Interleukin-27-adipose-derived mesenchymal stromal cell-based gene therapy attenuates inflammation in lipopolysaccharide-induced acute respiratory distress syndrome
Acute respiratory distress syndrome (ARDS) is a lung inflammatory condition associated with the accumulation of fluid edema and cell infiltrates into the alveolar space along with the dysregulation in the immune response. Current therapeutics are limited to palliative care, i.e., mechanical ventilators, thus highlighting the need of developing targeted therapeutic for ARDS. Interleukin-27 (IL-27) is a multifunctional cytokine with the capability for immune modulation. Our interest lies in exploring the properties of IL-27, particularly as an anti-inflammatory cytokine that functions as an antagonist of IL-6 signaling, as an inducer of anti-viral genes, as a promoter of tissue repair, and as a regulator of both the innate and adaptive immune responses, possessing promising potential as a therapeutic for ARDS. To overcome the challenge of repeated administration due to the short half-life of cytokines, we utilized a cell-based gene therapy approach. IL-27-expressing plasmid was transfected into adipose mesenchymal stromal cells (ASC) that serve as the gene therapy carriers. For in vitro studies, we treated mono- and co-culture lung lipopolysaccharide (LPS)-induced lung epithelial and monocytes/macrophages cell line with IL-27-expressing ASC (IL-27) conditioned media (CM) to determine the effects on pro-inflammatory gene expression. For in vivo studies, male C57BL/6 mice was intratracheally injected with LPS (5mg/kg) and treated either PBS, ASC, or IL-27 ASC (5 x 105 cells/mouse) after 24h of LPS instillation. Measurements of gene expression, histopathological changes, cell infiltrations, and protein content in the bronchoalveolar lavage fluid and RNA-seq analysis were performed for the in vivo studies.  Results indicated that IL-27ASC CM were able to reduce pro-inflammatory gene expression of lung epithelial and macrophages cultured in both mono- and co-culture systems. Additionally, IL-27 ASC were able to reduce pro-inflammatory markers, decrease cell infiltration into the lungs, promote genes and immune cells involved in tissue repair, and rebalance innate and adaptive immunity in an LPS-induced in vivo model. Collectively, our in vitro and in vivo results showed promising potential for IL-27 cell-based gene therapy as a treatment for ARDS. Overall design: In this study, male C57BL/6 mice (8-10 weeks old) were used to create an in vivo LPS-induced ARDS model. Anesthesia was administered using ketamine and xylazine, and LPS (E. coli O111:B4) was given intratracheally at 5 µg/g body weight. Mice were randomly assigned to treatment groups, including 1X PBS (control), LPS, LPS + hASC with empty vector, and LPS + hASC with IL-27 vector. At 24 hours post-LPS instillation, treatments were given, followed by air infusion to ensure distribution. Animals were sacrificed at 72 hours post-LPS instillation, and samples (BALF, serum, lung, and organ tissues) were collected for analysis.