Activation of GPR40 attenuates neuroinflammation and improves neurological function via PAK4/CREB/KDM6B pathway in an experimental GMH rat model

Abstract

Background Germinal matrix hemorrhage (GMH) is defined by the rupture of immature blood vessels in the germinal matrix, where subsequent hemorrhage enters the subependymal zone and the cerebral lateral ventricles. The consequent blood clot has been identified as the causative factor of secondary brain injury, which triggers a series of complex parallel and sequential harmful mechanisms, including neuroinflammation. The orphan G-protein-coupled receptor 40 (GPR40), a free fatty acid (FFA) receptor 1, has been shown to exert anti-inflammatory effects when activated and improved outcomes in animal models of stroke. We aimed to investigate the anti-inflammatory effects of GPR40 and its underlying mechanisms after GMH. Methods GMH model was induced in 7-day-old rat pups by an intraparenchymal injection of bacterial collagenase. GPR40 agonist, GW9508, was administered intranasally 1 h, 25 h, and 49 h after GMH induction. CRISPR targeting GPR40, PAK4, and KDM6B were administered through intracerebroventricular injection 48 h before GMH induction. Neurologic scores, microglia polarization, and brain morphology were evaluated by negative geotaxis, right reflex, rotarod test, foot fault test, Morris water maze, immunofluorescence staining, Western blots, and nissl staining respectfully. Results The results demonstrated that GW9508 improved neurological and morphological outcomes after GMH in the short (24 h, 48 h, 72h) and long-term (days 21–27). However, the neuroprotective effects of treatment were abolished by GW1100, a selective GPR40 antagonist. GW9508 treatment increased populations of M2 microglia and decreased M1 microglia in periventricular areas 24 h after GMH induction. GW9508 upregulated the phosphorylation of PAK4, CREB, and protein level of KDM6B, CD206, IL-10, which was also met with the downregulation of inflammatory markers IL-1β and TNF-α. The mechanism study demonstrated that the knockdown of GPR40, PAK4, and KDM6B reversed the neuroprotective effects brought on by GW9508. This evidence suggests that GPR40/PAK4/CREB/KDM6B signaling pathway in microglia plays a role in the attenuation of neuroinflammation after GMH. Conclusions In conclusion, the present study demonstrates that the activation of GPR40 attenuated GMH-induced neuroinflammation through the activation of the PAK4/CREB/KDM6B signaling pathway, and M2 microglia may be a major mediator of this effect. Thus, GPR40 may serve as a potential target in the reduction of the inflammatory response following GMH, thereby improving neurological outcomes in the short- and long-term.

Candesartan Reduces Neuronal Apoptosis Caused by Ischemic Stroke via Regulating the FFAR1/ITGA4 Pathway

“…GPR40, as the first identified G protein-coupled receptor, delivered nonenzyme-generated nitrooxidation products, particularly TAAs, and was involved in HI encephalopathy [ 7 ]. The activation of GPR40 attenuated the neuroinflammation induced by GMH through activating the PAK4/CREB/KDM6B signaling pathway [ 15 ]. DHA significantly induced growth inhibition and apoptosis of human androgen-dependent prostate cancer cells via inactivating YAP by promoting phosphorylation and cytoplasmic translocation pathways (FFAR1/FFAR4-Gas-PKA-Hippo) [ 16 ].…”

Electroacupuncture of Baihui and Shenting ameliorates cognitive deficits via Pten/Akt pathway in a rat cerebral ischemia injury model

“…For instance, Won et al demonstrated that Pak4 could protect dopaminergic neurons and preserve motor function in Parkinson's disease rat models (33). PAK4 loss abrogated the neuroprotective activity induced by the Gprotein-coupled receptor 40 agonist GW9508 (34). Aberrant Akt3 expression is closely linked with brain and cortical development and neurogenesis (35)(36)(37).…”

Adiponectin Ameliorates GMH-Induced Brain Injury by Regulating Microglia M1/M2 Polarization Via AdipoR1/APPL1/AMPK/PPARγ Signaling Pathway in Neonatal Rats

“…Stimulation of cannabinoid receptor 2 (CB2R) suppressed neuroinflammation via regulation of M1/M2 polarization via cAMP/PKA signaling pathway in an experimental rat model of GMH (26). Recently, we reported that GW9508-mediated GPR40 activation ameliorates neuroinflammation and improves neurological function through PAK4/CREB/KDM6B signaling pathway after GMH (20). Our data from this study showed that intranasal administration of rh-APN promoted M2 microglia polarization with increased anti-inflammatory cytokine (IL-10 and Arginase-1) release.…”

“…GMH induction was conducted in unsexed P7 rats and was accomplished via collagenase infusion as previously described (19,20). Briefly, rat pups weighting between 12g-15g were anesthetized using 3% isoflurane, and anesthesia maintenance was conducted using 1.5% isoflurane intraoperatively.…”

“…Double Immunofluorescence staining was conducted on the sections of fixed frozen brains as previously described (19,20). The 8-mm thick slides were rinsed using phosphate-buffered saline (PBS), followed by permeabilization using 0.3% Triton X-100 for 15 min at RT.…”