Abstract: OBJECTIVE：To investigate neurotoxic effects of chronic cobalt exposure on human cerebral organoids and to provide experimental evidence for assessing potential health risks of heavy metal cobalt. METHODS：Human iPSC-derived cerebral organoids at day 28 of differentiation were randomly divided into the control group， 5， 10， and 20 μmol/L CoCl₂ exposure groups， with continuous exposure for 28 days. Samples of human cerebral organoids were collected on day 56. Immunofluorescence staining was used to systematically characterize the differentiation status (SOX2/TBR2/CTIP2/TUJ1/NeuN/SYN1)，expression of hypoxia-inducible factor-1α，cellular hypoxia status (Hypoxyprobe)，proliferation and apoptosis levels (Ki-67/TUNEL)， and astrocyte damage (GFAP) of human cerebral organoids.RESULTS： Day56 organoids maintained high proliferative activity. Neural progenitor cells self-organized into ventricular zone-like structures， forming physiological lamellar architectures with intermediate progenitors and deep-layer cortical neurons. Chronic cobalt exposure significantly upregulated HIF-1α protein expression (P<0.01) and increased hypoxic cells versus controls. TUNEL-positive apoptotic cells showed dose-dependent elevation (P<0.01)，predominantly localized in intermediate progenitor/neuron interface regions. Bright-field monitoring revealed progressive disintegration of ventricular zone structures in medium/high-dose groups. GFAP fluorescence intensity was markedly attenuated in exposure groups， indicating astrocyte impairment. CONCLUSION： Chronic cobalt exposure stabilizes HIF-1α to induce pseudohypoxia， leading to three neurotoxic outcomes： increased neuronal apoptosis，ventricular zone collapse，and astrocyte damage.

Key words: cobalt, human cerebral organoids, HIF-1α, neurotoxicity