Research on cellular damages and astrocyte activation after cerebral ischemia and reperfusion / 中国药理学通报

Aim To observe cellular damage and astrocyte activation at different time points of cerebral ischemia and reperfusion. Methods The middle cerebral artery of male SpragueDawley rats was occluded for 90 min followed by different time points of reperfusion. Eighty-five SPF male SD rats were randomly divided into control group (Sham), IR3, 6, 12, 24 and IR48h (MCAO followed by 48 h of reperfusion) group. Cerebral ischemia and reperfusion injury was observed by HE staining, and the structure of astrocytes was estimated with transmission electron microscopy (TEM). GFAP expression was detected by immunofluorescence staining and Western blot. Results Cerebral ischemia following by different time points of reperfusion led to different degrees of cellular damage, which was the most serious at 24 h of reperfusion. TEM showed destruction of astrocytes structure, swollen organelles and broken mitochondrial ridge. After cerebral ischemia-reperfusion, the expression levels of GFAP were significant up-regulated in the ischemic penumbra cortex and the highest was at 48 h of reperfusion, indicating astrocytes were activated. In addition, the results showed the gradual decrease in GFAP expression in the infarct core. Conclusions After cerebral ischemia-reperfusion, cellular damage is aggravated, and astrocytes are gradually activated in the ischemic penumbra. With the extension of reperfusion time, the boundaries of infarct area and ischemic area are gradually clear, and scarring may occur.

Genetic Characteristics of Porcine Hemagglutinating Encephalomyelitis Coronavirus: Identification of Naturally Occurring Mutations Between 1970 and 2015.

Porcine hemagglutinating encephalomyelitis virus (PHEV) is a Betacoronavirus characterized by neurological symptoms and a worldwide prevalence. Although PHEV is one of the earliest discovered porcine coronaviruses, it remains poorly studied. The full-length genome of the earliest PHEV strain collected in 1970 in the United States (PHEV/67 N/US/1970) was determined in October 2020. Using this virus as a prototype, we comparatively analyzed all available PHEV full-length sequences during 1970-2015. In phylogenetic trees based on PHEV full-length or spike glycoprotein open reading frame genomic sequences, PHEV/67 N/US/1970 was sorted into a clade different from that of viruses isolated in the United States in 2015. Intriguingly, United States and Belgium viruses isolated in 2015 and 2005, respectively, revealed multiple deletion mutation patterns compared to the strain PHEV/67 N/US/1970, leading to a truncated or a non-functional NS2A coding region. In addition, the genomic similarity analysis showed a hypervariability of the spike glycoprotein coding region, which can affect at least eight potential linear B cell epitopes located in the spike glycoprotein. This report indicates that PHEVs in the United States underwent a significant genetic drift, which might influence PHEV surveillance in other countries.

The Characteristics of Spike Glycoprotein Gene of Swine Acute Diarrhea Syndrome Coronavirus Strain CH/FJWT/2018 Isolated in China.

Swine acute diarrhea syndrome (SADS) is a highly contagious infectious disease characterized by acute vomiting and watery diarrhea in neonatal piglets. The causative agent for SADS is the swine acute diarrhea syndrome coronavirus (SADS-CoV), an alphacoronavirus in the family Coronaviridae. Currently, SADS-CoV was identified only in Guangdong and Fujian provinces of China, not in any other regions or countries in the world. To explore the genetic diversity of SADS-CoV isolates, herein we comparatively analyzed 44 full-length genomes of viruses isolated in Guangdong and Fujian provinces during 2017-2019. The spike glycoprotein gene of SADS-CoV strain CH/FJWT/2018 isolated in Fujian province is distinct from that of other viral isolates in either spike glycoprotein gene-based phylogenetic analysis or whole genome-based gene similarity analysis. Moreover, at least 7 predicted linear B cell epitopes in the spike glycoprotein of CH/FJWT/2018 would be affected by amino acid variations when compared with a representative virus isolated in Guangdong province. The spike glycoprotein of coronaviruses determines viral host range and tissue tropism during virus infection via specific interactions with the cellular receptor and also plays critical roles in eliciting the production of neutralizing antibodies. Since SADS-CoVs have a broad cell tropism, the results in this report further emphasize that the spike glycoprotein gene is a pivotal target in the surveillance of SADS-CoV.

The spike glycoprotein genes of porcine epidemic diarrhea viruses isolated in China.

The porcine epidemic diarrhea virus (PEDV) causes a highly contagious disease in pigs, which is one of the most devastating viral diseases of swine in the world. In China, PEDV was first confirmed in 1984 and PEDV infections occurred sporadically from 1984 to early 2010. From late 2010 until present, PEDV infections have swept every province or region in China. In this study, we analyzed a total of 186 full-length spike genes and deduced proteins of all available complete genomes of PEDVs isolated in China during 2007-2019. A total of 28 potential recombination events were identified in the spike genes of PEDVs in China. Spike gene recombination not only expanded the genetic diversity of PEDVs in the GII genogroup, but also resulted in the emergence of a new evolutional branch GI-c during 2016-2018. In addition, comparative analysis of spike proteins between GI-a prototype virulent CV777 and GII strain AJ1102 reveals that the amino acid variations could affect 20 potential linear B cell epitopes, demonstrating a dramatic antigen drift in the spike protein. These results provide a thorough view of the information about the genetic and antigenic diversity of PEDVs circulating in China and therefore could benefit the development of suitable strategies for disease control.

Genetic drift of MERS-CoV in Saudi Arabia during 2012-2019.

Middle East respiratory syndrome (MERS) is caused by MERS-CoV that infects both human and camel. Camel is supposed to be the natural reservoir for human infection while the sources for most of the primary human infection cases are still not known. We identified two conserved pyrimidine nucleotides that flank UAAU element in MERS-CoV 5'-UTR. These conserved pyrimidine nucleotides distinguish MERS-CoVs into 3 types, that is, UUAAUU, CUAAUU and CUAAUC (referred to as U----U, C----U, and C----C types, respectively). Human MERS-CoV displays a genetic drift from U----U, C----U, to C----C from 2012 to 2019. Camel virus displays a genetic drift from U----U to C----U with a time lag when compared with human virus. The discrepancy in genetic drift seems not to support the notion that camel serves as the only natural reservoir for human infection.

RNA Helicase A Regulates the Replication of RNA Viruses.

The RNA helicase A (RHA) is a member of DExH-box helicases and characterized by two double-stranded RNA binding domains at the N-terminus. RHA unwinds double-stranded RNA in vitro and is involved in RNA metabolisms in the cell. RHA is also hijacked by a variety of RNA viruses to facilitate virus replication. Herein, this review will provide an overview of the role of RHA in the replication of RNA viruses.

Mechanism of Kaixinsan on Learning and Memory Ability and Synaptic Function of APP/PS1 Mice / 中国实验方剂学杂志

Objective:To explore the underlying protective mechanism of Kaixinsan on learning， memory， and synaptic function in APP/PS1 mice. Method:Sixty APP/PS1 mice were randomly divided into a model group， a donepezil （2 mg·kg^-1·d^-1） group， and low- （0.7 g·kg^-1·d^-1）， medium- （1.4 g·kg^-1·d^-1）， and high-dose （2.8 g·kg^-1·d^-1） Kaixinsan groups， and the wild-type mice of the same age in the same litter were assigned to the normal group， with 12 mice in each group. After continuous intragastric administration for two months， the Morris water maze experiment was performed. The ultrastructure of hippocampal neurons was observed by transmission electron microscopy. The colorimetric assay was used to detect serum content of acetylcholine （ACh）， choline acetyltransferase （ChAT）， acetylcholinesterase （AChE）， and levels of hippocampal reactive oxygen species （ROS）， malondialdehyde （MDA）， superoxide dismutase （SOD）， and glutathione peroxidase （GSH-Px）. Real-time fluorescence-based quantitative polymerase chain reaction （Real- time PCR） was used to detect the mRNA expression of hippocampal brain-derived neurotrophic factor （BDNF）， beta-nerve growth factor （NGFB）， discs large homolog (DLG)2， DLG4， and synaptophysin （SYP）. Result:Compared with the normal group， the model group showed prolonged escape latency， reduced number of crossing platforms， shortened stay in the target quadrant （<italic>P</italic><0.01）， decreased number of mitochondria with different shapes and irregular arrangement， some swollen and deformed mitochondria with broken mitochondrial cristae， endolysis， and cytoplasm vacuole， and more cell debris. Additionally， the model group also displayed reduced serum levels of ACh and ChAT， increased AChE （<italic>P</italic><0.01）， elevated hippocampal ROS and MDA （<italic>P</italic><0.05，<italic>P</italic><0.01）， declining SOD and GSH-Px （<italic>P</italic><0.01）， and diminished hippocampal BDNF， NGFB， DLG2， DLG4， and SYP mRNA levels （<italic>P</italic><0.05，<italic>P</italic><0.01）. Compared with the model group， the donepezil group， and the medium- and high-dose Kaixinsan groups showed shortened escape latency， increased number of crossing platforms， prolonged stay in the target quadrant （<italic>P</italic><0.05，<italic>P</italic><0.01）， improved mitochondrial damage with a regular shape （mainly oval shape）， relieved mitochondrial swelling and deformation， and clear mitochondrial cristae. Furthermore， the donepezil group， and the medium- and high-dose Kaixinsan groups also exhibited increased serum ACh and ChAT levels （<italic>P</italic><0.05，<italic>P</italic><0.01）， blunted AChE activity （<italic>P</italic><0.05）， reduced hippocampal ROS level （<italic>P</italic><0.05，<italic>P</italic><0.01）， declining MDA level （<italic>P</italic><0.05）， potentiated SOD and GSH-Px activities， and up-regulated hippocampal BDNF， NGFB， DLG2， DLG4， and SYP mRNA levels （<italic>P</italic><0.05，<italic>P</italic><0.01）. In the low-dose Kaixinsan group， the stay time in the target quadrant was prolonged and the expression of hippocampal SYP mRNA was elevated significantly （<italic>P</italic><0.05）. There was no statistical difference in swimming speed between the groups. Conclusion:Kaixinsan can improve the learning and memory ability of APP/PS1 mice by increasing the expression of synaptic plasticity-related proteins， reducing the ultrastructural damage to hippocampal neurons， resisting oxidative stress， and regulating cholinergic neurotransmitters， thereby exerting neuroprotective effects.