Establishment, characterization, and exploration of the first cell line of Qingtian paddy field carp (Cyprinus carpio var. qingtianensis) and its response to hypoxia mechanism

Abstract

To explore the stress resistance mechanisms in Qingtian paddy field carp (PF-carp) Cyprinus carpio var. qingtianensis under hypoxic conditions, we established the first fin-derived cell line for this species, termed cells of Qingtian paddy field carp fin (CQF). The CQF cells, primarily fibroblasts, were cultured at 27 °C with 5% CO₂ in the L-15 medium, supplemented with 1% antibiotics and 15% fetal bovine serum. To examine cellular responses to hypoxia, CQF cells were subjected to four conditions: normoxia and hypoxia exposure for 6, 12, and 24 h. With increasing hypoxia duration, abnormal cell morphology, disrupted redox homeostasis, and severe apoptosis were observed. These effects were characterized by elevated reactive oxygen species (ROS) levels, loss of mitochondrial membrane potential (ΔΨm), and upregulation of caspase-3. CQF cells adapted to hypoxia by upregulating antioxidant enzyme activity. Furthermore, hypoxia-inducible factor 1α (HIF-1α) overexpression plasmids were constructed using pcDNA3.1 as a vector, with fluorescent tagging via eGFP-N3 for subcellular localization. Under normoxic conditions, HIF-1α was located in both the cytoplasm and nucleus, but translocated to the nucleus under hypoxia, where it regulated adaptive transcription. Overexpression of HIF-1α in hypoxic CQF cells reduced apoptosis and preserved cell integrity. Thus, the CQF cell line provides a valuable model for investigating autophagy, apoptosis, and gene expression during hypoxia in PF-carp, offering novel insights into hypoxia response mechanisms in fish.