Objective: To investigate the effect of PIAS1 (protein inhibitor of activated STAT1) on regulating the migration of macrophage via transgenic zebrafish (Lyz-EGFP) with green fluorescent protein labelling macrophages, so as to provide evidences for basic research on anti-inflammatory role of PIAS1. Methods: Morpholino-PIAS1 (checking plasmid PIAS1 (213bp) GFP), PIAS1-mRNA (mRNA expressing plasmid PIAS1), and mismatch plasmid (as negative control) were established. The morpholino-PIAS1, PIAS1-mRNA, and mismatch plasmid were injected separately or together into zebrafish embryo using micro-injection method. When cultured for 6 h, the fluorescent protein expression was assessed under fluorescence microscopy to determine the efficiency and specificity of transfection. After cultured for 60 h, the tail of zebrafish was cut with a sterile scapel, and the migration and count of macrophages at the cutting site were observed by confocal microscope. The activation of ERK/JNK/p38MAPK/MMPs signal protein and MMP-9/TIMP-1 protein expression was detected by western blotting. Results: In single cell of 6 h embryo, the green fluorescence was reduced in morpholino-PIAS1 injected group, while no change was seen in mismatch control group. There was no significant difference in red fluo-rescence between these groups. And in the cutting tail of zebrafish, the migration of green fluorescent cells was increased in morpholino-PIAS1 injected group, mismatch control group and rescue group (morpholino-PIAS1+PIAS1-mRNA), while it was reduced in PIAS1-mRNA injected group, the difference was statistically significant(P<0.05). Western blotting showed that the phosphorylation levels of ERK/JNK/p38MAPK proteins and the ratio of MMP-9/TIMP1 proteins were increased in morpholino-PIAS1 injected group, mismatch control group and the rescue group, while were decreased in the PIAS1-mRNA injected group(P<0.05). Conclusions: PIAS1 is an inhibitor for macrophage activation, and is involved in ERK/JNK/p38MAPK/MMPs signaling transduction pathways. The up-regulation of PIAS1 expression could inhibit activation of macrophage and to ameliorate the severity of inflammatory diseases.
WANG Weiyi, ZHANG Yongping, YUAN Yaozong, WU Yunlin, CHEN Ping
. Experimental study on effect of PIAS1 in regulating migration of macrophage and its mechanism[J]. Journal of Diagnostics Concepts & Practice, 2017
, 16(01)
: 60
-65
.
DOI: 10.16150/j.1671-2870.2017.01.012
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