Abstract
Ischemic stroke has been reported to cause significant changes to memory, thinking, and behavior. Intriguingly, recently reported studies have indicated the association of Trimethylamine N-oxide (TMAO) with the acute phase of ischemic stroke. However, the comprehensive underlying mechanism remained unknown. The objective of the present study was to investigate the association between TMAO and recovery of neurological function after ischemic stroke. For this purpose, a middle cerebral artery occlusion/reperfusion (MCAO/R) rat model was established and treated with TMAO or/and sh-ALK5, followed by the neurological function evaluation. Behaviors of rats were observed through staircase and cylinder tests. Moreover, the expression of Smurf2 and ALK5 was detected by immunohistochemistry while expression of GFAP, Neurocan, and Phosphacan in brain tissues was determined by immunofluorescence. Thereafter, gain- and loss-of-function assays in astrocytes, the proliferation, viability, and migration were evaluated by the EdU, CCK-8, and Transwell assays. Besides, Smurf2 mRNA expression was determined by the RT-qPCR, whereas, Smurf2, ALK5, GFAP, Neurocan, and Phosphacan expression was evaluated by the Western blotting. Finally, the interaction of Smurf2 with ALK5 and ALK5 ubiquitination was assessed by the co-immunoprecipitation. Notably, our results showed that TMAO promoted the proliferation of reactive astrocyte and formation of glial scar in MCAO/R rats. However, this effect was abolished by the Smurf2 overexpression or ALK5 silencing. We further found that TMAO upregulated the ALK5 expression by inhibiting the ubiquitination role of Smurf2. Overexpression of ALK5 reversed the inhibitory effect of Smurf2 on astrocyte proliferation, migration, and viability. Collectively, our work identifies the evolutionarily TMAO/Smurf2/ALK5 signaling as a major genetic factor in the control of reactive astrocyte proliferation and glial scar formation in ischemic stroke, thus laying a theoretical foundation for the identification of ischemic stroke.
Highlights
Stroke is considered a leading cause of disability and death across the globe whilst ischemic stroke is a refractory brain injury resulting in substantial loss of function (Roy-O’Reilly and McCullough, 2018; Ozaki et al, 2019)
Compared with middle cerebral artery occlusion/reperfusion (MCAO/R) rats, the neurological function score of MCAO/R rats treated with Trimethylamine N-oxide (TMAO) was distinctly reduced (Figure 1A)
Similar results were observed in the MCAO/R rats treated with TMAO when compared to MCAO/R rats (Figures 1B,C)
Summary
Stroke is considered a leading cause of disability and death across the globe whilst ischemic stroke is a refractory brain injury resulting in substantial loss of function (Roy-O’Reilly and McCullough, 2018; Ozaki et al, 2019). Stroke can be induced by various modifiable and non-modifiable risk factors, including age, gender, hypertension, diet, and inflammatory disorders (Boehme et al, 2017). The onset and clinical outcomes of ischemic stroke have been reported to be closely correlated with age; the effects of currently available treatment modalities are limited in elderly patients (Chen et al, 2010). For young adults aged between 18 and 50 years, optimal management has been indicated to be distinctive from elder patients, which requires further exploration. Effective therapeutic strategies are prerequisite for the improvement of prognosis
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