Preliminary report on the hemagglutinating activity of the Scorpaena plumieri fish venom.

The scorpionfish Scorpaena plumieri is one of the most venomous fish species in the Brazilian coast. Amongst many biological activities, the S. plumieri fish venom (SpV) promotes hemagglutination. Although this activity appears to be associated to the presence of C-type lectins in the venom, it has not yet been chemically or functionally characterized. In the present work we sought to advance the characterization of the hemagglutinating activity associated to this venom. By fractionating SpV through saline precipitation followed by size exclusion chromatography we obtained two purified fractions - HF1 and HF3 - with Ca2+-dependent agglutinating activity against rabbit erythrocytes, which remained stable upon storage at 4 and -80oC. HF1 and HF3 were bacteriostatic against Gram-positive bacteria (Staphylococcus aureus), displaying minimum inhibitory concentration (MIC) of 50 and 200 µg/mL, respectively. In addition, a resazurin-based viability assay revealed that both fractions, at doses up to 370 µg/mL, were cytotoxic against tumor and non-tumor cell lines. Finally, a tendency towards edema formation could be detected when the fractions - particularly HF1 - were injected into mice footpads. We believe our data contribute to a better understanding of the biological properties of the so often neglected fish venoms.

Antiosteoporosis effects of a marine antimicrobial peptide pardaxin via regulation of the osteogenesis pathway.

Antimicrobial peptides (AMPs) are known to play an important role in natural immunity. Moreover, the diverse biological activities of AMPs showed great potency in treating many diseases. Thus, in this study, we used an AMP, that is, pardaxin, from a marine fish (Pardachirus marmoratus), which has been reported to possess antibacterial and antitumor activities. We first investigated the mechanisms of pardaxin in promoting osteogenic differentiation in vitro and in vivo. As per our data, it was determined that pardaxin could stimulate bone morphogenetic protein-2 (BMP-2) and downstream cascade. The activation of BMP-2 could further induce the phosphorylation of Akt and extracellular signal-regulated kinase (ERK). Additionally, the activation of p-Akt and p-ERK could prompt the elevation and translocation of runt-related transcription factor 2 (runx-2), which is associated with osteoblast differentiation. The translocation of runx-2 initiated transcription and translation of osteogenesis-related markers, including alkaline phosphatase (ALP), osterix, and osteocalcin. Pardaxin significantly facilitated preosteoblast cells in mineralization and reversed dexamethasone- (DM-) induced zebrafish bone formation deficiency by activating the osteogenesis pathway. Therefore, we suggest that pardaxin could be a possible candidate for osteoporosis treatment and a promising therapeutic agent.

Anti-Glioma Effect of Pseudosynanceia melanostigma Venom on Isolated Mitochondria from Glioblastoma Cells

Background: Glioblastoma is the most common primary malignant tumor of the central nervous system that occurs in the spinal cord or brain. Pseudosynanceia melanostigma is a venomous stonefish in the Persian Gulf, which our knowledge about is little. This study's goal is to investigate the toxicity of stonefish crude venom on mitochondria isolated from U87 cells. Methods: In the first stage, we extracted venom stonefish and then isolated mitochondria have exposed to different concentrations of venom. Finally, mitochondrial toxicity parameters (Succinate dehydrogenase (SDH) activity, Reactive oxygen species (ROS), cytochrome c release, Mitochondrial Membrane Potential (MMP), and mitochondrial swelling) have evaluated. Results: To determine mitochondrial parameters, we used 115, 230, and 460 µg/ml concentrations. The results of our study show that the venom of stonefish selectively increases upstream parameters of apoptosis such as mitochondrial swelling, cytochrome c release, MMP collapse and ROS. Conclusion: This study suggests that Pseudosynanceia melanostigma crude venom has selectively caused toxicity by increasing active mitochondrial oxygen radicals. This venom could potentially be a candidate for the treatment of glioblastoma.

In Silico Target Prediction of Overexpressed microRNAs from LPS-Challenged Zebrafish (Danio rerio) Treated with the Novel Anti-Inflammatory Peptide TnP.

miRNAs regulate gene expression post-transcriptionally in various processes, e.g., immunity, development, and diseases. Since their experimental analysis is complex, in silico target prediction is important for directing investigations. TnP is a candidate peptide for anti-inflammatory therapy, first discovered in the venom of Thalassophryne nattereri, which led to miRNAs overexpression in LPS-inflamed zebrafish post-treatment. This work aimed to predict miR-21, miR-122, miR-731, and miR-26 targets using overlapped results of DIANA microT-CDS and TargetScanFish software. This study described 513 miRNAs targets using highly specific thresholds. Using Gene Ontology over-representation analysis, we identified their main roles in regulating gene expression, neurogenesis, DNA-binding, transcription regulation, immune system process, and inflammatory response. miRNAs act in post-transcriptional regulation, but we revealed that their targets are strongly related to expression regulation at the transcriptional level, e.g., transcription factors proteins. A few predicted genes participated concomitantly in many biological processes and molecular functions, such as foxo3a, rbpjb, rxrbb, tyrobp, hes6, zic5, smad1, e2f7, and npas4a. Others were particularly involved in innate immunity regulation: il17a/f2, pik3r3b, and nlrc6. Together, these findings not only provide new insights into the miRNAs mode of action but also raise hope for TnP therapy and may direct future experimental investigations.

Deteriorating insulin resistance due to WL15 peptide from cysteine and glycine-rich protein 2 in high glucose-induced rat skeletal muscle L6 cells.

This study investigates the antioxidant and antidiabetic activity of the WL15 peptide derived from Channa striatus on regulating the antioxidant property in the rat skeletal muscle cell line (L6) and enhancing glucose uptake via glucose metabolism. Increased oxidative stress plays a major role in the development of diabetes and its complications. Strategies are needed to mitigate the oxidative stress that can reduce these pathogenic processes. Our results showed that with treatment with WL15 peptide, the reactive oxygen species significantly decreased in L6 myotubes in a dose-dependent manner, and increased antioxidant enzymes help to prevent the formation of lipid peroxidation in L6 myotubes. The cytotoxicity of WL15 is evaluated in the L6 cells and found to be non-cytotoxic at the tested concentration. Also, for the analysis of glucose uptake activity in L6 cells, the 2-(N-[7-nitrobenz-2-oxa-1,3-diazol-4-yl]amino)-2-deoxy- d -glucose assay was performed in the presence of wortmannin and genistein inhibitors. WL15 demonstrated antidiabetic activities through a dose-dependent increase in glucose uptake (64%) and glycogen storage (7.8 mM). The optimal concentration for the maximum activity was found to be 50 µM. In addition, studies of gene expression in L6 myotubes demonstrated upregulation of antioxidant genes and genes involved in the pathway of insulin signaling. In cell-based assays, WL15 peptide decreased intracellular reactive oxygen species levels and demonstrated insulin mimic activity by enhancing the primary genes involved in the insulin signaling pathway by increased glucose uptake indicating that glucose transporter type 4 (GLUT4) is regulated from the intracellular pool to the plasma membrane.

Stingray Venom Proteins: Mechanisms of Action Revealed Using a Novel Network Pharmacology Approach.

Animal venoms offer a valuable source of potent new drug leads, but their mechanisms of action are largely unknown. We therefore developed a novel network pharmacology approach based on multi-omics functional data integration to predict how stingray venom disrupts the physiological systems of target animals. We integrated 10 million transcripts from five stingray venom transcriptomes and 848,640 records from three high-content venom bioactivity datasets into a large functional data network. The network featured 216 signaling pathways, 29 of which were shared and targeted by 70 transcripts and 70 bioactivity hits. The network revealed clusters for single envenomation outcomes, such as pain, cardiotoxicity and hemorrhage. We carried out a detailed analysis of the pain cluster representing a primary envenomation symptom, revealing bibrotoxin and cholecystotoxin-like transcripts encoding pain-inducing candidate proteins in stingray venom. The cluster also suggested that such pain-inducing toxins primarily activate the inositol-3-phosphate receptor cascade, inducing intracellular calcium release. We also found strong evidence for synergistic activity among these candidates, with nerve growth factors cooperating with the most abundant translationally-controlled tumor proteins to activate pain signaling pathways. Our network pharmacology approach, here applied to stingray venom, can be used as a template for drug discovery in neglected venomous species.

Molecular Characterization and Functional Analysis of the Nattectin-like Toxin from the Venomous Fish Thalassophryne maculosa.

TmC4-47.2 is a toxin with myotoxic activity found in the venom of Thalassophryne maculosa, a venomous fish commonly found in Latin America whose envenomation produces an injury characterized by delayed neutrophil migration, production of major pro-inflammatory cytokines, and necrosis at the wound site, as well as a specific systemic immune response. However, there are few studies on the protein structure and functions associated with it. Here, the toxin was identified from the crude venom by chromatography and protein purification systems. TmC4-47.2 shows high homology with the Nattectin from Thalassophryne nattereri venom, with 6 cysteines and QPD domain for binding to galactose. We confirm its hemagglutinating and microbicide abilities independent of carbohydrate binding, supporting its classification as a nattectin-like lectin. After performing the characterization of TmC4-47.2, we verified its ability to induce an increase in the rolling and adherence of leukocytes in cremaster post-capillary venules dependent on the α5ß1 integrin. Finally, we could observe the inflammatory activity of TmC4-47.2 through the production of IL-6 and eotaxin in the peritoneal cavity with sustained recruitment of eosinophils and neutrophils up to 24 h. Together, our study characterized a nattectin-like protein from T. maculosa, pointing to its role as a molecule involved in the carbohydrate-independent agglutination response and modulation of eosinophilic and neutrophilic inflammation.

A Novel Mutant of rLj-RGD3 (rLj-112) Suppressed the Proliferation and Metastasis of B16 Cells through the EGFR Signaling Pathway.

Lj-RGD3, which contains three Arg⁻Gly⁻Asp (RGD) motifs, was first identified from the buccal glands of Lampetra japonica and has been shown to suppress the tumor progression in the previous studies. Apart from the three RGD motifs, Lj-RGD3 is also characterized by its high content of histidine in its amino acid sequence. In order to clarify whether the histidine-rich characterization of Lj-RGD3 is also associated with its anti-tumor activity, mutants were designed in which the three RGD motifs (Lj-112), or all histidines (Lj-27) or both (Lj-26) were deleted. Furthermore, a mutant (Lj-42) in which all histidines and three RGD motifs were respectively substituted with alanines and three Ala⁻Gly⁻Asp (AGD) motifs, as well as a mutant (Lj-41) in which all histidines were substituted with alanines was synthesized to avoid alterations in structure which might further cause changes in the peptides' functions. After recombination and purification, recombinant Lj-112 (rLj-112), recombinant Lj-27 (rLj-27), recombinant Lj-41 (rLj-41), and recombinant Lj-RGD3 (rLj-RGD3) exhibited anti-proliferative activity in B16 cells, respectively; while recombinant Lj-26 (rLj-26) and recombinant Lj-42 (rLj-42) did not affect the proliferation of B16 cells significantly. In addition, the anti-proliferative activity of rLj-112 in B16 cells was due to apoptosis. Typical apoptosis features were observed, including chromatin condensation, fragmented DNA, and increased levels of cleaved caspase 3/caspase 7/nuclear enzyme poly (ADP-ribose) polymerase (PARP) in B16 cells. Similar to rLj-RGD3, rLj-112 was also capable of suppressing the migration and invasion of B16 cells by disturbing the F-actin arrangement. After labeling with FITC, rLj-112 was found localized in the cytoplasm of B16 cells, which induced the internalization of epidermal growth factor receptor (EGFR), suggesting that rLj-112 might block the EGFR mediated signaling pathway. Actually, the phosphorylation level of EGFR and its downstream signal molecules including Akt, PI3K, p38, and ERK1/2 was reduced in the rLj-112 treated B16 cells. In vivo, rLj-112 also inhibited the growth, weight, and volume of the tumors in B16 xenografted C57BL/6 mice without reducing their body weight, indicating that rLj-112 might be safe and might be used as an effective anti-tumor drug in the near future.

rLj-RGD3 Suppresses the Growth of HeyA8 Cells in Nude Mice.

In the previous study, rLj-RGD3, a recombinant toxin protein which contains three RGD motifs, was reported to not only inhibit the proliferation of an ovarian cancer cell line, HeyA8 cells, by inducing apoptosis, but also block their adhesion, migration and invasion processes. However, whether rLj-RGD3 could also suppress the tumor growth in HeyA8 xenografted mice has not been reported yet. In the present study, rLj-RGD3 was intraperitoneally injected in the nude mice bearing HeyA8 tumors. Compared with the control group (normal saline), rLj-RGD3 inhibited the tumor growth significantly in the HeyA8 xenografted mice in a dose-dependent manner without affecting their body weights. Based on the H&E, Hoechst 33258 and TUNEL staining assays, as well as western blot analysis, rLj-RGD3 reduced the weight and volume of the solid tumors, probably by disturbing the tissue structure, inducing apoptosis and suppressing the FAK/PI3K/AKT pathway. Most importantly, rLj-RGD3 was found to prolong the survival days of the ovarian tumor xenografted mice, which suggested rLj-RGD3 might act as an effective and safe drug to treat ovarian cancer patients.

Inhibition of Colon Carcinoma Cell Migration Following Treatment with Purified Venom from Lesser Weever Fish (Trachinus Vipera).

BACKGROUND: Injury by the sting of Lesser weever fish (Trachinus vipera) may lead to severe pain, edema or tissue necrosis. Cellular effects of the venom are still incompletely understood. Previous observations revealed that purified Lesser weever fish venom (LWFV) induces suicidal death of erythrocytes and HCT116 human colon carcinoma cells. The present study addressed the effect of the venom on colon carcinoma cell toxicity, shape and migration both in p53+/+ and/or p53-/- conditions. METHODS: Cells were exposed to medium without or with 500 µg/ ml LWFV. Cell shape, cell area and circularity were visualized and quantified by fluorescence microscopy. Cell volume, granularity and cells toxicity were assessed via the apoptotic parameters dissipation of mitochondrial inner transmembrane potential, phosphatidylserine surface exposure and cell membrane permeabilization were measured utilizing flow cytometry. Cell migration was evaluated using wound healing assay and two-dimensional migration assay. RESULTS: LWFV treatment was followed by a marked change of cell shape and size, significant decrease of cell area and circularity, significant impairment of cell migration, as well as induction of apoptosis after long exposition. CONCLUSIONS: LWFV exposure leads to cell shrinkage, increased granularity, apoptosis and impairment of cell migration, effects presumably contributing to LWFV-induced tissue injury.

Purified Lesser weever fish venom (Trachinus vipera) induces eryptosis, apoptosis and cell cycle arrest.

Accidents caused by the sting of Trachinus vipera (known as Lesser weever fish) are relatively common in shallow waters of the Mediterranean. Symptoms after the sting vary from severe pain to edema or even tissue necrosis in some cases. Here we show that purified Lesser weever fish venom induces eryptosis, the suicidal erythrocyte death, and apoptosis of human colon carcinoma cells. The venom leads to erythrocyte shrinkage, phosphatidylserine translocation and increased intracellular Ca2+, events typical for eryptosis. According to mitochondrial staining cancer cells dyed after the activation of the intrinsic apoptotic pathway. Trachinus vipera venom further causes cell cycle arrest.

rLj-RGD3, a Novel Recombinant Toxin Protein from Lampetra japonica, Protects against Cerebral Reperfusion Injury Following Middle Cerebral Artery Occlusion Involving the Integrin-PI3K/Akt Pathway in Rats.

BACKGROUND: The RGD-toxin protein Lj-RGD3 is a naturally occurring 118 amino acid peptide that can be obtained from the salivary gland of the Lampetra japonica fish. This unique peptide contains 3 RGD (Arg-Gly-Asp) motifs in its primary structure. Lj-RGD3 is available in recombinant form (rLj-RGD3) and can be produced in large quantities using DNA recombination techniques. The pharmacology of the three RGD motif-containing peptides has not been studied. This study investigated the protective effects of rLj-RGD3, a novel polypeptide, against ischemia/reperfusion-induced damage to the brain caused by middle cerebral artery occlusion (MCAO) in a rat stroke model. We also explored the mechanism by which rLj-RGD3 acts by measuring protein and mRNA expression levels, with an emphasis on the FAK and integrin-PI3K/Akt anti-apoptosis pathways. METHODS: rLj-RGD3 was obtained from the buccal secretions of Lampetra japonica using gene recombination technology. Sprague Dawley (SD) rats were randomly divided into the following seven groups: a sham group; a vehicle-treated (VT) group; 100.0 µg·kg-1, 50.0 µg·kg-1 and 25.0 µg·kg-1 dose rLj-RGD3 groups; and two positive controls, including 1.5 mg·kg-1 Edaravone (ED) and 100.0 µg·kg-1 Eptifibatide (EP). MCAO was induced using a model consisting of 2 h of ischemia and 24 h of reperfusion. Behavioral changes were observed in the normal and operation groups after focal cerebral ischemia/reperfusion was applied. In addition, behavioral scores were evaluated at 4 and 24 h after reperfusion. Brain infarct volumes were determined based on 2,3,5-triphenyltetrazolium chloride (TTC) staining. Pathological changes in brain tissues were observed using hematoxylin and eosin (H&E) staining. Moreover, neuronal apoptosis was detected using terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling (TUNEL) assays. We determined the expression levels of focal adhesion kinase (FAK), phosphatidyl inositol 3-kinase (PI3K), protein kinase B (Akt, PKB), caspase-3 and Bcl-2 in the brain using western blot analysis and RT-PCR assays. The research protocol was approved by the Institutional Ethics Committee of Dalian Medical University. RESULTS: The behavioral scores and cerebral infarct volumes of the rLj-RGD3 groups were markedly lower at 4 and 24 h/RF. The rLj-RGD3 protein significantly ameliorated pathological changes in the brain and reduced the number of apoptotic neurons. Furthermore, the FAK and PI3K/Akt pathways were activated. rLj-RGD3 significantly increased the expression of FAK, p-FAK and Bcl-2 proteins. In contrast, caspase-3 expression was inhibited. CONCLUSION/SIGNIFICANCE: We conclude that recombinant Lampetra japonica RGD-peptide (rLj-RGD3) exerts a protective effect against cerebral ischemia/reperfusion injury in the brain. In addition, the mechanism of this protection is associated with the activation of the integrin-PI3K/Akt pathway. These results provide a theoretical foundation and an experimental basis for using RGD peptides as novel drugs for treating ischemic cerebral vascular diseases in addition to promoting the research and development of marine biotechnology drugs.

A review on the Scorpaena plumieri fish venom and its bioactive compounds

The most poisonous fish species found along the Brazilian coast is the spotted scorpionfish Scorpaena plumieri. Though hardly ever life-threatening to humans, envenomation by S. plumieri can be quite hazardous, provoking extreme pain and imposing significant socioeconomic costs, as the victims may require days to weeks to recover from their injuries. In this review we will walk the reader through the biological features that distinguish this species as well as the current epidemiological knowledge related to the envenomation and its consequences. But above all, we will discuss the challenges involved in the biochemical characterization of the S. plumieri venom and its compounds, focusing then on the successful isolation and pharmacological analysis of some of the bioactive molecules responsible for the effects observed upon envenomation as well as on experimental models. Despite the achievement of considerable progress, much remains to be done, particularly in relation to the non-proteinaceous components of the venom. Therefore, further studies are necessary in order to provide a more complete picture of the venoms chemical composition and physiological effects. Given that fish venoms remain considerably less studied when compared to terrestrial venoms, the exploration of their full potential opens a myriad of possibilities for the development of new drug leads and tools for elucidating the complex physiological processes.

In Vitro and in Vivo Anticancer Activity of Pardaxin against Proliferation and Growth of Oral Squamous Cell Carcinoma.

Pardaxin (H-GFFALIPKIISSPLFKTLLSAVGSALSSSGGQE-OH), a 33-amino-acid polypeptide, is an antimicrobial peptide (AMP) isolated from the marine fish species Pardachirus marmoratus. Pardaxin shows antibacterial and antitumor activities. However, pardaxin-induced inhibition of oral cancer and the mechanism of tumor reduction in buccal pouch carcinogenesis after pardaxin painting remain undetermined. Additionally, the toxic effects of pardaxin on normal tissue remain unclear. The present study investigated the anticancer activity of pardaxin in oral squamous cell carcinoma (OSCC) cells in the hamster buccal pouch model with or without 7,12-dimethylbenz[a]anthracene (DMBA) pretreatment. This is the first study to confirm the effects of pardaxin on normal tissue and its nontoxic effects in vivo. Cell viability assays and colony formation tests in OSCC cell lines (SCC-4) demonstrated that pardaxin reduced cell viability in a dose-dependent manner. Immunofluorescence staining of cleaved caspase-3 in SCC-4 cells revealed that expression of activated caspase-3 in SCC-4 cells significantly increased after 24-h treatment with pardaxin. Additionally, a cell cycle analysis indicated that pardaxin treatment resulted in the cell cycle arrest of SCC-4 cells in the G2/M phase, thereby limiting cell proliferation. Furthermore, pardaxin treatment substantially alleviated carcinogenesis in the DMBA-induced hamster buccal pouch model by lowering prostaglandin E2 levels. These results suggest that pardaxin is a potential marine drug for adjuvant chemotherapy for human OSCC and oral cancer.

The antimicrobial peptide pardaxin exerts potent anti-tumor activity against canine perianal gland adenoma.

Pardaxin is an antimicrobial peptide of 33 amino acids, originally isolated from marine fish. We previously demonstrated that pardaxin has anti-tumor activity against murine fibrosarcoma, both in vitro and in vivo. In this study, we examined the anti-tumor activity, toxicity profile, and maximally-tolerated dose of pardaxin treatment in dogs with different types of refractory tumor. Local injection of pardaxin resulted in a significant reduction of perianal gland adenoma growth between 28 and 38 days post-treatment. Surgical resection of canine histiocytomas revealed large areas of ulceration, suggesting that pardaxin acts like a lytic peptide. Pardaxin treatment was not associated with significant variations in blood biochemical parameters or secretion of immune-related proteins. Our findings indicate that pardaxin has strong therapeutic potential for treating perianal gland adenomas in dogs. These data justify the veterinary application of pardaxin, and also provide invaluable information for veterinary medicine and future human clinical trials.

The mechanisms by which pardaxin, a natural cationic antimicrobial peptide, targets the endoplasmic reticulum and induces c-FOS.

Pardaxin is a cationic antimicrobial peptide derived from Red Sea Moses sole. Previous studies have shown that pardaxin selectively triggers the death of cancer cells, initiating the development of a pardaxin-based cancer vaccine; however, the underlying mechanism by which pardaxin kills cancer cells has not yet been elucidated. Here, we demonstrate that this mechanism involves endoplasmic reticulum (ER) targeting and c-FOS induction. Transcriptiome analysis of pardaxin-treated HT-1080 cells revealed induction of the gene encoding c-FOS, an AP-1 transcription factor. Pardaxin mediates cell death by activating c-FOS, but not other AP-1 transcription factors. Overexpression of c-FOS caused a dramatic increase in cell death, while knockdown of c-FOS induced pardaxin resistance; such effects were observed in both an in vitro cell model and an in vivo xenograft tumor model. Treatment with pardaxin also increased the level of calcium, and blockage of cellular calcium signaling disrupted pardaxin-induced cell death. Immunocytochemistry was used to demonstrate targeting of pardaxin to the endoplasmic reticulum, but not to the Golgi apparatus or mitochondria. Importantly, pardaxin treatment or c-FOS overexpression induced cell death in diverse cancer cell lines, indicating that pardaxin and c-FOS may possess therapeutic potential for use in cancer treatment.

Proteomic analysis reveals that pardaxin triggers apoptotic signaling pathways in human cervical carcinoma HeLa cells: cross talk among the UPR, c-Jun and ROS.

Pardaxin, an antimicrobial peptide secreted by the Red Sea flatfish Pardachirus marmoratus, inhibits proliferation and induces apoptosis of human cancer cell lines. However, the underlying molecular mechanisms are only partially understood at present. In this study, we used proteomic approaches and network reconstruction to clarify the mechanism of pardaxin-induced apoptosis in human cervical carcinoma HeLa cells. We identified that pardaxin-regulated proteins predominantly function in the unfolded protein response, oxidative stress and cytoskeletal distribution. Molecular examination of signal transduction and cellular localization demonstrated that the activator protein-1 transcription factor was activated, which eventually caused apoptosis via both caspase- and apoptosis-inducing factor-dependent pathways. Scavenging of reactive oxygen species (ROS) alleviated c-Jun activation, and small interfering RNA knockdown of c-Jun abrogated pardaxin-induced caspase activation and cell death, thereby implicating ROS and c-Jun in pardaxin-induced apoptosis signaling. In summary, this study provides the first protein-interacting network maps and novel insights into the biological responses and potential toxicity of pardaxin.

Truncated antimicrobial peptides from marine organisms retain anticancer activity and antibacterial activity against multidrug-resistant Staphylococcus aureus.

Antimicrobial peptides (AMPs) were recently determined to be potential candidates for treating drug-resistant bacterial infections. The aim of this study was to develop shorter AMP fragments that combine maximal bactericidal effect with minimal synthesis cost. We first synthesized a series of truncated forms of AMPs (anti-lipopolysaccharide factor from shrimp, epinecidin from grouper, and pardaxin from Pardachirus marmoratus). The minimum inhibitory concentrations (MICs) of modified AMPs against ten bacterial species were determined. We also examined the synergy between peptide and non-peptide antibiotics. In addition, we measured the inhibitory rate of cancer cells treated with AMPs by MTS assay. We found that two modified antibacterial peptides (epinecidin-8 and pardaxin-6) had a broad range of action against both gram-positive and gram-negative bacteria. Furthermore, epinecidin and pardaxin were demonstrated to have high antibacterial and anticancer activities, and both AMPs resulted in a significant synergistic improvement in the potencies of streptomycin and kanamycin against methicillin-resistant Staphylococcus aureus. Neither AMP induced significant hemolysis at their MICs. In addition, both AMPs inhibited human epithelial carcinoma (HeLa) and fibrosarcoma (HT-1080) cell growth. The functions of these truncated AMPs were similar to those of their full-length equivalents. In conclusion, we have successfully identified shorter, inexpensive fragments with maximal bactericidal activity. This study also provides an excellent basis for the investigation of potential synergies between peptide and non-peptide antibiotics, for a broad range of antimicrobial and anticancer activities.

Pardaxin, a fish antimicrobial peptide, exhibits antitumor activity toward murine fibrosarcoma in vitro and in vivo.

The antitumor activity of pardaxin, a fish antimicrobial peptide, has not been previously examined in in vitro and in vivo systems for treating murine fibrosarcoma. In this study, the antitumor activity of synthetic pardaxin was tested using murine MN-11 tumor cells as the study model. We show that pardaxin inhibits the proliferation of MN-11 cells and reduces colony formation in a soft agar assay. Transmission electron microscopy (TEM) showed that pardaxin altered the membrane structure similar to what a lytic peptide does, and also produced apoptotic features, such as hollow mitochondria, nuclear condensation, and disrupted cell membranes. A qRT-PCR and ELISA showed that pardaxin induced apoptosis, activated caspase-7 and interleukin (IL)-7r, and downregulated caspase-9, ATF 3, SOCS3, STAT3, cathelicidin, p65, and interferon (IFN)-γ suggesting that pardaxin induces apoptosis through the death receptor/nuclear factor (NF)-κB signaling pathway after 14 days of treatment in tumor-bearing mice. An antitumor effect was observed when pardaxin (25 mg/kg; 0.5 mg/day) was used to treat mice for 14 days, which caused significant inhibition of MN-11 cell growth in mice. Overall, these results indicate that pardaxin has the potential to be a novel therapeutic agent to treat fibrosarcomas.

Pardaxin, an antimicrobial peptide, triggers caspase-dependent and ROS-mediated apoptosis in HT-1080 cells.

Pardaxin is an antimicrobial peptide (AMP) that was first isolated from secretions of the Red Sea Moses sole. The role of pardaxin in inducing apoptosis for preventing cancer has not yet been investigated. In the present study, we examined the antitumor activity of pardaxin against human fibrosarcoma HT-1080 cells; pardaxin inhibited cell proliferation by inducing apoptosis, as demonstrated by an increase in the externalization of plasma membrane phosphatidylserine and the presence of chromatin condensation. Additionally, pardaxin-treated cells showed elevation of caspase-3/7 activities, disruption of the mitochondrial membrane potential, and accumulation of reactive oxygen species (ROS) production. Inhibition of ROS production and caspase-3/7 activities reduced pardaxin-induced effects. Taken together, these findings suggest that pardaxin may be a potential anticancer agent for selectively inducing apoptosis in cancer cells.