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. 2023 Feb 22;13(8):862–879. doi: 10.1016/j.jpha.2023.02.007

Fig. 7.

Fig. 7

The combination of insulin like growth factor 1 (IGF1) and osteopontin (OPN) was the necessary and sufficient condition for repopulating microglia (RM) function. (A–F) Reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) analysis of complement C3 (C3) messenger RNA (mRNA) expression in astrocytes (A) and tumor necrosis factor-α (TNF-α) (B), interleukin-1a (IL-1a) (C), complement C1q (C1q) (D), IGF1 (E), and OPN (F) in microglia. ∗∗∗P < 0.001, ∗∗P < 0.01, and ∗P < 0.05 vs. sham group. #P < 0.05 vs. intracerebral hemorrhage (ICH) + control diet group (n = 8/group). (G) Quantification of lesion area at 7 days post-injury (dpi). ∗P < 0.05 vs. ICH + adeno-associated virus (AAV)-control (n = 6/group). (H) Representative fluorescent images of brain sections at 7 dpi (left: 0.75 mm anterior to bregma; right: 0.25 mm prior to bregma). (I, J) Representative Western-blot bands show the expression of phospho-S6 (pS6) (a recognized marker of mechanistic target of rapamycin (mTOR) activation) in brain tissues at 7 dpi, ∗∗∗P < 0.001, ∗∗P < 0.01, and ∗P < 0.05 vs. sham group. #P < 0.05 vs. ICH + AAV-control (n = 6/group). (K) Representative images of glial fibrillary acidic protein (GFAP) and pS6 double immunostaining at 7 dpi (left). Histograms show that pS6 largely increased and colocalized with GFAP in AAV-OPN/IGF1-treated mice at 7 dpi (right). (L) Representative images of GFAP and C3 double immunostaining in cultured primary astrocytes. (M) C3 mRNA expression in cultured astrocytes. ∗∗∗P < 0.001 and ∗P < 0.05 vs. control group. #P < 0.05 vs. microglia conditional medium (MCM) treated group (n = 6/group). (N, O) Representative images of F-actin immunostaining in cultured primary astrocytes. (P) Methylthiazole tetrazolium (MTT) analysis. ##P < 0.01 vs. MCM treated group (n = 6/group). MBP: myelin basic protein; DAPI: 4′,6-diamidino-2-phenylindole.