Objective To discover the host factor interacting with severe acute respiratory syndrome coronavirus-2(SARS-CoV-2)papain-like protease(PLpro)and explore the potential mechanism.Methods The second-generation proximity-dependent biotin identification(BioID2)approach combined with mass spectrometry analysis was used to search for the potential host factors.Immunofluorescence and co-immunoprecipitation(Co-IP)assay were used to verify the interactions between DEAD-box helicase 3(DDX3)and PLpro.The influence of PLpro on DDX3-inhibitor of kappa B kinase ε(IKKε)-TANK-binding kinase 1(TBK1)and DDX3-mitochondrial antiviral signaling protein(MAVS)complexes was also investigated by Co-IP.The effect of PLpro on interferon-β(IFN-β)immune pathway and the protease activity on substrates were studied via luciferase activity assay.Results DDX3 could co-locate and interact with PLpro intracellularly.PLpro might possibly inhibit both the formation of DDX3-MAVS complex and the interactions between DDX3-IKK-ε-TBK1.PLpro could negatively regulate type Ⅰ interferon pathway.Overexpression of DDX3 could lead to a significant increase in the cleavage activity of PLpro/PLP-TM that might be significantly decreased in case of inventions with DDX3 expressions.Conclusion DDX3 may be one of the host factors that interact with SARS-CoV-2 PLpro.PLpro negatively regulates IFN-β immune pathway induced by DDX3,which may provide a favorable immune environment for virus replication.

OBJECTIVE: To explore the coagulation deficit and genetic basis for a Chinese pedigree affected with Congenital dysfibrinogenemia (CD). METHODS: Peripheral venous blood samples of the proband and her family members (including 4 individuals from three generations) were subjected to routine blood test and assays of liver and kidney functions and viral hepatitis to exclude related diseases. Clauss method and DFg-PT method were used to determine the fibrinogen activity (Fg:C), and an immunoturbidimetric assay was used to determine the level of fibrinogen antigen (Fg:Ag). All of the exons (22 in total) and their flanking sequences of the FGA, FGB and FGG genes were amplified by PCR and directly sequenced. Variants in the coding regions of the three genes and transcriptional splicing sites were screened by using Mutation Surveyor^TM software. RESULTS: The Clauss method showed that Fg:C was significantly reduced in the proband and her father, whilst her mother and son were normal. With the DFg-PT method, the proband, her parents and son were all within the normal range. The Fg:C/Fg:Ag ratio of the proband and her father was lower than 0.7, whilst her mother and son were above 0.7. No significant change in the prothrombin time, activated partial thromboplastin clotting time and thrombin time was noted. Two genetic variants were detected, which included a homozygous missense variant in the FGA gene [c.991A>G (p.Thr331Ala)], which was predicted to be benign, and a heterozygous missense variant of the γ chain of the FGG gene [c.1211C>G (p.Ser404Phe)], which is located in a conserved region and unreported in the CLINVAR/HGMD/EXAC/1000G databases and literature. CONCLUSION This pedigree has conformed to the autosomal dominant inheritance of CD. The c.1211C>T (p.Ser404Phe) missense variant of the γ chain of the FGG gene probably underlay the pathogenesis of CD in this pedigree. The variant was unreported previously and named as "Fibrinogen Harbin II Ser404Phe".
Female ; Humans ; Afibrinogenemia/congenital* ; East Asian People ; Fibrinogen/genetics* ; Mothers ; Mutation ; Pedigree

Rabdosia serra(R.serra),an important component of Chinese herbal tea,has traditionally been used to treat hepatitis,jaundice,cholecystitis,and colitis.However,the chemical composition of R.serra and its effect against colitis remain unclear.In this study,the chemical composition of the water extract of R.serra was analyzed using ultra performance liquid chromatography coupled with a hybrid linear ion trap quadrupole-orbitrap mass spectrometer(UPLC-LTQ-Orbitrap-MS).A total of 46 compounds,comprising ent-kaurane diterpenoids,flavonoids,phenolic acids,and steroids,were identified in the water extract of R.serra,and the extract could significantly alleviate dextran sulfate sodium salt-induced colitis by improving colon length,upregulating anti-inflammatory factors,downregulating proinflammatory fac-tors,and restoring the balance of T helper 17/T regulatory cells.R.serra also preserved intestinal barrier function by increasing the level of tight junction proteins(zonula occludens 1 and occludin)in mouse colonic tissue.In addition,R.serra modulated the gut microbiota composition by increasing bacterial richness and diversity,increasing the abundance of beneficial bacteria(Muribaculaceae,Bacteroides,Lactobacillus,and Prevotellaceae_UCG-O01),and decreasing the abundance of pathogenic bacteria(Turi-cibacter,Eubacterium_fissicatena_group,and Eubacterium_xylanophilum_group).Gut microbiota depletion by antibiotics further confirmed that R.serra alleviated colitis in a microbiota-dependent manner.Overall,our findings provide chemical and biological evidence for the potential application of R.serra in the management of colitis.

Objective:To investigate endoscopic thyroidectomy in the treatment of thyroid cancer and its effects on patient's quality of life.Methods:A total of 98 patients with thyroid cancer who received treatment in Zhoushan Hospital from March 2019 to March 2021 were included in this study. They were randomly divided into observation and control groups, with 49 patients in each group. Patients in the observation group underwent endoscopic thyroidectomy, and those in the control group underwent traditional surgery. Perioperative indicators and postoperative complications were compared between the two groups. Stress response measured before and 2 months after surgery and quality of life before and 3 months after surgery were compared between the two groups.Results:Operative time in the observation group was significantly longer than that in the control group [(89.82 ± 18.49)] minutes vs. (63.24 ± 17.28) minutes, t = 7.35, P < 0.05]. Intraoperative blood loss and postoperative drainage in the observation group were (10.32 ± 2.28) mL and (29.25 ± 3.17) mL, which were significantly lower than (15.42 ± 3.56) mL and (40.52 ± 4.95) mL in the control group ( t = 8.44, 13.42, both P < 0.05). There was no significant difference in length of hospital stay between the two groups ( P > 0.05). The incidence of postoperative complications in the observation group was significantly lower than that in the control group [4.08% (2/49) vs. 18.37% (9/49), χ2 = 5.01, P < 0.05]. At 2 days after surgery, white blood cell count, fasting plasma glucose, C-reactive protein in the observation group were (5.62 ± 0.41) × 10 9/L, (4.77 ± 0.38) mmol/L, and (13.25 ± 2.79) mg/L, which were significantly lower than (8.71 ± 0.98) × 10 9/L, (5.75 ± 0.45) mmol/L, (20.84 ± 3.86) mg/L in the control group ( t = 20.36, 11.64, 11.26, all P < 0.05). At 3 months after surgery, Karnofsky performance scale score in the observation group was significantly higher than that in the control group [(87.64 ± 4.35) points vs. (81.29 ± 4.02) points, t = 7.58, P < 0.05). Conclusion:Endoscopic thyroidectomy is highly effective on thyroid cancer, has few postoperative complications, produces little effect on stress response, and can improve the quality of life of patients.

Objective:Our study aims to determine histological regression and clinical improvement after long-term antiviral therapy in hepatitis B virus-related cirrhosis patients.Methods:Treatment-na?ve chronic hepatitis B patients with histologically or clinically diagnosed liver cirrhosis were enrolled. Liver biopsies were performed after 5 years entecavir-based antiviral treatment. Patients were followed up every 6 months. Cirrhosis regression was evaluated based on Metavir system and P-I-R score. Clinical improvement was evaluated before and after the long-term treatment. Kruskal Wallis test and Wilcoxon signed-rank test were used for continuous variables, Fisher's exact test was used for categorical variables and multivariate analysis was performed using logistic regression analysis.Results:Totals of 73 patients with HBV-related liver cirrhosis were enrolled. Among them, 30 (41.1%) patients were biopsy proved liver cirrhosis and the remaining 43 (58.9%) cirrhotic patients were diagnosed by clinical features. Based on Metavir system and P-I-R score, 72.6% (53/73) patients attained histological regression. Furthermore, 30.1% (22/73) were defined as significant regression (Metavir decrease ≥2 stage), 42.5% (31/73) were mild regression (Metavir decrease 1 stage or predominantly regressive by P-I-R system if still cirrhosis after treatment) and 27.4% (20/73) were the non-regression. Compared to levels of clinical characteristics at baseline, HBV DNA, ALT, AST, liver stiffness(decreased from 12.7 to 6.4 kPa in significant regression, from 18.1 to 7.3 kPa in mild regression and from 21.4 to 11.2 kPa in non-regression)and Ishak-HAI score significantly decreased after 5 years of anti-HBV treatment, while serum levels of platelets and albumin improved remarkably ( P<0.05). In multivariate analysis, only the pre-treatment liver stiffness level was associated with significant regression ( OR=0.887, 95% CI: 0.802-0.981, P=0.020). Conclusions:After long-term antiviral therapy, patients with HBV-related cirrhosis are easily to attain improvements in clinical parameters, while a certain percentage of these patients still cannot achieve histological reversal.

Objective To investigate the clinical application value of the levels of heparin-binding protein (HBP) in cerebrospinal fluid (CSF) for intracranial infectious diseases. Methods A case-control study was conducted. 150 patients after craniotomy(73 in the postoperative bacterial intracranial infection group, 77 in the postoperative non-infection group) admitted to the Department of Neurology of the People's Hospital of Liaoning Province from December 2016 to May 2018 were collected. At the same time, 46 patients without operation (14 in the non-bacterial intracranial infection group, 32 patients without intracranial infection were selected as control group whose white blood cell count (WBC) values in CSF were all below 10 × 106/L) in the same period were also collected. According to the diagnostic criteria for severe intracranial infection, the patients with bacterial intracranial infection were divided into 26 cases of mild intracranial infection group and 47 cases of severe intracranial infection group. According to the Glasgow Outcome Scale (GOS) score at the time of discharge, the patients were divided into 30 cases of good prognosis group (GOS score 4-5 points) and 43 cases of poor prognosis group (GOS score 1-3 points). The concentrations of HBP in CSF were tested with latex immunoturbidimetry, and the concentrations of procalcitonin(PCT) in cerebrospinal fluid and serum were tested with electrochemiluminescence, and cerebrospinal fluid routine were tested with instrument method, and the concentrations of total protein(TP) in cerebrospinal fluid were tested with turbidimetry. The differences of the laboratory test indicators in each group were statistically analyzed, and the levels of HBP in CSF of patients with different degrees of intracranial infection and different prognosis were compared. Comparison of two independent samples was performed using the Mann-Whitney U test. Results The HBP levels in cerebrospinal fluid were 187.00 (73.00, 635.00) ng/ml, 10.00 (3.50, 32.00) ng/ml, 1.50 (0, 4.00) ng/ml, 3.00 (1.00, 4.00) ng/ml in post-craniotomy bacterial intracranial infection group, uninfected group after craniotomy, non-bacterial intracranial infection group and control group respectively. The cerebrospinal fluid levels of WBC count were 1280.00 (363.00, 4327.00)×106/L, 63.00 (18.50, 300.00)×106/L, 5.00 (3.00, 14.75)×106/L, 3.00 (2.00, 5.75)×106/L. The absolute value of cerebrospinal fluid neutrophils were 1216.00 (225.50, 3895.50)×106/L, 24.00 (2.00, 209.50)×106/L, 1.00 (1.00, 3.00)×106/L, 1.00 (1.00, 1.00)×106/L. The cerebrospinal fluid levels of PCT were 0.16 (0.10, 0.32) ng/ml, 0.09 (0.07, 0.14) ng/ml, 0.07 (0.06, 0.12) ng/ml, 0.07 (0.06, 0.13) ng/ml. The serum levels of PCT were 0.36 (0.15, 1.09) ng/ml, 0.09 (0.04, 0.16) ng/ml, 0.08 (0.04, 0.13) ng/ml, 0.07 (0.03, 0.11) ng/ml. The levels of HBP, WBC, neutrophils, PCT in CSF and serum PCT in the post-craniotomy bacterial intracranial infection group were significantly higher than those in the uninfected group after craniotomy (Z=-9.246,-6.759,-6.741,-4.477,-6.202, P<0.05), non-bacterial intracranial infection group(Z=-5.840,-5.412,-5.259,-2.923,-5.104,P<0.05) and the control group (Z=-7.905,-7.919,-7.335,-4.397,-5.474, P<0.05). There were significant differences in the levels of HBP, WBC and neutrophils in CSF(Z=-3.763,-3.444,-3.041,P<0.05) and no significant differences in CSF and serum PCT (Z=- 0.869, - 1.850, P>0.05)between the uninfected group after craniotomy and the non-bacterial intracranial infection group. There were significant differences in the levels of HBP, WBC and neutrophils in CSF(Z=-4.496,-6.685,-4.842,P<0.05) and no significant differences in CSF and serum PCT(Z=-0.676,-1.303, P>0.05)between the uninfected group after craniotomy and the control group. There were no significant differences in the levels of HBP, PCT in CSF and serum PCT (Z=-0.861,-0.514,-0.273, P>0.05)and significant differences in the levels of WBC and neutrophils in CSF(Z=-2.756,-3.060, P<0.05) between the non-bacterial intracranial infection group and the control group. The levels of HBP in CSF in the severe intracranial infection group were significantly higher than those in the mild intracranial infection group(Z=-6.267, P<0.05). The levels of HBP in CSF in the poor prognosis group were significantly higher than those in the good prognosis group(Z=-7.064, P<0.05). The area under the ROC curve for the diagnosis of bacterial intracranial infection by HBP, WBC, neutrophils, TP, PCT in CSF and PCT in serum was 0.986, 0.987, 0.945, 0.945, 0.770 and 0.914, respectively. The area under the ROC curve for differential diagnosis of bacterial intracranial infection and non-bacterial intracranial infection was 0.994, 0.958, 0.961, 0.929, 0.747 and 0.936, respectively. Conclusions HBP in CSF is an ideal indicator for the diagnosis of bacterial intracranial infection. It is important to distinguish between bacterial intracranial infection and non-bacterial intracranial infection. The extent of increase is related to the severity of infection and prognosis of the disease.

Objective: To analysis the genotype of Japanese encephalitis virus (JEV) in mosquitoes from Shandong province. Methods: Mosquitoes were collected between August and September in Weishan county, Junan county, and Kenli county of Shandong province in 2016. Viruses were isolated by BHK-21 cell and identified by molecular method . Real-Time RT-PCR was conducted to detect the Japanese encephalitis virus carried by the mosquitoes. Results: A total of 8418 mosquitoes divided into 81 pools including 3 species, Culex tritaeniorhynchus, Anopheles sinensis and Armigeres obturbans. Eight Japanese encephalitis viruses were isolated; 23 pools were positive by JEV specific real-time RT-PCR. Phylogenetic analysis on E sequence of JEV showed all JEV strains belonged to genotype Ⅰ JEV, and new strains that were homogenous with previous JEV strains isolated from Shandong. Conclusions Genotype Ⅰ JEV was the dominant genotype in Shandong province.

Objective To explore the application of nanodisc in functional and drug discovery research of G protein-coupled receptor (GPCR).Methods The purified recombinant 5-Hydroxytryptamine 2B receptor (5-HT2BR) was reconstituted into nanodisc complex.Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and size exclution chromatography were performed to evaluate the reconstitution reaction,followed by the use of surface plasmon resonance to validate the ligand-binding activity of 5-HT2BR after reconstitution.Results 5-HT2B R was effectively self-assembled into nanodisc while maintained its binding activity toward the antagonist SB204741.Conclusions The presented study provided potential application of 5-HT2B R-nanodisc for the development of subtype-selective drugs against 5-HT2B R and the fundamental of utilizing nanodisc for GPCR structural and functional studies as well as drug discovery.

Background:Nonvariceal vascular upper gastrointestinal bleeding is a special type of nonvariceal upper gastrointestinal bleeding (NVUGIB) with serious disease course and high mortality rate, which should be paid more attention by clinicians.Aims:To explore the etiological factors and therapeutic strategies of vascular NVUGIB for improving the diagnosis and treatment of the disease.Methods:Clinical data of 111 cases of vascular NVUGIB admitted from Jan.2012 to Dec.2016 at Daping Hospital, the Third Military Medical University were retrospectively analyzed.All cases were diagnosed by gastroscopy, abdominal CT or angiography.Results:One hundred and five patients underwent a gastroscopy within 24 hours of hospital admission.The major causes of bleeding were peptic ulcer involving blood vessels (62.2%) and vascular malformation (22.5%);other causes included Dieulafoy disease, gastrointestinal stromal tumor, malignant tumor, intra-abdominal infection, trauma and gastric angiotelectasis.Seventy-eight patients (70.3%) received endoscopic hemostasis, 19 received conservative medical therapy only, 13 were treated by interventional embolization and 5 underwent surgical operation.Hemostasis was achieved in 96.4% of the patients (107 cases);in four deceased, 3 were failures of endoscopic and interventional therapies.Conclusions:Peptic ulcer and vascular malformation are the major causes of vascular NVUGIB.Endoscopic therapy has generally been recommended as the first-line treatment, however, interventional embolization or surgical operation should be used directly if necessary.

Objective: To investigate the methods for qualitative pathological assessment of dynamic changes in liver fibrosis/cirrhosis after antiviral therapy in patients with chronic hepatitis B (CHB), since antiviral therapy can partially reverse liver fibrosis and cirrhosis caused by hepatitis B and semi-quantitative, rather than qualitative, pathological assessment is often used for the research on liver fibrosis regression. Methods: Previously untreated CHB patients with liver fibrosis and cirrhosis were enrolled, and liver biopsy was performed before treatment and at 78 weeks after the antiviral therapy based on entecavir. The follow-up assessment was performed once every half a year. Based on the proportion of different types of fibrous septum, we put forward the new qualitative criteria called P-I-R classification (predominantly progressive, predominantly regressive, and indeterminate) for evaluating dynamic changes in liver fibrosis. This classification or Ishak fibrosis stage was used to evaluate the change in liver fibrosis after treatment and Ishak liver inflammation score was used to evaluate the change in liver inflammation after treatment. Results: A total of 112 CHB patients who underwent liver biopsy before and after treatment were enrolled, and among these patients, 71 with an Ishak stage of ≥3 and qualified results of live biopsy were included in the final analysis. Based on the P-I-R classification, 58% (41/71) were classified as predominantly progressive, 29% (21/71) were classified as indeterminate, and 13% (9/71) were classified as predominantly regressive; there were no significant differences between the three groups in alanine aminotransferase, aspartate aminotransferase, albumin, HBeAg positive rate, HBV DNA, and liver stiffness (P < 0.05). After treatment, the proportion of predominantly progressive, indeterminate, or predominantly regressive patients changed to 11% (8/71), 11% (8/71), and 78% (55/71), respectively. Among the 35 patients who had no change in Ishak stage after treatment, 72% (25/35) were classified as predominantly regressive and had certain reductions in the Laennec score, percentage of collagen area, and liver stiffness. Conclusion This new P-I-R classification can be used to assess the dynamic changes in liver fibrosis after antiviral therapy in CHB patients.