Transplanted Senescent Cells Induce an Osteoarthritis-Like Condition in Mice

Abstract

Osteoarthritis (OA) is the leading form of arthritis in the elderly, causing pain, disability, and immobility. OA has been associated with accumulation of senescent cells in or near joints. However, evidence for a causal link between OA and cellular senescence is lacking. Here, we present a novel senescent cell transplantation model involving injection of small numbers of senescent or nonsenescent cells from the ear cartilage of luciferase-expressing mice into the knee joint area of wild-type mice. By using bioluminescence and 18FDG PET imaging, we could track the injected cells in vivo for more than 10 days. Transplanting senescent cells into the knee region caused leg pain, impaired mobility, and radiographic and histological changes suggestive of OA. Transplanting nonsenescent cells had less of these effects. Thus, senescent cells can induce an OA-like state and targeting senescent cells could be a promising strategy for treating OA.

Keywords: Bioluminescence imaging; Cell transplantation; Fluorodeoxyglucose; Mechanical allodynia; Senolytics.

Figure 1.

Establishing a senescent cell transplantation model. Primary mouse ear cartilage fibroblasts were isolated from CAG-luc transgenic mice. (A) Control nonsenescent (CON) and radiation-induced senescent fibroblasts (SEN) were assayed for cellular senescence-associated β-galactosidase (SABG) activity and stained with DAPI. (B) The relative mRNA abundance of p16^Ink4a and p21^Cip1 is shown. Results (N = 6) are expressed as mean ± SEM. *p < .05. (C) Conditioned medium (CM) was collected from CON and SEN fibroblasts. Cytokine protein levels in CM were measured by multiplex assay and are expressed as a function of cell number. Selected cytokine levels are shown as the fold change relative to the average level in the CON group. Results (N = 6) are expressed as mean ± SEM. *p < .05. Right knees from 7-month-old C57BL/6 female mice were injected with PBS and left knees were injected with 2 × 105 senescent or nonsenescent control CAG-luc mouse primary ear fibroblasts in PBS. (D) One day later, bioluminescence was assayed using a Xenogen Ivis 200 Imaging System. Representative images are shown. (E) A further 3 days later, mice were injected with ¹⁸fluorodeoxyglucose (¹⁸FDG) and images focused on knee region were acquired by PET/CT scanning. Representative images are shown. ¹⁸FDG standard uptake value (F) and uptake ratios of the left to right knees within the same mice (G) are shown as mean ± SEM (N = 7).

Figure 2.

Senescent cell transplantation induces osteoarthritis-like phenotypes and impairs function. Seven-month-old C57BL/6 female mice were subjected to control nonsenescent (CON) or senescent (SEN) primary ear fibroblast transplantation in the knee region. (A) Safranin O/Fast Green staining was performed 3 months after transplantation. Representative images (5× magnification of the knee joint) are shown. (B) Histology scores for the knee joints are shown as mean ± SEM (N = 5). (C) Representative radiographs are shown. (D and E) The von Frey filament assay was performed 3 months after transplantation. Paw withdrawal frequencies using both 0.16g (D) and 0.4g filaments (E) are shown as mean ± SEM (N = 7). (F) Rotarod assay was performed before and 1 month after transplantation. Results are shown as percent change in time to falling relative to baseline. Results (N = 7) are expressed as mean ± SEM. (G and H) Locomotor activity during 20 minutes of evaluation was monitored 3 months after transplantation. (G) Stationary time (ST) and active time (AT) are shown as mean ± SEM (N = 7). (H) Distance traveled is shown as mean ± SEM (N = 7).