Differential Sphingosine‐1‐Phosphate Receptor 1 Expression in Male and Female Murine Heart and Brain Following Acute Ischemia and Reperfusion
Authors: Shi Samuel , Mohamed Awad R. , Shi Kaibin , Jin Weina , Liu Qiang , Shi Fu‐Dong , Gonzales Rayna J. 
Published: June 2021

We and others demonstrate that nonselective sphingosine‐1 phosphate receptor (S1PR) modulators provide neuroprotection after stroke. However, non‐selective actions of S1PR modulators have been shown to induce off target effects such as bradycardia as a result of binding to multiple S1PR subtypes. Thus, identifying selective S1PR modulators with neuroprotective potential will reveal better drug development for treatment of acute ischemic stroke. S1PR, a GPCR, is expressed in a number of organ systems including the immune and cardiovascular systems. We observed in pre‐clinical studies that selective S1PR type 1 modulation can reduce parenchymal damage via attenuation of peripheral nodal egress. However, the role of S1PR1 in the brain following acute ischemic stroke remains to be investigated. Therefore, the present study investigates selective S1PR1 expression profile in the brain and if levels are altered following ischemic injury. Adult C57BL/6 black male and female mice were subjected to 60‐min transient middle cerebral artery occlusion (MCAO). Post 24h, whole heart and brain were removed and brain was divided into ipsilateral (lesion side) and contralateral hemispheres. Tissues were homogenized, and analyzed for total protein content. Immunoblotting was performed for S1PR1 receptor expression in sham vs. MCAO. S1PR1 is basally expressed in brain and heart under normal conditions. Following acute ischemic injury, S1PR1 protein expression increased in male and female brain as compared to sham controls and greatest S1PR1expression was observed on the ipsilateral side compared to the contralateral side. Conversely, in the heart, S1PR1 protein levels were decreased in MCAO injured females as compared to sham; although a trend was observed, there was no difference in S1PR1 expression in male heart. Flow cytometry (BD FACSARIA) was conducted to determine S1PR1 expression in peripheral and CNS cells after MCAO. Cell suspensions were prepared from MCAO male mouse brain; cell subtypes were isolated by fluorescent tagging then laser sorted. Quantitative S1PR1 expression was determined in purified cell sub‐types using a mouse S1PR1/EDG‐1 conjugated fluorescent antibody. We observed S1PR1 expression in endothelial and multiple brain cell subtypes following ischemic injury. RP101075 (0.6 mg/kg), a selective S1PR1 with improved cardiovascular safety profile, or vehicle was injected i.p. to recipient mice immediately after MCAO. At 24 and 72h post reperfusion, brains were imaged with MRI (7T‐Bruker) and lesion volume analyzed. RP101075 treatment decreased lesion volume compared to vehicle control at both time points. In conclusion, selective S1PR1 modulation may be a significant therapeutic target for treatment of ischemic stroke since expression increases in the brain and decreases in the heart, an organ with adverse effects following nonselective S1PR modulation. Future studies will investigate the beneficial effect of selective S1PR1 modulation in the brain on mechanisms associated with protective neurological outcomes following acute ischemic injury.

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Valley Research Partnership P2 VPR37 (RJG)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.