Synergistic effect of sildenafil combined with controlled hypothermia to alleviate microglial activation after neonatal hypoxia–ischemia in rats

SUMMARY

The only validated treatment to prevent brain damage associated with hypoxia-ischemia (HI) encephalopathy of the newborn is controlled hypothermia with limited benefits. Additional putative neuroprotective drug candidates include sildenafil citrate, a phosphodiesterase-type 5 inhibitor. The main objective of this preclinical study, led by GCIR Professor Olivier Baud, was to assess the ability of Sildenafil to reduce HI-induced neuroinflammation, in particular through its potential effect on microglial activation.

HI was induced in P10 Sprague-Dawley rats by unilateral carotid permanent artery occlusion and hypoxia (HI) and treated by either hypothermia (HT) alone, Sildenafil (Sild) alone or combined treatment (SildHT). Lesion size and glial activation were analyzed by immunohistochemistry, qRT-PCR, and proteomic analyses performed at P13.

None of the treatments was associated with a significant early reduction in lesion size 72h after HI, despite significant changes in tissue loss distribution. Significant reductions in both Iba1 + (within the ipsilateral hemisphere) and GFAP + cells (within the ipsilateral hippocampus) were observed in SildHT group, but not in the other treatment groups. In microglia-sorted cells, pro-inflammatory markers, i.e. Il1b, Il6, Nos2, and CD86 were significantly downregulated in SildHT treatment group only. These changes were restricted to the ipsilateral hemisphere, were not evidenced in sorted astrocytes, and were not sex-dependent. Proteomic analyses in sorted microglia refined the pro-inflammatory effect of HI and confirmed a biologically relevant impact of SildHT on specific molecular pathways including genes related to neutrophilic functions.

These findings suggest that Sildenafil combined with controlled hypothermia produces maximum effect in mitigating microglial activation induced by HI through complex proteomic regulation. The reduction of neuroinflammation induced by Sildenafil may represent an interesting therapeutic strategy for neonatal neuroprotection.

Full article: https://doi.org/10.1186/s12974-024-03022-w

Why is it important?

Neonatal hypoxic-ischaemic encephalopathy (HI) is a leading cause of death or disability in newborns, affecting between 1 and 4 out of every 1,000 live births. HI encephalopathy is more common and severe in disadvantaged areas, affecting approximately 1 million newborns per year worldwide. Controlled hypothermia (HT) is the only proven treatment to prevent brain damage in newborns with hypoxia-ischaemia (HI) encephalopathy, but its benefits are limited.

For this reason, recent studies have evaluated the efficacy of different strategies in combination with HT, including Sildenafil, a vasodilator commonly used to treat persistent pulmonary hypertension in neonates. Sildenafil is increasingly recognised as conferring multiple neuroprotective effects, but the mechanism of neuroprotection is unknown.

The authors investigated whether Sildenafil could decrease neuroinflammation caused by HI by modulating microglial activation (the activation of resident immune cells of the central nervous system - CNS). To test this hypothesis, they induced hypoxic ischaemia in rats and then treated them with hypothermia alone, Sildenafil alone or a combination of both. The researchers then examined lesion size and microglial activation using immunohistochemistry, qRT-PCR and proteomic analysis. They observed that the combination of Sildenafil and controlled hypothermia has a maximal effect on reducing microglial activation caused by HI by regulating specific molecular pathways of the inflammatory response.

The potential of Sildenafil to decrease neuroinflammation suggests that it could be a compelling therapeutic option for safeguarding neonatal brains.

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