Controlled induction and targeted elimination of p16^INK4a‐expressing chondrocytes in cartilage explant culture

Abstract

Cellular senescence is a phenotypic state that contributes to age‐related diseases through the secretion of matrix‐degrading and inflammatory molecules. An emerging therapeutic strategy for osteoarthritis (OA) is to selectively eliminate senescent cells by initiating apoptosis. This study establishes a cartilage explant model of senescence induction and senolytic clearance using p16Ink4a expression as a biomarker of senescence. Growth‐factor stimulation of explants increased the expression of p16Ink4a at both the mRNA and protein levels. Applying this culture system to cartilage from p16tdTom reporter mice (a knockin allele with tdTomato fluorescent protein regulated by the endogenous p16Ink4a promoter) demonstrated the emergence of a p16‐high population that was quantified using flow cytometry for tdTomato. Cell sorting was used to separate chondrocytes based on tdTomato fluorescence and p16‐high cells showed higher senescence‐associated β‐galactosidase activity and increased gene expression of the senescence‐associated secretory phenotype as compared with p16‐low cells. The potential for effective senolysis within the cartilage extracellular matrix was assessed using navitoclax (ABT‐263). Navitoclax treatment reduced the percentage of p16‐high cells from 17.9 to 6.1% (mean of 13 matched pairs; P < 0.001) and increased cleaved caspase‐3 confirmed apoptotic activity. Together, these findings establish a physiologically relevant cartilage explant model for testing the induction and elimination of senescent chondrocytes, which will support investigations of senolytic therapy for OA.—Sessions, G. A., Copp, M. E., Liu, J.‐Y., Sinkler, M. A., D'Costa, S., Diekman, B. O. Controlled induction and targeted elimination of p16INK4a‐expressing chondrocytes in cartilage explant culture. FASEB J. 33, 12364–12373 (2019). http://www.fasebj.org

Eliminating senescent chondrogenic progenitor cells enhances chondrogenesis under intermittent hydrostatic pressure for the treatment of OA

“…Senolysis has been proved very useful to efficiently remove the senescent cells from live organs and beneficial results had been reported in many diseases such as osteoarthritis, atherosclerosis, diabetics, and so on [7,[32][33][34][35][36][37]. Jeon et al reported that UBX0101, a smallmolecule senolytic, could effectively induce senescent chondrocytes toward apoptosis and attenuate the progression of OA in both posttraumatic and age-related spontaneous OA model [7].…”

Deletion of JNK Enhances Senescence in Joint Tissues and Increases the Severity of Age‐Related Osteoarthritis in Mice

“…Lysates were also generated from mouse embryonic fibroblast (MEF) cultures established from p16 INK4a intact (MEF2) and p16 INK4a /p19 Arf−/− germline‐knockout (MEF6) mice. These MEF samples had previously been used as positive and negative controls for assessing p16 INK4a by immunoblotting . The immunoblots were stained with a total protein stain (catalog no.…”

Senolytics for Cancer Therapy: Is All that Glitters Really Gold?

“…Notably, the senolytic action of ABT-263 appears to depend on inhibition of BCL-X L and/or BCL-W, while BCL-2 inhibition is dispensable [84,86]. Similar to D+Q, ABT-263 was successful in eliminating senescent cell populations in several disease models including aging-associated bone loss [87], radiation-induced lung fibrosis [88], lung emphysema [89], uterine leiomyoma [90], tau-dependent neurodegenerative disease [91], radiation-induced neurodegeneration [92], myocardial infarction (including ischemia-reperfusion injury) [93,94], heart failure [95], pulmonary hypertension [96], insulin resistance [97], osteoarthritis [98,99], synthetic implant-mediated fibrosis [100], and Duchenne muscular dystrophy [101]. ABT-263 is currently a cornerstone in preclinical studies of senolysis and remains a promising drug for use against both liquid and solid tumors [102][103][104].…”