HSF1 in macrophages suppressed the progression of asthma via modulating SIRPα/SHP2-Dectin-1/ SYK mediated ROS and inflammatory responses

Abstract

HSF1, SIRPα, and Dectin-1 play crucial roles in immune regulation and inflammatory responses, their rols in asthma remained unclear, thereby the study was carried on. Twenty-one SPF-grade C57BL/6 mice were randomly divided into three groups: sham group, Model group, and Model + HSF1A group, with seven mice in each group. Except for the sham group, the other two groups were induced with OVA to establish an asthma model. The Model + HSF1A group was additionally treated with HSF1A. General conditions of the mice were observed. Lung tissue damage was assessed with Masson staining. RAW264.7 cells were divided into NC group, OVA + LPS group, HSF1A + OVA + LPS group, KRIBB11 + OVA + LPS group, SIRPα-OE + KRIBB11 + OVA + LPS group, SIRPα-OE + OVA + LPS group and SIRPα-KD + KRIBB11 + OVA + LPS group. The levels of GSH, MDA, IL-1β and TNFα in serum and cell supernatants were determined by ELISA. Protein expression in lung tissue and RAW264.7 cells was detected by Western blotting. In in vivo experiments, OVA-induced asthmatic mice exhibited severe airway resistance, collagen deposition, and elevated ROS and pro-inflammatory cytokines.HSF1A treatment improved lung function, reduced fibrosis, and restored redox balance. In vitro, HSF1A enhanced SIRPα expression and inhibited SYK/Dectin-1 signaling in LPS/OVA-stimulated macrophages, whereas HSF1 knockdown exacerbated inflammation. overexpression of SIRPα reversed KRIBB11-induced SYK activation, confirming the regulatory role of HSF1. HSF1 in macrophages regulates ROS and inflammatory responses by modulating the SIRPα/Dectin-1/SYK balance, thereby inhibiting the progression of asthma.

Keywords: Asthma; HSF1; Inflammation; ROS; SIRPα/Dectin-1/SYK.

Fig. 1

Bioinformatics analysis. A: Box plot for sample correction. B: Density plot of sample expression. C: Sample quantile plot. D: Volcano plot of differentially expressed genes (DEGs). E: Mean difference plot of DEGs. F: Bar chart of GO enrichment analysis. G: Chord diagram of KEGG enrichment analysis. H: Protein-Protein Interaction (PPI) network diagram. I: Scatter plot of ROS and SIRT1 expression levels. J: Scatter plot of ROS and P38 expression levels. K: Scatter plot of SIRT1 and HSF1 expression levels.

Fig. 2

HSF1 improves lung function and lung injury in asthmatic mice. A: Masson staining of lung tissue changes in the Sham, Model, and Model + HSF1A groups. Scale: 100 μm. B: Immunofluorescence showing CD68 levels in the Sham, Model, and Model + HSF1A groups. Scale: 50 μm. C: Assessment of lung function in mice from the Sham, Model, and Model + HSF1A groups, including FEV1, FVC, FEV1/FVC, FEF 25%, and Ri changes. Data are expressed as mean ± standard deviation. N = 7; Sham vs. Model: *P < 0.05; **P < 0.01; Model vs. Model + HSF1A: ^#P < 0.05; ^##P < 0.01.

Fig. 3

HSF1 promotes SIRPα activity and inhibits p-SYK/t-SYK and Dectin-1 function. A: ELISA measuring levels of GSH, MDA, IL-1β, and TNFα in the Sham, Model, and Model + HSF1A groups. B: Western blot analysis of protein expression for P47 (ROS), p-P38/t-P38, p-SIRT1/t-SIRT1, HSF1, SIRPα, p-SYK/t-SYK, and Dectin-1 in the Sham, Model, and Model + HSF1A groups. GAPDH as control protein. Data are expressed as mean ± standard deviation. N = 7; Sham vs. Model: *P < 0.05; **P < 0.01; Model vs. Model + HSF1A: ^#P < 0.05; ^##P < 0.01.

Fig. 4

In macrophages, HSF1 mediates ROS and inflammatory responses by regulating the SIRPα/Dectin-1/SYK balance, thereby inhibiting asthma progression. A: Western blot analysis of HSF1, SIRPα, p-SYK/t-SYK, Dectin-1, P22, P47, and p-P38/t-P38 protein expression in the NC, OVA + LPS, HSF1A + OVA + LPS, KRIBB11 + OVA + LPS, and SIRPα-OE + KRIBB11 + OVA + LPS groups. B: ELISA measurements of GSH, MDA, IL-1β, and TNFα levels in the NC, OVA + LPS, HSF1A + OVA + LPS, KRIBB11 + OVA + LPS, and SIRPα-OE + KRIBB11 + OVA + LPS groups. GAPDH as control protein. Data are expressed as mean ± standard deviation. N = 3; NC vs. OVA + LPS: **P < 0.01; ^nsP>0.05; OVA + LPS vs. HSF1A + OVA + LPS: ^#P < 0.05; ^##P < 0.01; OVA + LPS vs. KRIBB11 + OVA + LPS: ^&P < 0.05; ^&&P < 0.01; ^nsP>0.05; KRIBB11 + OVA + LPS vs. SIRPα + KRIBB11 + OVA + LPS: ^††P < 0.01; ^nsP>0.05.

Fig. 5

SIRPα inhibits activation of SYK/ROS/ERK1/2 signaling pathway in asthma. A: Western blot analysis of protein banding plots of p-SYK, t-SYK, P22, P47, p-ERK1/2, and t-ERK1/2 in NC, OVA + LPS, OVA + LPS + SIRPα-OE, and OVA + LPS + SIRPα-KD groups. B: Western blot analysis of p-SYK/t-SYK, P22, P47, and p-ERK1/2/t-ERK1/2 protein expression in the NC, OVA + LPS, OVA + LPS + SIRPα-OE, and OVA + LPS + SIRPα-KD groups. GAPDH as control protein. Data are expressed as mean ± standard deviation. N = 3; NC vs. OVA + LPS: **P < 0.01; OVA + LPS vs. OVA + LPS + SIRPα-OE: ^##P < 0.01; OVA + LPS vs. OVA + LPS + SIRPα-KD: ^&&P < 0.01.

Fig. 6

Mechanistic study of how HSF1 inhibits asthma progression through the SIRPα/Dectin-1/SYK balance-mediated ROS and inflammatory responses in macrophages (Created with BioRender.com).