Kidney Week

Abstract: TH-PO561

Analysis of Kidney Biopsies from Patients with Glomerulonephritis Using Imaging Mass Cytometry Reveals Increase in Immune Cells with Associated Dedifferentiation and Injury of Tubular Cells

Session Information

• Glomerular Diseases: Omics, Biomarkers, and Tools
  October 24, 2024 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Glomerular Diseases

• 1401 Glomerular Diseases: Mechanisms, including Podocyte Biology

Authors

• Weiss, Marlene, Charite Universitatsmedizin Berlin, Berlin, Berlin, Germany

Marlene Weiss, DrMed

• Kakade, Vijayakumar R., Yale University School of Medicine, New Haven, Connecticut, United States

Vijayakumar R. Kakade, PhD

• Baker, Megan Leila, Yale University School of Medicine, New Haven, Connecticut, United States

Megan Leila Baker, MD

• Budiman, Tifanny, Yale University School of Medicine, New Haven, Connecticut, United States

Tifanny Budiman

• Shelar, Ashish, Yale University, New Haven, Connecticut, United States

Ashish Shelar, PhD

• Cantley, Lloyd G., Yale University School of Medicine, New Haven, Connecticut, United States

Lloyd G. Cantley, MD, FASN

Background

This study was conducted to characterize and develop mechanistic understanding of the tubulointerstitial impact of acute glomerulonephritis. For both ANCA-associated vasculitis (AAV) and Lupus nephritis (LN), there is a lack of patient-targeted therapies.

Methods

30 FFPE cortex samples from 15 patients (5 with AAV, 5 with proliferating LN, and 5 tumor-remote nephrectomy samples (reference) were analyzed in a Tissue Microarray with imaging mass cytometry (IMC) after staining it with 37 metal-tagged antibody markers. Analysis was performed using scripts in Python and RStudio. Cells were segmented with mesmer, a deep learning platform. Clustering was performed using the Phenograph algorithm.

Results

Analysis of all 30 samples resulted in identification and segmentation of 106,455 cells (37.2% from reference, 31.2% from AAV, 31.6% from LN) with total analyzed tissue areas of 9.7, 5.1 and 5.8 mm², respectively. Thirty-nine cell clusters were identified with annotation revealing 17 different cell types. 95.6% of cells were successfully annotated. Total cell counts per mm² tissue area showed that reference kidneys had 3.1±1.8% immune cells whereas AAV and LN kidneys contained 28±17% and 21.9±20% immune cells, respectively, including T cells, B cells and macrophages. Analysis of the resident cell types revealed the expected predominance of proximal tubule (PT) cells (38.4±5.2% in reference, 32.7±8.2% in AAV, 30.9±7.5% in LN). Injury markers analyzed included kidney-injury molecule 1 (KIM-1), vimentin, FACL4 (ferroptosis) and pRIPK3 (necroptosis). Initial clustering resulted in 9 different PT clusters, of which 2 were notable for high expression of injury markers. Cells in cluster iPT were increased in both AAV and LN and had high expression of vimentin, pRIPK3 and KIM-1, while cells in cluster aavPT were found only in AAV and had high expression of KIM-1 and FACL4.

Conclusion

AAV and LN kidney samples reveal a marked increase in interstitial immune cells with associated dedifferentiation and injury of resident tubular cells. The tubular injury is highlighted by increased KIM-1, ferroptosis and necroptosis marker expression in injured PT with concurrent downregulation of resident markers Megalin and Aquaporin 1.