The collagen receptor integrin alpha1beta1 slows cartilage degradation in mouse models of osteoarthritis. Epidermal growth factor receptor (EGFR) signalling appears to affect multiple aspects of cartilage health, although whether integrin alpha1beta1 regulates EGFR and downstream superoxide production has not been demonstrated in chondrocytes. We examined how integrin alpha1beta1/EGFR interplay regulates superoxide levels in ex vivo articular chondrocytes of male and female mice. Through confocal microscopy, we determined that chondrocytes of male mice had elevated levels of superoxide compared to female mice. Only male mice demonstrated a condyle effect, with greater superoxide levels in the medial condyles, whereas female mice showed a genotype effect, whereby integrin alpha1beta1 restricted superoxide abundance. Inhibition of EGFR with erlotinib dampened superoxide levels in the medial condyles of male mice but exacerbated superoxide abundance in wild-type female mice. These findings indicate a need for the consideration of sex and compartment when assessing cell signalling in cartilage.