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Abstract: PO0689

Insulin Receptor Signaling Is Necessary for NFκB-Activated Host Defense Responses in Murine Intercalated Cells

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Schwartz, Laura, Nationwide Children's Hospital, Columbus, Ohio, United States
  • Eichler, Tad, Nationwide Children's Hospital, Columbus, Ohio, United States
  • Yan, Pearlly, The Ohio State University, Columbus, Ohio, United States
  • Spencer, John David, Nationwide Children's Hospital, Columbus, Ohio, United States
Background

Urinary tract infection (UTI) disproportionately affects select groups, especially those with insulin resistance and diabetes mellitus. Kidney intercalated cells (IC) play a key role in preventing UTI by regulating urine pH and secreting cytokines and antimicrobial peptides (AMP). Our data show that insulin receptor (IR) deletion in murine IC causes insulin resistance and increases UTI risk in vivo while having no impact on glucose homeostasis or urine acidification. Here, we profile the transcriptomes of IC isolated from IR knockout (IRKO) mice and controls (WT) to identify IR-mediated host defenses.

Methods

Insr gene was deleted in murine IC by breeding Atp6v1b1-Cre transgenic mice with IR-floxed mice. A tdT reporter was added to aid fluorescence-assisted cell sorting (FACS) of IC. RNAseq was performed on IC and read count data were analyzed for differentially expressed genes (DEG) using edgeR. DEG were defined with FDR adjusted p-value < 0.05. Canonical pathway analysis of DEG was performed using Ingenuity Pathway Analysis. Sorted IC were cultured and challenged in vitro with uropathogenic E.coli (UPEC) to assess response and susceptibility to infection. To define the contributions of NFκB to UTI defense, NFKB1 was silenced in human medullary cells using siRNA. UPEC attachment and invasion assays were performed.

Results

FACS-enriched IC express IC-specific genes like Aqp6 and Atp6v0d2. Differential expression analysis reveals suppression of 138 genes and upregulation of 232 genes in IRKO vs WT IC. In IRKO IC, a decrease in Insr as well as downstream IR-regulated targets and host defense genes such as AMPs were observed. While diverse pathways implicated in innate immunity are suppressed in IRKO IC, many converge on one target: NFκB. When cultured IC from these mice are challenged with UPEC, IRKO IC exhibit suppressed NFκB activation and UPEC were more likely to invade them. Silencing NFKB1 results in decreased AMP expression and increased UPEC attachment and invasion of human medullary cells.

Conclusion

These data suggest IR signaling impacts the IC host defense transcriptome and identifies IR-sensitive pathways that aid in UPEC defense by activating NFκB signaling and expressing AMPs. A greater understanding of the factors that predispose diabetics to UTI may reveal novel, targeted therapies to prevent/treat diabetes-associated UTI.

Funding

  • NIDDK Support