3.4. Effects of α-Klotho on GCM -induced cytotoxicity of mouse primary neuronal culture
Primary glia culture was pretreated with serum-free medium (GCM) or with 1nM α-Klotho (GCM-KL) for 24 hours, and then challenged with 1 µg/mL LPS for 24 hours. Initially, it was necessary to determine the amount of GCM coming from LPS-challenged glia cell (GCM group) which caused neuronal death. Thus, we replaced 10, 25, and 50% of the cultured medium of the neuronal culture by GCM. The results showed that when the concentration of 25 and 50% of the neuronal culture medium were switched to GCM, there was an increase in neuronal toxicity when compared to the control group (Fig.5).
After determining the concentration of GCM that induced neurotoxicity, we investigated whether the GCM from α-Klotho pretreated glial culture (1nM) and stimulated with LPS (GCM-KL) would be able to decrease the neuronal death when compared to GCM at both 25 and 50% concentrations (Fig.6). The results showed that 25% of GCM-KL reversed the increase in neuronal death caused by GCM (Fig.6A), but α-Klotho pretreatment was not able to induce any effect to a high concentration (50% of GCM-KL) (Fig.6B).