Objective To investigate the effects of autologous blood withdrawal-reinfusion on the perioperative coagulation function in patients undergoing cardiac surgery with cardiopulmonary bypass (CPB ) . Methods Eighty-four ASA physical status Ⅱ-Ⅳ patients ,without impairment of coagulation function ,scheduled for cardiac surgery with CPB ,were equally and randomly divided into 2 groups using a random number table :autologous blood withdrawal-reinfusion group (group ABWR , n= 44 ) and control group (group C , n= 40 ) . Decreased coagulation function was diagnosed based on the following two criteria :laboratory standard of decreased coagulation function and clinical signs .After anesthesia and before the beginning of operation (T1 ) ,at 5 min after heparin was reversed with protamine (T2 ) ,at the end of operation (T3 ) and at 24 h after the end of operation (T4 ) ,venous blood samples were obtained to measure the blood routine and parameters of coagulation function . Blood routine included the red blood cell (RBC ) , hemoglobin (Hb ) , hematocrit (Hct ) , platelet count , and plasma fibrinogen concentration (Fib) .The parameters of coagulation function included thrombelastography (TEG) variables and prothrombin time (PT ) ,activated partial thromboplastin time (APTT ) ,international normalized ratio (INR ) , and activated clotting time (ACT ) . The volume of intraoperative blood loss , amount of mediastinal drainage at 6 and 24 h after operation , consumption of tranexamic acid and heparin during operation , and consumption of fibrinogen after operation ,and requirement for transfusion of allogeneic RBCs ,fresh frozen plasma (FFP) and platelet during operation and within 24 h after operation were recorded .The development of decreased coagulation function during operation and within 24 h after operation .Results Compared with group C , perioperative consumption of allogeneic RBCs were decreased ,reaction time (R) measured by celite-activated TEG was increased at T3 (P<0.05) ,and no significant change was found in the blood routine index ,incidence of decreased coagulation function ,volume of intraoperative blood loss ,amount of mediastinal drainage ,consumption of tranexamic acid and heparin , and postoperative consumption of fibrinogen in group ABWR ( P> 0.05 ) . Conclusion Autologous blood withdrawal-reinfusion provides similar effects on coagulation function with allogeneic blood transfusion ,and does not increase the development of decreased coagulation function in patients undergoing cardiac surgery with CPB .

Objective To design and construct a new non-fusion soluble expression vector pTIG-mSUMO(small ubiq-uitin-related modifier) using the widely used solubility promoting protein SUMO and based on the translational coupling phenomenon in order to enable the non-fusion soluble expression of the broad-spectrum antiviral protein RA in Escherichia coli by pTIG-mSUMO.Methods The smt3 gene coding for SUMO protein was cloned from yeast genome DNA by PCR. After directed-site silent mutation to eliminate the EcoRⅠsite, the mutant mSUMO was inserted into pET-22b to obtain the translational coupling expression vector pTIG-mSUMO.The RA was subject to PCR amplification and cloned into the pTIG-mSUMO to obtain the expression plasmid pTIG-mSUMO/RA which was supposed to direct the soluble expression of RA by the translational coupling with mSUMO.Results A translational coupling expression vector pTIG-mSUMO which could di-rect/drive the SUMO and heterogonous protein non-fusion expression simultaneously was designed and constructed.The Western blotting result indicated that pTIG-mSUMO could direct the high-level expression of RA, around 40%of which was soluble.Conclusion A translational coupling expression vector pTIG-mSUMO is obtained.After coupling with SUMO, RA is highly expressed in E.coli and both the expression level and solubility are greatly improved.pTIG-mSUMO might contrib-ute to soluble expression of other proteins.

Objective:To investigate the expression and distribution of human dermal papillary fibroblasts (Fp) , reticular fibroblasts (Fr) , and myofibroblasts (MFB) in keloid tissues.Methods:Keloid tissues were collected from 15 outpatients (including 8 males and 7 females) aged 20-50 years, who were diagnosed in the Department of Dermatology, Renmin Hospital of Wuhan University from May to December 2019. Normal skin tissues were taken from 15 age-matched women who underwent mammoplasty, and served as controls. The distribution of fibroblast activation protein (FAP) , CD90 and alpha-smooth muscle actin (α-SMA) was observed in the keloid tissues and normal skin tissues by dual immunofluorescence staining. Furthermore, fibroblasts were isolated from 3 normal skin and 3 keloid tissue samples, and subjected to primary culture. Subsequently, the fibroblasts were treated with 10 ng/ml transforming growth factor-β1 (TGF-β1) for 48 hours in vitro, during which, changes in fibroblast phenotypes were observed in the 2 groups. Fluorescence-based quantitative RT-PCR and Western blot analysis were performed to determine the mRNA and protein expression of FAP, CD90 and α-SMA. Measurement data were compared between 2 groups by using t test. Results:Immunofluorescence staining of the normal skin tissues revealed that FAP +/CD90 - fibroblasts were predominantly distributed in the superficial dermis, FAP -/CD90 + fibroblasts in the deep dermis, and CD90 + cells hardly expressed α-SMA; however, a large number of FAP + fibroblasts and CD90 + fibroblasts were observed in the deep keloid tissues, and many CD90 + fibroblasts also expressed α-SMA. Dual immunofluorescence staining showed that normal tissue-derived fibroblasts hardly expressed α-SMA, and keloid-derived fibroblasts expressed α-SMA. The fluorescence intensity of α-SMA + cells significantly increased in the normal tissue-and keloid-derived fibroblasts after 24-hour treatment with TGF-β1 (21.058 ± 0.709, 27.112 ± 0.097, respectively) compared with that in the corresponding untreated fibroblasts (11.312 ± 0.636, 21.306 ± 0.464, t=22.430, 13.370, respectively, both P < 0.05) . RT-PCR and Western blot analysis showed that the mRNA and protein expression of FAP, CD90 and α-SMA significantly increased in the keloid-derived fibroblasts after 48-hour treatment with TGF-β1 (mRNA: 92.610 ± 3.667, 1.366 ± 0.105, 3.240 ± 0.141; protein: 0.652 ± 0.073, 1.046 ± 0.119, 0.946 ± 0.117, respectively) compared with the untreated keloid-derived fibroblasts (all P < 0.05) . Conclusion:CD90 + Fr aberrantly proliferated in the deep dermis of keloid tissues, suggesting that directional intervention in aberrantly proliferating FAP -/CD90 + Fr in the deep dermis may promote the efficacy for keloids.

Objective To analyze the bio-mechanics when hemiplegics reach with the unaffected upper limb.Methods Thirty post-stroke hemiplegics were selected into the patient group,while 23 healthy counterparts were chosen for the control group.Both groups completed a reaching test of their upper limbs which divided reaching into a moving stage and a holding stage.Surface electromyography (sEMG) data were recorded during the tests along with the degree of torso twist,the range of motion of the shoulder,movement velocity,smoothness of movement and angle divergence collected using a wearable micro-sensor motion capture system.Results For the stroke patients whose dominant upper limb was unaffected,the average root mean square (RMS) signal from the upper trapezius (34.3 μV) and the average torso twist (-1.4°) in the moving phase were significantly larger than among the control subjects (19.7 μV and-2.3°),but their average movement velocity was significantly slower.In the holding phase the average RMS signal from the upper trapezius (55.4 μV) was still significantly higher than in the control group,but their average pectoralis major signal and the integrated EMG ratio of the anterior segments of the deltoid and upper trapezius muscle pairs were significantly lower.For the stroke patients whose dominant upper limbs were affected,in thc moving phase their average signal from the middle segments of the deltoid were significantly greater than those of the controls,but their movement velocity was significantly slower.For the control subjects,in the moving phase the average signal from the upper trapezius on their non-dominant side was significantly higher than that from the dominant upper limb.The integrated EMG ratio from the anterior segments of the deltoid and upper trapezius muscle pairs on that side was smaller throughout the whole reaching movement.Conclusion The bio-mechanical characteristics in reaching of the unaffected upper limbs of stroke patients are not the same as those of the corresponding upper limbs of healthy subjects.It is more reasonable to select the corresponding upper limbs of healthy subjects as controls when a bio-mechanical study of the affected upper limbs is conducted.

Objective To document the kinematics of upper-limb motor dysfunction among hemiplegic stroke survivors.Methods Thirty-nine stroke survivors with hemiplegia were selected as the experimental group,while twenty-five healthy counterparts were chosen as the control group.Reaching movements performed in the sagittal plane were divided into an anteflexion phase and a holding phase.Three-dimensional kinematics data were captured using a micro-sensor motion capture system,and surface electromyograms (sEMGs) were recorded synchronously from the upper trapezius (UT),the anterior (AD) and middle (MD) segments of the deltoid,the biceps brachii (BB) and the triceps brachii (TB).The torso twist (TTD),the range of motion (ROM) of the shoulder,movement velocity (MV),isotonic instability degree (IT) and isometric instability degree (IM) were extracted.Integrated electromyography (iEMG) and work ratios were chosen as indicators to compare the two groups.The experimental group's kinematic indicators were correlated with that group's sEMG parameters.Results The average TTD,IT and IM in the experimental group were significantly larger than those of the control group,while the ROM of the shoulder and the MV were significantly smaller.During the anteflexion phase,the average iEMG from the UT in the experimental group was significantly larger than that of the control group,while the average iEMGs from their AD and TB were significantly smaller;The BB/TB work ratios in the experimental group were significantly greater than those of the conrol group,while the AD/UT and AD/MD ratios were significantly smaller.The results during the holding phase were similar.In the experimental group,torso twist was found to be positively correlated with the iEMG of the UT,and the ROM of the shoulder and movement velocity were also positively correlated with the iEMG of the AD.Conclusions Kinematics variables and sEMG features can be used to evaluate the motor dysfunction of hemiplegic stroke patients' affected upper limbs quantitatively and provide guidance for rehabilitation.

Objective:To study the effect of p38 mitogen-activated protein kinase (MAPK) gene silencing on expression of apoptotic genes and oncogenes in hepatocytes treated with PM 2.5. Methods:From June to September 2019, according to the p38MAPK gene mRNA sequence provided by GenBank, three interfering sequences were designed and synthesized, ligated into PLVX-shRNA2-puro after annealing, and the recombinant lentiviral vector was transfected into L02 hepatocytes. The p38MAPK silencing cells were identified by real-time fluorescent quantitative PCR and western blotting. The normal L02 cells and p38MAPK silencing cells were treated with 50 μg/mL PM 2.5 water soluble solution, 10 μmol/L positive control Cr 6+, and a blank control group was set up, the treatment time was 24 h. The mRNA levels of oncogenes (c-fos, c-myc, k-ras) , tumor suppressor gene (p53) and apoptotic genes (Caspase-3, Caspase-8, Caspase-9) were detected by real-time PCR. The protein levels of oncogenes and apoptotic genes were detected by Western blotting. Results:The expression levels of p38MAPK mRNA and protein in p38MAPK gene silencing cells were significantly lower than those in L02 hepatocytes ( P<0.05) , and the p38MAPK gene silencing cell line was successfully constructed. Compared with the blank control group, the expression levels of the oncogenes c-fos, c-myc, k-ras and the apoptosis genes Caspase-3, Caspase-8 and Caspase-9 increased, the expression level of tumor suppressor gene p53 decreased in the L02 hepatocyte group treated with PM 2.5 water soluble matter, and the differences were statistically significant ( P<0.05) . Compared with the L02 hepatocytes group treated with PM 2.5 water soluble matter, the expression levels of the oncogenes c-fos, c-myc, k-ras and apoptosis genes Caspase-3, Caspase-8 and Caspase-9 decreased, the expression level of tumor suppressor gene p53 increased in the p38MAPK gene silencing cells group treated with PM 2.5 water soluble matter, and the differences were statistically significant ( P<0.05) . Conclusion:PM 2.5 has effects on the expression of oncogenes, tumor suppressor genes and apoptotic genes in L02 hepatocytes, while p38MAPK gene silencing can inhibit the effects of PM 2.5 on L02 hepatocytes.

Objective:To summarize the clinical treatment of complete left bundle branch block (CLBBB) after the transcatheter closure of ventricular septal defect (VSD).Methods:A case series study was conducted on the treatments and outcomes of 25 children with CLBBB after transcatheter VSD closure in Guangdong Provincial People′s Hospital from January 2010 to December 2023.Paired sample t test was used to evaluate the effect of occlude removal. Results:Among the 25 patients, 12 were males (48%), and 13 were females (52%).The age at surgery was 3.18 (2.51-3.86) years, the height before surgery was 95.0 (90.0-97.5) cm, and the weight before surgery was 13 (12-15) kg.Fourteen children were early-onset cases (≤ 1 month), while the other 11 were late-onset cases (> 1 month).The mean follow-up time was (6.63±3.93) years.Of the 14 early-onset cases, 6 children underwent occluder removal within 1 month and restored normal heart rhythm or incomplete right bundle branch block; 4 children underwent occluder removal after 1 month, of whom 2 recovered, 1 remained CLBBB, and 1 had complete atrioventricular block (CAVB); the other 4 children received drug treatment, of whom 2 had normal heart rhythm, 1 had left anterior fascicular block, and 1 died of cardiac shock and heart failure.All the 11 late-onset cases were first treated by drugs, of whom 3 recovered, and the other 8 remained CLBBB.One of the 8 cases received occluder removal at 8 months after surgery and recovered, 1 had CAVB, and the other 6 remained CLBBB.Conclusions:For patients with CLBBB after transcatheter closure of VSD, drug therapy is not always effective, and CLBBB is easy to recur.Therefore, occluder removal is recommended to be done immediately after CLBBB is discovered.Patients with persistent CLBBB should be followed up regularly, and pacemaker implantation may be performed if necessary.

Objective:To study the effect of p38 mitogen-activated protein kinase (MAPK) gene silencing on expression of apoptotic genes and oncogenes in hepatocytes treated with PM 2.5. Methods:From June to September 2019, according to the p38MAPK gene mRNA sequence provided by GenBank, three interfering sequences were designed and synthesized, ligated into PLVX-shRNA2-puro after annealing, and the recombinant lentiviral vector was transfected into L02 hepatocytes. The p38MAPK silencing cells were identified by real-time fluorescent quantitative PCR and western blotting. The normal L02 cells and p38MAPK silencing cells were treated with 50 μg/mL PM 2.5 water soluble solution, 10 μmol/L positive control Cr 6+, and a blank control group was set up, the treatment time was 24 h. The mRNA levels of oncogenes (c-fos, c-myc, k-ras) , tumor suppressor gene (p53) and apoptotic genes (Caspase-3, Caspase-8, Caspase-9) were detected by real-time PCR. The protein levels of oncogenes and apoptotic genes were detected by Western blotting. Results:The expression levels of p38MAPK mRNA and protein in p38MAPK gene silencing cells were significantly lower than those in L02 hepatocytes ( P<0.05) , and the p38MAPK gene silencing cell line was successfully constructed. Compared with the blank control group, the expression levels of the oncogenes c-fos, c-myc, k-ras and the apoptosis genes Caspase-3, Caspase-8 and Caspase-9 increased, the expression level of tumor suppressor gene p53 decreased in the L02 hepatocyte group treated with PM 2.5 water soluble matter, and the differences were statistically significant ( P<0.05) . Compared with the L02 hepatocytes group treated with PM 2.5 water soluble matter, the expression levels of the oncogenes c-fos, c-myc, k-ras and apoptosis genes Caspase-3, Caspase-8 and Caspase-9 decreased, the expression level of tumor suppressor gene p53 increased in the p38MAPK gene silencing cells group treated with PM 2.5 water soluble matter, and the differences were statistically significant ( P<0.05) . Conclusion:PM 2.5 has effects on the expression of oncogenes, tumor suppressor genes and apoptotic genes in L02 hepatocytes, while p38MAPK gene silencing can inhibit the effects of PM 2.5 on L02 hepatocytes.

Objective:To investigate the long-term safety and effectiveness of stent implantation for residual pulmonary artery stenosis after complicated congenital heart disease.Methods:The symptoms, signs, echocardiography, cardiac CT, cardiac catheterization, six-minute walking distance, and BNP of 41 patients diagnosed from January 1996 to January 2020. In this group, 41 patients, 30 males and 11 females, aged 1.3-14.5 years old, mean (6.1±3.6) years old, and weighed 8-43 kg, mean (18.9±9.4)kg, compared the diameter of the target vessel, pressure difference across stenosis, cardiac function before and postoperative follow-up, and evaluated the long-term effect of stent implantation in the treatment of pulmonary artery stenosis.Results:All 41 patients were not lost to follow-up, no death, and there were no serious adverse events such as stent fracture, artery dissection and pulmonary embolism during follow-up. The median follow-up time was 7.1 years (3.1 to 13.8 years). As of January 2023, the echocardiographic results showed that the diameter of the target vessels in 41 patients increased from preoperative (3.9±1.5) mm to (6.0±1.5) mm ( P<0.05), the pressure difference across the stenosis decreased from preoperative (51.4±19.1) mmHg to (33.1±19.7) mmHg (1 mmHg=0.133 kPa, P<0.05); Heart spiral CT showed that the ratio of target vessel diameter to distal vessel diameter increased from preoperative 0.4±0.2 to 0.9±0.3( P<0.05). All patients had no slow growth and development, no recurrent lung infection, 39 patients (95.1%) had gradeⅠcardiac function, and 2 patients (4.9%) had gradeⅡcardiac function.As children in school age, the walking distance of 6 min was 462 to 633 m, mean( 529.9±57.1)m, the respiratory score was 0.5-1, and the lower limb force score was 6-12. There were 5 long-term adverse events, including 4 cases of target vessel restenosis (9.7%), and 1 case (2.4%), two of the patients with restenosis with repeated target vessel stenosis and lateral pulmonary hypertension were surgically intervention: stent removing and pumonary expanding, after 4, 13 years of stent implantation.And the others were still in follow-up, and no further intervention was made. The Cox multivariate survival analysis suggested that right ventricular systolic blood pressure was a risk factor for endpoint events before stent implantation ( P<0.05). Conclusion:The treatment of residual pulmonary artery stenosis after complicated congenital heart disease after percutaneous stent implantation can effectively relieve the right heart pressure overload, improve pulmonary blood flow, stabilize cardiac function, improve the long-term prognosis of patients with complicated congenital heart disease, reduce the chest opening rate of reoperation, and have stable long-term curative effect.

OBJECTIVES: Graves' ophthalmopathy (GO) is a multifactorial disease, and the mechanism of non coding RNA interactions and inflammatory cell infiltration patterns are not fully understood. This study aims to construct a competing endogenous RNA (ceRNA) network for this disease and clarify the infiltration patterns of inflammatory cells in orbital tissue to further explore the pathogenesis of GO. METHODS: The differentially expressed genes were identified using the GEO2R analysis tool. The Kyoto encyclopedia of genes and genomes (KEGG) and gene ontology analysis were used to analyze differential genes. RNA interaction relationships were extracted from the RNA interactome database. Protein-protein interactions were identified using the STRING database and were visualized using Cytoscape. StarBase, miRcode, and DIANA-LncBase Experimental v.2 were used to construct ceRNA networks together with their interacted non-coding RNA. The CIBERSORT algorithm was used to detect the patterns of infiltrating immune cells in GO using R software. RESULTS: A total of 114 differentially expressed genes for GO and 121 pathways were detected using both the KEGG and gene ontology enrichment analysis. Four hub genes (SRSF6, DDX5, HNRNPC,and HNRNPM) were extracted from protein-protein interaction using cytoHubba in Cytoscape, 104 nodes and 142 edges were extracted, and a ceRNA network was identified (MALAT1-MIR21-DDX5). The results of immune cell analysis showed that in GO, the proportions of CD8⁺ T cells and CD4⁺ memory resting T cells were upregulated and downregulated, respectively. The proportion of CD4 memory resting T cells was positively correlated with the expression of MALAT1, MIR21, and DDX5. CONCLUSIONS This study has constructed a ceRNA regulatory network (MALAT1-MIR21-DDX5) in GO orbital tissue, clarifying the downregulation of the proportion of CD4⁺ stationary memory T cells and their positive regulatory relationship with ceRNA components, further revealing the pathogenesis of GO.
Humans ; CD8-Positive T-Lymphocytes ; RNA, Long Noncoding/genetics* ; Algorithms ; CD4-Positive T-Lymphocytes ; Down-Regulation ; Graves Ophthalmopathy/genetics* ; Gene Regulatory Networks ; MicroRNAs/genetics* ; Serine-Arginine Splicing Factors ; Phosphoproteins