| Amyotrophic lateral sclerosis(ALS)is a neurodegenerative disease that affects motor neurons(MN)in the spinal cord and brain.It is characterized by degeneration of upper(UMN)and lower(LMN)motor neurons in patients during the course of the disease,which can lead to functional loss of motor neuron in multiple brain areas(including the motor area of the cerebral cortex,the brain stem and the spinal cord),The specific manifestations are muscle weakness,muscle atrophy,respiratory failure,dysphagia,etc.,which eventually lead to paralysis and death of the patient.Studies on cortical dysfunction caused byALShave reported abnormalities of inhibitory synaptic transmission in neurons,but its effect on spinal motor neurons still less be ascertained.The balance between the excitability and inhibition of neurons,as well as the proper functioning of neurotransmission,are vital to neurons.Therefore,to understand how spinal motor neurons are affected during the course of ALS,whether it can be compensated by other ways to change the course of the disease,can provide ideas for the advancement of disease treatment.The Sigmal receptor is a small,highly conserved transmembrane protein located in the subsurface cisternae of the C-terminal of the post-synaptic motor neuron.It is usually enriched specifically in the endoplasmic reticulum membrane that is in contact with mitochondria,that is,the mitochondrial-associated membrane(MAM)area.In animals,Sigmal receptors are selectively and highly expressed in the nervous system,especially in the motor neurons of the spinal cord.Studies have shown that deletion or mutation of the Sigmal receptor can lead to a neurodegenerative disease called ALS 16 or other similar motor neuron defects.In addition,even in cases that do not contain SigmalR mutations,Sigma1R has been found to be involved in the pathogenesis of ALS.We believe that Sigma1R may be involved in the regulation of a series of processes such as neurogenesis and extensively participate in various cellular processes in the course of the nervous system disease.Our previous experiments have confirmed that SigmalR knockout rats showed phenotypes related to nerve injury,and then we isolated the dorsal spinal cord of rats and used RNA-seq technique to study the transcriptome of the control group(wild type group)and gene knockout group(SigmalR Knockout group).Finally,551 genes were up-regulated and 674 genes were down-regulated in Sigma1R-/-.Through the KEGG function enrichment analysis of up-regulated genes after SigmalR knockout,we found that the functions of genes related to Human papillomavirus infection and PI3K-Akt signaling pathway were enriched.Through the analysis of KEGG functional enrichment of down-regulated genes after SigmalR knockout,it was found that the calcium signal pathway and GABAergic synapse and other related functions were enriched.Through the GO functional enrichment analysis of up-regulated genes after SigmalR knockout,we found that ensheathment of neurons and axon ensheathment related functions were enriched.The enrichment of a large number of neurotransmitter and synaptic functions can be detected by GO functional enrichment analysis of down-regulated genes after SigmalR knockout.In the process of researching "the effect of SigmalR knockout on rat spinal cord motor neurons",we used the number of spinal ventral horn motor neurons as an important indicator of the degree of neuronal damage.The results of Nissl staining show that after knocking out the SigmalR gene,the number of rat spinal motor neurons has decreased at the age of six months,and the cell volume has shrunk.This situation will become more and more serious with the increase of age.At the age of one year,the number of neurons was significantly reduced,and the cell body volume of neurons was also significantly reduced,suggesting that the spinal motor neurons of SigmalR gene-deficient rats will undergo progressive degeneration.This phenomenon may be one of the important shreds of evidence that the SigmalR gene is involved in the pathophysiological process of neurodegenerative diseases.The labeling of synaptic vesicle glycoprotein(SV2B)showed that after knocking out the SigmalR gene,the number of excitatory synaptic connections on the soma of rat spinal cord motor neurons was significantly reduced,which may be due to the overall decrease in the number of neurons.At the same time,the weakening of neuronal synapse formation and damage repair function may also cause obstacles to the development and maintenance of neuronal synaptic terminals,so that the number of excitatory synapses cannot be maintained at the original level.Then we used transmission electron microscopy to observe the ultrastructure of neuron synapses.The results showed that after knocking out the SigmalR gene,there was no significant change in the length of the presynaptic active zone in the rat spinal motor neuron synapse,the synaptic gap became wider,and the postsynaptic density was significantly thickened.It suggests that SigmalR knockout has little effect on the ability of neurons to release transmitters,but it will reduce synaptic plasticity.Electrophysiological technology is a direct method to study the physiological activities of neurons.Through the patch-clamp recording of acutely separated spinal cord brain slices,we draw the following conclusions:comparison of SigmalR knockout rats and wild-type rats:(1)The excitability of the motor neurons in the ventral horn of the spinal cord was significantly increased,the resting membrane potential was significantly reduced,the membrane resistance did not change significantly,and the maximum amplitude of the action potential generated after the current stimulation was significantly increased.(2)The amplitude of the action potential is slightly smaller,the peak shape is wider,the time required for repolarization is longer,and the post-hyperpolarization potential is smaller,indicating a slower process of the potassium ion outflow.(3)The frequency of action potentials is generally higher,indicating that Sigma 1R knockout can make neurons more easily excited,and the level of excitement is higher.(4)The frequency of PSC is significantly higher,indicating that there are more vesicles released under normal physiological conditions,suggesting more frequent presynaptic activity in SigmalR knockout neurons;no significant difference in the amplitude of PSC,indicating that there is little difference in the levels of postsynaptic receptors under normal physiological conditions.(5)The mEPSC frequency of SigmalR knockout neurons is slightly higher than that of the wild type,indicating that there are more vesicles released without the action of action potentials,suggesting more frequent presynaptic activity.The mEPSC amplitude increased significantly,indicating that the level of excitatory postsynaptic receptors and the response-ability of neurons increased.The larger current peak area indicates that the post-synaptic signal acceptability is stronger.In addition to the above research,we consolidated and mining the GDC data of patients with hepatocellular carcinoma(HCC),looking for the key lncRNA-miRNA-mRNA network and competitive endogenous RNA(ceRNA).We found that SNHG1 was overexpressed and often amplified in HCC patients.In addition,SNHG1 upregulation was associated with the promotion of several primary biological functions,including cell proliferation,transcription and protein binding.Moreover,we found similar trends of small nucleolar RNA host gene 1(SNHG1),E2F8(E2F transcription factor 8),FANCE(FA complementation group E)and LMNB2(encodes lamin B2))expression.In the SNHG1-associated network,high expression levels of SNHG1(log-rank P value=0.0643),E2F8(log-rank P value=0.000048),FANCE(log-rank P value=0.00125)and LMNB2(log-rank P value=0.0392)were significantly associated with poor survival.Single-cell analysis showed that E2F8 may play an important role in tumorigenesis or cancer development. |
PDF Full Text Request