Free fatty acids and insulin secretion in humans

Acute increases in plasma levels of long-chain fatty acids raise plasma insulin levels by stimulating insulin secretion or by decreasing insulin clearance. In normal subjects, longterm elevations of fatty acids also stimulate insulin secretion. In fact, they increase insulin precisely to the degree needed to compensate for the fatty acid-induced insulin resistance. In contrast, in individuals who are genetically predisposed to develop type 2 diabetes (prediabetic subjects), the free fatty acid (FFA) stimulation of insulin secretion is not sufficient to fully compensate for the FFAinduced insulin resistance. Therefore, obesity, if associated with elevated fatty acid levels, may lead to hyperglycemia in prediabetic but not in normal individuals.     

Free fatty acids decrease circulating ghrelin concentrations in humans

OBJECTIVE: Concentrations of the orexigenic peptide ghrelin is affected by a number of hormones, which also affect circulating levels of free fatty acids (FFAs). The present study was therefore designed to determine the direct effect of FFAs on circulating ghrelin. DESIGN: Eight lean, healthy men were examined for 8 h on four occasions using variable infusion rates (0, 3, 6 and 12 microl/kg per min) of intralipid to create different plasma FFA concentrations. Constant levels of insulin and GH were obtained by administration of acipimox (250 mg) and somatostatin (300 microg/h). At the end of each study day a hyperinsulinaemic-euglycaemic clamp was performed. RESULTS: Four distinct levels of FFAs were obtained at the end of the lipid infusion period (FFA(LIPID): 0.03 +/- 0.00 vs: 0.49 +/- 0.04, 0.92 +/- 0.08 and 2.09 +/- 0.38 mmol/l; ANOVA P < 0.0001) and during hyperinsulinaemia (FFA(LIPID+INSULIN): 0.02 +/- 0.00 vs: 0.34 +/- 0.03, 0.68 +/- 0.09 and 1.78 +/- 0.32 mmol/l; ANOVA P < 0.0001). Whereas, somatostatin infusion alone reduced ghrelin concentration by approximately 67%, concomitant administration of increasing amounts of intralipid reduced circulating ghrelin by a further 14, 19 and 19% respectively (change in ghrelin: 0.52 +/- 0.05 vs: 0.62 +/- 0.06, 0.72 +/- 0.09 and 0.71 +/- 0.05 microg/l; ANOVA P = 0.04). No further reduction in ghrelin concentration was observed during hyperinsulinaemia. CONCLUSION: FFA exposure between 0 and 1 mmol/l significantly suppresses ghrelin levels independent of ambient GH and insulin levels.     

Free fatty acids increase hepatic glycogen content in obese males

Obesity is associated with increased hepatic glycogen content. In vivo and in vitro data suggest that plasma free fatty acids (FFA) may cause this increase. In this study we investigated the effect of physiological plasma FFA levels on hepatic glycogen metabolism by studying intrahepatic glucose pathways in lean and obese subjects. Six lean and 6 obese males were studied twice during a 16- to 22-hour fast, once with and once without acipimox, an inhibitor of lipolysis. Intrahepatic glucose fluxes were measured by infusion of [2-(13C1)]glycerol, [1-(2H1)]galactose, and [U-(13C6)]glucose. Acetaminophen was administered as a glucuronate probe. In both lean and obese control studies, plasma FFA levels increased progressively, whereas acipimox completely suppressed plasma FFA levels for the whole study period. In lean males glycogenolysis did not change in the acipimox study, but decreased in the control study (P < .01). In lean males, neither glycogen synthesis, glycogen synthesis retained as glycogen, nor glycogen balance differed between control and acipimox studies. In obese males glycogenolysis did not change in the acipimox study, but decreased in the control study (P < .01). Glycogen synthesis did not change in either study. Glycogen synthesis retained as glycogen did not change in acipimox study, but increased in the control study (P = .03). Glycogen balance did not change in the acipimox study, but increased in the control study (P < .01). This study demonstrates that in obese males physiological levels of FFA contribute to the retention of hepatic glycogen during short-term fasting by inhibiting breakdown of glycogen and increasing glycogen synthesis retained as glycogen, whereas in lean males this effect was absent due to unaltered glycogen synthesis retained as glycogen.     

Hypoglycemia-induced hemolysis in glucose-6-phosphate dehydrogenase deficiency

We present a young man with Mediterranean type glucose-6-phosphate dehydrogenase (G6PD) deficiency and insulin-dependent diabetes mellitus whose brittle course was characterized by recurrent bouts of hypoglycemia and diabetic ketoacidosis (DKA). While neither of the episodes of DKA was complicated by hemolysis, hemolytic anemia consistently followed the recurrent attacks of hypoglycemia. Stringent control of the patient's blood glucose levels in the upper limit and slightly above the normal range successfully prevented recurrence of hypoglycemia and recrudescence of hemolytic anemia. Hypoglycemia is proposed as capable of inducing hemolysis in G6PD deficiency.     

Cortisol metabolism in glucose-6-phosphate dehydrogenase deficiency

Ten subjects with glucose-6-phosphate dehydrogenase deficiency (G6PD Canton) and ten G6PD normal subjects matched for sex and age distribution were studied with respect to their plasma cortisol levels and 24-hr urinary total 17-oxogenic steroids excretion before and after maximal stimulation with B^1–24 corticotrophin (Synacthen depot). In five patients and five controls, the study included the metabolic clearance rate of cortisol which increased twofold after B^1–24 corticotrophin administration. No significant differences were found in these two groups of individuals. This indicates that in G6PD-deficient subjects, under basal conditions and with maximal stimulation, both cortisol production and metabolism proceed at a normal rate.     

Molecular heterogeneity of glucose-6-phosphate dehydrogenase A-

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is probably the most common disease-producing genetic polymorphism of humans. Virtually all G6PD-deficient Africans show the G6PD A- phenotype, an electrophoretically rapid, deficient enzyme. The recently acquired ability to identify the point mutations producing the different variants has given us new insights into the population genetics of G6PD variants. Twenty-nine males with the G6PD A- phenotype were studied. They were of African, Mexican, Spanish, and US white ethnic origin. All had the A---G transition at nucleotide 376 characteristic of G6PD A. In each case, one of three additional mutations was present, at nucleotides 202, 680, or 968. That in this population second mutations producing G6PD deficiency occurred only on the genetic background of G6PD A suggests that G6PD A was at one time the most common type of G6PD in Africa. However, the nucleotide sequence of the chimpanzee (Pan troglodytes) G6PD indicates that the primordial human type of G6PD was G6PD B.     

The genetics of glucose-6-phosphate dehydrogenase deficiency

In the past 35 years, G-6-PD and its genetic variants have been a valuable resource for the geneticist and cell biologist. Over 380 putative variants have been described and have served as a paradigm, showing how many ways mutations can affect an enzyme. With the cloning and sequencing of G-6-PD, a new chapter has been opened in our understanding of G-6-PD. Variants that were thought to be different have proven to be identical, and those that were thought to be the same are now seen to be heterogeneous. Importantly, the ability to deduce quickly the amino acid substitution in a G-6-PD variant makes feasible, for the first time, the unraveling of relationships between the structure and function of this enzyme.     

Detection of glucose-6-phosphate dehydrogenase deficient heterozygotes

Heterozygotes for the Mediterranean type of severe G-6-PD deficiencywere investigated by a variety of tests.

The methemoglobin reduction test was most successful in detecting heterozygotes (about 80 per cent). Enumeration of methemoglobin containingcells on blood films (Kleihauer-Betke technic) did not improve these results.Quantitation of enzyme level was less successful (65 per cent), and determination of decolorization time by the BCB technic was least sensitivein heterozygote detection.

Methemoglobin reduction technics reflect a more indirect effect of themutant gene than enzyme assay. The superiority of these technics in heterozygote detection is probably caused by the genetically determined presenceof both normal and enzyme-deficient cells in G-6-PD deficient heterozygotes.Since methemoglobin reduction is carried out by individual cells, the population of enzyme deficient cells does not reduce methemoglobin, and thereforeeven a minority of deficient cells leads to abnormal test results. In contrast,enzyme assay is less successful for heterozygote detection, since measurementof enzyme level is carried out on hemolyzed red cells, where cellular mosaicismno longer exists.

An additional source of variation of enzyme levels in heterozygotes iscaused by the existence of genetically determined control of normal enzymelevel. Possession of a high capacity allele for G-6-PD activity may placea heterozygote in the normal range of enzyme activity.

The various tests were also applied to subjects with the mild Greek typeof G-6-PD deficiency. Males with this mutation had enzyme levels varyingbetween 12-45 per cent of the mean of normal males. Methemoglobin reduction test results were considerably less abnormal in hemizygotes with themild type of Mediterranean deficiency than in heterozygotes with the severedeficiency. Fewer heterozygotes with the mild deficiency were detected.

Submitted on February 4, 1966 Accepted on May 31, 1966     

Free fatty acids exert an inhibitory effect on adrenocorticotropin and cortisol secretion in humans

Free fatty acid (FFA) administration stimulates the hypothalamic-pituitary-adrenal (HPA) axis in rats, suggesting that the HPA axis and lipolysis may be linked by a positive-feedback loop. To clarify the influence of FFA on the HPA axis in humans, we studied the effect of lipid load on both basal and stimulated ACTH and cortisol secretion in normal subjects. In six young female volunteers [(mean +/- SEM) age, 24.4 +/- 2.1 yr; body mass index, 23.1 +/- 1.2 kg/m(2)), ACTH, cortisol, FFA, glucose, and insulin levels were measured every 30 min for 330 min during the following procedures: 1) i.v. saline infusion (from 0 to 330 min); 2) i.v. FFA infusion (Intralipid 10%, from 0 to 210 min) followed by saline infusion (from 210 to 330 min); 3) human CRH (hCRH) administration (2 microg/kg i.v. at 90 min) during saline infusion (from 0 to 330 min); and 4) hCRH administration during FFA infusion (Intralipid 10%, from 0 to 210 min, followed by saline infusion from 210 to 330 min). During saline infusion, ACTH and cortisol levels progressively declined. Lipid-heparin emulsion (LHE) infusion strikingly increased circulating FFA levels and, simultaneously, amplified the ACTH and cortisol decrease (P < 0.05). After LHE withdrawal, FFA decrease was associated with an increase (P < 0.05) in ACTH and cortisol levels (restored to baseline values within 60 min). The ACTH and cortisol responses to hCRH, however, were unaffected by LHE that, concomitantly, induced an increase (P < 0.05) in glucose but not in insulin levels. This study shows that an LHE-induced increase in FFA levels has an inhibitory effect on spontaneous ACTH and cortisol secretion in humans. Lipid load, however, does not affect the ACTH and cortisol responses to hCRH; this evidence would indicate that the negative influence of FFA on the HPA axis in humans takes place at the suprapituitary level.     

Free fatty acids inhibit adrenocorticotropin and cortisol secretion stimulated by physical exercise in normal men

BACKGROUND: The basal circulating levels of ACTH and cortisol, but not the ACTH/cortisol response to hCRH, are significantly reduced by free fatty acid (FFA) infusion. OBJECTIVE: To verify whether FFA infusion modifies the ACTH/cortisol response to physical exercise, a well-known activator of the HPA axis at suprapituitary level. DESIGN: Exercise tests on a bicycle ergometer during infusion of a lipid-heparin emulsion (LHE) (experimental test) or normal saline (NaCl 0.9%) (control test). SETTING: Department of Cardiology at the University-Hospital. SUBJECTS: Seven healthy male subjects aged 25-33 years. INTERVENTIONS: On two mornings, at weekly intervals, LHE or saline were infused for 60 min; infusion started 10 min before exercise test on a bicycle ergometer, which lasted about 15 min. MAIN OUTCOME MEASURES: Circulating ACTH/cortisol levels and physiological variables during physical exercise. RESULTS: FFA levels (0.4 +/- 0.1 mEq/l) remained constant during control test, whereas they progressively rose (peak at 60 min, 2.7 +/- 1.0 mEq/l) during LHE infusion. Neither basal nor exercise-induced changes in physiological variables were modified by LHE infusion. Both ACTH and cortisol increased during exercise, with peak levels at 20 min and 30 min (control test: 103% and 42%, P < 0.001; experimental test: 28.5% and 18.6%, P < 0.05 higher than baseline, respectively). Both ACTH and cortisol responses were significantly lower in the experimental than in the control test (at 20 min P < 0.002 and at 30 min P < 0.05 for ACTH; at 20 min P < 0.05 and at 30 min, 40 min and 50 min P < 0.001 for cortisol). CONCLUSIONS: These data represent the first demonstration of an inhibitory action of increased circulating FFA levels on the HPA axis under stimulatory conditions (i.e. physical exercise, a challenge acting at suprapituitary level). In contrast, previous studies did not show FFA effects on the CRH-induced ACTH/cortisol response. Therefore, our data suggest negative effects of FFAs on the HPA axis at hypothalamic or higher centres in the central nervous system.