Screening carbonyl reductases with the ability to catalyze the reduction of complex carbonyl compounds is of great significance for the biosynthesis of R-tolvaptan(R-TVP). In this study, the target carbonyl reductase in the crude enzyme extract of rabbit liver was separated, purified, and identified by ammonium sulfate precipitation, gel-filtration chromatography, ion exchange chromatography, affinity chromatography, and protein mass spectrometry. With the rabbit liver genome as the template, the gene encoding the carbonyl reductase rlsr5 was amplified by PCR and the recombinant strain was successfully constructed. After RLSR5 was purified by affinity chromatography, its enzymatic properties were characterized. The results indicated that the gene sequence of rlsr5 was 972 bp, encoding a protein with a molecular weight of 40 kDa. RLSR5 was a dimeric protein, and each monomer was composed of a (α/β)₈-barrel structure. RLSR5 could asymmetrically reduce 7-chloro-1-[2-methyl-4-[(2- methylbenzoyl)amino]benzoyl]-5-oxo-2,3,4,5-tetrahydro-1H-1-benzazepine (prochiral ketone, PK) to synthesize R-TVP. The specific activity of the enzyme was 36.64 U/mg, and the optical purity of the product was 99%. This enzyme showcased the optimal performance at pH 6.0 and 30 °C. It was independent of metal ions, with the activity enhanced by Mn²⁺. This study lays a foundation for the biosynthesis of tolvaptan of optical grade.
Animals ; Rabbits ; Alcohol Oxidoreductases/biosynthesis* ; Recombinant Proteins/metabolism* ; Escherichia coli/metabolism* ; Liver/enzymology*

Objective:To determine the risk factors of reoperation and mortality after complete atrioventricular septal defect repair, and to evaluate the medium and long-term prognosis.Methods:From March 2008 to March 2022, a total of 266 children were selected from the Department of Thoracic and Cardiovascular Surgery, Nanjing Children's Hospital, who underwent the complete atrioventricular septal defect repair. Exclusion of children with conotrucal anomaly such as tetralogy of Fallot, transposition of the great arteries, and right ventricular double outlet. Demographic characteristics, surgical data, postoperative follow-up and associated risk factors were analyzed.Results:All the children were repaired with modified single-piece method for the first time, and 26 children were reoperated because of severe left atrioventricular valve regurgitation, left ventricular outflow tract obstruction and atrioventricular block. The 1-year, 3-year and 5-year overall survival rate and freedom from reoperation rate of all children were (98.1±0.8)%, (97.3±1.0)%, (96.2±1.2)%, (96.6±1.1)%, (93.9±1.5)% and (92.2±1.7)%, respectively. A total of 11 (42.3%) early reoperations and 15 (57.7%) late reoperations were performed, of which 1-year, 3-year and 5-year survival rates were (92.3±5.2)%, (82.1±8.3)% and (76.6±9.4)% respectively. Multifactorial analysis showed that age <3 months and left atrioventricular regurgitation >grade 2 at 24 hours postoperatively were independent risk factors for reoperation, whereas age <3 months and experience of reoperation were independent risk factors for death of children.Conclusion:Complete atrial septal defects have excellent surgical outcomes, but some children still require reoperation, and age <3 months and postoperative left atrioventricular valve regurgitation(LAVVR)>2 grades remain important predictors of their surgical prognosis.

Objective: To obtain chimeric antigen receptor macrophages ( CAR-M) targeting HER2 stably transfected． Methods : CAR lentivirus vector targeting HER2 was constructed and infected with human monocytic leukemia cell line (THP-1) ．CAR THP-1 cells with green fluorescent labeling were selected by sorting flow cytometry and continued to be cultured in vitro．The CAR THP-1 cells targeting HER2 were co-cultured with the endometrial cancer cell line Ishikawa with negative and positive HER2 expression，and their targeted phagocytosis of CAR-M to HER2 positive tumor cells was detected by imaging flow cytometry ，and the targeted phagocytosis efficiency of CAR-M to HER2 positive tumor cells was detected by flow cytometry. Results : CAR lentivirus infection with THP- 1 cells was less efficient ; After co-culture with cancer cells，flow cytometry and imaging flow cytometry showed that CAR THP-1 cells had enhanced phagocytosis of HER2 positive Ishikawa cells compared with the empty body group (P＜0. 01) ． Conclusion In this experiment，CAR THP-1 cell line targeting HER2 was established by constructing CAR lentivirus vector and transfecting THP-1 cells ，and it was proved that CAR THP-1 could phagocytize HER2 positive Ishikawa cells through specific targeting.

Objective:To analyze the early and middle term clinical effects of mitral valve repair in children with mitral insufficiency.Methods:From January 2012 to January 2019, a total of 202 cases of children with mitral insufficiency treated by mitral valve repair were selected from the department of cardiothoracic surgery of Nanjing Children's Hospital, patients with atrioventricular septal defect, single ventricle and ischemic mitral regurgitation were excluded. Echocardiography was used to compare the preoperative and postoperative left ventricular function and degree of regurgitation in children to evaluate the early and middle term efficacy of mitral valvuloplasty.Results:There were 5 cases of early death(5/202, 2.5%) and 3 cases of late death(3/202, 1.5%). The mean follow-up time was(19.49±17.48) months(1-68 months). Postoperative echocardiography showed that the left heart function and mitral regurgitation were significantly improved.Conclusion:Mitral valvuloplasty can significantly correct mitral insufficiency in children, and it has satisfactory mid-term efficacy and good clinical value.

Objective:To establish a nomogram prediction model for the prognosis of patients with septic cardiomyopathy (SCM) based on afterload-corrected cardiac performance (ACP), in order to identify septic patients with poor outcomes and treatment.Methods:The data of patients admitted to the department of critical medicine of the Second Affiliated Hospital of Guangzhou Medical University from June 2016 to June 2019 were analyzed. All patients were monitored by pulse indication continuous cardiac output (PiCCO) monitor more than 24 hours and diagnosed as SCM with ACP less than 80%. The predictors of 30-day death risk of SCM patients were screened by univariate Cox regression analysis. Multivariate Cox regression analysis was used to establish the prediction model for 30-day death risk of SCM patients, which was displayed by the nomogram. Finally, the discrimination and calibration of the model were analyzed by receiver operator characteristic curve (ROC curve) and consistency index (C-index).Results:A total of 102 patients with SCM were included and the 30-day mortality was 60.8% (62 cases). Among 102 patients with SCM, 57 patients (55.9%) had mild impairment of cardiac function (60%≤ACP < 80%), and the 30-day mortality was 43.9% (25/57); 39 patients (38.2%) had moderate impairment of cardiac function (40%≤ACP < 60%), and the 30-day mortality was 79.5% (31/39); 6 patients (5.9%) had severe impairment of cardiac function (ACP < 40%), and the 30-day mortality was 100% (6/6). There was significantly difference in mortality among the three groups (χ 2 = 24.156, P < 0.001). The potential risk factors for 30-day death of SCM patients screened by univariate Cox regression analysis were included in multivariate Cox regression analysis. The results showed that the independent risk factors for 30-day death of SCM patients were acute physiology and chronic health evaluation Ⅱ [APACHEⅡ, risk ratio ( HR) = 1.031, 95% confidence interval (95% CI) was 1.002-1.061, P = 0.039], vasoactive inotropic score (VIS, HR = 1.003, 95% CI was 1.001-1.005, P = 0.012), continuous renal replacement therapy (CRRT; HR = 2.106, 95% CI was 1.089-4.072, P = 0.027), and ACP ( HR = 0.952, 95% CI was 0.928-0.977, P < 0.001). The nomogram model was established based on the above independent risk factors and age, and the area under the curve (AUC) was 0.865 (95% CI was 0.795-0.935), P < 0.001; C-index was 0.797 (95% CI was 0.747-0.847), P > 0.05. Conclusions:The nomogram model based on age, APACHEⅡ score, VIS score, CRRT and ACP has a certain clinical reference significance for the prediction of 30-day mortality of SCM patients. The discrimination and calibration are good, however, further verification is needed.

Objective:To investigate the common types, surgical treatment and effects of tracheal stenosis in children.Methods:A total of 23 children with tracheal stenosis in our hospital from December 2017 to August 2020 were retrospectively reviewed, including 14 males and 9 females. The mean age at operation was(8.9±5.8)months(range: 2-3 months) and the mean weight was(6.4±2.3)kg(range: 4.2-10.5 kg). The common types of tracheal stenosis were complete tracheal ring in 9 children, tracheomalacia in 10 and subglottic membranous annular hyperplasia in 4. The type of congenital heart diseases included 10 patients of pulmonary artery sling, 1 of tetralogy of Fallot, 5 of ventricular septal defect, 1 of pulmonary atresia, and 1 of right aortic arch with aberrant left subclavian artery. Slide tracheoplasty was performed in 9 patients, external splint in 8, endotracheal stent in 2 and tracheal dilation in 4. All children were followed up after 1, 3, 6, and 12 months of operation with CT and bronchoscopy.Results:There was 1 death in all 23 patients and the mortality was 4.3%, which died of granulation tissue hyperplasia after slide tracheoplasty. Reoperation was performed in 1 patient with endotracheal stent. All patients were followed for 1 to 24 months. Clinical symptoms of tracheal stenosis disappeared and the results of CT and bronchoscopy were satisfied.Conclusion:Slide tracheoplasty is the effective surgical method for complete trachea ring. 3D printing bioresorbable external splint is a promising method for the treatment of tracheomalacia.

Based on the SWOT analysis, this paper studied the advantages and disadvantages, external opportunities and challenges of foreign affairs work in Chongqing TCM Hospital. It also discusseds and presented the development strategy of hospital foreign affairs work. Chongqing Traditional Chinese Medicine Hospital developed its own advantages and reduces pressures by seizing external opportunities. It made use of external opportunities to overcome shortcomings, solve internal problems, and respond to external challenges. The hospital gradually regulates the foreign affairs work, enhances the international communication capability of TCM, and expands the influence and core competitiveness.

The parachute mitral valve is a rare group of heart valve diseases that are typically characterized by only one papillary muscle.All the chordae tendineaes are connected to the papillary muscle and shaped like a para-chute.It mainly causes mitral stenosis and occasionally leads to mitral regurgitation. Due to the early appearance of clinical symptoms and rapid development of the disease,early accurate diagnosis and surgical treatment are needed. Common surgical treatments are mitral valve repair and mitral valve replacement.Now,the progress of surgical treatment of ＂parachute－like＂ mitral valve malformation was reviewed in order to provide a meaningful reference for its surgical treatment.

OBJECTIVETo compare the efficacy of thoracoscopic surgery versus thoracotomy in repairing esophageal atresia type Ⅲ with tracheoesophageal fistula (EA/TEF) in neonates.

METHODSA retrospective analysis was conducted in 97 neonates who underwent EA/TEF repair between January 2012 and December 2017 in the Affiliated Children's Hospital of Nanjing Medical University, including 75 patients receiving thoracotomy and 22 patients receiving thoracoscopic surgery. The perioperative data and the incidence of early postoperative complications were compared between two groups.

RESULTSThe operations were completed in all patients. One child (4.5%) in thoracoscopic surgery group was converted to thoracotomy with modified Livaditis procedure due to the long distance of two blind ends (>4 cm) and thinner distal end. The operation time was longer in thoracoscopic surgery group[(143±48) min vs. (120±40) min, <0.05], but the postoperative ventilation time was shorter[(55±22) h vs. (65±19) h, <0.05] and the first oral feeding was earlier in thoracoscopic surgery group[(3.2±1.1) d vs (3.9±1.3) d, <0.05]. No statistical difference was observed in the ratio of red blood cell transfusion, length of hospital stay and drainage tube indwelling time between two groups (all >0.05). The incidence of lung complications in thoracotomy group was higher than that in thoracoscopic surgery group (20.0% vs. 9.1%, <0.01), while there were no significant differences in the incidence of other postoperative complications between two groups. There was no death in thoracoscopic surgery group, while 2 patients died in thoracotomy group.

CONCLUSIONSThoracoscopic repair is a preferred surgical procedure for EA/TEF in neonates.