Esculentin-2PLa, a frog skin antimicrobial peptide, causes necrotic cell death in breast cancer cell lines

Background: Antimicrobial peptides are molecules which play important roles in the natural immune system from insects to mammals. These peptides show significant differences in terms of amino acid sequence and secondary structure. The net charges of these peptides vary between +2 to +9 in neutral pH and their cationic characters based on the basic amino acids such as arginine and lysine. On the other hand, these peptides are short peptides composed of 5−40 amino acids and 30% or more of these amino acids have hydrophobic character. Beside bacteria killing and immunomodulatory effects, these peptides have gained attention in recent years due to their anticancer activities. In this study we aimed to research the anticancer activity of Esculentin-2PLa on breast cancer cell lines, MCF-7 and MDA-MB 231. Esculentin-2PLa is a 37-mer peptide that is firstly defined in the skin secretion of Rana palustris and the net charge and hydrophobicity is +5 and 48%, respectively. Material and Methods: Estrogen receptor (ER) positive cell line, MCF-7 and ER(−) cell line, MDA-MB 231 were used in this study. Antitumor activity was determined with cell proliferation assay and cell cytotoxicity assay after treatment with 0–200 mg/ml Esculentin-2PLa for 24 hours. The cell death mechanism was investigated by measuring activity of caspase-3 and counting stained the apoptotic and necrotic cells with acridine orange-ethidium bromide. The cell morphology after the peptide treatment was observed by scanning electron microscopy. In addition, possible electrostatic interactions between cell membranes and the peptide were researched by solid-phase heparan sulfate and chondroitin sulfate binding assay and by measuring the affinity of peptide to cancer cell membranes. Results: Esculentin-2PLa reduced cell viability in a dose dependent manner in the two cell lines and MDA-MB 231 cell line was more sensitive. Beside this, the peptide caused the release of lactate dehydrogenase (LDH) from the cells. In addition, necrotic cell death was detected in both cell lines by observing membrane pores with scanning electrone microscope and this finding was consistent with the increase in the number of cell nuclei stained with etidium bromide. Moreover, Esculentin-2PLa attached to heparan sulfate and chondroitin sulfate proteoglycans which are abundant on cancer cell membranes in a dose dependent manner. Furthermore, this peptide has a high affinity to cancer cell membranes via electrostatic interaction. Conclusions: Overall, these results suggest that Esculentin-2PLa induced cell death through membranolytic effects and its anticancer activity may arise from its cationic and hydrophobic characters which facilitate the interaction with cancer cell membranes. Our findings indicate that Esculentin-2PLa has the potential for development as anticancer agent. No conflict of interest

Exendin-4 partly ameliorates - hyperglycemia-mediated tissue damage in lungs of streptozotocin-induced diabetic mice

Glucagon-like peptide-1 (GLP-1) stimulates insulin secretion, - plays anti-inflammatory role in atherosclerosis, and has surfactant-releasing effects in lungs. GLP-1 analogues are used in diabetes therapy. This is the first study to investigate the effects of exendin-4, a GLP-1 receptor agonist, on lung injury in diabetic mice. BALB/c male mice were divided into four groups. The first group was given only citrate buffer, the second group was given only exendin-4, the third group was given only streptozotocin (STZ), and the fourth group was given both exendin-4 and STZ. Exendin-4 (3 mu g/kg) was administered daily by subcutaneous injection for 30 days after mice were rendered diabetic with a single dose of STZ (200 mg/kg). Structural alterations, oxidative stress, apoptosis, insulin signaling and expressions of prosurfactant-C, alpha-smooth muscle actin, collagen-I and fibronectin were evaluated in lung tissue. Diabetic mice lungs were characterized by induced oxidative stress, apoptosis, edema, and cell proliferation. They had honeycomb-like alveoli, thicker alveolar walls, and hypertrophic pneumocytes. Although exendin-4 treatment improved pulmonary edema, apoptosis, oxidative stress, and lung injury, it led to the disrupted insulin signaling and interstitial collagen accumulation in the lungs of diabetic mice. Exendin-4 ameliorates hyperglycemia-mediated lung damage by reducing glucose, - oxidative stress and stimulating cell proliferation. However, exendin-4 led to increased lung injury partly by reducing insulin signaling - and collagen accumulation around pulmonary vasculature in diabetic mice