Reduced Life Expectancy in Rats After Neonatal Dexamethasone Treatment

0031-3998/07/6101-0072 PEDIATRIC RESEARCH Copyright © 2006 International Pediatric Research Foundation, Inc. Vol. 61, No. 1, 2007 Printed in U.S.A. Reduced Life Expectancy in Rats After Neonatal Dexamethasone Treatment PATRICK J. G. H. KAMPHUIS, WILLEM B. DE VRIES, JOOST M. BAKKER, ANNEMIEKE KAVELAARS, JAAP E. VAN DIJK, MARGUERITE E. SCHIPPER, MATTHIJS F. M. VAN OOSTERHOUT, GERDA CROISET, COBI J. HEIJNEN, FRANK VAN BEL, AND VICTOR M. WIEGANT Department of Pharmacology and Anatomy, Rudolph Magnus Institute of Neuroscience [P.J.G.H.K., G.C., V.M.W.], Department of Neonatology [P.J.G.H.K., W.B.V., J.M.B., F.B.], Laboratory for Psychoneuroimmunology [P.J.G.H.K., J.M.B., A.K., C.J.H.], Department of Pathology [M.E.S., M.F.M.O.], University Medical Center Utrecht, 3508 AB Utrecht, The Netherlands; Department of Pathobiology [J.E.D.], Veterinary Faculty, Utrecht University, 3508 TD Utrecht, The Netherlands of prematurely born children treated with Dex during the neonatal period (15,16). Although the available data leave no doubt that neonatal GC treatment can have long-term adverse effects on a wide variety of brain and bodily functions in animals, it remains unclear whether or not this treatment results in higher rates of mortality and cardiovascular morbidity in the long run. In rat pups we have found that neonatal Dex suppresses proliferation of cardiomyocytes leading to a lower number of cardiomyocytes (17). This reduced cardiomyocyte number was associated with a substantial cardiomyocyte hypertrophy and increased fibrosis at adult age (18). Furthermore, Le Cras et al. (19) described that GC treatment of neonatal rats causes lung hypoplasia and increased pulmonary arterial pressures. We hypothesized that the myocardial changes previously found in rats induced by neonatal Dex treatment, whether or not in combination with hemodynamic instability, may have implications for the life expectancy. To test this hypothesis, we compared the survival of rats neonatally treated with Dex with that of saline (Sal) controls. Subsequently, we investigated if the observed effects on mortality were associated with hypertension and/or histopathological alterations in organ tissues. ABSTRACT: The glucocorticoid dexamethasone (Dex) is widely used in preterm infants for the prevention of chronic lung disease. However, major concern has arisen about the long-term sequelae of this therapy. Here we report that neonatal treatment with dexamethasone significantly shortens the lifespan by 25% of male rats (28.6 ⫾ 1.1 to 21.3 ⫾ 0.8 mo) and by 18% of female rats (26.9 ⫾ 1.8 to 22.0 ⫾ 0.7 mo). Histopathological examination indicated end-stage cardiac and renal failure as the cause of premature death. Furthermore, Dextreated rats showed symptoms of hypertension at young adult age, which worsened with increasing age. Thus, a brief period of glucocorticoid treatment during early life results in untimely death presumably due to cardiovascular and renal disease later in life. These serious, adverse long-term consequences call for prudence with glucocorticoid treatment of human preterm infants and careful follow-up of young adults with a history of neonatal glucocorticoid treatment. (Pediatr Res 61: 72–76, 2007) S evere respiratory distress syndrome with concomitant chronic lung disease carries a high incidence of morbidity and mortality (1,2). Inflammation is thought to be an important factor in its pathogenesis. Therefore, treatment with glucocorticoids (GCs), particularly dexamethasone (Dex), was introduced in the eighties resulting in a reduction in chronic lung disease (3–5). Because of these potential benefits in relation to short-term effects on lung function and mechanics (3–5), neonatal Dex became common practice in many centers around the world despite the fact that there had been previous animal studies that showed potential adverse CNS effects of this treatment (6,7). More recently, animal studies indicated that neonatal treatment with GCs affects neurodevelopment and has long lasting adverse effects on cognition and behavior (8,9). In 2000, two papers were published that cautioned the use of Dex during the neonatal period in humans (10,11). Since then, more adverse effects of neonatal GC have been shown in rodents on the immune (12) and neuroendocrine systems (13), on the NMDA receptor (14) and in human follow-up studies MATERIALS AND METHODS Animals. Ten days pregnant Wistar rats (250 –280 g, Central Animal Laboratory, University Medical Center Utrecht, The Netherlands) were housed individually. For the experiments described in this manuscript animals from 22 litters were used. Pups were born on day 22–23 of gestation. On the day of birth (day 0) the pups were removed from their nest and 8 pups (4 females and 4 males) were randomly placed back with each dam. Pups were weaned at day 21 of age and then group-housed with same-sex littermates until experimentation. At weaning animals were randomly assigned to the various experimental conditions. Rats had ad libitum access to food and water. Light/dark cycle (dark phase 1900 – 0700 h), temperature (21°C) and humidity (60%) were kept constant. All experimental procedures were approved by the Committee for Animal Experimentation of the University Medical Center Utrecht. Glucocorticoid treatment design. Rat pups were injected intraperitoneally with dex 21-phosphate (Dex; Organon International B.V., Oss, The Netherlands) on neonatal day 1 (0.5 mg/kg body weight), day 2 (0.3 mg/kg) and day 3 (0.1 mg/kg) or with equal volumes (10 mL/kg) of sterile pyrogen free saline (Sal) for a total of 3 d. All pups in a nest received the same treatment, i.e., either Dex or Sal. The dose and duration of the treatment was based on a 21-d tapering course of Dex (starting dose, 0.5 mg/kg) to prevent or reduce chronic Received May 3, 2006; accepted August 17, 2006. Correspondence: Frank van Bel, M.D., Ph.D., Department of Neonatology, University Medical Center Utrecht/Wilhelmina Children’s Hospital, Roomnr. KE.04.123.1, PO Box 85090, 3508 AB Utrecht, The Netherlands; e-mail: Abbreviations: Dex, dexamethasone; GCs, glucocorticoids; Sal, saline DOI: 10.1203/01.pdr.0000249980.95264.dd 72 REDUCED LIFESPAN AFTER NEONATAL DEX 73 Neonatal dex treatment reduces lifespan. Fig. 1 shows the survival curve of male rats neonatally treated with Dex (n ⫽ 17) or Sal (n ⫽ 18). Dex-treated rats died at considerably younger age than Sal-treated rats (p ⬍ 0.0001). While Saltreated rats on average reached an age of 28.6 ⫾ 1.1 mo, Dex-treated rats reached an age of 21.3 ⫾ 0.8 mo. Thus, neonatal treatment with Dex shortened the average lifespan by approximately 25% (p ⬍ 0.0001). The last Dex-treated male rat died at 27 mo of age. At that age, 50% of the Sal (control) male rats were still alive. The last Sal-treated rat died at 36 mo. Figure 2 shows the survival of female rats neonatally treated with Dex (n ⫽ 13) or Sal (n ⫽ 12). As was the case with Dex-treated males, female Dex (...truncated)