The Expression Feature Of Ubiquitin Protein In Nerve Tissue At Different Age Stages

Objective: Neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD) and amyotrophic lateral sclerosis (ALS) are very common in clinic, most of which are unclear in etiology, complex in pathogenesis and pathology, and vary in clinical manifestations. However, there are two common features of these diseases. The onset stage of these diseases is in old age and its incidence increases with aging; The common pathological hallmark of many neurodegenerative diseases is the presence of ubiquitin-positive, intra or extracellular inclusion bodies in affected regions of the brain. So people pay close attention to the role of the ubiquitin-proteasome system in the neurodegenerative diseases.Ubiquitin-proteasome system (UPS) is the main path of the protein degradation within cells, which can degrade 80% of the intracellular misfolded protein or mutein. Previous studies suggested that disorders of ubiquitin-proteasome degradation pathway can lead to the accumulation of abnormal proteins in cells, leading to cell dysfunction and degeneration. UPS dysfunction can occur in any part of UPS pathway, such as the imbalance in the quantity of ubiquitin, dysfunction of E1, E2 or E3 in ubiquitin-binding process, disorder of proteasome degradation or deubiquitination. Therefore, it is significant to observe the expression of ubiquitin-proteasome system in the areas of liability to neurodegenerative diseases in different age stages.The study for the correlation of ubiquitin protein expression and age is less in the past, and there is no such reaserch in this field in our country. Therefore, this experiment aims to observe the expression and distribution features of the ubiquitin protein in susceptible areas of neurodegenerative diseases in different age stages, approach the correlation of the level of ubiquitin protein and neurodegenerative diseases and thus provide a theoretical basis for understanding age-relateness of neurodegenerative diseases.Methods: 3 healthy ICR mice were used in each group of aged 3 months, 6 months, 9 months and 12 months. After anesthesia, animal tissues were fixated via heart perfusion by 4% paraformaldehyde for 20min. Brain tissue from posterior of olfactory bulb to anterior of optic chiasma, brain tissue from 1mm after optic chiasma to midbrain inferior colliculus were cut coronarily; intumescentia lumbalisa of spinal cord and a part of liver tissue(as control group tissue) were cut. All the tissues cut above were fixated in 4% paraformaldehyde for 48h. Then they were dehydrated by gradient ethanol, transparentized by xylene, embedded by paraffin and made into conventional histological sections futher (5μm thick). 5μm thick paraffin sections were used to Immunohistostain for detection of ubiquitin-protein expression and distribution.True color medical image analysis software (Midia Cybernetic Corpo- ration Image-pro Plus5.0 image analysis system) and manual counting methods were used to capture images from Betz's cell layer of cerebral motor cortex, hippocampal CA1 pyramidal cell layer, the pars compacta of midbrain nigral, spinal cord anterior horn of lumbar intumescentia and the liver tissue, to analysis optical density and to counts ubiquitin-positive cell.Results:1 Mean integral opitcal density (MIOD) of Ubiquitin-positive staining regionThe MIOD values of the ubiquitin-positive region of Betz's cell layer of cerebral motor cortex, hippocampal CA1 pyramidal cell layer, the pars compacta of midbrain nigral and liver tissue showed that there was no significant differencese (P>0.05). The MIOD values of ubiquitin-positive region in spinal cord anterior horn of lumbar intumescentia were decreased with aging (P<0.05). The MIOD value of the 12 months old group was significantly lower than that of 3 months old group (P<0.05), 6 months old group and 9 months old group. 9 months old group was also significantly lower than the 3 months old group (P<0.05).2 Count of ubiquitin-positive cell2.1 The Betz's cell layer of cerebral motor cortexThere was no significant diference in number of ubiquitin-positive cell between the adjacent groups of 3 months old group, 6 months old group, 9 months old group and 12 months old group (P>0.05); 12 months old group was lower than 6 months old group and 3 months old group (P<0.05). 9 months old group was lower than 3 months old group (P<0.05).2.2 The pyramidal cell layer of hippocampal CA1 subfieldComparisons showed significantly and gradually decrease in the number of ubiquitin-positive cell among 3 months old group, 6 months group, 9 months group and 12 months group (P<0.05).2.3 The pars compacta of midbrain nigralThere was no significant diference in number of ubiquitin-positive cell between 6 months old group and 3 months old group (P>0.05). 9 months old group was significantly lower in the number of ubiquitin-positive cell compared with both 6 months old group (P<0.05) and 3 months old group (P<0.05). 12 months old group was significantly lower in the number of ubiquitin-positive cell compared with both 6 months old group (P<0.05) and 3 months old group (P<0.05).2.4 Spinal cord anterior horn of lumbar intumescentiaThere was no significant diference in number of ubiquitin-positive cell between 6 months old group and 3 months old group (P>0.05). 9 months old group was significantly lower in the number of ubiquitin-positive cell compared with both 6 months old group (P<0.05) and 3 months old group (P<0.05). 12 months old group was significantly lower in the number of ubiquitin-positive cell compared with both 6 months old group (P<0.05) and 3 months old group (P<0.05).2.5 Liver tissueThe ubiquitin-positive cell number of each month age group showed no significant difference (P>0.05). Conclusions:The number of ubiquitin-positive cell in the areas of liability to neurodegenerative diseases, such as cerebral motor cortex, hippocampus, midbrain nigral and spinal cord anterior horn of lumbar intumescentia, decreased with aging. The MOID of reservated ubiquitin-positive neuron has no significant difference except the spinal cord anterior horn of lumbar intumescentia.