Co-exposure to fluoride and arsenic induces endoplasmic reticulum stress in C2C12 myoblasts and MC3T3 preosteoblasts

Environmental exposure to fluoride and arsenic is a major public health concern in many regions worldwide. Although both toxicants are known to impair skeletal development, the cellular mechanisms underlying their combined effects on skeletal muscle and bone cells remain poorly understood. This study investigated the impact of sodium fluoride (NaF) and arsenic trioxide (As₂O₃) co-exposure, for the first time, in C2C12 myoblasts (at 0.14 mM NaF + 2.4 µM As₂O₃) and MC3T3 preosteoblasts (at 0.64 mM NaF + 13 µM As₂O₃). Co-exposure significantly reduced cell proliferation and decreased the expression of the lineage specific transcription factors such as MYOD and RUNX2 in C2C12 and MC3T3 cells, respectively. Both individual and combined treatments increased ROS production, with co-exposure producing the highest oxidative stress. Further, fluoride and arsenic co-exposure exerted its effects by inducing endoplasmic reticulum stress in both cell types which was witnessed by the activation of the unfolded protein response (UPR) pathways. Specifically, in C2C12 myoblasts, co-exposure predominantly activated the PERK-Atf4-Chop pathway while suppressing IRE1α-Xbp1 expression, indicating unresolved ER stress. In contrast, MC3T3 preosteoblasts activated both PERK-Atf4-Chop and IRE1α-Xbp1 pathways, suggesting engagement of adaptive ER stress mechanisms. Collectively, these findings demonstrate that fluoride and arsenic co-exposure disrupts cellular homeostasis by inducing ER stress in both myoblasts and preosteoblasts. The differences in activation of different UPR pathways suggests C2C12 myoblasts to be more susceptible to fluoride and arsenic co-exposure compared to MC3T3 preosteoblasts.