Elimination of p19ARF-expressing cells enhances pulmonary function in mice
Michihiro Hashimoto, Azusa Asai, Hiroyuki Kawagishi, Ryuta Mikawa, Yuji Iwashita, Kazuki Kanayama, Kazushi Sugimoto, Tadashi Sato, Mitsuo Maruyama, Masataka Sugimoto
Michihiro Hashimoto, Azusa Asai, Hiroyuki Kawagishi, Ryuta Mikawa, Yuji Iwashita, Kazuki Kanayama, Kazushi Sugimoto, Tadashi Sato, Mitsuo Maruyama, Masataka Sugimoto
View: Text | PDF
Research Article Aging Cell biology

Elimination of p19ARF-expressing cells enhances pulmonary function in mice

Abstract

Senescent cells accumulate in many tissues as animals age and are considered to underlie several aging-associated pathologies. The tumor suppressors p19ARF and p16INK4a, both of which are encoded in the CDKN2A locus, play critical roles in inducing and maintaining permanent cell cycle arrest during cellular senescence. Although the elimination of p16INK4a-expressing cells extends the life span of the mouse, it is unclear whether tissue function is restored by the elimination of senescent cells in aged animals and whether and how p19ARF contributes to tissue aging. The aging-associated decline in lung function is characterized by an increase in compliance as well as pathogenic susceptibility to pulmonary diseases. We herein demonstrated that pulmonary function in 12-month-old mice was reversibly restored by the elimination of p19ARF-expressing cells. The ablation of p19ARF-expressing cells using a toxin receptor-mediated cell knockout system ameliorated aging-associated lung hypofunction. Furthermore, the aging-associated gene expression profile was reversed after the elimination of p19ARF. Our results indicate that the aging-associated decline in lung function was, at least partly, attributed to p19ARF and was recovered by eliminating p19ARF-expressing cells.

Authors

Michihiro Hashimoto, Azusa Asai, Hiroyuki Kawagishi, Ryuta Mikawa, Yuji Iwashita, Kazuki Kanayama, Kazushi Sugimoto, Tadashi Sato, Mitsuo Maruyama, Masataka Sugimoto

×

Figure 8

Changes in senescence-associated gene expression in DT-treated lungs.

Options: View larger image (or click on image) Download as PowerPoint
Changes in senescence-associated gene expression in DT-treated lungs. (A...
(A) Total RNA was extracted from 2- and 12-month-old female ARF–diphtheria toxin receptor (ARF-DTR) mouse lung tissues. Twelve-month-old mice were treated with PBS or diphtheria toxin (DT), as depicted in Figure 4A. The expression of the indicated genes was analyzed by real-time PCR. mRNA levels were normalized to GAPDH mRNA in each sample. Values represent the mean ± SEM. (B) Wild-type MEFs were infected with retroviruses encoding shRNA against MMP-10. Two-independent shRNA were used to knockdown MMP-10. Scrambled short hairpin (sh-SCR) was used as a control in each group. Infected cells were selected with puromycin for 3 days, exposed to ionizing radiation (10 Gy), and cultured under 3% oxygen conditions for 10 days. The expression of MMP-10 was analyzed by immunoblotting. Lamin A/C was used as a loading control. (C) shRNA-infected senescent MEFs were analyzed for the amount of elastin. In the chemical inhibition of MMPs, senescent MEFs were cultured in the presence of an MMP inhibitor [N-isobutyl-N-(4-methoxyphenylsulfonyl)-glycylhydroxamic acid {NNGH}]. Media were replaced every 3 days. The amount of elastin was measured and then normalized to the total protein content in each sample. Values represent the mean ± SD of triplicate samples. Data are representative of 2 independent experiments. Data were analyzed 1-way ANOVA followed by post-hoc Tukey-Kramer multiple comparison test (for comparison of 3 groups) or unpaired Student’s t test. *P < 0.05, **P < 0.01, ***P < 0.001.