- Email: [email protected]
Myoplasmic Ca2+ concentration during exertional rhabdomyolysis
intake. However, in half the subjects (heterozygotes Bb) the rate of change was related to overall calcium intake. Thus the observed variability in elderly, and presumably younger, people in the response of bone mass to calcium intake and supplementation may be explained partly by the interaction between environmental and genetic factors.’° Analysis of VDR-gene polymorphisms may allow better definition and prediction of these responses. We thank Mrs M N Cerutti for data collection. This work was supported by a grant from the Swiss National Science Foundation (number
5
6 7
8
1637-42.
32-2415-91). 9
10
Smith DA, Nance WE, Won Kang K, Christian JC, Johnston CC Jr. Genetic factors in determining bone mass. J Clin Invest 1973; 52: 2800-08. 2 Pocock NA, Eisman JA, Hopper JL, Yeates MG, Sambrook PN, Eberl S. Genetic determinants of bone mass in adults: a twin study. J Clin Invest 1987; 80: 706-10. 3 Dequeker J, Nijs J, Verstraeten A, Geusens P, Gevers G. Genetic determinants of bone mineral content at the spine and the radius: a twin study. Bone 1987; 8: 207-09. 4 Seeman E, Hopper JL, Bach LA, et al. Reduced bone mass in daughters of women with osteoporosis. N Engl J Med 1989; 320: 554-58.
Myoplasmic Ca2+ concentration during exertional rhabdomyolysis
Exertional rhabdomyolysis can destroy muscle but the pathophysiology is unknown. Using intracellular selective microelectrodes, we found that intracellular Ca2+ concentration ([Ca2+]i) was 1·27 (0·17) µmol/L (median and interquartile range) in skeletal-muscle biopsy from specimens patients with exertional rhabdomyolysis compared with 0·12 (0·01) µmol/L in controls. 3 days treatment with dantrolene, a drug that inhibits Ca2+ release from the sarcoplasmic reticulum, decreased [Ca2+]i to 0·22 (0·05) µmol/L and accelerated patients’ recovery. This study demonstrated that exertional rhabdomyolysis is associated with elevated [Ca2+]i, and that dantrolene has a beneficial effect in this syndrome. Lancet 1995; 345: 424-25
The
most common cause of life-threatening is Exertional exercise. rhabdomyolysis physical rhabdomyolysis is characterised by muscle stiffness, rigidity, pain, and increased serum creatine kinase and myoglobin.1,2 Although intracellular Ca2+ concentration ([Ca2+]i) may be disturbed, the pathogenesis is not
understood and treatment is limited to the symptoms of the acute episode. We have directly measured myoplasmic Ca2+ in muscle fibres from patients during an acute episode of exertional rhabdomyolysis. Eleven patients (two female and three male athletes, and six male army recruits; median age 22, interquartile range 3) were with exertional rhabdomyolysis based on history (prolonged strenuous exercise), clinical presentation (generalised muscle stiffness, rigidity, pain, decreases in exercise tolerance), and laboratory studies (serum creatine kinase 8965 [4426] IU/L, normal <70; serum myoglobin 5430 [1951] ng/mL, normal <80; admitted
424
Chevalley T, Rizzoli R, Nydegger V, et al. Effects of calcium supplements on femoral bone mineral density and vertebral fracture rate in vitamin D-replete elderly patients. Osteoporosis Int 1994; 4: 245-52.
References 1
Morrison NA, Yeoman R, Kelly PJ, Eisman JA. Contribution of transacting factor alleles to normal physiological variability: vitamin D receptor polymorphisms and circulating osteocalcin. Proc Natl Acad Sci USA 1992; 89: 6665-69. Morrison NA, Qi J-C, Tokita A, et al. Prediction of bone density by vitamin D receptor alleles. Nature 1994; 367: 284-87. Dawson-Hughes B, Dallal GE, Krall EA, Sadowski L, Sahyoun N, Tannenbaum S. A controlled trial of the effect of calcium supplementation on bone density in post-menopausal women. N Engl J Med 1990; 323: 878-83. Chapuy MC, Arlot ME, Duboeuf F, et al. Vitamin D3 and calcium prevent hip fractures in elderly women. N Engl J Med 1992; 327:
Kelly PJ, Sambrook PN, Eisman JA. Interaction of genetic and environmental influences on peak bone density. Osteoporosis Int 1990; 1: 56-60.
Division of Clinical Pathophysiology, WHO Collaborating Centre for Osteoporosis and Bone Disease, Department of Medicine (S Ferrari MD, R Rizzoli MD, T Chevalley MD, Prof J A Eisman PhD, Prof J-P Bonjour MD); and Division of Nuclear Medicine, Department of Radiology (D Slosman MD), University Hospital of Geneva, 1211 Geneva, Switzerland
Correspondence to:
Dr R Rizzoli
ng/mL). Electromyography showed fasciculation and decreased amplitude of motor units. Serum electrolytes, rectal temperature, respiratory function, and renal output were normal. The subjects had no previous episodes of rhabdomyolysis, trauma, infection, malnutrition, or intoxication. The average ambient temperature at the time of the acute episode was 23°C (SE 1). Because of the association between rhabdomyolysis and malignant hyperthermia, ten of the eleven patients were tested for susceptibility to malignant hyperthermia with the caffeine-contracture test; seven were positive. Eight subjects with no neuromuscular disease were used as controls (one female and seven males; age 28 [5] years). Intact external intercostal muscle fibres were obtained under local anaesthesia from patients at admission (day 1) and 3 days later (day 4). In six controls, muscle biopsy specimens were obtained during thoracotomy for different surgical procedures; in two, muscle biopsies were done under local anaesthesia. In all cases, muscle biopsy specimens were collected with consent and approval of our human ethics committee. Intracellular resting [Ca=*]i was measured with doublebarrelled Ca2+ -selective microelectrodes that were impaled into a single muscle fibre stretched to 2-4 o.m sarcomere length.3 [Ca2+]i was measured only in polarised fibres with a resting membrane potential above -80 mV. Microelectrodes were calibrated individually with solutions of known Ca2* concentrations and those microelectrodes that gave an almost Nernstian response between 10-3 and 10-7 mol/L (30-5 mV per tenfold change in [Ca2*] at 37°C) were used. We have used Ca2+-selective microelectrodes to measure [Ca2*]i in other human muscle diseases.3-5 The experimental chamber was perfused with Krebs’ solution (37°C [1]) which was bubbled with 95% O2 and 5% CO, (pH 7-4). Results are expressed as medians and interquartile
myoglobinuria >14634
ranges.
Patients with exertional rhabdomyolysis were treated with isotonic intravenous fluid (250 mL/h) and 25% mannitol (100 mL per day) which were administered on days 1-3. Analgesics were provided on patient’s request. If increased [Ca2*]i is important in the pathogenesis of rhabdomyolysis, an agent that decreases [Ca 2,]might be beneficial in the treatment of exertional rhabdomyolysis. Six patients received dantrolene 2-5 mg/kg intravenously on day 1 and 2-0 mg/kg orally on days 2-4. Patients were discharged on day 6 after admission.
[Ca2+]i was eleven times higher in fibres from patients during acute rhabdomyolysis (day 1) than in those from
References 1
2 3
4
5
6
7
8
Controls
Patients
Figure: [Ca2+]i in intercostal muscles from patients in exertional rhabdomyolysis and from controls
acute
n=number of muscle fibres.
controls. Resting [Ca2*]i was 0-12 (0-01) )JLmoI/L in 35 fibres from eight control subjects and 1-27 (0-17 fJ.mol/L in 60 fibres from eleven patients during the
rhabdomyolytic episode (figure). Dantrolene reversed the increase in resting [Ca2+]iin fibres from patients with rhabdomyolysis (figure). On day 4, [Ca2+]iwas 0-22 (0-05) )JLmol/L in 32 fibres from six patients after dantrolene compared with 0-79 (0-09) j.1mol/L in 40 fibres from five patients who did not receive dantrolene. The 83% reduction in [Ca2*]i after dantrolene treatment was associated with an improvement of clinical symptoms (muscle stiffness, rigidity, and pain) and of laboratory values: on day 4, serum creatine kinase was 875 (172) and 5025 (2811) IU/L in dantrolene-treated versus non-treated patients. Serum myoglobin was 598 (322) and 3960 (1604) ng/mL, respectively. Two controls were treated with dantrolene at the same dose and schedule as the patients. [Ca 2+]in 13 fibres decreased from 0-10 (0-01) jjbmol/L on day 1 to 0-05 (0-01) µmol/L on day 4 in 12 fibres. Because of the short-term use of dantrolene, the only side-effect was weakness in arms and legs reported by four of six treated patients and in one control. This weakness lasted up to 48 h after withdrawal of dantrolene. We found that [Ca2*]i was increased in muscle fibres from patients with exertional rhabdomyolysis. The mechanism is unknown, but may include deficiencies in Ca2+-ATPase from the sarcoplasmic reticulum6 and sarcolemma (data from JRL). Whether the increase in [Ca2*]i precedes the episode of rhabdomyolysis also remains to be determined. The observation that dantrolene, a known inhibitor of Ca2* release from the sarcoplasmic reticulum,7,8 has a beneficial effect on the course of the rhabdomyolytic episode further suggests that myoplasmic Ca2* has an important role in the pathophysiology of this syndrome. A correction in myoplasmic Ca2+ should be considered in the treatment of exertional rhabdomyolysis. We thank Procter and Gamble for providing dantrolene sodium. AT receives a faculty developmental award in clinical pharmacology from the PhRMA Foundation and grant-in-aid from the AHA.
Milne JC. Rhabdomyolysis, myoglobinuria and exercise. Sports Med 1988; 6: 93-106. Knochel JP. Mechanism of rhabdomyolysis. Curr Opin Rheumatol 1993; 5: 725-31 López JR, Gerardi A, López MJ, Allen PD. Effect of dantrolene on myoplasmic free [Ca2+], measured in vivo in patients susceptible to malignant hyperthermia. Anesthesiology 1992; 76: 711—19. López JR, Sánchez V, López MJ. Sarcoplasmic ionic calcium concentration in neuroleptic malignant syndrome. Cell Calcium 1989; 10: 223-33. López JR. Free calcium concentration in skeletal muscle of malignant hyperthermia susceptible subjects: effects of ryanodine. In: Ohnishi ST, Ohnishi T, eds. Malignant hyperthermia: a genetic membrane disease. Boca Raton: CRC Press, 1994; 134-50. Poels PJE. Wevers RA, Braakhekke JP, Benders AAGM, Veerkamp JH, Joosten EMG. Exertional rhabdomyolysis in a patient with calcium adenosine triphosphatase deficiency. J Neurol Neurosurg Psychiatry 1993; 56: 823-26. Desmedt JE, Hainaut K. Inhibition of intracellular release of calcium by dantrolene in barnacle giant muscle fibers. J Physiol 1977; 265: 565-85. Van Winkle WB. Calcium release from skeletal muscle sarcoplasmic reticulum: site of action of dantrolene sodium? Science 1979; 193: 1130-31.
Centro de Biofisica y
Bioquimica, Instituto Venezolano de Investigaciones Científicas, Caracas, Venezuela (J R López MD, M A Gonzalez MD); Departamento de Medicina Interna, Hospital Militar Carlos Arvelo, Caracas, Venezuela (B Rojas MD); and Division of Cardiovascular Diseases and Clinical Pharmacology Unit, Departments of Internal Medicine and Pharmacology, Mayo Clinic, Mayo Foundation, Rochester, MN 55905, USA (J R López, A Terzic MD)
Correspondence to: Dr José R López
Reduced experimental contact sensitivity in squamous cell but not basal cell carcinomas of skin
Basal cell and squamous cell carcinomas (BCC) carcinomas (SCC) show clinical and epidemiological differences not accounted for by different ultraviolet radiation exposure. We have studied experimentally induced contact sensitivity to dinitrochlorobenzene by measuring increases in skin-fold thickness. Patients (n=37) with squamous tumours had impaired responses compared with controls (33) and patients with BCCs (31) (mean increase 4·5 vs 7·8 and 8·6 mm, respectively; p=0·002). This diminished immunological response may be causally related to the development of SCC. Because glutathione S-transferase (GST) metabolises dinitrochlorobenzene and polymorphisms of GST are associated with multiple skin tumours, variations in GST may underlie these differences. Lancet 1995; 345: 425-26
Ultraviolet radiation is a major cause of non-melanoma skin cancer (NMSC).’ However, the differences in bodysite distribution of basal cell carcinomas (BCCs) and squamous cell cancers (SCC), Bowen’s disease, and actinic keratoses, and changes in the ratio of the numbers of BCC to SCC in different risk groups and populations, suggest that there are other causes.2,3 An intact immune system may be important in preventing skin neoplasms: patients on immunosuppressive therapy after organ 425
5
6 7
8
1637-42.
32-2415-91). 9
10
Smith DA, Nance WE, Won Kang K, Christian JC, Johnston CC Jr. Genetic factors in determining bone mass. J Clin Invest 1973; 52: 2800-08. 2 Pocock NA, Eisman JA, Hopper JL, Yeates MG, Sambrook PN, Eberl S. Genetic determinants of bone mass in adults: a twin study. J Clin Invest 1987; 80: 706-10. 3 Dequeker J, Nijs J, Verstraeten A, Geusens P, Gevers G. Genetic determinants of bone mineral content at the spine and the radius: a twin study. Bone 1987; 8: 207-09. 4 Seeman E, Hopper JL, Bach LA, et al. Reduced bone mass in daughters of women with osteoporosis. N Engl J Med 1989; 320: 554-58.
Myoplasmic Ca2+ concentration during exertional rhabdomyolysis
Exertional rhabdomyolysis can destroy muscle but the pathophysiology is unknown. Using intracellular selective microelectrodes, we found that intracellular Ca2+ concentration ([Ca2+]i) was 1·27 (0·17) µmol/L (median and interquartile range) in skeletal-muscle biopsy from specimens patients with exertional rhabdomyolysis compared with 0·12 (0·01) µmol/L in controls. 3 days treatment with dantrolene, a drug that inhibits Ca2+ release from the sarcoplasmic reticulum, decreased [Ca2+]i to 0·22 (0·05) µmol/L and accelerated patients’ recovery. This study demonstrated that exertional rhabdomyolysis is associated with elevated [Ca2+]i, and that dantrolene has a beneficial effect in this syndrome. Lancet 1995; 345: 424-25
The
most common cause of life-threatening is Exertional exercise. rhabdomyolysis physical rhabdomyolysis is characterised by muscle stiffness, rigidity, pain, and increased serum creatine kinase and myoglobin.1,2 Although intracellular Ca2+ concentration ([Ca2+]i) may be disturbed, the pathogenesis is not
understood and treatment is limited to the symptoms of the acute episode. We have directly measured myoplasmic Ca2+ in muscle fibres from patients during an acute episode of exertional rhabdomyolysis. Eleven patients (two female and three male athletes, and six male army recruits; median age 22, interquartile range 3) were with exertional rhabdomyolysis based on history (prolonged strenuous exercise), clinical presentation (generalised muscle stiffness, rigidity, pain, decreases in exercise tolerance), and laboratory studies (serum creatine kinase 8965 [4426] IU/L, normal <70; serum myoglobin 5430 [1951] ng/mL, normal <80; admitted
424
Chevalley T, Rizzoli R, Nydegger V, et al. Effects of calcium supplements on femoral bone mineral density and vertebral fracture rate in vitamin D-replete elderly patients. Osteoporosis Int 1994; 4: 245-52.
References 1
Morrison NA, Yeoman R, Kelly PJ, Eisman JA. Contribution of transacting factor alleles to normal physiological variability: vitamin D receptor polymorphisms and circulating osteocalcin. Proc Natl Acad Sci USA 1992; 89: 6665-69. Morrison NA, Qi J-C, Tokita A, et al. Prediction of bone density by vitamin D receptor alleles. Nature 1994; 367: 284-87. Dawson-Hughes B, Dallal GE, Krall EA, Sadowski L, Sahyoun N, Tannenbaum S. A controlled trial of the effect of calcium supplementation on bone density in post-menopausal women. N Engl J Med 1990; 323: 878-83. Chapuy MC, Arlot ME, Duboeuf F, et al. Vitamin D3 and calcium prevent hip fractures in elderly women. N Engl J Med 1992; 327:
Kelly PJ, Sambrook PN, Eisman JA. Interaction of genetic and environmental influences on peak bone density. Osteoporosis Int 1990; 1: 56-60.
Division of Clinical Pathophysiology, WHO Collaborating Centre for Osteoporosis and Bone Disease, Department of Medicine (S Ferrari MD, R Rizzoli MD, T Chevalley MD, Prof J A Eisman PhD, Prof J-P Bonjour MD); and Division of Nuclear Medicine, Department of Radiology (D Slosman MD), University Hospital of Geneva, 1211 Geneva, Switzerland
Correspondence to:
Dr R Rizzoli
ng/mL). Electromyography showed fasciculation and decreased amplitude of motor units. Serum electrolytes, rectal temperature, respiratory function, and renal output were normal. The subjects had no previous episodes of rhabdomyolysis, trauma, infection, malnutrition, or intoxication. The average ambient temperature at the time of the acute episode was 23°C (SE 1). Because of the association between rhabdomyolysis and malignant hyperthermia, ten of the eleven patients were tested for susceptibility to malignant hyperthermia with the caffeine-contracture test; seven were positive. Eight subjects with no neuromuscular disease were used as controls (one female and seven males; age 28 [5] years). Intact external intercostal muscle fibres were obtained under local anaesthesia from patients at admission (day 1) and 3 days later (day 4). In six controls, muscle biopsy specimens were obtained during thoracotomy for different surgical procedures; in two, muscle biopsies were done under local anaesthesia. In all cases, muscle biopsy specimens were collected with consent and approval of our human ethics committee. Intracellular resting [Ca=*]i was measured with doublebarrelled Ca2+ -selective microelectrodes that were impaled into a single muscle fibre stretched to 2-4 o.m sarcomere length.3 [Ca2+]i was measured only in polarised fibres with a resting membrane potential above -80 mV. Microelectrodes were calibrated individually with solutions of known Ca2* concentrations and those microelectrodes that gave an almost Nernstian response between 10-3 and 10-7 mol/L (30-5 mV per tenfold change in [Ca2*] at 37°C) were used. We have used Ca2+-selective microelectrodes to measure [Ca2*]i in other human muscle diseases.3-5 The experimental chamber was perfused with Krebs’ solution (37°C [1]) which was bubbled with 95% O2 and 5% CO, (pH 7-4). Results are expressed as medians and interquartile
myoglobinuria >14634
ranges.
Patients with exertional rhabdomyolysis were treated with isotonic intravenous fluid (250 mL/h) and 25% mannitol (100 mL per day) which were administered on days 1-3. Analgesics were provided on patient’s request. If increased [Ca2*]i is important in the pathogenesis of rhabdomyolysis, an agent that decreases [Ca 2,]might be beneficial in the treatment of exertional rhabdomyolysis. Six patients received dantrolene 2-5 mg/kg intravenously on day 1 and 2-0 mg/kg orally on days 2-4. Patients were discharged on day 6 after admission.
[Ca2+]i was eleven times higher in fibres from patients during acute rhabdomyolysis (day 1) than in those from
References 1
2 3
4
5
6
7
8
Controls
Patients
Figure: [Ca2+]i in intercostal muscles from patients in exertional rhabdomyolysis and from controls
acute
n=number of muscle fibres.
controls. Resting [Ca2*]i was 0-12 (0-01) )JLmoI/L in 35 fibres from eight control subjects and 1-27 (0-17 fJ.mol/L in 60 fibres from eleven patients during the
rhabdomyolytic episode (figure). Dantrolene reversed the increase in resting [Ca2+]iin fibres from patients with rhabdomyolysis (figure). On day 4, [Ca2+]iwas 0-22 (0-05) )JLmol/L in 32 fibres from six patients after dantrolene compared with 0-79 (0-09) j.1mol/L in 40 fibres from five patients who did not receive dantrolene. The 83% reduction in [Ca2*]i after dantrolene treatment was associated with an improvement of clinical symptoms (muscle stiffness, rigidity, and pain) and of laboratory values: on day 4, serum creatine kinase was 875 (172) and 5025 (2811) IU/L in dantrolene-treated versus non-treated patients. Serum myoglobin was 598 (322) and 3960 (1604) ng/mL, respectively. Two controls were treated with dantrolene at the same dose and schedule as the patients. [Ca 2+]in 13 fibres decreased from 0-10 (0-01) jjbmol/L on day 1 to 0-05 (0-01) µmol/L on day 4 in 12 fibres. Because of the short-term use of dantrolene, the only side-effect was weakness in arms and legs reported by four of six treated patients and in one control. This weakness lasted up to 48 h after withdrawal of dantrolene. We found that [Ca2*]i was increased in muscle fibres from patients with exertional rhabdomyolysis. The mechanism is unknown, but may include deficiencies in Ca2+-ATPase from the sarcoplasmic reticulum6 and sarcolemma (data from JRL). Whether the increase in [Ca2*]i precedes the episode of rhabdomyolysis also remains to be determined. The observation that dantrolene, a known inhibitor of Ca2* release from the sarcoplasmic reticulum,7,8 has a beneficial effect on the course of the rhabdomyolytic episode further suggests that myoplasmic Ca2* has an important role in the pathophysiology of this syndrome. A correction in myoplasmic Ca2+ should be considered in the treatment of exertional rhabdomyolysis. We thank Procter and Gamble for providing dantrolene sodium. AT receives a faculty developmental award in clinical pharmacology from the PhRMA Foundation and grant-in-aid from the AHA.
Milne JC. Rhabdomyolysis, myoglobinuria and exercise. Sports Med 1988; 6: 93-106. Knochel JP. Mechanism of rhabdomyolysis. Curr Opin Rheumatol 1993; 5: 725-31 López JR, Gerardi A, López MJ, Allen PD. Effect of dantrolene on myoplasmic free [Ca2+], measured in vivo in patients susceptible to malignant hyperthermia. Anesthesiology 1992; 76: 711—19. López JR, Sánchez V, López MJ. Sarcoplasmic ionic calcium concentration in neuroleptic malignant syndrome. Cell Calcium 1989; 10: 223-33. López JR. Free calcium concentration in skeletal muscle of malignant hyperthermia susceptible subjects: effects of ryanodine. In: Ohnishi ST, Ohnishi T, eds. Malignant hyperthermia: a genetic membrane disease. Boca Raton: CRC Press, 1994; 134-50. Poels PJE. Wevers RA, Braakhekke JP, Benders AAGM, Veerkamp JH, Joosten EMG. Exertional rhabdomyolysis in a patient with calcium adenosine triphosphatase deficiency. J Neurol Neurosurg Psychiatry 1993; 56: 823-26. Desmedt JE, Hainaut K. Inhibition of intracellular release of calcium by dantrolene in barnacle giant muscle fibers. J Physiol 1977; 265: 565-85. Van Winkle WB. Calcium release from skeletal muscle sarcoplasmic reticulum: site of action of dantrolene sodium? Science 1979; 193: 1130-31.
Centro de Biofisica y
Bioquimica, Instituto Venezolano de Investigaciones Científicas, Caracas, Venezuela (J R López MD, M A Gonzalez MD); Departamento de Medicina Interna, Hospital Militar Carlos Arvelo, Caracas, Venezuela (B Rojas MD); and Division of Cardiovascular Diseases and Clinical Pharmacology Unit, Departments of Internal Medicine and Pharmacology, Mayo Clinic, Mayo Foundation, Rochester, MN 55905, USA (J R López, A Terzic MD)
Correspondence to: Dr José R López
Reduced experimental contact sensitivity in squamous cell but not basal cell carcinomas of skin
Basal cell and squamous cell carcinomas (BCC) carcinomas (SCC) show clinical and epidemiological differences not accounted for by different ultraviolet radiation exposure. We have studied experimentally induced contact sensitivity to dinitrochlorobenzene by measuring increases in skin-fold thickness. Patients (n=37) with squamous tumours had impaired responses compared with controls (33) and patients with BCCs (31) (mean increase 4·5 vs 7·8 and 8·6 mm, respectively; p=0·002). This diminished immunological response may be causally related to the development of SCC. Because glutathione S-transferase (GST) metabolises dinitrochlorobenzene and polymorphisms of GST are associated with multiple skin tumours, variations in GST may underlie these differences. Lancet 1995; 345: 425-26
Ultraviolet radiation is a major cause of non-melanoma skin cancer (NMSC).’ However, the differences in bodysite distribution of basal cell carcinomas (BCCs) and squamous cell cancers (SCC), Bowen’s disease, and actinic keratoses, and changes in the ratio of the numbers of BCC to SCC in different risk groups and populations, suggest that there are other causes.2,3 An intact immune system may be important in preventing skin neoplasms: patients on immunosuppressive therapy after organ 425