Peptides (v.40, #C)

Characterization of thymosin β4 in mammals’ saliva by Patrícia de Sousa-Pereira; Joana Abrantes; Bruno Colaço; Massimo Castagnola; Francisco Amado; Pedro J. Esteves; Rui Vitorino (1-7).
► Tβ4 amino acid sequences showed a high degree of conservation between species. ► Tβ4 was identified by MS for the first time in saliva from dog, horse and bovine. ► Tβ4 amino acid conservation suggest an important role of Tβ4 in the homeostasis of the mammalian oral cavity.Thymosin β4 (Tβ4) is a low molecular weight peptide found in several mammalian tissues and is known mainly by its ability to bind cytoskeletal actin, influencing cell migration and differentiation, and promoting tissue repair. Considering the functional role of this peptide, the main goal of this work was to characterize Tβ4 in mammals’ saliva by using evolutionary and proteomic tools. For this, mammalian Tβ4 sequences were retrieved from NCBI, SwissProt and Ensembl databases. The alignment of Tβ4 amino acid sequences showed a high degree of conservation between species. The gene seems to be evolving under negative selection as indicated by a dN/dS ratio of 0.05. Whole saliva was collected from dog, human, rabbit, cow, horse and sheep and the salivary peptides were isolated through filtration and analyzed by LC–MS/MS. Spectra was processed against the database constructed with the retrieved Tβ4 sequences. For the first time, the identification of this peptide was achieved in rat, dog, horse and bovine saliva. Detection in these mammal species and its amino acid conservation suggest an important role of Tβ4 in the homeostasis of the mammalian oral cavity.
Keywords: Saliva; Evolution; Phylogeny; Peptidomics; Proteomics;

► Visfatin genetic polymorphism is associated with plasma visfatin level. ► Visfatin genetic polymorphism is associated with plasma C-reactive protein level. ► Plasma visfatin level is highly correlated with plasma C-reactive protein level. ► Visfatin genetic polymorphism may be involved in the inflammatory process of TBI.Visfatin is a newly identified pro-inflammatory adipokine and a genetic polymorphism −1535 C > T located in the visfatin gene promoter has been suggested to be associated with the regulation of visfatin expression in some inflammatory illness. However, there were some conflicting results regarding whether this variant is functional or not. This study aimed to examine the relations of the −1535 C > T single nucleotide polymorphism (SNP) of visfatin gene to the plasma visfatin and C-reactive protein concentrations in traumatic brain injury (TBI). 318 Chinese Han patients with TBI were recruited in this study. Plasma visfatin and C-reactive protein levels were significantly different between the genotypes in the SNP-1535 C > T even after adjustment for age, sex and body mass index. The genotype C–C had the highest plasma visfatin and C-reactive protein concentrations. The plasma visfatin and C-reactive protein concentrations between the variant genotypes C–T and T–T did not differ significantly. Plasma visfatin level was significantly associated with plasma C-reactive protein level using multivariate linear regression. Thus, the SNP-1535 C > T of visfatin gene seemed to be potentially involved in the inflammatory component of TBI through a decreased production of visfatin.
Keywords: Visfatin; Polymorphism; C-reactive protein; Traumatic brain injury; Inflammatory reaction;

► Host defense cathelicidin peptides reverse ABCG2-mediated multidrug resistance (MDR). ► Cathelicidin peptides alone are not cytotoxic at MDR reversal concentrations. ► ABCG2 transport activity was inhibited in an uncompetitive manner by cathelicidin. ► Cathelicidin decreases ABCG2 expression in resistant cells by lysosomal degradation.Multidrug resistance (MDR) of cancer cells to a wide spectrum of anticancer drugs is a major obstacle to successful chemotherapy. It is usually mediated by the overexpression of one of the three major ABC transporters actively pumping cytotoxic drugs out of the cells. There has been great interest in the search for inhibitors toward these transporters with an aim to circumvent resistance. This is usually achieved by screening from natural product library and the subsequent structural modifications. This study reported the reversal of ABCG2-mediated MDR in drug-selected resistant cancer cell lines by a class of host defense antimicrobial peptides, the human cathelicidin LL37 and its fragments. The effective human cathelicidin peptides (LL17-32 and LL13-37) were found to increase the accumulation of mitoxantrone in cancer cell lines with ABCG2 overexpression, thereby circumventing resistance to mitoxantrone. At the effective concentrations of the cathelicidin peptides, cell proliferation of the parental cells without elevated ABCG2 expression was not affected. Result from drug efflux and ATPase assays suggested that both LL17-32 and LL13-37 interact with ABCG2 and inhibit its transport activity in an uncompetitive manner. The peptides were also found to downregulate ABCG2 protein expression in the resistant cells, probably through a lysosomal degradation pathway. Our data suggest that the human cathelicidin may be further developed for sensitizing resistant cancer cells to chemotherapy.
Keywords: Multidrug resistance; ABCG2; Efflux transporter; Cathelicidin LL37; Host defense peptides;

Comparative distribution of central neuropeptide Y (NPY) in the prairie (Microtus ochrogaster) and meadow (M. pennsylvanicus) vole by Caroline M. Hostetler; Leah N. Hitchcock; Allison M.J. Anacker; Larry J. Young; Andrey E. Ryabinin (22-29).
► Comparison of brain NPY-ir of promiscuous meadow vole and monogamous prairie vole. ► Meadow voles have higher NPY-ir in the extended amygdala and septal areas. ► Meadow voles also have higher NPY-ir in regions involved in circadian regulation. ► There were no species differences in the striatum or hippocampus. ► Findings support a role for NPY in regulating species-typical social behavior.Neuropeptide Y (NPY) has been implicated as a modulator of social behavior, often in a species-specific manner. Comparative studies of closely related vole species are particularly useful for identifying neural systems involved in social behaviors in both voles and humans. In the present study, immunohistochemistry was performed to compare NPY-like immunoreactivity (-ir) in brain tissue of the socially monogamous prairie vole and non-monogamous meadow vole. Species differences in NPY-ir were observed in a number of regions including the cortex, extended amygdala, septal area, suprachiasmatic nucleus, and intergeniculate leaf. Meadow voles had higher NPY-ir in all these regions as compared to prairie voles. No differences were observed in the striatum or hippocampus. The extended amygdala and lateral septum are regions that play a key role in regulation of monogamous behaviors such as pair bonding and paternal care. The present study suggests NPY in these regions may be an additional modulator of these species-specific social behaviors. Meadow voles had moderately higher NPY-ir in a number of hypothalamic regions, especially in the suprachiasmatic nucleus. Meadow voles also had much higher levels of NPY-ir in the intergeniculate leaflet, another key region in the regulation of circadian rhythms. Overall, species differences in NPY-ir were observed in a number of brain regions implicated in emotion, stress, circadian, and social behaviors. These findings provide additional support for a role for the NPY system in species-typical social behaviors.
Keywords: Neuropeptide Y; Prairie vole; Meadow vole; Social behavior; Circadian patterns;

Insights into scorpion venom peptides: Alternative processing of β-KTx propeptide from Tityus serrulatus venom results in a new naturally occurring thimet oligopeptidase inhibitor by Daniela Cajado Carvalho; Bruno Duzzi; Alexandre Kazuo Kuniyoshi; Mariana Fioramonte; Fábio Cesar Gozzo; Robson L. Melo; Denise Vilarinho Tambourgi; Vanessa Rioli; Fernanda Calheta Portaro (30-33).
► New pentapeptide KEILG from Tityus serrulatus venom is a EP24.15 inhibitor. ► A selective tool to differentiate EP 24.15 from EP24.16. ► Substitution of one residue affects the mechanism of inhibition. ► The sequence is homologue to the propeptide portion of βKTx. ► The presence of KEILG suggests a new processing of propeptides.Most functions attributed to Tityus serrulatus venom (TsV) are related to active molecules on ion-channels; however, here we describe a new pentapeptide that was discovered through enzymatic assay selection using EP24.15. The primary structure analysis revealed the sequence KEXXG (X means Ile or Leu), similar to the sequence present in the β-KTX propeptide described from the venom of Tityus spp. We confirmed through HPLC analysis that KEILG is the peptide present in TsV, but that KELLG also inhibits EP24.15 although through different mechanisms.

► gAd promoted proliferation and wound repair via Ca2+/CaM pathway in 16HBE14-cells, not leptin nor resistin. ► gAd inhibited ozone induced apoptosis which attributes to inhibit the ROS production. Leptin or resistin had no effect. ► 16HBE14-cells express adiponectin and its receptors, particularly adipoR1, there may be a feedback regulation between gAd and adipoR1.Epidemiological data indicate an increasing incidence of asthma in the obese individuals recent decades, while very little is known about the possible association between them. Here, we compared the roles of adipocyte-derived factors, including leptin, adiponectin and resistin on proliferation, wound repair and apoptosis in human bronchial epithelial cells (HBECs) which play an important role in the pathogenesis of asthma. The results showed that exogenous globular adiponectin (gAd) promoted proliferation, cell-cycle and wound repair of HBECs. This effect may be relevant to Ca2+/calmodulin signal pathway. Besides, gAd inhibited apoptosis induced by ozone and release of lactate dehydrogenase (LDH) of HBECs via regulated adipoR1 and reactive oxygen species. No effects of leptin or resistin on proliferation, wound repair and apoptosis of HBECs were detectable. These data indicate that airway epithelium is the direct target of gAd which plays an important role in protecting HBECs from mechanical or oxidant injuries and may have therapeutic implications in the treatment of asthma.
Keywords: Adipokines; HBECs; Proliferation; Wound repair; Apoptosis;

► SALF inhibited the production of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, IL-1α and monocyte chemoattractant protein (MCP)-1 according to an ELISA assay. ► SALF might regulate vaginal epithelial cell immune responses through the MAPK and NF-κB pathways by LPS treatment. ► SALF is thus a potential drug candidate for treating chronic inflammatory diseases, such as urethritis, vaginitis, cervicitis, and pelvic inflammatory diseases.Recently, an antimicrobial peptide (AMP), the shrimp anti-lipopolysaccharide factor (SALF), was shown to act against vaginal pathogens as demonstrated by a minimum inhibitory concentration (MIC) assay and suggested that the SALF might play a protective role in orchestrating various defensive responses. The demonstration of a protective role of the SALF in cervical cancer epithelial cells (HeLa cells) led us to investigate the anti-inflammatory effects of the SALF by determining its inhibitory effects on proinflammatory markers in LPS-stimulated cervical cancer HeLa cells. The SALF was shown to inhibit the production of inflammatory cytokines including tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, IL-1α, and monocyte chemoattractant protein (MCP)-1 according to an ELISA analysis. The SALF also suppressed mRNA levels of il-6, il-8, il-1α, and mcp-1 according to an RT-PCR. We also found that the SALF might regulate vaginal epithelial cell immune responses through the MAPK and NF-κB pathways. These findings suggest that the SALF is a potential drug candidate for treating chronic inflammatory diseases, such as urethritis, vaginitis, cervicitis, and pelvic inflammatory diseases.
Keywords: Antimicrobial peptide; LPS; Cervical cancer; Inflammatory response;

Distinct morphological and electrophysiological properties of an elk prion peptide by John Paul Glaves; Przemek A. Gorski; Kwai Alier; Li Ma; Ludovic Renault; Joseph O. Primeau; Jack H. Jhamandas; Howard S. Young (49-56).
► Prion peptides based on the 127–147 region of bovine, hamster and elk were studied. ► All of the peptides self-assembled into fibrillar states. ► Fibril conformations were highlighted by electron microscopy 3D reconstructions. ► The elk peptide had distinct electrophysiological and conformational properties.A key event in prion diseases is the conversion of the prion protein (PrP) from its native α-helical conformation to a misfolded, β-sheet rich conformation. Thus, preventing or reversing PrP misfolding could provide a means to disrupt prion disease progression and transmission. However, determining the structure of misfolded PrP has been notoriously difficult due to its inherent heterogeneity and aggregation behavior. For these reasons, simplified peptide fragments have been used as models that recapitulate characteristics of full-length PrP, such as amyloid-like aggregation and fibril formation, and in vitro toxicity. We provide a biochemical and structural comparison of PrP(127–147) peptides from elk, bovine and hamster using electrophysiology, electron microscopy and fluorescence. Our results demonstrate that the PrP(127–147) peptides adopt distinct populations of fibril structures. In addition, the elk PrP(127–147) peptide is unique in its ability to enhance Thioflavin T fluorescence and its ability to modulate neuronal ion channel conductances.
Keywords: Prions; Prion peptide aggregation; Prion peptide fibrils;

High-dose versus low-dose octreotide in the treatment of acute pancreatitis: A randomized controlled trial by Rui Wang; Fan Yang; Hao Wu; Yufang Wang; Zhiyin Huang; Bing Hu; Mingguang Zhang; Chengwei Tang (57-64).
► We conducted a RCT to find an efficient octreotide regimen to prevent or treat SAP. ► No significant difference was between control and low-dose octreotide in efficacy. ► Compared to low-dose, high-dose octreotide efficiently reduce and attenuate SAP. ► Superior effect of high-dose was due to return of plasma SST to normal level. ► High-dose was also better in decreasing plasma level of IL-6 and TNF-α.To evaluate the therapeutic efficacy of high-dose octreotide in patients with predicted severe acute pancreatitis (SAP) or SAP, two hundred and thirty-six patients with predicted SAP and 136 patients with SAP were randomized into control, high-dose octreotide (High-O) and low-dose octreotide (Low-O) groups. In addition to the conventional managements administrated in control group, High-O group received an intravenous infusion of octreotide at 50 μg/h × 3d + 25 μg/h × 4d, and Low-O group received octreotide at 25 μg/h × 7d. The major primary outcomes included the numbers of predicted SAP patients which developed SAP after intervention and the number of patients with SAP amelioration. Secondary outcomes included APACHE II, SIRS scores, plasma levels of somatostatin (SST), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6). There were no significant differences between the control and Low-O groups in terms of prevention and treatment for SAP. The incidence of SAP in patients with predicted SAP who received High-O was significantly lower than the Low-O group: 37.5% vs. 59.8%, p  = 0.005. Compared with Low-O group, the number of SAP patients in the SAP arm in the High-O group was reduced by 29.8%. Plasma levels of SST in both predicted SAP and the SAP patients were efficiently recovered (from 132.71 ± 31.40 pg/ml to 180.00 ± 23.50 pg/ml, p  < 0.05) after high-dose octreotide supplementation, which concomitantly reduced TNF-α and IL-6 levels. High-dose octreotide administration within 48 h after AP onset may efficiently reduce the risk of SAP developing and partly attenuate SAP through raising plasma SST to a normal level and decreasing IL-6 and TNF-α.
Keywords: Octreotide; Acute pancreatitis; Somatostatin; Inflammation;

An immunomodulatory peptide related to frenatin 2 from skin secretions of the Tyrrhenian painted frog Discoglossus sardus (Alytidae) by J. Michael Conlon; Milena Mechkarska; Jelena M. Pantic; Miodrag L. Lukic; Laurent Coquet; Jérôme Leprince; Per F. Nielsen; Andrea C. Rinaldi (65-71).
► Skin secretions of Discoglossus sardus do not contain cationic antimicrobial peptides. ► Frenatin 2D (DLLGTLGNLPLPFI.NH2) is present in high concentration. ► Frenatin 2D stimulates IL-1β, IL-12, and TNF-α release by macrophages. ► Frenatin 2D may act to stimulate the adaptive immune system following infection.Norepinephrine-stimulated skin secretions of the Tyrrhenian painted frog Discoglossus sardus Tschudi, 1837 (Alytidae) did not contain any peptide with antimicrobial or hemolytic activity. However, peptidomic analysis of the secretions revealed the presence of an abundant peptide with structural similarity to frenatin 2, previously isolated from the Australian frog Litoria infrafrenata (Hylidae). The primary structure of the peptide, termed frenatin 2D, was established as DLLGTLGNLPLPFI.NH2 by automated Edman degradation and mass spectrometry with electron-transfer dissociation (ETD)-based fragmentation and confirmed by chemical synthesis. The structure of a second frenatin 2-related peptide, termed frenatin 2.1D, that was present in much lower abundance was established as GTLGNLPAPFPG. Frenatin 2D (20 μg/ml) significantly stimulated production of the proinflammatory cytokines TNF-α (P  < 0.05) and IL-1β (P  < 0.01) by mouse peritoneal macrophages but the peptide did not potentiate the stimulation produced by lipopolysaccharide (LPS). The peptide increased IL-12 production in both unstimulated (P  < 0.01) and LPS-stimulated (P  < 0.05) cells but stimulatory effects on IL-6 production were not significant. The biological role of frenatin 2D is unknown but it is speculated that the peptide acts on skin macrophages to produce a cytokine-mediated stimulation of the adaptive immune system in response to invasion by microorganisms.
Keywords: Frenatin; Discoglossus sardus; Alytidae; Immunomodulatory peptide; Cytokine; Macrophage;

► We differentially extracted Urotensin II and Urotensin related peptide from plasma. ► We used a novel immunoluminometric assay to measure the peptides. ► We measured these peptides in controls and acute heart failure patients. ► Both peptides were elevated in heart failure patients compared to controls. ► This further supports a role of the Urotensin system in heart failure.Urotensin II (UTN) is a cyclic eleven amino acid peptide that can induce endothelial independent vasoconstriction and endothelial dependent vasodilatation in human vasculature. The cyclic part of the peptide is composed of six amino acids. Similarly, Urotensin Related Peptide (URP) is only eight amino acids long but shares the identical ring structure to UTN. Plasma UTN has been shown to be raised in patients with chronic heart failure (CHF) suggesting a potential role of the peptide system in the pathophysiology of heart failure. Given their similar structures, techniques measuring plasma UTN may also be simultaneously detecting URP and could provide a misrepresentation of true UTN and URP levels in patients’ plasma. Thus we describe the development of a solid phase extraction technique that can differentially extract UTN and URP from human plasma so that they can be assayed separately using non-radioactive immunoluminometric assays. This reliable and sensitive protocol was utilized to characterise the plasma of 20 healthy controls and 20 patients admitted with acute heart failure (AHF). The groups were age and sex matched. Plasma UTN was significantly raised in patients with AHF on admission when compared to controls (median 1.29 [range 0.50–5.55] pmol/L vs 0.50 [0.50–3.33] pmol/L, p  = 0.019). Likewise plasma URP was significantly higher in the heart failure group on admission (8.38 [1.30–66.80] pmol/L vs 2.25 [1.30–14.40] pmol/L, p  < 0.005). This suggests a role for both members of the Urotensin peptide system in acute heart failure.
Keywords: Urotensin II; Urotensin related peptide; Heart failure; Plasma;

Quorum sensing inhibitor FS3-coated vascular graft enhances daptomycin efficacy in a rat model of staphylococcal infection by Oscar Cirioni; Federico Mocchegiani; Ivana Cacciatore; Jacopo Vecchiet; Carmela Silvestri; Leonardo Baldassarre; Claudio Ucciferri; Elena Orsetti; Pamela Castelli; Mauro Provinciali; Marco Vivarelli; Erika Fornasari; Andrea Giacometti (77-81).
► Daptomycin without FS3 showed MIC and MBC values of 4.00 and 16.00 mg/l. ► Daptomycin in presence of FS3 showed MICs and MBCs fourfold lower. ► In in vivo studies, singly daptomycin or FS3 have good efficacies. ► Their combination was significantly higher than that of each single compound.The aim of the study was to investigate the efficacy of the quorum sensing inhibitor FS3 and daptomycin in preventing prosthesis biofilm in a rat model of staphylococcal vascular graft infection. Graft infections were established in the back subcutaneous tissue of adult male Wistar rats by implantation of Dacron prostheses followed by topical inoculation with 2 × 107 colony-forming units of Staphylococcus aureus, strain Smith diffuse. The study included a control group, a contaminated group that did not receive any antibiotic prophylaxis and three contaminated groups that received: (i) intraperitoneal daptomycin, (ii) FS3-soacked graft, and (iii) daptomycin plus FS3-soaked graft, respectively. Each group included 15 animals. The infection burden was evaluated by using sonication and quantitative agar culture. Moreover, an in vitro binding-study was performed to quantify the how much FS3 was coated to the surface of the prosthesis. The in vitro studies showed, that minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values for daptomycin were lower in presence of FS3. In in vivo studies, when tested alone, daptomycin and FS3 showed good efficacies. Their combination showed efficacies significantly higher than that of each single compound. Daptomycin is an important candidate for prevention of staphylococcal biofilm related infection and FS3 could serve as an interesting anti-staphylococcal antibiotic enhancer.
Keywords: Lipopeptides; Daptomycin; FS3; Vascular graft infection; Bacterial biofilm;

The protective effect of oxytocin on ischemia/reperfusion injury in rat urinary bladder by G. Erkanli Senturk; K. Erkanli; U. Aydin; D. Yucel; N. Isiksacan; F. Ercan; S. Arbak (82-88).
► Oxytocin played role as an antioxidant on I/R injury in urinary bladder. ► Oxytocin protected the urothelium histopathologically against I/R injury. ► Oxytocin reduced the activation and mast cell number in urinary bladder after I/R.Oxytocin (OXY), a well-known nonapeptide, plays a crucial role in reproduction, and has effects on modulating the immune and inflammatory processes in living organisms as well. Recently it is also known as an antioxidant in several organs. The present study aims to demonstrate the protective effect of OXY against ischemia/reperfusion (I/R) injury in urinary bladder tissue. Abdominal aorta of rats, were clamped to perform urinary bladder ischemia. OXY (0.5 μg/kg) was injected intraperitoneally before ischemia in I/R + OXY group, whereas the vehicle solution was injected to I/R group. At the end of reperfusion, tissue samples from urinary bladder were processed for histochemical, ultrastructural and biochemical analysis. Tissue sections were stained by toluidine blue for mast cell counting and hematoxylin–eosin for histopathology. In addition, malondialdehyde (MDA) and glutathione (GSH) levels were determined biochemically. The results demonstrated that there was an extreme damage at urothelium, dilatation of intercellular junctions, inflammatory cell infiltration in I/R group. I/R + OXY group demonstrated a reduction in the severity of urinary bladder damage. According to mast cell counting results, both granulated and degranulated mast cells were decreased in I/R + OXY group compared to I/R group. The mean MDA level was higher in I/R group compared to control and lower in I/R + OXY group compared to I/R group. GSH level reduced in I/R group compared to the control and increased in I/R + OXY group compared to I/R group. In conclusion, oxytocin, as confirmed by histological evaluation and biochemical assays has a potential protective effect in the urinary bladder tissue against ischemia/reperfusion injury.
Keywords: Ischemia/reperfusion; Oxytocin; Urinary bladder;

Treatment with pioglitazone is associated with decreased preprandial ghrelin levels: A randomized clinical trial by Shervin Taslimi; Alireza Esteghamati; Armin Rashidi; Hosein Moin Tavakkoli; Manouchehr Nakhjavani; Abbas Kebriaee-Zadeh (89-92).
► The effect of pioglitazone on ghrelin has not been clearly established. ► Pioglitazone was associated with preprandial ghrelin reduction in type 2 diabetes. ► This effect was independent of changes in body weight, BMI, glucose, and insulin.The effects of metformin and pioglitazone on ghrelin, a physiologic regulator of appetite and food intake, have not been clearly established. In a randomized clinical trial, we randomly assigned 60 type 2 diabetic patients to either metformin (Group A; n  = 30) or pioglitazone (Group B; n  = 30) treatment groups. The groups were similar in their baseline characteristics. A standard fasting 75 g oral glucose tolerance test was performed at time zero before starting metformin or pioglitazone, and 3 months later. After 3 months of treatment, pioglitazone, but not metformin, was significantly associated with weight gain. Both groups experienced a significant reduction in fasting plasma glucose (p  < 0.01), hemoglobin A1c (p  < 0.01 in Group A and p  < 0.05 in Group B), and insulin resistance (p  < 0.01). The effect of metformin on preprandial ghrelin and its response to glucose challenge was not significant, while the pioglitazone group had a significant reduction in preprandial ghrelin levels after treatment (p  < 0.05). The effect of pioglitazone on ghrelin was independent of changes in body weight, body mass index, glucose control, insulin resistance, and plasma insulin. In conclusion, treatment with pioglitazone is associated with a decrease in preprandial ghrelin levels and therefore, the weight gain and increased food intake related to pioglitazone use cannot be explained by its effects on ghrelin. The effect of pioglitazone on ghrelin is independent of changes in body weight, body mass index, plasma insulin, insulin resistance, or glucose control.
Keywords: Glucose tolerance; Metformin; Leptin; Ghrelin;

Display Omitted► Role of CNP in heart failure and myocardial infarction. ► Increased CNP mRNA expression and plasmatic concentrations in humans and in animal models. ► CNP expression in cardiomyocytes and its evaluation in human leukocytes. ► CNP and its specific receptor, NPR-B, play a very important role in regulating cardiac hypertrophy and remodeling. ► NPR-B could be a new potential drug target for the treatment of cardiovascular disease.Natriuretic peptides are endogenous hormones released by the heart in response to myocardial stretch and overload. While atrial and brain natriuretic peptides (ANP, BNP) were immediately considered cardiac hormones and their role was well-characterized and defined in predicting risk in cardiovascular disease, evidence indicating the role of C-type natriuretic peptide (CNP) in cardiovascular regulation was slow to emerge until about 8 years ago. Since then, considerable literature on CNP and the cardiovascular system has been published; the aim of this review is to examine current literature relating to CNP and cardiovascular disease, in particular its role in heart failure (HF) and myocardial infarction (MI). This review retraces the fundamental steps in research that led understanding the role of CNP in HF and MI; from increased CNP mRNA expression and plasmatic concentrations in humans and in animal models, to detection of CNP expression in cardiomyocytes, to its evaluation in human leukocytes. The traditional view of CNP as an endothelial peptide has been surpassed by the results of many studies published in recent years, and while its physiological role is still under investigation, information is now available regarding its contribution to cardiovascular function. Taken together, these observations suggest that CNP and its specific receptor, NPR-B, can play a very important role in regulating cardiac hypertrophy and remodeling, indicating NPR-B as a new potential drug target for the treatment of cardiovascular disease.
Keywords: C-type natriuretic peptide; Natriuretic peptide; Chronic heart failure; Natriuretic peptide receptors; Myocardial infarction;

Salivary vasopressin increases following intranasal oxytocin administration by Omri Weisman; Inna Schneiderman; Orna Zagoory-Sharon; Ruth Feldman (99-103).
► Peripheral vasopressin increases following intranasal oxytocin administration in humans. ► This pattern is limited to the first hour following administration. ► Vasopressin demonstrates high degree of individual stability over four hours and across six months. ► Plasma and salivary AVP are moderately correlated, supporting analysis of salivary AVP using EIA. ► Endogenous levels of AVP shape response to intranasal manipulation.Extant research has documented the effects of intranasal administration of oxytocin (OT), and to a lesser degree Arginine Vasopressin (AVP) – two structurally-related neuropeptides – on brain and behaviour, yet the effects of exogenous manipulation of one on circulating levels of the other remain unknown. Studies have shown that OT administration impacts the peripheral levels of numerous hormones; however, whether OT administration also increases AVP concentrations has not been explored. Utilizing a double-blind placebo-controlled within-subject design, ten male and female subjects provided ten saliva samples over four consecutive hours: at baseline and nine times following OT administration. Results indicate that salivary AVP increased in the first hour following OT manipulation (OT condition: mean AVP = 2.17 pg/ml, SE  = 28, placebo condition: mean AVP = 1.42 pg/ml, SE  = .18) but returned to baseline levels at the next assessment (80 min) and remained low for the remaining period. Similar to OT, AVP showed high degree of individual stability and baseline levels of AVP correlated with AVP concentrations at the first and second post-administration hours regardless of drug condition (Pearson r  = .85–.93). Validity of salivary AVP ELISA measurement was verified by demonstrating individual stability of salivary AVP over a six-month period (r  = .70, p  < .000) as well correlation with plasma levels over the same period (r  = .32, p  = .037) in a sample of 45 young adults who did not participate in the current study. Overall, findings suggest a potential crosstalk between OT and AVP and indicate that baseline levels of the two neuropeptides may shape the degree to which these systems respond to exogenous manipulation.

Diabetes-induced loss of gastric ICC accompanied by up-regulation of natriuretic peptide signaling pathways in STZ-induced diabetic mice by Yi-Song Wu; Hong-Li Lu; Xu Huang; Dong-Hai Liu; Xiang-Min Meng; Xin Guo; Young-chul Kim; Wen-Xie Xu (104-111).
► We found that natriuretic peptide (NP) signaling pathway are up-regulated in diabetic gastric smooth muscle of mouse. ► NP/NPR-A, B/cGMP signaling pathway down-regulates mSCF/c-Kit signaling pathway in gastric smooth muscle. ► cANF/NPR-C signaling pathway shows an anti-proliferation effect in cultured gastric smooth muscle cells.Our previous study demonstrated that natriuretic peptides (NPs) play an inhibitory role in regulation of gastric smooth muscle motility. However, it is not clear whether NPs are involved in diabetics-induced loss of gastric interstitial cell of Cajal (ICC). The present study was designed to investigate the relationship between diabetics-induced loss of gastric ICC and natriuretic peptide signaling pathway in streptozotocin (STZ)-induced diabetic mice. The results showed that the protein expression levels of c-Kit and membrane-bound stem cell factor (mSCF) in gastric smooth muscle layers were decreased in STZ-induced diabetic mice. However, both mRNA and protein expression levels of natriuretic peptide receptor (NPR)-A, B and C were increased in the same place of the diabetic mice. The amplitude of spontaneous contraction in gastric antral smooth muscles was inhibited by C-type natriuretic peptide (CNP) dose-dependently and the inhibitory effect was potentiated in diabetic mice. Pretreatment of the cultured gastric smooth muscle cells (GSMCs) with different concentration of CNP can significantly decrease the mSCF expression level. 8-Bromoguanosine-3′,5′-cyclomo-nophosphate (8-Br-cGMP), a membrane permeable cGMP analog, mimicked the effect of CNP but not cANF (a specific NPR-C agonist). Methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay showed that high concentration of cANF (10−6  mol/L) inhibited cell proliferation in cultured GSMCs. These findings suggest that up-regulation of NPs/NPR-A, B/cGMP and NPs/NPR-C signaling pathways may be involved in diabetes-induced loss of gastric ICC.
Keywords: Natriuretic peptide; Diabetic gastroparesis; Interstitial cell of Cajal (ICC); Membrane-bound stem cell factor (mSCF); Gastric smooth muscle cells (GSMCs);

wrwyrggrywrw is a single-chain functional analog of the Holliday junction-binding homodimer, (wrwycr)2 by Marc C. Rideout; Ilham Naili; Jeffrey L. Boldt; America Flores-Fujimoto; Sukanya Patra; Jason E. Rostron; Anca M. Segall (112-122).
► wrwyrggrywrw inhibits RecG- and RuvABC- mediated processing of Holliday junctions in vitro. ► wrwyrggrywrw causes DNA damage and loss of membrane potential in E. coli in vivo. ► wrwyrggrywrw is a potent antimicrobial with minimum inhibitory concentrations in the low μg/mL range. ► wrwyrggrywrw has minimal toxicity to eukaryotic cell types at the concentrations necessary to inhibit bacterial growth.DNA repair pathways in bacteria that use homologous recombination involve the formation and subsequent resolution of Holliday junction (HJ) intermediates. We have previously identified several hexameric peptides that bind to HJs and interfere with HJ processing enzymes in vitro. The peptide WRWYCR and its D-amino acid stereoisomer wrwycr, are potent antibacterial agents. These hexapeptides must form homodimers in order to interact stably with HJs, and inhibit bacterial growth, and this represents a potential limitation. Herein we describe a disulfide bond-independent inhibitor, WRWYRGGRYWRW and its D-stereoisomer wrwyrggrywrw. We have characterized these single-chain, linear analogs of the hexapeptides, and show that in addition to effectively binding to HJs, and inhibiting the activity of DNA repair enzymes that process HJs, they have equal or greater potency against Gram-positive and Gram-negative bacterial growth. The analogs were also shown to cause DNA damage in bacteria, and disrupt the integrity of the bacterial cytoplasmic membrane. Finally, we found that they have little toxicity toward several eukaryotic cell types at concentrations needed to inhibit bacterial growth.
Keywords: Antibacterial peptides; Homodimer; Holliday junction-binding peptides; RecG helicase; RuvABC resolvase; DNA repair inihbitors;

Long-term treatment with EXf, a peptide analog of Exendin-4, improves β-cell function and survival in diabetic KKAy mice by Guo-jiang Hou; Cai-na Li; Shuai-nan Liu; Yi Huan; Quan Liu; Su-juan Sun; Lin-yi Li; Shao-cong Hou; Zhu-fang Shen (123-132).
► EXf is a novel peptide analog of Exendin-4 with two amino acid substitutions. ► EXf improved glycemic control and β-cell function, restored islet morphology. ► EXf stimulated proliferation and inhibited apoptosis of β-cells. ► EXf improved islet β-cell survival by PI3 K/Akt1 signal pathway. ► EXf may slow down the diabetes progress in diabetic KKAy mice.EXf is a C-terminally truncated fragment of Exendin-4 with two amino acid substitutions. Previous studies showed that EXf controls plasma glucose level acting as a glucagon-like peptide 1 (GLP-1) receptor agonist. The purpose of this study was to evaluate the effects of EXf on β-cell function and survival in diabetic KKAy mice. EXf treatment significantly improved the glucose intolerance and reduced non-fasting and fasting plasma glucose levels, as well as plasma triglyceride levels in diabetic KKAy mice. In hyperglycemic clamp test, EXf-treated mice displayed an increased glucose infusion rate and first-phase insulin secretion. Treatment with EXf also led to a significant restoration of islet morphology, an increase in Ki67 expression in β-cells, and a reduction in the number of TUNEL positive β-cells. In the pancreas, comparative transcription analysis showed up-regulation of Akt1. The up-regulation of phosphorylated Akt1 was confirmed by Western blot, and changes in the protein levels of members of the Akt1 pathway, such as PI3K, Bim, Bcl-2, Bax, Caspase-3, and Caspase-9, PDX-1, were observed as well. Therefore, EXf treatment could improve β-cell function and survival in diabetic KKAy mice, likely as a result of islet morphology restoration, stimulation of β-cell proliferation, and inhibition of β-cell apoptosis.
Keywords: EXf; β-Cell function; β-Cell survival; Proliferation; Apoptosis; Diabetic KKAy mice;

► Bradykinin attenuated basal and LPS-induced NO production in BV2 microglial cells. ► A protein kinase A inhibitor mimicked the effects of Bradykinin, while cAMP elevating agents antagonized Bradykinin-mediated NO decrease. ► Moreover, Bradykinin inhibited the activation of cAMP responsive element binding protein expression. ► Bradykinin protected microglial cells from death triggered by a combination of LPS and each of the cAMP elevating agents. ► Gαi protein inhibitor abrogated the effects of BK on NO release, and the expression of Gαi protein in the plasma membrane was induced by BK.Bradykinin (BK) is a major potent inflammatory mediator outside the central nervous system. In Alzheimer's disease, BK release and BK receptor expression in brain tissues are upregulated relatively early during the course of the disease. Hence, BK was believed to promote neuroinflammation. However, BK was recently reported to possess anti-inflammatory and neuroprotective roles. Exposure of BV2 microglial cell line to BK lead to a decrease in NO release from unstimulated cells as well as a dose-dependent attenuation, mediated by both B1 and B2 receptors, in lipopolysaccharide (LPS)-induced NO production. In this study we examined whether cyclic adenosine monophosphate (cAMP) signaling is involved in BK-mediated effect in microglial nitric oxide (NO) production. A protein kinase A (PKA) inhibitor mimicked the effects of BK, while cAMP elevating agents antagonized BK-mediated NO decrease. Moreover, BK inhibited the activation of cAMP responsive element binding protein (CREB). In addition, BK protected microglial cells from death triggered by combinations of LPS and each of the cAMP elevating agents. Finally, the addition of Gαi protein inhibitor abrogated the effects of BK on NO release, and the expression of Gαi protein in the plasma membrane was induced by BK. These results suggest that BK-mediated reduction in microglial NO production depends on coupling to Gi protein and also involves inhibition of cAMP-PKA-CREB signaling.
Keywords: Bradykinin; Microglia; Nitric oxide; Cyclic adenosine monophosphate; Gαi protein;

Oxytocin inhibits pentylentetrazol-induced seizures in the rat by Oytun Erbas; Mustafa Yılmaz; Huseyin Anil Korkmaz; Saylav Bora; Vedat Evren; Gonul Peker (141-144).
► Epilepsy is characterized as recurrent seizures arising from abnormal electrical activity. ► PTZ-induced seizures result in electrographic, molecular and endocrine responses in the brain. ► Oxytocin (OT) is a neurohypophysial nonapeptide synthesized in hypothalamus. ► It was found that OT has an inhibitory effect on seizures via EEG recording. ► OT could be used to treat epilepsy patient.We aimed to reveal the anti-convulsant effects of oxytocin (OT) in pentylenetetrazol (PTZ)-induced seizures in rats. Thirty rats were randomly divided into 5 equal groups. Using stereotaxy, we implanted electroencephologram (EEG) electrodes in the left nucleus of the posterior thalamus. After 2 days, the first and second groups were used as the control and PTZ (35 mg/kg) groups, respectively. We administered 40, 80 and 160 nmol/kg OT + 35 mg/kg PTZ to the rats, constituting the third, fourth, and fifth groups, respectively, for 5 days. At the end of 5 days, we recorded EEGs via bipolar EEG electrodes. After 12 h, all groups except the first received 70 mg/kg PTZ and we determined the dose–response ratio. Racine's Convulsion Scale was used to evaluate seizures. The spike–wave complex percentage in the EEG was determined as 0% for the first group, 38.6% ± 7.2 for the second group, 36.4% ± 5.6 for the third group, 4.3% ± 1.8 for the fifth group and 4.1% ± 1.1 for the fifth group. The fourth and fifth groups had significantly decreased spike–wave complex percentages compared to the second group (p  < 0.0001). OT may prevent PTZ-induced epilepsy on an EEG. OT may also be considered for use in the treatment of epilepsy in the future.
Keywords: Oxytocin; Pentylenetetrazol; Epilepsy; Rat;

► Food intake and body weight were reduced by acupuncture ST36 in obese prone rats. ► Acupuncture ST36 increased POMC mRNA level in NTS and hypoglossal nucleus regions. ► TRPV1 and nNOS mRNAs were increased by acupuncture ST36 in the NTS/GN regions. ► Acupuncture ST36 increased TRPV1 and nNOS mRNAs in the skin acupoints. Objective: The purpose of this study was to determine the effects of electroacupuncture (EA) ST36 on food intake and body weight in obese prone (OP) rats compared to obese resistant (OR) strain on a high fat diet. The influences of EA on mRNA levels of pro-opiomelanocortin (POMC), transient receptor potential vanilloid type-1 (TRPV1), and neuronal nitric oxide synthase (nNOS) were also examined in the medulla regions and ST36 skin tissue. Methods: Advanced EA ST36 was conducted in two sessions of 20 min separated by an 80 min interval for 7 days. Food intake and body weight were recorded in conscious rats every day. Real time PCR was conducted in the micropunches of the medulla regions and skin tissues at the end of the treatment. Results: Food intake and body weight were significantly reduced by advanced EA ST36 in OP rats, but slightly decreased in OR strain and sham-EA rats. Advanced EA ST36 produced a marked increase in POMC mRNA level in the nucleus tractus solitarius (NTS) and hypoglossal nucleus (HN) regions. TRPV1 and nNOS mRNAs were simultaneously increased in the NTS/gracile nucleus regions and in the ST36 skin regions by the EA treatment in OP rats. Conclusions: We conclude that advanced EA ST36 produces an up-regulation of anorexigenic factor POMC production in the NTS/HN, which inhibits food intake and reduces body weight. EA-induced expression of TRPV1-nNOS in the ST36 and the NTS/gracile nucleus is involved in the signal transduction of EA stimuli via somatosensory afferents-medulla pathways.
Keywords: Appetite regulation; POMC; TRPV; Medulla; Acupuncture;