The Impact Of NAC On Physiological Activity Of SEl1L Deficiency In The Central Nervous System Of Mice

Objective:1.To investigate the role of endoplasmic reticulum protein SEl1L in the maintenance of physiological activities of the Central Nervous System(CNS)in mice.2.To investigate the correlation of NAC and SEl1L deficiency-mediated neurodegeneration.Methods:1.Neuron-specific SEl1L knockout(NKO)and wild type control mice were generated by crossing Floxed Selll mice with Synapsin-Cre mice.Both NKO and wild type mice(WT)were further grouped by sex with n = 15.The average postnatal survival rate,body weight,skeletal muscle strength,balance/coordination,locomotion,cognition along with brain mitochondrial correlation index(ATP,MDA)of NKO mice were examined,and compared to those of wild-type(WT)control mice.2.To assess the correlation of ROS and SEL1L deficiency-mediated neurodegeneration,NKO and WT mice were randomly divided into 4 groups(WT,WT+NAC,NKO,NKO+NAC).with drug intervention.The NAC drug was fed with drinking water of 0.1 g/ml.The postnatal survival rate,body weights,skeletal muscle strength,balance/coordination,locomotion、cognition and anxiety of NKO mice were assessed,the brain tissues of the mice were examined for mitochondrial correlation index(ATP,MDA),and compared to control mice.3.Primary cerebral cortex neurons were isolated from new-born WT and NKO pups(1-3 days post birth).The cells were cultured for up to 14 days and their viability,neurite length,neurite branch points were analyzed using Incucyte Zoom,mitochondria functions of the cultured neurons were examined by Seahorse;For NAC rescue experiments,three different concentrations of NAC(0.5 μM,5μM and 50μM)were added to the media of the cultured NKO cells.After 8 days of treatment,cell viability,ATP and MDA of the cultured neurons were analyzed.Results:1.Physiological impact on Neuron-specific SEl1L knockout mice.Comparied with WT mice,the average lifespan of NKO mice was only(6±3)weeks.Both the male and female NKO mice were found to be significantly growth retarded than their WT counterparts as judged by their body weights.The muscle strength,balance/coordination,cognition abilities of NKO mice were remarkedly weaker than WT mice.In addition,NKO mice exhibited higher anxiety than WT mice(P<0.001).The ATP content of brain tissue of NKO mice was significantly decreased,whereas the MDA content was significantly increased(P<0.001).2.The rescuing effect of NAC on SEl1L deficient mice.The average lifespan of NKO fed with NAC was longer than untreated NKO mice.Additionally,the muscle strength,balance/coordination,cognition abilities of NAC-treated NKO mice were significantly improved(P<0.05),but NAC-treated NKO mice still had a great difference with WT mice(P<0.05).After treatment with NAC,the body weight did not change either(P>0.05).The brain ATP content was significantly increased,whereas the MDA content was significantly decreased(P<0.05).NAC had no effect on the physiological activity of the mice(P>0.05).3.Morphology and function of neuronal primary cells in the cerebral cortex of mice.Comparied with WT cell,NKO cell viability was week,its neurith length was short,the number of NKO neurith branch was small,and NKO cell had mitochondrial dysfunction.The ATP content of NKO cell was significantly decreased,whereas the MDA content was significantly increased(P<0.01).Then Cells were treated with NAC.Compared with WT cell and untreated NKO cell,the cell viability and ATP content of NAC-treated NKO cell were increased,whereas the MDA content was decreased(P<0.05).And 5uM NAC treatment significantly improved the viability and mitochondrial dysfunction of NKO neurons.Conclusions:1.We showed through in vitro and in vivo experiments that SEL1L deficiency in the CNS leads to neurodegeneration.Thus,it is concluded that SE11L plays an important role in the maintenance of normal physiological functions of the central nervous system2.NAC treatment partially reverse the balance/coordination,locomotion and cognition,muscle strength and other symptoms of NKO mice.Thus,it provides evidence that mitochondrial dysfunction,or more specifically overproduction of ROS,may be an underlying mechanism of the SEL1L deficieny-mediated neurodegeneration.