Objective To investigate the correlation between serum levels of prostaglandin-endoperoxide synthase 2(PTGS2),chitinase-3-like protein 1(CHI3L1)and cognitive impairment caused by cerebrovascular disease.Methods From October 2020 to October 2022,96 inpatients with cerebrovascular diseases admitted to the Third People's Hospital of Chengdu were regarded as the study subjects.The basic clinical data of the patients were recorded,the serum levels of PTGS2 and CHI3L1 were detected by enzyme-linked immunosorbent assay,and these patients were grouped into normal group(n=60)and impaired group(n=36)based on the presence or absence of cognitive impairment.The correlation between serum PTGS2 and CHI3L1 levels and fasting blood glucose(FBG)and homocysteine(Hcy)was analyzed by Pearson method.Logistic regression model was used to determine whether serum PTGS2 and CHI3L1 were independent risk factors for predicting cognitive impairment.Receiver operating characteristic(ROC)curve was drawn,and the predictive value of CHI3L1 and serum PTGS2 expression level in cognitive impairment in patients with cerebrovascular disease was analyzed according to the area under the curve(AUC).Results Compared with the normal group,the levels of serum PTGS2(29.30±9.46 pg/ml vs 17.86±5.40 pg/ml)and CHI3L1(13.04±4.06 pg/ml vs 7.51±2.66 pg/ml)in the disorder group were increased,and the differences were statistically significant(t=7.553,8.065,all P＜0.05).Multivariate Logistic regression analysis showed that FBG(OR=3.612,95%CI:2.324～5.614),Hcy(OR=2.584,95%CI:1.351～4.944),PTGS2(OR=1.964,95%CI:1.194～3.231)and CHI3L1(OR= 1.556,95%CI:1.023～2.367)were independent risk factors of cognitive impairment(all P＜0.05).PTGS2 was positively correlated with FBG and Hcy(r=0.368,0.551,all P＜0.05),and CHI3L1 was positively correlated with FBG and Hcy(r=0.510,0.376,all P＜0.05).The ROC curve showed that the area under curve(AUC)of PTGS2 and CHI3L1 in predicting cognitive impairment was 0.819 and 0.829,respectively.The AUC of the combined prediction of cognitive impairment was 0.902,which was obviously higher than that of the independent prediction of the two(Z =2.089,2.293;P=0.037,0.021),with sensitivity and specificity of 77.78%and 98.33%,respectively.Conclusion PTGS2 and CHI3L1 were highly expressed in the serum of patients with cognitive impairment of cerebrovascular disease,indicating that both were related to cognitive impairment of patients with cerebrovascular disease.

Objective:To discuss the damage effect of vesicular stomatitis virus(VSV)on the vascular endothelial(VE)barrier,and to clarify its mechanism.Methods:The canine kidney cells were used to amplify VSV.The half tissue culture infective dose(TCID50)of VSV was determined using mouse brain endothelial tumor bEnd.3 cells,and subsequent experiment was conducted using 300 times the TCID50.The bEnd.3 cells were divided into infection 0 h group,infection 4 h group,infection 8 h group,and infection 12 h group for VE barrier damage experiments due to VSV infection.The bEnd.3 cells were also divided into control group,infection group,and correction group for experiments to inhibit the VSV replication and restore the VE barrier.The bEnd.3 cells were inoculated into Transwell chambers to construct an in vitro VE barrier model.Cell voltage resistance meter was used to detect the transepithelial resistance(TER)in various groups after the bEnd.3 cells were infected with VSV at different time points;fluorescein isothiocyanate-dextran leakage assay was used to detect the permeability coefficients of the cells in various groups;immunofluorescence staining was used to observe the localization changes of VE-cadherin,β-catenin,and phosphorylated β-catenin(p-β-catenin)in cytoskeleton and adherens junctions(AJs)of the bEnd.3 cells after VSV infection;real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression levels of Wnt and β-catenin mRNA in the cells in various groups;Western blotting method was used to detect the expression levels of Wnt,β-catenin,and p-β-catenin proteins in the cells in various groups.Results:The TCID50 of VSV was 10-4.5·100 μL-1.TheTranswell chamber experiment results showed that compared with infection 0 h group,the TERs in the cells in the other groups were significantly decreased(P＜0.05),and the permeability coefficients were significantly increased(P＜0.05).The immunofluorescence staining results showed that compared with control group,the cytoskeleton of the bEnd.3 cells in infection group was disordered,the cell gaps was increased,the linear index of AJs was significantly decreased(P＜0.05),and β-catenin and p-β-catenin translocated from the cell membrane to the perinuclear area.The RT-qPCR results showed that compared with infection 0 h group,the expression levels of Wnt mRNA in the cells in the other groups were significantly decreased(P＜0.05),while the expression levels of β-catenin mRNA showed no statistically significant difference(P＞0.05).The Western blotting results showed that compared with infection 0 h group,the expression levels of Wnt protein in the cells in the other groups were significantly decreased(P＜0.05),the expression levels of β-catenin showed no statistically significant differences(P＞0.05),and the expression levels of p-β-catenin were significantly increased(P＜0.05).After inhibiting the VSV replication and correcting the low density lipoprotein receptor(LDLR)abnormalities,the Transwell chamber experiment results showed that compared with infection group,the TER in the cells in correction group was significantly increased(P＜0.05),and the permeability coefficient was significantly decreased(P＜0.05).The immunofluorescence staining results showed that compared with infection group,the gaps in the cells in correction group were reduced,and the perinuclear aggregation of β-catenin and p-β-catenin in the cells was restrained.The RT-qPCR results showed that compared with infection group,the expression level of Wnt mRNA in the cells in correction group was significantly increased(P＜0.05).The Western blotting results showed that compared with infection group,the expression level of Wnt protein in the cells in correction group was significantly increased(P＜0.05),the expression level of β-catenin showed no statistically significant difference(P＞0.05),and the expression level of p-P-catenin was significantly decreased(P＜0.05).Conclusion:VSV infection can cause the LDLR inactivation,reduce the expression level of Wnt protein,increase the phosphorylation level of β-catenin and cause its internalization,disrupt the stability of AJs,and ultimately lead to VE barrier damage.

Objective:To establish a Plaque-reduction Neutralization Test (PRNT) for the detection of neutralizing antibody titers of Human Respiratory Syncytial Virus (HRSV) and optimize the conditions for preliminary application.Methods:The CHO expression system was used to produce palivizumab monoclonal antibody (palivizumab) and the influencing factors such as cell type, cell culture duration, fixation and permeabilization protocols, and blocking agents. The reproducibility of the method was verified and its correlation was verified with conventional PRNT. Finally, the optimized PRNT assay was further used to determine neutralizing antibody titers against HRSV subtypes A and B in BALB/c mouse serum (immunized by intramuscular injection of HRSV fusion proteins).Results:Palivizumab was expressed at approximately 50 mg/L. The optimal working conditions for PRNT were as follows: culturing HEp-2 cells for 2 days, fixing with 4% (V/V) paraformaldehyde at room temperature for 15 min followed by 0.2% (V/V) Triton X-100 permeabilization for 15 minutes as the optimal fixation-permeabilization and removing the blocking step. The overall coefficient of variation (CV) for the reproducibility validation of this method was <15%, showing a good linear relationship with the conventional PRNT. The Spearman correlation coefficient r s was 0.983. This method was used to detect neutralizing antibody titers in mouse sera against HRSV subtype A strain long and subtype B strain 9320, and the fusion proteins combined with AlOH and CpG adjuvant induced the highest neutralizing antibody titers in mice. Conclusion:The HRSV neutralizing antibody assay established in this study is rapid, reproducible, high-throughput, and can be used to detect neutralizing antibodies to HRSV subtypes A and B.

Objective To explore the mechanism of microRNA(miRNA)-365a-3p affecting the function of vascular endothelial cells involved in the pathogenesis of preeclampsia(PE).Methods Primary human umbilical vein endothelial cells(HUVECs)were set as a NC group(transfected miR-365a-3p NC),a mimics group(trans-fected miR-365a-3p mimics)and a inhibitor group(transfected miR-365a-3p inhibitor).Logarithmic HUVECs cells were set as the blank group.The cell proliferation,migration and angiogenesis in each group were detected.Dual luciferin assay verified the targeting relationship between miR-365a-3p and downstream gene.The protein expressions of TGF-β1,Smad4 and Smad7 in each group were detected.Results Compared with the blank group and the NC group,the absorbance value and mobility of 24,48 and 72 h were decreased(P＜0.05),the number of tubular structures per field were decreased in the mimics group(P＜0.05),the absorbance value and mobility of 24,48 and 72 h were increased(P＜0.05),and the number of tubular structures per field were increased in the inhibitor group(P＜0.05).Dual luciferin assay showed that Smad7 was a target gene of miR-365a-3p.Compared with the blank group and the NC group,the protein expressions of TGF-β1 and Smad4 in the mimics group were increased(P＜0.05),while the protein expression of Smad7 was decreased(P＜0.05).The protein expression levels of TGF-β1 and Smad4 in the inhibitor group were decreased(P＜0.05),while the protein expression levels of Smad7 were increased(P＜0.05).Conclusion miR-365a-3p may affect the function of vascular endothelial cells by regulating the downstream TGF-β signaling pathway,and thus participate in the pathogenesis of PE.

Objective:To explore the impact of rehabilitation exercise intervention mode based on cardiac function classification on clinical effect and quality of life(QOL)in patients with chronic heart failure(CHF).Methods:A total of 160 CHF patients who visited our hospital from Dec 2021 to Jan 2023 were selected,and 154 cases were fi-nally enrolled.According to the random number table method,patients were divided into study group and control group with 77 cases in each group.Control group received routine nursing program,while the study group received rehabilitation exercise intervention based on cardiac function classification on the basis of control group,both groups were intervened for three months.Clinical total effective rate,and cardiopulmonary function,serum oxidative stress indicators and MLHFQ score before and after intervention were compared between two groups.Results:Total effective rates of study subgroups of class Ⅱ and Ⅲ were significantly higher than those of control group(class Ⅱ:100.00％vs.83.78％;class Ⅲ:97.37％vs.80.00％)(P＜0.05 both).Compared with control subgroup of classⅢ after intervention,there were significant rise in peak VO2[(16.98±2.03)ml·min-1·kg-1 vs.(18.61±2.41)ml·min-1·kg-1],LVEF[(41.73±4.53)％vs.(48.03±5.22)％]and 6MWD[(351.34±61.00)m vs.(391.53±64.42)m](P＜0.01 all);and significant reductions in LVEDd[(57.55±3.91)mm vs.(53.18±3.07)mm],LVESd[(35.90±2.91)mm vs.(30.50±2.67)mm],levels of LPO[(6.00±0.99)mg/L vs.(3.95±0.61)mg/L],MPO[(3.83±0.58)mg/L vs.(2.03±0.28)mg/L],and MLHFQ total score[(57.05±4.57)points vs.(45.29±3.94)points]in study subgroup of class Ⅲ(P=0.001 all).Compared with control subgroup of class Ⅱ after intervention,there were significant rise in peak VO2,LVEF and 6MWD,and significant reductions in LVEDd,LVESd,levels of LPO,MPO and MLHFQ score in study subgroup of class Ⅱ,P＜0.05 or＜0.01.There was no significant difference in the incidence rate of adverse events during follow-up between two groups(3.90％vs.6.49％,P=0.717).Conclusion:Rehabilitation exercise intervention based on cardiac function classifi-cation can significantly improve cardiopulmonary function,inhibit oxidative stress response in vivo and improve quality of life in CHF patients,which is worthy of promotion and application in clinical practice.

Objective:To establish a Plaque-reduction Neutralization Test (PRNT) for the detection of neutralizing antibody titers of Human Respiratory Syncytial Virus (HRSV) and optimize the conditions for preliminary application.Methods:The CHO expression system was used to produce palivizumab monoclonal antibody (palivizumab) and the influencing factors such as cell type, cell culture duration, fixation and permeabilization protocols, and blocking agents. The reproducibility of the method was verified and its correlation was verified with conventional PRNT. Finally, the optimized PRNT assay was further used to determine neutralizing antibody titers against HRSV subtypes A and B in BALB/c mouse serum (immunized by intramuscular injection of HRSV fusion proteins).Results:Palivizumab was expressed at approximately 50 mg/L. The optimal working conditions for PRNT were as follows: culturing HEp-2 cells for 2 days, fixing with 4% (V/V) paraformaldehyde at room temperature for 15 min followed by 0.2% (V/V) Triton X-100 permeabilization for 15 minutes as the optimal fixation-permeabilization and removing the blocking step. The overall coefficient of variation (CV) for the reproducibility validation of this method was <15%, showing a good linear relationship with the conventional PRNT. The Spearman correlation coefficient r s was 0.983. This method was used to detect neutralizing antibody titers in mouse sera against HRSV subtype A strain long and subtype B strain 9320, and the fusion proteins combined with AlOH and CpG adjuvant induced the highest neutralizing antibody titers in mice. Conclusion:The HRSV neutralizing antibody assay established in this study is rapid, reproducible, high-throughput, and can be used to detect neutralizing antibodies to HRSV subtypes A and B.

AIM To study the chemical constituents from flower buds of Magnolia biondii Pamp.and their in vitro acetylcholinesterase inhibitory activities.METHODS The 50% acetone extract from the flower buds of M.biondii was isolated and purified by Diaion HP-20,Toyopearl HW-40C,ODS and semi-preparative HPLC,then the structures of obtained compounds were identified by physicochemical properties and spectral data.The in vitro acetylcholinesterase inhibitory activities of these compounds were determined according to previous method established by research group.RESULTS Seventeen compounds were isolated and identified as crassifolioside(1),magnoloside B(2),rutin(3),isoquercitrin(4),quercetin(5),northalifoline(6),cordysinin B(7),thymidine(8),indazole(9),dihydrodehydrodiconiferyl alcohol(10),aesculetin(11),C-veratroylglycol(12),3,4-dihydroxyphenylethanol(13),3-methoxy-4-hydroxyphenylethanol(14),3,4-dihydroxybenzoic acid(15),2,4,6-trimethoxyphenol(16),syringic acid(17).CONCLUSION Compounds 1-17 are isolated from this plant for the first time,none of which show acetylcholinesterase inhibitory activities at the concentration of 20 μmol/L.

Objective To explore the clinical effect of Liuwei Dihuang decoction on thrombocytopenia(HIT)of liver and kidney yin deficiency in hemodialysis patients.Methods 84 hemodialysis patients with liver and kidney yin deficiency HIT were randomly divided into observation group and control group,with 42 cases in each group.The control group was treated with conventional western medicine,while the observation group was treated with Liuwei Dihuang decoction on the basis of the control group.The scores of liver and kidney yin deficiency syndrome before and after treatment and the curative effect of traditional Chinese medicine syndrome were compared between the two groups,and the platelet count(Plt),P-selectin level,HIT antibody before and after treatment,the platelet recovery time and clinical efficacy and adverse reactions of the two groups were also compared.Results After 1 month of treatment,the scores of liver and kidney yin deficiency syndrome in the two groups were both lower than those before treatment(P<0.05),which in the observation group was lower than that of the control group(P<0.05).The total effective rate of traditional Chinese medicine syndromes in the observation group(97.62%)was higher than that in the control group(P<0.05).After 14 d of treatment,the Plt of the two groups were both higher than those before treatment,while the levels of P-selectin and the positive rate of HIT antibody of the two groups were lower than those before treatment(P<0.05),and the Plt in the observation group was higher than that of the control group,the levels of P-selectin and the positive rate of HIT antibody in the observation group were lower than those in the control group(P<0.05).The platelet recovery time in the observation group was shorter than that in the control group(P<0.05),the total effective rate of clinical treatment in the observation group(95.24%)was higher than that in the control group(P<0.05).There was no significant difference in the incidence of adverse reactions between the two groups during treatment(P>0.05).Conclusion On the basis of conventional western medicine treatment,the addition of Liuwei Dihuang decoction to treat HIT of liver and kidney yin deficiency in hemodialysis patients can improve the clinical efficacy,reduce the clinical symptoms of patients,improve the level of Plt,shorten the recovery time of platelet,and reduce the level of P-selectin and the positive rate of HIT antibody,with good safety,which is worthy of clinical promotion.

Objective:To analyze and summarize the clinical characteristics, endoscopic manifestations, pathology and treatment of gastric adenocarcinoma of the fundic gland(GA-FD)to promote clinicians' understanding of this disease.Methods:The clinical data of 3 patients with GA-FD admitted to Beijing Hospital from May 2020 to January 2022 were retrospectively analyzed.Among them, 2 presented with abdominal distension or abdominal pain.The other case was diagnosed during follow-up endoscopy after gastrectomy for gastric cancer.A diagnosis of GA-FD was made based on the results of white light imaging, magnifying endoscopy, pathology and immunohistochemistry.In addition, we reviewed the literature about GA-FD between January 1990 and April 2021 from both national and international reports of 195 cases.We analyzed and summarized the endoscopic and pathological characteristics of the disease and insights on its diagnosis and treatment.Results:Of all 198 cases, patients had a male/female ratio of 1.6/1.0 and an average age of 65.3 years(range: 39-87 years).79.82%(91/114)of the patients were negative for Helicobacter pylori.Gastroscopy showed that the average diameter was 8.97 mm, and 89.9%(178/198)of the patients had lesions infiltrating into the submucosa.91.98%(149/162)of the patients had gastric chief cells as the predominant cell type, 98.86%(130/140)had no lymphovascular invasion, and 97.6%(162/166)had no recurrence or metastasis.Conclusions:GA-FD is a rare gastric cancer, with low-grade atypia and highly differentiated tissues.The mechanisms of GA-FD are not clear and the disease is more commonly seen in the elderly.There is no obvious correlation between Helicobacter pylori infections and GA-FD.Gastric chief cells make up the predominant cell type.The results of endoscopy, pathology and immunohistochemistry can help confirm the diagnosis.The prognosis of most cases is good.

Neuropathic pain(NP) has similar phenotypes but different sequential neuroinflammatory mechanisms in the pathological process. It is of great significance to inhibit the initiation of neuroinflammation, which has become a new direction of NP treatment and drug development in recent years. Mongolian drug Naru-3 is clinically effective in the treatment of trigeminal neuralgia, sciatica, and other NPs in a short time, but its pharmacodynamic characteristics and mechanism of analgesia are still unclear. In this study, a spinal nerve ligation(SNL) model simulating clinical peripheral nerve injury was established and the efficacy and mechanism of Naru-3 in the treatment of NPs was discussed by means of behavioral detection, side effect evaluation, network analysis, and experimental verification. Pharmacodynamic results showed that Naru-3 increased the basic pain sensitivity threshold(mechanical hyperalgesia and thermal radiation hyperalgesia) in the initiation of SNL in animals and relieved spontaneous pain, however, there was no significant effect on the basic pain sensitivity threshold and motor coordination function of normal animals under physiological and pathological conditions. Meanwhile, the results of primary screening of target tissues showed that Naru-3 inhibited the second phase of injury-induced nociceptive response of formalin test in mice and reduced the expression of inflammatory factors in the spinal cord. Network analysis discovered that Naru-3 had synergy in the treatment of NP, and its mechanism was associated with core targets such as matrix metalloproteinase-9(MMP9) and interleukin-1β(IL-1β). The experiment further took the dorsal root ganglion(DRG) and the stage of patho-logical spinal cord as the research objects, focusing on the core targets of inducing microglial neuroinflammation. By means of Western blot, immunofluorescence, agonists, antagonists, behavior, etc., the mechanism of Naru-3 in exerting NP analgesia may be related to the negative regulation of the MMP9/IL-1β signaling pathway-mediated microglia p38/IL-1β inflammatory loop in the activation phase. The relevant research enriches the biological connotation of Naru-3 in the treatment of NP and provides references for clinical rational drug use.
Rats ; Mice ; Animals ; Matrix Metalloproteinase 9/metabolism* ; Rats, Sprague-Dawley ; Neuroinflammatory Diseases ; Interleukin-1beta/metabolism* ; Spinal Cord/metabolism* ; Signal Transduction ; Hyperalgesia/metabolism* ; Neuralgia/metabolism*