Transcriptomics Study Of VPA-induced Autism Model In Rats

Autism is a neurodevelopmental disorder characterized by impairment of communication,social interaction,abnormally restricted and repetitive behavior.These symptoms lasted until adulthood,which cause serious disease burden to the children,families and society.The etiology of autism suggest that the interaction of genetic and environmental factors may be the cause of autism.But so far,the pathogenesis of autism is unknown,which brought great difficulties to the prevention and treatment of this disease.Valproic acid(VPA)is a broad-spectrum anti-epileptic drugs,which can reduce the excitability of neurons to inhibit seizures,VPA exposure during early gestation is associated with higher incidence of autism in the offspring.Embryonic exposure to VPA in rats is a frequently studied model of autism,which reflects the interaction of genes and environment and with good repeatability.In current study,an VPA induced model of autism was established,total RNA was extracted from prefrontal cortex in VPA and control group.Then,Transcriptome data was produced on the platform of Illumina Hiseq 2000,autism specific gene expression profiles was set up,and enrichment pathway analysis was processed to help to explain the molecular mechanism in autism.We hope this research could provid experimental evidence and theoretical support for the clinical treatment.This research included the following contents:1.Establishment of animal model of VPA-induced.An VPA induced model of autism was obtained in offspring of the Female Sprague Dawley rat that received a single intraperitoneal injection of VPA(600mg/kg)on the 12.5 day after conception,Tail appearance,weight,timing of eye opening,swimming performance,olfactory discrimination,negative geotaxis and social interaction was processed to evaluate the model.Compared to control group,VPA rats showed lower body weight,delayer eyes opening,lower swimming performance,delayed nest seeking response mediated by olfactory system,abnormal negative geotaxis and social interaction deficits.Most of these behavioral abnormalities were similar to human autistic symptomes.These results indicate that animal model of autism were successfully replicated.2.Screening out specific gene expression profiles in VPA induced model of autism in rats.Sequenced on the platform of Illumina Hiseq 2000,we obtained 77,979,376,77,979,376,72,038,636 clean reads fragments from the 3 control samples(C1,C2 and C3)respectively,.and corresponding sequencing depth was 7.8 G bp,7.84G bp and 7.2Gbp.From 3 VPA samples(V1,V2 and V3),We obtained 74,108,162,70,693,152 and 76,305,116 clean reads fragments respectively,and corresponding sequencing depth was 7.42G bp,7.06G bp and 7.64G bp.The sample sequencing error rate,the percentage of Q20,Q30 and GC content,meet the quality control requirements of sequencing.We screened 3228 differentially expressed genes,including 1075 up-regulated genes and 2153 down-regulated genes in VPA compared to controls.3.By GO function and KEGG pathway significant enrichment of differentially expressed genes(3228 gene IDs)in prefrontal cortex,we preliminary understand differentially expressed genes main function and involved in nervous system relevant pathways of development which include glutamatergic synapses,GABAergic synaptic pathways etc..,which were related to synapse-related pathways of prefrontal cortex.4.Selection of reference genes in prefrontal cortex and hippocampus in VPA induced rat models of autism.In this study,we chose 9 commonly used reference genes such as Actin,Ywhaz,Hmbs,Ppia,Gapdh,Rps18,Tbp,Hprtl and Rpl13a was analyzed by qRT-PCR.We identified the stability of these reference genes in prefrontal cortex and hippocampus across time points(3wk,5wk and 80d)after VPA exposure.The results suggest that the expression of these 9 reference genes displayed temporal and regional variation after the VPA treatment.