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Background: Insulin dysregulation (ID) with basal or postprandial hyperinsulinemia is one of the key findings in horses and ponies suffering from the equine metabolic syndrome (EMS). Assessment of ID can easily be performed in clinical settings by the use of oral glucose challenge tests. Oral glucose test (OGT) performed with 1 g/kg bodyweight (BW) glucose administered via naso-gastric tube allows the exact administration of a defined glucose dosage in a short time. However, reliable cut-off values have not been available so far. Therefore, the aim of the study was to describe variations in insulin response to OGT via naso-gastric tubing and to provide a clinical useful cut-off value for ID when using the insulin quantification performed with an equine-optimized insulin enzyme-linked immunosorbent assay., Results: Data visualization revealed no clear separation in the serum insulin concentration of insulin sensitive and insulin dysregulated horses during OGT. Therefore, a model based clustering method was used to circumvent the use of an arbitrary limit for categorization. This method considered all data-points for the classification, taking into account the individual insulin trajectory during the OGT. With this method two clusters were differentiated, one with low and one with high insulin responses during OGT. The cluster of individuals with low insulin response was consistently detected, independently of the initialization parameters of the algorithm. In this cluster the 97.5% quantile of insulin is 110 µLU/mL at 120 min. We suggest using this insulin concentration of 110 µLU/mL as a cut-off value for samples obtained at 120 min in OGT., Conclusion: OGT performed with 1 g/kg BW glucose and administration via naso-gastric tubing can easily be performed under clinical settings. Application of the cut-off value of 110 µLU/mL at 120 min allows assessment of ID in horses.

The aim of the study was to analyze key proteins of the equine insulin signaling cascade and their extent of phosphorylation in biopsies from muscle tissue (MT), liver tissue (LT), and nuchal AT, subcutaneous AT, and retroperitoneal adipose tissues. This was investigated under unstimulated (B1) and intravenously insulin stimulated (B2) conditions, which were achieved by injection of insulin (0.1 IU/kg bodyweight) and glucose (150 mg/kg bodyweight). Twelve warmblood horses aged 15 ± 6.8 yr (yr), weighing 559 ± 79 kg, and with a mean body condition score of 4.7 ± 1.5 were included in the study. Key proteins of the insulin signaling cascade were semiquantitatively determined using Western blotting. Furthermore, modulation of the cascade was assessed. The basal expression of the proteins was only slightly influenced during the experimental period. Insulin induced a high extent of phosphorylation of insulin receptor in LT (P < 0.01) but not in MT. Protein kinase B and mechanistic target of rapamycin expressed a higher extent of phosphorylation in all tissues in B2 biopsies. Adenosine monophosphate protein kinase, as a component related to insulin signaling, expressed enhanced phosphorylation in MT (P < 0.05) and adipose tissues (nuchal AT P < 0.05; SCAT P < 0.01; retroperitoneal adipose tissue P < 0.05), but not in LT at B2. Tissue-specific variations in the acute response of insulin signaling to intravenously injected insulin were observed. In conclusion, insulin sensitivity in healthy horses is based on a complex concerted action of different tissues by their variations in the molecular response to insulin., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The aim of the present study was to investigate the effects of butyrate as a feed supplement on the expression of insulin signalling proteins as potent regulators of metabolism and growth in Ross 308 broiler chickens fed maize- or wheat-based diets. Both diets were supplemented with non-protected butyrate (1.5 and 3.0 g/kg of diet, respectively) or with protected butyrate (0.2 g/kg of diet); the diet of the control groups was prepared without any additives (control). On day 42 of life, systemic blood samples were drawn for analyses of glucose and insulin concentrations, and tissue samples (liver, gastrocnemius muscle and subcutaneous adipose tissue) were taken for Western blotting examinations. The expression of key insulin signalling proteins (IRβ, PKCζ and mTOR) was assessed by semiquantitative Western blotting from the tissues mentioned. The type of diet had a remarkable influence on the insulin homeostasis of chickens. The wheat-based diet significantly increased IRβ and mTOR expression in the liver as well as mTOR and PKCζ expression in the adipose tissue when compared to animals kept on a maize-based diet. IRβ expression in the liver was stimulated by the lower dose of non-protected butyrate as well, suggesting the potential of butyrate as a feed additive to affect insulin sensitivity. Based on the results obtained, the present study shows new aspects of nutritional factors by comparing the special effects of butyrate as a feed additive and those of the cereal type, presumably in association with dietary non-starch polysaccharide- (NSP-) driven enteric shortchain fatty acid release including butyrate, influencing insulin homeostasis in chickens. As the tissues of chickens have physiologically lower insulin sensitivity compared to mammals, diet-associated induction of the insulin signalling pathway can be of special importance in improving growth and metabolic health.

The glucose homeostasis in dairy cattle is very well controlled, in line with the metabolic adaptation during the periparturient period. Former studies showed that nicotinic acid (NA) lowered plasma non-esterified fatty acids (NEFA) concentrations and increased insulin sensitivity in dairy cows. Thus, the purpose of this study was to investigate whether the expression of proteins involved in hepatic and adipose insulin signaling and protein expression of hepatic glucose transporter 2 (GLUT2) were affected by dietary NA and dietary concentrate intake in periparturient dairy cows. Twenty pluriparous German Holstein cows were fed with the same diet from about 21 days before the expected calving date (d-21) to calving. After calving, cows were randomly assigned in 4 groups and fed with diets different in concentrate proportion ("HC" with 60:40% or "LC" with 30:70% concentrate-to-roughage ratio) and supplemented with NA (24 g/day) (NA) or without (CON) until d21. Biopsy samples were taken from the liver, subcutaneous (SCAT) and retroperitoneal (RPAT) adipose tissues at d-21 and d21. Protein expression of insulin signaling molecules (insulin receptor (INSR), phosphatidylinositol-3-kinase (PI3K), protein kinase Cζ (PKCζ)) and hepatic GLUT2 was measured by Western Blotting. The ratio of protein expression at d21/at d-21 was calculated and statistically evaluated for the effects of time and diet. Cows in HC had significantly higher dietary energy intake than cows in LC. In RPAT a decrease in PI3K and PKCζ expression was found in all groups, irrespectively of diet. In the liver, the GLUT2 expression was significantly lower in cows in NA compared with cows in CON. In conclusion, insulin signaling might be decreased in RPAT over time without any effect of diet. NA was able to modulate hepatic GLUT2 expression, but its physiological role is unclear.

Influencing the endocrine metabolic regulation of chickens by nutritional factors might provide novel possibilities for improving animal health and productivity. This study was designed to evaluate the impact of dietary cereal type (wheat-based (WB) vs. maize-based (MB) diets), crude protein level (normal (NP) vs. lowered (LP)), and sodium (n-)butyrate (1.5 g/kg diet) supplementation (vs. no butyrate) on the responsiveness of hepatic glucagon receptor (GCGR), insulin receptor beta (IRβ) and mammalian target of rapamycin (mTOR) in the phase of intensive growth of chickens. Liver samples of Ross 308 broiler chickens (Gallus gallus domesticus) were collected on day 21 for quantitative real-time polymerase chain reaction and Western blot analyses. Hepatic GCGR and mTOR gene expressions were up-regulated by WB and LP diet. GCGR and IRβ protein level decreased in groups with butyrate supplementation; however, the quantity of IRβ and mTOR protein increased in WB groups. Based on these data, the applied dietary strategies may be useful tools to modulate hepatic insulin and glucagon signaling of chickens in the period of intensive growth. The obtained results might contribute to the better understanding of glycemic control of birds and increase the opportunity of ameliorating insulin sensitivity, hence, improving the production parameters and the welfare of broilers. [ABSTRACT FROM AUTHOR]

Background: Insulin dysregulation (ID) goes along with lasting or transient hyperinsulinemia able to trigger equine laminitis, a painful and crippling foot condition. Promoting weight loss through dietary changes and physical activity is currently the main option to prevent this disease. This study aimed at describing the relationship between weight variations and the level of ID as determined by oral glucose tests (OGT). Therefore, the insulin response of 19 Icelandic horses to repeated OGTs was retrospectively analysed considering the variations in their body weight. Results: There was a strong linear relationship between variations in body weight and variations in the total insulin response to OGT as approximated by the area under the curve of insulin (p < 0.001). As indicated by a weighted least squares model, the insulin response decreased by 22% for 5% weight loss on average. However some horses did not respond to weight loss with a reduction of their insulin response to OGT. Additionally, a high correlation between 120 min serum insulin concentration and total insulin response was observed (r = 0.96, p < 0.001). Conclusions: The results corroborate that weight loss is effective against ID and allow for a better quantification of the expected improvement of the insulin response after weight loss. However, it is unclear why some horses did not respond as expected. The high correlation between the 120 min insulin concentration and total insulin response suggests that insulin status can be accurately determined and monitored with only few samples in a practical setting. [ABSTRACT FROM AUTHOR]

Different diagnostic tests to determine the insulin sensitivity in horses are commonly used in veterinary practice. However, endocrine and metabolic responses provoked by physiological processes during the respective test procedures are not well described. In the present study, oral glucose tests (OGTs) and combined iv glucose insulin tests (CGITs) were employed under standardized conditions. The OGTs and CGITs were performed in twelve healthy warmblood horses of different sex, age (15±6.5 years), weight (567±81kg) and body condition score (4.8±1.6). Horses were tested under fasting conditions. The OGT was performed with 1g/kgBW glucose administered via nasogastric intubation and CGIT was performed by injection of 150mg/kg BW glucose solution and 0.1IU/kgBW porcine zinc-insulin. Blood samples were taken for three hours at intervals of 15 minutes and were analysed for insulin, glucose, triglyceride, non-esterified fatty acids (NEFA), fructosamine and cortisol concentrations. Glucose concentrations increased in the OGTs and CGITs directly after administration. Insulin concentrations increased significantly in OGTs within 30 minutes and stayed elevated for three hours. Peak concentrations of 493.98±86.84μIU/mL were measured in the CGITs, followed by a continuous decline. Baseline NEFA concentrations varied between individual horses and declined in a comparable manner to similar minimum concentrations of 93.82±53.22μmol/L in OGTs and 91.97±56.89μmol/L in CGITs. Regarding the stress response of the test procedure, cortisol concentrations remained unaffected during CGITs, while the OGT procedure was accompanied by a significant initial rise in cortisol concentrations. To conclude, OGT and CGIT mirror different facets of the metabolic response to a glycemic stimulus, highlighting different aspects of glucose homeostasis and insulin regulation. During the CGIT, insulin dynamics with porcine zinc-insulin differ from insulin dynamics described in reports published previously using short-acting insulins. Furthermore, the antilipolytic effects of insulin during OGTs and CGITs via endogenous secretion or exogenous injection resulted in similar reduction of NEFA concentrations and unaffected triglyceride concentrations. This indicates a saturation of the suppression of lipolysis by insulin with already low concentrations and no induction of re-esterification in liver tissue. [ABSTRACT FROM AUTHOR]

Forkhead box protein O1 (FoxO1) is a transcription factor which promotes hepatic glucose production (HGP) by up-regulating the transcription of gluconeogenic enzymes in monogastric species. The activity of FoxO1 is inhibited by insulin-induced phosphorylation. The aims of the present study were to find associations between FoxO1 expression and variables associated with HGP as affected by feeding regimen in dairy cows during the transition period. Twenty one healthy German Holstein cows were allocated to four groups (LC-CON, HC-CON, LC-NA with 5 cows/group and HC-NA with 6 cows/group, respectively). Cows received 0 (LC-CON and HC-CON) or 24 (LC-NA and HC-NA) g/d nicotinic acid with high (HC) or low (LC) concentrate proportion from -42 days (-41.8 + 4.8; mean + standard deviation) relative to expected calving date (d-42) to d24. Liver biopsy was taken at d-42, 1, 21, and 100. The total protein expression of FoxO1 (tFoxO1) and the extent of phosphorylation of FoxO1 at serine 256 (pFoxO1) were analysed semiquantitatively by Western Blotting. The expression of hepatic mRNA of FoxO1 and seven genes associated with HGP was measured by real-time RT-PCR. Mixed model and Pearson’s correlation were used for statistical evaluation with the level of significance at P<0.05. No dietary effect was observed either on feed intake, energy balance, or on the concentration of blood metabolites. Neither time nor diet affected the expression of FoxO1 total protein and mRNA. A NA × concentrate interaction was found in pFoxO1. However, no corresponding dietary effect was found in the mRNA expression of investigated genes. Different patterns of correlations between FoxO1-related variables and investigated indicators for HGP were found at d21 and 100. The results indicated that the regulation of HGP did not take place on the levels of mRNA and protein expression and the phosphorylation of FoxO1 in dairy cows in early lactation. [ABSTRACT FROM AUTHOR]

Simple Summary: l-carnitine might limit the mitochondrial energy generation from fatty acids, particularly in periods of enhanced energy requirement such as during lipopolysaccharide (LPS) mediated innate immune responses. This study examined the influence of a dietary l-carnitine supplementation on dairy cows in mid-lactation which were challenged by an injection with LPS. Results suggest that this intervention supported the energy metabolism of the cows during LPS-induced acute phase reaction (APR). l-carnitine plays an important role in energy metabolism through supporting the transport of activated fatty acids to the subcellular site of β-oxidation. An acute phase reaction (APR) is known as an energy consuming process. Lipopolysaccharides (LPS) are often used in animal models to study intervention measures during innate immune responses such as APR. Thus, the aim of the study was to investigate the effects of dietary l-carnitine supplementation during an LPS-induced APR in mid-lactating German Holstein cows. Animals were assigned to a control (CON, n = 26) or l-carnitine group (CAR, n = 27, 25 g rumen-protected l-carnitine/cow/d) and received an intravenous injection of LPS (0.5 μg/kg body weight) at day 111 post-partum. Blood samples were collected from day 1 pre-injection until day 14 post-injection (pi). From 0.5 h pi until 72 h pi blood samplings and clinical examinations were performed in short intervals. Clinical signs of the APR were not altered in group CAR except rumen motility which increased at a lower level compared to the CON group after a period of atonia. Group CAR maintained a higher insulin level compared to group CON even up to 72 h pi which might support glucose utilization following an APR. [ABSTRACT FROM AUTHOR]