The Downregulation of LKB1 in Small Extracellular Vesicles Elucidates the Downstream Mechanisms Driving Acute Lung Allograft Rejection.

BACKGROUND

This study explored the role of a tumor suppressor gene, liver kinase B1 (LKB1), a key metabolic regulator in inflammatory and cellular processes associated with lung transplantation. Acute cellular rejection (ACR) after lung transplantation poses a significant threat, escalating the risk for chronic lung allograft dysfunction. We identified LKB1 as a pivotal regulator in lung transplant pathology, particularly in ACR.

METHODS

Using NanoSight, ExoView, and Western blot analysis, we characterized plasma-derived small extracellular vesicles (sEVs). Griess reactions were used to measure plasma nitrite/nitrate levels. To determine the role of LKB1 on inflammation, we genetically altered LKB1 in human alveolar basal epithelial cell line and human bronchial epithelial cell line (BEAS-2B) cells, and in vivo analysis, we treated C57BL/6 mice with lipopolysaccharide (LPS).

RESULTS

LKB1 levels in sEVs were significantly downregulated in lung transplant recipients, correlating with increased inducible nitric oxide synthase (iNOS). Furthermore, sEVs-LKB1 levels recovered only in 50% of lung transplant recipients after anti-rejection therapy; LKB1 downregulation in ACR was linked to elevated nitrite/nitrate. LKB1 overexpression inhibited LPS-induced iNOS, whereas LKB1 knockdown amplified it. Furthermore, LKB1 deficiency exacerbated transforming growth factor-beta-induced iNOS and tumor necrosis factor-alpha, confirming its protective role against pro-fibrotic signaling. Finally, we demonstrated wild-type LKB1 containing sEVs (W-LKB1-sEVs) activate AMP-activated protein kinase in BEAS-2B cells, and reduced LPS-induced iNOS and tumor necrosis factor-alpha in BEAS-2B cells. This protective effect is largely blocked by the AMP-activated protein kinase inhibitor Compound C.

CONCLUSIONS

The study demonstrates that LKB1 is a key suppressor of inflammation, and its downregulation in sEVs during ACR can serve not only as a biomarker but also offers new therapeutic targets.