Neuroinflammatory processes are augmented in mice overexpressing human heat-shock protein B1 following ethanol-induced brain injury

Abstract

Background Heat-shock protein B1 (HSPB1) is among the most well-known and versatile member of the evolutionarily conserved family of small heat-shock proteins. It has been implicated to serve a neuroprotective role against various neurological disorders via its modulatory activity on inflammation, yet its exact role in neuroinflammation is poorly understood. In order to shed light on the exact mechanism of inflammation modulation by HSPB1, we investigated the effect of HSPB1 on neuroinflammatory processes in an in vivo and in vitro model of acute brain injury. Methods In this study, we used a transgenic mouse strain overexpressing the human HSPB1 protein. In the in vivo experiments, 7-day-old transgenic and wild-type mice were treated with ethanol. Apoptotic cells were detected using TUNEL assay. The mRNA and protein levels of cytokines and glial cell markers were examined using RT-PCR and immunohistochemistry in the brain. We also established primary neuronal, astrocyte, and microglial cultures which were subjected to cytokine and ethanol treatments. TNFα and hHSPB1 levels were measured from the supernates by ELISA, and intracellular hHSPB1 expression was analyzed using fluorescent immunohistochemistry. Results Following ethanol treatment, the brains of hHSPB1-overexpressing mice showed a significantly higher mRNA level of pro-inflammatory cytokines (Tnf, Il1b), microglia (Cd68, Arg1), and astrocyte (Gfap) markers compared to wild-type brains. Microglial activation, and 1 week later, reactive astrogliosis was higher in certain brain areas of ethanol-treated transgenic mice compared to those of wild-types. Despite the remarkably high expression of pro-apoptotic Tnf, hHSPB1-overexpressing mice did not exhibit higher level of apoptosis. Our data suggest that intracellular hHSPB1, showing the highest level in primary astrocytes, was responsible for the inflammation-regulating effects. Microglia cells were the main source of TNFα in our model. Microglia isolated from hHSPB1-overexpressing mice showed a significantly higher release of TNFα compared to wild-type cells under inflammatory conditions. Conclusions Our work provides novel in vivo evidence that hHSPB1 overexpression has a regulating effect on acute neuroinflammation by intensifying the expression of pro-inflammatory cytokines and enhancing glial cell activation, but not increasing neuronal apoptosis. These results suggest that hHSPB1 may play a complex role in the modulation of the ethanol-induced neuroinflammatory response.

Distinct brain regional proteome changes in the rTg‐DI rat model of cerebral amyloid angiopathy

“…A heat map indicating relative expression of the downstream target molecules identified in our proteomic analysis for all three regions is depicted in Figure 3b. Many of the mapped strongly elevated proteins common to the hippocampus and thalamus are specifically expressed in astrocytes, including Hspb1, S100a6, Clu, Htra1, and GFAP (Dukay et al, 2021;Launay et al, 2008;Morgan et al, 1995;Nafar et al, 2016;Yamashita et al, 1999), while other markers such as Vim and ApoE can be expressed in both astrocytes and microglia (Graeber et al, 1988;O'Leary et al, 2020). Activation of TGF-β1 is consistent with our recent findings that its mRNA expression is significantly elevated in 12 M rTg-DI rats (Zhu, Hatfield, et al, 2020).…”

Small Heat Shock Proteins in Retinal Diseases

“…Previous studies have shown that sHsps are anti-inflammatory proteins. Overexpression of HspB1 ameliorates neuroinflammation caused by ethanol-induced brain injury in mice ( Dukay et al, 2021 ). HspB5 reduces inflammation by decreasing the secretion of proinflammatory cytokines in animal models of multiple sclerosis, ischemia, or nerve crush injury ( Kurnellas et al, 2012 ; Rothbard et al, 2012 ; Lim et al, 2021 ).…”

APOE4drives transcriptional heterogeneity and maladaptive immunometabolic responses of astrocytes

“…As_9, over-represented in the E3 LPS brain, exhibited hallmark genes associated with scar formation and wound healing ( Cspg5, Htra1 ), 25,26 , while E4 LPS-enriched As_8/11 subtypes clustered similarly together and expressed genes associated with pro-inflammatory response ( Lcn2, Cxcl10 ), as well as oxidative stress and metal cation buffering (Fig. 1d; Hsbp1, S100a6, Mt1, Mt2 ) 27–29 . Using Monocle2 as an alternate analysis pipeline, we demonstrate via pseudotime trajectory that several of the hallmark genes from As_8/11 are highly expressed in the E4_LPS condition (Fig 1e; purple).…”