C70-38 Cftr Mutation Linked Immunometabolic Alterations in Monocytes

Rationale

Mutations in the CFTR gene cause cystic fibrosis (CF), and the prevalent ΔF508 variant results in posttranslational destabilization, misfolding, and degradation of the CFTR protein, impairing airway host defense, mucociliary clearance, and microbicidal activity. Although chronic bacterial infection and purulent airway obstruction in CF stem largely from dysfunctional innate immunity, the lack of robust cellular models for CF monocytes has hindered investigation of their role in CF airway pathology. Recent studies indicate that CFTR modulators enhance CFTR expression in monocytes, revealing metabolically regulated activities that influence inflammatory responses. Because ATP is the central cellular energy currency, ATP production rates—derived from mitochondrial oxidative phosphorylation (OXPHOS) and glycolysis—offer dynamic, quantitative insight into cellular metabolic state and critically govern immune cell activation, proliferation, and effector function.

Methods

Promyelocytic HL-60 cells, CFTRmutant HL-60-ΔF508 cells, and leukemic monocytelike THP-1 cells were cultured in RPMI640 supplemented with 2mM glutamine, 2 mM LalanylLglutamine, and 10% FBS. Cells were differentiated into monocytes using LPS (2µg/mL) and 1α,25dihydroxyvitamin-D3 (150µM), and differentiation was validated by flow cytometry using monocyte surface markers. Bioenergetic activity was assessed using the Agilent Seahorse XFp RealTime ATP Rate Assay, which simultaneously measures oxygen consumption rate (OCR) and extracellular acidification rate (ECAR).

Results

Monocyte (CD14⁺) differentiation of THP-1, HL60, and HL60-ΔF508 cells produced distinct bioenergetic outcomes compared with controls. THP-1 cells showed moderate ATP shifts (Glyco-ATP +18% and mitoATP −26%) with LPS and balanced increases in glycolytic (+113%) and mitochondrial ATP (+60%) with Vitamin D3, reflecting intact metabolic flexibility. HL60 cells displayed strong activation, with LPS driving a robust rise in glycolytic ATP (+257%) and Vitamin D3 inducing a mitochondrialdominant increase (+142% mitoATP). In contrast, HL-60-ΔF508 cells showed baseline glycolytic suppression with elevated mitochondrial dependence and failed to elevate total ATP under either treatment (+30% glycolATP and −32% mitoATP for LPS; −62% glycolATP and −65% mito-ATP for VitD3), resulting in overall ATP decreases of − 16% and −57%, respectively. HL-60-ΔF508 produced only ∼27-30% of the ATP generated by HL-60 under identical conditions, indicating that the CFTR mutation disrupts normal metabolic reprogramming and impairs ATP biogenesis.

Conclusion

The reduced ATP production rate observed in differentiated HL-60-ΔF508 monocytes suggests impaired metabolic function associated with CFTR dysfunction. Further evaluation of metabolic parameters—including proliferation, activation, and mitochondrial susceptibility—will deepen understanding of monocytedriven phagocytic defects in CF onset and progression and may guide development of therapeutic strategies that complement existing CFTR modulator therapies.

This abstract is funded by: The Rosenau Family Research Foundation, UW-SMPH