Degenerative effects of cobalt-chloride treatment on neurons and microglia in a porcine retina organ culture model

摘要

Abstract In order to understand the pathological processes of retinal diseases, experimental models are necessary. Cobalt, as part of the vitamin B12 complex, is important for neuronal integrity. However, it is known that high quantities of cobalt induce cytotoxic mechanisms via hypoxia mimicry. Therefore, we tested the degenerative effect of cobalt chloride (CoCl 2 ) on neurons and microglia in a porcine retina organ culture model. Organotypic cultures of porcine retinas were cultured and treated with different concentrations of CoCl 2 (0, 100, 300 and 500?μM) for 48?h. After four and eight days, CoCl 2 induced a strong degeneration of the porcine retina, starting at 300?μM. A loss of retinal ganglion cells (RGCs, Brn-3a), amacrine cells (calretinin) and bipolar cells (PKCα) was observed. Additionally, a high expression of hypoxia induced factor-1a (HIF-1a) and heat shock protein 70 (HSP70) was noted at both points in time. Also, the Caspase 3 protein was activated and P21 expression was induced. However, only at day four, the Bax/Bcl-2 ratio was increased. The effect of CoCl 2 was not restricted to neurons. CoCl 2 concentrations reduced the microglia amount (Iba1) and activity (Iba1?+?Fc γ -Receptor) at both points in time. These damaging effects on microglia were surprising, since CoCl 2 causes hypoxia and a pro-inflammatory environment. However, high concentrations of CoCl 2 also seem to be toxic to these cells. Similar degenerative mechanisms as in comparison to retinal ischemia animal models were observed. In summary, an effective and reproducible hypoxia-mimicking organotypic model for retinal degeneration was established, which is easy to handle and ready for drug studies. Highlights ? CoCl 2 stabilized HIF-1α during the cultivation period. ? CoCl 2 induced a neuronal degeneration of the porcine retina; starting at 300?μM. ? High concentrations of CoCl 2 are generally toxic for neurons and microglia. ? Retinal degeneration occurred via apoptosis and cellular senescence mechanisms. ? The intrinsic pathway via Bax played a role only at the beginning of the cultivation.