Transient antibiotic-induced changes in the neonatal swine intestinal microbiota impact islet expression profiles reducing subsequent function.

Neonatal antibiotics administered to human infants initiate gut microbiota dysbiosis that may have long-term effects on body weight and metabolism. We examined antibiotic-induced adaptations in pancreatic islets of the piglet, a well-accepted model of human infant microbiota and pancreas development. Neonatal piglets randomized to amoxicillin [30 mg/kg body wt/day; n = 7, antibiotic (ANTI)] or placebo [vehicle control; n = 7, control (CON)] from postnatal day (PND)0-13 were euthanized at PND7, 14, and 49. The metabolic phenotype along with functional, immunohistological, and transcriptional phenotypes of the pancreatic islets were studied. The gut microbiome was characterized by 16S rRNA gene sequencing, and microbial metabolites and microbiome-sensitive host molecules were measured. Compared with CON, ANTI PND7 piglets had elevated transcripts of genes involved in glucagon-like peptide 1 ((GLP-1) synthesis or signaling in islets (P < 0.05) coinciding with higher plasma GLP-1 (P = 0.11), along with increased tumor necrosis factor α (Tnf) (P < 0.05) and protegrin 1 (Npg1) (P < 0.05). Antibiotic-induced relative increases in Escherichia, Coprococcus, Ruminococcus, Dehalobacterium, and Oscillospira of the ileal microbiome at PND7 normalized after antibiotic withdrawal. In ANTI islets at PND14, the expression of key regulators pancreatic and duodenal homeobox 1 (Pdx1), insulin-like growth factor-2 (Igf2), and transcription factor 7-like 2 (Tcf7l2) was downregulated, preceding a 40% reduction of ß-cell area (P < 0.01) and islet insulin content at PND49 (P < 0.05). At PND49, a twofold elevated plasma insulin concentration (P = 0.07) was observed in ANTI compared with CON. We conclude that antibiotic treatment of neonatal piglets elicited gut microbial changes accompanied by phasic alterations in key regulatory genes in pancreatic islets at PND7 and 14. By PND49, reduced ß-cell area and islet insulin content were accompanied by elevated nonfasted insulin despite normoglycemia, indicative of islet stress.

Agonists and protein kinase C-activation induce phosphorylation and internalization of FFA1 receptors.

FFA1 (previously known as GPR40) is a free fatty acid receptor involved in the regulation of inflammatory processes and insulin secretion. The cellular actions resulting from FFA1 activation have received considerable attention. However, little is known on the regulation of the receptor function. In the present work, using cells transfected with this receptor, docosahexaenoic acid and α-linolenic acid increased intracellular calcium concentration and ERK 1/2 phosphorylation. It was also observed that FFA1 is a phosphoprotein whose phosphorylation state was increased (2- to 3-fold) by agonists (i.e., free fatty acids) and also by phorbol myristate acetate. Agonist- and phorbol ester-mediated FFA1 phosphorylation was markedly reduced by inhibitors of protein kinase C. Receptor stimulation by free fatty acids and protein kinase C activation also induced receptor internalization as evidenced by confocal microscopy. In summary, our data show that FFA1 is a phosphoprotein whose phosphorylation state is modulated by agonists and protein kinase C activation; such covalent modification is associated with receptor internalization.