Peripheral Pain Modulation of Chrysaora pacifica Jellyfish Venom Requires Both Ca2+ Influx and TRPA1 Channel Activation in Rats.

The venom of jellyfish triggers severe dermal pain along with inflammation and tissue necrosis, and occasionally, induces internal organ dysfunction. However, the basic mechanisms underlying its cytotoxic effects are still unknown. Here, we report one of the mechanisms involved in peripheral pain modulation associated with inflammatory and neurotoxic oxidative signaling in rats using the venom of jellyfish, Chrysaora pacifica (CpV). This jellyfish is identified by brown tentacles carrying nematocysts filled with cytotoxic venom that induces severe pain, pruritus, tentacle marks, and blisters. The subcutaneous injection of CpV into rat forepaws in behavioral tests triggered nociceptive response with a decreased threshold for mechanical pain perception. These responses lasted up to 48 h and were completely blocked by verapamil and TTA-P2, T-type Ca2+ channel blockers, or HC030031, a transient receptor potential cation ankyrin 1 (TRPA1) channel blocker, while another Ca2+ channel blocker, nimodipine, was ineffective. Also, treatment with Ca2+ chelators (EGTA and BaptaAM) significantly alleviated the CpV-induced pain response. These results indicate that CpV-induced pain modulation may require both Ca2+ influx through the T-type Ca2+ channels and activation of TRPA1 channels. Furthermore, CpV induced Ca2+-mediated oxidative neurotoxicity in the dorsal root ganglion (DRG) and cortical neurons dissociated from rats, resulting in decreased neuronal viability and increased intracellular levels of ROS. Taken together, CpV may activate Ca2+-mediated oxidative signaling to produce excessive ROS acting as an endogenous agonist of TRPA1 channels in the peripheral terminals of the primary afferent neurons, resulting in persistent inflammatory pain. These findings provide strong evidence supporting the therapeutic effectiveness of blocking oxidative signaling against pain and cytotoxicity induced by jellyfish venom.

Stemness characteristics of human corneal endothelial cells cultured in various media.

PURPOSE: To investigate stemness characteristics of human corneal endothelial cells (HCECs) cultured in various media. METHODS: Human corneal endothelial cells were isolated using a sphere-forming assay. Cells were allowed to attach to the bottom of culture plates and were cultured in different media designated as medium A (Opti-MEM I with 8% fetal bovine serum), medium B (DMEM/F12 with B27 supplement), medium E (DMEM/F12 with epidermal growth factor [EGF]), and medium BE (DMEM/F12 with B27 supplement and EGF), respectively. Cell morphology was evaluated with an phase-contrast inverted microscope. Immunofluorescence staining and western blotting of nestin, octamer-binding transcription factor (OCT3/4), glial fibrillary acidic protein (GFAP), zonula occludens-1 (ZO-1), collagen VIII alpha2, and Na-K ATPase was performed. Cell proliferation was assessed with a cell counting kit-8 assay. RESULTS: A few cultured cells stained with nestin. The cells cultured in medium A expressed high levels of GFAP, OCT3/4, and nestin, and higher levels of ZO-1 were expressed in the cells cultured in medium A and medium B compared with cells cultured in the other media. The cells cultured in medium A assumed a fibroblast-like shape, whereas the cells cultured in medium B and medium BE appeared as mosaics. Cell proliferation was highest in medium A compared with those cultured in the other media. CONCLUSIONS: Cultured HCECs expressed stem cell markers, including nestin, OCT3/4, and GFAP. The expression of stem cell markers differed according to the culture media and associated proliferation rate.