Syringin improves myocardial mitochondrial dynamics and alleviates oxidative stress injury in septic mice by activating SIRT3

doi:10.12122/j.issn.1673-4254.2026.10.21

Abstract

Abstract:

Objective To investigate the protective effects of syringin against septic myocardial injury and explore the underlying mechanisms. Methods Thirty C57 mice were randomly assigned to receive sham operation with or without subsequent syringin treatment or sepsis modeling by cecal ligation and puncture (CLP) followed by intraperitoneal injection of saline, syringin, or both syringin and 3-TYP (an SIRT3 inhibitor). Cardiac function and myocardial injury of the mice were assessed with echocardiography and HE staining, and serum cTnT, LDH, MDA, SOD, and GSH-Px levels were detected using ELISA. ROS production and expressions of p-DRP1, MFN1, and NOX4 were detected with immunohistochemistry and immunofluorescence staining, and the expressions of the SIRT3/NOX2/p-DRP1 pathway proteins were determined with Western blotting. In primary culture of neonatal mouse cardiomyocytes (NMCMs) treated with lipopolysaccharide (LPS), the effects of syringin, syringin combined with 3-TYP, and honokiol on mitochondrial membrane potential and expression of the SIRT3/NOX2/p-DRP1 pathway were analyzed by immunofluorescence staining and Western blotting. Results CLP mice exhibited significant impairment of cardiac function, obvious myocardial injury, elevated serum cTnT and LDH levels, and increased expression of oxidative stress- and mitochondrial injury-related proteins. Syringin treatment significantly improved myocardial structure and cardiac function, attenuated oxidative stress, and reduced mitochondrial damage in the CLP mice. Treatment with 3-TYP significantly abolished the protective effects of syringin and resulted in increased MDA and cTnT levels, decreased SOD activity, elevated ROS production, and increased expressions of oxidative stress and mitochondrial injury markers. In NMCMs, LPS stimulation obviously reduced mitochondrial membrane potential, increased AC-SOD2, NOX2, and p-DRP1 expressions, and decreased MFN1 levels. Syringin significantly alleviated LPS-induced oxidative stress and mitochondrial dysfunction, and these effects were abolished by 3-TYP treatment or SIRT3 silencing. Conclusion Syringin attenuates sepsis-induced myocardial injury by activating SIRT3 to improve cardiac function, suppress oxidative stress, and alleviate mitochondrial damage.

Key words: syringin, sepsis, mitochondria, oxidative stress, SIRT3

Yan LIU, Hongqi ZHANG. Syringin improves myocardial mitochondrial dynamics and alleviates oxidative stress injury in septic mice by activating SIRT3[J]. Journal of Southern Medical University, 2026, 46(10): 2536-2547.

Figures/Tables 8

Tab.1 Primer sequences for RT-qPCR

Gene	Forward primer	Reverse primer
NOX2	TGCCAACTTCCTCAGCTACAATA	GTGTTCACTTGCAATGGTCTTGA
NRF2	TGGCAGAGACATTCCCATTTGTA	CTTGCTCCATGTCCTGCTCTATG
NOX4	CCTCCTGGCTGCATTAGTCTTAA	AAAGGCACAAAGGTCCAGAAATC
GAPDH	GGCATTGTGGAAGGGCTCAT	AGATCCACGACGGACACATT

Fig.1 Syringin (SYR) alleviates myocardial injury in mice with sepsis. A: Representative left ventricular M-mode echocardiograms of the mice in different groups. B: Statistical analysis of heart rate (n=6). C: Statistical analysis of LVEF (n=6). D: Statistical analysis of LVFS (n=6). E: Serum cTnt levels in the mice. F: LDH activity in mouse serum. G: Survival rates of the mice with or without SYR therapy 24 h after cecal ligation and puncture (CLP) (n=20). Time (h) refers to the survival hours after sepsis. H: HE staining (Scale bar=100 μm). *P<0.05, **P<0.01, ****P<0.0001.

Fig.2 SYR treatment alleviates sepsis-induced aberrant expressions of oxidative stress-related genes and proteins in mice. A: NOX2, NRF2 and NOX4 mRNA levels in mouse myocardial tissue. B: Target proteins examined by Western blotting. C: Statistical analysis of NRF2/GAPDH (n=6). D: Statistical analysis of SIRT3/GAPDH (n=6). E: Statistical analysis of SOD2/GAPDH (n=6). F: Statistical analysis of AC-SOD2/GAPDH (n=6). G: Statistical analysis of NQO1/GAPDH (n=6). H: Statistical analysis of NOX2/GAPDH (n=6). Comparisons among multiple groups were made using one-way ANOVA. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

Fig.3 SYR treatment reduces the levels of oxidative stress in the serum and tissues of septic mice. A: MDA content in mouse serum (n=6). B: GSH-Px content in mouse serum (n=6). C: SOD content in mouse serum (n=6). D: Expression and location of NOX4 determined by immunohistochemistry (Scale bar=100 μm). E: Statistical analysis of NOX4 area (n=6). F: Expression and location of 8-OHdG determined by immunofluorescence staining (Scale bar=100 μm). G: Statistical analysis of fluorescence intensity (n=6). *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

Fig.4 SYR treatment alleviates mitochondrial damage in septic mice. A: Expression and location of MFN1 determined by immunofluorescence staining (Scale bar=100 μm). B: Statistical analysis of fluorescence intensity (n=6). C: Expression and location of p-DRP1 were determined by immunohistochemistry (Scale bar=100 μm). D: Statistical analysis of p-DRP1 area (n=6). E: Target proteins examined by Western blotting. F: Statistical analysis of p-DRP1(S616)/DRP1 (n=6). G: Statistical analysis of MFN1/GAPDH (n=6). *P<0.05, ***P<0.001, ****P<0.0001.

Fig.5 SYR attenuates oxidative stress and mitochondrial damage in primary culture of neonatal mouse cardiomyocytes (NMCMs) by activating SIRT3. A: Expression of DCFH-DA determined by immunofluorescence staining (Scale bar=200 μm). B: Statistical analysis of fluorescence intensity (n=6). C: JC-1 staining for measuring the mitochondrial membrane potential in NMCMs showing depolarization (green) and hyperpolarization (red) of the mitochondria. D: Statistical analysis of fluorescence intensity (n=6). E: Target proteins examined by Western blotting. F: Statistical analysis of SIRT3/GAPDH (n=6). G: Statistical analysis of p-DRP1/DRP1 (n=6). H: Statistical analysis of MFN1/GAPDH (n=6). I: Statistical analysis of AC-SOD2/GAPDH (n=6). J: Statistical analysis of NOX2/GAPDH (n=6). *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

Fig.6 Inhibiting SIRT3 eliminates the inhibitory effects of SYR on oxidative stress in neonatal mouse cardiomyocytes and its protective effect on mitochondria. A: Expression of DCFH-DA determined by immunofluorescence staining (Scale bar=200 μm). B: Statistical analysis of fluorescence intensity (n=6). C: Target proteins examined by Western blotting. D: Statistical analysis of SIRT3/GAPDH (n=6). E: Statistical analysis of p-DRP1/DRP1 (n=6). F: Statistical analysis of MFN1/GAPDH (n=6). G: Statistical analysis of AC-SOD2/GAPDH (n=6). H: Statistical analysis of NOX2/GAPDH (n=6). ***P<0.001, ****P<0.0001.

Fig.7 SYR produces protective effect on the myocardium in septic mice by activating SIRT3. A: Representative electron microscopy images showing damaged mitochondrial with loss of cristae in mouse cardiomyocytes (Scale bar=500 nm). B: Quantification of damaged mitochondria in mouse cardiomyocytes (n=6). C: MDA content in mouse serum (n=6). D: cTnt content in mouse (n=6). E: SOD content in mouse serum (n=6). F: Target proteins examined by Western blotting. G: Statistical analysis of SIRT3/GAPDH (n=6). H: Statistical analysis of SOD2/GAPDH (n=6). I: Statistical analysis of p-DRP1/DRP1 (n=6). J: Statistical analysis of NOX2/GAPDH (n=6). K: Statistical analysis of NQO1/GAPDH (n=6). *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

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