Effect of Dietary Nonstructural Carbohydrate Content on Activation of 5′‐Adenosine Monophosphate‐Activated Protein Kinase in Liver, Skeletal Muscle, and Digital Laminae of Lean and Obese Ponies

Abstract

BackgroundIn EMS‐associated laminitis, laminar failure may occur in response to energy failure related to insulin resistance (IR) or to the effect of hyperinsulinemia on laminar tissue. 5′‐Adenosine‐monophosphate‐activated protein kinase (AMPK) is a marker of tissue energy deprivation, which may occur in IR.Hypothesis/ObjectivesTo characterize tissue AMPK regulation in ponies subjected to a dietary carbohydrate (CHO) challenge.AnimalsTwenty‐two mixed‐breed ponies.MethodsImmunohistochemistry and immunoblotting for total AMPK and phospho(P)‐AMPK and RT‐qPCR for AMPK‐responsive genes were performed on laminar, liver, and skeletal muscle samples collected after a 7‐day feeding protocol in which ponies stratified on body condition score (BCS; obese or lean) were fed either a low‐CHO diet (ESC + starch, approximately 7% DM; n = 5 obese, 5 lean) or a high‐CHO diet (ESC + starch, approximately 42% DM; n = 6 obese, 6 lean).Results5′‐Adenosine‐monophosphate‐activated protein kinase was immunolocalized to laminar keratinocytes, dermal constituents, and hepatocytes. A high‐CHO diet resulted in significantly decreased laminar [P‐AMPK] in lean ponies (P = .03), but no changes in skeletal muscle (lean, P = .33; obese, P = .43) or liver (lean, P = .84; obese, P = .13) [P‐AMPK]. An inverse correlation existed between [blood glucose] and laminar [P‐AMPK] in obese ponies on a high‐CHO diet.Conclusions and Clinical ImportanceLaminar tissue exhibited a normal response to a high‐CHO diet (decreased [P‐AMPK]), whereas this response was not observed in liver and skeletal muscle in both lean (skeletal muscle, P = .33; liver, P = .84) and obese (skeletal muscle, P = .43; liver, P = .13) ponies.

Continuous digital hypothermia prevents lamellar failure in the euglycaemic hyperinsulinaemic clamp model of equine laminitis

“…The association between insulin dysregulation and laminitis has been well established but the exact pathophysiology remains unclear. Proposed mechanisms include lamellar ischaemia [5,23,24], energy failure at the level of the lamellar epithelial cell [25] and activation of growth factor signalling pathways via IGF-1R in lamellar epithelial cells [12,26]. Following the discovery that hyperinsulinaemia can cause laminitis in the absence of insulin resistance in the EHC model [7,10], there has been growing recognition of the role of hyperinsulinaemia in naturally-occurring cases [1,2,21].…”

Association of sustained supraphysiologic hyperinsulinemia and inflammatory signaling within the digital lamellae in light‐breed horses

“…[24][25][26][27] Laminitis that occurs in equine patients with systemic insulin dysregulation and hyperinsulinemia is likely to be associated with disordered nutrient sensing and metabolism within the digital lamellae. [28][29][30] Molecular mechanisms for nutrient and energetic sensing are highly conserved among eukaryotic organisms 31 ; 2 centrally important signaling pathways linking nutrient substrate flux to cellular processes (such as differentiation, proliferation, intercellular adhesion, inflammation, etc.) are mammalian target of rapamycin complex-1 (mTORC1) signaling 24,32 and adenosine 5 0 -monophosphate-activated protein kinase (AMPK) signaling.…”

Laminar inflammatory events in lean and obese ponies subjected to high carbohydrate feeding: Implications for pasture‐associated laminitis

“…their laminae did not contain higher concentrations of cytokine mRNA or leucocytes than nonlaminitic individuals with the same diet exposure). Finally, additional investigations by our laboratory into the regulation of several signalling pathways within the digital laminae of these ponies suggest that this dietary model does affect laminar metabolism; increased dietary NSC content resulted in decreased phosphorylation of 5′‐adenosine monophosphate activated protein kinase (AMPK) and markedly increased phosphorylation of RPS6 (a protein downstream of growth factor signalling) in the laminae, events which may be involved in the degree of adherence of laminar basal epithelial cells (LBEC) to their basement membrane .…”

“…overdose of pentobarbital sodium and phenytoin sodium (Fatal‐Plus d ; 20 mg/kg bwt i.v.). Immunohistochemistry of laminar tissue was performed as previously described . Briefly, laminar tissue sections were deparaffinised and incubated in either a mouse monoclonal anti‐CD163 primary antibody e (Cosmo; 1:100 ), a mouse monoclonal anti‐calprotectin primary antibody f (1:100 ), or a goat polyclonal anti‐cyclo‐oxygenase‐2 (COX‐2) primary antibody g (1:100 ).…”