SHJH hr 小鼠的心脏衰老表型研究

doi:10.12300/j.issn.1674-5817.2024.100

实验动物与比较医学 ›› 2025, Vol. 45 ›› Issue (1): 13-20.DOI: 10.12300/j.issn.1674-5817.2024.100

• 实验动物资源开发与利用 • 上一篇下一篇

SHJH ^hr 小鼠的心脏衰老表型研究

刘荣乐¹(), 程灏¹, 尚付生², 常书福¹()(), 徐平³

^1.复旦大学附属中山医院, 上海 200032
^2.上海大学转化医学研究院, 上海 200444
^3.上海吉辉实验动物饲养有限公司, 上海 201611

收稿日期:2024-07-19 修回日期:2024-12-11 出版日期:2025-03-12 发布日期:2025-02-25
通讯作者: 常书福（1981—），男，博士，副主任医师，研究方向：心脏衰老和冠心病。E-mail：chang.shufu@zs-hospital.sh.cn。 ORCID： 0000-0003-4190-3655
作者简介:刘荣乐（1988—），男，博士，主治医师，研究方向：动脉粥样硬化和衰老。E-mail：liu.rongle@zs-hospital.sh.cn
基金资助:
上海市科技创新行动计划项目“自发性早衰老小鼠的心血管衰老相关表型分析及机制研究”(21140900103)

Study on Cardiac Aging Phenotypes of SHJH ^hr Mice

LIU Rongle¹(), CHENG Hao¹, SHANG Fusheng², CHANG Shufu¹()(), XU Ping³

^1.Zhongshan Hospital, Fudan University, Shanghai 200032, China
^2.Institute of Translational Medicine, Shanghai University, Shanghai 200444, China
^3.Shanghai Jihui Laboratory Animal Care Co. , Ltd. , Shanghai 201611, China

Received:2024-07-19 Revised:2024-12-11 Published:2025-02-25 Online:2025-03-12
Contact: CHANG Shufu (ORCID: 0000-0003-4190-3655), E-mail: chang.shufu@zs-hospital.sh.cn

摘要/Abstract

摘要：

目的探讨自发性早衰SHJH ^hr 小鼠心脏的衰老现象。方法对不同周龄（10周龄和24周龄）的SHJH ^hr 小鼠（SHJH ^hr 组）和野生型ICR小鼠［用作野生组（wild-type， WT组）］进行对比研究。用小动物活体超声影像系统分析心脏功能，安死术后采集各脏器并称重，了解心脏萎缩情况；苏木精-伊红（hematoxylin-eosin，HE）染色后观察心脏病理损伤情况；Masson染色后分析心脏纤维化情况；麦胚凝集素（wheat-germ agglutinin，WGA）染色后分析心脏心肌细胞面积；酶联免疫吸附法检测心肌组织中氧化损伤指标包括超氧化物歧化酶（superoxide dismutase，SOD）、谷胱甘肽过氧化物酶（glutathione peroxidase，GPX）和过氧化氢酶（catalase，CAT）的活性以及8-羟基-2'-脱氧鸟苷（8-hydroxy-2'-deoxyguanosine，8-OHdG）含量；实时荧光定量PCR法检测炎症、纤维化、氧化应激相关标志因子的mRNA表达水平；比色法检测丙二醛（malondialdehyde，MDA）水平。结果与同周龄的WT组小鼠相比，10周龄SHJH ^hr 组小鼠的每搏输出量（stroke volume，SV）、射血分数（ejection fraction，EF）、缩短分数（fractional shortening，FS）和心肺重量无明显差异；但是当小鼠达到24周龄时，SHJH ^hr 组小鼠的SV、EF和FS值明显低于同周龄WT组小鼠（均P<0.05），肺脏重量无显著变化，但是心脏重量显著降低（P<0.05）。选取24周龄小鼠心脏组织进行组织学分析发现：与WT组小鼠相比，SHJH ^hr 组小鼠的心脏纤维化水平无明显差异，但WGA染色显示心肌细胞面积显著减少（P<0.05）。PCR检测发现氧化应激标志因子Sod2、Gpx1和Cat基因的mRNA水平显著下调（均P<0.05）。生化检测发现心肌组织中氧化损伤相关酶SOD、GPX和CAT活性显著降低（均P<0.05），而氧化应激标志物8-OHdG和MDA水平显著增加（均P<0.05）。结论 SHJH ^hr 小鼠心脏出现早衰现象，可能与心肌组织氧化应激损伤有关。

关键词: SHJH ^hr 小鼠, 心脏衰老, 氧化损伤

Abstract:

Objective To investigate the spontaneous premature cardiac aging in SHJH ^hr mice. Methods A comparative study was conducted between SHJH ^hr mice (SHJH ^hr group) and wild-type ICR mice (WT group) at different ages (10 and 24 weeks). Cardiac function was analyzed using a small animal in vivo ultrasound imaging system. After euthanasia, organs were collected and weighed to assess the extent of cardiac atrophy. Cardiac pathological damage was observed using hematoxylin-eosin (HE) staining. Cardiac fibrosis was analyzed using Masson staining. Myocardial cell area was analyzed after wheat germ agglutinin (WGA) staining. The activities of oxidative damage indicators in myocardial tissue, including superoxide dismutase (SOD), glutathione peroxidase (GPX), and catalase (CAT), as well as the content of 8-hydroxy-2'-deoxyguanosine (8-OHdG), were measured using enzyme-linked immunosorbent assay. Real-time fluorescence quantitative PCR was used to measure the mRNA expression levels of factors associated with inflammation, fibrosis, and oxidative stress. Colorimetric assay was used to measure malondialdehyde (MDA) levels. Results Compared to WT group mice of the same age, 10-week-old mice in the SHJH ^hr group showed no significant differences in stroke volume (SV), ejection fraction (EF), fractional shortening (FS), or heart and lung weights. However, at 24 weeks of age, mice in the SHJH ^hr group had significantly lower SV, EF, and FS values compared to mice of the same age in the WT group (P<0.05), with no significant change in lung weight but a significant reduction in heart weight (P<0.05). Histological analysis of heart tissue from 24-week-old mice revealed no significant difference in cardiac fibrosis levels between SHJH ^hr and WT groups, but WGA staining showed a significant reduction in myocardial cell area in mice in the SHJH ^hr group (P<0.05). PCR analysis revealed a significant downregulation of mRNA levels of oxidative stress factors Sod2, Gpx1, and Cat genes (P<0.05). Biochemical assays indicated significantly reduced activities of oxidative damage-related enzymes SOD, GPX, and CAT in myocardial tissue (P<0.05), while the levels of oxidative damage markers 8-OHdG and MDA significantly increased (P<0.05). Conclusion Mice in the SHJH ^hr group exhibit premature cardiac aging, which may be associated with oxidative stress damage in myocardial tissue.

Key words: SHJH ^hr mice, Cardiac aging, Oxidative damage

中图分类号:

R-332

刘荣乐,程灏,尚付生,等. SHJH ^hr 小鼠的心脏衰老表型研究[J]. 实验动物与比较医学, 2025, 45(1): 13-20. DOI: 10.12300/j.issn.1674-5817.2024.100.

LIU Rongle,CHENG Hao,SHANG Fusheng,et al. Study on Cardiac Aging Phenotypes of SHJH ^hr Mice[J]. Laboratory Animal and Comparative Medicine, 2025, 45(1): 13-20. DOI: 10.12300/j.issn.1674-5817.2024.100.

图/表 5

表1 目的基因的PCR引物序列

Table 1 Primer sequences of target genes used for real-time fluorescent quantitative PCR

基因名称 Name of genes	引物序列 Sequence (5’→3’)
Sod2	CAGACCTGCCTTACGACTATGG; CTCGGTGGCGTTGAGATTGTT
Sod3	CCTTCTTGTTCTACGGCTTGC; TCGCCTATCTTCTCAACCAGG
Gpx1	CCACCGTGTATGCCTTCTCC; AGAGAGACGCGACATTCTCAAT
Cat	GGAGGCGGGAACCCAATAG; GTGTGCCATCTCGTCAGTGAA
Il1β	GCAACTGTTCCTGAACTCAACT; ATCTTTTGGGGTCCGTCAACT
Il6	CCAAGAGGTGAGTGCTTCCC; CTGTTGTTCAGACTCTCTCCCT
Tnf	CACGCTCTTCTGTCTACTGAACTTC; CTTGGTGGTTTGTGAGTGTGAGG
Ccl2	TAAAAACCTGGATCGGAACCAAA; GCATTAGCTTCAGATTTACGGGT
Tgfβ1	CCACCTGCAAGACCATCGAC; CTGGCGAGCCTTAGTTTGGAC
Col1α1	GCTCCTCTTAGGGGCCACT; ATTGGGGACCCTTAGGCCAT
Actα2	GTCCCAGACATCAGGGAGTAA; TCGGATACTTCAGCGTCAGGA
GAPDH	AGAAGGTGGTGAAGCAGGCATC; CGAAGGTGGAAGAGTGGGAGTTG

图1 心脏超声检测不同周龄SHJH hr 小鼠的心脏功能注：A，多普勒超声图；B～E，心功能指标的组间比较：每搏输出量（SV）、射血分数（EF）、缩短分数（FS）和二尖瓣前向血流e峰与a峰的比值（E'/A'）。以非早衰的野生型ICR小鼠（WT组）作为对照，n=6；*P<0.05。

Figure 1 Cardiac ultrasound examination of SHJH hr mice cardiac function at different agesNote： A， Doppler ultrasound images； B to E， Intergroup comparison of cardiac function indicators： stroke volume （SV）， ejection fraction （EF）， fractional shortening （FS）， and the ratio of the early （E'） to late （A'） mitral inflow velocities （E'/A'）. Wild-type ICR mice without premature aging （WT group） were used as controls， n=6； *P<0.05.

图2 不同周龄SHJH hr 小鼠的肺脏和心脏重量注：A，肺脏重量/胫骨长度比率；B，心脏重量/胫骨长度比率。以非早衰的野生型ICR小鼠（WT组）作为对照，n=6，*P<0.05。

Figure 2 Lung and heart weights of SHJH hr mice at different agesNote： A， Lung weight to tibia length ratio； B， Heart weight to tibia length ratio. Wild-type ICR mice without premature aging （WT group） were used as controls， n=6， *P<0.05.

图3 24周龄SHJH hr 小鼠的心脏组织病理和相关因子转录水平分析注：A，HE、Masson 和WGA染色的代表性图片；B，Masson染色显示心肌纤维化水平统计；C，WGA染色显示心肌WGA表达细胞所占面积分析；D，实时荧光定量PCR（RFQ-PCR）检测心肌细胞中Il1β、Il6、Tnf和Ccl2基因的mRNA表达水平；E，RFQ-PCR检测心肌细胞中Tgfβ1、Col1α1和Actα2的mRNA表达水平。以非早衰的野生型ICR小鼠（WT组）作为对照，n=6；与WT组比较，*P <0.05。

Figure 3 Analysis of cardiac histopathology and related factor transcriptional levels in 24-week-old SHJH hr miceNote： A， Representative images of HE， Masson， and WGA staining； B， Statistical analysis of myocardial fibrosis levels shown by Masson staining； C， Analysis of the area occupied by WGA-positive cells in myocardial tissue as shown by WGA staining； D， Real-time fluorescent quantitative PCR （RFQ-PCR） measurement of mRNA expression levels of Il1β， Il6， Tnf， and Ccl2 in cardiomyocytes； E， RFQ-PCR measurement of mRNA expression levels of Tgfβ1， Col1α1， and Actα2 in cardiomyocytes. Wild-type ICR mice without premature aging （WT group） were used as controls， n=6. Compared with the WT group， *P <0.05.

图4 24周龄SHJH hr 小鼠的心脏组织氧化损伤相关因子分析注：A，RT-PCR检测氧化应激因子超氧化物歧化酶（Sod）2、Sod3、谷胱甘肽过氧化物酶（Gpx）1和过氧化氢酶（Cat）的mRNA表达水平；B~D，ELISA法检测SOD、GPX和 CAT活性； E，ELISA法检测8-羟基-2'-脱氧鸟苷（8-OHdG）含量表达；F，ELISA法检测丙二醛（MDA）水平。以非早衰的野生型ICR小鼠（WT组）作为对照，n=6，与WT组比较，*P <0.05。

Figure 4 Analysis of oxidative damage-related factors in cardiac tissue of 24-week-old SHJH hr miceNote： A， RT-PCR was used to measure the mRNA expression levels ofoxidative stress factors superoxide dismutase（Sod）2， Sod3， glutathione peroxidase（Gpx）1， and catalase（Cat）； B to D， ELISA was used to determine the activities of SOD， GPX， and CAT； E， ELISA was used to measure the expression level of 8-hydroxy-2'-deoxyguanosine （8-OHdG）； F， ELISA was used to assess the level of malondialdehyde （MDA）. Wild-type ICR mice without premature aging （WT group） were used as controls， n=6. Compared with the WT group， *P <0.05.

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SHJH ^hr 小鼠的心脏衰老表型研究

Study on Cardiac Aging Phenotypes of SHJH ^hr Mice

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SHJH ^hr 小鼠的心脏衰老表型研究

Study on Cardiac Aging Phenotypes of SHJH ^hr Mice