Recently, we developed highly fluorescent Ti3C2 and Nb2C Mxene quantum dots (QDs) for labeling of in vitro models. However, the mechanism of the toxicity of the prepared QDs was not explored before. In this study, we addressed the possible mechanism associated with cytotoxicity of the QDs to human umbilical vein endothelial cells (HUVECs). Exposure to up to 100 μg/ml Ti3C2 but not Nb2C QDs for 24 h significantly induced cytotoxicity. The exposure also increased intracellular Ti and Nb elements, indicating the internalization of both types of QDs. None of the QDs promoted interleukin 6 (IL‐6) and IL‐8 releases. Rather, Ti3C2 QDs significantly reduced IL‐6 and IL‐8 release, indicating that the toxicity of Ti3C2 QDs was not due to elevated inflammatory responses. Exposure to Ti3C2 but not Nb2C QDs resulted in increased LC3B‐II/I ratio and beclin‐1 proteins, biomarkers of autophagy, as well as the accumulation of autophagic substance p62. Ti3C2 QDs also more effectively promoted pro‐caspase 3 but not pro‐caspase 8 compared with Nb2C QDs. Furthermore, pre‐treatment with autophagic modulators altered the cytotoxicity of Ti3C2 QDs, which further confirmed the role of autophagic dysfunction in Ti3C2 QD‐induced toxicity to HUVECs. In conclusion, the results from this study suggested that high levels of Ti3C2 QDs could induce cytotoxicity to HUVECs by inducing the dysfunction of autophagy. Nb2C QDs appeared to be more biocompatible to HUVECs compared with Ti3C2 QDs at the same mass concentrations, which suggested a role of composition of Mxene QDs to determine their toxicity to human endothelial cells.