Flow based single cell analysis of the immune landscape distinguishes Barrett’s esophagus from adjacent normal tissue

// Moen Sen 1 , * , Friedrich Hahn 2 , * , Taylor A. Black 1 , * , Maureen DeMarshall 3 , Warren Porter 2 , Eileen Snowden 2 , Stephanie S. Yee 1 , Frances Tong 2 , Mitchell Ferguson 2 , Emylee N. Fleshman 1 , Hiroshi Nakagawa 3 , Gary W. Falk 3 , Gregory G. Ginsberg 3 , Michael L. Kochman 3 , Rainer Blaesius 2 , Anil K. Rustgi 3 and Erica L. Carpenter 1 1 Division of Hematology and Oncology, Department of Medicine, Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA 2 Department of Genomic Sciences, BD Technologies and Innovation, Research Triangle Park, Durham, North Carolina, USA 3 Division of Gastroenterology, Department of Medicine, Abramson Cancer Center, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA * These co-first authors have contributed equally to this work Correspondence to: Erica L. Carpenter, email: erical@upenn.edu Anil K. Rustgi, email: anil2@pennmedicine.upenn.edu Keywords: Barrett’s esophagus; microenvironment; immune cells; single cell; flow cytometry Received: January 17, 2019     Accepted: April 14, 2019     Published: June 04, 2019 ABSTRACT Barrett’s esophagus (BE) is metaplasia of the squamous epithelium to a specialized columnar epithelium. BE progresses through low- and high-grade dysplasia before developing into esophageal adenocarcinoma. The BE microenvironment is not well defined. We compare 12 human clinical BE and adjacent normal squamous epithelium biopsies using single cell immunophenotyping by flow cytometry. A cassette of 19 epithelial and immune cell markers was used to detect differences between cellular compartments in normal and BE tissues. We found that the BE microenvironment has an immunological landscape distinct from adjacent normal epithelium. BE has an increased percentage of epithelial cells with a concomitant decrease in the percentage of immune cells, accompanied by a shift in the immune landscape from a predominantly T cell rich microenvironment in normal tissue to a B cell rich landscape in BE tissue. Hierarchical clustering separates BE and normal samples into two discrete groups based upon our 19-marker panel, but also reveals unexpected, shared phenotypes for three patients. Our results suggest that flow based single cell analysis may have the potential for revealing clinically relevant differences between BE and normal adjacent tissue, and that surface immunophenotypes could identify specific subpopulations from dysplastic tissue for further investigation.