PMC8298819_01
In vivo imaging of the patient with an esophageal bronchus. (A) Pre-repair/resection of (left) chest X-ray and (right) esophageal contrast study reveals opacification of the right upper lobe and the presence of an esophageal bronchus. (B,C) Selected (B) coronal and transverse (C) cross-sections from UTE MRI. Yellow arrowheads point to the fistula connecting from the esophagus to the right upper lobe of the lung. (D) 3D rendering of the chest from UTE MRI. The foregut (esophagus, trachea, and bronchi) is highlighted in green. The esophageal bronchus is seen with a superior branch proceeding to the right upper lobe. There is also a caudal branch inferior to the crossing of the right main stem pulmonary artery that is narrowed shortly after branching. The narrowing does not appear to be a result of compression by the right main stem pulmonary artery and is presumed to most likely be an intrinsic stricture.
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F1
Hospital on day of life 11 with concerns of poor feeding and choking during feeding. He was found to have right upper lobe consolidation on chest X-ray (Figure 1A) and was started empirically on ampicillin and cefotaxime with concerns for pneumonia, although urine, blood, and cerebrospinal fluid (CSF) cultures were all negative. Over the next 2 days, an esophageal contrast study (Figure 1A), 3D UTE chest MRI, and chest CT revealed a broncho-esophageal fistula arising from the distal esophagus to a segment of the right upper lobe (Figures 1B-D).
Histological and molecular analysis of the identity of the broncho-esophageal fistula and more distal airway. (A) H&E staining of the patient vs. control tissues from the "proximal" epithelium (broncho-esophageal fistula vs. trachea, respectively), bronchi, and distal bronchioles and alveoli. (B) Immunofluorescence staining of patient tissue for transcription factors SOX2 and NKX2-1 in sections from the proximal epithelium (broncho-esophageal fistula vs. trachea, respectively), bronchi, and distal bronchioles and alveoli. Scale bars represent 50 mum.
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F2
Histological analysis of the surgically resected esophageal bronchus demonstrated an increased stratification of the epithelium proximally (fistula and bronchi) when compared to control proximal airway epithelium samples which were obtained postmortem from infant/neonatal patients without esophageal, tracheal, or lung abnormalities (Figure 2A). The distal airway epithelium (bronchioles and alveoli) of both control and patient tissue appeared to be similar (Figure 2A). Immunofluorescence staining for the transcription factors SOX2 (normally expressed in both the esophageal and proximal respiratory epithelia) and NKX2-1 (normally expressed only in the respiratory epithelium) suggested that the identity of the fistula and airways distal to the fistula was primarily respiratory (Figure 2B).
Molecular analysis for differentiation markers of the broncho-esophageal fistula and more distal airways. (A) Immunofluorescence staining for cytokeratins KRT5 and KRT8, as well as acetylated tubulin in sections from the proximal epithelium (broncho-esophageal fistula vs. trachea, respectively), bronchi, and distal bronchioles and alveoli. (B) Immunofluorescence staining for cytokeratins KRT8 and KRT13 in sections from the proximal epithelium (broncho-esophageal fistula vs. trachea, respectively), bronchi, and distal bronchioles and alveoli. Scale bars represent 50 mum.
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F3
However, additional staining for the cytokeratin proteins revealed heterogeneity in the fistula and more proximal airways of the current infant, which was not seen in the control tissue (Figure 3). However, the proximal airway of the current infant lacked acetylated tubulin, typically a marker of cilia in the airway epithelium, while the distal airways had similar expression patterns between patient and control tissue samples (Figure 3A). Additional staining for keratin (KRT13), a cytokeratin expressed in esophageal but not respiratory epithelium, revealed foci of the epithelium with some esophageal identity (Figure 3B).