RESUMEN

BACKGROUND: The liver-expressed antimicrobial peptide 2 (LEAP2) is essential in host immunity against harmful pathogens and is only known to act as an extracellular modulator to regulate embryonic development in amphibians. However, there is a dearth of information on the antimicrobial function of amphibian LEAP2. Hence, a LEAP2 homologue from Leptobrachium liui was identified, characterized, and chemically synthesized, and its antibacterial activities and mechanisms were determined. RESULTS: In this study, LEAP2 gene (Ll-LEAP2) cDNA was cloned and sequenced from the Chong'an Moustache Toad (Leptobrachium liui). The predicted amino acid sequence of Ll-LEAP2 comprises a signal peptide, a mature peptide, and a prodomain. From sequence analysis, it was revealed that Ll-LEAP2 belongs to the cluster of amphibian LEAP2 and displays high similarity to the Tropical Clawed Frog (Xenopus tropicalis)'s LEAP2. Our study revealed that LEAP2 protein was found in different tissues, with the highest concentration in the kidney and liver of L. liui; and Ll-LEAP2 mRNA transcripts were expressed in various tissues with the kidney having the highest mRNA expression level. As a result of Aeromonas hydrophila infection, Ll-LEAP2 underwent a noticeable up-regulation in the skin while it was down-regulated in the intestines. The chemically synthesized Ll-LEAP2 mature peptide was selective in its antimicrobial activity against several in vitro bacteria including both gram-positive and negative bacteria. Additionally, Ll-LEAP2 can kill specific bacteria by disrupting bacterial membrane and hydrolyzing bacterial gDNA. CONCLUSIONS: This study is the first report on the antibacterial activity and mechanism of amphibian LEAP2. With more to uncover, the immunomodulatory functions and wound-healing activities of Ll-LEAP2 holds great potential for future research.

Asunto(s)

Antibacterianos , Péptidos Catiónicos Antimicrobianos , Animales , Péptidos Catiónicos Antimicrobianos/genética , Péptidos Catiónicos Antimicrobianos/farmacología , Péptidos Catiónicos Antimicrobianos/química , Secuencia de Aminoácidos , Secuencia de Bases , ARN Mensajero

RESUMEN

Objective To investigate the expression level of nuclear factor-kappa(NF-κB),substance P(SP)and aquaporin 4(AQP4)gene and the change of discharge frequency of pain sensitive neurons (PSN)in the ventral posteriolateral thalamic nucleus(VPL)of acute gout(AG)rats.The neuro-genic inflammation of AG was explored.Methods Forty-eight rats were randomly divided into two groups:the control group and the AG group. According to the time interval after injection of monosodiumurate(MSU)into the the unilateral ankle joint,the AG group was subdivided into seven groups,ie.0.5 hours group,2 hours group,6 hours group,12 hours group,24 hours group,48 hours group and 72 hours group.There were 6 rats in each group.After recording of the discharge frequency of PSN in rats VPL nucleus,the expression level of NF-κB,SP and AQP4 gene in the rats joint synovium were evaluated at mRNA level by PCR in above mentioned time.Results In the 0.5 hours group,the discharge frequency of PSN in the rats VPL nueleus and the expression level of SP gene in the rats joint synovium had increased immediately after the injection of MSU(P＜0.05).In the 6 hours group,they reached the peak level(P＜0.05),and in the 12 hours group,they began to decrease gradually(P＜0.05).In the 0.5 hours group,the expression level of NF-κB and AQP4 gene increased after the injection of MSU(P＜0.05).However,their peak level presented at the 12 hours(P＜0.05),and they decreaged after 24 hours(P＜0.05).The statistical analysis of correlation had shown that there were positive correlation among the expression level of NF-κB,substance P.AQP4 gene and the change of discharge frequency of PSN in the rats VPL nucleus.Conclusion The discharge frequency of PSN in the rats VPL and the expression level of SP gene in the rats ioint synovium can be used to evaluate the Severity of pain in the AG rats.The expression level of NF-κB and SP gene can reflect the severity of neurogenic intlammation.We can know the severity of edema of the joint synovium by detecting the expression level of AQP4 gene.Pain and neurogenic inflammation play an important role in the pathogenesis of AG.Combingelectrophysiology and biochemical technique can shade light on the pathogenesis of AG from different aspects.In the meantime,it may provide a new method for developing new drugs and new approaches for clinical treatment.