Single-Cell Pathological Landscape Analysis of Vascular Calcification in CKD by Imaging Mass Cytometry

Background: Vascular calcification (VC) is regarded as an important pathological change which associated with high mortality in patient with chronic kidney disease (CKD). It is necessary to study changes of cell communication and their PTMs in situ. Methods: A highly multiplexed imaging mass cytometry (IMC) panel was designed to simultaneously quantify 34 biomarkers of tissues from 38 patients with CKD and 2 donors to generate 64 highly multiplexed images at single-cell resolution. All cells were divided into different cell types according to the expression of markers. Neighboring cells were defined as the top 15th nearest cells to a cell. K-means clustering algorithm was then applied to cluster cell neighborhoods into numerous community clusters (CCs). Relevant to the stage of vascular calcification by Von Kossa staining was identified as a specific target for preventing vascular calcification. Results: 8 cell types and 16 community clusters were defined. We found that contractile VSMCs and progenitor cells decrease with progression of vascular calcification. The osteochondrogenic VSMCs showed a trend of decreasing first and then increasing, which may be caused by the proliferation of synthetic VSMCs in the early stage of calcification. At the same time, community clusters changed with progression of vascular calcification. Contractile VSMCs and osteochondrogenic VSMCs mixed community clusters decreased with the progression of vascular calcification. On the contrary, osteochondrogenic VSMCs dominant community clusters osteochondrogenic VSMCs elevated in severely calcified vessels. In addition, expression of H3K27me3 of community cluster 10 that mainly composed of osteochondrogenic VSMCs and fibroblasts decreased with the progression of vascular calcification. Conclusions: Our findings reveal for the first time the various topological function units of VC in CKD, which also presents the single-cell pathological landscape for VC in CKD. This work highlights the potential of a community cluster with a mixture of osteogenic VSMC and fibroblasts in driving vascular calcification, which may cause by the decreasing of H3K27me3. Funding: Government Support - Non-U.S.