α 1-antitrypsin modulates microglial-mediated neuroinflammation and protects microglial cells from amyloid-β-induced toxicity

Abstract

BackgroundOne hallmark of Alzheimer disease is microglial activation. Therapeutic approaches for this neurodegenerative disease include the modulation of microglial cells. α1-antitrypsin (A1AT) has been shown to exert anti-inflammatory effects on macrophages and lung epithelial cells and an inhibition of calpain activity in neutrophil granulocytes. Nothing is known about the effect of A1AT on microglial-mediated neuroinflammation. Our aim was to investigate the effect of A1AT on amyloid-β (Aβ)- and LPS-treated microglial cells in vitro with respect to cytokine production, stress pathways, cell viability, phagocytotic abilities and the underlying mechanisms.MethodsPrimary microglial cells were isolated from Swiss Webster mouse embryos on embryonic day 13.5. Cytokines in the supernatants of treated primary microglial cells were analyzed with ELISAs, and accumulated nitrite was detected with Griess reagents. Intracellular stress pathways were investigated in cell lysates using western blotting. Intracellular calcium levels were detected in BV-2 microglial cells loaded with the Ca2+-sensitive (fluorescent) dye Fluo-4. Calpain activity in primary microglial cells was assessed by using a calpain activity assay. Cell viability of Aβ-treated microglial cells was analyzed using MTT assay. Phagocytosis of Aβ was evaluated with western blot analysis.ResultsUpon co-administration, A1AT reduced pro-inflammatory mediators induced by LPS or Aβ. Interestingly, we detected a reduction in calpain activity and in the concentration of intracellular calcium that might mediate the anti-inflammatory effects of A1AT. Inhibition of the classic activation pathways, such as phosphorylation of mitogen-activated protein kinases or activation of protein kinase A were excluded as a mechanism of A1AT-mediated effects. In addition, A1AT increased the viability of Aβ-treated microglial cells and reduced Aβ phagocytosis.ConclusionsWe provide evidence on the mechanism of action of A1AT on microglial-mediated neuroinflammation in vitro. Our in vitro data indicate that A1AT treatment modulates microglial cells in inflammatory conditions and that this modulation is due to an inhibition of calpain activity and intracellular calcium levels. The underlying mechanisms of the effects observed here are promising for future therapeutic strategies and should thus be further pursued in transgenic mouse models of Alzheimer disease.

CSF SerpinA1 in Creutzfeldt–Jakob disease and frontotemporal lobar degeneration

“…Alternatively, SerpinA1 post‐translational modifications might be detrimental by (1) blocking specific serine proteases and lead to impaired clearance of prion‐like proteins or (2) promoting the formation of a misfolded and self‐aggregate‐prone structure of SerpinA1 . On another issue, SerpinA1 is emerging as an important modulator of neuroinflammation …”

“…In this regard, overexpression of the protein or its RNA has been described in the brains of patients with Alzheimer’s disease (AD) and frontotemporal lobar degeneration (FTLD), suggesting that SerpinA1 overexpression may be detrimental to neuronal function . Moreover, the protein may play an anti‐inflammatory role by attenuating microglial activation, causing a reduction of neuroinflammation . It is also known that the protein might diffuse out of venous blood and be released from the brain tissue into the cerebrospinal fluid (CSF) .…”

Cerebrospinal Fluid Proteins as Regulators of Beta‐amyloid Aggregation and Toxicity

“…Despite the failure to inhibit fibril formation, A1AT protected human red blood cells from Aβ fibril‐induced lysis . Additionally, in primary microglial cells, A1AT increased cell viability and reduced the release of both TNF‐α and IL‐6 stimulated by Aβ oligomer treatment . This protective effect was not attributable to inhibition of aggregation, as western blot analysis showed no change in Aβ oligomer formation upon incubation with A1AT.…”

“…These results suggest that A1AT attenuates Aβ toxicity indirectly, possibly through its anti‐inflammatory function. However, inhibition of oligomer formation was assessed at a 1 : 13 ratio A1AT/Aβ, whereas the neuroprotective effects by A1AT were obtained at a 9 : 1 ratio . This large discrepancy precludes any firm conclusions connecting inhibition of aggregation to inhibition of toxicity.…”

Commentary: Effect of Probiotic Supplementation on Cognitive Function and Metabolic Status in Alzheimer's Disease: A Randomized, Double-Blind and Controlled Trial

“…Further, a strong correlation was found between anti-inflammatory α1-antitrypsin and pro-inflammatory S100A12 ( p < 0.001) in the fecal specimens of our cognitively impaired patients. The anti-inflammatory action of α1-antiptrypsin on microglial mediated neuroinflammation could be shown in vitro (Gold et al, 2014). In our series, α1-antitrypsin was also correlated with zonulin ( p < 0.01), a protein modulating tight junction permeability between cells of the digestive tract.…”