1 Section of Obstetrics and Gynaecology, Imperial College School of Medicine, Chelsea and Westminster Hospital, 369 Fulham Road, London SW10 9NH, 2 Department of Medicine, King's College School of Medicine and Dentistry and 3 Department of Chemical Pathology, Imperial College School of Medicine, Charing Cross Hospital, London W6 9RF, UK
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Abstract |
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Key words: antenatal/dexamethasone/IGF
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Introduction |
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Short-term administration of dexamethasone leads to a reduction of mean IGF bioactivity in the human male, despite an increase in the concentrations of IGF-I (Miell et al., 1993). The change in IGF bioactivity is probably secondary to changes in circulating binding protein concentrations as IGF binding protein (IGFBP)-1 and IGFBP-2 were reduced and IGFBP-3 increased (Miell et al., 1993
). The dexamethasone-induced reduction in IGF bioactivity in the rat is thought to be due to increases in the circulating concentrations of IGFBP-1 (Price et al., 1992
). It has been reported previously that IGFBP-1 was not altered by the antenatal administration of dexamethasone (Ogueh et al., 1998
). However, it is possible that IGFBP-3 may be altered by dexamethasone administration as described above (Miell et al., 1993
). IGFBP-3 inhibits the mitogenic actions of IGF-I in the perfused rat heart, fat cells and chick embryo fibroblasts (Martin and Baxter, 1992
; Lalou et al., 1996
). IGFBP-3 is proteolysed in the circulation, by serine proteases, to two major fragments. The larger 22/25-kDa fragment has low affinity for IGF-I and weakly inhibits IGF-I mitogenic effects, whilst the smaller 16-kDa fragment does not bind to IGF-I, but inhibits IGF-I bioactivity to a similar extent as intact IGFBP-3 (Lalou et al., 1996
, 1997
). Hence, IGFBP-3 proteolysis may inhibit IGF-I-stimulated mitogenesis by a mechanism independent of IGF concentration.
Antenatal dexamethasone administration in the human has only been associated with fetal growth restriction with repeated treatments (French et al., 1999). However, if the fetal growth restriction is mediated through an effect on the growth hormone (GH)IGF axis, it is likely that an effect would be seen following a single treatment course. Indeed, using a total dose of 16 mg in the human male the effects on IGF-bioactivity were profound and persisted for more than 24 h (Miell et al., 1993
). Therefore, given the available human data, to test the hypothesis that antenatal dexamethasone administration to pregnant women is associated with reduced activity of the GHIGF axis, blood samples were obtained pre- and at 24 and 48 h after the administration of 24 mg dexamethasone. In addition, given the relationship between insulin and the GHIGF axis (Holly et al., 1988
; Suikkari et al., 1989
), the concentrations of glucose and insulin were also measured.
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Materials and methods |
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All women gave informed consent to the study, which was approved by the local research ethics committee. Blood samples were collected before starting the dexamethasone therapy, and 24 h and 48 h after completing therapy. The subjects were fasting at the time samples were obtained 24 and 48 h after completing therapy. Samples were centrifuged for 15 min at ~250g at 4°C for the separation of plasma which were stored at 20°C until analysed in a single batch. Statistical analysis was performed using the Statistical Package for Social Sciences (SPSS). Statistical significance was assumed where P < 0.05.
IGF-I was measured by an in-house radioimmunoassay (Morrel et al., 1986) following an acid ethanol extraction. Interassay coefficient of variation (CV) (concentration) was 19.1% (6.7 nmol/l) and 8.7% (21.7 nmol/l). The normal ranges of IGF-I for individuals of <60 years is 3060 nmol/l and for those >60 years it is 1030 nmol/l.
IGF bioactivity was measured using a porcine costal cartilage bioassay as previously described (Miell et al., 1993).
The amount of proteolytic activity directed against recombinant IGFBP-3 was assessed (Lamson et al., 1993). Plasma samples were diluted 1 in 10 with 0.1 mol/l Tris-HCl pH 7.4. A total of 30 µl of dilute sample was incubated for 5h with 10µl 125I-IGFBP-3 (30 000 cpm) and the reaction quenched with 30 µl sample buffer. The samples were subjected to sodium dodecyl sulphatepolyacrylamide gel electrophoresis (SDSPAGE) and the resultant gels dried and autoradiographed. Autoradiographs resulting from the above three methods were assessed by densitometry (SW2000; Ultra-Violet Products Ltd, Cambridge, UK). The density of cleavage fragments appearing as a result of proteolytic activity was calculated and expressed as a percentage of the total density of all bands in each lane to give a percentage protease activity.
GH was measured using the Omnia® radioimmunometric assay (IDS Ltd, Tyne and Wear, UK). Interassay CV (concentration) was 6.4% (1.6 mIU/l) and 8.0% (24.2 mIU/l).
Insulin was measured using an enzyme immunoassay on Boehringer ES700® (Boehringer Mannheim Immunodiagnostics, Lewes, Sussex, UK). Interassay CV (concentration) was 6.0% (88.6 mIU/l) and 10.8% (17.9 mIU/l). Blood glucose was measured by an enzymatic colorimetric assay (glucose oxidase) on the BM/Hitachi 747® (Boehringer Mannheim).
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Results |
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The post-therapy values of IGF bioactivity, IGFBP-3 protease activity, and the plasma concentrations of IGF-I and GH were not different from the pre-therapy concentrations, but the concentrations of glucose and insulin increased 24 h after dexamethasone therapy, and fell towards the pre-therapy value 48 h after therapy (Table I).
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Discussion |
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In pregnant animals, dexamethasone administration is consistently associated with impaired fetal growth due to reduced IGF bioactivity (Luo and Murphy, 1989; Luo et al., 1990
; Price et al., 1992
). In the human, only multiple courses of antenatal corticosteroids have been associated with significant reduction in birth weight (French et al., 1999
). The data presented here are confined to a single treatment course and samples obtained at 24 and 48 h after dexamethasone administration and thus an effect on the GHIGF axis of multiple treatments or a subtle effect of less than 24 h duration cannot be excluded. Nevertheless, the non-pregnant human study used a lower total dosage (16 versus 24 mg) and the pregnant animal data are based on a single treatment (Price et al., 1992
; Miell et al., 1993
). In both studies, profound, protracted effects were reported on the GHIGF axis. If the fetal growth restriction induced in the human by multiple steroid treatments does involve the GHIGF axis it would have been expected that an effect would be apparent in the current study. Moreover, in contrast to Miell et al.'s data, there was a trend for IGF bioactivity actually to increase in the current data. Although this did not reach statistical significance, it supports the idea that fetal growth restriction in the human following dexamethasone administration is not induced through changes in the GHIGF axis.
An alternative explanation for the fetal growth restriction is altered placental function as it has previously been found that the circulating concentrations of human chorionic gonadotrophin were reduced following dexamethasone administration (Ogueh et al., 1999). Indeed, dexamethasone administration has also been reported to cause fetal hypoxia independent of increases in fetal glucose concentrations (Bennett et al., 1999).
Glucocorticoid therapy has been suggested to reduce GH secretion or to alter it in a time dependent mechanism (Hartog et al., 1964; Casanueva et al., 1990
). Casanueva and colleagues found that after administration of 4 mg dexamethasone to eight volunteers, the basal GH concentrations started to rise after 2 h, reached a peak after 33.5 h, and declined after 5 h (Casanueva et al., 1990
). However, in pregnancy, most circulating GH is placental and not pituitary in origin and is secreted in a tonic fashion (Alsat et al., 1998
). Therefore, the lack of change in the concentrations of GH following dexamethasone therapy in this study may be because of the difference in origin or because the samples were obtained only 24 and 48 h after therapy.
Acute and chronic administration of glucocorticoids increases plasma insulin concentrations (Miell et al., 1993). This rise is likely to be secondary to a slight elevation of plasma glucose concentrations (Lambillotte et al., 1997
). In the current study a transient increase in the circulating concentrations of glucose and a more prolonged elevation in insulin concentrations after dexamethasone administration were found. The fact that the glucose concentrations had fallen to close to baseline by 48 h post-treatment suggests that the insulin concentrations would also fall towards baseline thereafter. However, a further study of longer duration is required to establish this. These data contrast with those of a study of 12 women who were given dexamethasone 1 mg orally four times daily for an average of 7.8 days; the circulating concentrations of insulin and glucose did not change during the treatment period when compared to pre-therapy concentrations (Tuimala et al., 1975
). Although the average total dexamethasone dosage of 31.2 mg used is higher than the current dosage of 24 mg, the duration of treatment was longer (Tuimala et al., 1975
). Therefore, the current treatment regimen is likely to have attained a higher concentration of dexamethasone and so induced a more profound, if short lived, effect. Given the overall lack of effect of dexamethasone on the GHIGF axis, the increase in insulin seems not to be significant.
In conclusion, antenatal dexamethasone did not alter the GHIGF axis at 24 and 48 h after administration and had a transient effect on glucose and insulin concentrations. These data do not exclude an effect of antenatal dexamethasone administration on the GHIGF axis, but show that in contrast to non-pregnant human and animal data any effect must be short lived.
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Notes |
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References |
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Submitted on November 11, 1999; accepted on February 29, 2000.