Biomedical subjects
H Oftebro
Publications and source records attributed to H Oftebro.
Evaluating an abnormal urinary steroid profile.
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Detection of testosterone administration by increased ratio between serum concentrations of testosterone and 17 alpha-hydroxyprogesterone.
An increased ratio between urinary testosterone (T) and epitestosterone (epiT) has been accepted by the International Olympic Committee as a marker for T doping. However, in a few subjects, we and others have observed constantly above-normal urinary T/epiT ratios that are unlikely to be related to exogenous T administration. To find a better test for T doping, we studied several serum and urinary androgens and androgen precursors, estrogens, and luteinizing hormone (LH) in seven healthy volunteers for 35 days after an intramuscular injection of 250 mg of testosterone enanthate. Among urinary analyses, only the T/epiT ratio was a suitable marker of T doping; of the serum assays, 17 alpha-hydroxyprogesterone (17OHP), T/17OHP ratio, LH, and T/LH ratio were fair to good markers of T doping. The serum T/17OHP ratio was the best marker of those tested, with all seven subjects having above-normal values for this in the first 3 days of the observation period. No other marker showed abnormal values in all subjects at any time. Moreover, the T/17OHP ratio was affected by neither diurnal variation nor physical stress. The value of this marker for T doping was further supported by the finding of normal T/17OHP ratios in a subject with increased urinary T/epiT ratios caused by an abnormally low testicular epiT production, probably related to genetic factors.
Comparison of changes in testosterone concentrations after strength and endurance exercise in well trained men.
Changes in the testosterone concentrations after single sessions of endurance and strength training were measured in seven well trained men, experienced in both forms of training. Both training sessions were rated as hard to very hard on the Borg scale. Blood samples for testosterone measurements were taken before, immediately after, and 2, 4 and 6 h after the training sessions as well as the next morning. The mean testosterone concentration increased 27% (P less than 0.02) and 37% (P less than 0.02) during the strength and endurance training session, respectively. Two hours after the training sessions the mean testosterone concentration had returned to the pre-training level and remained at that level for the length of the observation period. There were no significant differences in the changes in testosterone concentration after strength and endurance training but there were large differences in the testosterone response at the level of the individual. A high correlation (r = 0.98; P less than 0.001) for individuals was found between increases in testosterone concentration after strength and after endurance training. It was concluded that the changes in mean testosterone values followed the same timecourse after single sessions of strength and endurance training of the same duration and perceived exertion. The interindividual differences in testosterone response may be of importance for individual adaptation to training.
Athletics and doping--effects on clinical laboratory analyses.
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Reconstitution of C27-steroid 26-hydroxylase activity from bovine brain mitochondria.
We have previously demonstrated the presence of cytochrome P-450 in a soluble preparation of bovine brain mitochondria (Oftebro, H., Størmer, F.C., and Pedersen, J.I. (1979) J. Biol. Chem. 254, 4331). In the present work we show that this preparation, in the presence of NADPH, adrenodoxin and adrenodoxin reductase catalyzes omega-hydroxylation of a number of C27-steroids that are intermediates in bile acid biosynthesis. The rates of hydroxylation were 1-2 order of magnitudes lower than reported for similar preparations from rat and human liver. No significant activity was detected with cholesterol as substrate. The physiological significance of brain mitochondrial cytochrome P-450 is discussed.
Validation of a radioreceptor assay for 1,25-dihydroxyvitamin D using selected ion monitoring GC-MS.
This report describes two methods for the measurement of 1,25-dihydroxyvitamin D [1,25(OH)2D] in serum: A modified radio receptor assay (RRA), employing a 1,25(OH)2D receptor from calf thymus, and selected ion monitoring (SIM) with combined capillary gas chromatography (GC)-mass spectrometry (MS). The intra-assay coefficient of variation was close to 13% for both methods, and the inter-assay coefficients of variation were 14.0 and 6.5% for RRA and SIM (GC-MS), respectively. Aliquots of 2 ml (RRA) and 20 ml (SIM) serum were used, and the limits of detection were 10 and 6 pmol/l, respectively. The analytical recovery of each method was assessed, and a maximum deviation from the expected value of 10 and 2% was found for RRA and SIM, respectively. A correlation coefficient of 0.93 (slope 0.97) was obtained when 27 different serum samples were analyzed by both methods. Included in this study were serum samples from healthy subjects and patients with subnormal as well as supranormal 1,25(OH)2D levels. This result showed that the RRA accurately measured the serum levels of 1,25(OH)2D and therefore should be useful in the diagnosis and control of vitamin D dependent diseases.
Biochemical changes and endocrine responses in cystic fibrosis in relation to incremental maximal exhaustive exercise.
The purpose of this study was to compare biochemical changes and endocrine responses during an incremental maximal bicycle test in three well-trained 18-year-old patients with cystic fibrosis (CF) and in three healthy controls. Although the blood concentration at rest of the white cell count, haptoglobin, phosphorus, urea, creatinine, and uric acid were somewhat different in the two groups, the CF patients had similar biochemical changes in response to the exercise compared with the healthy men. The endocrine responses to exercise seemed to be different between the two groups with regard to changes in cortisol, growth hormone, and testosterone concentrations. The differences, however, were probably caused by differences in age, training situation, and psychological stress reaction rather than by pathological mechanisms.
Biochemical changes and endocrine responses in cystic fibrosis in relation to a marathon race.
Biochemical changes and endocrine responses during the New York Marathon (42195 m) were investigated in three 18-year-old male adolescents with cystic fibrosis (CF) and three healthy men who accompanied the CFs during the race. The ambient temperature was 20 degrees-28 degrees C and the relative humidity 98%-75% during the run. The CF patients, who had Shwachman scores of 60, 85 and 95 completed the run without major problems in 6.10, 4.42, and 4.32 h, respectively. Serum concentrations of Na and Cl decreased slightly, but the values were still within normal range. Na and Cl excretions in the urine decreased to very low levels in the CF adolescents during the run. All the other biochemical changes were similar to the changes in the controls. Aldosterone concentration increased to a higher level and maintained this increase for a longer time after the race in the CFs. Testosterone concentration decreased more in the CFs during the race compared with the controls. Growth hormone concentration showed individually varying changes in response to the exercise. This study demonstrates that patients with CF may participate in strenuous prolonged exercise even in humid and hot conditions, without untoward effects. The observed differences in hormonal responses to exercise might be explained by differences in age, training status, and relative exercise intensity rather than by hormonal or other disturbances in CF.
Vigorous exercise in a female with cystic fibrosis: spirometric, biochemical, and endocrine responses during four types of intensive exercise.
Spirometric, biochemical, and endocrine responses during a maximal ergometer cycle test and during three runs (10 km, 21.1 km, and 42.2 km) were investigated in one female with cystic fibrosis (CF) 27 years of age and in two healthy control females 26 and 29 years of age. One of the controls ran as a companion to the CF woman, while the other ran at her own speed. The CF woman has a chronic respiratory Pseudomonas aeruginosa infection, and her spirometric values were 50%-70% of predicted values at the time of the study. For the last years she has been training almost daily with aerobics, running, cycling, or skiing. She completed the four types of exercise without major problems. Her spirometric values increased transiently following the cycle test and for several hours following the three races (maximal 20%-30% increase of spirometric values), while the controls had transient decreases of the same variables in response to the runs. The biochemical and the hormonal changes were similar in the CF woman and the control who ran at her own speed, while the control who was a less stressed companion showed smaller changes. This study demonstrates that well-trained females with CF may participate in strenuous prolonged exercise without untoward effects.
Calcium status and calcium-regulating hormones in alcoholics.
To elucidate effects of chronic ethanol consumption on clinical chemical parameters reflecting overall calcium homeostasis 34 hospitalized male alcoholics and 35 age-matched controls were studied during the winter season. Serum concentrations of 25-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D3 were reduced by 28% (p less than 0.01) and 24% (p less than 0.02) among the alcoholics as compared to the controls, respectively. Dietary intake of vitamin D3 did not differ significantly between the groups. The calcium level was below lower limit of reference in nine alcoholics (26%). Serum concentrations of parathyroid hormone and phosphorus were within normal ranges in both groups, and no differences were observed in levels of magnesium, vitamin D-binding protein, calcitonin, or alkaline phosphatase. In conclusion, it is possible that the activities of enzymes crucial in vitamin D3 metabolism may be altered in alcoholics.
Early postmenopausal bone loss is not associated with a decrease in circulating levels of 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D, or vitamin D-binding protein.
To study changes in the rate of bone loss before and after the menopause, 19 normal premenopausal women of the same age were followed with annual measurements of appendicular bone mass for 8 yr. Their serum estrone, estradiol, 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D, and vitamin D-binding protein levels also were measured annually. The serum estrogen levels declined before the menopause, and the rate of bone loss more than doubled at the time of the menopause. There were no significant changes in the serum 25-hydroxyvitamin D, 1,25-dihydroxyvitamin LD or vitamin D-binding protein levels from the pre- to the postmenopausal period. We conclude that changes in vitamin D metabolite levels are not associated with early postmenopausal bone loss.
Subcellular localization of vitamin D3 25-hydroxylase in human liver.
Vitamin D3 25-hydroxylase activity was measured in subcellular and submitochondrial fractions of human liver. Quantitation of 25-hydroxyvitamin D3 was based on high performance liquid chromatography. Vitamin D3 25-hydroxylase activity was detected in the mitochondrial fraction only. The mitochondrial 25-hydroxylase activity was linear with time up to 60 min and with mitochondrial protein up to 1 mg/ml. An apparent Km value of about 10(-5) M was found. Substrate satuation level was not reached. In the presence of 2.4 X 10(-4) M vitamin D3, the rate of 25-hydroxyvitamin D3 formation was 0.19 nmol X mg of protein-1 X h-1 After fractionation of the mitochondria, 86% of the 25-hydroxylase activity was recovered in the mitoplast fraction. The outer membrane fraction was devoid of activity. It is concluded that human liver contains only one detectable vitamin D3 25-hydroxylase enzyme localized to the mitochondrial inner membrane.
Role of the 26-hydroxylase in the biosynthesis of bile acids in the normal state and in cerebrotendinous xanthomatosis. An in vivo study.
On the basis of different in vitro studies, we have previously suggested that the basic metabolic defect in the rare inherited disease cerebrotendinous xanthomatosis (CTX) is a lack of a hepatic mitochondrial C27-steroid 26-hydroxylase, involved in the normal biosynthesis of bile acids (1980. J. Clin. Invest. 65: 1418-1430; 1981. J. Lipid Res. 22: 191-200; 22: 632-640). In the present work, this hypothesis was tested in vivo. One patient with CTX and two control subjects received intravenously a mixture of [4-14C]7 alpha-hydroxy-4-cholesten-3-one and [6 beta-3H]7 alpha,26-dihydroxy-4-cholesten-3-one, steroids believed to be important precursors of chenodeoxycholic acid. The ratio between 14C and 3H in cholic acid and chenodeoxycholic acid isolated from bile of the CTX-patient was approximately 1/40 and 1/60 of those of the control subjects, respectively. Another patient with CTX and one control subject received a mixture of [4-14C]5 beta-cholestane-3 alpha,7 alpha-diol and [1,2-3H]5 beta-cholestane-3 alpha,7 alpha,26-triol, both possible precursors to chenodeoxycholic acid. In this case the 14C/3H ratio in cholic acid and chenodeoxycholic acid from the patient with CTX was 1/10 and 1/15, respectively, compared with that of the control subject. The most likely explanation for these findings is that very little of the 14C-precursors, i.e. without a 26-hydroxyl group, can be converted into cholic acid and chenodeoxycholic acid because of a defect of the 26-hydroxylase step. The results obtained are in accord with our previous findings in vitro. The results further underline the importance of the 26-hydroxylase pathway in the normal biosynthesis of cholic acid and chenodeoxycholic acid in man.
Side chain hydroxylation of C27-steroids and vitamin D3 by a cytochrome P-450 enzyme system isolated from human liver mitochondria.
The present study was undertaken to obtain information on the involvement of cytochrome P-450 in the 26-hydroxylation on bile acid intermediates and in the 25-hydroxylation of vitamin D3 in human liver mitochondria. Cytochrome P-450 was solubilized from human liver mitochondria and purified two times to a specific content of 0.125 nmol per mg protein. Furthermore, a ferredoxin was isolated from the mitochondria and partly purified. This iron-sulfur protein had properties similar to bovine adrenal ferredoxin. A mitochondrial NADPH-ferredoxin reductase was also isolated and purified to homogeneity. This enzyme was a flavoprotein with properties very similar to the bovine adrenal NADPH-ferredoxin reductase. The cytochrome P-450 preparation catalyzed 26-hydroxylation of C27-steroids and 25-hydroxylation of vitamin D3 when reconstructed with NADPH, the ferredoxin and the ferredoxin reductase. With different substrates the following turnover numbers (nmol product X nmol P-450(-1) X min-1) were found: cholesterol, 8; 5-cholestene-3 beta, 7 alpha-diol, 10; 7 alpha-hydroxy-4-cholesten-3-one, 23; 7 alpha, 12 alpha-dihydroxy-4-cholesten-3-one, 27; 5 beta-cholestane-3 alpha, 7 alpha-diol, 28; 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol, 41; and vitamin D3, 0.16. The hydroxylation reactions were inhibited by CO and metyrapone. The human liver mitochondrial ferredoxin and ferredoxin reductase could be replaced by adrenal ferredoxin and adrenal ferredoxin reductase without reduction of activity, but they could not be replaced by microsomal NADPH-cytochrome P-450 reductase. It is concluded that human liver mitochondria contain cytochrome P-450 involved in the oxidation of the side chain of C27-steroids and vitamin D3.
Cerebrotendinous xanthomatosis: defective liver mitochondrial hydroxylation of chenodeoxycholic acid precursors.
Oxidation of the side chain of 5 beta-cholestane-3 alpha, 7 alpha-diol, 7 alpha-hydroxy-4-cholesten-3-one, and 5-cholestene-3 beta, 7 alpha-diol has been studied in subcellular fractions of liver from a patient with cerebrotendinous xanthomatosis (CTX) and a control subject. All intermediates were efficiently 26-hydroxylated and further converted to the corresponding 26-carboxylated derivatives by the mitochondrial fraction of normal human liver. No such conversion was observed with the mitochondria from the liver of the CTX patient and the control subject. 12 alpha-Hydroxylation of the patient and the control subject. 12 alpha-Hydroxylation of the substrates was very efficient with the microsomal fractions from both subjects. Bases on these and previous findings (Oftebro, H., I. Björkhem, S. Skrede, A. Schreiner, and J. I. Pedersen. 1980. J. Clin. Invest. 65: 1481-1430), it i concluded that the metabolic defect in CTX is a complete lack of mitochondrial C27-steroid 26-hydroxylase. In CTX the precursors of chenodeoxycholic acid are first attacked by the microsomal 12 alpha-hydroxylase and subsequently by the microsomal 25-hydroxylase as an alternate route to cholic acid formation. This explains the increased ratio of cholic acid to chenodeoxycholic acid observed in the bile of these patients. In the normal liver the formation of both cholic acid and chenodeoxycholic acid involves a mitochondrial 26-hydroxylation.
Properties of a reconstituted vitamin D3 25-hydroxylase from rat liver mitochondria.
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