α-NETA down-regulates CMKLR1 mRNA expression in ileum and prevents body weight gains collaborating with ERK inhibitor PD98059 in turn to alleviate hepatic steatosis in HFD-induced obese mice but no impact on ileal mucosal integrity and steatohepatitis progression.

BACKGROUND: Studies on chemerin/chemokine-like receptor-1 have mainly focused on adipose and liver with the intestinal tissues largely overlooked. In this study conducted on obese mice, we have explored: 1) CMKLR1 expression in the ileums; 2) CMKLR1 inhibitor α-NETA on body weight and intestinal mucosa integrity hence the impact on hepatic steatosis and pathway involved. METHODS: Nineteen male C57BL/6 mice were randomly divided into five groups: normal diet group (ND), high-fat diet group (HFD), HFD + α-NETA group (NETA), HFD + PD98059 group (PD) and HFD + α-NETA + PD98059 group (NETA + PD). Mice were fed either with a chow diet or HFD for 12 weeks. At 12th week, mice of ND were put on the diet as before; mice of NETA received daily treatments of α-NETA (30 mg/kg) via gavage; mice of PD received daily treatment of PD98059 via tail vein injection; mice of NETA + PD received daily treatment of α-NETA + PD98059, all for another 4 weeks. At the time intervention ended, mice were sacrificed. The body weight, the liver pathologies were assessed. Ileal CMKLR1 mRNA was evaluated by rtPCR; ZO-1, ERK1/2 protein expression of ileal tissues by western blotting; liver TNF-α and serum endotoxin by Elisa. RESULTS: More weight gains in mice of HFD than ND (37.90 ± 3.00 g) vs (24.47 ± 0.50 g), P = 0.002; α-NETA reduced the body weight (33.22 ± 1.90 g) vs (37.90 ± 3.00 g), P = 0.033; and further reduced by NETA + PD98059: (31.20 ± 1.74 g) vs (37.30 ± 4.05 g), P = 0.032. CMKLR1 mRNA expression was up-regulated in ileum in group HFD compared with ND and down-regulated by α-NETA. Steatosis was only alleviated in group PD + NETA with less weight gain. No impact of α-NETA on ileal ZO-1 or pERK with western blotting, and no endotoxin level changes were detected. TNF-α was higher in group HFD than in group ND, while no significant difference between other groups. CONCLUSIONS: CMKLR1 mRNA was up-regulated in the ileum of obese mice and down-regulated by α-NETA along with a body weight control collaborating with ERK inhibitor PD98059. Steatosis was alleviated in a weight dependent way. α-NETA has no influence on intestinal mucosal integrity and no impact on steatohepatitis progression.

GDF-15 Level Correlates with CMKLR1 and VEGF-A in Tumor-free Margin in Colorectal Cancer.

Colorectal cancer (CRC) is the third most frequently diagnosed cancer worldwide, responsible for over 880 000 deaths each year. Growth/differentiation factor 15 (GDF-15) is reported to be a promising diagnostic and prognostic factor in CRC. It induces pleiotropic effects in tumor cells: proliferation, stemness, invasion and metastasis. Some studies indicate that GDF-15 may stimulate angiogenesis in malignant neoplasms. However, it has not been investigated in CRC yet. The aim of our study was to determine the level of GDF-15 and the concentrations of hypoxia-inducible factor-1α (HIF-1α), VEGF-A and chemokine-like receptor 1 (CMKLR1) in tumor and margin specimens of CRC in relation to histological grade and TNM staging. The study comprised 33 samples of tumor and margin tissues obtained from CRC patients. To assess the concentration of GDF-15, HIF-1α, VEGF-A and CMKLR1, commercially available enzyme-linked immunosorbent assay (ELISA) kits were used. We found significantly increased levels of GDF-15 and CMKLR1 in tumor tissue compared to margin tissue and higher concentrations of HIF-1α and VEGF-A in margin tissue than in tumor tissue. The levels of GDF-15 and HIF-1α were significantly correlated with VEGF-A and CMKLR1 in margin tissue. In CRC, the increased level of GDF-15 might stimulate angiogenesis through upregulation of HIF-1α, VEGF A and CMKLR1 expression. Our study is the first one to reveal the correlation between the levels of GDF-15 and CMKLR1 in CRC. The elevated levels of HIF-1α and VEGF-A in tumor-free margin tissues suggest that noncancer cells in the tumor microenvironment are an important source of proangiogenic factors.

Chemerin promotes angiogenesis in vivo.

Chemerin acts as a chemotactic factor for leukocyte populations expressing the G protein-coupled receptor CMKLR1 (ChemR23). It is also an adipocytokine involved in obesity and metabolic syndromes. Previous studies have demonstrated that chemerin promotes angiogenesis in vitro, although the precise mechanism has not been elucidated. In this study, we have investigated whether chemerin regulates angiogenic processes and validated the associated mechanisms. In this study, chemerin stimulated angiogenesis in mice, which was demonstrated using Matrigel plug implantation assay, mouse corneal models of angiogenesis, and ex vivo rat aortic ring assay. To explore the mechanisms by which chemerin induced angiogenesis, we examined the effects of chemerin in human umbilical vein endothelium cells (HUVECs). Chemerin stimulated the differentiation of HUVECs into capillary-like structures, promoted the proliferation of HUVECs, and functioned as a chemoattractant in migration assays. Chemerin induced the phosphorylation of Akt and p42/44 extracellular signal-regulated kinase (ERK) in HUVECs and chemerin promotes angiogenesis via Akt and ERK. SiRNA against the chemerin receptor CMKLR1 but not that against another chemerin receptor, CCRL2, completely inhibited the chemerin-induced migration and angiogenesis of HUVECs, which indicates that chemerin promotes the migration and angiogenic activities of HUVECs mainly through CMKLR1.

CMKLR1 activation ex vivo does not increase proportionally to serum total chemerin in obese humans.

Prochemerin is the inactive precursor of the adipokine chemerin. Proteolytic processing is obligatory for the conversion of prochemerin into active chemerin and subsequent regulation of cellular processes via the chemokine-like receptor 1 (CMKLR1). Elevated plasma or serum chemerin concentrations and differential processing of prochemerin have been reported in obese humans. The impact of these changes on CMKLR1 signalling in humans is unknown. The objective of this pilot study was to develop a cellular bioassay to measure CMKLR1 activation by chemerin present in human serum and to characterise how obesity modifies serum activation of CMKLR1 under fasted and fed conditions. Blood samples were collected from control (N = 4, BMI 20-25) and obese (N = 4, BMI >30) female subjects after an overnight fast (n = 2) and at regular intervals (n = 7) following consumption of breakfast over a period of 6 h. A cellular CMKLR1-luminescent reporter assay and a pan-chemerin ELISA were used to determine CMKLR1 activation and total chemerin concentrations, respectively. Serum total chemerin concentration (averaged across all samples) was higher in obese vs control subjects (17.9 ± 1.8 vs 10.9 ± 0.5 nM, P < 0.05), but serum activation of CMKLR1 was similar in both groups. The CMKLR1 activation/total chemerin ratio was lower in obese vs control subjects (0.33 ± 0.04 vs 0.58 ± 0.05, P < 0.05). After breakfast, serum total chemerin or CMKLR1 activation did not differ from baseline values. In conclusion, the unexpected observation that obese serum activation of CMKLR1 did not match increased total chemerin concentrations suggests impaired processing to and/or enhanced degradation of active chemerin in serum of obese humans.