Kidney Week

Abstract: SA-PO1107

Lineage-Tracing of p16INK4a-Positive Cells Reveals Nephron-by-Nephron Heterogeneity in Kidney Aging

Session Information

• Geriatric Nephrology
  November 08, 2025 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Geriatric Nephrology

• 1300 Geriatric Nephrology

Authors

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

Koji Muro, MD

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

Ryo Yamada, MD

• Konishi, Ryo, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Kyoto Prefecture, Japan

Ryo Konishi

• Morita, Keisuke, Kyoto Daigaku Daigakuin Igaku Kenkyuka Igakubu, Kyoto, Kyoto Prefecture, Japan

Keisuke Morita, MD

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

Takuya Morinishi, MD

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

Yohei Iwashige, MD

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

Shigenori Yamamoto, MD

• Kimura, Tomonori, Osaka Daigaku, Suita, Osaka Prefecture, Japan

Tomonori Kimura, MD, PhD

• Nakanishi, Makoto, Tokyo Daigaku, Bunkyo, Tokyo, Japan

Makoto Nakanishi, MD, PhD

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

Motoko Yanagita, MD, PhD

Background

Senescent cells, marked by cell cycle arrest and secretory phenotypes, are implicated in tissue aging and repair. While p16^INK4a is a common senescence marker, its in vivo detection remains challenging. Most studies used fibroblasts in vitro, leaving epithelial senescence unclear. Here we performed a lineage tracing of p16^INK4a-positive cells using a new transgenic mouse line.

Methods

We traced p16^INK4a-positive cells using p16CreERT2:R26-tdTomato mice. Kidneys from aged (2-year-old), young, gonadectomized and Klotho^-/- mice were analyzed focusing on proximal tubules (PTs). Spatial transcriptomics via photo-isolation chemistry (PIC-RNAseq) compared proximal tubular cells (PTCs) by tdTomato (tdT) expression. Young mice underwent tamoxifen labeling followed by unilateral ischemia-reperfusion injury (IRI).

Results

tdT⁺ cells represent p16^INK4a expression at tamoxifen labeling, with 70% in PTs. tdT⁺ PTCs were more abundant in aged females than young females (14.0% vs. 6.5%, p < 0.0001), with no difference in males. Gonadectomy and Klotho^-/- increased tdT⁺ PTCs. 3D imaging showed tdT⁺ clusters in PTs. PIC-RNAseq of tdT⁺ PTCs (“Tomato⁺ PTC”), tdT- PTCs within tdT+ PTs (“Neighbor^- PTC”), and tdT- PTCs within tdT- PTs (“Control^- PTC”) revealed upregulation of DNA damage and repair pathways in “Tomato⁺ PTC”, and mitochondrial and ER stress pathways in “Neighbor^- PTC” (Figure), which were confirmed by immunostaining. Based on inter-tubular differences, whole-PT PIC-RNAseq showed distinct profiles between PTs with and without tdT+ cells (Figure). In kidneys with IRI, tdT⁺ PTCs detached from basement membrane acutely and were scarce in VCAM1+ failed-repair PTs chronically.

Conclusion

This study showed preferential localization and injury-associated behavior of p16^INK4a-positive cells in PTs. These cells show damage-related signatures, form clusters, and detach upon injury. Distinct profiles of PTs with p16^INK4a-positive cells suggest p16^INK4a as a marker of nephron vulnerability, supporting strategies to improve kidney resilience.

Digital Object Identifier (DOI)

• doi: 10.1681/ASN.2025j0rff0mk