Journal: Scientific Reports

Article Title: Anti PD-L1 immunotherapy alters macrophage phenotypes via EGR1 and HSP90AB1 supported by integrated methodologies

doi: 10.1038/s41598-025-20826-9

Figure Lengend Snippet: Anti-PD-L1 immunotherapy drives foam cell formation through EGR1 and HSP90AB1. EGR1 and HSP90AB1 were key genes responsible for circulating macrophage phenotypes in in the CHEMO and ANTI-PD-L1 + CHEMO groups. ( A ) The percentage of M1 and M2 macrophage clustering relative to all clustered cells was compared. (B) A violin plot was utilized to display the standardized expression levels of EGR1 and HSP90AB1 in macrophage clusters between both groups. ( C ) A volcano plot was used to visualize the differential gene expression of macrophage clusters in the CHEMO group and ANTI-PD-L1 + CHEMO group, where red indicates upregulated genes and blue indicates downregulated genes. ( D ) Western blots indicating total EGR1, HSP90AB1, and β-Actin levels in whole cell lysates. ( E ) Quantification of EGR1 protein levels from panel ( D ). *** denotes p-value < 0.001, ** denotes p-value < 0.01 by Two-way ANOVA ( n = 4, > 1000 cells per trial). ( F ) Quantification of HSP90AB1 protein levels from panel ( D ). ***denotes p-value < 0.01, * denotes p-value < 0.05 by Two-way ANOVA ( n = 4, > 1000 cells per trial).

Article Snippet: Western blotting was performed as previously described by our laboratory using the antibodies listed below: anti-β-Actin (13E5) rabbit mAb (CST, MA, USA, #4970,1:1000 dilution), anti-human CD86 (Huabio, Hangzhou, Zhejiang, China, ET1606-50, 1:1000 dilution), anti-human CD80 (Huabio, Hangzhou, Zhejiang, China, M1007-10, 1:1000 dilution), anti-human CD206 (Huabio, Hangzhou, Zhejiang, China, ET1702-04, 1:1000 dilution), Rabbit anti-human CD163 (abcam, MA, USA, ab182422, 1:1000 dilution), early growth response-1 (EGR‐1) Rabbit pAb (ABclone, MA, USA, A2722, 1:1000 dilution), and heat shock protein 90 kDa alpha family class B member 1 (HSP90AB1) Polyclonal antibody (Proteintech, Wuhan, Hubei, China, 11405-1-AP, 1:1000 dilution).

Techniques: Expressing, Gene Expression, Western Blot