Characterizing the immune and inflammatory microenvironments of Hunner lesions in interstitial cystitis/bladder pain syndrome using imaging mass cytometry: A new look at an old disease
Tiziana Cotechini1, Nathalia Kim1, Charles C.T. Hindmarch2,3,4, Michael Morra5, Sindhura Thirumal6, Parvin Mousavi6, David Berman7, J. Curtis Nickel8, Kerri-Lynn Kelly8, Amber Simpson1, D. Robert Siemens8, Charles Graham1, R. Christopher Doiron8.
1Biomedical & Molecular Sciences, Queen's University , Kingston, ON, Canada; 2School of Medicine, Queen's University , Kingston, ON, Canada; 3Translational Institute of Medicine, Department of Medicine, Queen's University , Kingston, ON, Canada; 4Queen's Cardiopulmonary Unity, Queen's University , Kingston, ON, Canada; 5School of Medicine, Queen's University , Kingston, ON, Canada; 6School of Computing, Queen's University , Kingston, ON, Canada; 7Department of Pathology, Queen's University , Kingston, ON, Canada; 8Department of Urology, Queen's University , Kingston, ON, Canada
CUA Astellas Research Grant.
Introduction: What exactly are Hunner lesions (HL) identified in patients with interstitial cystitis (IC)/bladder pain syndrome (BPS)? We provide a deep-dive of HLs with an in-depth characterization of the cellular immune microenvironment using state-of-the-art imaging mass cytometry (IMC), Hyperion™ Imaging System in a cohort of HL-IC/BPS patients.
Methods: Bladder biopsies from patients with HL-IC/BPS were used to create formalin-fixed, paraffin-embedded tissue sections (5 mm). These were stained with 23 metal-conjugated antibodies and processed using the Hyperion™ IMC System — a technology that enables the detection of up to 37 markers on a single formalin-fixed, paraffin-embedded tissue slide. Multiplexed images were visualized and data were processed and analyzed using various computational methods to identify individual cell populations.
Results: A panel of 23 markers (Figure 1) was designed to evaluate cellular microenvironment of HL biopsies. Analysis of HL tissue samples from our discovery cohort of 10 patients, with a median age at time of tissue biopsy of 63 years and median duration of disease of 7.5 years, provided a total of 113 500 single cells. On average, 16% of all cells within HLs were identified as CD45+ immune cells. Within the immune cell compartment, clustering analysis identified various populations including B cells, CD8+ cytotoxic T cells, FoxP3+ T regulatory cells, CD4+ T helper cells, monocytes, classical macrophages, CD163+ macrophages, and dendritic cells. Tertiary lymphoid structures were observed in three of 10 patient samples. Further analyses include stratification based on proliferation (Ki67+), activation (Granzyme B+), and/or polarization states.
Conclusions: We report the first quantification and spatial resolution of single cells in tissue samples collected from HL-IC/BPS patients using IMC. The immune complexity of HLs uncovered with this most in-depth look at the heterogeneous phenotypes of immune cells and their in situ organization lays the groundwork for further understanding of these enigmatic lesions.
