Department of Pathophysiology, Wannan Medical College, Wuhu 241002, Anhui Province, China;
2. Clinical Colloge of Wannan Medical College, Wuhu 241002, Anhui Province, China; 3. Department of Ultrasonic Medicine, Yijishan Hospital of Wannan Medical College, Wuhu 241001, Anhui Province, China
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Abstract  

Objective:To investigate the effects of salt-inducible kinase 2 (SIK2) on energy metabolism in rats with cerebral ischemia-reperfusion. Methods: Adult SD male rats (240-260?g) were divided into 5 groups: sham group, ischemia group, reperfusion group, adenovirus no-load group, and SIK2 overexpression group with 5 animals in each group. The middle cerebral artery occlusion (MCAO) was induced with the modified Zea-Longa line thrombus method to establish the cerebral ischemia reperfusion model. Eight days before the MCAO, SIK2 overexpression was induced by injecting 7 μL adenovirus in the right ventricle, then MCAO was performed for 2?h, followed by reperfusion 24?h. HE staining was used to observe the pathological changes of cerebral tissue in rats; TTC staining was used to observe the volume of cerebral infarct. The levels of adenosine triphosphate (ATP) and adenosine diphosphate (ADP) in rat brain tissue were detected by ELISA; the levels of SIK2 and hypoxia-inducible factor 1α (HIF-1α) in the rat brain tissues were detected by RT-qPCR and Western blotting. Results:Compared with the sham group, SIK2 level was decreased in the ischemia group, and it was further declined in the reperfusion group (P<0.05). Compared with the sham group and ischemic group, the pathological injury in reperfusion group were more severe, and the infarct size was larger; compared with the reperfusion group and adenovirus no-load group, the pathological injury of the SIK2 overexpression group was milder, and the infarct size is less. Compared with the sharn group, HIF-1α was increased in both ischemia group and reperfusion group, especially in ischemia group (allP<0.05); HIF-1α level in the SIK2 overexpression group was higher than that in the reperfusion group and adenovirus no-load group (allP<0.05). ATP level in ischemia group and reperfusion group was lower than that in the sham group, and the reperfusion group decreased more significantly than the ischemia group (P<0.05); ADP content was increased in the ischemia and reperfusion group, and the ADP content in reperfusion group was significantly higher than that in the ischemia group (P<0.05). ATP level in the SIK2 overexpression group was higher than that in the reperfusion group and adenovirus no-load group (allP<0.05), and ADP was decreased in the SIK2 overexpression group (allP<0.05).Conclusion:SIK2 can up-regulate the ATP level and down-regulate the ADP level in rat brain tissue and alleviate cerebral ischemia-reperfusion injury by increase the level of HIF-1α.



Key wordsIschemia reperfusion      Energy metabolism      Salt-inducible kinase 2      Hypoxia-inducible factor-1α      Adenosine triphosphate      Adenosine diphosphate      SD rat     
Received: 11 January 2021      Published: 16 August 2021
CLC:  R363  
Corresponding Authors: LI Shu,HONG Yun     E-mail: 18255366693@163.com;yxx2003@126.com
Cite this article:

ZHANG Ran,LIU Yun,ZHANG Cui,MA Mengyao,LI Shu,HONG Yun. Salt-inducible kinase 2 regulates energy metabolism in rats with cerebral ischemia-reperfusion. J Zhejiang Univ (Med Sci), 2021, 50(3): 352-360.

URL:

http://www.zjujournals.com/med/10.3724/zdxbyxb-2021-0164     OR     http://www.zjujournals.com/med/Y2021/V50/I3/352


盐诱导激酶2对脑缺血再灌注大鼠脑组织能量代谢的影响

目的:探究盐诱导激酶2(SIK2)对脑缺血再灌注大鼠能量代谢相关物质表达的影响。方法:选用成年SD雄性大鼠(240~260?g),参照改良Zea-Longa线栓法建立大鼠短暂性中动脉栓塞模型(MCAO),在构建MCAO模型前8?d予以右侧脑室注射7?μL腺病毒使大鼠脑组织SIK2过表达,随后建立缺血2?h再灌注24?h模型。实验分为手术对照组、缺血对照组、缺血再灌注组、腺病毒空载组及SIK2过表达组。苏木精-伊红(HE)染色观察大鼠神经细胞损伤的病理学改变;氯化三苯基四氮唑(TTC)染色观察大鼠脑组织梗死情况;酶联免疫吸附试验(ELISA)检测各组大鼠脑组织中腺苷三磷酸(ATP)、腺苷二磷酸(ADP)的含量;荧光定量聚合酶链反应和蛋白质印迹法检测各组大鼠脑组织中SIK2及缺氧诱导因子1α(HIF-1α)的表达量。结果:与手术对照组比较,缺血对照组及缺血再灌注组SIK2表达减少,且缺血再灌注组较缺血对照组减少更明显(P<0.05);与手术对照组和缺血对照组比较,缺血再灌注组病理损伤较重,梗死体积较大;与缺血再灌注组和腺病毒空载组比较,SIK2过表达组病理损伤较轻,梗死体积减少。与手术对照组比较,缺血对照组和缺血再灌注组HIF-1α表达均增加,其中缺血对照组增加更明显(均P<0.05);SIK2过表达组HIF-1α表达比缺血再灌注组和腺病毒空载组均增多(均P<0.05)。与手术对照组比较,缺血对照组和缺血再灌注组ATP含量均减少,且缺血再灌注组较缺血对照组减少更为显著(P<0.05);ADP含量在缺血对照组和缺血再灌注组均增多,且缺血再灌注组较缺血对照组明显增多(P<0.05);与缺血再灌注组和腺病毒空载组比较,SIK2过表达组ATP含量增加(均P<0.05),ADP含量减少(均P<0.05)。结论:SIK2可通过上调HIF-1α的表达,增加大鼠脑组织中ATP的含量,减少ADP含量,减轻大鼠脑缺血再灌注损伤。


关键词: 缺血再灌注,  能量代谢,  盐诱导激酶2,  缺氧诱导因子1α,  腺苷三磷酸,  腺苷二磷酸,  SD大鼠 
Figure 1 Electrophoretogram of SIK2 expression in brain tissues of sham group, ischemia group and ischemia-reperfusion group

组 别

n

mRNA

蛋白

手术对照组

4

0.73±0.09

0.98±0.05

缺血对照组

4

0.58±0.05

0.96±0.04

缺血再灌注组

4

0.46±0.10*

0.85±0.02*#

Table 1 Comparison of SIK2 levels among brain tissues of sham group, ischemia group and ischemia-reperfusion group
Figure 2 HE staining of brain tissues in each groups
Figure 3 Infarction of brain tissues in each group
Figure 4 Electro phoretogram of HIF-1α expression in each group

组 别

n

mRNA

蛋 白

手术对照组

5

0.92±0.11

0.77±0.10

缺血对照组

5

1.29±0.62*

1.00±0.00*

缺血再灌注组

5

1.03±0.10#

0.86±0.08#

Table 2 The level of HIF-1α in ischemia and reperfusion rats

组 别

n

mRNA

蛋白

缺血再灌注组

5

0.92±0.14

0.73±0.09

腺病毒空载组

5

1.00±0.07

0.75±0.08

SIK2过表达组

5

1.27±0.07*#

1.00±0.09*#

Table 3 The level of HIF-1α in ischemic-reperfusion rat brain tissue with SIK2 overexpression

组 别

n

腺苷三磷酸

腺苷二磷酸

手术对照组

5

6187±306

0.98±0.14

缺血对照组

5

5877±244

2.08±1.18

缺血再灌注组

5

5122±233*#

7.38±3.25*#

腺病毒空载组

5

5491±295

7.08±2.45

SIK2过表达组

5

7296±214△▲

0.58±0.28△▲

Table 4 The content of ATP and ADP in brain tissues of each group
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