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목적 : To evaluate the effects of nerve growth factor (NGF), which is activated during inflammatory episodes of ocular diseases, on the apoptotic response in cultured human primary conjunctival epithelial cells (pHCECs)
방법 : Levels of NGF transcripts and NGF protein in pHCEC grown in medium with normal-osmolarity (307 mOsm/L) or hyperosmolar media (350, 400, and 450 mOsm/L) were determined using RT-PCR and ELISA, respectively. To assess apoptosis, pHCEC were cultured in normal- or 400 mOsm/L hyperosmolar medium with NGF neutralizing antibody or recombinant human NGF for six hours, before analysis by flow cytometry. Levels of Bcl-xL, Bax, phospho-JNK, and cleaved caspase-3 were detected using western blotting. Levels of the inflammatory cytokine IL-6 was analyzed using ELISA.
결과 : Hyperosmolar conditions increased NGF levels in cultured pHCECs. Hyperosmolarity and exposure to NGF neutralizing antibody significantly increased the number of apoptotic cells. Addition of recombinant human NGF to 400 mOsm/L osmolar medium decreased the number of apoptotic cells, reduced the expression of phospho-JNK, Bax, and cleaved caspase-3 expression, and increased the expression of Bcl-xL. Levels of IL-6 were increased by hyperosmotic stress but decreased by exposure to recombinant human NGF.
결론 : Hyperosmolarity induces apoptosis of pHCECs by activating JNK signaling. Increased levels of NGF under hyperosmolar conditions may contribute, at least in part, to the reduced apoptosis of pHCECs, and may be beneficial in recovering conjunctival damage due to chronic hyperosmolar stress.
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