Hypoxia-preconditioned mesenchymal stem cells attenuate microglial pyroptosis after intracerebral hemorrhage

Abstract

Background: Microglia plays a vital role in neuroinflammation, contributing to the pathogenesis of intracerebral hemorrhage (ICH)-induced brain injury. Mesenchymal stem cells (MSCs) hold great potential for treating ICH. We previously revealed that MSCs ameliorate the microglial pyroptosis caused by an ischemic stroke. However, whether MSCs can modulate microglial pyroptosis after ICH remains unknown.This study aimed to investigate the neuroprotective effects of hypoxia-preconditioned olfactory mucosa MSCs (OM-MSCs) on ICH and the possible mechanisms.Methods: ICH was induced in mice via administration of collagenase IV. At 6 h post-ICH, 2-4×10 5 normoxic/hypoxic OM-MSCs or saline were intracerebrally administered. To evaluate the neuroprotective effects, the behavioral outcome, apoptosis, and neuronal injury were measured. Microglia activation and proinflammatory cytokines were applied to detect neuroinflammation. Microglial pyroptosis was determined by western blotting, immunofluorescence staining, and transmission electron microscopy (TEM). Results:The two OM-MSC-transplanted groups exhibited significantly improved functional recovery and reduced neuronal injury, especially the hypoxic OM-MSCs group. Hypoxic OM-MSCs attenuated microglial activation as well as the levels of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α). Moreover, we found that hypoxia-preconditioned OM-MSCs ameliorated pyroptosis by diminishing the levels of pyroptosis-associated proteins in peri-hematoma brain tissues, decreasing the expression of the microglial nod-like receptor family protein 3 (NLRP3) and caspase-1, and reducing the membrane pores on microglia post-ICH.Conclusions: Our study showed that hypoxic preconditioning augments the therapeutic efficacy of OM-MSCs, and hypoxia-preconditioned OM-MSCs alleviate microglial pyroptosis in the ICH model.

Modes of Brain Cell Death Following Intracerebral Hemorrhage

“…NLRP3and caspase-1-positive cells were microglia, as determined through double immunofluorescence labeling. The authors also observed bubbles and large holes in the plasma membrane of microglia from the ICH group, in contrast to the linear and intact membrane from the sham-operated group, using a transmission electron microscope (Liu J. et al, 2021). Similar results were reported by Lin et al (2018), who demonstrated that NLRP3, caspase-1, and mature IL-1β expression were increased 24 h after ICH.…”

“…NLR pyrin domain containing 3 (NLRP3) and NLRP1 inflammasome activate caspase-1 to initiate the cleavage and activation of interleukin-1β (IL-1β) and IL-18 leading to neuroinflammation and cell death after ICH (Bobinger et al, 2018; Figure 4). Liu J. et al (2021) found that the levels of NLRP3, cleaved caspase-1, cleaved caspase-8, and IL-1β were significantly increased 3 days post-ICH in mice. NLRP3and caspase-1-positive cells were microglia, as determined through double immunofluorescence labeling.…”

“…Nucleotide-binding oligomerization domain-like receptor pyrin domain-containing 1-dependent neuronal pyroptosis can be decreased by CCR5 antagonist maraviroc and melanocortin receptor 4 agonist RO27-3225; this is reported to be through CCR5/PKA/CREB pathway and apoptosis signalling-regulating kinase 1 (ASK1)/jun N-terminal kinase (JNK)/p38 mitogenactivated protein kinase (MAPK) pathways after ICH in mice, respectively (Chen S. et al, 2019;Yan et al, 2021). Microglia pyroptosis induced by ICH was suppressed by hypoxiapreconditioned olfactory mucosa mesenchymal stem cells or pharmacological treatment with andrographolide, by inhibiting the NLRP3 inflammasome and caspase-1 activity (Li et al, 2018d;Liu J. et al, 2021). AC-YVAD-CMK, a selective inhibitor of caspase-1, reduced caspase-1 activation and inhibited IL-1 production and maturation, but has no effect on NLRP3 expression.…”

Mesenchymal Stem Cell Application and Its Therapeutic Mechanisms in Intracerebral Hemorrhage

“…Recent studies have confirmed that MSCs are effective modifiers maintaining a resting microglial phenotype, preventing microglial activation by downregulating pro-inflammatory cytokines/chemokines and upregulating hypoxia-inducible factor 1-alpha (HIF-1α) and growth factors including VEGF, BDNF, GDNF, stromal-derived factor-1 (SDF-1), and erythropoietin (EPO) following stroke ( Wei et al, 2012 ; Yan et al, 2013 ). Other studies have shown that hypoxic preconditioning augmented MSC survival and tissue-protective capability, displaying better therapeutic efficacy than the single MSC transplantation by improving the miR-326/PTBP1/PI3K-mediated autophagy and alleviated microglial activation to downregulate IL-1β and TNF-α expression levels and microglial pyroptosis after ICH ( Liu et al, 2021a , b ). Human ADMSCs improved the neurological deficits in the ICH mice model by suppressing the acute inflammation mediated by the CD11 + CD45 + subpopulation of cells ( Kuramoto et al, 2019 ).…”

Mesenchymal Stromal Cells Preconditioning: A New Strategy to Improve Neuroprotective Properties

“…Probably, hypoxic preconditioning led to the reduction in microglial pyroptosis through a mechanism mediated by NLRP3 inflammasome to promote the passage of microglia into the anti-inflammatory phenotype. Moreover, mice transplanted with preconditioned OM-MSCs showed an improvement in behavioral deficits [69].…”