Visual screening and efficacy evaluation of high-performance fluorescent probe DAF-FM in esophagitis cancer transformation.

BACKGROUND: Chronic esophagitis can progress to esophageal cancer via "inflammation-dysplasia-cancer" transformation, with nitric oxide (NO) serving as a critical mediator in this process. Traditional diagnostic methods (e.g., endoscopic biopsy) for esophageal cancer transformation have low sensitivity and require long detection time, while existing fluorescent probes lack specificity and stability for real-time NO monitoring. High-performance fluorescent probes like DAF-FM, with NO-targeting ability, show potential for visual screening and efficacy evaluation but need systematic validation in esophageal cancer models. AIM: To validate the applicability of the fluorescent probe DAF-FM for visual screening of esophageal cancer transformation, explore the underlying mechanism of NO-regulated transformation, and evaluate the probe's efficacy in monitoring therapeutic responses. METHODS: Laser confocal imaging and flow cytometry were used to analyze DAF-FM's NO concentration/time-dependent fluorescence response, lysosomal targeting (via Pearson coefficient), and cytotoxicity (with cholecystokinin-8 assay) in esophageal cells. Sprague-Dawley rat esophageal cancer models (normal, esophagitis, esophageal cancer, and drug/radiotherapy intervention) were established to monitor NO dynamics and tumor volume correlation. Clinical diagnostic comparison (50 suspected patients) with endoscopic biopsy/histopathology was conducted using Kolmogorov-Smirnov test and Student's t-test (P < 0.05). Western blot and quantitative real-time polymerase chain reaction were used to explore NO's role in the nuclear factor-kappa B (NF-κB) pathway. RESULTS: DAF-FM exhibited concentration/time-dependent fluorescence with NO (300 μM NO: 60-minute fluorescence intensity 458 ± 15 arbitrary units, P < 0.05) and specific lysosomal targeting (Pearson's coefficient = 0.82 ± 0.03). It had low cytotoxicity (82.3% ± 4.1% cell viability at 50 μM). In rat models, DAF-FM showed that NO was correlated with tumor volume (R² = 0.87). Clinically, its sensitivity (92.5%) outperformed endoscopic biopsy (78.3%), with shorter detection time (30 minutes vs 48 hours, P < 0.05). Mechanistically, NO regulated transformation via the NF-κB pathway (Pearson's coefficient = 0.78 ± 0.05 between DAF-FM and NF-κB). CONCLUSION: DAF-FM is a feasible tool for visual screening of esophageal cancer transformation, enabling real-time NO monitoring, high-sensitivity diagnosis, and therapeutic efficacy evaluation. It provides a new approach for esophageal cancer diagnosis and mechanism research.