Kidney Week

Abstract: FR-PO1075

Lcn2 Plays a Key Role in Dot1l-HDAC2-ET1-Mediated Kidney Fibrosis

Session Information

• CKD Mechanisms: Progression, Fibrosis, and Beyond
  November 03, 2023 | Location: Exhibit Hall, Pennsylvania Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: CKD (Non-Dialysis)

• 2303 CKD (Non-Dialysis): Mechanisms

Authors

• Tsilosani, Akaki, Albany Medical Center, Albany, New York, United States

Akaki Tsilosani,

• Gao, Chao, Albany Medical Center, Albany, New York, United States

Chao Gao,

• Guo, Chenchun, Albany Medical Center, Albany, New York, United States

Chenchun Guo,

• Chen, Enuo, Albany Medical Center, Albany, New York, United States

Enuo Chen,

• Zhang, Wenzheng, Albany Medical Center, Albany, New York, United States

Wenzheng Zhang,

Background

Kidney fibrosis is a hallmark of chronic kidney disease. The genetic and epigenetic factors regulating kidney fibrosis are largely unknown. Kidney injury marker Lipocalin 2 (Lcn2) is involved in kidney fibrosis. We reported that Aqp2Cre mice express Cre specifically in Aqp2⁺ progenitor cells, which generates at least 5 types of cells forming distal convoluted tubule 2 (DCT2), connecting tubule (CNT) and collecting duct (CD). Our recent studies also indicated that disruption of histone H3 K79 methyltransferase Dot1l in Aqp2⁺ progenitor cells led to upregulation of endothelin 1 (ET1) and kidney fibrosis through histone deacetylase 2 (HDAC2) during normal aging. Here, we hypothesize that Lcn2 is a new component of this process and upregulates HDAC2, which in turn increases ET1 and facilitates kidney fibrosis.

Methods

Using the Aqp2Cre driver, we developed three conditional knockout mice: 1) DCT2/CNT/CD-specific Dot1l conditional knockout mice (Dot1^f/f Aqp2Cre or Dot1l^AC); 2) DCT2/CNT/CD-specific Dot1l and Edn1 double conditional knockout mice (Dot1l^f/f Edn1^f/f Aqp2Cre or DE^AC); 3) DCT2/CNT/CD-specific Dot1l conditional knockout plus Lcn2 global knockout mice (Dot1^f/f Aqp2Cre Lcn2^-/- or DL^-/-). WT and the three knockout mice were analyzed at the age of 14 months. A variety of approaches including immunofluorescence staining coupled by high-resolution confocal microscopy, metabolic assays, real-time RT-qPCR, and Western blotting were used to assess kidney fibrosis, kidney function, and/or expression of HDAC2, Lcn2, and ET1 in mice, in IMCD3 cells overexpressing Lcn2, or in IMCD3 cells treated with various doses of recombinant Lcn2.

Results

Like DE^AC vs. Dot1^AC mice, DL^-/- vs. Dot1^AC mice displayed less pronounced kidney fibrosis and improved kidney function. While high Lcn2 expression remained in DE^AC vs. Dot1^AC mice, significantly reduced HDAC2 and ET1 levels were observed in DL^-/- vs. Dot1^AC. In IMCD3 cells, silencing Dot1l in IMCD3 cells led to upregulation of Lcn2, and addition of recombinant Lcn2 or overexpression of Lcn2 increased expression of HDAC2 and ET1.

Conclusion

Dot1l-Lcn2-HDAC2-ET1-kidney fibrosis may be considered as a new fibrogenesis pathway. That is, disruption of Dot1l in Aqp2⁺ progenitor cells not only directly upregulates ET1, but also indirectly through Lcn2, which positively regulates HDAC2, further facilitating ET1-mediated kidney fibrosis.

Funding

• NIDDK Support