Anna K. Bassil

Growth Hormone Secretagogue Receptors in Rat and Human Gastrointestinal Tract and the Effects of Ghrelin

The peptide hormone ghrelin is known to be present within stomach and, to a lesser extent, elsewhere in gut. Although reports suggest that gastric function may be modulated by ghrelin acting via the vagus nerve, the gastrointestinal distribution and functions of its receptor, the growth hormone secretagogue receptor (GHS-R), are not clear and may show signs of species-dependency. This study sought to determine the cellular localisation and distribution of GHS-R-immunoreactivity (-Ir) using immunofluorescent histochemistry and explore the function of ghrelin in both human and rat isolated gastric and/or colonic circular muscle preparations in which nerve-mediated responses were evoked by electrical field stimulation. The expression of GHS-R-Ir differed to a greater extent between species than between gut regions of the same species. Both the human and rat gastric and colonic preparations (n=3 each) expressed GHS-R-Ir within neuronal cell bodies and fibres, cells associated with gastric glands and putative entero-endocrine and/or mast cells. Smooth muscle cells and epithelia were devoid of GHS-R-Ir and only rat preparations expressed GHS-R-Ir on nerve fibres associated with the muscle layers. GHS-R-Ir was fully competed in all cases in pre-adsorption studies and antiserum specificity was confirmed using a cell line transiently expressing the rat GHS-R. In rat isolated forestomach circular muscle, ghrelin 0.1-10 microM had no effect on smooth muscle tension but concentration-dependently facilitated the amplitude of contractions evoked by excitatory nerve stimulation (n=4-7; P<0.05 for each concentration versus vehicle; n=18). When examined under similar conditions, in both rat distal colon (n=4-6, P>0.05 each) and human ascending (n=3) and sigmoid (n=1) colon, these concentrations of ghrelin were without effect (P>0.05 each). The data suggest that ghrelin has the potential to profoundly affect gastrointestinal functions in both species and at least one of these functions is to exert a gastric prokinetic activity. Moreover, we suggest that this activity of ghrelin is mediated via the enteric nervous system, in addition to known vagus nerve-dependent mechanisms.

Little or no ability of obestatin to interact with ghrelin or modify motility in the rat gastrointestinal tract

Obestatin, encoded by the ghrelin gene may inhibit gastrointestinal (GI) motility. This activity was re‐investigated.

The relationship between the effects of short‐chain fatty acids on intestinal motility in vitro and GPR43 receptor activation

Abstract  The G protein‐coupled receptors, GPR41 and GPR43, are activated by short‐chain fatty acids (SCFAs), with distinct rank order potencies. This study investigated the possibility that SCFAs modulate intestinal motility via these receptors. Luminal SCFA concentrations within the rat intestine were greatest in the caecum (c. 115 mmol L−1) and proximal colon. Using similar concentrations (0.1–100 mmol L−1), SCFAs were found to inhibit electrically evoked, neuronally mediated contractions of rat distal colon, possibly via a prejunctional site of action; this activity was independent of the presence or absence of the mucosa. By contrast, SCFAs reduced the amplitude but also reduced the threshold and increased the frequency of peristaltic contractions in guinea‐pig terminal ileum. In each model, the rank‐order of activity was acetate (C2) ≈ propionate (C3) ≈ butyrate (C4) > pentanoate (C5) ∼ formate (C1), consistent with activity at the GPR43 receptor. GPR43 mRNA was expressed throughout the rat gut, with highest levels in the colon. However, the ability of SCFAs to inhibit neuronally mediated contractions of the colon was similar in tissues from wild‐type and GPR43 gene knockout mice, with identical rank‐orders of potency. In conclusion, SCFAs can modulate intestinal motility, but these effects can be independent of the GPR43 receptor.

Potent antimyeloma activity of the novel bromodomain inhibitors I-BET151 and I-BET762.

The bromodomain and extraterminal (BET) protein BRD2-4 inhibitors hold therapeutic promise in preclinical models of hematologic malignancies. However, translation of these data to molecules suitable for clinical development has yet to be accomplished. Herein we expand the mechanistic understanding of BET inhibitors in multiple myeloma by using the chemical probe molecule I-BET151. I-BET151 induces apoptosis and exerts strong antiproliferative effect in vitro and in vivo. This is associated with contrasting effects on oncogenic MYC and HEXIM1, an inhibitor of the transcriptional activator P-TEFb. I-BET151 causes transcriptional repression of MYC and MYC-dependent programs by abrogating recruitment to the chromatin of the P-TEFb component CDK9 in a BRD2-4-dependent manner. In contrast, transcriptional upregulation of HEXIM1 is BRD2-4 independent. Finally, preclinical studies show that I-BET762 has a favorable pharmacologic profile as an oral agent and that it inhibits myeloma cell proliferation, resulting in survival advantage in a systemic myeloma xenograft model. These data provide a strong rationale for extending the clinical testing of the novel antimyeloma agent I-BET762 and reveal insights into biologic pathways required for myeloma cell proliferation.

Activation of prostaglandin EP receptors by lubiprostone in rat and human stomach and colon

Background and purpose: Lubiprostone (Amitiza), a possible ClC‐2 channel opener derived from prostaglandin E1 and indicated for the treatment of constipation, increases chloride ion transport and fluid secretion into the intestinal lumen. As lubiprostone may also directly modulate gastrointestinal motility, we investigated its actions and the possible involvement of prostaglandin EP receptor activation on rat and human isolated gastrointestinal preparations.

5-HT4 receptor agonists enhance both cholinergic and nitrergic activities in human isolated colon circular muscle

Abstract  Previous studies have demonstrated mixed inhibitory and facilitatory effects of 5‐hydroxytryptamine‐4 (5‐HT4) receptor agonists on electrical field stimulation (EFS)‐induced responses in human isolated colon. Here we report three types of responses to EFS in human isolated colon circular muscle: monophasic cholinergic contraction during EFS, biphasic response (nitrergic relaxation during EFS followed by cholinergic contraction after termination of EFS) and triphasic response (cholinergic contraction followed by nitrergic relaxation during EFS and a tachykininergic contraction after EFS). The effects of two 5‐HT4 receptor agonists, prucalopride and tegaserod were then investigated on monophasic responses only. Each compound inhibited contractions during EFS in a concentration‐dependent manner. In the presence of Nω‐nitro‐l‐arginine methyl ester (l‐NAME) however, prucalopride and tegaserod enhanced the contractions in a concentration‐dependent manner. In strips where the tone was elevated with substance‐P and treated with scopolamine, EFS‐induced relaxations were enhanced by the two agonists. The above observed effects by the two agonists were abolished by 5‐HT4 receptor antagonist SB‐204070. The two agonists did not alter the tone raised by substance‐P in the presence of scopolamine and l‐NAME and did not affect carbachol‐induced contractions in the presence of tetrodotoxin. These results suggest that in the circular muscle of human colon, 5‐HT4 receptor agonists simultaneously facilitate the activity of neurones which release the inhibitory and excitatory neurotransmitters, nitric oxide and acetylcholine respectively.

Differences between the abilities of tegaserod and motilin receptor agonists to stimulate gastric motility in vitro

Motilin or 5‐HT4 receptor agonists stimulate gastrointestinal motility. Differences in activity are suggested but direct comparisons are few. A method was devised to directly compare the gastric prokinetic activities of motilin, the motilin receptor agonist, erythromycin, and the 5‐HT4 receptor agonist, tegaserod.

UDP-glucose modulates gastric function through P2Y14 receptor-dependent and -independent mechanisms

P2Y receptors have been reported to modulate gastrointestinal functions. The newest family member is the nucleotide-sugar receptor P2Y14. P2ry14 mRNA was detected throughout the rat gut, with the highest level being in the forestomach. We investigated the role of the receptor in stomach motility using cognate agonists and knockout (KO) mice. In rat isolated forestomach, 100 microM UDP-glucose and 100 muM UDP-galactose both increased the baseline muscle tension (BMT) by 6.2+/-0.6 and 1.6+/-0.6 mN (P<0.05, n=3-4), respectively, and the amplitude of contractions during electrical field stimulation (EFS) by 3.7+/-1.7 and 4.3+/-2.5 mN (P<0.05, n=3-4), respectively. In forestomach from wild-type (WT) mice, 100 microM UDP-glucose increased the BMT by 1.0+/-0.1 mN (P<0.05, n=6) but this effect was lost in the KO mice (change of -0.1+/-0.1 mN, n=6). The 100 microM UDP-glucose also increased the contraction amplitude during EFS in this tissue from the WT animals (0.9+/-0.4 mN, P < 0.05, n=6) but not from the KO mice (0.0+/-0.2 mN, n=6). In vivo, UDP-glucose at 2,000 mg/kg ip reduced gastric emptying in rats by 49.7% (P<0.05, n=4-6) and in WT and KO mice by 56.1 and 66.2%, respectively (P<0.05, n=7-10) vs. saline-treated control animals. There was no significant difference in gastric emptying between WT and KO animals receiving either saline or d-glucose. These results demonstrate a novel function of the P2Y14 receptor associated with contractility in the rodent stomach that does not lead to altered gastric emptying after receptor deletion and an ability of UDP-glucose to delay gastric emptying without involving the P2Y14 receptor.

Neuromedin U can exert colon‐specific, enteric nerve‐mediated prokinetic activity, via a pathway involving NMU1 receptor activation

The neuromedin U (NMU) receptors, NMU1 and NMU2, are expressed in the gut but their functions are unclear. This study explores the role of NMU in gastrointestinal motility.

Inhibition of colonic motility and defecation by RS-127445 suggests an involvement of the 5-HT2B receptor in rodent large bowel physiology

Background:  5‐HT2B receptors are localized within the myenteric nervous system, but their functions on motor/sensory neurons are unclear. To explore the role of these receptors, we further characterized the 5‐HT2B receptor antagonist RS‐127445 and studied its effects on peristalsis and defecation.