Skeletal muscle metabolism is impaired during exercise in glycogen storage disease type III Nicolai Preisler, MD, PhD Pascal Laforêt, MD, PhD Karen Lindhardt Madsen, MD Kira Philipsen Prahm, MD Gitte Hedermann, MD Christoffer Rasmus Vissing, BSc Henrik Galbo, MD, PhD John Vissing, MD, PhD
Correspondence to Dr. Preisler: [email protected]
ABSTRACT
Objective: Glycogen storage disease type IIIa (GSDIIIa) is classically regarded as a glycogenosis with fixed weakness, but we hypothesized that exercise intolerance in GSDIIIa is related to muscle energy failure and that oral fructose ingestion could improve exercise tolerance in this metabolic myopathy.
Methods: We challenged metabolism with cycle-ergometer exercise and measured substrate turnover and oxidation rates using stable isotope methodology and indirect calorimetry in 3 patients and 6 age-matched controls on 1 day, and examined the effect of fructose ingestion on exercise tolerance in the patients on another day. Results: Total fatty acid oxidation rates during exercise were higher in patients than controls, 32.1 (SE 1.2) vs 20.7 (SE 0.5; range 15.8–29.3) mmol/kg/min (p 5 0.048), and oxidation of carbohydrates was lower in patients, 1.0 (SE 5.4) vs 38.4 (SE 8.0; range 23.0–77.1) mmol/kg/ min (p 5 0.024). Fructose ingestion improved exercise tolerance in the patients.
Conclusion: Similar to patients with McArdle disease, in whom muscle glycogenolysis is also impaired, GSDIIIa is associated with a reduced skeletal muscle oxidation of carbohydrates and a compensatory increase in fatty acid oxidation, and fructose ingestion improves exercise tolerance. Our results indicate that GSDIIIa should not only be viewed as a glycogenosis with fixed skeletal muscle weakness, but should also be considered among the glycogenoses presenting with exercise-related dynamic symptoms caused by muscular energy deficiency.
Classification of evidence: This study provides Class IV evidence that ingestion of fructose improves exercise tolerance in patients with GSDIIIa. Neurology® 2015;84:1767–1771 GLOSSARY FFA 5 free fatty acids; GSDIIIa 5 glycogen storage disease type IIIa; Ra 5 rate of appearance; RER 5 respiratory exchange ratio; ROX 5 rate of oxidation; RPE 5 rate of perceived exertion.
Editorial, page 1726
Glycogen storage disease type III (OMIM #232400) is caused by mutations in the AGL gene, which lead to deficiency of the glycogen debranching enzyme (GDE).1 GDE works in concert with phosphorylase to facilitate the degradation of glycogen to glucose-1-phosphate.2 In glycogen storage disease type IIIa (GSDIIIa), the GDE activity is deficient in both skeletal muscle and liver.3 In this study, we exclusively examined patients with GSDIIIa. In the early stages of the disease, the liver is usually primarily affected in patients with GSDIIIa, even though GDE levels are reduced in skeletal muscle as well.1,4 With aging, weakness and atrophy of limb muscles develops, usually in the 3rd to 4th decade.1,4 Recently, however, glucose infusion has been shown to improve exercise tolerance, suggesting exerciserelated energy deficiency in GSDIIIa.5 Inspired by these findings, we investigated substrate turnover during exercise in patients with GSDIIIa to test our hypothesis that exercise intolerance is related to muscle energy failure caused by the metabolic block in glycogen breakdown. We also hypothesized and tested whether oral fructose ingestion can improve exercise tolerance in this metabolic myopathy.
Supplemental data at Neurology.org From the Neuromuscular Research Unit, Department of Neurology (N.P., K.L.M., K.P.P., G.H., C.R.V., J.V.), and the Department of Inflammation Research (H.G.), Rigshospitalet, University of Copenhagen, Denmark; and the Centre de Référence de Pathologie Neuromusculaire Paris-Est (P.L.), Institut de Myologie, GH Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, France. Go to Neurology.org for full disclosures. Funding information and disclosures deemed relevant by the authors, if any, are provided at the end of the article. © 2015 American Academy of Neurology
1767
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
5.4–7.2 5.0–6.4 220–370 2,722–4,643 — 54–88 1.66–1.86 Range
19–27
— 69 6 11.3 1.78 6 0.1 24.3 6 2.3 Controls 6 SD
p Value
Peak exercise testing (day 1). The subjects performed an incremental exercise test to exhaustion on a cycle-ergometer to determine peak oxidative capacity (VO2peak) and workload (Wpeak).
Abbreviations: RER 5 respiratory exchange ratio measured at time of exhaustion; VO2 peak 5 peak oxidative capacity; Wpeak 5 peak workload. a Female. A detailed description of the patients has been presented previously (patients 1, 2, and 55). Controls, n 5 6. b Significantly different from controls at time of exhaustion.
276 6 53 3,414 6 711
1.09–1.22
0.024 0.024
112 6 21 2,052 6 444 — 78 6 3.2 1.75 6 0.0 Mean 6 SD
23.7 6 8.1
2,117
b
2,459
c.66411G.A, c.66411G.A 79 1.77
c.66411G.A, c.66411G.A 80 1.78
20 Patient 3
Patient 2
18
1,579 c.2229delT, c.3486_3488delGGA 74 1.71 33 Patient 1a
Neurology 84
Subjects. Three patients with genetically and biochemically verified GSDIIIa and 6 healthy sex- and age-matched controls (2:1) were included (table 1). The patients have been described in detail in a previous article (patients 1, 2, and 5).5 For a detailed description of the exercise interventions, see e-Methods on the Neurology® Web site at Neurology.org.
0.7–1.2
6.4–13.3
6.1 6 0.7 5.5 6 0.6
0.024
0.82 6 0.0
0.80 105
b
0.84 135
1.15 6 0.1
0.85 6 0.2
0.024
10.2 6 2.7
0.024
4.5 6 0.1b 4.9 6 0.5
4.6
4.6
0.82 95
b
1.0 6 0.2
1.6 6 0.2
b
4.4
4.8 1.7
1.8 1.0
0.9
4.4 5.4 1.4 1.2
Glucose peak, mmol/L Glucose rest, mmol/L Lactate peak, mmol/L Lactate rest, mmol/L RER Wpeak, watts mL/min peak,
VO2 AGL gene, mutation Weight, kg Height, m Age, y Participants
Demographic and peak exercise data from patients with glycogen storage disease type IIIa and healthy controls Table 1 1768
METHODS Our aim was to examine substrate turnover during exercise and to examine the effect of substrate supplementation therapy on exercise tolerance in patients with GSDIIIa. The study provides Class IV evidence that fructose ingestion improves exercise tolerance in patients with GSDIIIa.
Substrate turnover during exercise (day 2). We used stable isotope methodology ([U-13C]-palmitate [primed by a NaH13CO3 bolus], [6,6-D2 ]-glucose, and [1,1,2,3,3-D5 ]-glycerol) in combination with indirect calorimetry to examine substrate turnover during submaximal ergometer cycling (60% of VO2peak). The subjects were tested after an overnight fast, and substrate turnover, heart rate, and rate of perceived exertion (RPE) were determined at rest and during exercise. Samples were collected as shown in figure 1. Blood samples were analyzed for concentration and enrichment of plasma palmitate, glucose, and glycerol, and for concentrations of lactate, free fatty acids (FFA), and insulin. Rates of appearance (Ra), disappearance, and oxidation (ROX) were calculated as described in e-Methods. Oral fructose supplementation during exercise (day 3). On day 3, the patients exercised as on day 2, however, this time drinking apple juice (500 mL, containing 9.5 g of carbohydrate per 100 mL) 10 minutes before exercise. Every 20 minutes during exercise, the patients drank an additional 250 mL of juice. Stable isotopes were not used. The effect of fructose was evaluated comparing results obtained on day 2 (stable isotopes 5 placebo) and day 3, respectively, so that patients served as their own controls.
Standard protocol approvals, registrations, and patient consents. Ethics and statistics. The Regional Ethical Committee of the Capital Region of Denmark approved the study (#H-22010-008, with amendment #39559), and it was conducted in accordance with the ethical standards of the Declaration of Helsinki. The subjects gave written informed consent prior to inclusion. Median resting and exercise (averages from the whole period) values were compared between patients and controls with a Mann-Whitney rank-sum test, and a p value , 0.05 was considered significant. Results are reported as mean 6 SE unless otherwise stated. The effect of fructose ingestion was described quantitatively.
The VO2peak, the Wpeak, the respiratory exchange ratio (RER), and the lactate and glucose concentrations at time of exhaustion were significantly lower in the patients compared to controls (table 1). RESULTS Peak exercise testing.
Substrate turnover during exercise. Substrate turnover during exercise is detailed in figures 1 and 2 and table e-1. Total FFA and palmitate ROX at rest were similar between groups. However, during exercise, FFA ROX was higher in the patients. In line with this, the
April 28, 2015
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
Figure 1
Substrate metabolism during exercise in patients with glycogen storage disease type IIIa
(A) Palmitate rate of oxidation (ROX), (B) rate of appearance (Ra) of glucose, (C) total free fatty acid (FFA) ROX, (D) plasma FFA concentration, (E) total carbohydrate (Carb) ROX. and (F) Ra of glycerol in patients with glycogen storage disease type IIIa (GSD, n 5 3) and healthy controls (CON, n 5 6) during submaximal exercise at 60% of VO2peak in the patients. The controls exercised at the same absolute workload as the patients with whom they were matched. Data in A, B, and F were only collected during the stable isotope trial (placebo) on day 2. Patients were also tested with an oral fructose supplement (FRU). *Significantly different from controls (mean of the whole exercise period). **Significant difference between the fructose ingestion trial and the stable isotope (placebo) study (patients only, mean of the whole exercise period, quantitative assessment). Error bars are standard error.
Ra of palmitate and Ra of glycerol and palmitate and FFA concentrations were significantly higher during exercise in the patients. In contrast, the oxidation of carbohydrates during exercise was significantly lower in the patients.
In accordance with this, the RER was also lower in the patients, and, paradoxically, the RER declined during exercise. In patient 1, RER dropped to 0.67, and in patient 3, to 0.68, indicative of ketone body production. Neurology 84
April 28, 2015
1769
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
Figure 2
Workload and Borg score, and lactate and glucose concentrations, placebo vs fructose
(A, B) The workload and duration of exercise and the rate of perceived exertion (RPE) in the placebo (nonfructose experiment, stable isotope) trial. (C, D) The workload and duration of exercise and the RPE after fructose ingestion. Oral fructose ingestion improved exercise tolerance, and 2 patients exercised longer, and all patients rated exercise as being easier with fructose ingestion, and heart rate tended to be lower; 124 6 1.1 vs 119 6 0.7 beats 3 minute21 on fructose. (E) Lactate concentration. (F) Glucose concentration. CON 5 controls (n 5 6); FRU 5 data from the fructose ingestion trial; GSD 5 patients with glycogen storage disease type IIIa (n 5 3). *Significantly different from controls (mean of the whole exercise period). **Significant difference between the fructose ingestion trial and the stable isotope (placebo) study (patients only, mean of the whole exercise period, quantitative assessment). Error bars are standard error.
Glucose levels were consistently lower at all times in the patients and tended to drop during exercise. In line with this, insulin levels were also low in patients. Oral fructose supplementation during exercise. Exercise capacity improved with oral fructose ingestion, which 1770
Neurology 84
was evident from the increased duration that the patients could sustain exercise, a lower heart rate during exercise (in 2 patients), and a lower RPE during exercise in all patients (figures 1 and 2 and table e-2). Fructose ingestion increased blood glucose and lactate levels, RER, and oxidation of carbohydrates.
April 28, 2015
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
DISCUSSION The main finding of the present study is that patients with GSDIIIa have a severely limited ability to break down glycogen during exercise, which results in energy deficiency and a compensatory increase in the oxidation of fat. These findings are similar to what has been observed in patients with McArdle disease, who also have blocked muscle glycogen breakdown.6 Our findings indicate that the exercise intolerance, which was reported by our patients and other patients, and the exercise-induced symptoms of pain and excessive fatigue that we recently demonstrated in patients with GSDIIIa, most likely can be explained by an energy shortage in muscle during exercise.5,7–9 Similar to findings in patients with McArdle disease, we demonstrate that exercise capacity in GSDIIIa can be improved by raising blood glucose levels.10 This supports energy deficiency as cause of exercise intolerance and suggests a dynamic component of the exerciseinduced symptoms. The findings are not surprising considering the nature of the enzyme defect, but are at variance with the general notion that GSDIIIa is a glycogenosis associated only with static symptoms due to fixed muscle weakness.1,4 More severe exercise-induced muscular symptoms like contractures and rhabdomyolysis are not features of GSDIIIa, unlike McArdle disease, because myophosphorylase can release small amounts of glucose-1-phosphate from glycogen early during exercise, which probably protects skeletal muscle in patients with GSDIIIa. This is supported by the presence of abnormal glycogen, phosphorylase limit-dextrin, in muscle of patients with GSDIIIa.8 However, exercise capacity and CNS function may be severely compromised by hypoglycemia, which evolves rapidly at higher intensities of exercise, due to liver involvement in GSDIIIa.5 The lower glucose and insulin levels could explain the high resting FFA concentrations that we observed in our patients in the present study. The data on muscle substrate turnover consistently indicated a significant block in skeletal muscle glycogenolytic capacity during exercise; however, exercise intolerance differed between the patients, and the effect of fructose ingestion on work capacity and RPE in patient 1 was minimal. Residual enzyme activity or other factors could account for this difference. Therefore it is important to test substrate supplementation therapy in larger populations of patients, also when they are in their habitual metabolic state and not overnight fasted as in previous trials, to establish if our results apply to all patients with GSDIIIa. Still, patients should be cautioned to use oral sucrose/fructose supplements carefully and only in situations preceding exercise, to avoid overuse, weight gain, and associated complications.
AUTHOR CONTRIBUTIONS N. Preisler: design of study, analysis, acquisition, and interpretation of data, and drafting the manuscript. P. Laforet: acquisition of data and critical revision of manuscript. K.L. Madsen: design of study, acquisition of data, and critical revision of manuscript. K. Prahm: acquisition of data and critical revision of manuscript. G. Hedermann: acquisition of data and critical revision of manuscript. C.R. Vissing: acquisition of data and critical revision of manuscript. H. Galbo: interpretation of data and critical revision of the manuscript. J. Vissing: design of study, analysis, interpretation of data, and critical revision of the manuscript.
ACKNOWLEDGMENT The authors thank Danuta Goralska-Olsen, Thomas Lauridsen, Nina Pluszek, and Lene Foged for technical assistance.
STUDY FUNDING Supported by the Aase and Ejnar Danielsens Foundation, Merchant L.F. Foghts Foundation, The A.P. Møller Foundation for the Advancement of Medical Science, and the Sara and Ludvig Elssas Foundation.
DISCLOSURE N. Preisler reports having received research support, honoraria, and travel funding from the Genzyme Corporation. P. Laforêt reports having received research support and honoraria from the Genzyme Corporation. Dr Laforêt is a member of the Genzyme Pompe Disease Advisory Board. K. Madsen, K. Prahm, G. Hedermann, C. Vissing, and H. Galbo report no disclosures relevant to the manuscript. J. Vissing reports having received research support and honoraria from the Genzyme Corporation. He is a member of the Genzyme Pompe Disease Advisory Board. Go to Neurology.org for full disclosures.
Received August 27, 2014. Accepted in final form December 10, 2014. REFERENCES 1. Kishnani PS, Austin SL, Arn P, et al. Glycogen storage disease type III diagnosis and management guidelines. Genet Med 2010;12:446–463. 2. Harris RA. Carbohydrate metabolism I: major metabolic pathways and their control. In: Devlin TM, ed. Textbook of Biochemistry with Clinical Correlations, 6th ed. Wilmington, DE: Wiley-Liss; 2006:581–635. 3. Van HF, Hers HG. The subgroups of type 3 glycogenosis. Eur J Biochem 1967;2:265–270. 4. Laforet P, Weinstein DA, Smit PA. The glycogen storage diseases and related disorders. In: Saudubray JM, van den Berghe G, Walter JH, eds. Inborn Metabolic Diseases, 5th ed. Berlin: Springer; 2012:115–139. 5. Preisler N, Pradel A, Husu E, et al. Exercise intolerance in glycogen storage disease type III: weakness or energy deficiency? Mol Genet Metab 2013;109:14–20. 6. Orngreen MC, Jeppesen TD, Andersen ST, et al. Fat metabolism during exercise in patients with McArdle disease. Neurology 2009;72:718–724. 7. Cornelio F, Bresolin N, Singer PA, DiMauro S, Rowland LP. Clinical varieties of neuromuscular disease in debrancher deficiency. Arch Neurol 1984;41:1027–1032. 8. DiMauro S, Hays AP, Tsujino S. Metabolic disorders affecting muscle. In: Engel AG, Franzini-Armstrong C, eds. Myology, 3rd ed. New York: McGraw-Hill; 2004: 1535–1558. 9. DiMauro S, Hartwig GB, Hays A, et al. Debrancher deficiency: neuromuscular disorder in 5 adults. Ann Neurol 1979;5:422–436. 10. Vissing J, Haller RG. The effect of oral sucrose on exercise tolerance in patients with McArdle’s disease. N Engl J Med 2003;349:2503–2509. Neurology 84
April 28, 2015
1771
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
Correspondence to Dr. Preisler: [email protected]
ABSTRACT
Objective: Glycogen storage disease type IIIa (GSDIIIa) is classically regarded as a glycogenosis with fixed weakness, but we hypothesized that exercise intolerance in GSDIIIa is related to muscle energy failure and that oral fructose ingestion could improve exercise tolerance in this metabolic myopathy.
Methods: We challenged metabolism with cycle-ergometer exercise and measured substrate turnover and oxidation rates using stable isotope methodology and indirect calorimetry in 3 patients and 6 age-matched controls on 1 day, and examined the effect of fructose ingestion on exercise tolerance in the patients on another day. Results: Total fatty acid oxidation rates during exercise were higher in patients than controls, 32.1 (SE 1.2) vs 20.7 (SE 0.5; range 15.8–29.3) mmol/kg/min (p 5 0.048), and oxidation of carbohydrates was lower in patients, 1.0 (SE 5.4) vs 38.4 (SE 8.0; range 23.0–77.1) mmol/kg/ min (p 5 0.024). Fructose ingestion improved exercise tolerance in the patients.
Conclusion: Similar to patients with McArdle disease, in whom muscle glycogenolysis is also impaired, GSDIIIa is associated with a reduced skeletal muscle oxidation of carbohydrates and a compensatory increase in fatty acid oxidation, and fructose ingestion improves exercise tolerance. Our results indicate that GSDIIIa should not only be viewed as a glycogenosis with fixed skeletal muscle weakness, but should also be considered among the glycogenoses presenting with exercise-related dynamic symptoms caused by muscular energy deficiency.
Classification of evidence: This study provides Class IV evidence that ingestion of fructose improves exercise tolerance in patients with GSDIIIa. Neurology® 2015;84:1767–1771 GLOSSARY FFA 5 free fatty acids; GSDIIIa 5 glycogen storage disease type IIIa; Ra 5 rate of appearance; RER 5 respiratory exchange ratio; ROX 5 rate of oxidation; RPE 5 rate of perceived exertion.
Editorial, page 1726
Glycogen storage disease type III (OMIM #232400) is caused by mutations in the AGL gene, which lead to deficiency of the glycogen debranching enzyme (GDE).1 GDE works in concert with phosphorylase to facilitate the degradation of glycogen to glucose-1-phosphate.2 In glycogen storage disease type IIIa (GSDIIIa), the GDE activity is deficient in both skeletal muscle and liver.3 In this study, we exclusively examined patients with GSDIIIa. In the early stages of the disease, the liver is usually primarily affected in patients with GSDIIIa, even though GDE levels are reduced in skeletal muscle as well.1,4 With aging, weakness and atrophy of limb muscles develops, usually in the 3rd to 4th decade.1,4 Recently, however, glucose infusion has been shown to improve exercise tolerance, suggesting exerciserelated energy deficiency in GSDIIIa.5 Inspired by these findings, we investigated substrate turnover during exercise in patients with GSDIIIa to test our hypothesis that exercise intolerance is related to muscle energy failure caused by the metabolic block in glycogen breakdown. We also hypothesized and tested whether oral fructose ingestion can improve exercise tolerance in this metabolic myopathy.
Supplemental data at Neurology.org From the Neuromuscular Research Unit, Department of Neurology (N.P., K.L.M., K.P.P., G.H., C.R.V., J.V.), and the Department of Inflammation Research (H.G.), Rigshospitalet, University of Copenhagen, Denmark; and the Centre de Référence de Pathologie Neuromusculaire Paris-Est (P.L.), Institut de Myologie, GH Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, France. Go to Neurology.org for full disclosures. Funding information and disclosures deemed relevant by the authors, if any, are provided at the end of the article. © 2015 American Academy of Neurology
1767
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
5.4–7.2 5.0–6.4 220–370 2,722–4,643 — 54–88 1.66–1.86 Range
19–27
— 69 6 11.3 1.78 6 0.1 24.3 6 2.3 Controls 6 SD
p Value
Peak exercise testing (day 1). The subjects performed an incremental exercise test to exhaustion on a cycle-ergometer to determine peak oxidative capacity (VO2peak) and workload (Wpeak).
Abbreviations: RER 5 respiratory exchange ratio measured at time of exhaustion; VO2 peak 5 peak oxidative capacity; Wpeak 5 peak workload. a Female. A detailed description of the patients has been presented previously (patients 1, 2, and 55). Controls, n 5 6. b Significantly different from controls at time of exhaustion.
276 6 53 3,414 6 711
1.09–1.22
0.024 0.024
112 6 21 2,052 6 444 — 78 6 3.2 1.75 6 0.0 Mean 6 SD
23.7 6 8.1
2,117
b
2,459
c.66411G.A, c.66411G.A 79 1.77
c.66411G.A, c.66411G.A 80 1.78
20 Patient 3
Patient 2
18
1,579 c.2229delT, c.3486_3488delGGA 74 1.71 33 Patient 1a
Neurology 84
Subjects. Three patients with genetically and biochemically verified GSDIIIa and 6 healthy sex- and age-matched controls (2:1) were included (table 1). The patients have been described in detail in a previous article (patients 1, 2, and 5).5 For a detailed description of the exercise interventions, see e-Methods on the Neurology® Web site at Neurology.org.
0.7–1.2
6.4–13.3
6.1 6 0.7 5.5 6 0.6
0.024
0.82 6 0.0
0.80 105
b
0.84 135
1.15 6 0.1
0.85 6 0.2
0.024
10.2 6 2.7
0.024
4.5 6 0.1b 4.9 6 0.5
4.6
4.6
0.82 95
b
1.0 6 0.2
1.6 6 0.2
b
4.4
4.8 1.7
1.8 1.0
0.9
4.4 5.4 1.4 1.2
Glucose peak, mmol/L Glucose rest, mmol/L Lactate peak, mmol/L Lactate rest, mmol/L RER Wpeak, watts mL/min peak,
VO2 AGL gene, mutation Weight, kg Height, m Age, y Participants
Demographic and peak exercise data from patients with glycogen storage disease type IIIa and healthy controls Table 1 1768
METHODS Our aim was to examine substrate turnover during exercise and to examine the effect of substrate supplementation therapy on exercise tolerance in patients with GSDIIIa. The study provides Class IV evidence that fructose ingestion improves exercise tolerance in patients with GSDIIIa.
Substrate turnover during exercise (day 2). We used stable isotope methodology ([U-13C]-palmitate [primed by a NaH13CO3 bolus], [6,6-D2 ]-glucose, and [1,1,2,3,3-D5 ]-glycerol) in combination with indirect calorimetry to examine substrate turnover during submaximal ergometer cycling (60% of VO2peak). The subjects were tested after an overnight fast, and substrate turnover, heart rate, and rate of perceived exertion (RPE) were determined at rest and during exercise. Samples were collected as shown in figure 1. Blood samples were analyzed for concentration and enrichment of plasma palmitate, glucose, and glycerol, and for concentrations of lactate, free fatty acids (FFA), and insulin. Rates of appearance (Ra), disappearance, and oxidation (ROX) were calculated as described in e-Methods. Oral fructose supplementation during exercise (day 3). On day 3, the patients exercised as on day 2, however, this time drinking apple juice (500 mL, containing 9.5 g of carbohydrate per 100 mL) 10 minutes before exercise. Every 20 minutes during exercise, the patients drank an additional 250 mL of juice. Stable isotopes were not used. The effect of fructose was evaluated comparing results obtained on day 2 (stable isotopes 5 placebo) and day 3, respectively, so that patients served as their own controls.
Standard protocol approvals, registrations, and patient consents. Ethics and statistics. The Regional Ethical Committee of the Capital Region of Denmark approved the study (#H-22010-008, with amendment #39559), and it was conducted in accordance with the ethical standards of the Declaration of Helsinki. The subjects gave written informed consent prior to inclusion. Median resting and exercise (averages from the whole period) values were compared between patients and controls with a Mann-Whitney rank-sum test, and a p value , 0.05 was considered significant. Results are reported as mean 6 SE unless otherwise stated. The effect of fructose ingestion was described quantitatively.
The VO2peak, the Wpeak, the respiratory exchange ratio (RER), and the lactate and glucose concentrations at time of exhaustion were significantly lower in the patients compared to controls (table 1). RESULTS Peak exercise testing.
Substrate turnover during exercise. Substrate turnover during exercise is detailed in figures 1 and 2 and table e-1. Total FFA and palmitate ROX at rest were similar between groups. However, during exercise, FFA ROX was higher in the patients. In line with this, the
April 28, 2015
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
Figure 1
Substrate metabolism during exercise in patients with glycogen storage disease type IIIa
(A) Palmitate rate of oxidation (ROX), (B) rate of appearance (Ra) of glucose, (C) total free fatty acid (FFA) ROX, (D) plasma FFA concentration, (E) total carbohydrate (Carb) ROX. and (F) Ra of glycerol in patients with glycogen storage disease type IIIa (GSD, n 5 3) and healthy controls (CON, n 5 6) during submaximal exercise at 60% of VO2peak in the patients. The controls exercised at the same absolute workload as the patients with whom they were matched. Data in A, B, and F were only collected during the stable isotope trial (placebo) on day 2. Patients were also tested with an oral fructose supplement (FRU). *Significantly different from controls (mean of the whole exercise period). **Significant difference between the fructose ingestion trial and the stable isotope (placebo) study (patients only, mean of the whole exercise period, quantitative assessment). Error bars are standard error.
Ra of palmitate and Ra of glycerol and palmitate and FFA concentrations were significantly higher during exercise in the patients. In contrast, the oxidation of carbohydrates during exercise was significantly lower in the patients.
In accordance with this, the RER was also lower in the patients, and, paradoxically, the RER declined during exercise. In patient 1, RER dropped to 0.67, and in patient 3, to 0.68, indicative of ketone body production. Neurology 84
April 28, 2015
1769
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
Figure 2
Workload and Borg score, and lactate and glucose concentrations, placebo vs fructose
(A, B) The workload and duration of exercise and the rate of perceived exertion (RPE) in the placebo (nonfructose experiment, stable isotope) trial. (C, D) The workload and duration of exercise and the RPE after fructose ingestion. Oral fructose ingestion improved exercise tolerance, and 2 patients exercised longer, and all patients rated exercise as being easier with fructose ingestion, and heart rate tended to be lower; 124 6 1.1 vs 119 6 0.7 beats 3 minute21 on fructose. (E) Lactate concentration. (F) Glucose concentration. CON 5 controls (n 5 6); FRU 5 data from the fructose ingestion trial; GSD 5 patients with glycogen storage disease type IIIa (n 5 3). *Significantly different from controls (mean of the whole exercise period). **Significant difference between the fructose ingestion trial and the stable isotope (placebo) study (patients only, mean of the whole exercise period, quantitative assessment). Error bars are standard error.
Glucose levels were consistently lower at all times in the patients and tended to drop during exercise. In line with this, insulin levels were also low in patients. Oral fructose supplementation during exercise. Exercise capacity improved with oral fructose ingestion, which 1770
Neurology 84
was evident from the increased duration that the patients could sustain exercise, a lower heart rate during exercise (in 2 patients), and a lower RPE during exercise in all patients (figures 1 and 2 and table e-2). Fructose ingestion increased blood glucose and lactate levels, RER, and oxidation of carbohydrates.
April 28, 2015
ª 2015 American Academy of Neurology. Unauthorized reproduction of this article is prohibited.
DISCUSSION The main finding of the present study is that patients with GSDIIIa have a severely limited ability to break down glycogen during exercise, which results in energy deficiency and a compensatory increase in the oxidation of fat. These findings are similar to what has been observed in patients with McArdle disease, who also have blocked muscle glycogen breakdown.6 Our findings indicate that the exercise intolerance, which was reported by our patients and other patients, and the exercise-induced symptoms of pain and excessive fatigue that we recently demonstrated in patients with GSDIIIa, most likely can be explained by an energy shortage in muscle during exercise.5,7–9 Similar to findings in patients with McArdle disease, we demonstrate that exercise capacity in GSDIIIa can be improved by raising blood glucose levels.10 This supports energy deficiency as cause of exercise intolerance and suggests a dynamic component of the exerciseinduced symptoms. The findings are not surprising considering the nature of the enzyme defect, but are at variance with the general notion that GSDIIIa is a glycogenosis associated only with static symptoms due to fixed muscle weakness.1,4 More severe exercise-induced muscular symptoms like contractures and rhabdomyolysis are not features of GSDIIIa, unlike McArdle disease, because myophosphorylase can release small amounts of glucose-1-phosphate from glycogen early during exercise, which probably protects skeletal muscle in patients with GSDIIIa. This is supported by the presence of abnormal glycogen, phosphorylase limit-dextrin, in muscle of patients with GSDIIIa.8 However, exercise capacity and CNS function may be severely compromised by hypoglycemia, which evolves rapidly at higher intensities of exercise, due to liver involvement in GSDIIIa.5 The lower glucose and insulin levels could explain the high resting FFA concentrations that we observed in our patients in the present study. The data on muscle substrate turnover consistently indicated a significant block in skeletal muscle glycogenolytic capacity during exercise; however, exercise intolerance differed between the patients, and the effect of fructose ingestion on work capacity and RPE in patient 1 was minimal. Residual enzyme activity or other factors could account for this difference. Therefore it is important to test substrate supplementation therapy in larger populations of patients, also when they are in their habitual metabolic state and not overnight fasted as in previous trials, to establish if our results apply to all patients with GSDIIIa. Still, patients should be cautioned to use oral sucrose/fructose supplements carefully and only in situations preceding exercise, to avoid overuse, weight gain, and associated complications.
AUTHOR CONTRIBUTIONS N. Preisler: design of study, analysis, acquisition, and interpretation of data, and drafting the manuscript. P. Laforet: acquisition of data and critical revision of manuscript. K.L. Madsen: design of study, acquisition of data, and critical revision of manuscript. K. Prahm: acquisition of data and critical revision of manuscript. G. Hedermann: acquisition of data and critical revision of manuscript. C.R. Vissing: acquisition of data and critical revision of manuscript. H. Galbo: interpretation of data and critical revision of the manuscript. J. Vissing: design of study, analysis, interpretation of data, and critical revision of the manuscript.
ACKNOWLEDGMENT The authors thank Danuta Goralska-Olsen, Thomas Lauridsen, Nina Pluszek, and Lene Foged for technical assistance.
STUDY FUNDING Supported by the Aase and Ejnar Danielsens Foundation, Merchant L.F. Foghts Foundation, The A.P. Møller Foundation for the Advancement of Medical Science, and the Sara and Ludvig Elssas Foundation.
DISCLOSURE N. Preisler reports having received research support, honoraria, and travel funding from the Genzyme Corporation. P. Laforêt reports having received research support and honoraria from the Genzyme Corporation. Dr Laforêt is a member of the Genzyme Pompe Disease Advisory Board. K. Madsen, K. Prahm, G. Hedermann, C. Vissing, and H. Galbo report no disclosures relevant to the manuscript. J. Vissing reports having received research support and honoraria from the Genzyme Corporation. He is a member of the Genzyme Pompe Disease Advisory Board. Go to Neurology.org for full disclosures.
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