Effect Of CMKLR1 Signaling On LPS-Induced Orchitis In Mice

Testis is mainly composed of seminiferous tubules and interstitial, both reproductive system and part of the endocrine system, its functions include spermatogenesis and steroidogenesis. Orchitis is an inflammation lesion with leukocyte infiltration and seminiferous tubule injury in testis. Chemerin is a newly identified adipokine in 2007, and plays an important role in inflammation, adipogenesis, energy metabolism, reproductive regulation and so on. CMKLR1(chemokine like receptor 1) is one of chemerin's receptors. In the present study, we established model of mouse inflammation to investigate effects of CMKLR1 deficiency on lipopolysaccharide (LPS) induced mouse orchitis with time-dependence.Materials and methods:Male CMKLR1 deficient (CMKLR1-/-) mice (8-10 wk old) and corresponding C57BL/6 wild type (WT) mice (8-10 wk old). Mice were injected intraperitoneally of LPS solution for 0.5 mg/Kg-BW to establish mouse orchitis model. Respectively, mice were killed by decapitation in the processing 6 h,12 h and 24 h later, serum, testes and epididymis were been cryopreservation for next experiment. The methods, including RT-qPCR, H.E. staining and radibimmunoassay (RIA), were used to detect pro-inflammatory factors, steroidogenic enzymes, hormones and morphological changes.Results:1. Judgment of model:H.E. staining results show that LPS induced accumulation of immature germ cells in lumen and epithelium damage after 6 h in WT mice; seminiferous epithelium damage and spermatogenic cell shedding after 12 h; while it was gradually restored after 24 h. RT-qPCR shows the relative expression of TNFa and IL6 is significantly increased in testes after 6 h?12 h and 24 h. The above results suggest that the model of mouse orchitis is successful after 6 h LPS treatment, strongest inflammation reaction in 12 h and gradually restored in 24 h.2. Effect of CMKLR1 on LPS induced mouse orchitis:RT-qPCR shows significant inflammatory response in WT mice exposed to LPS, this response is attenuated in CMKLR1 deficient mice, mainly for the smaller increase of pro-inflammatory factors mRNA expression. It suggests the pro-inflammatory role of CMKLR1 in mouse acute orchitis.3. Effect of CMKLR1 on steroidogenesis and spermatogenesis in mouse testis exposed to LPS:H.E. staining results show slowly recovery in CMKLR1 deficient mice. RT-qPCR shows mRNA expressions of StAR, P450scc,3?-HSD and LHR are further decreased in CMKLRl deficient mice, but FSHR between WT and CMKLR1-/- are similar. RT-qPCR shows mRNA expressions of chemerin are decreased exposed to saline in CMKLR1-/- mice, meanwhile, GPR1 are smaller decreased and CCRL2 are further. RIA shows further decrease in serum testosterone and estradiol. The above results suggest:(1) CMKLR1 affects spermatogenesis in mice; (2) CMKLR1 decreased the inhibition of LPS on steroidogenesis in mice; (3) CMKLR1 attenuated the inhibition of LPS on serum testosterone and estradiol secrestion in mice.4. Effect of CMKLRl on LPS induced mouse epididymitis:H.E. staining results show that CMKLR1-/- induced sperm stasis in cauda exposed to LPS 12 h later, but no significant changes in caput or corpus. RT-qPCR shows further increase of TNFa and IL6 mRNA expression in CMKLRl-/- mice. It suggests the anti-inflammatory role of CMKLR1 in LPS induced inflammation in mouse epididymitis.Conclusion:1:CMKLR1 is advantage to steroidogenesis and probably affects spermatogenesis in mouse.2:CMKLR1 shows the pro-inflammatory role in mouse testis but anti-inflammatory role in mouse epididymis. It requires further study.