Abstract:

Objective To discuss the effect of hirudin on post-stroke depression （PSD） in the mice， and to clarify its potential mechanism. Methods Sixty male C57BL/6 mice were randomly divided into control group， PSD group， PSD+low dose of hirudin group， PSD+medium dose of hirudin group， and PSD+high dose of hirudin group， and there were 12 mice in each group. The stroke model was established by middle cerebral artery occlusion （MCAO）， and the depression model was induced by chronic unpredictable mild stress （CUMS） combined with solitary housing. The mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were intravenously injected with 10， 20， and 40 U·kg^-1 hirudin， respectively， while the mice in control and PSD groups received equal volumes of saline. The body weights of the mice were recorded on days 0， 7， 14， and 21 of CUMS. The LONGA neurological function score was calculated. Sucrose preference test， tail suspension test， and forced swimming test were used to detect the sucrose preference rate， immobility time in tail suspension， and forced swimming in various groups， respectively； HE staining was used to observe the histopathological changes in the medial prefrontal cortex （mPFC）； biochemical kits were used to detect the levels of malondialdehyde （MDA） and reduced glutathione （GSH） as well as superoxide dismutase （SOD） activity in mPFC tissue of the mice in various groups； 2'，7'-dichlorodihydrofluorescein diacetate （DCFH-DA） fluorescence probe method was used to detect the reactive oxygen species （ROS） positive rate in mPFC tissue of the mice in various groups； real-time fluorescence quantitative PCR （RT-qPCR） and Western blotting method were used to detect the expression levels of nucleoredoxin （NXN） mRNA and protein in mPFC tissue of the mice in various groups. Results Compared with control group， the body weight of the mice in PSD group was significantly decreased on days 0， 7， 14， and 21 of CUMS （P<0.05 or P<0.01）. Compared with PSD group， the body weights of the mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were significantly increased on days 14 and 21 of CUMS （P<0.05 or P<0.01）， and the neurological function scores were significantly decreased （P<0.05 or P<0.01）. The sucrose preference test， tail suspension test， and forced swimming test results showed that compared with control group， the sucrose preference rate of the mice in PSD group was significantly decreased （P<0.01）， while the immobility times in tail suspension and forced swimming were significantly increased （P<0.01）. Compared with PSD group， the sucrose preference rates of the mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were significantly increased （P<0.05 or P<0.01）， and the immobility times were significantly decreased （P<0.05 or P<0.01）. The HE staining showed normal cell morphology， clear structure， and uniform size distribution in mPFC tissue in control group. In PSD and PSD+low dose of hirudin groups， the number of the cells in mPFC tissue was significantly reduced， with severe vacuolar degeneration and pyknotic nuclei. Compared with PSD group， the numbers of the cells in PSD+medium dose of hirudin and PSD+high dose of hirudin groups were significantly increased， and the vacuolar degeneration and nuclear pyknosis were alleviated. The Biochemical and DCFH-DA fluorescence probe assays results showed that compared with control group， the GSH level and SOD activity in mPFC tissue of the mice in PSD group were significantly decreased （P<0.01）， while the MDA level and ROS positive rate were significantly increased （P<0.01）. Compared with PSD group， the GSH levels and SOD activities of the mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were significantly increased （P<0.05 or P<0.01）， while the MDA levels and ROS positive rates were significantly decreased （P<0.05 or P<0.01）. The RT-qPCR and Western blotting results showed that compared with control group， the expression levels of NXN mRNA and protein in mPFC tissue of the mice in PSD group were significantly decreased （P<0.01）. Compared with PSD group， the expression levels of NXN mRNA and protein in mPFC tissue of the mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were significantly increased （P<0.05 or P<0.01）. Conclusion Hirudin promotes redox balance in mPFC of the PSD mice， repairs neurological damage， and improves PSD.

Key words: Hirudin, Stroke, Depression, Medial prefrontal cortex, Nucleoredoxin