Oral Semaglutide and Cardiovascular Outcomes in Patients with Type 2 Diabetes.

BACKGROUND: Establishing cardiovascular safety of new therapies for type 2 diabetes is important. Safety data are available for the subcutaneous form of the glucagon-like peptide-1 receptor agonist semaglutide but are needed for oral semaglutide. METHODS: We assessed cardiovascular outcomes of once-daily oral semaglutide in an event-driven, randomized, double-blind, placebo-controlled trial involving patients at high cardiovascular risk (age of ≥50 years with established cardiovascular or chronic kidney disease, or age of ≥60 years with cardiovascular risk factors only). The primary outcome in a time-to-event analysis was the first occurrence of a major adverse cardiovascular event (death from cardiovascular causes, nonfatal myocardial infarction, or nonfatal stroke). The trial was designed to rule out 80% excess cardiovascular risk as compared with placebo (noninferiority margin of 1.8 for the upper boundary of the 95% confidence interval for the hazard ratio for the primary outcome). RESULTS: A total of 3183 patients were randomly assigned to receive oral semaglutide or placebo. The mean age of the patients was 66 years; 2695 patients (84.7%) were 50 years of age or older and had cardiovascular or chronic kidney disease. The median time in the trial was 15.9 months. Major adverse cardiovascular events occurred in 61 of 1591 patients (3.8%) in the oral semaglutide group and 76 of 1592 (4.8%) in the placebo group (hazard ratio, 0.79; 95% confidence interval [CI], 0.57 to 1.11; P<0.001 for noninferiority). Results for components of the primary outcome were as follows: death from cardiovascular causes, 15 of 1591 patients (0.9%) in the oral semaglutide group and 30 of 1592 (1.9%) in the placebo group (hazard ratio, 0.49; 95% CI, 0.27 to 0.92); nonfatal myocardial infarction, 37 of 1591 patients (2.3%) and 31 of 1592 (1.9%), respectively (hazard ratio, 1.18; 95% CI, 0.73 to 1.90); and nonfatal stroke, 12 of 1591 patients (0.8%) and 16 of 1592 (1.0%), respectively (hazard ratio, 0.74; 95% CI, 0.35 to 1.57). Death from any cause occurred in 23 of 1591 patients (1.4%) in the oral semaglutide group and 45 of 1592 (2.8%) in the placebo group (hazard ratio, 0.51; 95% CI, 0.31 to 0.84). Gastrointestinal adverse events leading to discontinuation of oral semaglutide or placebo were more common with oral semaglutide. CONCLUSIONS: In this trial involving patients with type 2 diabetes, the cardiovascular risk profile of oral semaglutide was not inferior to that of placebo. (Funded by Novo Nordisk; PIONEER 6 ClinicalTrials.gov number, NCT02692716.).

Incorporating and interpreting regulatory guidance on estimands in diabetes clinical trials: The PIONEER 1 randomized clinical trial as an example.

Regulatory guidelines describe the use of estimands in designing and conducting clinical trials. Estimands ensure alignment of the objectives with the design, conduct and analysis of a trial. An estimand is defined by four inter-related attributes: the population of interest, the variable (endpoint) of interest, the way intercurrent events are handled and the population level summary. A trial may employ multiple estimands to evaluate treatment effects from different perspectives in order to address different scientific questions. As estimands may be an unfamiliar concept for many clinicians treating diabetes, this paper reviews the estimand concept and uses the PIONEER 1 phase 3a clinical trial, which investigated the efficacy and safety of oral semaglutide vs placebo, as an example of the way in which estimands can be implemented and interpreted. In the PIONEER 1 trial, two estimands were employed for each efficacy endpoint and were labelled as: (a) the treatment policy estimand, used to assess the treatment effect regardless of use of rescue medication or discontinuation of trial product, and provides a broad perspective of the treatment effect in the population of patients with type 2 diabetes in clinical practice; and (b) the trial product estimand, used to assess the treatment effect if all patients had continued to use trial product for the planned duration of the trial without rescue medication, thereby providing information on the anticipated treatment effect of the medication. Both approaches are complementary to understanding the effect of the studied treatments.

Cardiovascular and renal outcomes of GLP-1 receptor agonists vs. DPP-4 inhibitors and basal insulin in type 2 diabetes mellitus: A systematic review and meta-analysis.

OBJECTIVE: To compare the cardiovascular and renal outcomes of GLP-1 RA versus DPP4i and basal insulin in the management of T2DM. METHODS: Data from 22 studies involving over 200,000 participants were pooled using the inverse variance method and random-effects meta-analysis. The review was reported in accordance with PRISMA. RESULTS: Compared with DPP4i, treatment with GLP-1 RA was associated with a greater benefit on composite cardiovascular outcomes (HR:0.77, 95% CI:0.69-0.87), myocardial infarction (HR:0.82, 95% CI:0.69-0.97), stroke (HR:0.83, 95% CI: 0.74-0.93), cardiovascular mortality (HR:0.76, 95% CI:0.68-0.85) and all-cause mortality (HR:0.65, 95% CI:0.48-0.90). There was no difference in effect on heart failure (HR:0.97, 95% CI:0.82-1.15). Compared with basal insulin, GLP-1 RA was associated with better effects on composite cardiovascular outcomes (HR:0.62, 95% CI:0.48-0.79), heart failure (HR:0.57, 95% CI:0.35-0.92), myocardial infarction (HR:0.70, 95% CI:0.58-0.85), stroke (HR:0.50, 95% CI:0.40-0.63) and all-cause mortality (HR:0.31, 95% CI:0.20-0.48). Evidence from a small number of studies suggests that GLP-1 RA had better effects on composite and individual renal outcomes, such as eGFR, compared with either DPP4i and basal insulin. CONCLUSION: Available evidence suggests that treating T2DM with GLP-1 RA can yield better benefits on composite and specific cardiorenal outcomes than with DPP4i and basal insulin. PROSPERO REGISTRATION NUMBER: CRD42022335504.

Effect of Various Dosing Conditions on the Pharmacokinetics of Oral Semaglutide, a Human Glucagon-Like Peptide-1 Analogue in a Tablet Formulation.

INTRODUCTION: Oral semaglutide is a novel tablet formulation of the human glucagon-like peptide-1 analogue semaglutide. In two trials, the effects of prior food ingestion (food effect), post-dose fasting period and water volume with dosing (dosing conditions) on oral semaglutide pharmacokinetics were investigated. METHODS: Subjects received once-daily oral semaglutide for 10 days. In the food-effect trial, 78 healthy subjects were randomised 1:1:1 to fed (meal 30 min pre-dose; 240 mL water with dosing), fasting (overnight until 4 h post-dose; 240 mL) or reference (fasting overnight until 30 min post-dose; 120 mL) arms. In the dosing conditions trial, 161 healthy men were randomised into eight dosing groups (overnight fasted with 50/120 mL water and 15/30/60/120 min post-dose fasting). Semaglutide plasma concentrations were measured frequently until 504 h after the 10th dose. RESULTS: In the food-effect trial, limited or no measurable semaglutide exposure was observed in the fed arm, while all subjects in the fasting arm had measurable semaglutide exposure. Area under the semaglutide concentration-time curve (AUC0-24h,semaglutide,day10) and maximum semaglutide concentration (Cmax,semaglutide,day10) were numerically greater by approximately 40% for the fasting versus reference arm (p = 0.082 and p = 0.080, respectively). In the dosing conditions trial, AUC0-24h,semaglutide,day10 and Cmax,semaglutide,day10 were not different between water volumes (p = 0.541 and p = 0.676), but increased with longer post-dose fasting (p < 0.001). CONCLUSION: Administration of oral semaglutide in the fasting state with up to 120 mL water and at least 30 min post-dose fasting results in clinically relevant semaglutide exposure. These dosing conditions have been used in the oral semaglutide phase 3 trials and are part of the approved label. TRIAL REGISTRATION: ClinicalTrials.gov identifiers NCT02172313, NCT01572753.

Relationship Between Oral Semaglutide Tablet Erosion and Pharmacokinetics: A Pharmacoscintigraphic Study.

Semaglutide, a glucagon-like peptide-1 (GLP-1) analogue, has been coformulated in a tablet with the absorption enhancer, sodium N-(8-[2-hydroxybenzoyl] amino) caprylate (SNAC). We investigated tablet erosion and the pharmacokinetics of oral semaglutide administered with 2 different water volumes and evaluated the relationships between these parameters. In a randomized, single-center (Quotient Sciences, UK), open-label, 2-period crossover trial, 26 healthy men received single doses of 10 mg oral semaglutide with 50 or 240 mL water while fasting. Tablet erosion and gastrointestinal transit were assessed by gamma scintigraphy. Semaglutide and SNAC plasma concentrations were measured until 24 and 6 hours, respectively, after administration. Complete tablet erosion (CTE) occurred in the stomach irrespective of water volume administered with the tablet (primary end point). Mean time to CTE was 85 versus 57 minutes with 50 versus 240 mL water (ratio 50/240 mL, 1.51; 95% confidence interval, 0.96-2.37; P = .072). Area under the semaglutide concentration-time curve from 0 to 24 hours (AUC0-24h,semaglutide ) and maximum semaglutide concentration (Cmax,semaglutide ) were ∼70% higher with 50 versus 240 mL water (P = .056 and P = .048, respectively). Median time to maximum semaglutide concentration (tmax,semaglutide ) was 1.5 hours independent of water volume with dosing. Higher AUC0-24h,semaglutide and Cmax,semaglutide and longer tmax,semaglutide were significantly correlated with longer time to CTE and later gastric emptying of tablet and water (all P < .05). The safety profile was as expected for the GLP-1 receptor agonist drug class. In conclusion, the oral semaglutide tablet erodes in the stomach irrespective of water volume with dosing. Slower tablet erosion in the stomach results in higher semaglutide plasma exposure.

Safety and Pharmacokinetics of Single and Multiple Ascending Doses of the Novel Oral Human GLP-1 Analogue, Oral Semaglutide, in Healthy Subjects and Subjects with Type 2 Diabetes.

BACKGROUND: Oral semaglutide is a novel tablet containing the human glucagon-like peptide-1 (GLP­1) analogue semaglutide, co-formulated with the absorption enhancer sodium N-(8-[2-hydroxybenzoyl] amino) caprylate (SNAC). The safety and pharmacokinetics of oral semaglutide were investigated in two randomised, double-blind, placebo-controlled trials. METHODS: In a single-dose, first-in-human trial, 135 healthy males received oral semaglutide (2-20 mg semaglutide co-formulated with 150-600 mg SNAC) or placebo with SNAC. In a 10-week, once-daily, multiple-dose trial, 84 healthy males received 20 or 40 mg oral semaglutide (with 300 mg SNAC), placebo, or placebo with SNAC, and 23 males with type 2 diabetes (T2D) received 40 mg oral semaglutide (with 300 mg SNAC), placebo, or placebo with SNAC. RESULTS: Oral semaglutide was safe and well-tolerated in both trials. The majority of adverse events (AEs) were mild, with the most common AEs being gastrointestinal disorders. In the single-dose trial, semaglutide exposure was highest when co-formulated with 300 mg SNAC. In the multiple-dose trial, semaglutide exposure was approximately twofold higher with 40 versus 20 mg oral semaglutide in healthy males, in accordance with dose proportionality, and was similar between healthy males and males with T2D. The half-life of semaglutide was approximately 1 week in all groups. CONCLUSION: The safety profile of oral semaglutide was as expected for the GLP-1 receptor agonist drug class. Oral semaglutide co-formulated with 300 mg SNAC was chosen for further clinical development. The pharmacokinetic results supported that oral semaglutide is suitable for once-daily dosing. CLINICALTRIALS. GOV IDENTIFIERS: NCT01037582, NCT01686945.

Impact of Liraglutide on Amylase, Lipase, and Acute Pancreatitis in Participants With Overweight/Obesity and Normoglycemia, Prediabetes, or Type 2 Diabetes: Secondary Analyses of Pooled Data From the SCALE Clinical Development Program.

OBJECTIVE: To describe amylase/lipase activity levels and events of acute pancreatitis (AP) in the SCALE (Satiety and Clinical Adiposity-Liraglutide Evidence in individuals with and without diabetes) weight-management trials. RESEARCH DESIGN AND METHODS: Secondary analyses were performed on pooled data from four trials (N = 5,358 with BMI ≥30, or 27 to <30 kg/m2 with ≥1 comorbidity). Of these, 1,723 had normoglycemia, 2,789 had prediabetes, and 846 had type 2 diabetes. Participants were randomized to liraglutide 3.0 mg (n = 3,302), liraglutide 1.8 mg (n = 211, only type 2 diabetes), or placebo (n = 1,845). Relationships between baseline characteristics and amylase/lipase activity at baseline and during treatment were investigated. RESULTS: Over 56 weeks, liraglutide 3.0 mg versus placebo was associated with increases in mean levels of 7% (amylase) and 31% (lipase), respectively. Similar changes in amylase/lipase levels were observed with liraglutide 1.8 mg. More participants receiving liraglutide 3.0 mg versus placebo experienced amylase (9.4% vs. 5.9%) and lipase (43.5% vs. 15.1%) elevations greater than or equal to the upper limit of normal (ULN); few had elevations ≥3 × ULN for amylase (<0.1% with liraglutide 3.0 mg or placebo) or lipase (2.9% vs. 1.5%, respectively). After liraglutide discontinuation, enzymes returned to baseline levels. Thirteen participants developed AP: 12 on (n = 9, 0.3%) or after (n = 3, 0.1%) liraglutide 3.0 mg treatment and one (0.1%) with placebo. A total of 6/13 participants with AP (5/12 liraglutide; 1 placebo) had gallstone disease evident at AP onset. Amylase/lipase elevations either 1 × ULN or ≥3 × ULN before AP onset had very low positive predictive value for AP (<1%). CONCLUSIONS: Liraglutide resulted in dose-independent, reversible increases in amylase/lipase activity, unrelated to baseline characteristics, not predicting AP onset. Gallstones possibly contributed to 50% of AP cases. Data provide no basis for amylase/lipase level monitoring in liraglutide treatment except in suspected AP.

Reductions in systolic blood pressure with liraglutide in patients with type 2 diabetes: insights from a patient-level pooled analysis of six randomized clinical trials.

AIMS: To quantify the effect of liraglutide on systolic blood pressure (SBP) and pulse in patients with type 2 diabetes (T2D), and assess the influence of covariates on observed SBP reductions. METHODS: A patient-level pooled analysis of six phase 3, randomized trials was conducted. RESULTS: The analysis included 2792 randomized patients. In the intention-to-treat population (n=2783), mean [±SE] SBP reductions from baseline with liraglutide 1.2 mg (2.7 [0.8] mmHg) and 1.8 mg (2.9 [0.7] mmHg) once daily were significantly greater than with placebo (0.5 [0.9] mmHg; P=0.0029 and P=0.0004, respectively) after 26 weeks, and were evident after 2 weeks. Liraglutide was also associated with significantly greater SBP reductions than glimepiride and, at a dose of 1.8 mg, insulin glargine and rosiglitazone. SBP reductions with liraglutide weakly correlated with weight loss (Pearson's correlation coefficient: 0.08-0.12; P≤0.0148). No dependence of these reductions on concomitant antihypertensive medications was detected (P=0.1304). Liraglutide 1.2 and 1.8 mg were associated with mean increases in pulse of 3 beats per minute (bpm), versus a 1 bpm increase with placebo (P<0.0001 for each dose versus placebo). CONCLUSIONS: Liraglutide reduces SBP in patients with T2D, including those receiving concomitant antihypertensive medication.

Liraglutide achieves A1C targets more often than sitagliptin or exenatide when added to metformin in patients with type 2 diabetes and a baseline A1C <8.0%.

OBJECTIVE: Compare the safety and efficacy of liraglutide to that of sitagliptin or exenatide as add-on to metformin in patients with type 2 diabetes (T2D) and glycated hemoglobin (A1C) <8.0%. METHODS: Post hoc analysis of 26-week data from liraglutide 1.8 mg once daily (OD) versus exenatide 10 µg twice daily (LEAD-6) and liraglutide 1.8 mg OD versus sitagliptin 100 mg OD (LIRA-DPP-4); only patients treated as add-on to metformin with baseline A1C <8.0% were included. Efficacy analysis was performed on the intention-to-treat population with missing values imputed by last observation carried forward. RESULTS: More patients achieved A1C targets (<7.0% and ≤6.5%) with liraglutide versus exenatide or sitagliptin; the difference was greatest for A1C ≤6.5% (LEAD-6: 65% versus 35%; odds ratio [OR]=3.37, 95% confidence interval [CI]: 1.31-8.63; P = .01 or LIRA-DPP-4: 53% versus 19%; OR = 4.78, 95% CI 2.10 to 10.87; P = .0002). Significantly more patients achieved a composite endpoint of A1C <7.0% with no weight gain or hypoglycemia with liraglutide compared with exenatide (78% versus 42%; OR = 4.99, 95% CI: 1.77 to 14.04; P = .0023) or sitagliptin (61% versus 21%; OR = 5.95, 95% CI: 2.66 to 13.29; P<.0001). All treatments were well tolerated, there was no major hypoglycemia and few patients (8 to 10%) experienced minor hypoglycemia. CONCLUSION: When added to metformin in patients with an A1C <8.0%, more patients using liraglutide 1.8 mg reached A1C targets than with exenatide or sitagliptin. Sitagliptin had particularly low efficacy in this analysis. These data support the use of liraglutide 1.8 mg as a safe and effective alternative to sitagliptin or exenatide following metformin failure in patients with an A1C <8.0%.

Defining the ideal injection techniques when using 5-mm needles in children and adults.

OBJECTIVE: We aimed to establish the ideal injection techniques using 5-mm needles to reliably inject insulin into the subcutaneous fat in both children and adults and to quantify the associated pain and leakage of the test medium. RESEARCH DESIGN AND METHODS: A total of 259 subjects (122 children/adolescents and 137 adults) were injected with sterile air corresponding to 20 IU insulin (200 microl) with 32-G 5-mm needles at 90 degrees or 45 degrees , in the abdomen and thigh, and with or without a pinched skin fold. Injection depth was assessed via ultrasonography. Subjects rated pain on a visual analog scale. Test medium injections into the abdomen and thigh (0.2-0.6 ml) were also administered to assess injection leakage. RESULTS: Among children, 5.5% of injections were intramuscular (IM) and 0.5% were intradermal, while in adults, the incidence was 1.3 and 0.6%, respectively. The frequency of IM injections was greater in boys and negligible among adult women. Subcutaneous fat thickness was the primary predictor of the likelihood of IM injections (P < 0.001). A third of all patients reported experiencing no pain during insulin injection, with children/adolescents experiencing considerably more discomfort than adults. Some leakage of medium was observed, but was unrelated to injection volume and was generally minimal. CONCLUSIONS: 5-mm needles are reliably inserted into subcutaneous fat in both adults and children. These needles were associated with reduced pain and minimal leakage. We recommend an angled injection with a pinched skin fold for children, while in adults, the technique should be left to patient preference.

Dose accuracy and durability of the NovoPen 4 insulin delivery device before and after simulation of 5 years of use and under various stress conditions.

OBJECTIVE: The dose accuracy of the NovoPen 4 (Novo Nordisk A/S, Bagsvaerd, Denmark) insulin delivery device was evaluated before and after simulation of 5 years of use and under conditions of mechanical and temperature stress. METHODS: To simulate 5 years of use of the NovoPen 4, > or =5475 injections were performed automatically, corresponding to 3 daily injections for 5 years. Accuracy in the delivery of 1-, 30-, and 60-U insulin doses before and after simulation of 5 years of use was assessed over the in-use temperature range (5 degrees C-40 degrees C [41 degrees F-104 degrees F]). The durability and dose accuracy of the NovoPen 4 were also assessed after exposure to extremes of temperature (-40 degrees C and 70 degrees C), high relative humidity (95% RH), and mechanical challenges (freefall from 1.5 m at ambient temperature and from 1 m at 5 degrees C, 40 degrees C, and 95% RH). RESULTS: The dose accuracy of the NovoPen 4 before and after 5 years of simulated use was within International Organization for Standardization 11608-specified limits for all doses studied. Dose accuracy remained within specifications for all doses after exposure to variations in temperature and humidity, and after freefall. CONCLUSION: The NovoPen 4 exhibited dose accuracy before and after simulation of 5 years of use, across a wide range of temperature and humidity conditions, and after mechanical challenges.

A comparison of injection force and dosage scale size between NovoPen 3 and NovoPen 4.

BACKGROUND: Easy-to-use insulin devices are an important tool in the treatment of diabetes. In this study, injection force requirements and dosage scale displays were evaluated for NovoPen 3 and NovoPen 4 (both from Novo Nordisk A/S, Copenhagen, Denmark). METHODS: To simulate 5 years of expected lifetime usage, 5,475 injections were performed automatically. Before and after lifetime testing, the force required to expel 60 U of insulin from NovoPen 3 and NovoPen 4 with 30-gauge or 31-gauge needles was measured. To compare dosage scale displays, digital images of multiple settings were made, and the total inked areas of digits were converted to mm(2). RESULTS: At baseline, the mean +/- SD injection force of NovoPen 4 was 9.14 +/- 0.87 N and 16.55 +/- 1.17 N, which was significantly lower (P < 0.001) than for NovoPen 3, at 18.36 +/- 1.06 N and 29.81 +/- 1.26 N, with the 30-gauge and 31-gauge needle, respectively. After simulated lifetime testing, mean +/- SD injection force was 10.93 +/- 0.77 N and 17.77 +/- 1.14 N for NovoPen 4, which was significantly lower than the injection force of 18.54 +/- 0.94 N and 31.69 +/- 1.98 N for NovoPen 3 (P < 0.001). The mean dosage scale digit size was 1.63 mm(2) for NovoPen 3 and 7.82 mm(2) for NovoPen 4, with a mean difference of 6.19 mm(2). The display for NovoPen 4 was 4.74 times larger (P < 0.001). CONCLUSIONS: The mean injection force required to operate NovoPen 4 was reduced up to 50% compared with NovoPen 3 (P < 0.001), and the mean dosage display for NovoPen 4 was over four times larger than for NovoPen 3 (P < 0.001). Based on these findings, patients with diabetes who have manual or visual impairment should find it easier to dose insulin with NovoPen 4.

Denervation and high-fat diet reduce insulin signaling in T-tubules in skeletal muscle of living mice.

OBJECTIVE: Insulin stimulates muscle glucose transport by translocation of GLUT4 to sarcolemma and T-tubules. Despite muscle glucose uptake playing a major role in insulin resistance and type 2 diabetes, the temporal and spatial changes in insulin signaling and GLUT4 translocation during these conditions are not well described. RESEARCH DESIGN AND METHODS: We used time-lapse confocal imaging of green fluorescent protein (GFP) ADP-ribosylation factor nucleotide-binding site opener (ARNO) (evaluation of phosphatidylinositide 3-kinase activation) and GLUT4-GFP-transfected quadriceps muscle in living, anesthetized mice either muscle denervated or high-fat fed. T-tubules were visualized with sulforhodamine B dye. In incubated muscle, glucose transport was measured by 2-deoxy-D-[(3)H]-glucose uptake, and functional detubulation was carried out by osmotic shock. Muscle fibers were immunostained for insulin receptors. RESULTS: Denervation and high-fat diet reduced insulin-mediated glucose transport. In denervated muscle, insulin-stimulated phosphatidylinositol 3,4,5 P(3) (PIP3) production was abolished in T-tubules, while PIP3 production at sarcolemma was increased 2.6-fold. Correspondingly, GLUT4-GFP translocation to T-tubules was abolished, while translocation to sarcolemma was increased 2.3-fold. In high fat-fed mice, a approximately 65% reduction in both insulin-induced T-tubular PIP3 production and GLUT4-GFP translocation was seen. Sarcolemma was less affected, with reductions of approximately 40% in PIP3 production and approximately 15% in GLUT4-GFP translocation. Access to T-tubules was not compromised, and insulin receptor distribution in sarcolemma and T-tubules was unaffected by denervation or high-fat feeding. Detubulation of normal muscle reduced basal and abolished insulin-induced glucose transport. CONCLUSIONS: Our findings demonstrate, for the first time, that impaired insulin signaling and GLUT4 translocation is compartmentalized in muscle and primarily localized to T-tubules and not sarcolemma during insulin resistance.

Decrease in intramuscular lipid droplets and translocation of HSL in response to muscle contraction and epinephrine.

A better understanding of skeletal muscle lipid metabolism is needed to identify the molecular mechanisms relating intramuscular triglyceride (IMTG) to muscle metabolism and insulin sensitivity. An increasing number of proteins have been reported to be associated with intracellular triglyceride (TG), among them the PAT family members: perilipin, ADRP (for adipocyte differentiation-related protein), and TIP47 (for tail-interacting protein of 47 kDa). Hormone-sensitive lipase (HSL) is thought to be the major enzyme responsible for IMTG hydrolysis in skeletal muscle. In adipocytes, regulation of HSL by intracellular redistribution has been demonstrated. The existence of such regulatory mechanisms in skeletal muscle has long been hypothesized but has never been demonstrated. The aim of this study was to characterize the PAT family proteins associated with IMTG and to investigate the effect of epinephrine stimulation or muscle contraction on skeletal muscle TG content and HSL intracellular distribution. Rat soleus muscles were either incubated with epinephrine or electrically stimulated for 15 min. Single muscle fibers were used for morphological analysis by confocal and transmission electron microscopy. We show a decrease in IMTG in response to both lipolytic stimuli. Furthermore, we identify two PAT family proteins, ADRP and TIP47, associated with IMTG. Finally, we demonstrate HSL translocation to IMTG and ADRP after stimulation with epinephrine or contraction.

Muscle triacylglycerol and hormone-sensitive lipase activity in untrained and trained human muscles.

During exercise, triacylglycerol (TG) is recruited in skeletal muscles. We hypothesized that both muscle hormone-sensitive lipase (HSL) activity and TG recruitment would be higher in trained than in untrained subjects in response to prolonged exercise. Healthy male subjects (26 +/- 1 years, body moss index 23.3 +/- 0.5 kg m(-2)), either untrained (N = 8, VO(2max) 3.8 +/- 0.2 l min(-1)) or trained (N = 8, VO(2max) 5.1 +/- 0.1 l min(-1)), were studied. Before and after 3-h exercise (58 +/- 1% VO(2max)), a biopsy was taken. Muscle citrate synthase (32 +/- 2 vs. 47 +/- 6 mumol g(-1) min(-1) d.w.) and beta-hydroxy-acyl-CoA-dehydrogenase (38 +/- 3 vs. 52 +/- 5 mumol g(-1) min(-1) d.w.) activities were lower in untrained than in trained subjects (p < 0.05). Throughout the exercise, fat oxidation was higher in trained than in untrained subjects (p < 0.05). Muscle HSL activity was similar at rest (0.72 +/- 0.08 and 0.74 +/- 0.03 mU mg(-1) protein) and after exercise (0.71 +/- 0.1 and 0.68 +/- 0.03 mU mg(-1) protein) in untrained and trained subjects. At rest, the chemically determined muscle TG content (37 +/- 8 and 26 +/- 5 mmol g(-1) d.w.) was similar (p > 0.05), and after exercise it was unchanged in untrained and lower (p < 0.05) in trained subjects (41 +/- 9 and 10 +/- 2 mmol g((1) d.w.). Determined histochemically, TG was decreased (p < 0.05) after exercise in type I and II fibres. Depletion of TG was not different between fibre types in untrained, but tended to be higher (p = 0.07) in type I compared with type II fibres in trained muscles. In conclusion, HSL activity is similar in untrained and trained skeletal muscles both before and after prolonged exercise. However, the tendency to higher muscle TG recruitment during exercise in the trained subjects suggests a difference in the regulation of HSL or other lipases during exercise in trained compared with untrained subjects.

Sex differences in hormone-sensitive lipase expression, activity, and phosphorylation in skeletal muscle at rest and during exercise.

Women have been shown to use more intramuscular triacylglycerol (IMTG) during exercise than men. To investigate whether this could be due to sex-specific regulation of hormone-sensitive lipase (HSL) and to use sex comparison as a model to gain further insight into HSL regulation, nine women and eight men performed bicycle exercise (90 min, 60% Vo(2peak)), and skeletal muscle HSL expression, phosphorylation, and activity were determined. Supporting previous findings, basal IMTG content (P < 0.001) and net IMTG decrease during exercise (P < 0.01) were higher in women than in men and correlated significantly (r = 0.72, P = 0.001). Muscle HSL mRNA (80%, P = 0.11) and protein content (50%, P < 0.05) were higher in women than in men. HSL total activity increased during exercise (47%, P < 0.05) but did not differ between sexes. Accordingly, HSL specific activity (HSL activity per HSL protein content) increased during exercise (62%, P < 0.05) and was generally higher in men than in women (82%, P < 0.05). A similar pattern was observed for HSL Ser(659) phosphorylation, suggesting a role in regulation of HSL activity. Likewise, plasma epinephrine increased during exercise (P < 0.05) and was higher in men than in women during the end of the exercise bout (P < 0.05). We conclude that, although HSL expression and Ser(659) phosphorylation in skeletal muscle during exercise is sex specific, total muscle HSL activity measured in vitro was similar between sexes. The higher basal IMTG content in women compared with men is therefore the best candidate to explain the higher IMTG net hydrolysis during exercise in women.

Hormone-sensitive lipase as mediator of lipolysis in contracting skeletal muscle.

The authors propose that the enzyme hormone-sensitive lipase (HSL), which is the rate-limiting enzyme for hydrolysis of triacylglycerol in adipocytes, also regulates the intramyocellular triacylglycerol mobilization and is controlled by mechanisms similar to those regulating glycogen phosphorylase. From an exercise perspective, it is fascinating that the primary enzymatic setting allows simultaneous mobilization of all major extramuscular and intramuscular energy stores.