Epinecidin-1 has immunomodulatory effects, facilitating its therapeutic use in a mouse model of Pseudomonas aeruginosa sepsis.

Antimicrobial peptides (AMPs) are garnering attention as possible alternatives to antibiotics. Here, we describe the antimicrobial properties of epinecidin-1 against a multidrug-resistant clinical isolate of P. aeruginosa (P. aeruginosa R) and a P. aeruginosa strain from ATCC (P. aeruginosa ATCC 19660) in vivo. The MICs of epinecidin-1 against P. aeruginosa R and P. aeruginosa ATCC 19660 were determined and compared with those of imipenem. Epinecidin-1 was found to be highly effective at combating peritonitis infection caused by P. aeruginosa R or P. aeruginosa ATCC 19660 in mouse models, without inducing adverse behavioral effects or liver or kidney toxicity. Taken together, our results indicate that epinecidin-1 enhances the rate of survival of mice infected with the bacterial pathogen P. aeruginosa through both antimicrobial and immunomodulatory effects.

Stable expression in a Chinese hamster ovary (CHO) cell line of bioactive recombinant chelonianin, which plays an important role in protecting fish against pathogenic infection.

Chelonianin, originally isolated from the shrimp (Penaeus monodon), exhibits antimicrobial effects in vitro and in vivo and is used to treat infectious fish diseases. Herein, we report that the recombinant chelonianin protein fused to a fluorescent protein (rcf protein) was expressed from a stably transfected Chinese hamster ovary (CHO) cells. The in vitro experiments showed that the rcf protein exhibited antimicrobial activity against several bacteria, while the recombinant fluorescent protein alone did not. In addition, pretreatment and post-treatment with the rcf protein were both effective in promoting a significant decrease in fish mortality and decreasing the number of infectious bacteria. We utilized the quantitative reverse-transcriptase polymerase chain reaction technique to survey the levels of gene expressions of tumor necrosis factor-alpha (TNF-alpha) and nitric oxide synthase 1 induced in response to bacterial infection in experiments with tilapia (Oreochromis mossambicus). Our results indicated that the rescue of fish treated with the rcf protein may involve regulation of TNF-alpha expression. Collectively, chelonianin inhibited the production of an inflammatory mediator and reduced mortality in fish during bacterial challenge, suggesting that it has potential as a therapeutic or prophylactic drug for use against bacterial infectious diseases.

Epinecidin-1, an antimicrobial peptide from fish (Epinephelus coioides) which has an antitumor effect like lytic peptides in human fibrosarcoma cells.

Epinecidin-1, a synthetic 21-mer antimicrobial peptide originally identified from grouper (Epinephelus coioides), specifically exhibited high antimicrobial activities against both Gram-negative and Gram-positive bacteria. In the current study we report on the in vitro cytotoxicity of the peptide, an important factor before it can be considered for further applications in cancer therapy. The cytotoxicity of epinecidin-1 was investigated against several cancer cells (A549, HA59T/VGH, HeLa, HepG2, HT1080, RAW264.7, and U937) and normal cells (AML-12, NIH3T3, and WS-1) with the MTT assay, and the inhibition of cancer cell growth was confirmed by a soft agar assay and scanning electron microscopy. However, cell variations were detected with AO/EtBr staining, while apoptosis and necrosis gene expressions in HT1080 cells after treatment with the epinecidin-1 peptide and Nec-1 showed that epinecidin-1 had an anti-necrosis function in HT1080 cells. The data presented here indicate that epinecidin-1 has in vitro antitumor activity against the HT1080 cell line, and functions like lytic peptides. In addition, our results suggest that epinecidin-1 may prove to be an effective chemotherapeutic agent for human fibrosarcoma cells in the future.

In vitro activities of three synthetic peptides derived from epinecidin-1 and an anti-lipopolysaccharide factor against Propionibacterium acnes, Candida albicans, and Trichomonas vaginalis.

The synthetic epinecidin-1(22-42) peptide was derived from positions 22-42 of Epinephelus coioides epinecidin-1. The synthetic SALF(55-76) cyclic peptide (csSALF(55-76)) and SALF(55-76) linear peptide (lsSALF(55-76)) contained sequences from positions 55 to 76 of the Penaeus monodon anti-lipopolysaccharide factor (ALF), respectively. We studied the in vitro activities of epinecidin-1(22-42), csSALF(55-76), and lsSALF(55-76) against Propionibacterium acnes, Candida albicans, and Trichomonas vaginalis. The minimum inhibitory concentrations (MICs) of epinecidin-1(22-42) for the test pathogen strains ranged 12.5-200 microg/ml, those of csSALF(55-76) ranged 100-200 microg/ml, and those of lsSALF(55-76) ranged 25-200 microg/ml. epinecidin-1(22-42) exhibited cytotoxicity towards P. acnes, C. albicans, and T. vaginalis (one strain of which was a metronidazole-resistant strain, while the other strain was not), suggesting that epinecidin-1 functions like a lytic peptide. Similar cytotoxicity was identified against T. vaginalis treated with the csSALF(55-76) and lsSALF(55-76) peptides. The antimicrobial activities of these peptides were confirmed by scanning electron microscopy (SEM), transmission electron microscopy (TEM), a viable cell count assay, and flow cytometric analysis. TEM and SEM examinations of T. vaginalis treated with these three peptides showed that severe swelling preceded cell death and breakage of the outer membrane, and the intracellular inclusion was found to have effluxed extracellularly. This phenomenon was also found with epinecidin-1(22-42) treatment of P. acnes and C. albicans. Our results suggest that the epinecidin-1(22-42), csSALF(55-76), and lsSALF(55-76) peptides may be good candidates for treating trichomoniasis, and epinecidin-1(22-42) may have potential as a drug supporting therapy for acne and candidiasis.

A fish antimicrobial peptide, tilapia hepcidin TH2-3, shows potent antitumor activity against human fibrosarcoma cells.

As part of a continuing search for potential anticancer drug candidates from antimicrobial peptides of marine organisms, tilapia (Oreochromis mossambicus) hepcidin TH2-3 was evaluated in several tumor cell lines. The results indicated that TH2-3, a synthetic 20-mer antimicrobial peptide, specifically inhibited human fibrosarcoma cell (HT1080 cell line) proliferation and migration. The way in which TH2-3 inhibited HT1080 cell growth was then studied. TH2-3 inhibited HT1080 cell growth in a concentration-dependent manner according to an MTT analysis, which was confirmed by a soft-agar assay and AO/EtBr staining. Scanning electron microscopy revealed that TH2-3 caused lethal membrane disruption in HT1080 cancer cells, and a wound healing assay supported that TH2-3 decreased the migration of HT1080 cells. In addition, c-Jun mRNA expression was downregulated after treatment with TH2-3 for 48-96 h compared to the untreated group. These findings suggest a mechanism of cytotoxic action of TH2-3 and indicate that TH2-3 may be a promising chemotherapeutic agent against human fibrosarcoma cells.

Phosphatidylserine receptor is required for the engulfment of dead apoptotic cells and for normal embryonic development in zebrafish.

During development, the role of the phosphatidylserine receptor (PSR) in the removal of apoptotic cells that have died is poorly understood. We have investigated this role of PSR in developing zebrafish. Programmed cell death began during the shield stage, with dead cells being engulfed by a neighboring cell that showed a normal-looking nucleus and the nuclear condensation multi-micronuclei of an apoptotic cell. The zebrafish PSR engulfing receptor was cloned (zfpsr), and its nucleotide sequence was compared with corresponding sequences in Drosophila melanogaster (76% identity), human (74%), mouse (72%) and Caenorhabditis elegans (60%). The PSR receptor contained a jmjC domain (residues 143-206) that is a member of the cupin metalloenzyme superfamily, but in this case serves an as yet unknown function(s). psr knockdown by a PSR morpholino oligonucleotide led to accumulation of a large number of dead apoptotic cells in whole early embryo. These cells interfered with embryonic cell migration. In addition, normal development of the somite, brain, heart and notochord was sequentially disrupted up to 24 hours post-fertilization. Development could be rescued in defective embryos by injecting psr mRNA. These results are consistent with a PSR-dependent system in zebrafish embryos that engulfs apoptotic cells mediated by PSR-phagocytes during development, with the system assuming an important role in the normal development of tissues such as the brain, heart, notochord and somite.