Abstract This study aimed to investigate the cytotoxicity and pro-inflammatory responses induced by tungsten disulphide (WS2) and molybdenum disulphide (MoS2) nanoparticles (NPs) in human bronchial cells (BEAS-2B). For cytotoxicity assessment, the cells were exposed to different concentrations (2.5–200 µg/mL) of WS2-NPs or MoS2-NPs for 24 and 48 h and then the MTT assay was performed. Afterwards, long-term toxicity was assessed by the colony forming efficiency assay (CFEA) during a 10 days’ exposure of the cells. For pro-inflammatory responses, the expression of interleukin-6 (IL-6) and interleukin-1β (IL-1β) mRNA was estimated by the real-time PCR method. Both nanomaterials showed similar cytotoxic effects on BEAS-2B cells assessed by the MTT assay, i.e. reduction in cell viability to approx. 60–70% at concentrations of 2.5 and 5 μg/mL after 24 and 48 h. The percentage viability remained relatively constant at this level across all concentrations above 5 μg/mL. In long-term exposure, both nanomaterials inhibited colony formation in a wide range of concentrations up to 100 µg/mL. MoS2-NPs were slightly more cytotoxic than WS2-NPs. Additionally, MoS2-NPs caused an increase in mRNA levels of cytokines, IL-1β, and IL-6 at concentration of 50 µg/mL, while WS2-NPs did not cause any changes in the level of mRNA for both cytokines. We also visualised the changes in the cells as a result of WS2-NPs or MoS2-NPs exposure (2.5 and 25 µg/mL) via holotomographic microscopy. This work demonstrates the hazardous potential of both nanomaterials and indicate that WS2 and MoS2 nanoparticles should be included in the occupational risk assessment.