Overwork damages myocardial energy metabolism homeostasis in mice

doi:10.12122/j.issn.1673-4254.2025.12.07

Abstract

Abstract:

Objective To investigate the effect of overwork on myocardial energy metabolism in mice. Methods Thirty-two C57BL/6J mice were randomized equally into a control group and 3 overwork groups with overwork for 2, 4, and 6 weeks (W2, W4, and W6 groups, respectively). The mice in overwork groups were subjected to daily forced water standing and restraint. The changes in body weight and general condition of the mice were observed weekly. After successful modeling, the mice were examined for changes in echocardiography, blood glucose/lipid profiles, myocardial pathologies, myocardial TG and ATP levels, and expressions in CD36, GLUT1, CPT1B, PPARα, PFKM, and PKM2 using immunohistochemistry, RT-qPCR or Western blotting. Results The mice with prolonged overwork exhibited reduced activity with hair loss, dull fur, and slowed body weight gain without significant changes in cardiac index or function. Blood glucose levels increased significantly in W2 and W4 groups but decreased in W6 group. Serum TG level increased significantly while TC, HDL, and LDL decreased in W4 and W6 groups. HE staining revealed myocardial swelling, disorganization, and vacuolation in the mouse models. Myocardial TG was elevated in W4 and W6 groups and ATP level decreased in W6 group. The mRNA and protein expressions of CPT1B and PPARα were downregulated in W4 and W6 group, and CD36 expression increased significantly in W4 group. GLUT1 and PFKM/PKM2 expressions decreased obviously in W2 group but increased in W4 and W6 group compared with that in W2 group. Conclusion Short-term overwork causes elevation of blood glucose and suppresses glycolysis in mice, while prolonged overwork reduces glucose, increases TG, impairs fatty acid oxidation, and limits glycolytic compensation to eventually result in myocardial damage, lipid accumulation, and ATP deficiency.

Key words: overwork, myocardium, energy metabolism, fatty acid oxidation, glycolysis

Junjie CUI, Ruiyin LAI, Suheng CHEN, Shanshan QU, Yue LIAO, Xue MA, Yulan LI. Overwork damages myocardial energy metabolism homeostasis in mice[J]. Journal of Southern Medical University, 2025, 45(12): 2598-2606.

Figures/Tables 9

Tab.1 Primer sequence for RT-qPCR

Gene	Sequence of primers
CPT1B	F: GCACACCAGGCAGTAGCTTT
CPT1B	R: CAGGAGTTGATTCCAGACAGGTA
PPARα	F: AGAGCCCCATCTGTCCTCTC
PPARα	R: ACTGGTAGTCTGCAAAACCAAA
PFKM	F: TGTGGTCCGAGTTGGTATCTT
PFKM	R: GCACTTCCAATCACTGTGCC
PKM2	F: GCCGCCTGGACATTGACTC
PKM2	R: CCATGAGAGAAATTCAGCCGAG
ACTB	F: GGCTGTATTCCCCTCCATCG
ACTB	R: CCAGTTGGTAACAATGCCATGT

Tab.2 Changes of body weight of the mice in each group during modeling (g, Mean±SD, n=8)

Group	Modeling time (weeks)
Group	0	1	2	3	4	5	6
CON	20.45±0.29	21.13±0.23	22.02±0.31^a	22.54±0.47^a	23.18±0.47^a	23.96±0.37^a	24.3±0.38^a
W2	20.00±0.37	19.94±0.50	20.13±0.40^d
W4	20.50±0.19	20.24±0.24	20.41±0.19^d	20.90±0.24^b	21.14±0.30^c
W6	19.89±0.19	20.51±0.20	20.85±0.25^c	20.35±0.48^c	20.80±0.49^d	21.29±0.52^d	21.54±0.45^d

Fig.1 Heart weight index (A) and echocardiography (B) in each group (mg/g, Mean±SD, n=8). #P>0.05 vs CON group. CON: Control group. W2: overwork for 2 weeks group; W4: overwork for 4 weeks group; W6: overwork for 6 weeks group.

Tab.3 Cardiac function changes in each group (Mean±SD, n=3)

Group	LVIDs (mm)	LVIDd (mm)	EF%	FS%	LVPWd (mm)
CON	2.06±0.52	3.03±0.02	65.29±0.32	35.01±0.50	0.84±0.03
W2	2.01±0.33	3.06±0.03	65.76±0.44	35.14±0.79	0.84±0.01
W4	2.10±0.09	3.11±0.06	64.99±0.79	36.26±0.21	0.85±0.05
W6	2.11±0.06	3.18±0.01	64.87±0.55	35.28±0.43	0.85±0.01

Fig.2 Changes of blood glucose and blood lipids in each group (Mean±SD, n=6). A-E: Blood glucose (Glu), triglycerides (TG), cholesterol (TC), high-density lipoprotein cholesterol (HDL), and low-density lipoprotein cholesterol (LDL) levels, respectively. **P<0.01, ***P<0.001 vs CON group; ##P<0.01 vs W2 group; &&&P<0.001 vs W4 group.

Fig.3 Histopathological changes of the myocardial tissues and changes in myocardial TG and ATP levels in each group. A: HE staining (Scale bar=50 μm). B: Myocardial TG level (n=6; ***P<0.001 vs CON group). C: Myocardial ATP level (n=6; *P<0.05 vs CON group).

Fig.4 Relative expression levels of CPT1B (A), PPARα (B), PFKM (C), and PKM2 (D) mRNA in each group (n=3). *P<0.05, **P<0.01, ***P<0.001 vs CON group; #P<0.05, ###P<0.001 vs W2 group.

Fig.5 Expression of CD36 and GLUT1 protein and the area of positive cells (%) in the myocardium in each group. A, B: Immunohistochemical staining (Scale bar=100 μm). C: CD36 and GLUT1 protein expression levels. *P<0.05 vs CON group.

Fig.6 Relative protein expressions of CPT1B, PPARα, PFKM and PKM2 in the myocardium in each group. A, B: Protein bands in Western blotting. C, D: CPT1B, PPARα, PFKM, and PKM2 protein expression levels in each group. *P<0.05, **P<0.01, ***P<0.001 vs CON group; ##P<0.01, ###P<0.001 vs W2 group.

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