Recombinant milk fat globule-EGF factor-8 reduces apoptosis via integrin β3/FAK/PI3K/AKT signaling pathway in rats after traumatic brain injury

Abstract

Accumulating evidence suggests neuronal apoptosis has the potential to lead to more harmful effects in the pathological processes following traumatic brain injury (TBI). Previous studies have established that milk fat globule-EGF factor-8 (MFG-E8) provides neuroprotection through modulation of inflammation, oxidative stress, and especially apoptosis in cerebral ischemia and neurodegenerative disease. However, the effects of MFG-E8 on neuronal apoptosis in TBI have not yet been investigated. Therefore, we explored the role of MFG-E8 on anti-apoptosis and its potential mechanism following TBI. In the first set of experiments, adult male Sprague–Dawley (SD) rats were randomly divided into Sham and TBI groups that were each further divided into five groups representing different time points (6 h, 24 h, 72 h, and 7 days) (n = 9 each). Western blotting, quantitative real-time PCR, and immunofluorescence staining were performed to identify the expression and cellular localization of MFG-E8. In the second set of experiments, four groups were randomly assigned: Sham group, TBI + Vehicle group, and TBI + rhMFG-E8 (1 and 3 µg) (n = 15). Recombinant human MFGE8 (rhMFG-E8) was administrated as two concentrations through intracerebroventricular (i.c.v.) injection at 1 h after TBI induction. Brain water content, neurological severity score, western blotting, and immunofluorescence staining were measured at 24 and 72 h following TBI. In the final set of experiments, MFG-E8 siRNA (500 pmol/3 µl), integrin β3 siRNA (500 pmol/3 µl), and PI3K inhibitor LY294002 (5 and 20 µM) were injected i.c.v. and thereafter rats exposed to TBI. Western blotting, immunofluorescence staining, brain water content, neurological severity score, and Fluoro-Jade C (FJC) staining were used to investigate the effect of the integrin-β3/FAK/PI3K/AKT signaling pathway on MFG-E8-mediated anti-apoptosis after TBI. The expression of MFG-E8 was mainly located in microglial cells and increased to peak at 24 h after TBI. Treatment with rhMFG-E8 (3 µg) markedly decreased brain water content, improved neurological deficits, and reduced neuronal apoptosis at 24 and 72 h after TBI. rhMFG-E8 significantly enhanced the expression of integrin-β3/FAK/PI3K/AKT pathway-related components. Administration of integrin-β3 siRNA and LY294002 (5 and 20 µM) abolished the effect of rhMFG-E8 on anti-apoptosis and neuroprotection after TBI. This study demonstrated for the first time that rhMFG-E8 inhibits neuronal apoptosis and offers neuroprotection. This is suggested to occur through the modulation of the integrin-β3/FAK/PI3K/AKT signaling pathway, highlighting rhMFG-E8 as a potentially promising therapeutic strategy for TBI patients.

WITHDRAWN: Combination of Paeoniflorin and Calycosin-7-glucoside Promotes Ischemic Stroke Recovery via the PI3K/AKT Signaling Pathway

“…Activation of the signaling pathway can inhibit a variety of stimulationinduced apoptosis mechanisms and promote cell cycle progression, thereby promoting cell survival and proliferation while participating in angiogenesis 14 . The current study found that activation of the PI3K/AKT pathway can signi cantly reduce the area of brain damage and the rate of apoptosis 15 . Activated AKT can also protect rat hippocampal neurons and cortical neurons against hypoxia/reoxygenation-induced apoptosis [16][17][18] . In this study, we hypothesized that PF + CG may confer protection from brain damage following cerebral ischemia via the PI3K/AKT pathway.…”

Brain-Derived Microparticles (BDMPs) Contribute to Neuroinflammation and Lactadherin Reduces BDMP Induced Neuroinflammation and Improves Outcome After Stroke

“…Several studies have confirmed the therapeutic effects of Lactadherin in stroke (23,24,26,52). Lactadherin exerts neuroprotection against cerebral injury by suppressing inflammation, reducing neuronal cell death, promoting apoptotic cell clearance (26). Additionally, Lactadherin improves subarachnoid hemorrhage (SAH) outcome via anti-oxidation which may be dependent on integrin β3/ nuclear factor erythroid 2-related factor 2/HO pathway (24).…”

“…Lactadherin mediates cell-cell interactions and is involved in various physiological and pathophysiological functions including angiogenesis (49), fertilization (50), inflammation (23) and clearance of apoptotic cells (51). Several studies have confirmed the therapeutic effects of Lactadherin in stroke (23,24,26,52). Lactadherin exerts neuroprotection against cerebral injury by suppressing inflammation, reducing neuronal cell death, promoting apoptotic cell clearance (26).…”

“…Lactadherin (milk fat globule-epidermal growth factor 8, Lactadherin) is a multifunctional glycoprotein originally identified as part of the milk fat globule membrane. Lactadherin couples apoptotic cells with monocytes/macrophages to facilitate phagocytosis (20)(21)(22) and clearance of apoptotic cells, and regulates immune response after stroke (23)(24)(25)(26). In this study, we are the first to investigate that BDMPs contribute to neuroinflammation after stroke, while Lactadherin promotes the clearance of BDMPs and reduces inflammation and thereby improves ischemic stroke outcome.…”

MFG-E8 attenuates neuro-inflammation in subarachnoid hemorrhage through driving microglial M2 polarization via modulating Integrin β3/SOCS3/STAT3 signaling pathway

“…Administration of MFG-E8 could alleviate pathological lesion of Alzheimer's disease (AD) via modulating the alteration of M1/M2 polarization [14]. Recently our laboratory have confirmed that recombinant human MFG-E8 (rhMFG-E8) improve neurological function in an animal model of traumatic brain injury (TBI) by inhibiting neuronal apoptosis [17]. These studies indicated that MFG-E8 could provide neuroprotection in CNS.…”