Autofluorescence Imaging of Living Pancreatic Islets Reveals Fibroblast Growth Factor-21 (FGF21)-Induced Metabolism

Abstract

Fibroblast growth factor-21 (FGF21) has therapeutic potential for metabolic syndrome due to positive effects on fatty acid metabolism in liver and white adipose tissue. FGF21 also improves pancreatic islet survival in excess palmitate; however, much less is known about FGF21-induced metabolism in this tissue. We first confirmed FGF21-dependent activity in islets by identifying expression of the cognate coreceptor Klothoβ, and by measuring a ligand-stimulated decrease in acetyl-CoA carboxylase expression. To further reveal the effect of FGF21 on metabolism, we employed a unique combination of two-photon and confocal autofluorescence imaging of the NAD(P)H and mitochondrial NADH responses while holding living islets stationary in a microfluidic device. These responses were further correlated to mitochondrial membrane potential and insulin secretion. Glucose-stimulated responses were relatively unchanged by FGF21. In contrast, responses to glucose in the presence of palmitate were significantly reduced compared to controls showing diminished NAD(P)H, mitochondrial NADH, mitochondrial membrane potential, and insulin secretion. Consistent with the glucose-stimulated responses being smaller due to continued fatty acid oxidation, mitochondrial membrane potential was increased in FGF21-treated islets by using the fatty acid transport inhibitor etomoxir. Citrate-stimulated NADPH responses were also significantly larger in FGF21-treated islets suggesting preference for citrate cycling rather than acetyl-CoA carboxylase-dependent fatty acid synthesis. Overall, these data show a reduction in palmitate-induced potentiation of glucose-stimulated metabolism and insulin secretion in FGF21-treated islets, and establish the use of autofluorescence imaging and microfluidic devices to investigate cell metabolism in a limited amount of living tissue.

Fibroblast Growth Factor Receptor Like-1 (FGFRL1) Interacts with SHP-1 Phosphatase at Insulin Secretory Granules and Induces Beta-cell ERK1/2 Protein Activation

“…The islets of these mice contained a diminished number of beta-cells with reduced glucose sensitivity and impaired insulin processing, demonstrating the importance of FGF signaling in maintenance of beta-cell physiology and glucose homeostasis. We previously confirmed FGFR1 activity in adult murine pancreatic beta-cells, and we further showed changes in islet glucose-stimulated metabolism and insulin secretion in the presence of the FGFR1-ligand FGF21 (2,22). Overall, these data suggest that FGFR1-induced signaling regulates beta-cell biology via classical intracellular signaling responses and may be associated with the onset and progression of metabolic disorders such as diabetes.…”

“…Pancreatic islets were isolated from 8-to 12-week-old C57BL6 male mice by collagenase digestion (Roche Applied Science) (22). Islets were equilibrated in full RPMI 1640 medium supplemented with 11 mM glucose, 10% FBS, and 5 units/ml penicillin/streptomycin prior to mRNA harvest or dispersion to single cells using Accutase (Sigma) for fixation and immunostaining as described previously (2).…”

The control of insulin secretion by adipokines: current evidence for adipocyte-beta cell endocrine signalling in metabolic homeostasis

“…The mechanism of FGF21 action suggested by this study was via activation of AKT and ERK pathways in beta cells. Another study showed that FGF21 did not affect GSIS from normal mouse islets, but disrupted the potentiating effect of NEFA on GSIS, potentially by altering lipid metabolism and signalling (Sun et al 2012). In addition to a direct effect on beta cells, FGF21 may act indirectly by stimulating secretion of adiponectin from adipocytes (Holland et al 2013;Lin et al 2013) which may in turn stimulate insulin secretion (as reviewed above).…”

“…In adipocytes, FGF21 signals through the fibroblast growth factor receptor 1 (FGFR1) and beta-klotho co-receptor Ogawa et al 2007), and both of these key receptors are expressed in islets (Hart et al 2000;Sun et al 2012). Acute FGF21 administration in mice decreases blood glucose and insulin levels , suggesting a primary role in improving insulin sensitivity.…”

The Pancreatic β-Cell: A Bioenergetic Perspective

“…Leucine enhanced the mitochondrial fluorescence with no significant effect on the cytosol, while 20 mM glucose roughly doubled the autofluorescence in each compartment. Rocheleau et al (501) attempted to distin-guish changes in matrix NADH from total islet NAD(P)H, by simultaneously monitoring lipoamide dehydrogenase (LipDH) flavin autofluorescence. The enzyme is a component of pyruvate and 2-oxoglutarate dehydrogenases, transferring electrons to NAD ϩ .…”