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The effect of glucose and insulin on the susceptibility of cultured photoreceptor‐like cells to hypoxia
Author(s) -
Layton C.,
De Moraes G.
Publication year - 2015
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/j.1755-3768.2015.0711
Subject(s) - apoptosis , medicine , insulin , wortmannin , endocrinology , annexin , programmed cell death , necrosis , biology , protein kinase b , hypoxia (environmental) , chemistry , biochemistry , organic chemistry , oxygen
Purpose Diabetes causes a retinal neuropathy, however whether glucose itself is beneficial to stressed retinal neurons is controversial. This investigation aimed to assess the effect of glucose on the resilience of cultured photoreceptor‐like cells to hypoxia. Methods 661W cells were cultured in serum free media and 5 mM glucose. Cellular insult and damage after CoCl 2 induced hypoxia was measured by caspase 3/7 activation, annexin V binding and propridium iodide marking on flow cytometry and confirmed by immunofluorescence. hif 1α, VEGF expression and akt phosphorylation were assessed with primary conjugated antibodies and flow cytometry. Results In 5 mM glucose, hypoxia increased hif 1α activation and led to cell death by both apoptosis and necrosis in a dose dependent manner. Higher glucose levels increased hypoxia induced apoptosis and cell death. Photoreceptor‐like cells expressed the insulin receptor, and the increase in apoptosis associated with 25 mM glucose was blocked by insulin treatment, which increased intracellular levels of phosphorylated akt, decreased VEGF production and decreased caspase 3 activation and Annexin binding. However in increased necrosis resulted in no significant difference in hypoxic cell death compared with 5 mM glucose. These effects of insulin were partially blocked by the PI 3K inhibitor wortmannin, but not by PKC inhibition with chelerythrine. Conclusions 661W photoreceptor‐like cells were less resilient to hypoxia in “diabetic” glucose conditions. Insulin decreased apoptosis to levels equivalent to “non‐diabetic” hypoxic cells, but increased necrosis led to no change in the percentage of dead cells. An insulin induced shift to cell death by necrosis in hypoxic photoreceptors may be one mechanism of the initial exacerbation of diabetic retinopathy with intensive treatment.