Combined Statin and Glucocorticoid Therapy for the Safer Treatment of Preterm Birth

Abstract

Background: Prematurity is strongly associated with poor respiratory function in the neonate. Rescue therapies include treatment with glucocorticoids due to their anti-inflammatory and maturational effects on the developing lung. However, glucocorticoid treatment in the infant can increase the risk of long-term cardiovascular complications including hypertension, cardiac, and endothelial dysfunction. Accumulating evidence implicates a molecular link between glucocorticoid excess and depletion of nitric oxide (NO) bioavailability as a mechanism underlying the detrimental effects of postnatal steroids on the heart and circulation. Therefore, combined glucocorticoid and statin therapy, by increasing NO bioavailability, may protect the developing cardiovascular system while maintaining beneficial effects on the lung.

Methods: We investigated combined glucocorticoid and statin therapy using an established rodent model of prematurity and combined experiments of cardiovascular function in vivo, with those in isolated organs as well as measurements at the cellular and molecular levels.

Results: We show that neonatal glucocorticoid treatment increases the risk of later cardiovascular dysfunction in the offspring. Underlying mechanisms include decreased circulating NO bioavailability, sympathetic hyper-reactivity, and NO-dependent endothelial dysfunction. Combined neonatal glucocorticoid and statin therapy protects the developing cardiovascular system by normalizing NO and sympathetic signaling, without affecting pulmonary maturational or anti-inflammatory effects of glucocorticoids.

Conclusions: Therefore, combined glucocorticoid and statin therapy may be safer than glucocorticoids alone for the treatment of preterm birth.

Keywords: cardiovascular diseases; glucocorticoids; hypertension; infant; newborn; premature birth.

Figure 1.

Experimental protocol and postnatal growth. A, On postnatal day 1 (P1) to P3 all male pups received either saline (10 µL g⁻¹ IP) or a 3-day, tapering course of dexamethasone (Dex; 10 µL g⁻¹ IP of solution equating to 0.5, 0.3, and 0.1 mg kg⁻¹ dissolved in saline IP). In addition, on postnatal days P1 to P6 all male pups received either saline or pravastatin (10 mg kg⁻¹, pravastatin sodium; Sigma-Aldrich, United Kingdom). B, Body weight and (C) fractional growth rate in pups from control (, C, n=12 litters), Dex-treated group (, D, n=12 litters), Dex and pravastatin–treated group Dex/pravastatin (, DP, n=12 litters) and pravastatin-treated group (, P, n=12 litters) treatment groups. The data represent one male rat per litter per outcome variable. For outcome variables, 1 male rat per litter was used for the in vivo studies, 1 for the isolated organ experiments, 1 was perfusion fixed and 1 was used to harvest tissues that were frozen for molecular studies. Therefore, each experimental group has biometry data from 4 pups per litter. Values are mean±SEM. Black bar signifies the 6-day treatment period. The P values for the main factors and interaction in the 2- or 3-way ANOVA are shown. Significant differences (P<0.05) for the post hoc Student-Newman-Keuls are *vs control. †D vs DP.

Figure 2.

In vivo cardiovascular function and NO bioavailability. Values are mean± SEM for basal mean arterial pressure (A), heart rate (B), heart rate variability in the time (RMMSD), and frequency (low frequency [LF]/ high frequency [HF] ratio) domains (C and D, respectively), baroreflex gain (E) and circulating plasma NO_x (NO species; nitrates and nitrites; F) in P21 pups from control (, C, n=8), dexamethasone (Dex)-treated group (, D, n=9), Dex and pravastatin–treated group (, DP, n=9) and pravastatin-treated group (, P, n=9) treatment groups. The P values for the main factors and interaction in the 2-way ANOVA are shown. Significant differences (P<0.05) for the post hoc Student-Newman-Keuls are *vs control. †D vs DP.

Figure 3.

Isolated cardiac function. Values are the mean±SEM for left ventricular developed pressure (LVDP; A), coronary flow rate (B), the LVDP following 15-minute ischemia (C), the maximum first derivatives of the left ventricular pressure with respect to time (dP/dt_max; D); the minimum first derivatives of the left ventricular pressure with respect to time (dP/dt_min; E); and the heart rate following 15-minute ischemia (F) in Langendorff preparations from control (, C, n=7), dexamethasone (Dex)-treated group (, D, n=7), Dex and pravastatin–treated group (, DP, n=7) and pravastatin-treated group (, P, n=7) treated animals. The P values for the main factors and interaction in the 2- or 3-way ANOVA are shown. Significant differences (P<0.05) for the post hoc Student-Newman-Keuls are *vs control. †D vs DP.

Figure 4.

Isolated peripheral vascular reactivity. Values are the mean±SEM for the dose-response constrictor curve to phenylephrine (A), the dose-response dilator curve to methacholine following preconstriction (B), and the nitric oxide (NO)-dependent and NO-independent vasodilator components to the methacholine-induced relaxation in femoral arteries isolated from control group (, C, n=8), dexamethasone (Dex)-treated group (, D, n=7), Dex and pravastatin–treated group (, DP, n=7) and pravastatin-treated group (, P, n=7) treated animals. The P values for the main factors and interaction in the 2- or 3-way ANOVA are shown. For A and B, significant differences (P<0.05) for the post hoc Student-Newman-Keuls are also detailed on the figures. For C, significant differences (P<0.05) for the post hoc Student-Newman-Keuls are *vs control. †D vs DP, #C vs P. AUC indicates area under the curve; and pD2(−logEC₅₀), negative logarithm of an EC₅₀.

Figure 5.

Lung structure and surfactant expression. Values are the mean±SEM for secondary crest formation (A), lung tissue to air space ratio (B), pulmonary tissue expression of SP-C (surfactant protein C; C), and pulmonary tissue expression of SP-D (surfactant protein D; D) in control group (, C, n=7), dexamethasone (Dex)-treated group (, D, n=7), Dex and pravastatin–treated group (, DP, n=7) and pravastatin-treated group (, P, n=7) animals. The P values for the main factors and interaction in the 2-way ANOVA are shown. Significant differences (P<0.05) for the post hoc Student-Newman-Keuls are *vs control. PonC indicates ponceau.