Selective cerebral hypothermia alleviates focal cerebral ischemia/reperfusion injury via enhancing SUMO2/3 modification of Drp1 in rats

Background

Selective Cerebral Hypothermia (SCH) has been demonstrated to potentiate SUMO2/3 modification, a native cellular safeguard against Cerebral Ischemia/Reperfusion Injury (CIRI). Dynamin-Related Protein 1 (Drp1), a pivotal regulator in the mitochondrial fission pathway, is an important substrate for SUMO2/3 modification. However, effects of SCH on SUMO2/3 modification of Drp1 remain undefined. Herein, the current study posits that SCH augments the SUMO2/3 modification of Drp1, thereby preserving mitochondrial integrity and mitigating CIRI.

Methods

A focal CIRI model was established in Sprague-Dawley rats, with 20°C saline perfused via the transcarotid artery to induce SCH condition, and 37°C saline serving as a control. The modified Neurological Severity Score (mNSS) was used to quantitate the degree of neurological deficits. Staining of 2,3–5-triphenyltetrazolium chloride (TTC) was performed to detect cerebral infarction volume. Histological change of neurocyte was observed through Hematoxylin-eosin (HE) staining. Neurocyte apoptosis was evaluated using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) immunofluorescence staining. Western blot (WB) was utilized to evaluated the expressions of Drp1 and Cytochrome C. Co-immunoprecipitation was performed to evaluate the level of SUMO2/3 modification of Drp1. And transmission electron microscopy was used to observe the mitochondrial ultrastructure. The ratio of M-Drp1 to T-Drp1 and mitochondria morphological changes were observed under confocal microscopy.

Results

Research data revealed that SCH significantly enhanced the SUMO2/3 modification of Drp1 when CIRI occurred. Concurrently, mNSSs, cerebral infarct volume, and apoptotic rates were notably attenuated in the SCH group, corroborating SCH's protective role. Expression levels of mitochondrial outer membrane Drp1 (M-Drp1), cytoplasmic cytochrome C (C-CytC), and ratio of M-Drp1 to T-Drp1 were reduced, and changes of mitochondrial ultrastructural and morphology were mitigated, underscoring SCH's inhibitory effect on mitochondrial fission. In contrast, 37°C saline displayed negligible protective impact while compare with 20°C saline perfusion.

Conclusions

The findings support that SCH amplifies SUMO2/3 modification of Drp1, curtails excessive mitochondrial fission, and consequently ameliorates focal CIRI in a rat model.