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. Author manuscript; available in PMC: 2023 Jan 3.
Published in final edited form as: Cell Rep. 2022 Nov 29;41(9):111744. doi: 10.1016/j.celrep.2022.111744

Figure 6. Inhibition of GLS1 improves mitochondrial function in senescent mesenchymal stem cells.

Figure 6.

Senescent MSCs were treated with GLS1 inhibitor, BPTES (10 μM), or CB-839 (4 μM) or for 10 days. (A) Intracellular urea concentration measurement, normalized to S; data shown as mean ± SD.

(B) Representative images of Mitotracker Red live stain of Y, S, S-BPTES, and S-CB cells; scale bar represents 100 μm.

(C) Quantification of Mitotracker Red intensity per cell, normalized to S; data shown as mean ± 95% CI for >200 cells.

(D) Measurement of oxygen consumption rate (OCR) using Seahorse extracellular flux analyzer.

(E–H) Basal respiration (E), ATP-linked respiration (F), maximal respiration (G), and spare respiratory capacity (H) calculated from the OCR data in (D).

(I) Immunostaining for the phosphorylated form of histone H2AX (γ-H2AX); scale bar represents 50 μm.

(J) Quantification of γ-H2AX intensity per cell, normalized to S; data shown as mean ± 95% CI for >100 cells.

(K–N) Quantification of gene expression of inflammatory cytokines via quantitative RT-PCR normaliszed to S and internally normalized to RPL32 cycle number.