Corrigendum: Exogenous Ketone Supplements Reduce Anxiety-Related Behavior in Sprague-Dawley and Wistar Albino Glaxo/Rijswijk Rats.

[This corrects the article on p. 137 in vol. 9, PMID: 27999529.].

Exogenous Ketone Supplements Reduce Anxiety-Related Behavior in Sprague-Dawley and Wistar Albino Glaxo/Rijswijk Rats.

Nutritional ketosis has been proven effective for seizure disorders and other neurological disorders. The focus of this study was to determine the effects of ketone supplementation on anxiety-related behavior in Sprague-Dawley (SPD) and Wistar Albino Glaxo/Rijswijk (WAG/Rij) rats. We tested exogenous ketone supplements added to food and fed chronically for 83 days in SPD rats and administered sub-chronically for 7 days in both rat models by daily intragastric gavage bolus followed by assessment of anxiety measures on elevated plus maze (EPM). The groups included standard diet (SD) or SD + ketone supplementation. Low-dose ketone ester (LKE; 1,3-butanediol-acetoacetate diester, ~10 g/kg/day, LKE), high dose ketone ester (HKE; ~25 g/kg/day, HKE), beta-hydroxybutyrate-mineral salt (ßHB-S; ~25 g/kg/day, KS) and ßHB-S + medium chain triglyceride (MCT; ~25 g/kg/day, KSMCT) were used as ketone supplementation for chronic administration. To extend our results, exogenous ketone supplements were also tested sub-chronically on SPD rats (KE, KS and KSMCT; 5 g/kg/day) and on WAG/Rij rats (KE, KS and KSMCT; 2.5 g/kg/day). At the end of treatments behavioral data collection was conducted manually by a blinded observer and with a video-tracking system, after which blood ßHB and glucose levels were measured. Ketone supplementation reduced anxiety on EPM as measured by less entries to closed arms (sub-chronic KE and KS: SPD rats and KSMCT: WAG/Rij rats), more time spent in open arms (sub-chronic KE: SPD and KSMCT: WAG/Rij rats; chronic KSMCT: SPD rats), more distance traveled in open arms (chronic KS and KSMCT: SPD rats) and by delayed latency to entrance to closed arms (chronic KSMCT: SPD rats), when compared to control. Our data indicates that chronic and sub-chronic ketone supplementation not only elevated blood ßHB levels in both animal models, but reduced anxiety-related behavior. We conclude that ketone supplementation may represent a promising anxiolytic strategy through a novel means of inducing nutritional ketosis.

Effects of exogenous ketone supplementation on blood ketone, glucose, triglyceride, and lipoprotein levels in Sprague-Dawley rats.

BACKGROUND: Nutritional ketosis induced by the ketogenic diet (KD) has therapeutic applications for many disease states. We hypothesized that oral administration of exogenous ketone supplements could produce sustained nutritional ketosis (>0.5 mM) without carbohydrate restriction. METHODS: We tested the effects of 28-day administration of five ketone supplements on blood glucose, ketones, and lipids in male Sprague-Dawley rats. The supplements included: 1,3-butanediol (BD), a sodium/potassium ß-hydroxybutyrate (ßHB) mineral salt (BMS), medium chain triglyceride oil (MCT), BMS + MCT 1:1 mixture, and 1,3 butanediol acetoacetate diester (KE). Rats received a daily 5-10 g/kg dose of their respective ketone supplement via intragastric gavage during treatment. Weekly whole blood samples were taken for analysis of glucose and ßHB at baseline and, 0.5, 1, 4, 8, and 12 h post-gavage, or until ßHB returned to baseline. At 28 days, triglycerides, total cholesterol and high-density lipoprotein (HDL) were measured. RESULTS: Exogenous ketone supplementation caused a rapid and sustained elevation of ßHB, reduction of glucose, and little change to lipid biomarkers compared to control animals. CONCLUSIONS: This study demonstrates the efficacy and tolerability of oral exogenous ketone supplementation in inducing nutritional ketosis independent of dietary restriction.

Demonstration of the rat ischemic skin wound model.

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and unrelieved pressure due to immobility. Animal models have been developed that attempt to mimic these conditions for the purpose of furthering our understanding of the complexity of chronic wounds. The model described herein is a rat ischemic skin flap model that permits a prolonged reduction of blood flow resulting in wounds that become ischemic and resemble a chronic wound phenotype (reduced vascularization, increased inflammation and delayed wound closure). It consists of a bipedicled dorsal flap with 2 ischemic wounds placed centrally and 2 non-ischemic wounds lateral to the flap as controls. A novel addition to this ischemic skin flap model is the placement of a silicone sheet beneath the flap that functions as a barrier and a splint to prevent revascularization and reduce contraction as the wounds heal. Despite the debate of using rats for wound healing studies due to their quite distinct anatomic and physiologic differences compared to humans (i.e., the presence of a panniculus carnosus muscle, short life-span, increased number of hair follicles, and their ability to heal infected wounds) the modifications employed in this model make it a valuable alternative to previously developed ischemic skin flap models.

Critical role of 5-lipoxygenase and heme oxygenase-1 in wound healing.

Lipid mediators derived from 5-lipoxygenase (5-LO) metabolism can activate both pro- and anti-inflammatory pathways, but their role in wound healing remains largely unexplored. In this study we show that 5-LO knockout (5-LO(-/-)) mice exhibited faster wound healing than wild-type (WT) animals, and exhibited upregulation of heme oxygenase-1 (HO-1). Furthermore, HO-1 inhibition in 5-LO(-/-) mice abolished the beneficial effect observed. Despite the fact that 5-LO(-/-) mice exhibited faster healing, in in vitro assays both migration and proliferation of human dermal fibroblasts (HDFs) were inhibited by the 5-LO pharmacologic inhibitor AA861. No changes were observed in the expression of fibronectin, transforming growth factor (I and III), and α-smooth muscle actin (α-SMA). Interestingly, AA861 treatment significantly decreased ROS formation by stimulated fibroblasts. Similar to 5-LO(-/-) mice, induction of HO-1, but not superoxide dismutase-2 (SOD-2), was also observed in response to 5-LO (AA861) or 5-LO activating protein (MK886) inhibitors. HO-1 induction was independent of nuclear factor (erythroid derived-2) like2 (Nrf-2), cyclooxygenase 2 (COX-2) products, or lipoxin action. Taken together, our results show that 5-LO disruption improves wound healing and alters fibroblast function by an antioxidant mechanism based on HO-1 induction. Overexpression of HO-1 in wounds may facilitate early wound resolution.