Objective To construct an eukaryotic expression vetor of ring finger protein 6 (RNF6)and to investigate its ubiqu-itylation on insulin receptor substrate-1(IRS-1).Methods Human RNF6 coding sequence was amplified by polymerase chain reac-tion (PCR)method with human cDNA as template.The pcDNA3.1-CHA-RNF6 was constructed with routine molecular methods and transfected into hepatocarcinoma cell HepG2.Relative quantification of IRS-1 mRNA was preformed by real-time reverse tran-scription PCR.Western blot was applied to detect the expression levels of IRS-1.Results 48 h after the plasmid carring RNF6 gene transfected HepG2 cells,the mRNA level of ISR-1 gene reduced to 69% of control group.The expression of IRS-1 was markedly lower than control group (P <0.01).Conclusion The expression of IRS-1 is markedly down-regulated in HepG2 and enhancement of the ubiquitylation level of IRS-1 contribute to the disorder in insulin signal transferring.

Objective To investigate the risk factors of severe intrahepatic cholestasis during early period after liver transplantation.Methods The clinical data of 225 patients who received orthotopic liver transplantation at the Nanfang Hospital of Southern Medical University from August 2004 to February 2011 were retrospectively analyzed.All patients were divided into positive group (60 patients with intrahepatic cholestasis) and negative group (165 patients without intrahepatic cholestasis).Preoperative,intraoperative and postoperative factors of the 2 groups were compared via t test,chi-square test,Wilcoxon test or Logistic regression analysis.Results The proportion of patients with hepatic cirrhosis,hepatic encephalopathy integral,ascites integral,international normalized ratio,and the levels of prothrombin time (PT),total bilirubin (TBil),aspartate aminotransferase of the positive group before operation were significantly higher than those in the negative group (x2 =6.09,Z =2.22,2.60,2.46,2.84,4.81,3.42,P ＜ 0.05),while the levels of albumin,Na +,K +,hemoglobin,platelet (PLT) of the positive group in the operation were significantly higher than those in the negative group (t =2.10,4.97,Z =2.49,t =3.51,Z =3.66,P ＜ 0.05).The ratio of compatible blood type of the donors and recipients,ratio of fatty liver graft,cold ischemia time,relative warm ischemia time,intraoperative blood loss,intraoperative transfusion of red blood cells,PLT,and cryoprecipitate of the positive group after the operation were significantly higher than those in the negative group (x2 =4.29,13.11,Z =2.45,2.61,3.75,3.20,2.89,3.95,P ＜0.05).The incidences of acute rejection,hepatic artery embolism,pulmonary infection,bacteraemia,fungal infection and cytomegalovirus (CMV) infection were significantly higher than those in the negative group (x2 =9.87,4.91,8.21,6.29,3.92,9.26,P ＜0.05).The results of multivariate analysis revealed that preoperative level of TBil ＞ 51.3 μmol/L,fatty of the liver graft,intraoperative transfusion of cryoprecipitate,postoperative acute rejection,hepatic artery embolism,postoperative pulmonary infection,bacteraemia,CMV infection were independent risk factors of severe inrahepatic cholestasis (OR =15.82,7.99,2.88,3.03,53.20,3.34,4.11,3.22,P ＜ 0.05).The incidence of severe intrahepatic cholestasis was significantly lower in patients with higher level of PLT and longer PT (OR =0.33,0.25,P ＜ 0.05).The mortality rates of the positive group and negative group at 6 months after the operation were 41.7％ (25/60) and 19.4％ (32/165),and the mortality rate of the positive group was significantly higher (x2 =11.54,P ＜ 0.05).Conclusion Correction of poor clinical status before liver transplantation,reinforcement of infection control and anti-rejection may reduce the incidences of complications and decrease the associated early mortality.

Objective To construct an objective,comprehensive core competence evaluation index system for nursing postgraduates,and to provide evidence for postgraduates nursing education.Methods According to literature review,expert interviews,focus group discussions,Delphi and other research methods,we conducted two rounds of 32 experts consultation in 18 cities from 10 provinces to establish the final core competence evaluation index system.Results The two experts consultation derived core competence evaluation index system including 7 level-1 indexes,20 level-2 indexes and 53 entries.Level-1 indexes:theoretical knowledge and skills,critical thinking ability,research ability,clinical care ability,teaching skills,interpersonal communication skills,professional development skills.Conclusions This program is rigorous and reliable,with some theoretical and practical significance of its results,which provide a reference for the training program,evaluation of the quality,assessment of postgraduate nursing education.

Objective To judge the prognosis of adult recipients after first liver transplantation by stepwise discriminant analysis,and screen out the main influencing factors.Methods The clinical data of 221 patients who received liver transplantation at the Nanfang Hospital of Southern Medical University were retrospectively analyzed.A total of 218 patients who met the criteria were divided into the training samples (188 patients admitted from August 2004 to June 2010) and checking samples (30 patients admitted from July 2010 to February 2011),and then all patients were re-divided into dead group (survival time ≤ 90 days,34 patients) and surviving group (survival time ＞ 90 days,184 patients).Factors which had significant difference after the univariate analysis was further analyzed by the stepwise discriminant analysis method.All data were analyzed by the t test,rank sum test,chi-square test or Fisher exact probability test.Results The ages of the recipients in the dead group and the living group were (54 ± 11)years and (51 ± 11)years,respectively,with no significant difference between the 2 groups (t =-1.681,P ＞ 0.05).The preoperative levels of hemoglobin in the dead group and the living group were 106.7 g/L and 119.2 g/L,respectively,with a significant difference between the 2 groups (t =2.809,P ＜ 0.05).There were significant differences in the levels of serum creatinine,urea nitrogen,albumin,total bilirubin,indirect bilirubin,Na+,prothrombin time,activated partial thromboplastin time,international normalized ratio,fibrinogen,prothrombin activity,platelet,nutrition risk index,model for end-stage liver disease score,number of patients with preoperative hepatic encephalopathy (HE),preoperative hepatorenal syndrome (HRS),preoperative digestive tract bleeding,preoperative infection,preoperative diabetes,Child-Turcotte-Pugh score,cardiac function classification and anesthesia risk rating operation time,anhepatic time,volume of intraoperative blood transfusion,volume of peritoneal effusion ; intraoperative urine output,between the 2 groups (Z =-2.277,-2.595,-3.290,-3.486,-2.562,-2.577,-3.670,-3.882,-3.625,-3.557,-3.837,-1.974,-3.693,-3.815,x2 =19.632,9.756,28.143,Z =-4.175,-3.905,-4.865,-3.564,-5.822,P ＜ 0.05).Preoperative HE,preoperative HRS,duration of operation,intraoperative blood transfusion and intraoperative volume of urine were the independent influencing factors of early prognosis after liver transplantation.The standardized partial regression coefficients were 0.146,0.188,0.257,0.181,-0.340,89.9％ (169/188) of the training samples and 90.0％ (27/30) of the checking samples were correctly classified.Conclusion Based on factors including HRS,HE,intraoperative blood transfusion,intraoperative volume of urine and duration of operation,the early prognosis can be judged in adult recipients after first liver transplantation.

Objectives To evaluate the predictive value of the revised model for end-stage liver disease in the clinical early stage after liver transplantation.Methods The clinical data of 218 patients were retrospectively analyzed.After calculating the MELD score,ReFit MELD score and ReFit MELDNa score before transplantation,we compared the predictive accuracies of these scoring systems using the area under curve (AUC) of the receiver operating characteristic.The groups were categorized with the cut-offs of the MELD,ReFit MELD and ReFit MELDNa,and the early-stage complications and mortality in the different groups were analyzed.Results The AUC for the MELD,ReFit MELD and ReFit MELDNa were 0.737 (95％CI 0.621～0.854),0.727 (95％CI 0.663～0.785) and 0.735 (95％CI 0.671～0.792),respectively.There was no statistical difference is the AUC among the MELD,ReFit MELD and ReFit MELDNa.Elevated scores in the 3 models predicted higher rates of pulmonary infection,abdominal infection and acute renal dysfunction,as well as a higher mortality.Conclusions The ReFit MELD score and ReFit MELDNa score were relatively useful predictors of short-term survival rates after liver transplantation.The predictive accuracy was similar to the MELD score.Values of the score above the cutoff values indicated higher rates of complication and poorer prognosis.

Objective:To study on the effect of ultrasound-guided thoracic paravertebral nerve (TPVB) block on quality of recovery from general anesthesia in tuberculosis patients with fiberboard exfoliation in post anesthesia recovery unit (PACU).Methods:From May 2018 to December 2019, 40 tuberculosis patients in Changsha Central Hospital with pulmonary fibreboard exfoliation and focal abscess lesions cleaning were randomly divided into two groups, with 20 patients in each group. The patients in group A received endobronchial general anesthesia and in group B received ultrasound-guided TPVB combined with endobronchial general anesthesia. Patients in the two groups were maintained under anesthesia by propofol, and the bispectral index (BIS) was maintained within the range of 40-50. The dosage of propofol and sufentanil was adjusted according to changes in BIS and hemodynamics. The mean arterial pressure (MAP), heart rate (HR) in two groups of patients were recorded at before anesthesia induction (T 0), before cutting leather (T 1), cut skin after (T 2), the end of operation (T 3), extubation time (T 4), and T 5 (time of leaving PACU). The visual analogue scale (VAS) of all patients in resting and cough state was recorded at 5, 30 min after extubation and the time of leaving PACU. The dosage of propofol and sufentanil in the operation and the additional dosage of sufentanil in PACU were recorded in both two groups. And the respiratory recovery time, consciousness recovery time, extubation time and sedation agitation scale(SAS) were observed. The adverse reactions such as nausea, vomiting, drowsiness and hypotension were observed in PACU. Results:Compared with group A, MAP and HR of patients at T 2, T 3, T 4, T 5 in group B were more stable during anesthesia, and VAS of patients in group B were lower than that in group A at each time point after extubation ( P<0.05). The dosage of sufentanil and propofol in group B were (35.92±8.12)μg and (749.56±95.30)mg respectively, which were significantly lower than those in group A [(45.74±4.42)μg and (862.83±105.34)mg, P<0.05]; the dosage of sufentanil in postoperative anesthesia recovery room of group B was (5.26±2.10)μg, significantly less than that of group A (10.35±5.86)μg ( P<0.05). The respiratory recovery time, consciousness recovery time and extubation time in group B were (12.92±5.12) min, (20.56±5.10) min and (26.87 ± 6.16) min, which were shorter than those in group A [(15.74±4.72)min, (25.83±5.34)min and (35.35±5.80)min, P<0.05]. The incidence of postoperative nausea, vomiting, lethargy and hypotension in group B were 10%, 10%, 35% and 20%, which were significantly lower than those in group A (30%, 20%, 75% and 45%, P<0.05). Conclusions:Ultrasound-guided paravertebral nerve block may significantly reduce the dosage of opioid analgesics for general anesthesia in tuberculosis patients with fiberboard exfoliation, accelerate the speed of anesthesia recovery, reduce the agitation during recovery, and improve the quality of anesthesia recovery.

OBJECTIVETo study major human UGT isoforms involved in trans-resveratrol (TR) phase II metabolism.

METHODtrans-resveratrol and 12 major human UGT isoforms were incubated in vitro and then glucuronic acid metabolites were determined by HPLC-MS, in order to preliminarily analyze the structure and observe the effect of different UGT isoforms on the generation rate of glucuronic acid metabolites.

RESULTIn in vitro metabolic system, two metabolites-4'-O-monoglucuronide resveratrol (M-1) and 3-0-monoglucuronide resveratrol (M-2)-were generated from trans-resveratrol after being catalyzed by UGT. During the cause, generation of M-1 and M-2 were catalyzed by UGT1A1, UGT1A3, 1A8, 1A9 andlA10, whereas only UGT1A6 and 1A7 contributed to the forma-tion of M-2. Both the formation rate of M-1 and M-2 catalyzed by UGT1A1, 1A10 and the formation of M-2 catalyzed by UGT1A8 slowed down with the increasing concentration of substrates, causing the phenomenon of "substrate inhibition".

CONCLUSIONUGT1A1, 1A8, 1A9, 1A10 get involved in the formation of M-1, and of them UGTIA9 is the most important contributor. UGT1A3 also makes small contribution to the formation of M-1 and M-2, while other UGT isoforms show hardly any reaction with the trans-resveratrol phase II metabolites.

Glucuronic Acid ; metabolism ; Glucuronosyltransferase ; metabolism ; Humans ; Isoenzymes ; metabolism ; Kinetics ; Stilbenes ; chemistry ; metabolism

Objective To investigate the clinical efficacy of bone filling mesh bag combined with pedicle anchoring for the treatment of Stage Ⅲ reducible Kummell disease.Method The 35 paients with Stage Ⅲ reducible Kummell disease were treated with bone filling mesh bag combined with pedicle anchoring from January 2018 to December 2022.The operation Time,intraoperative blood lose,bone cement injection volume and surgical complications were recorded.The VAS score,ODI value,kyphosis Cobb angle and midline height of the injured vertebral were compared at preoperative,postoperative 1 day and last follow-up.Results All patients were followed up for 12-24 months[(15±3.5)months].Operation time was 35-63 min[(45±5.8)min],intraoperative blood loss was 10-35 ml[(20±5)ml],bone cement injection volume was 4.5-7.8 ml[(5.5±1.8)ml].There were 4 cases of bone cement leakage,there were 1 case of intervertebral leakage,2 cases of lateral leakage,1 case of anterior leakage and no patient with intracanal leakage.All bone cement leakage did not lead to clinical symptoms,bone cement poisoning and pulmonary embolism.No cement mass slip.All patients were followed up for 12 to 24 months[(15±3.5)months].VAS scores and Oswestry Disability Index(ODI)values were significantly lower on the first day after surgery than before surgery,with statistical significance(P＜0.05).The 3-month follow-up was slightly higher than that on the first day after surgery,and the difference was not statistically significant(P＞0.05).The midline height and Cobb Angle of the injured vertebra were measured by imaging.The height of the injured vertebra recovered significantly on the first day after operation,and the Cobb Angle decreased significantly,the difference was statistically significant(P＜0.05).The midline height of the injured vertebrae decreased and the Cobb Angle increased slightly at 3 months after the operation,but the difference was not statistically significant(P＞0.05).Conclusion In the the treatment of Stage Ⅲ reducible Kummell disease,Bone filling mesh bag combined with Pedicle anchoring have good clinical efficacy,which can significantly reduce the pain of patients,relieve clinical symptoms,improve spinal function,improve quality of life,and reduce the incidence of bone cement leakage and slippage.

OBJECTIVETo detect pharmacokinetics of Danshensu Sodium in Danshen dripping solution in Beagles.

METHODDanshen dripping solution was dripped intravenously into healthy Beagles at a dose of 10 mL x kg(-1). Their plasma samples were extracted with acetic ether, the blood concentrations were determined by HPLC method.

RESULTDanshensu Sodium showed a good linear relationship within the range of 0.225-18.000 mg x L(-1), with the lowest detectable limit of 0.113 mg x L(-1). Its pharmacokinetic parameters were as follows: tmax was 30 min, Cmax was (9.5742 +/- 2.3715) mg x L(-1), t1/2 was (19.23 +/- 2.97) min, CL was (0.0127 +/- 0.0030) L x min(-1) x kg(-1), AUC(0-tn) was (474.954 +/- 95.483) mg x min x L(-1) and AUC(0-infinity) was (482.494 +/- 95.353) mg x min x L(-1).

CONCLUSIONThe accurate, stable and reliable blood concentration method shows a one-compartment mode of Danshensu Sodium in Beagles.

Animals ; Chromatography, High Pressure Liquid ; Dogs ; Drug Stability ; Drugs, Chinese Herbal ; pharmacokinetics ; Female ; Lactates ; pharmacokinetics ; Male ; Solutions

Objective:To construct the expression vector of the fusion protein of human serum albumin (HSA) and thymopentin (TP5) and to express it in Pichia pastoris,and to elucidate the biological activity of fusion protein.Methods:The HSA-TP5 fusion gene was constructed by gene recombination and transfected into Pichia pastoris to construct the eukaryotic expression system of HSA-TP5.The recombinant eukaryotic expression plasmid of PPICZα-HSA-TP5 was obtained by agarose gel electrophoresis and purification reagent.The two step fermentation method was used to ferment gene engineering bacteria of HSA-TP5 in high density,and the fermentation supernatant protein was precipitated and concentrated;the purified fusion protein was obtained by cation exchange chromatography and hydrophobic chromatography and analyzed by SDS-polyacrylamide gel electrophoresis.The effect of the fusion protein on the proliferation of lymphocytes was detected by MTT assay.Results:The HSA target gene fragment with length of 1 845 bp was achieved by PCR method.The HSA-TP5-pPICZαC fusion plasmid was identified by restriction endonuclease digestion,and the fragment length was 707 bp.The sequence analysis showed that the HSA and TP5 sequences of the target genes were identical with the gene sequences reported in GenBank and were fused by forward fusion.PCR method confirmed that the eukaryotic recombinant plasmid PPICZ αC-HSA-TP5 was integrated into the yeast genome,and compared with control group,the target gene PCR product length was found to be 1 860 bp.SDS-PAGE analysis showed that the expression level of HSA-TP5 fusion protein was gradually increased with the induction time within 72 h.HSA-TP5 fusion protein was purified by cation exchange chromatography and AKTA multifunctional protein purification system.The MTT assay results showed that HSA-TP5 fusion protein was consistent with TP5 protein in promoting lymphocyte proliferation activity.Conclusion:HSA-TP5 fusion protein can be obtained by constructing the eukaryotic expression system of Pichia pastoris and owns the biological activity.