活化T淋巴细胞核因子5在高盐诱导小鼠平滑肌细胞衰老中的作用及其机制

doi:10.13481/j.1671-587X.20250302

Abstract

Abstract:

Objective To discuss the role of nuclear factor of activated T-cells 5 （NFAT5） inhibitor KRN5 in high salt-induced senescence of mouse vascular smooth muscle cells （VSMCs）， and to clarify its mechanism. Methods Thirty 8-week-old male ApoE^-/- mice were divided into normal group， senescence group and high-salt treatment senecence group， with 10 mice in each group； the mice in senescence group and high-salt treatment senecence group were used to establish natural senecence mouse models； the mouse VSMCs were isolated and cultured， and divided into normal group， senescence group， high-salt treatment senecence group and high-salt treatment senecence+KRN5 group. β-galactosidase （Sa-β-gal） staining was used to detect the senescence of aortic tissues and VSMCs in various groups； immunofluorescence method was used to detect the expressions of NFAT5 and phosphorylated histone H2A variant X （γ-H2AX） proteins in mouse aortic tissues and VSMCs in various groups； real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the mRNA expression levels of NFAT5， γ-H2AX， cyclin-dependent kinase inhibitor 2A （P16） and cyclin-dependent kinase inhibitor 1A （P21） in the cells in various groups； Western blotting method was used to detect the protein expression levels of NFAT5， γ-H2AX， P16 and P21 in VSMCs in various groups. Results The Sa-β-gal staining results showed that compared with normal group， the proportions of senescence-positive area in aortic tissues of the mice in senescence group and high-salt treatment senecence group were significantly increased （P<0.05）， and the proportion of senescence-positive cells in the VSMCs of the mice was significantly increased （P<0.05）； compared with senecence group， the proportion of senescence-positive cells in the VSMCs mice in high-salt treatment senecence group was significantly increased （P<0.05）； compared with high-salt treatment senecence group， the proportion of senescence-positive cells in the VSMCs of the mice in high-salt treatment senecence+KRN5 group was significantly decreased （P<0.01）. The immunofluorescence results showed that compared with normal group， the expression level of γ-H2AX protein in mouse VSMCs of the mice in senescence group was significantly increased （P<0.05）； compared with senescence group， the expression levels of SA-β-gal staining and NFAT5 protein in aortic tissue of the mice in high-salt treatment senecence group were significantly increased （P<0.05）； compared with normal group， the expression level of NFAT5 protein in the VSMCs of the mice in senecence group and high-salt treatment senecence group was significantly increased （P<0.05）； compared with senecence group， the expression level of NFAT5 protein in the VSMCs of the mice in high-salt treatment senecence group was significantly increased （P<0.05）. The RT-qPCR results showed that compared with normal group， the expression levels of NFAT5， γ-H2AX， P16， and P21 mRNA in the VSMCs of the mice in senescence group and high-salt treatment senecence group were significantly increased （P<0.05）； compared with senecence group， the mRNA expression levels of NFAT5， γ-H2AX， P16， and P21 mRNA in the VSMCs of the mice in high-salt treatment senecence group were significantly increased （P<0.05）； compared with senecence group， the expression levels of NFAT5， γ-H2AX， P16， and P21 mRNA in the VSMCs of the mice in high-salt treatment senecence group and high-salt treatment senecence+KRN5 group were significantly increased （P<0.05）； compared with high-salt treatment senecence group， the mRNA expression levels of NFAT5， γ-H2AX， P16， and P21 in the VSMCs of the mice in high-salt treatment senecence+KRN5 group were significantly decreased （P<0.05）. The Western blotting results showed that compared with normal group， the expression levels of NFAT5， γ-H2AX， P16， and P21 proteins in the VSMCs of the mice in senescence group and high-salt treatment senecence group were significantly increased （P<0.05）； compared with senescence group， the expression levels of NFAT5， γ-H2AX， P16， and P21 proteins in the VSMCs of the mice in high-salt treatment senecence group were significantly increased （P<0.05）； compared with senecence group， the expression levels of NFAT5， γ-H2AX， P16， and P21 proteins in the VSMCs of the mice in high-salt treatment senecence group and high-salt treatment senecence+KRN5 group were significantly increased （P<0.05）； compared with high-salt treatment senecence group， the expression levels of NFAT5， γ-H2AX， P16， and P21 proteins in the VSMCs of the mice in high-salt treatment senecence+KRN5 group were significantly decreased （P<0.05）. Conclusion NFAT5 may play a promoting role in high salt-induced senescence of the mouse VSMCs.

Key words: Vascular aging, Nuclear factor of activated T-cells 5, KRN5, Vascular smooth muscle cells, β-galactosidase

CLC Number:

R339.38

Wei ZHONG,Zhiyin DAI,Xinggang CUI,Bo LI,Yu JIANG. Effect of nuclear factor of activated T lymphocytes 5 on senescence of smooth muscle cells of mice induced by high-salt and its mechanism[J].Journal of Jilin University(Medicine Edition), 2025, 51(3): 567-575.

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References 19

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Group	Proportion of positive area
Normal	1.00±0.00
Senescence	0.22±0.04^*
High-salt treatment senescence	0.33±0.05^*

Group	Proportion of positive area ofsenescence cells
Normal	0.00±0.00
Senescence	0.44±0.04^*
High-salt treatment senescence	0.80±0.02^*

Group	Proportion of positive area
Senescence	0.42±0.02
High-salt treatment senescence	0.78±0.07^*
High-salt treatment senescence+KRN5	0.27±0.03^*△

Group	Expression of NFAT5
Normal	0.00±0.00
Senescence	0.52±0.04^*
High-salt treatment senescence	0.70±0.02^*△

Group	NFAT5	γ-H2AX	P16	P21
Normal	1.00±0.00	1.00±0.00	1.00±0.00	1.00±0.00
Senescence	2.34±0.08^*	2.69±0.22^*	1.91±0.18^*	1.84±0.13^*
High-salt treatment senescence	3.07±0.15^*△	3.42±0.09^*△	2.18±0.08^*△	2.35±0.19^*△

Effect of nuclear factor of activated T lymphocytes 5 on senescence of smooth muscle cells of mice induced by high-salt and its mechanism

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