Cathepsin H deficiency decreases hypoxia-ischemia-induced hippocampal atrophy in neonatal mice through attenuated TLR3/IFN-β signaling

Abstract

Background Cathepsin H (CatH) is a lysosomal cysteine protease with a unique aminopeptidase activity. Its expression level is increased in activated immune cells including dendritic cells, macrophages, and microglia. We have previously reported that CatH deficiency impairs toll-like receptor 3 (TLR3)-mediated activation of interferon regulatory factor 3 (IRF3), and the subsequent secretion of interferon (IFN)-β from dendritic cells. Furthermore, there is increasing evidence that IFN-β secreted from microglia/macrophages has neuroprotective effects. These observations prompted further investigation into the effects of CatH deficiency on neuropathological changes. Methods In this study, neuropathological changes were examined using histochemical staining (both hematoxylin-eosin (H&E) and Nissl) of the hippocampus of wild-type (WT) and CatH-deficient (CatH−/−) mice after hypoxia-ischemia (HI). The density and the localization of CatH and TLR3 were examined by immunofluorescent staining. CatH processing in microglia was assayed by pulse-chase experiments, while immunoblotting was used to examine TLR3 expression and IRF3 activation in microglia/macrophages in the presence of poly(I:C). Microglial cell death was examined by fluorescence-activated cell sorting (FACS), and primary astrocyte proliferation in the presence of IFN-β was examined using scratch wound assay. Results WT mice displayed severe atrophy in association with neuronal death and moderate astrogliosis in the hippocampus following neonatal HI. Somewhat surprisingly, CatH−/− mice showed marked neuronal death without severe atrophy in the hippocampus following HI. Furthermore, there was notable microglia/macrophages cell death and strong astrogliosis in the hippocampus. The TLR3 and phosphorylated IRF3 expression level in the hippocampus or splenocytes (mainly splenic macrophages); from CatH−/− mice was lower than in WT mice. In vitro experiments demonstrated that recombinant IFN-β suppressed HI-induced microglial cell death and astrocyte proliferation. Conclusion These observations suggest that CatH plays a critical role in the proteolytic maturation and stabilization of TLR3, which is necessary for IFN-β production. Therefore, impaired TLR3/IFN-β signaling resulting from CatH deficiency may induce microglial cell death after activation and astrogliosis/glial scar formation in the hippocampus following HI injury, leading to suppression of hippocampal atrophy.

Amanita muscaria extract potentiates production of proinflammatory cytokines by dsRNA-activated human microglia

“…For example, primary murine microglia can be activated with poly(I:C), a TLR3 ligand, that increases their proinflammatory response though activation of cathepsin X ( Pišlar et al, 2022 ), suggesting that this process is associated with inflammation-induced neurodegeneration. However, cathepsin H deficiency decreases hypoxia-ischemia-induced hippocampal atrophy in neonatal mice through attenuation of TLR3/IFN-β signaling, suggesting that this pathway may be neuroprotective ( Ni et al, 2021 ). As such, upregulation of TLR3 by AME-1 may participate in this pathway, although whether the balance is shifted toward a pro-inflammatory neurodegenerative or neuroprotective pathway is still unknown and requires further experimentation.…”

Zinc finger protein 335 mediates lipopolysaccharide‐induced neurodegeneration and memory loss as a transcriptional factor in microglia

“…This phenomenon could explain the specific increased expression of Zfp335 in microglia rather than in other cell types. Microglia activation has been reported to be controlled by various key transcription factors, such as NF‐κB (Meng et al, 2020; Wu et al, 2017), STAT1 (Ni et al, 2021), and PU.1 (Wu et al, 2017). Based on the regulation afforded by Zfp335 upon microglia activation, we propose that there may be interactions between Zfp335 and the transcription factors stated above because H3K4 methyltransferase has been reported to be essential for the transactivation of a subset of NF‐κB target genes (Redd et al, 2018; Yu et al, 2017).…”

Regulation of microglia polarization after cerebral ischemia

“…The proteolysis of TLR3 is essential for its function, and Cathepsin H and B play a crucial role in the regulation of this process ( Garcia-Cattaneo et al, 2012 ). In neonatal mice, the deficiency of Cathepsin H leads to prolonged neurotoxic M1-like polarization following hypoxia ischemia, which ultimately results in microglia death and subsequent neurotoxic A1 astrocyte proliferation and progressive neuronal death ( Liddelow et al, 2017 ; Ni et al, 2021 ). However, in mice injected with LPS, upregulated expression of Cathepsin H in microglia exacerbates neuronal death in neuroinflammation ( Fan et al, 2015 ).…”