Kidney Week

Abstract: SA-PO109

Establishment of a New Detection Method for Senescent Cells in Injured Kidneys and Proof of Their Relationship to Failed-Repair Proximal Tubules

Session Information

• AKI: Inflammation and Cell Cycle
  October 26, 2024 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Acute Kidney Injury

• 103 AKI: Mechanisms

Authors

• Yamada, Ryo, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Kyoto, Japan

Ryo Yamada, MD

• Iwashige, Yohei, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Kyoto, Japan

Yohei Iwashige, MD

• Morinishi, Takuya, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Kyoto, Japan

Takuya Morinishi, MD

• Muro, Koji, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Kyoto, Japan

Koji Muro, MD

• Yamamoto, Shigenori, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Kyoto, Japan

Shigenori Yamamoto, MD

• Kitamura, Toshio, Foundation of Biomedical Research and Innovation at Kobe, Kobe, Hyogo, Japan

Toshio Kitamura, MD, PhD

• Yanagita, Motoko, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Kyoto, Japan

Motoko Yanagita, MD, PhD

Group or Team Name

• Cell Cycle & Cellular Senescence Group.

Background

Cellular senescence occurs in response to repeated passages, persistent DNA damage, and oxidative stress and is characterized by permanent growth arrest. Recently, accumulation of senescent cells in vivo and their correlation with organ dysfunction has been reported; however, a highly sensitive method to identify senescent cells in vivo has not yet been established.

Methods

The cell cycle of the proximal tubular cells was evaluated in mice expressing the proximal tubule-specific G0 marker (Ndrg1CreER^{T2 /+}: R26-mVenus-p27K^-/+). The G0 marker is mVenus-p27K^- and is usually used to identify cells in the quiescent phase (G0 phase). We also performed spatial transcriptomics of G0 marker-positive cells using photo-isolation chemistry (PIC) based system.

Results

Using adult G0 marker mice, almost all proximal tubular cells (PTCs) were stayed at G0 phase under physiological conditions. In the chronic phase of kidney injury models, most PTCs became G0⁺, but some G0⁺ cells exhibited nuclear abnormalities with Cyclin D1 positivity (G0 double positive cells: G0^DP cells). G0^DP cells exhibited various features of cellular senescence in tissue. In addition, G0^DP cells were included within the VCAM1⁺ failed-repair PTCs (FR-PTCs), which were recently attracted attention for their inflammatory features. After injury, most FR-PTCs were G0⁺ (≥ 90%), and Cyclin D1 positivity rate gradually increased up to 20-30% depending on the injury type. The PIC-based RNA-seq study also revealed that G0^DP cells exhibited features not only of FR-PTC but also of cellular senescence compared to G0⁺ cells in the vehicle group and Cyclin D1^- G0⁺ cells (G0 single positive cells: G0^SP cells) within FR-PTCs. Timelapse imaging of primary PTCs from G0 marker mice, treated with doxorubicin (DOXO), continuously expressed the G0 marker, and their cell cycle was permanently arrested. DOXO-treated primary PTCs further obtained Cyclin D1 positivity with senescent phenotypes.

Conclusion

G0 marker with Cyclin D1 expression can be a useful tool for identifying senescent proximal tubular cells under DNA damage conditions, both in vivo and in vitro.