Sweet-taste receptors, glucose absorption and insulin release: Are LCS nutritionally active? Samuel V. Molinary, Ph.D. Consultant, Scientific & Regulatory Affairs ILSI/NA April 6, 2011 Washington, DC
Why is this question being asked now after a century of human exposure to low calorie sweeteners? Recent basic research on the molecular biology of taste receptors and related GI receptors has led to speculation that LCS may interact with the GI glucose sensing system to alter glucose uptake and, consequently, to undermine glucose homeostasis, increase appetite and lead to an increase in obesity.
Outline 1. Review the GI sweet receptor and its function in glucose absorption. 2. Briefly review the interaction of LCS with the GI sweet receptor. 3. Examine the speculation re LCS and adverse effects. 4. Review available clinical data that address the speculation. 5. Review recent clinical studies designed to test the hypothesis that LCS are nutritionally active.
GI Taste Receptors and Glucose Sensing and Transport
Useful Definitions Enterocyte GI cells that actually absorb nutrients from the gut lumen. Enteroendocrine cell GI cells (~1% of total) that sense the presence of nutrients in the gut lumen, release incretin hormones that prepare the enterocytes and body for a nutrient load. GLP-1 glucagon-like peptide, an incretin secreted from L cells. GIP glucose-dependent insulinotropic polypeptide, an incretin secreted from K cells.
Glucose absorption across the intestinal wall Glucose is a highly polar molecule that cannot simply diffuse across the gut wall Cells have transporters in their cell membranes for the uptake of glucose In the intestine these transporters occur on both the luminal (apical) and basolateral (blood) sides of the enterocytes
Enterocyte G Gut lumen SGLT1 G Cytosol G G ATP ADP GLUT2 GLUT2 G G Blood stream
Glucose absorption across the intestinal wall The glucose transporters in enterocyte cell membranes adapt to the amount of glucose available in the gut lumen How is the presence of glucose detected and signalled to the enterocytes? Enteroendocrine cells interspersed with the enterocytes have sweet-taste receptors activation of which produces 2 incretin hormones (GLP-1 and GIP)
Enteroendocrine cell Gut lumen Sweet-taste receptor Ca 2+ channel Adenylate cyclase α-g ATP c-amp Kinase activities Synthesis and release of GLP-1 and GIP
Glucose absorption across the intestinal wall GLP-1 and GIP are released when the tastereceptor is stimulated and produce 2 effects in the enterocytes: 1. The expression of SGLT-1 on the luminal membrane is increased most effective at low concentrations of glucose 2. GLUT2 is increased and now is also present in the luminal membrane to deal with high glucose concentrations
G G Gut lumen GLUT2 SGLT1 G Cytosol G ATP ADP G G GLUT2 GLUT2 G G Blood stream
Glucose absorption across the intestinal wall Intense sweeteners can stimulate the intestinal sweet-taste receptors in in vitro studies and cause the same changes in the intestinal epithelium as glucose. Release of GLP-1 and GIP Increase intestinal glucose transporters So why is this an issue? If LCS can have the same effect as glucose on the GI sensing system by increasing incretin secretion, there might be long term physiological consequences re glucose homeostasis and energy balance.
Intestinal sweet-taste receptors and insulin release The issue has received attention because GLP-1 and GIP have additional non-local functions, which prepare the body for a glucose load Stimulate gustatory nerves Delay gastric emptying Stimulate the release of insulin What might happen if intestinal sweet tastereceptors are stimulated by an intense sweetener?
The hypothesis of Egan and Margolskee (2008) i. stimulation of the sweet-taste receptors in the intestine by low-calorie sweeteners causes increased expression of glucose uptake transporters (SGLT1 and GLUT2) on the enterocytes to enhance glucose uptake, ii. the released GLP-1 and GIP enter the general circulation and increase insulin secretion which lowers blood sugar, iii. this affects glucose homeostasis and the risk of diabetes, iv. the lowered blood sugar increases appetite, leading to increased weight gain.
Egan and Margolskee (2008) went on to state Obesity and hyperlipidemias are linked to sugary soft drinks but low calorie diet drinks may further increase the incidence of obesity and/or metabolic syndrome Small studies have associated diet soft drinks with high blood pressure in women and increased weight gain in boys In the absence of calories the balance between taste receptor activation, nutrient assimilation and appetite may be disequilibrated leading to an increase in appetite and overeating when calories are available
Experimental basis The experiments on which this hypothesis is based are: 1. In vitro studies exposing mouse & human cell culture lines to various sweeteners. 2. Knockout mouse studies of short duration. 3. Short-term feeding studies in mice. 4. In situ studies in rats. 5. Really beautiful molecular biology studies aimed at defining the enteroendocrine cell to cell signaling mechanism.
BUT In vivo studies in humans have shown that ingestion of an intense sweetener does NOT i. Increase appetite ii. Increase food intake iii. Cause an increase in body weight iv. Cause the release of insulin v. Affect glucose homeostasis or control of diabetes Renwick 1994 review
The hypothesis as stated by Egan and Margolskee suggests that LCS are nutritionally active with respect to GLP-1 and GIP secretion by the enteroendocrine cells. Several other investigators have also raised this possibility
Reasons to question relevance of this hypothesis All modern LCS have large safety databases, which include animal studies of two-years duration at massive daily intakes. If glucose homeostasis is undermined as predicted by Egan/Margolskee, rats ingesting >3000 mg/kg/day for two years should have shown multiple adverse effects. Blood chemistries, histology and in-life data show no evidence of aberrations in glucose control; there is no evidence of obesity.
Long-term studies from the sucralose and rebaudioside A safety data bases
Sucralose Clinical Studies (1) Normal Volunteers: N=48, duration = 12 weeks + 4 week screening phase and a 4 week follow-up phase. The study was a double-blind, placebo-controlled, randomized, parallel group designed to assess the effect of sucralose on glucose homeostasis in healthy male volunteers. Study parameters: HbA1c, insulin, C-peptide, glucagon and glucose. OGTT s given during study. Capsules taken 3X per day with meals (1000 mg total). Achieved daily dose = 13.22 mg/kg/day vs. ADI of 15 mg/kg/day. (High daily intake is ~ 2.4 mg/kg/day) Result: No effect on glucose homeostasis, all values were within normal limits and all OGTT s were normal.
Sucralose Clinical Studies (1) The Egan/Margolskee hypothesis suggests that sucralose ingestion should have increased glucose transport by increasing incretin secretion by the enteroendocrine cells, thus modifying blood glucose and insulin levels. The C-peptide level, and possibly the blood glucose levels, should have increased even in these normal subjects over the three-month study duration. BUT no effects were observe in any parameter.
Sucralose Clinical Studies (2) Type II diabetic subjects: N =128, duration = three months + 8 week screening phase and a 4 week followup phase. The study was a double-blind, placebo-controlled, randomized, parallel group, multi-center trial designed to assess the effect of sucralose on glucose homeostasis in type II diabetics. Of the 128 subjects, 64 were being treated with oral hypoglycemic agents and 64 with insulin. Study parameters: HbA1c, insulin, C-peptide, glucagon and glucose. OGTT s given during study. Capsules taken 2X daily with meals. Achieved daily dose = 7.5 mg/kg/day.
Sucralose Clinical Studies (2) There was no difference between the cellulose placebo controls and the sucralose group. There was no change in medication during the study. Compliance was monitored by measuring sucralose in the urine of the subjects. There were no differences in any safety measure. There was no evidence suggesting that glucose control was undermined in this study of diabetic subjects.
Stevia Clinical Studies (1) Normotensive adults: N = 100, duration = 4 weeks + 4 week screening period. Study was a randomized, double-blind trial designed to evaluate the hemodymanic effects of 1000 mg/day (capsules 500mg 2X daily with meals) of rebaudioside A vs. placebo. Compared resting BP, mean BP, HR and 24 hour ambulatory BP responses. Result: no clinically important changes in BP in healthy, normotensive adults.
Stevia Clinical Studies (2) Type II diabetic subjects: N = 122, duration = 16 weeks + 2 week screening phase. The study was a double-blind, placebo controlled multicenter trial designed to assess the effect of rebaudioside A on glucose homeostasis and blood pressure. Study parameters: HbA1c, insulin, C-peptide, and glucose. Measurements of BP, BW and fasting lipids were also made. Capsules taken 2X daily with meals, total of 1000 mg steviol glycoside. Results: there were no differences between the rebaudioside A group and the placebo group in any parameter.
Recently published clinical studies
Recent Clinical Studies 1. Ma et al (2009): measured circulating levels of GLP-1, GIP, insulin, blood glucose and gastric emptying after intragastric infusions of sucrose, sucralose or saline control in 500 ml of iso-osmotic solutions in 7 healthy subjects. Result:: only sucrose elevated GLP-1, GIP, insulin and glucose and delayed gastric emptying. Sucralose (0.4mM* & 4mM) did not differ from the saline controls. *[= 168 ppm, approx. the amount in a diet soft drink]
Recent Clinical Studies 2. Ma et al (2010): measured GLP-1, glucose and OMG in 10 healthy subjects after intraduodenal infusion of glucose, sucralose (4mM) and OMG to evaluate whether sucralose exposure of the upper GI tract will increase the rate of glucose absorption and glycaemic response. Result:: sucralose did not modify the rate of glucose absorption or the glycaemic or incretin response to ID infusion when given acutely to healthy subjects.
Recent Clinical Studies 3. Ford et al (2011) investigated whether oral ingestion of sucralose, at a dose level consistent with a normal diet (2 mm), increases circulating GLP-1 or PYY concentrations in 8 healthy volunteers. This was a randomized, single-blind, crossover study conducted on 4 separate days. Results: sucralose ingestion did not increase plasma GLP- 1 or PYY concentrations. MSF of sucralose did not elicit a cephalic response for insulin or GLP-1. The control, maltodextrin, significantly increased insulin and blood glucose vs water. Appetite and energy intake was similar for all groups.
Recent Clinical Studies 4. Steinert et al (2011), investigated the human gut response to LCS (APM, AC-K, sucralose) and sugars (glu, fruc, 2-DG) using solutions that were iso-sweet. Solutions were administered IG in a placebo-controlled, double-blind, six-way, cross-over trial using 12 healthy subjects. Results: LCS did not affect GI peptide secretion (GLP-1, PYY & ghrelin) and had minimal effect on appetite. These investigators conclude that stimulation of gut sweet taste receptors per se is not sufficient to produce regulatory peptide responses.
Conclusions (1) The data used to support the speculation that LCS interact with the GI taste receptors to increase incretin secretion, and, consequently, increase glucose absorption are derived from elegant molecular biology experiments using short-term in vitro tissue culture systems and rodent model systems to define the receptor site of the enteroendocrine cells.
Conclusions (2) There are very extensive databases demonstrating the safety and efficacy of each LCS from which relevant data can be drawn. Had LCS induced the derangements in glucose homeostasis predicted by Egan/Margolskee hypothesis, they would have manifest themselves in the preclinical and clinical safety studies.
Conclusions (3) An increasing number of clinical studies have been recently published that specifically test the Egan/Margolskee hypothesis and none has, as of this date, supported the concept that LCS are nutritionally active in a way that is clinically significant. There is no consistent evidence that LCS cause insulin release or lower blood sugar when ingested.