Circular RNA(circRNA),as a kind of non-coding RNA,regulates a variety of biological functions.To explore the effect of circRNA on PEDV replication in the host porcine intestinal epi-thelial cells,this study screened and analyzed the differentially expressed circRNAs by bioinforma-tic software in African Green Monkey renal cells(Vero-E6 cells)infected by porcine epidemic di-arrhea virus(PEDV),the differentially expressed circRNA ssc_circ_PIK3C2A was identified and the secondary structure was analyzed.PCR was used to identify the ssc_circ_PIK3C2A circRNA structure,the model of PEDV-infected IPEC-J2 cells was constructed,the TCID50 test was used to validate the viral titer of PEDV.The expression of circ_PIK3C2A was detected by qRT-PCR in IPEC-J2 infected by PEDV.circ_PIK3C2A qRT-PCR was performed to detect the expression of N gene of PEDV when ssc_circ_PIK3C2A was over-expressed in IPEC-J2 cells.The results showed that ssc_circ_PIK3C2 A is a porcine circular RNA with a typical circular structure,the virus titer of PEDV reached 10-6/mL after PEDV infected IPEC-J2 cells for 48 h,the expression of circ_PIK3C2A increased extremely(P＜0.01)at 6 h after PEDV-infection,with the extension of infec-tion time,its expression gradually decreased,and the expression was the lowest at 24 h,but there was no time-dependent trend.The expression of PEDV N gene decreased significantly when ssc_circ_PIK3C2A was over-expressed in IPEC-J2 cells.In conclusion,when PEDV infects IPEC-J2 cells,the expression of porcine circ_PIK3C2A decreases,and replication of PEDV increases signifi-cantly in IPEC-J2 cells.our result provides a basis for further study of the mechanism of circular RNA on PEDV replication and its physiological activities in host cells in the future.

AIM:This study aims to investigate the mechanism by which Qingfei-Jiedu-Huatan Formula(QJHF)regulates autophagy in alveolar macrophages through mTOR in the treatment of severe pneumonia(SP)in rats.METHODS:Sixty SPF-grade male rats were randomly assigned to six groups:control,model,QJHF,moxifloxacin(MOX),rapamycin(RAPA),and QJHF+RAPA,with ten rats in each group.An SP rat model was established using Klebsiella pneumoniae.After seven days of treatment,changes in IL-33 and IFN-γ levels in bronchoalveolar lavage fluid(BALF)were measured using ELISA.Histopathological alterations in lung tissue were assessed via HE staining,and the autophagy of alveolar macrophages was detected using immunofluorescence co-localization methods.The expression levels of mTOR,beclin-1,and LC3 mRNA in lung tissue were analyzed using qPCR,while Western blot was employed to assess the protein levels of p-mTOR/mTOR,beclin-1,and LC3-II/LC3-I.RESULTS:Compared to the control group,the model group exhibited a deteriorated condition,characterized by alveolar wall rupture and thickening,significant inflammatory cell infiltration in the alveolar cavity,and extensive lung tissue damage(P<0.01).Elevated levels of IL-33 and IFN-γ in BALF were also observed(P<0.01),along with increased colocalization of CD68 and LC3 in immunofluorescence analy-sis.The mTOR mRNA expression in lung tissue decreased(P<0.01),while LC3 and beclin-1 mRNA expressions in-creased(P<0.01).Additionally,the protein expression ratio of p-mTOR/mTOR decreased(P<0.01),whereas LC3-II/LC3-I and beclin-1 protein levels increased(P<0.01).In comparison to the model group,significant improvements were noted after treatment with QJHF and MOX(P<0.01),while RAPA treatment led to a worsening of these indicators(P<0.05).A slight improvement was observed with the QJHF combined with RAPA intervention,though this was not statisti-cally significant.No significant differences were found between the MOX and QJHF groups.However,the QJHF+RAPA group displayed notable improvements in various indicators compared to the RAPA group(P<0.05).CONCLUSION:The QJHF can mitigate the inflammatory response associated with severe pneumonia,potentially by activating mTOR phos-phorylation activity,which in turn inhibits excessive autophagy in alveolar macrophages.

AIM:This study aims to investigate the mechanism by which Qingfei-Jiedu-Huatan Formula(QJHF)regulates autophagy in alveolar macrophages through mTOR in the treatment of severe pneumonia(SP)in rats.METHODS:Sixty SPF-grade male rats were randomly assigned to six groups:control,model,QJHF,moxifloxacin(MOX),rapamycin(RAPA),and QJHF+RAPA,with ten rats in each group.An SP rat model was established using Klebsiella pneumoniae.After seven days of treatment,changes in IL-33 and IFN-γ levels in bronchoalveolar lavage fluid(BALF)were measured using ELISA.Histopathological alterations in lung tissue were assessed via HE staining,and the autophagy of alveolar macrophages was detected using immunofluorescence co-localization methods.The expression levels of mTOR,beclin-1,and LC3 mRNA in lung tissue were analyzed using qPCR,while Western blot was employed to assess the protein levels of p-mTOR/mTOR,beclin-1,and LC3-II/LC3-I.RESULTS:Compared to the control group,the model group exhibited a deteriorated condition,characterized by alveolar wall rupture and thickening,significant inflammatory cell infiltration in the alveolar cavity,and extensive lung tissue damage(P<0.01).Elevated levels of IL-33 and IFN-γ in BALF were also observed(P<0.01),along with increased colocalization of CD68 and LC3 in immunofluorescence analy-sis.The mTOR mRNA expression in lung tissue decreased(P<0.01),while LC3 and beclin-1 mRNA expressions in-creased(P<0.01).Additionally,the protein expression ratio of p-mTOR/mTOR decreased(P<0.01),whereas LC3-II/LC3-I and beclin-1 protein levels increased(P<0.01).In comparison to the model group,significant improvements were noted after treatment with QJHF and MOX(P<0.01),while RAPA treatment led to a worsening of these indicators(P<0.05).A slight improvement was observed with the QJHF combined with RAPA intervention,though this was not statisti-cally significant.No significant differences were found between the MOX and QJHF groups.However,the QJHF+RAPA group displayed notable improvements in various indicators compared to the RAPA group(P<0.05).CONCLUSION:The QJHF can mitigate the inflammatory response associated with severe pneumonia,potentially by activating mTOR phos-phorylation activity,which in turn inhibits excessive autophagy in alveolar macrophages.

Circular RNA(circRNA),as a kind of non-coding RNA,regulates a variety of biological functions.To explore the effect of circRNA on PEDV replication in the host porcine intestinal epi-thelial cells,this study screened and analyzed the differentially expressed circRNAs by bioinforma-tic software in African Green Monkey renal cells(Vero-E6 cells)infected by porcine epidemic di-arrhea virus(PEDV),the differentially expressed circRNA ssc_circ_PIK3C2A was identified and the secondary structure was analyzed.PCR was used to identify the ssc_circ_PIK3C2A circRNA structure,the model of PEDV-infected IPEC-J2 cells was constructed,the TCID50 test was used to validate the viral titer of PEDV.The expression of circ_PIK3C2A was detected by qRT-PCR in IPEC-J2 infected by PEDV.circ_PIK3C2A qRT-PCR was performed to detect the expression of N gene of PEDV when ssc_circ_PIK3C2A was over-expressed in IPEC-J2 cells.The results showed that ssc_circ_PIK3C2 A is a porcine circular RNA with a typical circular structure,the virus titer of PEDV reached 10-6/mL after PEDV infected IPEC-J2 cells for 48 h,the expression of circ_PIK3C2A increased extremely(P＜0.01)at 6 h after PEDV-infection,with the extension of infec-tion time,its expression gradually decreased,and the expression was the lowest at 24 h,but there was no time-dependent trend.The expression of PEDV N gene decreased significantly when ssc_circ_PIK3C2A was over-expressed in IPEC-J2 cells.In conclusion,when PEDV infects IPEC-J2 cells,the expression of porcine circ_PIK3C2A decreases,and replication of PEDV increases signifi-cantly in IPEC-J2 cells.our result provides a basis for further study of the mechanism of circular RNA on PEDV replication and its physiological activities in host cells in the future.

Objective:To examine the clinical efficacy of endovascular treatment on symptomatic occlusion of intracranial vertebral artery (ICVA) in early non-acute stage.Methods:Nine consecutive patients who presented with aggressive ischemic events in the early non-acute stage of ICVA occlusion from January 2014 to December 2019 and received endovascular treatment at Department of Neurosurgery, Peking University First Hospital were retrospectively reviewed.There were 7 males and 2 females, aged 63.4 years old(range: 52 to 72 years).The average preoperative modified Rankin scale(mRS) was 4.3(range: 4 to 5), the National Institute of Health stroke scale(NIHSS) was 12.3(range: 8 to 18). Among them, 2 patients received a single stage endovascular treatment, and the other 7 patients received staged endovascular treatment.The strategy of staged treatment was as follows: firstly, the occlusion part was passed through by a micro-guidewire and dilated with balloons to maintain the blood flow above Thrombolysis In Cerebral Infarction grade 2b. Then, the intravascular large load thrombus was eliminated by the fibrinolytic system and strengthened antiplatelet drugs. After that, a second stage of angioplasty with stenting was performed on the severe residual stenosis part.The complications and the recanalization rate were collected, and the National NIHSS and mRS after endovascular treatment and in follow-up period were recorded.Results:In the 2 cases received single stage endovascular treatment, although revascularization was achieved lastly, one patient suffered embolus translocation and the other suffered re-occlusion after mechanical thrombectomy during the operation, respectively.Technical success was achieved in 6 of the 7 patients received staged endovascular treatment.On discharge, the average NIHSS scores was 5.7(range: 3 to 4) of the patients. Three months after operation,the average mRS was 1.6(range:0 to 3) and it was 0.9(range: 0 to 2) at the latest follow-up, which were better than preoperative status.Conclusions:Staged endovascular treatment might be a safe, efficient, viable option in carefully selected patients with symptomatic ICVA occlusion in early non-acute stage. It needs to be confirmed by further investigation, preferably in a large controlled setting.

Objective:To examine the clinical efficacy of endovascular treatment on symptomatic occlusion of intracranial vertebral artery (ICVA) in early non-acute stage.Methods:Nine consecutive patients who presented with aggressive ischemic events in the early non-acute stage of ICVA occlusion from January 2014 to December 2019 and received endovascular treatment at Department of Neurosurgery, Peking University First Hospital were retrospectively reviewed.There were 7 males and 2 females, aged 63.4 years old(range: 52 to 72 years).The average preoperative modified Rankin scale(mRS) was 4.3(range: 4 to 5), the National Institute of Health stroke scale(NIHSS) was 12.3(range: 8 to 18). Among them, 2 patients received a single stage endovascular treatment, and the other 7 patients received staged endovascular treatment.The strategy of staged treatment was as follows: firstly, the occlusion part was passed through by a micro-guidewire and dilated with balloons to maintain the blood flow above Thrombolysis In Cerebral Infarction grade 2b. Then, the intravascular large load thrombus was eliminated by the fibrinolytic system and strengthened antiplatelet drugs. After that, a second stage of angioplasty with stenting was performed on the severe residual stenosis part.The complications and the recanalization rate were collected, and the National NIHSS and mRS after endovascular treatment and in follow-up period were recorded.Results:In the 2 cases received single stage endovascular treatment, although revascularization was achieved lastly, one patient suffered embolus translocation and the other suffered re-occlusion after mechanical thrombectomy during the operation, respectively.Technical success was achieved in 6 of the 7 patients received staged endovascular treatment.On discharge, the average NIHSS scores was 5.7(range: 3 to 4) of the patients. Three months after operation,the average mRS was 1.6(range:0 to 3) and it was 0.9(range: 0 to 2) at the latest follow-up, which were better than preoperative status.Conclusions:Staged endovascular treatment might be a safe, efficient, viable option in carefully selected patients with symptomatic ICVA occlusion in early non-acute stage. It needs to be confirmed by further investigation, preferably in a large controlled setting.

Objective: To illustrate the epidemical characteristics of the dog-biting events and molecular evolution of rabies virus (RV) strains prevalent in Shuangbai county of Yunnan province, China. Methods: Epidemical investigation on the dog-biting events and human cases were conducted and the brain tissues of the biting dogs and human cases were sampled post-mortem. Nucleoprotein (N) genes of the RVs were sequenced. Homology and phylogenetic analysis were performed using the relevant bioinformatics software. Results: A total of 12 dog-biting events took place between 2011-2017 in Shuangbai county and 35 persons were bitten. Of the 12 events, 11 were investigated in time and 32 bitten persons received proper wound management and a full post-exposure vaccination course. Rabies has not developed in these wounded cases until now. However, due to failure to receive medical intervention and post-exposure treatment in time, 1 of 3 bitten persons in a single event died of rabies. RV N genes from 5 dogs and 1 person were sequenced. Phylogenetic tree showed that RV strains prevalent in Shuangbai county were closely related with the ones found in neighboring counties/cities such as Chuxiong, Lufeng, Jingdong and Xiangyun. All these strains were related to the ones denoted as clade China-I and prevalent in Sichuan province. Homology analysis showed 99.6%-100% homology in nucleotide and amino acid among the 6 RVs prevalent in Shuangbai county and those prevalent in Chuxiong, Lufeng, Xiangyun and clade China-I of Sichuan province. Compared with the China-I strains prevalent in Chuxiong, Zhaotong and Qujing prefectures between 2006-2007, the homology of nucleotide and amino acid were 97.1%-99.3% and 99.1%-99.6%, respectively. Conclusions Surveillance on the dog-biting events can prevent rabies in humans effectively. RV strains prevalent in Shuangbai county belong to clade China-I and have a close relationship with those of neighboring prefectures, cities, counties and the ones prevalent in Sichuan province.

Objective To optimize the fermentation conditions for Enterococcus faecalis (E.faecalis) and Saccharomyces cerevisiae (S.cerevisiae) mixed culture.Methods The optimum fermentation conditions for E.faecalis and S.cerevisiae mixed culture were identified by investigating the influence of initial pH,inoculum size,and ventilation rate of the culture broth on mixed microbial growth.Results The optimal initial pH of mixed microbial culture was 7.0 and the inoculation amounts of E.faecalis and S.cerevisiae were 4％ and 10％ respectively,when fermented in 600-mL pilot fermentor (liquid volume 40％) at 30 ℃,with ventilation rate of 0.2 L/min,for 20 h.At the end of the fermentation,the E.faecalis and S.cerevisiae counts were approximately 3.8 × 108 CFU/mL and 2.4 × 108 CFU/mL,respectively.Ventilation for the amount difference of E.faecalis was no significant (P ＞ 0.05),and for the amount difference of S.cerevisiae was statistically significant (P ＜ 0.05).Conclusion The E.faecalis and S.cerevisiae counts increased by 32.2％ and 31.5％ respectively,when the optimized conditions of fermentation culture were used.In this study,high mixed microbial counts were obtained,thus providing a reference for the preparation of large-scale production of mixed microbes.

Brain injury in preterm infants is an important reason for making the newborn disability.Neonatal cerebral injury of imaging examination method has a time lag.Looking for a simple,timely,accurate predictor of biological markers of brain injury in preterm infants is particularly important.In this paper,the role of S100B protein,glial fibrillary acidic protein and neuron-specific enolase of brain injury in preterm infants was reviewed,and the significance of early diagnosis of brain injury in preterm infants was discussed.

Chromosomal integration of heterologous genes or pathways is preferred over the use of episomal plasmids for its inherently stability and thus more desirable in the industrial setting. However, the position of integration of heterologous genes in the genome influences the expression levels. In combination of high throughput transformation of the Yeast Knock-out Collection (YKO) and FACS analysis, the position effect on heterologous reporter gene gfp was identified across the whole genome in yeast. In total 428 high-expressed sites and 444 low-expressed sites were spotted, providing massive data to analyze patterns and reasons for region dependency of gene expression on the genome-wide scale.
Gene Expression Regulation, Fungal ; Gene Knock-In Techniques ; Genes, Reporter ; Genome, Fungal ; Saccharomyces cerevisiae ; genetics