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Search for other papers by S. Poole in
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ABSTRACT
Activated cells of the monocyte—macrophage lineage produce two forms of the inflammatory cytokine interleukin-1 (IL-1), IL-1α and IL-1β, of which IL-1β is the predominant secreted form and has a wide range of modulatory effects on the endocrine system. Immunoassays of human IL-1β have been described, but are not suitable for measurement of rat and mouse IL-1β because of limited cross-reactivity. Polyclonal sheep anti-rat or sheep anti-mouse IL-1β antisera were used to develop sensitive and specific immunoradiometric assays for rat and mouse IL-1β. Secretion of IL-1β from endotoxin-activated monocytes or macrophages was measured in vitro or in vivo in both species. In vitro, rat monocytes and mouse macrophages produced IL-1β in response to endotoxin, with a relatively small proportion of total IL-1β being secreted. In vivo, endotoxin stimulated an increase in plasma IL-1β in both animals. The development of these assays will facilitate studies of the role of endogenous IL-1β in animal endocrine models.
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Search for other papers by A F Bristow in
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ABSTRACT
Regulation of a number of aspects of the acute-phase response, including induction of fever and activation of the hypothalamo-pituitary-adrenal axis, occurs within the hypothalamus. The acute-phase response appears to be co-ordinated by the inflammatory cytokine interleukin-1 (IL-1). A number of studies using hybridization techniques to measure IL-1 gene expression and immunocyto-chemistry to localize immunoactive IL-1 have established the concept that the central nervous system, and in particular the hypothalamus, is a site of IL-1 production, and that levels increase in response to inflammatory stimuli. In this report we present data on the levels of IL-1β produced in the rat hypothalamus using quantitative immunoassay techniques. Bacterial endotoxin, administered to rats in vivo, evoked increases in hypothalamic IL-1β levels which were significant within 1 h, and reached maximum levels at 5–10 h. The response to endotoxin was dose-related, and levels reached in hypothalamic extracts corresponded to intra-hypothalamic levels of the order of 20 ng/ml. During short-term in-vitro culture of rat hypothalami, endotoxin stimulated a dose-related increase in both the synthesis and the secretion of IL-1β, which reached similar levels to those seen after in-vivo stimulation. Hypothalami obtained from animals stimulated with endotoxin in vivo did not, however, show any evidence of persistent stimulation of IL-1β production when subsequently cultured in vitro.
These data support the concept that production of hypothalamic IL-1 is an essential step in regulating the activity of the hypothalamus during the acute-phase response, and provide for the first time quantitative data on the magnitude, dose—response relationships and time-courses of rat hypothalamic IL-1β production in vivo and in vitro.
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Search for other papers by Daniel P Poole in
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Department of Biochemistry and Molecular Biology, University of Melbourne, Melbourne, Victoria, Australia
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Department of Biochemistry and Molecular Biology, University of Melbourne, Melbourne, Victoria, Australia
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Department of Pharmacology, Monash University, Clayton, Victoria, Australia
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Search for other papers by Roger J Summers in
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Insulin-like peptide 5 (INSL5) is a newly discovered gut hormone expressed in colonic enteroendocrine L-cells but little is known about its biological function. Here, we show using RT-qPCR and in situ hybridisation that Insl5 mRNA is highly expressed in the mouse colonic mucosa, colocalised with proglucagon immunoreactivity. In comparison, mRNA for RXFP4 (the cognate receptor for INSL5) is expressed in various mouse tissues, including the intestinal tract. We show that the human enteroendocrine L-cell model NCI-H716 cell line, and goblet-like colorectal cell lines SW1463 and LS513 endogenously express RXFP4. Stimulation of NCI-H716 cells with INSL5 produced phosphorylation of ERK1/2 (Thr202/Tyr204), AKT (Thr308 and Ser473) and S6RP (Ser235/236) and inhibited cAMP production but did not stimulate Ca2+ release. Acute INSL5 treatment had no effect on GLP-1 secretion mediated by carbachol or insulin, but modestly inhibited forskolin-stimulated GLP-1 secretion in NCI-H716 cells. However, chronic INSL5 pre-treatment (18 h) increased basal GLP-1 secretion and prevented the inhibitory effect of acute INSL5 administration. LS513 cells were found to be unresponsive to INSL5 despite expressing RXFP4. Another enteroendocrine L-cell model, mouse GLUTag cells did not express detectable levels of Rxfp4 and were unresponsive to INSL5. This study provides novel insights into possible autocrine/paracrine roles of INSL5 in the intestinal tract.
