In the last years, macrophages (MΦ) have shown great potential for various cell-based therapies. We previously demonstrated beneficial effects of pulmonary MΦ transfer (PMT) in various lung diseases, including lung infection. Single-cell analysis of transferred bone marrow (BM)-derived MΦ post-PMT showed a stepwise adaptation process to an alveolar MΦ (AM)-like phenotype and transcriptome, underlined by a population expressing Ki67, indicating local proliferation.
However, to better understand this adaption process and promote MΦ adaption post PMT, we performed CRISPR/Cas9-mediated knockout (KO) of various candidate genes, including kinase p38 that regulates self-renewal in hematopoietic stem cells (HSC) but also differentiation and MΦ function. Performing a competitive BM transplantation, we observed superior engraftment of p38KO HSC compared to wildtype (WT) HSC in the BM, while AM in the lung were mainly WT-derived, suggesting a role of p38 in AM biology. Since p38KO did not affect myelopoiesis, we adoptively transferred a 1:1 mixture of WT and p38KO BM-MΦ into lungs of Csf2rb-deficient mice that lack endogenous AM. Donor cells adapted an AM-like phenotype and improved disease parameters in Csf2rb-deficient mice, such as improved turbidity, lower protein and cholesterol concentrations in the broncho-alveolar lavage, thus demonstrating proper AM function. However, in contrast to the competitive BM transplantation experiments the ratio of WT:p38KO cells shifted in favour of p38KO cells to 66,4%±4,8% 4 weeks post-PMT and 74,1%±9,7% 7-8 weeks post PMT. These results suggest a beneficial effect of p38KO in the overall fitness of AM under stress conditions, which may be abolished in steady state.