Multi-parameter comparison of a standardized mixed meal tolerance test in healthy and type 2 diabetic subjects: the PhenFlex challenge

Abstract

Background: A key feature of metabolic health is the ability to adapt upon dietary perturbations. Recently, it was shown that metabolic challenge tests in combination with the new generation biomarkers allow the simultaneous quantification of major metabolic health processes. Currently, applied challenge tests are largely non-standardized. A systematic review defined an optimal nutritional challenge test, the "PhenFlex test" (PFT). This study aimed to prove that PFT modulates all relevant processes governing metabolic health thereby allowing to distinguish subjects with different metabolic health status. Therefore, 20 healthy and 20 type 2 diabetic (T2D) male subjects were challenged both by PFT and oral glucose tolerance test (OGTT). During the 8-h response time course, 132 parameters were quantified that report on 26 metabolic processes distributed over 7 organs (gut, liver, adipose, pancreas, vasculature, muscle, kidney) and systemic stress.

Results: In healthy subjects, 110 of the 132 parameters showed a time course response. Patients with T2D showed 18 parameters to be significantly different after overnight fasting compared to healthy subjects, while 58 parameters were different in the post-challenge time course after the PFT. This demonstrates the added value of PFT in distinguishing subjects with different health status. The OGTT and PFT response was highly comparable for glucose metabolism as identical amounts of glucose were present in both challenge tests. Yet the PFT reports on additional processes, including vasculature, systemic stress, and metabolic flexibility.

Conclusion: The PFT enables the quantification of all relevant metabolic processes involved in maintaining or regaining homeostasis of metabolic health. Studying both healthy subjects and subjects with impaired metabolic health showed that the PFT revealed new processes laying underneath health. This study provides the first evidence towards adopting the PFT as gold standard in nutrition research.

Keywords: Biomarkers; Metabolic health; Nutrition; Nutritional challenge test; Phenotypic flexibility; Type 2 diabetes.

Fig. 1

Overview of study design and time points at which blood samples were collected in response to challenge tests for biomarker analysis. Twenty healthy male volunteers and 20 T2D subjects were given the PhenFlex test (PFT) drink or a glucose drink (OGTT) in randomized order on two different study days. The wash-out period between the two challenge tests was at least 2 days. On study days, blood samples were taken at t = 0 (≥ 10 h of fasting) and six time points (t = 0.5, t = 1, t = 2, t = 4, t = 6, and t = 8 h) after consumption of the challenge drinks for analysis of a total of 132 different metabolic markers

Fig. 2

Overview of processes that are modulated by the PhenFlex test based on biomarker response in n = 20 healthy male subjects. Black: modulated by PFT; gray: not responding

Fig. 3

The five different observed time course clusters in response to the PhenFlex test, based on the 110 plasma metabolites and proteins with a significant effect in time. The red line represents the average cluster time profile. The x-axes were expressed as time (minutes), the y-axes were expressed as relatively scaled concentrations. Parameters from cluster 1 increased during most of the 8-h time course. Parameters from cluster 2 decreased during most of the time course of 8 h. Parameters from clusters 3 decreased upon PFT, with a subsequent recovery phase. Parameters from cluster 4 showed a classical absorption profile. Parameters from cluster 5 reached maximum values around 1 h and minimum concentrations were reached at the final time point (8 h). Finally, the different average cluster curves are summarized in one figure

Fig. 4

Overview of markers that have a different PhenFlex test response between 20 healthy male and 20 male type 2 diabetic patients. Gray = no significant differences between T2D and healthy subjects; black = significant different postprandial levels between healthy and diabetic subjects; bold black = significantly different responses to PhenFlex challenge between healthy and type 2 diabetics; asterisk = significant different fasting levels

Fig. 5

a Average glucose response in OGTT vs PhenFlex test in 20 diabetic type 2 male patients and 20 healthy male volunteers. b Average insulin response in OGTT vs. PhenFlex test in 20 diabetic type 2 male patients and 20 healthy male volunteers. Red = average diabetes type 2 response. Blue = average healthy response. Solid line = PhenFlex test. Broken line = OGTT