## 4 Discussion  
The current study showed that MIA decreased the viability of chondrocytes and suppressed COL1\(\alpha\)1 and ADAMST4 levels, but increased the 8-isoprostane F2-\(\alpha\) level of chondrocytes. Concurrent incubation with AnTT and PT3 prevented this effect by preventing oxidative damage and improving COL2\(\alpha\)1/COL1\(\alpha\)1 indices. Prior incubation of chondrocytes with AnTT and PT3 did not achieve the same effects. The concurrent incubation of AnTT and MIA also increased the COL2\(\alpha\)1, aggrecan and SOX9 protein levels in chondrocytes. The inhibitory effects of MIA on chondrocytes' viability found in this study are consistent with findings in previous literature [34; 35]. [PERSON] et al. showed that MIA at a similar concentration as the current study (31.5 \(\upmu\)M) induced a decrease in the viability of SW1353 chondrocytes by proliferation arrest, not cell death [16]. As a glyceraldehyde-3-phosphate dehydrogenase inhibitor, MIA reduced the cellular energy production and expression of proteins related to glycolysis in SW1353 chondrocytes [36; 37]. It has also been suggested that MIA suppresses the mitophagy process, which clears dysfunctional mitochondria [36]. Furthermore, MIA has been shown to disrupt mitochondria membrane potential and trigger autophagy in chondrocytes in primary rat chondrocytes [35]. These findings might explain how MIA causes the number of viable and functioning chondrocytes to decrease. The chondrocytes effects of AnTT and PT3 were time-sensitive in this study. Concurrent incubation of AnTT or PT3 and MIA reversed the decrease in the viability of chondrocytes. This observation agrees with a previous animal study by [PERSON] et al. (2019), which reported that treating rats with AnTT immediately after MIA induction protected them from cartilage damage [25]. Another study showed that \(\alpha\)-tocopherol preserved chondrocytes' morphology in diabetic rats with MIA-induced cartilage damage [38]. It is hypothesised that toctorienols scavenge free radicals and maintain the intracellular defence against oxidative stress generated by MIA. Co-treatment with zinc, another antioxidant, has also been shown to ameliorate MIA-mediated reduction of chondrocyte viability [28]. However, the present study also showed that pre-treatment of toctorienol did not offer the same protection. At higher doses, locotrienol even potentiated the suppressive effects of MIA on cell viability. These findings are in contradiction with previous studies using \(\alpha\)-tocopherol. [PERSON] et al. (2007) reported that pre-incubation with \(\alpha\)-tocopherol for 24 h preserved the viability of the chondrocytes in human cartilage explants subjected to mechanical stress [39]. [PERSON] et al. (2013) treated rat primary chondrocytes with \(\alpha\)-tocopherol for 24 h before exposing them to hydrogen peroxide for 3 h [40]. The intervention preserved viability and the proliferating cell nuclear antigen expression of chondrocytes [40]. It is noteworthy that the MIA-induced chondrocyte damage is mechanistically different from other inducers, such as mechanical stress and hydrogen peroxide, so direct comparison is unattainable. Additionally, since SW1353 is a chondrosarcoma cell line, it is hypothesised that pre-treatment with toctorienols for 24 h, which are compounds with anticancer properties, could senstise the cells towards mitochondria dysfunction induced by MIA, independent of their antioxidant properties [41; 42]. Thus, the pre-treatment model was not suitable for the objective of this study due to this technical limitation, and the subsequent experiments were conducted using the co-treatment approach. Oxidative stress contributes to senescence and functional loss of chondrocytes, leading to dysregulation of cartilage haemostasis and subsequent damage [43; 44]. In this study, MIA induced significant oxidative stress in chondrocytes, as evidenced by an elevated level of 8-isoprostane F2-\(\alpha\), a stable lipid peroxidation marker [45].