Former-preterm lambs have persistent alveolar simplification at 2 and 5 months corrected postnatal age

Abstract

Preterm birth and mechanical ventilation (MV) frequently lead to bronchopulmonary dysplasia, the histopathological hallmark of which is alveolar simplification. How developmental immaturity and ongoing injury, repair, and remodeling impact completion of alveolar formation later in life is not known, in part because of lack of suitable animal models. We report a new model using former preterm lambs to test the hypothesis that they will have persistent alveolar simplification later in life. Moderately preterm lambs (~85% gestation) were supported by MV for ~6 days before being transitioned from all respiratory support to become former preterm lambs. Results are compared to term control lambs that were not ventilated, and between males (M) and females (F). Alveolar simplification was quantified morphometrically and stereologically at 2 months (4M:4F) or 5 months (4M:6F) corrected postnatal age (cPNA) compared to unventilated, age-matched term control lambs (4M:4F per control group). These postnatal ages in sheep are equivalent to human postnatal ages of 1-2 years and ~6 years, respectively. Multivariable linear regression results showed that former preterm lambs at 2 or 5 months cPNA had significantly thicker distal airspace walls (p<0.001 and p<0.009, respectively), lower volume density of secondary septa (p<0.007 and p<0.001, respectively), and lower radial alveolar count (p<0.003 and p<0.020, respectively) compared to term control lambs. Sex-specific differences were not detected. We conclude that moderate preterm birth and MV for ~6 days impedes completion of alveolarization in former preterm lambs. This new model provides opportunity to identify underlying pathogenic mechanisms that may reveal treatment approaches.

Preterm Birth With Neonatal Interventions Accelerates Collagen Deposition in the Left Ventricle of Lambs Without Affecting Cardiomyocyte Development

“…Details of the perinatal procedure and outcomes of this model (including the data related to weight gain, systematic blood pressure, and blood gas values) have been previously reported. 35 Briefly, the ewes chosen for preterm delivery received intramuscular injections of dexamethasone phosphate (6 mg/kg; Vedco, Inc, St. Joseph, MO) 48 hours and again at 24 hours before caesarean delivery. At 128 days’ gestation, the ewes received an intramuscular injection of ketamine hydrochloride (10-20 mg/kg; KetaVed, Vedco, Inc, St. Joseph, MO), followed by inhalation anaesthesia with isoflurane (2.5% isoflurane; Piramal Healthcare, Bethlehem, PA).…”

Microarchitecture of the hearts in term and former‐preterm lambs using diffusion tensor imaging

“…Date‐mated Rambouillet × Columbia ewes were randomly assigned for cesarean‐section delivery of preterm lambs at ~128 days' gestation (female = 2, male = 3) or were spontaneously born at term (~150 days' gestation) (female = 5, male = 2). Details of the perinatal procedures have been reported by our group (Dahl et al, 2018). Briefly, the ewes chosen for preterm delivery received an intramuscular injection of dexamethasone phosphate (6 mg/kg; Vedco, Inc., St. Joseph, MO) 48 and 24 hr before operative delivery.…”

“…At 5 months after TEA, the lambs were weighed, then euthanized via administration of Beuthanasia (0.25 mL/Kg, Intervet Inc., Madison, NJ) followed by potassium chloride (10 mEq; Hospira, IL) intravenously. The full terminal tissue collection procedure is reported by our group (Dahl et al, 2018). The hearts were excised, arrested in diastole via coronary perfusion with potassium chloride (10 mEq; Hospira, IL), then immersion fixed in 10% buffered formalin at 4°C for 24 hr before scanning.…”

l‐Citrulline treatment alters the structure of the pulmonary circulation in hypoxic newborn pigs

“…PECAM1 (CD‐31) immunostaining was used for endothelial detection for quantitative histological analysis of capillary surface density. Lung slides from normoxic, n = 4, untreated chronically hypoxic, n = 4, and l ‐citrulline treated chronically hypoxic, n = 4, piglets were incubated with 1:1000 to 1:2000 dilutions of anti‐PECAM1 primary antibody (custom‐made rabbit, polyclonal, antisheep antibody, antigenic sequence RHYGFNEDVGNHAMKP; Absea, Beijing, China 12 ) diluted in 0.5% casein solution. Antigen detection was performed by incubation in 1:500 dilution of biotinylated antirabbit IgG (Cat# BA‐1100, Vector Laboratories), rinsing with 0.01 M phosphate‐buffered saline (PBS), followed by a 1:500 dilution of streptavidin‐horseradish peroxidase conjugate (Tyramide Signal Amplification [TSA] Kit, Cat# NEL700; PerkinElmer, Boston, MA).…”