Peptides (v.91, #C)

Role of human lipocalin proteins in abdominal obesity after acute pancreatitis by Ruma G. Singh; Sayali A. Pendharkar; Lindsay D. Plank; Maxim S. Petrov (1-7).
Lipocalin proteins are small regulatory peptides implicated in metabolism, inflammation, and immunity. Although lipocalin proteins have been linked to various clinical conditions, their role in the acute inflammatory setting, such as acute pancreatitis (AP), has only been sparsely investigated. Two members of the lipocalin family, lipocalin-2 (LCN-2) and retinol binding protein −4 (RBP-4), play an important role in obesity and insulin resistance. In this study, we analysed circulating levels of LCN-2 and RBP-4 in 92 individuals after AP, of whom 41 individuals had abdominal obesity and 51 did not. Binary logistic regression analyses were performed to determine whether abdominal obesity was associated with the two lipocalin proteins. Lipocalin-2 was significantly associated with abdominal obesity in the unadjusted model (Odds ratio (OR) = 1.014 [95% confidence interval (CI): 1.000, 1.028], P = 0.05) and after adjusting for patient related (age, ethnicity, and diabetes mellitus) and pancreatitis related (aetiology, severity, recurrence, and duration of AP) characteristics (OR = 1.018 [95% CI: 1.001, 1.036], p = 0.04). Further, the association of LCN-2 with waist circumference was significant in individuals with alcohol aetiology of AP (β=1.082 [95% CI: 1.011, 1.158], p = 0.02]. The association between RBP-4 and abdominal obesity was not significant in both unadjusted and adjusted models. These findings indicate that circulating levels of LCN-2 in patients after AP may play a role in chronic low grade inflammation associated with abdominal adiposity and that alcohol consumption may further exacerbate adipose tissue dysfunction.
Keywords: Lipocalin proteins; Lipocalin-2; Retinol binding protein-4; Abdominal obesity; Acute pancreatitis; Inflammation;

Recent years have brought in an increased interest to develop improved polymyxins. The currently used polymyxins, i.e. polymyxin B and colistin (polymyxin E) are pentacationic lipopeptides that possess a cyclic heptapeptide part with three positive charges, a linear “panhandle” part with two positive charges, and a fatty acyl tail. Unfortunately, their clinical use is shadowed by their notable nephrotoxicity. We have previously developed a polymyxin derivative NAB739 which lacks the positive charges in the linear part. This derivative is better tolerated than polymyxin B in cynomolgus monkeys and is, in contrast to polymyxin B, excreted into urine in monkeys and rats. Here we have conducted further structure-activity relationship (SAR) studies on 17 derivatives with three positive charges only. We discovered a remarkably antibacterial class, as exemplified by NAB815, that carries two positive charges only in the cyclic part.
Keywords: NAB739, NAB815; Escherichia coli; Klebsiella pneumoniae; Acinetobacter baumannii; Pseudomonas aeruginosa; Excretion to urine; Nephrotoxicity; HK-2;

Discovery of three toxin peptides with Kv1.3 channel and IL-2 cytokine-inhibiting activities from Non-Buthidae scorpions, Chaerilus tricostatus and Chaerilus tryznai by Li Ding; Jing Chen; Jinbo Hao; Jiahui Zhang; Xuejun Huang; Fangfang Hu; Zheng Wu; Yaru Liu; Wenxin Li; Zhijian Cao; Yingliang Wu; Jian Li; Shan Li; Hongyan Liu; Wenlong Wu; Zongyun Chen (13-19).
Non-Buthidae venomous scorpions are huge natural sources of toxin peptides; however, only a few studies have been done to understand their toxin peptides. Herein, we describe three new potential immunomodulating toxin peptides, Ctri18, Ctry68 and Ctry2908, from two non-Buthidae scorpions, Chaerilus tricostatus and Chaerilus tryznai. Sequence alignment analyses showed that Ctri18, Ctry68 and Ctry2908 are three new members of the scorpion toxin α-KTx15 subfamily. Electrophysiological experiments showed that Ctri18, Ctry68 and Ctry2908 blocked the Kv1.3 channel at micromole to nanomole levels, but had weak effects on potassium channel KCNQ1 and sodium channel Nav1.4, which indicated that Ctri18, Ctry68 and Ctry2908 might have specific inhibiting effects on the Kv1.3 channel. ELISA experiments showed that Ctri18, Ctry68 and Ctry2908 inhibited IL-2 cytokine secretions of activated T lymphocyte in human PBMCs. Excitingly, consistent with the good Kv1.3 channel inhibitory activity, Ctry2908 inhibited cytokine IL-2 secretion in nanomole level, which indicated that Ctry2908 might be a new lead drug template toward Kv1.3 channels. Together, these studies discovered three new toxin peptides, Ctri18, Ctry68 and Ctry2908, with Kv1.3 channel and IL-2 cytokine-inhibiting activities from two scorpions, C. tricostatus and C. tryznai, and highlighted that non-Buthidae venomous scorpions are new natural toxin peptide sources.
Keywords: Chaerilidae; Kv1.3 channel; Scorpion toxin; IL-2; Potential immunoregulatory peptide;

The effect of CART on pituitary hormones release from cultured pituitary cells harvested from fasted and fed ad libitum male rats by M. Chmielowska; B. Baranowska; E. Wolinska-Witort; L. Martynska; M. Kalisz; A. Litwiniuk; W. Bik (20-25).
Cocaine and Amphetamine-Regulated Transcript (CART) is widely expressed in the central nervous system and in several endocrine organs. CART is an important factor in the regulation of energy homeostasis. The aim of the study was to assess the role of CART in physiological response of pituitary cells in a course of starvation. The pituitary cells harvested from starved and fed ad libitum male rats were cultured for 48 h and treated with: 0.1 nM, 1 nM, 10 nM or 100 nM doses of CART. The medium was collected after 60 min and stored at −70 °C until samples were further assayed for: LH, FSH, PRL, GH, TSH and ACTH. We revealed that in cultures of pituitary cells collected from fasted rats the basal levels of the examined hormones were reduced. Incubation of pituitary cells of non-starved rats with any dose of CART reduced the concentration of LH and TSH, while the levels of the other hormones were decreased after administration only specific doses of CART. In cells of fasted rats no change in the concentration of gonadotrophins was observed. The PRL level was increased only in the 1 nM dose of CART, while the 10 nM and 100 nM CART doses markedly enhanced GH and TSH. Moreover, administration of 1 nM, 10 nM and 100 nM of CART to cultured cells of fasted rats resulted in a significant rise of the ACTH.Our results indicate that CART can directly affect the physiological release of PRL, ACTH, TSH and GH in pituitary cells of starved animals. Moreover, CART did not alter the LH and FSH suppression level, which is correlated with food deprivation. This data stays in contrast with the already proposed role of CART as an anorexigenic hypothalamic factor.
Keywords: CART; Pituitary hormones; Starvation;

The effects of different exercise training modalities on plasma proenkephalin Peptide F in women by William H. DuPont; William J. Kraemer; Bradley C. Nindl; Elaine C. Lee; Maren S. Fragala; Disa L. Hatfield; Lydia K. Caldwell; Emily M. Post; Matthew K. Beeler; Jeff S. Volek; Carl M. Maresh (26-32).
Due to the important interactions of proenkephalin fragments (e.g., proenkephalin [107–140] Peptide F) to enhance activation of immune cells and potentially combat pain associated with exercise-induced muscle tissue damage, we examined the differential plasma responses of Peptide F to different exercise training programs. Participants were tested pre-training (T1), and after 8 weeks (T2) of training. Fifty-nine healthy women were matched and then randomly assigned to one of four groups: heavy resistance strength training (STR, n  = 18), high intensity endurance training (END, n  = 14), combined strength and endurance training (CMB, n  = 17), or control (CON, n  = 10). Blood was collected using a cannula inserted into a superficial vein in the antecubital fossa with samples collected at rest and immediately after an acute bout of 6 X 10 RM in a squat resistance exercise before training and after training. Prior to any training, no significant differences were observed for any of the groups before or after acute exercise. With training, significant (P  ≤ 0.95) elevations were observed with acute exercise in each of the exercise training groups and this effect was significantly greater in the CMB group. These data indicate that in untrained women exercise training will not change resting of plasma Peptide F concentrations unless both forms of exercise are performed but will result in significant increases in the immediate post-exercise responses. Such findings appear to indicate adrenal medullary adaptations opioid production significantly altered with exercise training.
Keywords: Opioid peptides; Physical training; Endurance exercise; Strength;

Semi-microbiological synthesis of an active lysinoalanine-bridged analog of glucagon-like-peptide-1 by Anneke Kuipers; Louwe de Vries; Marcel P. de Vries; Rick Rink; Tjibbe Bosma; Gert N. Moll (33-39).
Display OmittedSome modified glucagon-like-peptide-1 (GLP-1) analogs are highly important for treating type 2 diabetes. Here we investigated whether GLP-1 analogs expressed in Lactococcus lactis could be substrates for modification and export by the nisin dehydratase and transporter enzyme. Subsequently we introduced a lysinoalanine by coupling a formed dehydroalanine with a lysine and investigated the structure and activity of the formed lysinoalanine-bridged GLP-1 analog. Our data show: (i) GLP-1 fused to the nisin leader peptide is very well exported via the nisin transporter NisT, (ii) production of leader-GLP-1 via NisT is higher than via the SEC system, (iii) leader-GLP-1 exported via NisT was more efficiently dehydrated by the nisin dehydratase NisB than when exported via the SEC system, (iv) individual serines and threonines in GLP-1 are dehydrated by NisB to a significantly different extent, (v) an introduced Ser30 is well dehydrated and can be coupled to Lys34 to form a lysinoalanine-bridged GLP-1 analog, (vi) a lysinoalanine(30-34) variant's conformation shifts in the presence of 25% trifluoroethanol towards a higher alpha helix content than observed for wild type GLP-1 under identical condition, (vii) a lysinoalanine(30-34) GLP-1 variant has retained significant activity. Taken together the data extend knowledge on the substrate specificities of NisT and NisB and their combined activity relative to export via the Sec system, and demonstrate that introducing a lysinoalanine bridge is an option for modifying therapeutic peptides.
Keywords: Glucagon like peptide-1; Lanthipeptide; Nisin; Lysine; Dehydroalanine;

Structure-constrained endomorphin analogs display differential antinociceptive mechanisms in mice after spinal administration by Yuan Wang; Jingjing Zhou; Xin Liu; Long Zhao; Zhaojuan Wang; Xianghui Zhang; Kezhou Wang; Linqing Wang; Rui Wang (40-48).
Display OmittedWe previously reported a series of novel endomorphin analogs with unnatural amino acid modifications. These analogs display good binding affinity and functional activity toward the μ opioid receptor (MOP). In the present study, we further investigated the spinal antinociceptive activity of these compounds. The analogs were potent in several nociceptive models. Opioid antagonists and antibodies against several endogenous opioid peptides were used to determine the mechanisms of action of these peptides. Intrathecal pretreatment with naloxone and β-funaltrexamine (β-FNA) effectively inhibited analog-induced analgesia, demonstrating that activity of the analogs is regulated primarily through MOP. Antinociception induced by analog 2 through 4 was not reversed by δ opioid receptor (DOP) or κ opioid receptor (KOP) antagonist; antibodies against dynorphin-A (1–17), dynorphin-B (1–13), and Leu5/Met5-enkephalin had no impact on the antinociceptive effects of these analogs. In contrast, antinociceptive effects induced by a spinal injection of the fluorine substituted analog 1 were significantly reversed by KOP antagonism. Furthermore, intrathecal pretreatment with antibodies against dynorphin-B (1–13) attenuated the antinociceptive effect of analog 1. These results indicate that the antinociceptive activity exerted by intrathecally-administered analog 1 is mediated, in part, through KOP with increased release of dynorphin-B (1–13). The chemical modifications used in the present study may serve as a useful tool to gain insight into the mechanisms of endomorphins activity.
Keywords: Spinal antinociception; Endomorphin; Dynorphin; Antagonist; Antibody;

Glucagon-like petide-2 acts on colon cancer myofibroblasts to stimulate proliferation, migration and invasion of both myofibroblasts and cancer cells via the IGF pathway by Marianne Shawe-Taylor; J. Dinesh Kumar; Whitney Holden; Steven Dodd; Akos Varga; Olivier Giger; Andrea Varro; Graham J. Dockray (49-57).
Glucagon-like peptide (GLP)-2 stimulates intestinal epithelial proliferation by acting, in part, via IGF release from sub-epithelial myofibroblasts. The response of myofibroblasts to GLP-2 remains incompletely understood. We studied the action of GLP-2 on myofibroblasts from colon cancer and adjacent tissue, and the effects of conditioned medium from these cells on epithelial cell proliferation, migration and invasion. GLP-2 stimulated proliferation, migration and invasion of myofibroblasts and the proliferative and invasive responses of cancer-associated myofibroblasts were greater than those of myofibroblasts from adjacent tissue. The responses were inhibited by an IGF receptor inhibitor, AG1024. Conditioned medium from GLP-2 treated myofibroblasts increased proliferation, migration and invasion of SW480, HT29, LoVo epithelial cells and these responses were inhibited by AG1024; GLP-2 alone had no effect on these cells. In addition, when myofibroblasts and epithelial cells were co-cultured in Ibidi chambers there was mutual stimulation of migration in response to GLP-2. The latter increased both IGF-1 and IGF-2 transcript abundance in myofibroblasts. Moreover, a number of IGF binding proteins (IGFBP-4, −5, −7) were identified in myofibroblast medium; in the presence of GLP-2 there was increased abundance of the cleavage products of IGBBP-4 and IGFBP-5 suggesting activation of a degradation mechanism that might increase IGF bioavailability. The data suggest that GLP-2 stimulates cancer myofibroblast proliferation, migration and invasion; GLP-2 acts indirectly on epithelial cells partly via increased IGF expression in myofibroblasts and partly, perhaps, by increased bioavailability through degradation of IGFBPs.
Keywords: GLP-2; IGF; IGFBP; Myofibroblast migration; Proliferation;

Adiponectin: Serum-saliva associations and relations with oral and systemic markers of inflammation by Jenna L. Riis; Crystal I. Bryce; Thao Ha; Tracey Hand; John L. Stebbins; Marla Matin; Katrin M. Jaedicke; Douglas A. Granger (58-64).
This study addresses gaps in our understanding about the validity and utility of using salivary adiponectin to index serum adiponectin levels. Matched blood and saliva samples were collected on a single occasion from healthy adults (n  = 99; age 18–36 years, 53% male). Serum and saliva was assayed for adiponectin and inflammatory cytokines (IL-1β, IL-6, IL-8, TNFα), and saliva was also assayed for markers of blood contamination (transferrin), total protein (salivary flow rate) and matrix metalloproteinase-8 (MMP-8). We examined the extent to which salivary adiponectin was associated with serum adiponectin, and the influence of potential confounders on the serum-saliva correlation, including age, sex, body mass index, and markers of inflammation, oral health, salivary blood contamination, and flow rate. Findings revealed a modest serum-saliva association for adiponectin, and strong positive associations between salivary adiponectin and salivary levels of inflammatory cytokines, MMP-8, transferrin, and total protein. By contrast, salivary adiponectin was not related to serum levels of inflammatory activity. The magnitude of the serum-saliva association was strengthened when controlling for total protein in saliva, blood leakage into oral fluid, salivary inflammatory cytokines, and MMP-8. The pattern of findings extends our understanding of salivary adiponectin and its potential use as an index of circulating adiponectin levels.
Keywords: Saliva; Serum; Adiponectin; Cytokines; Inflammation; Oral health;