Fig. 3: CXCR3 knockdown contributes to mitochondrial dysfunction.

A Pathway enrichment analysis using RNA sequencing data from orthotopic xenografts illustrating key signaling pathways involved after CXCR3 ablation. n = 3 per group. B Heatmap analysis highlighting changes in expression of mitochondrial processes in shCtrl and shCXCR3 xenografts. C Representative fluorescence images from orthotopic xenografts showing the subcellular localization of CXCR3 receptor expression in normal brain tissue. Original magnification: ×40. D Representative fluorescence images stained with mitoSOX (red) and Hoechst 33342 (blue). E Flow cytometry analysis of mitochondrial reactive oxygen species production from U87 and U251 cells expressing shCXCR3 compared to control. Red indicates the MitoSOX red stain, which detects superoxide production within the mitochondria of live cells. F Representative fluorescence images in U87 and U251 cells. Green indicates the MitoTracker green dye, which labels mitochondria and provides a measure of total mitochondrial mass within live cells. Red indicates the Mitotracker red CMXROS dye, which accumulates in mitochondria based on their membrane potential. Blue indicates hoechst 33342. Reduced Mitotracker red staining indicates dysfunctional mitochondria with mitochondrial depolarization. G Seahorse mito stress analysis of mitochondrial respiration in U87 cells. (Top) Oxygen Consumption rate (OCR) of U87 cells expressing shCtrl and shCXCR3 under basal conditions, followed by sequential addition of mitochondrial inhibitors (right). Quantification of key respiratory parameters: Non-mit O2 cons non-mitochondrial O2 consumption, Basal basal respiration, Maximal maximal respiration, ATP-linked ATP-linked respiration, Spare Cap. spare capacity. n = 8 per group. Error bars indicate mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns (no statistical significance).