["The Journal of Physiology, Volume 604, Issue 11, Page 4225-4246, 1 June 2026. ", "\nAbstract figure legend The human placenta not only detects the presence of Pb and Cd but also carries hypoxia and inflammatory risks, which are associated with pregnancy complications. To understand the impact of these factors on the placental trophoblast, we evaluated the effect of Pb and Cd on mitochondrial function and the NLRP3 inflammasome in three trophoblast cell lines under normoxia, hypoxia and pro‐inflammatory conditions. We observed that hypoxia decreased ΔΨ and promoted apoptosis in JEG‐3 cells, increased ΔΨ and prevented apoptosis in BeWo cells, and did not change ΔΨ and apoptosis in HTR‐8/SVneo cells. Moreover, Pb under hypoxic conditions reduced ΔΨ and promoted apoptosis of BeWo cells. Exposure of BeWo and HTR‐8/SVneo cells to hypoxia, Pb or Cd alone upregulated the expression of NLRP3 and pro‐caspase 1 but did not activate the NLRP3 inflammasome. To conclude, Pb and Cd affected trophoblast mitochondrial function and NLRP3 proteins in trophoblast cell lines, but in a cell line‐specific way.\n\n\n\n\n\n\n\n\n\nAbstract\nHeavy metals disrupt mitochondrial function and activate the NOD‐like receptor pyrin‐containing 3 (NLRP3) inflammasome. We investigated the effect of lead (Pb)/cadmium (Cd) on mitochondrial function and NLRP3 inflammasome activation in human trophoblast under normoxic, hypoxic and pro‐inflammatory conditions. JEG‐3, BeWo and HTR‐8/SVneo cells were exposed to Pb or Cd for 24 h in the absence or presence of hypoxia or pro‐inflammatory lipopolysaccharide (LPS) or poly(I:C). Then, we evaluated cell viability, apoptosis, mitochondrial DNA copy number (mtDNAcn), mitochondrial membrane potential (ΔΨ), NLRP3 inflammasome proteins and interleukin (IL)‐1β secretion. Although our data showed that Pb, Cd, hypoxia, poly(I:C) and LPS decreased mtDNAcn in the three cell lines, the effects of these treatments on other biomarkers were different in the different cell lines. We found that hypoxia decreased ΔΨ and promoted apoptosis in JEG‐3 cells, increased ΔΨ and prevented apoptosis in BeWo cells, and did not change ΔΨ and apoptosis in HTR‐8/SVneo cells. Moreover, Pb under hypoxic conditions reduced ΔΨ and promoted apoptosis of BeWo cells. Exposure of BeWo and HTR‐8/SVneo cells to hypoxia, Pb or Cd alone upregulated the expression of NLRP3 and pro‐caspase 1 but did not activate the NLRP3 inflammasome since cleaved‐caspase 1 and IL‐1β were not increased. To conclude, Pb and Cd affected trophoblast mitochondrial function and NLRP3 proteins in trophoblast cell lines, but in a cell line‐specific way.\n\n\n\n\n\n\n\n\n\nKey points\n\nThe objective of this work was an understanding of the effect of lead (Pb) and cadmium (Cd) on mitochondrial function and NLRP3 inflammasome activation in human trophoblast cell lines under normoxic, hypoxic and pro‐inflammatory conditions.\nApoptosis of JEG‐3 cells was increased by hypoxia, while in BeWo cells, apoptosis was decreased by hypoxia, and in HTR‐8/SVneo, apoptosis was not affected by hypoxic treatment.\nExposure to either Pb or Cd decreased mtDNAcn in three human placental trophoblast cell lines. However, Pb under hypoxia induced a decrease of ΔΨ and promoted apoptosis of BeWo cells, but Cd did not induce a reduction in ΔΨ in the three trophoblast cell lines under any conditions.\nExposure to hypoxia, Pb or Cd increased NLRP3 and pro‐caspase 1 in BeWo and HTR‐8/SVneo cells.\nOur findings highlight that Pb and Cd affected trophoblast mitochondrial function and NLRP3 proteins in trophoblast cell lines but in a cell line‐specific way.\n\n\n"]