Observation On Growth And Development Of Mice With CTNND2 Deficiency In Autism Model

Autism spectrum disorder?ASD?,as a representative disease of generalized developmental disorder,has attracted worldwide attention.The incidence of ASD has increased over the past two decades,according to the new study in the United States,one in 54 children has ASD.ASD occurs frequently in infants and young children,and the prevalence rate of male and female patients is about 5:1.The core symptoms are speech developmental disorder,interpersonal communication disorder,narrow interest and rigid behavior pattern,accompanied by varying degrees of neurological and mental retardation.Unfortunately,apart from early behavioral interventions,there are no effective treatments or drugs,and the pathogenesis of ASD is still not fully understood.The onset of ASD is substantially heterogeneous,with some children showing signs of developmental retardation in the first 18 months of life.However,25 to 40 percent of children with autism initially show normal development and do not develop the developmental disorder of early autism until 18 to 24 months of age.The early onset and degenerative behavior of autism may be related to the disorder of neuronal dendritic development in the brain.The human cerebral cortex is the highest center of sensory analysis,and different brain regions have different functional localization.The temporal lobe mainly integrates and processes auditory,vision,linguistic and psychological signals,which is of great significance to the language communication and interpersonal communication of children with autism.The hippocampus is mainly responsible for the adjustment of cognition,learning,memory,emotions and directional functions.The hippocampal dentate gyrus is mainly related to learning and spatial memory capabilities,which has a profound impact on the spatial recognition,learning and memory of children with ASD.Catenin delta 2?CTNND2?is located on the short arm of chromosome 5 in humans and on chromosome 15 in mice and codes for?-catenin,which is expressed almost entirely in neurons.?-catenin expression begins on the 10th day of the embryo and peaks on the 7th day after birth.In mature neurons?-catenin is localized in dendrites and dendritic spines and is expressed primarily in the cerebral cortex,hippocampus,olfactory bulb,and to a lesser extent in the thalamus and cerebellum.Exome sequence analysis of 13 patients with ASDs revealed that CTNND2 is closely related to ASD,and its lack of function may be a major factor limiting the formation and maintenance of dendritic processes.In addition,some studies have shown that the interaction of?-catenin with N-cadherin and proteins containing PDZ domains is a key factor regulating neuronal synapse and dendritic spine development and maturation.However,there is no literature on the growth and development characteristics of CTNND2 deficient mice and the changes in neurons dendritic of their brain with age.Part One Observed the growth and development patternsand behavioral changes of CTNND2 deficient miceObjective:Observe the early physiological development,neurological development and behavioral changes of CTNND2 deficient mice.Methods:1.The mice genotypes were identified by Reverse Transcription Polymerase Chain Reaction?RT-pcr?.2.Grouping:Postnatal?PN?0 d mice were divided into CTNND2^-/-group and CTNND2^+/+group.3.Detection indicators:?1?Physiological development indicators:body weight,body length and tail length of mice;eyes open,tooth eruption,abdominal hair grows out,body hair grows out,body black hair grows out,auricle separation,open ears,testicular descent positive days.?2?Developmental indicators of the nervous system:pivoting,surface righting reflex,negative geotaxis reflex,cliff aversion,air righting reflex,crawl,front limb suspension,auditory surprise reflex,visual placing reflex,tail pinch reflex positive days,and homing cage emergence time,feet unfolded distance of PN15 d and PN30 d.?3?Behavior test:three chamber sociability test,open-field test,youth play test,Morris water maze test.Results:1.In terms of physiological development,compared with the CTNND2^+/+group,the body weight,body length and tail length of mice in the CTNND2^-/-group were delayed?P<0.05?,and there was no significant difference in other physiological development indicators?P>0.05?.2.In terms of neurological development,compared with the CTNND2^+/+group,the positive days of pivoting,surface righting reflex and auditory surprise reflex in the CTNND2^-/-group were significantly delayed?P<0.05?.Homing cage emergence time also increased significantly?P<0.05?.The feet unfolded distance of PN15 d and PN30 d was significantly shortened?P<0.01?.There was no obvious difference in the other nervous system development indicators?P>0.05?.3.Behavior detection results:?1?In the three chamber sociability test,compared with the CTNND2^+/+group,the CTNND2^-/-group in the first stage had a shorter stay in chamber 1?P<0.05?and a longer stay in chamber 2?P<0.01?.In the second stage,the CTNND2^-/-group stayed in chamber 1 for a longer time?P<0.05?,and had a longer time with stranger1?P<0.05?,and a shorter time with stranger2?P<0.001?.?2?In the open-field test,compared with the CTNND2^+/+group,the CTNND2^-/-group significantly reduced the total number of cross grids?P<0.001?,the number of cross central grid was also significantly reduced?P<0.001?,and the total number of vertical was also decreased significantly?P<0.001?,the number of jump was increased significantly?P<0.01?,and there was no significant difference in the number of defecation and urination?P>0.05?.?3?In the juvenil play test,compared with the CTNND2^+/+group,the social interaction time in the CTNND2^-/-group was significantly reduced?P<0.001?,the self-grooming time was significantly prolonged?P<0.001?,and the digging time was not significantly different?P>0.05?.?4?In the morris water maze test,during the positioning and navigation phase,compared with the CTNND2^+/+group,the CTNND2^-/-group significantly prolonged the escape latency on the 2-3 d?P<0.05?,and the swimming speed of the CTNND2^-/-group was significantly slower throughout the positioning navigation period?P<0.05?.In the period of space exploration,compared with the CTNND2^+/+group,the CTNND2^-/-group showed slower swimming speed?P<0.05?,fewer times of passing through the platform?P<0.01?and shorter time of staying in the quadrant of the platform?P<0.001?.Conclusion:CTNND2 deficient mice showed physical growth retardation in their physiological development,and there were neuroreflexivity disorders in the development of the nervous system,decreased muscle tone in the hind limbs and partial sensory dysfunction,behavioral tests showed impaired social preferences,decreased ability to explore,increased stereotyped behaviors and decreased ability to learn and remember in space,which suggested that CTNND2 deficient mice physical growth retardation,delayed the development of the nervous system and occurred that CTNND2 deficient mice would develop asautism like behavior disorders.Part Two Detection of changes in neuronal dendritesin CTNND2 deficient miceObjective:To explore whether the autistic-like behavior of CTNND2 deficient mice is caused by the abnormality of neuronal dendrites.Methods:1.The mice were divided into CTNND2^-/-group and CTNND2^+/+group,and brain tissues were obtained at PN7 d,PN21 d,PN42 d and PN70 d,respectively.2.Golgi staining was used to detect the length of the dendrites of the pyramidal neurons in the temporal lobe and hippocampal dentate gyrus of mice at the above time points and the number of primary and secondary dendritic branches.Results:1.In mice brain temporal lobe area,compared with CTNND2^+/+group,in the CTNND2^-/-group on PN7 d the mean number of dendrites has no obvious difference?P>0.05?,but the dendrites length increased significantly?P<0.001?;The mean number of dendrites in PN21 d and PN42 d increased significantly?P<0.05?,and the length of dendrites decreased significantly?P<0.05?.There was no significant difference in the average number of PN70 d dendritic branches?P>0.05?,but the length of dendrites was shortened?P<0.05?.2.In the dentate gyrus of mice brain,compared with CTNND2^+/+group,CTNND2^-/-group showed no significant difference in length and average number of branches on PN7 d and PN70 d.?P>0.05?.The mean number and length of dendrites of PN21 d and PN42 d decreased significantly?P<0.001?.Conclusion:CTNND2 deficient mice appeared autism-like behavior may related with the temporal lobe area and dentate gyrus of the hippocampus of the mice brain abnormal length and the number of neuron dendrites.