Introduction
Over the past few decades, the high prevalence of metabolic syndrome has become a worldwide phenomenon, associated with an increased risk and prevalence of type 2 diabetes mellitus [
1]. This increasing phenomenon has been found to be associated with dietary and lifestyle changes, which favor the consumption of phosphorus (P)-deficient food items [
2]. In parallel, plasma phosphate levels were found to be synergistically related to glucose tolerance and insulin sensitivity [
3‐
5]. The association between P and glucose is believed to be mediated by insulin [
6], as a result of its capacity to stimulate both P uptake by the muscles [
7] and phosphorylation of many compounds; thereby, creating a competition [
8] among these compounds for P.
Additionally, overweight and/or obesity, a known risk factor for type 2 diabetes mellitus, is reported to be inversely related to plasma P [
2,
3]. The intakes of high protein food choices, which are rich sources of P, have been reported to improve weight loss [
9] by a calcium independent mechanism [
10,
11]. Furthermore, the intake of dairy products has been also found to be associated with a lower risk for type 2 diabetes mellitus in three meta-analyses of large prospective epidemiological studies [
12‐
15]. Moreover, since post-prandial glycaemia is a risk factor for type 2 diabetes mellitus [
16,
17], it is reasonable to postulate that P is involved in its development. Several dietary, lifestyle and pharmaceutical approaches have been used to reduce postprandial glycaemia, including the enhancement of intracellular glucose trapping. Recently glucokinase activators have been developed and tested, but their capacity to improve glycaemia was associated with several side effects [
18‐
20] that may be attributed to hepatic intracellular P depletion, especially since a limited quantity of free P is present intracellularly.
In brief, increased incidence of glucose intolerance and type 2 diabetes mellitus is associated with dietary changes that favor a reduction in the consumption of P, an important mediator in glucose metabolism [
5,
21‐
23]. Despite the presence of evidence showing the inefficiency of fasting P to reflect its diurnal [
24] or postprandial status [
25], a positive association between fasting P levels and insulin sensitivity has been reported [
26].
Micronutrients enrichment of carbohydrate meals was reported to reduce post-prandial glycaemia [
27]. Although P is tightly integrated in the metabolism of glucose, the impact of P ingestion on glucose tolerance and insulin resistance of healthy subjects is not well documented. However, phosphate infusion of healthy subjects was reported to improve insulin sensitivity under euglycaemic conditions [
6] and phosphate supplementation of hypophosphatemic glucose intolerant patients improved glucose tolerance [
28]. The aim of this study was to investigate the effect of exogenous P on postprandial glucose and insulin status in healthy subjects. Based on that, this study holds significance in the advances of human nutrition in improving glucose tolerance and therefore preventing the development of type 2 diabetes mellitus in humans.
Discussion
In agreement with other studies, ingestion of P alone (experiment 1 and 2) increased postprandial serum P [
31], while ingestion of grape juice [
31], glucose alone [
32‐
34], or other type of carbohydrate [
35] reduced postprandial serum P levels. Glucose may indirectly affect P status through the stimulation of peripheral P uptake by insulin [
7,
36,
37], which in turn stimulates the phosphorylation of several compounds including carbohydrate [
35], fat and protein. About 60 % of infused P has been reported to be translocated from the extracellular to the intracellular compartment [
6], mainly in skeletal muscles [
21,
36]. This translocation is believed to be mediated via insulin action as glucose infusion in pancreatectomic dogs failed to induce a reduction in serum P except when insulin became available [
21,
37,
38]. In line with this, the parallel increase in postprandial glucose and insulin (peaks at 30–60 min) was followed by a decrease in serum P (dip at 90 min). Therefore, under conditions of P intake alone, intracellular P is likely to have been affected since insulin was not altered.
In the current study, both co- and pre-ingestion of P were able to halt the drop in serum P levels following glucose ingestion. In the G + P treatment, increased P uptake and availability might have contributed to the drop in glucose and insulin levels at time 60 min, due to an increase in intracellular glucose trapping (phosphorylation), especially since insulin release depends on glucose circulation. This process might have played a role in the observed improvement in the measures of insulin sensitivity following P ingestion with glucose (Experiment 1). Phosphorylation or P trapping seems to have been substantially stimulated or became dependent on extracellular P 30 min after glucose ingestion; as indicated by the drop in postprandial serum P levels. This may partially be behind the failure of P pre-ingestion to impact postprandial plasma glucose, since the majority of phosphorus is known to be absorbed within 60 min, as supported by the finding from experiment 1. When glucose was ingested alone, the low availability of P may have hindered insulin phosphorylation capacity through creating competition for P. Such competition may have deleterious effects since it can affect glucose clearance and trapping, glycolysis and gluconeogenesis [
39], and phospholipids and hepatic fat accumulation [
40]. Therefore, postprandial glycemia and insulinemia seem to be improved by exogenous P availability and this may partially explain the reported association between low serum P with insulin resistance and elevated blood glucose levels [
5,
23].
Ingestion of P before or with glucose was able to prevent the drop in postprandial serum P levels. The sustained high postprandial serum P in the G + P treatment in comparison to that of the glucose ingestion alone implies that intracellular P uptake may be controlled by a limited capacity for phosphorylation and/ or glucose uptake. In healthy subjects, peripheral glucose uptake, especially in skeletal muscles, is known to be triggered by insulin dependent Glut 4 stimulation [
41]. While, intracellular glucose phosphorylation is controlled by the activities of glucokinase (liver) and hexokinase (muscle), the latter has a low Vmax (maximum velocity) capacity and is highly inhibited by glucose-P production [
42]. The reduction in serum glucose of the G + P treatment argues against a defect in Glut 4 (glucose uptake), therefore the sustenance in plasma P may have been attributed to the low Vmax capacity of muscle hexokinase. Accordingly, ingestion of higher P doses would not be expected to further improve glucose, insulin, or insulin sensitivity.
The fact that P is absorbed along the entire intestinal tract [
22,
43] could be responsible for the observed high plasma P levels (above baseline value) in the G + P treatment beyond the time (120 min) of availability of glucose and insulin. Moreover, the difference in the magnitude of changes in postprandial glucose and insulin levels between the preload and the co-ingestion experiments implies that factors beyond the availability of circulating P, glucose and insulin may have been involved in improving of insulin sensitivity. The weak significant association seen between P preload on insulin sensitivity could be explained by the small sample size of the present study. On the other hand, glucose-phosphorus interaction in the proximal part of the small intestine may been involved in insulin sensitivity through incretin hormones. These hormones, especially glucagon like peptide-1 (GLP-1) and gastric inhibitory polypeptide (GIP) are known to be secreted in response to meal ingestion, especially high protein meals (rich in P) [
9,
42] and were reported to affect insulin status and to play an important role in regulating postprandial blood glucose [
9,
44,
45].
The observed improvement in the measures of insulin sensitivity following meal-phosphorus co-ingestion may have been partially involved in the reported synergic relationship between the intake of whole grains and glucose tolerance [
46]. Especially since this relationship was not explained by the function of dietary fiber [
47] and whole grains are rich in phosphors. Additionally, our observation may partially explain the observed parallel rise in metabolic syndrome with global urbanization and westernization of dietary habits, which favor low P intake [
3]. In comparison to other studies in the literature, which have used P injections to study its effect on glycaemia and insulinemia [
6], the current study used a different method that mimicked daily dietary habits, through the ingestion of 500 mg of P with a glucose load (approximately 1.7 mg of P per Kcal). Therefore, these findings highlight the role of P in improving the states of hyperglycemia and hyperinsulinemia in healthy individuals without the influence of serum calcium and FGF-23 which did not vary when P is used [
25].
On the other hand, elevated fasting serum P levels were reported to be associated with mortality among patients with chronic kidney [
48] and coronary [
49] diseases. The association was partially explained by the capacity of high P conditions to induce vascular calcification and endothelial injury using in vitro studies [
50‐
53]. Moreover, in a human (
in vivo) study endothelial function impairment was apparent under high (1200 mg P/meal) and not normal (400 mg P/meal) P ingestion [
54]. In fact, the negligible impact of dietary P intake on serum P levels implies that factors associated with increased serum P, rather than P intake, were probably behind the association between cardiovascular disease and serum P [
24]. Recently, a weak association between dietary P intake and all-cause mortality was reported [
55] and this was questioned since participants adopted different dietary patterns and P intake was not the only variable. Thus, the nature of the relation between P intake and cardiovascular disease and mortality is far from clear and requires further scrutiny [
56].
In the present study, sample size was based on the previously reported data of AUC for glucose [
57]. However, the observed large variations between subjects seem to have diluted the impact of the interventions. Further studies, using a larger sample size, would help in exploring the mechanisms by which the observed effects are mediated especially by examining the role of incretin hormones.
The limitation of this study lies in its small number of subjects enrolled into the experimental design. Moreover, although the dietary habits of these subjects were not assessed prior to the initiation of the study; however, as stated previously, fasting serum P status is not a good indicator of P intake. Postprandial status of the measured parameters is not likely to be affected by prior meal intake since all subjects were overnight fasted. Therefore, assessing dietary habits of subjects has no added implications on the study results.
Acknowledgements
The research was supported by Lebanese National Council for Scientific Research (LNCSR) and University Research Board (URB). The following sponsors did not participate in the design or conduct of this study, collection, management, analysis, or interpretation of the data, writing of the manuscript, or in the preparation, review, approval or decision to submit this manuscript for publication.
We thank Dr Mazen Al-Ghoul for his mathematical support for the calculation of insulin sensitivity.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
MK carried out the studies, sample analysis and writing of manuscript. CAR carried out the studies, sample analysis and writing of manuscript. GH contributed to data analysis and writing of the manuscript. SH performed the statistical analysis. OO made substantial contributions to conception, design, acquisition of data, analysis, and interpretation of data. All authors read and approved the final manuscript.