hGPR45

Fig.1 Detection of GPR45 expression in hGPR45 mice by WB. GPR45 was detectable in hypothalamus from both WT C57BL/6 and HO hGPR45 mice, as the antibody was cross-reactived between human and mouse GPR45. Under fed conditions, Jak2 (Tyr1007/1008) autophosphorylation showed no significant difference between the hypothalamus from hGPR45 knock-in mice and WT mice. Following a 6-hour fast, Jak2 (Tyr1007/1008) autophosphorylation was markedly reduced in hGPR45 knock-in mice. The hypothalamus tissue lysates were collected from 6-week-old male wild-type C57BL/6 mice and 6-week-old male homozygous hGPR45 mice, and then analyzed by western blot with anti-GPR45 antibody, anti-p-JAK2 (Tyr1007/1008) antibody and anti-JAK2 antibody. The gray value was calculated using Image J. (Mean±SEM, Unpaired t-test, *p<0.05)

Abbr. HO, homozygous; WT, wild type.

Fig.2 Detection of mouse Pomc mRNA expression in hypothalamus and cortex from hGPR45 mice by qPCR. Hypothalamic and cortical Pomc mRNA expression levels showed no significant differences between hGPR45 knock-in mice and WT mice under both fed and 6-hour fasted conditions. Tissues were collected from WT C57BL/6 mice and homozygous hGPR45 knock-in mice. Total RNA was extracted, then cDNA libraries were synthesized by reverse transcription, followed by qPCR with mouse Pomc and mouse Gapdh primers. Relative expression represents the mouse Pomc mRNA level relative to its average expression in hypothalamus and cortex of WT C57BL/6 mice in the fed group, respectively. 

Abbr. HO, homozygous; WT, wild type.

Fig.3 Body weight and organs weight of WT C57BL/6 mice and hGPR45 knock-in mice. Body weight and the ratios of eWAT, iWAT, BAT and liver weight to body weight showed no significant differences between hGPR45 knock-in mice and WT mice under 6-hour fasted conditions. Male, 6-week-old mice were fasted for 6 h prior to body weight measurement. Epididymal white adipose tissue (eWAT), inguinal WAT (iWAT), brown adipose tissue (BAT), and liver tissues were harvested and weighed. (n=4 in each group, Mean±SEM, Unpaired t-test, ns: no significance) 

Abbr. HO, homozygous; WT, wild type.

Fig.4 Detection of mouse insulin levels in plasma of WT C57BL/6 mice and hGPR45 knock-in mice by ELISA. There was no significant differences in insulin levels between WT C57BL/6 mcie and homozygous hGPR45 mice.

Male, 6-week-old micen (n=3/group) were fasted for 6 h prior to plasma colltection. The insulin levels were analyzed by ELISA with anti-mouse insulin ELISA kit. 

Abbr.  HO, homozygous; WT, wild type.

Tabel 1. Blod biochemical test of WT C57BL/6 mice and hGPR45 knock-in mice. ALT, AST, UREA, CREA, glucose, TG, T-CHO, HDL-C and LDL-C levels showed no significant differences between hGPR45 knock-in mice and WT mice under 6-hour fasted conditions. NEFA levels were sightly reduced in hGPR45 knock-in mice compared with WT mice. Male, 6-week-old mice were fasted for 6 h prior to body weight measurement. Serum was collected for the detection. (n=4 in each group, Mean±SEM, Unpaired t-test) 

Abbr. HO, homozygous; WT, wild type.