Diabetic nephropathy (DN), a common cause of chronic renal failure and end-stage renal disease, leads to a high mortality. However, the role of TTN-AS1 in extracellular matrix (ECM) accumulation during DN remains unclear. In our study, TTN-AS1 exhibited high expression in high glucose-treated mesangial cells, and TTN-AS1 silencing alleviated high glucose-induced ECM accumulation in mesangial cells. Additionally, animal study revealed that TTN-AS1 was upregulated in renal tissues of DN rats, and TTN-AS1 knockdown mitigated renal injury of DN rats. Mechanistically, TTN-AS1 was validated to bind to miR-493-3p, and miR-493-3p targeted forkhead box P2 (FOXP2) 3'untranslated region in mesangial cells. TTN-AS1 interacted with miR-493-3p to upregulate FOXP2 in vitro and in vivo. Moreover, FOXP2 overexpression counteracted the effects of TTN-AS1 silencing on the ECM accumulation. In conclusion, TTN-AS1 exacerbated ECM accumulation via the miR-493-3p/FOXP2 axis during DN development. This research may provide a potential new direction for DN treatment.