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Kidney Week

Abstract: FR-PO1210

Single Cell RNA-Seq profiling of renal endothelial cells in experimental diabetic nephropathy model reveals transcriptomic changes in separate endothelial subpopulations

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Zhou, Alex Xianghua, AstraZeneca Gothenburg, Mölndal, Sweden
  • Laerkegaard Hansen, Pernille B., Imed CVMD, Mölndal, Sweden
  • Betsholtz, Christer, Uppsala University, Uppsala, Sweden
  • Jeansson, Marie, Karolinska Institutet, Huddinge, Sweden
  • He, Liqun, Uppsala University, Uppsala, Sweden
  • Uhrbom, Martin, AstraZeneca, Mölndal, Sweden
  • Liu, Jianping, Karolinska Institutet, Huddinge, Sweden
  • Granqvist, Anna, AstraZeneca, Mölndal, Sweden
  • Tonelius, Pernilla, AstraZeneca R&D Gothenburg, Sweden, Mölndal, Sweden
Background

Endothelial dysfunction and vascular rarefaction are hallmarks of chronic kidney disease, while little is known about the transcriptomic changes of renal endothelial cells (ECs) during disease progression. Moreover, kidney contains heterogenous EC subpopulations that are structurally and functionally distinguishable. This study aims to determine and compare transcriptome profiles in separate renal EC subpopulations between healthy mice and mice of diabetic nephropathy (DN) using single cell RNA-seq (scRNA-Seq).

Methods

Kidneys of BTBR lean and ob/ob mice at 6, 11 and 20 weeks of age were enzymatically dissociated with LiberaseTM. After incubation with Pecam1 antibody and Calcein-AM, Pecam1+ and Calcein-AM+ single live cells were FACS sorted into 384-well plates. Single EC cDNA library was generated by Smart-seq2 technique and the sequencing was performed on Illumina HiSeq 3000. Unsupervised clustering of EC subpopulations was performed with Pagoda2 analysis.

Results

BTBR ob/ob mice develop vascular rarefaction with age. Compared to the lean mice, the proportion of renal single live ECs in the ob/ob mice showed no difference at 6 weeks of age, a 25% reduction at 11 weeks of age, and a 32% reduction at 20 weeks of age (P=0.01). The current Pagoda2 analysis on the 11-week-old lean and ob/ob mice revealed twelve EC subpopulations and 142 genes with significantly altered expression in ob/ob mice. Among differentially expressed genes (DEGs), certain redox genes were ubiquitously regulated. However, majority of the DEGs were altered in distinct EC subpopulations, likely owing to either the different sample sizes or the heterogeneity on gene expression/regulation in various EC subpopulations. The data of 6- and 20-week-old BTBR mice is under analysis to explore the time course of the transcriptomic changes.

Conclusion

The full length scRNA-seq on FACS sorted renal ECs provides a feasible approach to a high-resolution transcriptomic profiling of heterogenous EC populations in kidney and importantly the heterogenous transcriptomic changes in an experimental DN model.

Funding

  • Commercial Support