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 explore the effects of plantamajoside on the proliferation and invasion of human gastric cancer cell line BGC823.Methods BGC823 cells were randomly separated into a control group,a plantamajoside group,an AAV-NC(transfection of empty plasmids packaged with lentivirus)group and a plantamajoside+AAV-HIF-1α(transfection of HIF-1α overexpression plasmid packaged with lentivirus)group.Cell proliferation,inva-sion,apoptosis,the numbers of vascular mimicry(VM)lumens and vascular branches,the expression of prolifera-tion,apoptosis,epithelial mesenchymal transition(EMT)related proteins,HIF-1α/VEGF pathway proteins of cells were all examined.Results Compared with control group,the BGC823 cell viability,colony formation num-ber,invasion number,VM lumen number,vascular branch number,and expression of Ki-67,PCNA,vimentin,MMP9,Snail,VEGFA,VE-cadhering,HIF-1α and VEGF protein were all lower in plantamajoside group(P＜0.05).The apoptosis rate,the cleaved Caspase-3,Bax,and E-cadherin protein expression were significantly increased(P＜0.05).Compared with plantamajoside group,the BGC823 cell viability,colony formation number,invasion number,VM lumen number,vascular branch number,and expression of Ki-67,PCNA,vimentin,MMP9,Snail,VEGFA,VE-cadherin,HIF-1α and VEGF protein were higher in the plantamajoside+AAV-HIF-1α group(P＜0.05).The apoptosis rate,the cleaved caspase-3,Bax,and E-cadherin protein expression were lower(P＜0.05).Conclusions Plantamajoside inhibits proliferation,EMT,invasion,and VM of human gastric cancer cell line and induce its apoptosis.

BACKGROUND: Rheumatoid arthritis (RA), a chronic systemic autoimmune disease, is characterized by synovitis and progressive damage to the bone and cartilage of the joints, leading to disability and reduced quality of life. This study was a randomized clinical trial comparing the outcomes between withdrawal and dose reduction of tofacitinib in patients with RA who achieved sustained disease control. METHODS: The study was designed as a multicenter, open-label, randomized controlled trial. Eligible patients who were taking tofacitinib (5 mg twice daily) and had achieved sustained RA remission or low disease activity (disease activity score in 28 joints [DAS28] ≤3.2) for at least 3 months were enrolled at six centers in Shanghai, China. Patients were randomly assigned (1:1:1) to one of three treatment groups: continuation of tofacitinib (5 mg twice daily); reduction in tofacitinib dose (5 mg daily); and withdrawal of tofacitinib. Efficacy and safety were assessed up to 6 months. RESULTS: Overall, 122 eligible patients were enrolled, with 41 in the continuation group, 42 in the dose-reduction group, and 39 in the withdrawal group. After 6 months, the percentage of patients with a DAS28-erythrocyte sedimentation rate (ESR) of <3.2 was significantly lower in the withdrawal group than that in the reduction and continuation groups (20.5%, 64.3%, and 95.1%, respectively; P  < 0.0001 for both comparisons). The average flare-free time was 5.8 months for the continuation group, 4.7 months for the dose reduction group, and 2.4 months for the withdrawal group. CONCLUSION: Withdrawal of tofacitinib in patients with RA with stable disease control resulted in a rapid and significant loss of efficacy, while standard or reduced doses of tofacitinib maintained a favorable state. TRIAL REGISTRATION Chictr.org, ChiCTR2000039799.
Humans ; Quality of Life ; China ; Arthritis, Rheumatoid/drug therapy* ; Piperidines/therapeutic use* ; Treatment Outcome ; Antirheumatic Agents/therapeutic use* ; Pyrroles/therapeutic use*

Objective: To investigate the effects of Tim-3 on osteoclast-like cell (OLC) formation and bone resorption induced by peripheral blood mononuclear cells (PBMCs). Methods: The expression levels of Tim-3 in of rheumatoid arthritis (RA) patients and healthy controls were detected by flow cytometry. The OLCs were induced by human PBMCs in vitro. The expression levels of tartrate-resistant acid phosphatase (TRAP), cathepsin K (CTSK) and matrix metalloproteinase 9 (MMP9) mRNAs in the formation of OLCs were detected by real-time quantitative PCR. The morphology of OLCs was observed by Wright′s staining and G-actin staining, and the number of OLCs was counted by TRAP staining. The number and area of bone resorption pits in OLCs were detected by the Corning Osteo Assay Surface. Results: The expression levels of Tim-3 in PBMCs of RA patients ([77.31±10.66]%) were significantly higher than that of healthy controls ([51.72±16.69]%, t=7.593, P＜0.01). When PBMCs with different Tim-3 levels were induced into OLCs, the area of bone resorption pits in the high Tim-3 level group ([1.054±0.085] S/mm 2 ) were significantly lower than those in the intermediate Tim-3 level group ([1.889±0.053] S/mm 2 ) and the low Tim-3 level group ([2.763±0.066] S/mm 2 , F=9.318, P＜0.05). Conclusion Tim-3 may negatively regulate the bone resorption of OLCs.

Objective To investigate the clinical effect of caffeic acid tablets on hypocytosis induced by radiotherapy in patients with lung cancer. Methods From January 2017 to October 2015,one hundred and twenty patients with lung cancer treated by radiotherapy in No. 88 Hospital of PLA were selected and randomly divided into the observation group and the control group,60 cases in each group. Patients in the observation group were treated by oral caffeic acid tablets 0. 3 g/time,3 times/d until the end of radiotherapy. The control group was treated with radiotherapy alone and did not take any blood raising drugs. When myelosuppression appeared, recombinant human granulocyte colony-stimulating factor Injection or recombinant human interleukin 11 was given till it turned to normal blood. The incidence rate of bone marrow suppression,number of white blood cells (WBC),platelet (PLT) and the efficacy of radiotherapy were observed. Results The incidence rate of bone marrow suppression of the observation group was 16. 67％( 10/60) ,which was significantly lower than that of the control group ( 63. 33％( 38/60 ) ) . The difference was statistically significant (χ2 = 27. 22, P<0. 05 ); the incidence of bone marrow inhibition in the observation group was 0,while the control group was 15. 00％ (9/60) ,and the difference was statistically significant (χ2=9. 73,P<0. 05) . The number of white blood cells and platelets decreased significantly in the two groups after radiotherapy. The number of leukocytes and platelets in the observation group at 1,2,3 and 4 weeks after radiotherapy were significantly higher than those in the control group,the difference was statistically significant (P<0. 05). The effective rate of radiotherapy was 70. 00％ (42/60) in the observation group and 66. 67％ (40/60) in the control group,with no significant difference (χ2=0. 15,P>0. 05) . Conclusion Caffeic acid tablet is effective in preventing leukocyte and thrombocytopenia caused by bone marrow suppression in patients with lung cancer after radiotherapy.

Objective:To explore the predictive value of serum thyroglobulin autoantibody (TgAb) for recurrence / metastasisin of thyroglobulin (TG)-negative and TgAb-positive patients with differentiated thyroid carcinoma(DTC) after thyroid ablation.Methods:The clinical data of 57 patients with DTC,undergoing complete resection of thyroid tissue,TG negative and TgAb positive,who were reviewed in Liaocheng People's Hospital during April 2013 to April 2015,were selected and divided into recurrence/metastasis group (n=20) and no recurrence/metastasis group (n=37).The TG and TgAb levels in the two groups were detected and compared by the electrochemical luminescence method,the sensitivity,specificity,positive predictive value and negative predictive value of TgAb in the diagnosis of DTC recurrence / metastasis were analyzed;the independent risk factors of DTC recurrence / metastasis were analyzed by Logistic regression.Results:The TgAb levels(72～3850 IU/mL) in the recurrence/metastasis group was higher than that(18～3638 IU/mL) in the no recurrence/metastasis group,the difference was statistically significant (P＜0.05).The sensitivity,specificity,positive predictive value,negative preictive value of TgAb in the diagnosis of recurrence/metastasis of DTC were 85.71％,83.33％,75.00％,90.91％,respectively.Logistic Regression analysis showed that the 100≤ TgAb ＜204 IU/mL,204≤ TgAb≤ 1000IU/mL,and ＞ 1000IU/rnL levels of TgAb were the independent risk factors of rucurrence/metastasis ofDTC (OR=1.267,2.853,6.791,P＜0.05).Conclusion:TgAb can be used as evaluation of an important index of the recurrence/metastasis of patients with DTC when serum thyroglobulin (TG) was negative and TgAb was positive after thyroid ablation.The higher the TgAb levels,the more probability of the recurrence/metastasis.

Objective:To assess the accuracy of the European Organization of Research and Treatment of Cancer (EORTC) Risk Tables in predicting the prognosis of patients with T1 non-muscle-invasive bladder cancer (NMIBC) treated in the Tianjin Medical University Can-cer Institute and Hospital (TMUCIH). The prognostic factors of T1 NMIBC are also explored, and a new risk scoring model suitable for T1 NMIBC is determined. Methods:We retrospectively reviewed the clinicopathologic characteristics of 108 patients with T1 NMIBC who underwent transurethral resections in TMUCIH from January 2011 to June 2013. We scored patients based on the number of ad-verse factors. Afterwards, divided them into different risk groups by the limits determined using receiver operating characteristic curve (ROC) analysis, and created a new risk scoring model. Results:In a group of 108 patients, 90 (83%) were male and 18 were female (17%). The median age was 65 years old (ranging from 24 to 88). Furthermore, 21 patients (19.4%) had a recurrence and 11 cases (10.2%) progressed to muscle-invasive disease. Conclusion:The EORTC cannot accurately predict the recurrence and progressive rate of T1 NMIBC. The most important prognostic factors for recurrence were tumor size and prior recurrence rate. Tumor grade and prior recurrence rate are independent prognostic factors for tumor progression. The new risk scoring model is more accurate in predicting the recurrence risk and progression of T1 NMIBC.

Objective: To evaluate the post-operative mortality of left ventricular end-diastolic pressure (LVEDP) during primary percutaneous coronary intervention (PCI) in patients with acute ST-segment elevation myocardial infarction (STEMI). Methods: We retrospectively analyzed 255 patients with new onset of STEMI who received primary PCI in our hospital and all patients received LVEDP measurement before coronary artery opening. According to LVEDP value, the patients were divided into 2 groups: LVEDP≤14 mmHg group,n=155 and LVEDP>14 mmHg group,n=100. The post-operative mortality up to 6 months was observed, and the effect of LVEDP on death rate was studied by Cox regression analysis. Results: Compared with LVEDP≤14 mmHg group, the patients in LVEDP>14 mmHg group had the 6 months mortality at HR=4.26, 95% CI (1.13-16.08),P=0.03. Relevant study presented that LVEDP was slightly related to LVEF (r=-0.267, P=0.001) and BNP (r=-0.154,P=0.041). Multi-regression analysis indicated that with adjusted LVEF and BNP, LVEDP was the independent predictor for post-operative mortality up to 6 months in acute STEMI patients after PCI. Conclusion: The LVEDP value measured during PCI procedure is the independent predictor for mortality after PCI in patients with new onset of STEMI.

Objective To evaluate the outcome of ST-segment elevation myocardial infarction (STEMI) patients received different reperfusion therapies. Methods The 238 consecutive STEMI patients were enrolled from February 2012 to December 2012. According to the current guideline of PCI and the choice of patients, the patients were divided into the groups of percutaneous coronary intervention (PCI), ifbrinolysis, and conservative medication. The major adverse cardiac events (MACE) was analyzed in a follow up of 6 months. Results (1) The enrolled patients included the 210 patients received PCI (88.2%), 14 patients received fibrinolysis (5.9%) and 14 patients received conservative medication (5.9%).The Median time of D2B was 110minutes.(2) The rate of late stent thrombosis was signiifcant higher in BMS than DES (n=2, 2.8%vs 0, P < 0.05) . (3) The PCI group had a signiifcantly higher incidence of stroke than the ifbrinolysis group and the conservative medication group (1.0%vs 0, P < 0.05;1.0%vs 0, P<0.05). (4) The PCI group had a signiifcantly higher incidence of bleeding compared to the thrombolysis group and the medication group (1.0% vs 0, P < 0.05; 1.0% vs 0%, P < 0.05). Conclusions The majority of STEMI patients received PCI;The D2B time, which was required<90 minutes in guideline of PCI, was found delayed in our study;Compared to ifbrinolysis and conservative medication, PCI showed better clinical outcomes of STEMI patients.

OBJECTIVETo investigate the expressions of miR-101, protein kinase C-α (PKC-α), and cyclooxygenase-2 (COX-2) in gastric cancer (GC) tissue and their correlations.

METHODSRT-qPCR was used to examine miR-101 expression and Western blotting employed to detect PKC-α and COX-2 expressions in 57 cases of gastric cancer tissues and paired normal gastric mucosal tissues.

RESULTSThe gastric cancer tissues showed a significantly lower miR-101 expression (Z=6.102, P<0.05) but significantly higher expressions of COX-2 (Z=14.436, P<0.05) and PKC-α (Z=6.955, P<0.05) than the normal gastric tissues. The expression of COX-2 protein was significantly correlated with the degree of differentiation, invasion depth, lymph node metastasis and TNM stage (P<0.05); PKC-α protein expression was associated with lymph node metastasis and TNM stage (P<0.05). PKC-α expression was positively correlated (r=0.531, P<0.05) and miR-101 expression negatively correlated (r=-0.627, P<0.05) with COX-2 expression in gastric cancer tissues.

CONCLUSIONSmiR-101, PKC-α and COX-2 all play a role in the tumorigenesis and progression of gastric cancer. miR-101 and PKC-α might be new potential therapeutic targets for inhibiting COX-2 in gastric cancer.

Adult ; Aged ; Aged, 80 and over ; Cyclooxygenase 2 ; metabolism ; Female ; Gastric Mucosa ; metabolism ; Humans ; Male ; MicroRNAs ; metabolism ; Middle Aged ; Neoplasm Staging ; Protein Kinase C-alpha ; metabolism ; Stomach Neoplasms ; metabolism ; pathology