Fig. 3. Transient OSKMNL expression mitigates inflammatory phenotypes in diseased chondrocytes.

a Workflow summarizing the strategy adopted to mitigation of age-related disease. Chondrocytes were obtained from six distinct aged patients diagnosed late stage Osteoarthritis (OA) patients from cartilage biopsies. Healthy cells (blue), aged OA cells (red) and transiently reprogrammed OA cells (light blue) were evaluated for OA specific phenotypes. b qRT-PCR evaluation shows treatment diminishes of intracellular RNA levels of NF-κB ligand RANKL. c qRT-PCR evaluation shows treatment drops levels of iNOS for producing nitric oxide as a response and to propagate inflammatory stimulus. d Cytokine profiling of chondrocyte secretions shows an increase pro-inflammatory cytokines in OA chondrocytes that diminishes with treatment. e Cell proliferation rate as measured by cell-tracking dye. f Measurement of ATP concentration using glycerol based fluorophore shows elevation of ATP levels with treatment. g Live single-cell image of cells up taking superoxide triggered fluorescent dyes shows diminished signal after treatment. h qRT-PCR evaluation of RNA levels of antioxidant SOD2, elevated with treatment. i qRT-PCR levels of chondrogenic identity and function transcription factor SOX9 is retained after treatment. j qRT-PCR shows elevation RNA levels for extracellular matrix protein component. Young samples n = 3 individuals; aged OA samples treated and untreated n = 6 individuals. Pairwise statistical analysis was done by one-way ANOVA. For ROS (g) analysis was conducted by high-throughput imaging on 500–1000 cells per sample to allow population-wide studies with single-cell resolution. One-hundred cells per sample were randomly selected to do a statistical comparison across the three groups. Statistical analysis was then done by one-way ANOVA. *P < 0.05, **P < 0.01, ***P < 0.001. Statistical analysis by one-way ANOVA was conducted for all the other assays.