Effect of KIF5A mediated lysosomal functions in cadmium exposure-induced neurotoxicity in vitro

Abstract

Objective To investigate the effect of kinesin family member 5A (KIF5A) mediated lysosomal functions in cadmium (Cd) exposure-induced neurotoxicity. Methods Primary cortical neurons isolated and primarily cultured from specific pathogen free (SPF) C57BL/6J mice were used and divided into control and Cd-exposed groups. The 50% inhibitory concentration (IC50) of cortical neurons after 24 h of Cd exposure was measured. Furthermore, the neurons were treated with 1, 2 and 3 μmol/L cadmium chloride (CdCl2) for 24 h, and then cell viability, neuronal differentiation and lysosomal function were measured. RT-qPCR and Western blotting were used to detect the changes of KIF5A expression at mRNA and protein levels. The effects of overexpression of KIF5A by adenoviral vector on cell viability, neuronal differentiation and lysosomal function were also investigated in the Cd-exposed cortical neurons. Results The IC50 value in neurons exposure to CdCl2 for 24 h was determined to be 2.9 μmol/L. Compared with the control group, the cell viability was inhibited (P < 0.01) and the total length of protrusion growth and the number of branching points were obviously reduced (P < 0.01) after 2 and 3 μmol/L CdCl2 exposure for 24 h. In addition, 2 and 3 μmol/L CdCl2 treatment also resulted in dramatically decreased lysosomal activity of cathepsin B (CTSB) and altered acidic environment when compared with the control group (P < 0.01). However, the activity cathepsin D(CTSD) was not significantly changed in CdCl2-exposed neuronal cells. More importantly, the mRNA and protein levels of KIF5A were remarkably decreased after Cd exposure (P < 0.01). Overexpression of KIF5A effectively increased the CTSB activity, stabilized the lysosomal pH value, and antagonized the inhibitory effect on cell viability and protrusion growth induced by Cd exposure (P < 0.01). Conclusion Cd exposure significantly reduces KIF5A protein expression in neurons, and thereby impairs lysosomal function and inhibits neuronal cell viability and protrusion growth.

Keywords

• cdcl2
• neurotoxicity
• kinesin family member 5a
• lysosomal function