Objective:To establish a clinical laboratory diagnostic pathway for hepatitis C covering diagnosis, differential diagnosis, drug toxicity monitoring, and therapeutic and prognostic evaluation, and to explore a new teaching model for laboratory diagnostics based on the clinical laboratory diagnostic pathway.Methods:According to the clinical diagnosis and treatment guidelines for hepatitis C, laboratory testing strategies for different stages of diagnosis and treatment of the disease were formulated to establish a clinical laboratory diagnostic pathway for hepatitis C. The pathway was applied in the teaching for undergraduate medical students of the seven-year program of grade 2019 of The First Clinical College of Wuhan University, with those of grade 2018 as the control to receive traditional teaching. The teaching effect was compared through questionnaires and quizzes in class. The data were analyzed through the t test with the use of SPSS 19.0. Results:A clinical laboratory diagnostic pathway for hepatitis C recognized by clinicians was established, covering the entire process of clinical diagnosis, differential diagnosis, monitoring of drug side effects, and therapeutic and prognostic evaluation. The students of grade 2019 receiving the pathway-based teaching model had significant improvements in teaching quality evaluation indicators ( P<0.05), with the most marked improvement in "having mastered the key and difficult points of this lesson", with a score of (60.90±2.15) points for grade 2018 and (84.80±3.44) points for grade 2019. The total score for teaching evaluation was significantly higher in students of grade 2019 than in those of grade 2018 [(94.02±4.29) vs. (79.21±3.68)] points, P<0.05). Grade 2019 also had a significantly higher classroom quiz score than grade 2018 (94.60±5.63) vs. (78.10±4.92), P<0.01]. Conclusions:We established and applied a clinical laboratory diagnostic pathway of hepatitis C in the teaching model of laboratory diagnostics, which organically integrates laboratory diagnostics and clinical medicine, and significantly improves the quality of teaching.

Objective To explore the key active ingredients and action characteristics of Wenxing Jingjintong Gel Patch in the treatment of rheumatoid arthritis(RA)based on the systematic pharmacology and LC-MS/MS technology.Methods The information of active ingredient from Wenxing Jingjintong Gel Patch was established through LC-MS/MS analysis and literature retrieval.The targets of the active ingredients were predicted using Swiss Target Prediction platform and then mapped with the RA-related targets obtained from GeneCards,DrugBank,and OMIM databases to identify the intersecting targets.The"active ingredients-effective targets"network was constructed through the Cytoscape software.The shared targets were imported into STRING database to construct a protein-protein interaction network.GO function and KEGG pathway enrichment analysis were performed using the Metascape database.Molecular docking studies were conducted using AutoDock software to investigate the interactions between key ingredients and target proteins.Results A total of 142 active ingredients were identified in Wenxing Jingjintong Gel Patch by wsing LC-MS/MS,which were further supplemented to 174 through literature retrieval.There were 175 shared targets between the active ingredients and RA.It was anticipated that Wenxing Jingjintong Gel Patch exerted immune regulation and anti-inflammatory and analgesic effects through the interaction between key active ingredients such as berberine,neobavaisoflavone,and palmatine chloride with key targets,including TNF,IL6,and AKT1 to regulate PI3K/Akt1,JAK/STAT,and MAPK signaling pathways.In 1 152 molecular docking validation,94%of them had binding energies less than-5.0 kcal·mol-1,while 51%of them had binding energies less than-7.0 kcal·mol-1.It was indicated that there was a good binding affinity between the potential active ingredients and core targets.Conclusion This study predicted the active ingredients and action characteristics of Wenxing Jingjintong Gel Patch in the treatment of RA,which provided a theoretical basis for further clinical application and quality control.

ObjectiveTo explore the mechanism of Huangqisan （HQS） in regulating autophagy to alleviate hepatic steatosis and improve non-alcoholic fatty liver disease （NAFLD） based on adenosine 5'-monophosphate-activated protein kinase （AMPK）/mammalian target of rapamycin （mTOR） signaling pathway. MethodThe main chemical components and targets of HQS and NAFLD-related targets were collected from database and the intersection targets were used for Gene Ontology （GO） functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis. The protein-protein interaction （PPI） network was constructed， and in vivo experimental verification was conducted. Sixty C57BL/6J male mice were randomly divided into normal control group （NCD）， model group high-fat diet （HFD）， metformin group （MET， 0.25 g·kg^-1）， low-dose Huangqisan group （HQS-L， 0.5 g·kg^-1）， and the high-dose Huangqisan group （HQS-H， 1 g·kg^-1）， with 12 mice in each group after a one-week acclimatization period. NAFLD model was induced by HFD， and intragastric administration was performed at the same time， once a day for 13 weeks. Random blood glucose， serum total cholesterol （TC）， triglyceride （TG）， non-esterified fatty acid （NEFA）， low density lipoprotein-chdesterol （LDL-C） levels， and liver TG content were determined. The liver weight was weighed， and liver index was calculated. Hematoxylin-eosin （HE） staining， oil red O staining， transmission electron microscope （TEM）， real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）， and Western blot were used to verify the effect and reveal the potential mechanism of C57BL/6J mice in vivo. ResultThrough network pharmacology analysis， combined with previous studies， it was predicted that HQS may improve NAFLD by regulating autophagy via the AMPK/mTOR signaling pathway. The result of in vivo experiment showed that， as compared with NCD group， random blood glucose， body weight， serum TC， LDL-C， NEFA， liver weight， liver index， and liver TG content of mice in the HFD groups were significantly increased （P<0.01）. HE staining showed massive lipid droplets （LDs） vacuolated， oil red O staining showed lipid accumulation in liver cells， and no obvious autophagosomes and autolysosome were observed under TEM. The relative mRNA expression of LC3A、LC3B、AMPKα1 and protein expression of AMPK， phosphory phosphorylated（p）-AMPK， and p-AMPK/AMPK were significantly down-regulated （P<0.01）， while the protein expression of microtubule-associated protein 1 light chain 3 （LC3）Ⅱ/Ⅰ and p-mTOR was significantly up-regulated （P<0.01）. As compared with HFD groups， liver weight， serum TG， and NEFA levels in HQS-L and HQS-H groups were significantly deceased （P<0.05， P<0.01）. HE staining and oil red O staining showed the improvement of liver pathological changes after HQS administration. Under TEM， a small amount of autophagosome and autolysosome were observed. Besides， liver index was significantly decreased in the HQS-L group （P<0.01）， and random blood glucose， serum TC level and liver TG content were significantly decreased in the HQS-H group （P<0.05）. The results of Western blot and Real-time PCR showed that the mRNA expression of LC3A and LC3B and the protein expression of LC3Ⅱ/Ⅰ， p-AMPK， and p-AMPK/AMPK were significantly up-regulated （P<0.01）， while the mRNA expressions of p62 and protein expression of p62 and p-mTOR were significantly down-regulated （P<0.05， P<0.01）. ConclusionHQS may promote autophagy and restore autophagy flux via the AMPK/mTOR signaling pathway to alleviate hepatic steatosis improving NAFLD.

Objective:To analyze the risk factors of fatal outcome in patients with severe COVID-19.Methods:The clinical characteristics of 107 patients with severe COVID-19 admitted in Renmin Hospital of Wuhan University from February 12 to March 12, 2020 were retrospectively analyzed. During the hospitalization 49 patients died (fatal group) and 58 patients survived (survival group). The clinical characteristics, baseline laboratory findings were analyzed using R and Python statistical software. The risk factors of fatal outcome in patients with severe COVID-19 were analyzed with multivariate logistic regression.Results:Univariate analysis showed that the two groups had statistically significant differences in age, clinical classification, dry cough, dyspnea and laboratory test indicators ( P<0.05 or <0.01). The random forest model was used to rank the significance of the statistically significant variables in the univariate analysis, and the selected variables were included in the binary logistic regression model. After stepwise regression analysis, the patient’s clinical type, age, neutrophil count, and the proportion of CD3 cells are independent risk factors for death in severe COVID-19 patients. Dry cough is an independent protective factor for the death of severe COVID-19 patients. Conclusion:COVID-19 patients with fatal outcome are more likely to have suppressed immune function, secondary infection and inflammatory factor storm. These factors may work together in severe patients, leading to intractable hypoxemia and multiple organ dysfunction and resulting in fatal outcome of patients. The study indicates that timely intervention and treatment measures against above factors may be effective to save the lives of patients with severe COVID-19.

Objective To analyze the application value and clinical experience of intraoperative ultrasound in microsurgical resection of supratentorial cavernous malformations.Methods From March 2018 to March 2019,31 patients with supratentorial cavernous malformations were operated with assistance of intraoperative ultrasound in our hospital.Intraoperative ultrasound could clearly show the locations and sizes of the lesions,and relations of lesions with surrounding structures to choose appropriate surgical routes.The degrees of resection were judged and complications were recorded.The patients were followed up for 3-6 months,and modified Rankin scale (mRS) was used to evaluate the prognoses.Results In these 31 patients,35 cavernous hemangiomas were diagnosed by MR imaging before surgery;33 lesions were removed surgically,and the other two lesions located in the contralateral side of the operative field were not removed.The accuracy of intraoperative ultrasound guiding the lesions was up to 100％.There were no postoperative infections or deaths.During follow-up,mRS score of 28 patients was 0;the other 3 patients with postoperative neurological impairment recovered partially during follow-up.Conclusion With the help of real-time monitoring of intraoperative ultrasound and location technique of small pieces of hemostatic yam,patients with supratentorial cavernous malformations could acquire satisfactory prognosis through meticulous microsurgery.

Objective: To investigate the resuscitation effect of aortic balloon occlusion (ABO) on the traumatic cardiac arrest (TCA) in swine. Methods: Twenty-seven male domestic swine weighing (32.7±3.8)kg were utilized. After 40% of estimated blood volume was removed within 20 minutes, the animals were subjected to 5 minutes of untreated ventricular fibrillation and then 5 minutes of cardiopulmonary resuscitation. Additionally, fluid resuscitation was initiated coincident with the beginning of cardiopulmonary resuscitation. The animals were randomly divided into model group (n=12) and ABO group (n=15). Once cardiopulmonary resuscitation was implemented, aortic balloon was concurrently inflated to stop the blood flow of descending thoracic aorta at the level of the diaphragm in the ABO group. In the model group, aortic balloon was placed in the same position without inflation. During cardiopulmonary resuscitation, the changes of coronary perfusion pressure (CPP), forehead's regional cerebral oxygen saturation (rSO₂) and pressure of end-tidal CO₂ (PETCO₂) were continuously monitored, and the rate of return of spontaneous circulation (ROSC), duration of cardiopulmonary resuscitation, number of shocks and dose of epinephrine were recorded. At 5 minutes after successful resuscitation, the levels of arterial blood gas, lactate and jugular venous blood oxygen saturation (SjvO₂) were measured. Results: Compared with the model group, the values of CPP, rSO₂ and PETCO₂ during cardiopulmonary resuscitation were significantly increased in the ABO group [CPP: (33.5±5.6)mmHg vs. (23.1±5.2)mmHg at 1 minute, (35.3±6.0)mmHg vs. (26.8±7.4)mmHg at 2 minutes, (36.3±6.3)mmHg vs. (28.2±6.3)mmHg at 3 minutes, (40.1±7.1)mmHg vs. (30.5±6.2)mmHg at 4 minutes, (38.1±7.5)mmHg vs. (29.8±5.3)mmHg at 5 minutes; rSO₂: (45.4±5.2)% vs. (39.2±5.1)% at 1 minute, (47.2±3.6)% vs. (42.0±6.4)% at 2 minutes, (47.7±3.0)% vs. (41.5±5.4)% at 3 minutes, (47.0±2.5)% vs. (42.1±5.9)% at 4 minutes, (47.1±2.0)% vs. (41.5±7.4)% at 5 minutes; PETCO₂: (17.0±3.5)mmHg vs. (12.7±4.2)mmHg at 1 minute, (18.5±3.7)mmHg vs. (14.5±2.7)mmHg at 2 minutes, (20.7±5.3)mmHg vs. (15.5±3.2)mmHg at 3 minutes, (18.7±4.5)mmHg vs. (14.9±3.5)mmHg at 4 minutes, (18.2±3.2)mmHg vs. (14.5±4.2)mmHg at 5 minutes] (all P<0.05). The rate of ROSC was significantly higher in the ABO group than in the model group[100%(15/15) vs. 75%(9/12)] (P<0.05). Additionally, shorter duration of cardiopulmonary resuscitation, less number of shocks and lower doses of epinephrine were observed in the ABO group when compared with the model group[duration of cardiopulmonary resuscitation: 5(5, 5)minutes vs. 5(5, 12.5)minutes, number of shocks: 1(1, 1)times vs. 1(1, 4)times, dose of epinephrine: 0.62(0.62, 0.74)mg vs. 0.64(0.59, 2.59)mg] (all P<0.05). At 5 minutes after resuscitation, the level of arterial lactate was significantly decreased and the value of SjvO₂ was significantly increased in the ABO group compared with the model group[Lactate: (9.6±0.8)mmol/L vs. (10.8±1.4)mmol/L; SjvO₂: (50.0±8.6)% vs. (37.9±16.3)%] (both P<0.05). Conclusions In a swine model of TCA, ABO can increase cardiac and cerebral perfusion during cardiopulmonary resuscitation and improve the efficacy of cardiopulmonary resuscitation. It might provide a novel and effective method for the resuscitation of TCA in the clinical setting.

Objective To evaluate preoperative apparent diffusion coefficient (ADC) and exponential apparent diffusion coefficient (eADC) in diffusion weighted imaging (DWI) on microvascular invasion (MVI) of hepatocellular carcinoma (HCC).Methods We retrospectively reviewed 43 HCC patients with DWI data confirmed by surgery and pathology.ADC and eADC values were measured both in neoplasm and hepatic tissue,the ratio of tumor to liver (T/L) on ADC and eADC was calculated.The correlation between MVI and ADC value,T/L of ADC value,eADC value and T/L of eADC value was analysed,the best cut-off value of variables was analysed by ROC curve,multi factor analysis was carried out by Logistic regression model.Results The ADC value,T/L of ADC,eADC value and T/L of eADC were (1.458 ± 0.444) × 10-3 mm2/s,0.787 ± 0.186,0.289 ± 0.144 and 1.383 (1.179,1.655),respectively.Among them,the positive MVI group were (1.232 ± 0.480) × 10-3 mm2/s,0.683 ± 0.229,0.323 ±0.123 and 1.630(1.387,2.066),respectively.The negative MVI group were(1.545 ±0.404) × 10-3mm2/s,0.844 ±0.149,0.277 ±0.152 and 1.303(1.176,1.545),respectively.There was significant difference in ADC value,T/L of ADC value and T/L of eADC value (t =2.164,2.654,z =-2.058,all P ＜0.05),the area under the ROC curve were (1.085 × 10-3) mm2/s,0.685 and 1.475 using MVI as a diagnostic standard.Multivariate analysis showed that T/L of ADC value was an independent factor affecting MVI of HCC (OR=0.002,95％CI:1.380E-5-0.311,P＜0.05).Conclusions The ADC value in HCC patients with positive MVI is lower than in HCC patients with negative MVI,T/L of eADC value is higher than in HCC patients with negative MVI,and T/L of ADC value is an independent factor predicticing MVI of HCC.

Objective To evaluate the effect of mild hypothermia on inositol-requiring enzyme 1 (IRE1) signaling pathway during myocardial injury after cardiac arrest and resuscitation in swine.Methods Twenty-one healthy male white swine,weighing 33-41 kg,were divided into 3 groups using a random number table method:sham operation group (group S,n =5),cardiac arrest-cardiopulmonary resuscitation group (group CA-CPR,n=8),and mild hypothermia group (group MH,n=8).The model of cardiac arrest and resuscitation was established based on the previously reported method.The catheters placed in the left femoral artery and right internal jugular vein were connected to the PiCCO Monitor system,and another pacing catheter was advanced from the right external jugular vein into the right ventricle.Ventricular fibrillation was induced by using a 1 mA alternating current through the pacing catheter.Once ventricular fibrillation was successfully induced,mechanical ventilation was discontinued for 8 min,and then cardiopulmonary resuscitation was initiated.Epinephrine 20 μg/kg was administered at 2.5 min of resuscitation followed by repetition every 3 min.Defibrillation was delivered at 5 min of resuscitation,and then spontaneous circulation was evaluated.If return of spontaneous circulation was not achieved,cardiopulmonary resuscitation was immediately resumed for 2 min and then defibrillation was delivered again.Mechanical ventilation was continued for 30 h after successful resuscitation.Animals in group S only underwent surgical preparation without experiencing cardiac arrest and resuscitation.At 5 min after successful resuscitation,body temperature was cooled down to 33 ℃ by using a cooling blanket,and then maintained at this level until 24 h after resuscitation,followed by 5 h of re-warming at a rate of 1 ℃/h in group MH.The temperature was maintained at 37.5-38.5 ℃ with the aid of surface cooling blanket in the other two groups.At 1,6,12,24 and 30 h after resuscitation (T1-5),the values of stroke volume (SV) and global ejection fraction (GEF) were recorded,and meanwhile 2 ml of blood samples was obtained via the femoral vein to measure the concentration of serum cardiac troponin Ⅰ (cTnI) (by enzyme-linked immunosorbent assay) and activity of serum creatine kinase-MB (CK-MB) (by immunosuppression).The swine were sacrificed at 30 h after resuscitation,and then myocardial specimens from the left ventricle were obtained for determination of the expression of caspase-3 (by immunohistochemistry),cell apoptosis (by TUNEL),and expression of IRE1 and casepase-12 (by Western blot).Apoptosis index was calculated.Results Compared with group S,SV and GEF were significantly decreased and the serum CK-MB activity was increased at T1-5,the concentration of serum cTnI was increased at T2-5,the expression of IRE1,caspase-12 and caspase-3 in myocardium was up-regulated,and apoptosis index was increased in CA-CPR and MH groups (P＜0.05).Compared with group CA-CPR,the SV and GEF were significantly increased and the concentration of serum cTnI was decreased at T2-5,the activity of serum CK-MB was decreased at T3-5,the expression of IRE1,caspase-12 and caspase-3 in myocardium was down-regulated,and apoptosis index was decreased in group MH (P＜0.05).Conclusion The mechanism by which mild hypothermia mitigates myocardial injury after cardiac arrest and resuscitation may be related to inhibiting IRE1 signaling pathway in swine.

Objective To observe the clinical efficacy and its mechanism of Tongluoyuzufang Ⅰ in patients with diabetic foot (DF).Methods A total of 60 cases with DF was randomly divided into experimental group and control group,30 cases in each group.All patients were given insulin and/or hypoglycemic drugs and other Western medicine comprehensive treatment to control blood sugar steadily.The control group received methylcobalamin treatment,and the experimental group received Tongluoyuzufang Ⅰ therapy treatment based on the methylcobalamin treatment.Both groups were treated for 4 weeks for a course of treatment.The clinical efficacy,ankle-brachial index,nerve conduction velocity,hemorheology,blood lipid,serum cytokines and adverse reactions before and after treatment were compared after 3 courses of treatment.Results After treatment,the total effective rate was 90.0 ％ in the observation group,higher than that in the control group(x2 =5.963,P＜0.05).After treatment,the ankle-brachial index,nerve conduction velocity,hemorheology and blood lipid level in both groups were significantly improved(P＜0.05).After treatment,serum levels of tumor necrosis factor-α(TNF-α) and vascular endothelial growth factor(VEGF) in both groups were significantly decreased,while the levels of anti-thrombin Ⅲ (AT-Ⅲ) has increased(P＜0.05).The improvement of the indexes in the observation group was better than that in the control group(P＜0.05).There were no serious adverse events during treatment.Conclusion Tongluoyuzufang Ⅰ can significantly improve limb ischemia and nerve conduction function in patient with DF.

Objective To study 18F-deoxyglueose positron emission tomography computed tomo graphy(18 FDG PET-CT) in the diagnosis of cervical lymph node(LN) metastasis from rabbit nasopharyngeal VX2 carcinoma.Methods Nasopharyngeal VX2 carcinoma model using 30 rabbits was established. 18 FDG PET-CT,MRI and pathological diagnosis were performed and compared.ResultsFifty-three cervi cal LNs were picked up from resected specimens of 30 rabbits with nasopharyngeal VX2 carcinoma.There were 42 pathologically confirmed positive LNs.Positivity rate was significantly correlated with the volume and the shortest diameter of the LNs (r = 9.18,P =0.007 ; r = 2.77,P = 0.008).The diagnostic sensitivity of PET-CT was 96% (24/25) and 29% (5/17) for LNs with volume >0.5 cm3 and ≤0.5 cm3 ,83% (25/30) and 33% (4/12) for LNs with the shortest diameter ≥0.5 cm and < 0.5 cm,respectively.The diagnostic sensitivity,specificity and accuracy of PET-CT was 69% (29/42) ,100% (11/11) and 95% (40/42) ,com paring with 60% (25/42) ,91% (10/11) and 83% (35/42) of MRl,respectively.The volume measured by PET-CT images was not significantly different from the pathologically measured volume (t =-1.23,P = 0.233) ,while the volume measured by MRI was significantly different from the pathologically measured vol ume (t =-3.99,P = 0.001).Conclusions The sensitivity,specificity and accuracy of PET-CT are better than those of MRl,especially for the cervical lymph nodes with volume >0.5 cm3 or the shortest diameter ≤ 0.5 cm.PET-CT also can be used to detect the smaller metastatic lymph nodes,though the false negative rate is higher.