The simultaneous targeting of RXFP1 and RXFP3 improves motor function and reduces brain inflammation, apoptosis and glial scarring following photothrombotic stroke in male mice

Inflammation and apoptosis are two major contributors to post-stroke brain injury, but can be targeted by the H2 relaxin hormone, via its GPCR, Relaxin Family Peptide Receptor 1 (RXFP1), to provide neuroprotection. The current study determined whether the therapeutic targeting of the related GPCR, RXFP3, which is widely expressed in the brain, alone or in combination with RXFP1, provided any additional neuroprotection post-stroke. Male C57Bl/6 mice underwent sham or photothrombotic stroke, targeting the primary motor (M1) cortex. Stroke-injured mice were intravenously-treated with either vehicle, the selective RXFP3 agonists, A2 (0.2 mg/kg) or H3B10 (0.2 mg/kg), or H3B10 (0.2 mg/kg) combined with H2 relaxin (0.5 mg/kg) at 6, 24 and 48 h post-stroke. Motor function was assessed via the hanging wire test at 24 and 72 h post-stroke. Infarct volume was determined via thionin staining, and immunofluorescence was performed to assess measures of inflammation, apoptosis and reactive astrocytes (which can produce glial scars) at 72 h post-stroke. The targeting of RXFP3 alone led to reduced neuronal apoptosis and T cell infiltration, but an increased ratio of anti-inflammatory M2-like macrophages compared with vehicle-treated stroke-injured mice. However, this did not influence motor function or infarct size post-stroke. On the other hand, the simultaneous targeting of RXFP3 and RXFP1 significantly reduced or attenuated the stroke-induced motor function impairment, brain inflammation, apoptosis and glial scarring in the absence of any effects on infarct size. Hence, these findings suggested that the simultaneous targeting of RXFP1 and RXFP3 provided broader neuroprotection over that of RXFP3 alone against ischemic stroke.