NLRP3 Is Involved in the Maintenance of Cerebral Pericytes

Abstract

Pericytes play a central role in regulating the structure and function of capillaries in the brain. However, molecular mechanisms that drive pericyte proliferation and differentiation are unclear. In our study, we immunostained NACHT, LRR and PYD domainscontaining protein 3 (NLRP3)-deficient and wild-type littermate mice and observed that NLRP3 deficiency reduced platelet-derived growth factor receptor β (PDGFRβ)-positive pericytes and collagen type IV immunoreactive vasculature in the brain. In Western blot analysis, PDGFRβ and CD13 proteins in isolated cerebral microvessels from the NLRP3-deficient mouse brain were decreased. We further treated cultured pericytes with NLRP3 inhibitor, MCC950, and demonstrated that NLRP3 inhibition attenuated cell proliferation but did not induce apoptosis. NLRP3 inhibition also decreased protein levels of PDGFRβ and CD13 in cultured pericytes. On the contrary, treatments with IL-1β, the major product of NLRP3-contained inflammasome, increased protein levels of PDGFRβ, and CD13 in cultured cells. The alteration of PDGFRβ and CD13 protein levels were correlated with the phosphorylation of AKT. Inhibition of AKT reduced both protein markers and abolished the effect of IL-1β activation in cultured pericytes. Thus, NLRP3 activation might be essential to maintain pericytes in the healthy brain through phosphorylating AKT. The potential adverse effects on the cerebral vascular pericytes should be considered in clinical therapies with NLRP3 inhibitors.

Haploinsufficiency of microglial MyD88 ameliorates Alzheimer's pathology and vascular disorders in APP/PS1‐transgenic mice

“…In the 8‐day treatment experiment, withdrawal of IL‐1β for the last 3 days restored expression of LRP1 in cultured pericytes (Figure 6j,k; two‐way ANOVA, p <.05). The short‐term treatment of IL‐1β markedly increased expression of PDGFRβ and CD13, which corroborates our recent finding (Quan et al, 2020) (Figure 6f,h,i; one‐way ANOVA, p <.05). The long‐term treatment of IL‐1β only at a high concentration (e.g., 50 ng/ml) tended to decrease the expression of PDGFRβ and CD13; however, it was not statistically significant (shown in Figure 6l,m with solid lines; one‐way ANOVA, p >.05).…”

“…To quantify vasculature in the brain, our established protocol was used (Decker et al, 2018; Quan et al, 2020). Four serial paraffin‐embedded sections per mouse with 300 μm of distance in between were deparaffinized, heated at 80°C in citrate buffer (10 mM, pH = 6) for 1 hr and digested with Digest‐All 3 (Pepsin) (Thermo Fisher Scientific) for 20 min.…”

Not All Lectins Are Equally Suitable for Labeling Rodent Vasculature

“…Our findings also underline the importance of establishing a reliable and consistent method to implement lectin use in histological staining of blood vessels. Indeed, several recently published studies relied on the IB4 lectin to stain blood vessels in embryos [ 46 ], in the retina [ 47 , 48 , 49 ], and also in the brain [ 50 , 51 ]; however, our systematic comparison showed that the LEA lectin is preferable to IB4 in such applications, particularly in the brain of adult mice.…”

GTPBP4: A New Therapeutic Target Gene Promotes Tumor Progression in Non-Small Cell Lung Cancer via EMT

“…Western blotting analysis was performed as described [17]. Briefly, Frozen mouse lung tissue or NSCLC cells were homogenized on ice in 5 × radioimmunoprecipitation assay buffer (RIPA buffer; 50 mM Tris [pH 8.0], 150 mM NaCl, 0.1% SDS, 0.5% sodiumdeoxy-cholate, 1% NP-40, and 5 mM EDTA) supplemented with protease inhibitor cocktail (Sigma-Aldrich, St.Louis, MO, USA), followed by centrifugation at 16,000 × g for 30 minutes at 4 °C to collect the supernatants.…”