Prenatal lipopolysaccharide exposure programs the early-life lung development by modulating mesenchymal fibroblasts in a time- and dose-dependent manner.

backgroundBronchopulmonary dysplasia (BPD) is one common and severe complication of preterm births. Prenatal infection/inflammation is the major cause driving preterm deliveries, but its contribution to the early-life lung development remains unclear.

methodsPregnant C57BL/6J mice were randomized at 15.5 or 18.5 days post coitum (dpc) to receive intraperitoneal injection of sterile saline, lipopolysaccharide (LPS) 50 μg/kg, or LPS 100 μg/kg. Animal experiments were performed compliantly to the ARRIVE guidelines. Lungs of newborn pups of mixed genders were harvested at the first postnatal day (P1) for histopathology to investigate prenatal LPS-induced structural changes. Expression of key pro- and anti-inflammatory mediators was assessed at both transcriptional and translational levels. Based on bulk RNA sequencing and exploratory transcriptomic analysis, signature gene and phenotypical expression of two major mesenchymal fibroblasts subsets, myofibroblasts (MYFs) and lipofibroblasts (LIFs), were analyzed with qPCR, immunofluorescence, and western blot. Human embryo-derived WI-38 fibroblast cell line was used as an in vitro model to investigate LPS-induced fibroblast inflammatory responses and MYFs-LIFs differentiation.

resultsNewborn mice exposed to prenatal LPS exhibited enlarged air spaces and thinner septal walls, with an enhanced interleukin 6 (IL6) response in lung tissues. The degree of lung structural changes is LPS dose- and timing-dependent. Exploratory transcriptomic analysis revealed the enrichment in myofibrogenic pathways in LPS-exposed lungs. Platelet-derived growth factor receptor alpha (PDGFRA)-expressing progenitor lung fibroblasts were generally suppressed by prenatal LPS exposure. The expression of MYFs signature markers (ELN and ACTA2) in relation to LIFs (PLIN2 and FGF10) demonstrated a highly heterogeneous and dynamic pattern, depending on LPS doses and timing. Upon LPS exposure, human WI-38 fibroblasts upregulated a panel of pro-inflammatory mediators via nuclear factor kappa B signaling and displayed diverse MYFs and LIFs differentiation depending on the dose and duration of LPS stimulation.

conclusionsPrenatal LPS exposure induces heterogeneous structural and molecular changes in the newborn mice lung, showing LPS dose- and time-dependent modulation of mesenchymal fibroblast differentiation. These findings may contribute to refine the risk stratification of preterm infants exposed to prenatal infection/inflammation. Mesenchymal fibroblast plasticity and MYFs-LIFs phenotypic shifts may inspire preventive and therapeutic strategies for BPD.