Labordiagnostik bei Wachstumshormon-assoziierten Erkrankungen/Biochemical diagnosis of growth hormone related diseases

The symptoms of growth hormone deficiency and growth hormone excess manifest themselves clinically in different ways, before and after the completion of longitudinal growth. Always, however, biochemical diagnostics is based on the measurement of circulating concentrations of growth hormone and insulin-like growth factor I (IGF-I). Immunoassays are practical, sensitive, and mostly specific methods for measuring either hormone. Still, there are serious discrepancies between the measured results of different assays. These discrepancies are mainly due to differences in the isoform specificity of assays, the use of different standard preparations, as well as the interference of binding proteins. The method-related differences in measured results make the general application of published diagnostic decision limits more difficult. At an interdisciplinary consensus conference, with the participation of the Growth Hormone Research Society, the IGF Society, and the International Federal of Clinical Chemistry and Laboratory Medicine (IFCC), the existing problems were analyzed and possible strategies were highlighted to improve the comparability of the measured results of different GH and IFG-I assays. Currently, however, the use of method-specific reference ranges obtained from well-characterized cohorts continues to be essential in clinical practice.Die Krankheitsbilder des Wachstumshormonmangels und des Wachstumshormonexzesses imponieren klinisch unterschiedlich vor und nach dem Abschluss des Längenwachstums. Stets jedoch bilden die Messung der zirkulierenden Konzentrationen von Wachstumshormon und Insulinartigem Wachstumsfaktor I (Insulin-like growth-factor I, IGF-I) die Basis der laborchemischen Diagnostik. Mit den Immunoassays stehen praktikable, sensitive und meist auch spezifische Methoden zur Messung beider Hormone zur Verfügung. Trotzdem bestehen nach wie vor gravierende Diskrepanzen zwischen den Messergebnissen verschiedener Assays. Diese Diskrepanzen sind vor allem bedingt durch Unterschiede in der Isoformspezifität der Assays, die Verwendung unterschiedlicher Standardpräparationen sowie die Interferenz von Bindungsproteinen. Die methodenbedingten Unterschiede in den Messergebnissen erschweren die allgemeine Anwendung publizierter diagnostischer Entscheidungsgrenzen. Auf einer interdisziplinären Konsensuskonferenz unter Beteiligung der Growth Hormone Research Society, der IGF Society, und der International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) wurden die bestehenden Probleme analysiert und mögliche Strategien zu einer Verbesserung der Vergleichbarkeit der Messergebnisse verschiedener GH- und IGF-I-Assays aufgezeigt. Derzeit bleibt jedoch in der klinischen Praxis die Anwendung methodenspezifischer, an gut charakterisierten Kollektiven gewonnener Referenzbereiche unabdingba

Quantification of growth hormone in serum by isotope dilution mass spectrometry

Inter-assay variation of antibody based routine tests is hampering comparability of measurement results for growth hormone (GH) between different laboratories and decision making in clinical practice. Here it is demonstrated, that quantification of GH by isotope dilution mass spectrometry (IDMS) constitutes a way to precise and reliable results which can be referred to in evaluation of performance of commercial test kits. With the IDMS method developed, tryptic cleavage products YSFLQNPQTSLCFSESIPTPSNR (T6) and LEDGSPR (T12) of GH are quantified by LC/MS-MS using the isotopically labeled forms of the peptides as internal standards. The GH cleavage fragments are obtained by whole-serum tryptic proteolysis and then extracted from the resulting mixture by semi-preparative reversed phase liquid chromatography followed by strong cation-exchange chromatography. Method validation basing on recovery of recombinant 22 kDa GH spiked to blank serum in defined amounts covering the intended concentration range (3-30 µg/L) would yield mean recoveries of 101.6% (100.7%), standard deviations of 2.5% (2.4%) and combined uncertainties (_u~c~_) of 3.0% (2.5%) if quantifying T6 (T12) as GH derived fragments, while the LOQ were 1.7 µg/L (2.7 µg/L). Potential to acquisition of reference values is exemplified by application to serum materials used in a recent quality assessment exercise for routine laboratories

Contribution of the adrenal gland to the production of androstenedione and testosterone during the first two years of life

Androstenedione and testosterone were measured in whole adrenal glands of 56 previously healthy boys who died suddenly between birth and 2 yr of age. In each adrenal gland, the concentration of androstenedione considerably exceeded that of testosterone. The highest concentrations were found during the first week of life (median, 295 ng/g; range, 98- 320 ng/g). Thereafter, values decreased rapidly until the end of the first year of life (median, 10 ng/g; range, 4.4-22.7 ng/g). Adrenal testosterone concentrations averaged 15% of those of androstenedione in the same gland and similarly decreased until the end of the first year. The decrease of adrenal androgen concentrations paralleled the involution of the fetal adrenal zone. A close correlation existed between the concentration of androstenedione in adrenal tissue and plasma. However, no correlation existed between adrenal and plasma testosterone. When the adrenals and testes of the same infant were compared, there was 10 times more androstenedione in the adrenals than in the testes during the first 2 yr of life. The testes contained more testosterone than the adrenals only during the first 4 months. Thus, in infant boys the adrenals are the main source of androstenedione during the first 2 yr. After the sixth month of life, they also are the main source of testosterone

Testosterone and androstenedione concentrations in human testis and epididymis during the first two years of life

Testosterone and androstenedione were measured in testicular and epididymal tissue of 37 previously healthy infants between 1 and 24 months of age who died suddenly. In half of the patients elevated plasma levels of cortisol and androstenedione suggested preterminal stress. Plasma testosterone levels, however, did not differ from those in healthy infants. Testicular testosterone concentrations were maximal in boys from 1-3 months of age (median, 36.6 ng/g; range, 7-380 ng/g) with peak values similar to those found in pubertal or even adult testes. Thereafter testicular testosterone concentrations decreased and after the age of 6 months all values were below 12.5 ng/g, which corresponds to the low normal range of older prepubertal boys. Plasma testosterone and testicular testosterone correlated significantly (P less than 0.001). On average the testicular concentrations were 36.4 times higher than the corresponding plasma concentrations. Testicular androstenedione was low but correlated significantly with testicular testosterone (P less than 0.001). Epididymal testosterone concentrations were surprisingly high (1-3 months: median, 10.3 ng/g; range, 4-42.7 ng/g) and averaged 30% of the testicular testosterone concentration. Thus, epididymal testosterone concentrations were significantly higher than the circulating plasma testosterone levels, indicating the capacity of the infant epididymis to accumulate androgens. These findings suggest that high local testosterone concentrations during early infancy are important not only for the testis itself but particularly for the developing epididymi

Decreasing resistance in the maternal uterine and peripheral arterial system is apparently unrelated to plasma and urinary levels of nitrite/nitrate and cyclic-guanosinmonophosohate during the course of normal pregnancies

Aims: The aim of the presented study was to clarify the relationship between the pulsatility index of the uterine arteries and the maternal cubital artery and peripheral concentrations of the metabolites of nitric oxide (NO) and its second messenger cyclic guanosinmonophophate (cGMP) during the normal course of pregnancy and postpartum. Methods: 49 uncomplicated pregnancies were investigated every 46 weeks until delivery, 29 of them were additionally investigated postpartum. Paralleling each Doppler sonografic investigation maternal blood and urine samples were taken. The measurements of nitrite/ nitrate and cGMP were performed with a colorimetric and radio immuno assay. We demonstrate a significant decrease of the PI of the uterine arteries and of the cubital artery with inverse correlation to advancing gestational age. Results: The concentrations of nitrite/nitrate and cGMP remain stable during gestation and do not correlate to the PI of the uterine and cubital artery. Postpartum a reincrease in the uterine and peripheral resistance can be shown. The concentrations of urinary cGMP and nitrite/ nitrate as well as plasma cGMP remain unchanged, whereas plasma nitrite/nitrate decreases postpartum. Conclusions: The status of NO biosyntheses in normal pregnancy remains controversial. We hypothesize further systemically acting mediators which contribute to the decreasing vascular resistance

Estrone and estradiol concentrations in human ovaries, testes, and adrenals during the first two years of life

To determine the origin of estrogens in infant blood, we measured estrone (E1) and estradiol (E2) in the gonads of 50 girls and 64 boys who died suddenly between birth and 2 yr of age as well as in the adrenals of 18 of these infant girls and 16 of the boys. In the adrenals, E1 [median, 2.8 ng/g (10.4 pmol/g); range, 1.1-4.8 ng/g (4.1- 17.8 pmol/g)] and E2 [median, 3.0 ng/g (10.9 pmol/g); range, 1.2-5.3 ng/g (4.4-19.5 pmol/g)] were found in similar concentrations and were independent of age and sex. In the gonads, E2 was the major estrogen, but the concentrations differed markedly between the sexes; E2 exceeded E1 almost 10-fold in the ovaries and 2-fold in the testes. On the average, the gonads of the infant girls had 5 times more E2 and 2 times more E1 than those of the boys. As in plasma, E2 concentrations were highest in the ovaries of 1- to 6-month-old girls [median, 10.5 ng/g (38.5 pmol/g); range, 1.1-55.1 ng/g (4.0-202.0 pmol/g)] and in testes of 1- to 3-month-old boys [median, 1.8 ng/g (6.6 pmol/g); range, 0.6- 6.4 ng/g (2.3-23.5 pmol/g)]. Ovarian E2 concentrations declined to less than 3.0 ng/g (11.0 pmol/g) by the end of the first year of life, and testicular E2 declined to less than 1.0 ng/g (3.7 pmol/g) after only 6 months of age. Gonadal estrogen concentrations paralleled changes in gonadal morphology. Ovarian weights varied in a pattern of rise and fall similar to that of ovarian E2 concentrations; the biggest ovaries contained multiple macroscopic cysts. Testicular E2 closely correlated with Leydig cell development and testicular testosterone concentrations. We infer, therefore, that the surge of plasma E2 in infant girls originates from ovarian follicles and that of boys from testicular Leydig cells, and that these both occur as a result of the postnatal surge in gonadotropin secretion. The basal plasma E1 and E2 pool, however, is derived from the adrenals and remains at a comparatively constant level in both sexe