Peptides (v.78, #C)

Nesfatin-1 improves oxidative skin injury in normoglycemic or hyperglycemic rats by Ali Solmaz; Elif Bahadır; Osman B. Gülçiçek; Hakan Yiğitbaş; Atilla Çelik; Ayça Karagöz; Derya Özsavcı; Serap Şirvancı; Berrak Ç. Yeğen (1-10).
Hyperglycemia is one of the major causes of suppressed angiogenesis and impaired wound healing leading to chronic wounds. Nesfatin-1 a novel peptide was reported to have antioxidant and anti-apoptotic properties. This study is aimed to investigate the potential healing-promoting effects of nesfatin-1 in non-diabetic or diabetic rats with surgical wounds. In male Sprague-Dawley rats, hyperglycemia was induced by intraperitoneal (ip) injection of streptozotocin (55 mg/kg). Under anesthesia, dorsum skin tissues of normoglycemic (n  = 16) and hyperglycemic rats were excised (2 × 2 cm, full-thickness), while control rats (n  = 16) had neither hyperglycemia nor wounds. Half of the rats in each group were treated ip with saline, while the others were treated with nesfatin-1 (2 μg/kg/day) for 3 days until they were decapitated. Plasma interleukin-1-beta (IL-1β), transforming growth factor-beta (TGF-β-1), IL–6 levels, and dermal tissue malondialdehyde (MDA), glutathione (GSH) levels, myeloperoxidase (MPO) and caspase-3 activity were measured. For histological examination, paraffin sections were stained with hematoxylin-eosin or Masson’s trichrome and immunohistochemistry for vascular endothelial growth factor (VEGF) was applied. ANOVA and Student’s t-tests were used for statistical analysis. Compared to control rats, skin MPO activity, MDA and caspase-3 levels were increased similarly in saline-treated normo- and hyperglycemic rats. Nesfatin-1 depressed MDA, caspase-3, MPO activity and IL-1β with concomitant elevations in dermal GSH and plasma TGF-β-1 levels. Histopathological examination revealed regeneration of epidermis, regular arrangement of collagen fibers in the dermis and a decrease in VEGF immunoreactivity in the epidermal keratinocytes of nesfatin-1-treated groups. Nesfatin-1 improved surgical wound healing in both normo- and hyperglycemic rats via the suppression of neutrophil recruitment, apoptosis and VEGF activation.
Keywords: Nesfatin-1; Vascular endothelial growth factor (VEGF); Hyperglycaemia; Surgical wound; Neutrophil infiltration; Oxidative injury;

Inhibition of malaria parasite Plasmodium falciparum development by crotamine, a cell penetrating peptide from the snake venom by S. El Chamy Maluf; C. Dal Mas; E.B. Oliveira; P.M. Melo; A.K. Carmona; M.L. Gazarini; M.A.F. Hayashi (11-16).
Schematic figure showing the preferential affinity of crotamine for P. falciparum infected erythrocytes, which may have increased negative charge exposure compared to the liquid net neutral surface of uninfected erythrocytes due to the extensive host cell remodeling mediated by parasites, for instance maure’s clefts, plasma membrane knobs complexes and TVN (tubovesicular network). The localization of crotamine in parasitophorous vacuole (PV), as well as in acidic digestive vacuole (DV) and nucleus are also indicated.Display OmittedWe show here that crotamine, a polypeptide from the South American rattlesnake venom with cell penetrating and selective anti-fungal and anti-tumoral properties, presents a potent anti-plasmodial activity in culture. Crotamine inhibits the development of the Plasmodium falciparum parasites in a dose-dependent manner [IC50 value of 1.87 μM], and confocal microscopy analysis showed a selective internalization of fluorescent-labeled crotamine into P. falciparum infected erythrocytes, with no detectable fluorescence in uninfected healthy erythrocytes. In addition, similarly to the crotamine cytotoxic effects, the mechanism underlying the anti-plasmodial activity may involve the disruption of parasite acidic compartments H+ homeostasis. In fact, crotamine promoted a reduction of parasites organelle fluorescence loaded with the lysosomotropic fluorochrome acridine orange, in the same way as previously observed mammalian tumoral cells. Taken together, we show for the first time crotamine not only compromised the metabolism of the P. falciparum, but this toxin also inhibited the parasite growth. Therefore, we suggest this snake polypeptide as a promising lead molecule for the development of potential new molecules, namely peptidomimetics, with selectivity for infected erythrocytes and ability to inhibit the malaria infection by its natural affinity for acid vesicles.
Keywords: Plasmodium; Parasites; Crotamine; Antimalarial; Acidic compartments; Peptide trafficking;

The functional interaction between abaecin and pore-forming peptides indicates a general mechanism of antibacterial potentiation by Mohammad Rahnamaeian; Małgorzata Cytryńska; Agnieszka Zdybicka-Barabas; Andreas Vilcinskas (17-23).
Long-chain proline-rich antimicrobial peptides such as bumblebee abaecin show minimal activity against Gram-negative bacteria despite binding efficiently to specific intracellular targets. We recently reported that bumblebee abaecin interacts with Escherichia coli DnaK but shows negligible antibacterial activity unless it is combined with sublethal doses of the pore-forming peptide hymenoptaecin. These two bumblebee peptides are co-expressed in vivo in response to a bacterial challenge. Here we investigated whether abaecin interacts similarly with pore-forming peptides from other organisms by replacing hymenoptaecin with sublethal concentrations of cecropin A (0.3 μM) or stomoxyn (0.05 μM). We found that abaecin increased the membrane permeabilization effects of both peptides, confirming that it can reduce the minimal inhibitory concentrations of pore-forming peptides from other species. We also used atomic force microscopy to show that 20 μM abaecin combined with sublethal concentrations of cecropin A or stomoxyn causes profound structural changes to the bacterial cell surface. Our data indicate that the potentiating functional interaction between abaecin and pore-forming peptides is not restricted to specific co-expressed peptides from the same species but is likely to be a general mechanism. Combination therapies based on diverse insect-derived peptides could therefore be used to tackle bacteria that are recalcitrant to current antibiotics.
Keywords: Proline-rich peptides; Combination therapy; Membrane permeabilization; Therapeutic leads; Atomic force microscopy; β-galactosidase assay;

The antimicrobial peptide, LP5, is a lysine-peptoid hybrid, with antimicrobial activity against clinically relevant bacteria. Here, we investigated how various environmental conditions affect the antimicrobial activity of LP5 against Staphylococcus aureus (S. aureus). We found that LP5 maintained activity under host physiological conditions of NaCl, MgCl2 and pH. However, when exposed to serum, LP5 lost activity. Furthermore, when increasing NaCl concentration and lowering pH, the peptide showed reduces activity. When investigating the tolerance mechanisms of S. aureus toward antimicrobial peptides, we found that LP5 was protease resistant. However, the dltA and vraF genes, involved in reducing the net anionic charge of the bacterial cell envelope and sensing of antimicrobial peptides, respectively, played a role in the tolerance of S. aureus against LP5. In addition, the exposure of S. aureus to sub-inhibitory concentrations of LP5 affected the expression of the major virulence factors of S. aureus, revealing a potential as anti-virulence compound. Thus, these results show how environmental factors affect the peptide efficiency and further add to the knowledge on how the peptide affects S. aureus, which is crucial information for designing new peptides for optimizing antimicrobial therapy.
Keywords: Antimicrobial peptide; Lysine-peptoid hybrid; LP5 activity; LP5 stability; S. aureus; Anti-virulence factor;

VIP and PACAP analogs regulate therapeutic targets in high-risk neuroblastoma cells by Madryssa de Boisvilliers; Florian Perrin; Salima Hebache; Annie-Claire Balandre; Souheyla Bensalma; Agnès Garnier; David Vaudry; Alain Fournier; Franck Festy; Jean-Marc Muller; Corinne Chadéneau (30-41).
Neuroblastoma (NB) is a pediatric cancer. New therapies for high-risk NB aim to induce cell differentiation and to inhibit MYCN and ALK signaling in NB. The vasoactive intestinal peptide (VIP) and the pituitary adenylate cyclase-activating polypeptide (PACAP) are 2 related neuropeptides sharing common receptors. The level of VIP increases with NB differentiation. Here, the effects of VIP and PACAP analogs developed for therapeutic use were studied in MYCN-amplified NB SK-N-DZ and IMR-32 cells and in Kelly cells that in addition present the F1174L ALK mutation. As previously reported by our group in IMR-32 cells, VIP induced neuritogenesis in SK-N-DZ and Kelly cells and reduced MYCN expression in Kelly but not in SK-N-DZ cells. VIP decreased AKT activity in the ALK-mutated Kelly cells. These effects were PKA-dependent. IMR-32, SK-NDZ and Kelly cells expressed the genes encoding the 3 subtypes of VIP and PACAP receptors, VPAC1, VPAC2 and PAC1. In parallel to its effect on MYCN expression, VIP inhibited invasion in IMR-32 and Kelly cells. Among the 3 PACAP analogs tested, [Hyp2]PACAP-27 showed higher efficiency than VIP in Kelly cells. These results indicate that VIP and PACAP analogs act on molecular and cellular processes that could reduce aggressiveness of high-risk NB.
Keywords: MYCN; ALK; AKT; PKA; Invasion; Differentiation;

Secretoneurin (SN) is a conserved peptide derived by proteolytic processing from the middle domain of the ∼600 amino acid precursor secretogranin-II (SgII). Secretoneurin is widely distributed in secretory granules of endocrine cells and neurons and has important roles in reproduction as it stimulates luteinizing hormone release from the pituitary. A potential new role of SN in goldfish feeding is the subject of this study. Firstly, we established that acute (26 h; p  < 0.0001) and short-term (72 h; p  = 0.016) fasting increased SgIIa precursor mRNA levels 1.25-fold in the telencephalon, implicating SN in the control of feeding. Secondly, we determined that intracerebroventricular injections of the type A SN (SNa; 0.2 and 1 ng/g BW) increased food intake and locomotor behavior by 60 min. Fish injected with the lower and higher doses of SNa (0.2 and 1 ng/g) respectively exhibited significant 1.77- and 2.58-fold higher food intake (p  < 0.0001) than the saline-injected control fish. Locomotor behavior was increased by 1.35- and 2.26-fold for 0.2 ng/g SNa (p  = 0.0001) and 1 ng/g SNa (p  < 0.0001), respectively. Injection of 1 ng/g SNa increased mRNA levels of hypothalamic neuropeptide Y 1.36-fold (p  = 0.038) and decreased hypothalamic cocaine-and amphetamine-regulated transcript by 33% (p  = 0.01) at 2 h and 5 h post-injection, respectively. These data suggest interactions of SNa with stimulatory and inhibitory pathways of food intake control in fish.
Keywords: Feeding; Locomotion; Secretogranin-II; Secretoneurin; Fish;

Naturally-occurring TGR5 agonists modulating glucagon-like peptide-1 biosynthesis and secretion by Laila Jafri; Samreen Saleem; Danielle Calderwood; Anna Gillespie; Bushra Mirza; Brian D. Green (51-58).
Selective GLP-1 secretagogues represent a novel potential therapy for type 2 diabetes mellitus. This study examined the GLP-1 secretory activity of the ethnomedicinal plant, Fagonia cretica, which is postulated to possess anti-diabetic activity. After extraction and fractionation extracts and purified compounds were tested for GLP-1 and GIP secretory activity in pGIP/neo STC-1 cells. Intracellular levels of incretin hormones and their gene expression were also determined. Crude F. cretica extracts stimulated both GLP-1 and GIP secretion, increased cellular hormone content, and upregulated gene expression of proglucagon, GIP and prohormone convertase. However, ethyl acetate partitioning significantly enriched GLP-1 secretory activity and this fraction underwent bioactivity-guided fractionation. Three isolated compounds were potent and selective GLP-1 secretagogues: quinovic acid (QA) and two QA derivatives, QA-3β-O-β-d-glycopyranoside and QA-3β-O-β-d-glucopyranosyl-(28 → 1)-β-d-glucopyranosyl ester. All QA compounds activated the TGR5 receptor and increased intracellular incretin levels and gene expression. QA derivatives were more potent GLP-1 secretagogues than QA. This is the first time that QA and its naturally-occurring derivatives have been shown to activate TGR5 and stimulate GLP-1 secretion. These data provide a plausible mechanism for the ethnomedicinal use of F. cretica and may assist in the ongoing development of selective GLP-1 agonists.
Keywords: GLP-1; GIP; Incretin; Secretagogue; TGR5;

Metabolism of peptide YY 3–36 in Göttingen mini-pig and rhesus monkey by Jørgen Olsen; Jacob Kofoed; Søren Østergaard; Birgitte S. Wulff; Flemming S. Nielsen; Rasmus Jorgensen (59-67).
Peptide YY 3–36-amide (PYY3–36) is a peptide hormone, which is known to decrease appetite and food-intake by activation of the Y2 receptor. The current studies were designed to identify the metabolites of PYY3–36 in mini-pig and rhesus monkey. Plasma samples were analyzed by high resolution LC–MS (and MS/MS) in order to unambiguously identify the metabolites of PYY3–36. In summary, the metabolism of PYY3–36 was similar in mini-pig and rhesus monkey. Several metabolites were identified and PYY3–34 was identified at the highest levels in plasma. In addition, mini-pigs were also dosed with PYY1–36-amide, PYY3–35, PYY3–34 and [N-methyl 34Q]-PYY3–36-amide in order to investigate the mechanisms by which PYY was metabolized. PYY3–35 was rapidly converted to PYY3–34 whereas dosing of PYY3–34 to mini-pigs only showed circulating degradation products at low levels, i.e., PYY3–34 was metabolically more stable than PYY3–36 and PYY3–35. [N-methyl 34Q]-PYY3–36-amide was hypothesized to be stable toward cleavage between 34Q and 35R and after i.v. administration to mini-pigs, one major cleavage product was identified as [N-methyl 34Q]-PYY3–35. Overall, this showed that cleavage between 35R and 36Y was possible as well as between 34Q and 35R (as shown for PYY3–35), which indicated that metabolism of PYY3–36 to PYY3–34 may be a two-step process. PYY1–36 was also dosed to mini-pigs, which showed that PYY1–36 was metabolized in the C-terminal as PYY3–36. The overall degradation pattern of PYY1–36 was more complex due to the simultaneous enzymatic degradation in the N-terminal to form PYY2–34/36 and PYY3–34/36. In vitro incubations with heparin stabilized plasma showed that PYY3–36 was degraded with a half-life of 175 min, whereas incubations with PYY3–35 (half-life of 6 min) showed a rapid formation of PYY3–34. In conclusion, the present studies showed that PYY3–36 underwent enzymatic degradation in the C-terminal part and that the major circulating metabolite was PYY3–34. Furthermore, it may be a sequential two-step process leading to the formation of PYY3–35 and subsequently the metabolically more stable PYY3–34.
Keywords: PYY3–36; In vivo metabolism; Liquid chromatography mass spectrometry (LC–MS);

Effect of secondary anchor amino acid substitutions on the immunogenic properties of an HLA-A*0201-restricted T cell epitope derived from the Trypanosoma cruzi KMP-11 protein by Paola Lasso; Constanza Cárdenas; Fanny Guzmán; Fernando Rosas; María Carmen Thomas; Manuel Carlos López; John Mario González; Adriana Cuéllar; Josep Maria Campanera; F. Javier Luque; Concepción Judith Puerta (68-76).
The TcTLE peptide (TLEEFSAKL) is a CD8+ T cell HLA-A*0201-restricted epitope derived from the Trypanosoma cruzi KMP-11 protein that is efficiently processed, presented and recognized by CD8+ T cells from chagasic patients. Since the immunogenic properties of wild-type epitopes may be enhanced by suitable substitutions in secondary anchor residues, we have studied the effect of introducing specific mutations at position 3, 6 and 7 of the TcTLE peptide. Mutations (E3L, S6V and A7F) were chosen on the basis of in silico predictions and in vitro assays were performed to determine the TcTLE-modified peptide binding capacity to the HLA-A*0201 molecule. In addition, the functional activity of peptide-specific CD8+ T cells in HLA-A2+ chagasic patients was also interrogated. In contrast to bioinformatics predictions, the TcTLE-modified peptide was found to have lower binding affinity and stability than the original peptide. Nevertheless, CD8+ T cells from chronic chagasic patients recognized the TcTLE-modified peptide producing TNF-α and INF-γ and expressing CD107a/b, though in less extension than the response triggered by the original peptide. Overall, although the amino acids at positions 3, 6 and 7 of TcTLE are critical for the peptide affinity, they have a limited effect on the immunogenic properties of the TcTLE epitope.
Keywords: CD8+ T cells; Chagas disease; Peptides; MHC class I;

A chirality change in XPC- and Sfi1-derived peptides affects their affinity for centrin by Dora Grecu; Victor Paul Raj Irudayaraj; Juan Martinez-Sanz; Jean-Maurice Mallet; Liliane Assairi (77-86).
The Ca2+-binding protein centrin binds to a hydrophobic motif (W1xxL4xxxL8) included in the sequence of several cellular targets: XPC (xeroderma pigmentosum group C protein), Sfi1 (suppressor of fermentation-induced loss of stress resistance protein1), and Sac3 [the central component of the transcription and mRNA export (TREX-2) complex]. However, centrin binding occurs in a reversed orientation (L8xxxL4xxW1) for Sfi1 and Sac3 compared with XPC. Because d-peptides have been investigated for future therapeutic use, we analyzed their centrin-binding properties. Their affinity for centrin was measured using isothermal titration calorimetry. The chirality change in the target-derived peptides affected their ability to bind centrin in a specific manner depending on the sequence orientation of the centrin-binding motif. In contrast to l-XPC-P10, d-XPC-P10 bound C-HsCen1 in a Ca2+-dependent manner and to a lesser extent. d-XPC-P10 exhibited a reduced affinity for C-HsCen1 (K a  = 0.064 × 106  M−1) by a factor of 2000 compared with l-XPC-P10 (K a  = 132 × 106  M−1). d-peptides have a lower affinity than l-peptides for centrin, and the strength of this affinity depends on the sequence orientation of the target-derived peptides. The residual affinity observed for d-XPC suggests that the use of d-peptides represents a promising strategy for inhibiting centrin binding to its targets.
Keywords: d-Peptides; Retro-inverso peptides; Centrin; XPC; Sfi1; Isothermal titration calorimetry; Circular dichroism;

The influence of intracellular angiotensin II on the regulation of potassium current and membrane potential of smooth muscle cells of mesenteric arteries and its relevance for the regulation of vascular tone was reviewed. The presence of components of the renin angiotensin system (RAS) in different cells of the cardiovascular system, was discussed including their presence in the nuclei and mitochondria. Emphasis was given to the opposite effects of intracellular and extracellular angiotensin II (Ang II) on the regulation of potassium current, membrane potential and contractility of vascular resistance vessels and its implication to vascular physiology and pathology and the possible role of epigenetic factors on the expression of angiotensin II (Ang II) and renin in vascular resistance vessels as well as its possible pathophysiological role in hypertension and other cardiovascular diseases.
Keywords: Intracellular; Extracellular; Angiotensin II; Potassium current; Mesenteric artery myocyte; Rat;

Glucagon-like peptide-1 regulates calcium homeostasis and electrophysiological activities of HL-1 cardiomyocytes by Jen-Hung Huang; Yao-Chang Chen; Ting-I Lee; Yu-Hsun Kao; Tze-Fan Chazo; Shih-Ann Chen; Yi-Jen Chen (91-98).
Glucagon like-peptide-1 (GLP-1) is an incretin hormone with antidiabetic effects through stimulating insulin secretion, β cell neogenesis, satiety sensation, and inhibiting glucagon secretion. Administration of GLP-1 provides cardioprotective effects through attenuating cardiac inflammation and insulin resistance. GLP-1 also modulates the heart rate and systolic pressure, which suggests that GLP-1 may have cardiac electrical effects. Therefore, the purposes of this study were to evaluate whether GLP-1 has direct cardiac effects and identify the underlying mechanisms. Patch clamp, confocal microscopy with Fluo-3 fluorescence, and Western blot analyses were used to evaluate the electrophysiological characteristics, calcium homeostasis, and calcium regulatory proteins in HL-1 atrial myocytes with and without GLP-1 (1 and 10 nM) incubation for 24 h. GLP-1 (1 and 10 nM) and control cells had similar action potential durations. However, GLP-1 at 10 nM significantly increased calcium transients and sarcoplasmic reticular Ca2+ contents. Compared to the control, GLP-1 (10 nM)—treated cells significantly decreased phosphorylation of the ryanodine receptor at S2814 and total phospholamban, but there were similar protein levels of sarcoplasmic reticular Ca2+-ATPase and the sodium–calcium exchanger. Moreover, exendin (9–39) amide (a GLP-1 receptor antagonist, 10 nM) attenuated GLP-1-mediated effects on total SR content and phosphorylated ryanodine receptor S2814. This study demonstrates GLP-1 may regulate HL-1 cell arrhythmogenesis through modulating calcium handling proteins.
Keywords: Atrial arrhythmogenesis; Calcium handling protein; Glucagon like-peptide-1; Ryanodine receptor;

Display OmittedIn a previous report, seven Cry1Ab-resistant strains were identified in the silkworm, Bombyx mori; these strains were shown to have a tyrosine insertion at position 234 in extracellular loop 2 of the ABC transporter C2 (BmABCC2). This insertion was confirmed to destroy the receptor function of BmABCC2 and confer the strains resistance against Cry1Ab and Cry1Ac. However, these strains were susceptible to Cry1Aa. In this report, we examined the mechanisms of the loss of receptor function of the transporter by expressing mutations in Sf9 cells. After replacement of one or two of the five amino acid residues in loop 2 of the susceptible BmABCC2 gene [BmABCC2_S] with alanine, cells still showed susceptibility, retaining the receptor function. Five mutants with single amino acid insertions at position 234 in BmABCC2 were also generated, resulting in loop 2 having six amino acids, which corresponds to replacing the tyrosine insertion in the resistant BmABCC2 gene [BmABCC2_R(+234Y)] with another amino acid. All five mutants exhibited loss of function against Cry1Ab and Cry1Ac. These results suggest that the amino acid sequence in loop 2 is less important than the loop size (five vs. six amino acids) or loop structure for Cry1Ab and Cry1Ac activity. Several domain-swapped mutant toxins were then generated among Cry1Aa, Cry1Ab, and Cry1Ac, which are composed of three domains. Swapped mutants containing domain II of Cry1Ab or Cry1Ac did not kill Sf9 cells expressing BmABCC2_R(+234Y), suggesting that domain II of the Cry toxin is related to the interaction with the receptor function of BmABCC2. This also suggests that different reactions against Bt-toxins in some B. mori strains, that is, Cry1Ab resistance or Cry1Aa susceptibility, are attributable to structural differences in domain II of Cry1A toxins.
Keywords: Bacillus thuringiensis; ABC transporter C2; BmABCC2; Receptor; Cry1A; Domain II; Bombyx mori;

Defensin γ-thionin from Capsicum chinense has immunomodulatory effects on bovine mammary epithelial cells during Staphylococcus aureus internalization by Violeta Díaz-Murillo; Ivan Medina-Estrada; Joel E. López-Meza; Alejandra Ochoa-Zarzosa (109-118).
β-Defensins are members of the antimicrobial peptide superfamily that are produced in various species from different kingdoms, including plants. Plant defensins exhibit primarily antifungal activities, unlike those from animals that exhibit a broad-spectrum antimicrobial action. Recently, immunomodulatory roles of mammal β-defensins have been observed to regulate inflammation and activate the immune system. Similar roles for plant β-defensins remain unknown. In addition, the regulation of the immune system by mammalian β-defensins has been studied in humans and mice models, particularly in immune cells, but few studies have investigated these peptides in epithelial cells, which are in intimate contact with pathogens. The aim of this work was to evaluate the effect of the chemically synthesized β-defensin γ-thionin from Capsicum chinense on the innate immune response of bovine mammary epithelial cells (bMECs) infected with Staphylococcus aureus, the primary pathogen responsible for bovine mastitis, which is capable of living within bMECs. Our results indicate that γ-thionin at 0.1 μg/ml was able to reduce the internalization of S. aureus into bMECs (∼50%), and it also modulates the innate immune response of these cells by inducing the mRNA expression (∼5-fold) and membrane abundance (∼3-fold) of Toll-like receptor 2 (TLR2), as well as by inducing genes coding for the pro-inflammatory cytokines TNF-α and IL-1β (∼14 and 8-fold, respectively) before and after the bacterial infection. γ-Thionin also induces the expression of the mRNA of anti-inflammatory cytokine IL-10 (∼12-fold). Interestingly, the reduction in bacterial internalization coincides with the production of other antimicrobial products by bMECs, such as NO before infection, and the secretion into the medium of the endogenous antimicrobial peptide DEFB1 after infection. The results from this work support the potential use of β-defensins from plants as immunomodulators of the mammalian innate immune response.
Keywords: Defensins; Immunomodulation; Staphylococcus aureus; Innate immune response; Bovine mammary epithelium; Antimicrobial peptides;