Confocal laser scanning fluorescence microscopy for in vivo determination of microvessel density in Barrett's esophagus

Background and study aims: Angiogenesis has been reported to be an essential step in the progression of cancers arising from Barrett's esophagus. Confocal laser scanning microscopy (CLM) has the potential to perform in vivo microscopy to detect angiogenesis and determine microvessel density (MVD). We aimed therefore to use this new promising imaging tool for the evaluation of MVD in Barrett's esophagus and associated neoplasia. Patients and methods: We enrolled 20 patients with Barrett's esophagus. CLM sequences were recorded from pre-marked areas using argon beamer coagulation spots after intravenous application of fluorescein. Sequences had to be recorded within the first 8 minutes of injection. Biopsies were taken from the same areas for standard histopathology. All CLM sequences were put into a random order and analyzed by a single investigator who was blinded to any clinical or histopathological data. Five still images per sequence were analyzed for MVD using a specially designed software algorithm. The primary endpoint was determination of vessel diameter and MVD in relation to neoplastic or non-neoplastic Barrett's esophagus. Results: We evaluated 750 still CLM images from 150 sequences/biopsy sites. Histopathology revealed 69 biopsies as non-neoplastic Barrett's esophagus (46.0%), 11 as neoplastic Barrett's esophagus (7.3%), 64 as cardiac mucosa (42.7%), and six as squamous mucosa (4.0%). Mean vessel diameter as determined by CLM was similar in all four groups (P = 0.2). However, MVD was significantly higher in CLM sequences of neoplastic Barrett's esophagus compared with benign conditions (neoplastic Barrett's esophagus 23.6%: Barrett's esophagus 14.2%; cardiac mucosa 15.8%; squamous epithelium 20.6 %; neoplastic Barrett's esophagus vs. Barrett's esophagus P < 0.001, t-test). Conclusion: Fibered fluorescein-guided CLM helps to detect angiogenesis in malignant and non-malignant Barrett's esophagus in vivo. These data might help to improve the diagnostic yield of detecting Barrett's neoplasia but also to facilitate monitoring of antiangiogenetic therapy.