Retrospective analysis of insulin responses to standard dosed oral glucose tests (OGTs) via naso-gastric tubing towards definition of an objective cut-off value

Abstract

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.

Keywords: ELISA; Equine metabolic syndrome; Horse; Insulin; Insulin dysregulation; Naso-gastric tubing; Oral glucose test.

Fig. 1

Serum insulin concentrations during oral glucose test (OGT), n = 56. Model based clustering algorithm detected an intrinsic structure to the data and grouped individuals based on their similarities in all insulin measurements in an unsupervised manner. This figure shows the two clusters detected by the algorithm (cluster 1—dark blue; cluster 2—light blue). The calculated limit between both clusters at 120 min was 105 µLU/mL (red lines)

Fig. 2

Principal component analysis (PCA) plot of the two clusters. A PCA plot is a 2D representation of high-dimensional data. In this case, the dimensions consist in measurements of serum insulin at different time points of the oral glucose test (OGT). Data-points that are close show a similar insulin response during the OGT (cluster 1—dark blue points; cluster 2—light blue triangles)

Fig. 3

Pseudomedian (solid line) and 95% confidence interval (dashed line) of the insulin response in oral glucose test (OGT) for both clusters (cluster 1—dark blue; cluster 2—light blue). As the distribution of the insulin levels in each cluster for each time-point are not normal, the median was chosen as a better representation of how the insulin response differs between the clusters

Fig. 4

Median serum insulin concentration (solid line) of the cluster 1 classified as insulin sensitive during oral glucose test (OGT) with calculated 2.5 and 97.5% quantile (dashed lines). Calculated cut-off at 120 min is 110 µLU/mL (grey lines)