Abstract Rationale Pericytes play a key role in the development and maturation of the lung vasculature. In the developing lung, angiogenesis is intricately linked to alveolarization. Pericytes are an extremely heterogenous population which differ in phenotype and gene expression signature. We hypothesize that developmental stage specific pulmonary pericytes have distinct functions involved in angiogenesis-driven alveolarization. Methods Pulmonary pericytes from postnatal days 3, 7, and 28 (P3, P7, and P28) were isolated from C57BL/6J wild-type mice. Cell identity was confirmed by expression of pericyte specific markers. Pericytes were interrogated for differences in protein expression, cell morphology, cell contraction, cell migration, cell proliferation, and the capacity to remodel the extracellular matrix (ECM) and promote tube formation. Results We found that pulmonary pericytes differ relative to cell morphology, ECM remodeling, contraction, proliferation, migration, tube formation, and protein expression across postnatal lung development. In brief, P3 pericytes (late saccular stage) express significantly more α-SMA and SM-MHC11, and are more contractile than at later time points as assessed by collagen gel contraction assays (mean 72.9% vs. 96.3% surface area, p 0.001), pointing to a role for contractile pericytes during the late saccular stage of lung development, a period of enhanced ECM remodeling and reorganization. P7 pericytes (early alveolar stage) express significantly more Profilin1 and nuclear Profilin1, a protein essential for cell motility. Cell migration assays demonstrated that P7 pericytes are more migratory than at other time points (mean 40.2 vs. 16.8 cells migrated, p 0.0001). In addition, cell proliferation assays demonstrated that at P7, pericytes are more proliferative (mean 164 P7 vs. 79.3 P3 cells, p 0.0001). Treatment with Delta-Like 4 (DLL4), a Notch3 ligand, enhances P7 pericyte proliferation. Together, these data suggest that early alveolar pericytes may play a role in the rapid expansion of the microvasculature during a period of heightened angiogenesis within the lung. Tube formation assays demonstrated that P28 pericytes (late alveolar stage) are capable of driving microvasculature maturity. We found that DLL4 treatment of P28 pericytes increases proliferation, albeit less than in P7 pericytes (1.80-fold vs. 2.27-fold). P28 pericytes are capable of ECM remodeling, yet to a lesser degree than P3 pericytes. Conclusion Pulmonary pericyte phenotype changes across development. Pulmonary pericyte contractility peaks during the late saccular stage, migration and proliferation are greatest during early alveolarization, and stabilization of the pulmonary circulation is highest during late alveolarization. The results from this study suggest that developmental differences in pericyte phenotype and function facilitate lung morphogenesis. This abstract is funded by: None
Barnes et al. (Fri,) studied this question.