Objective:To investigate the effect of Lamins B2 (LMNB2) on the migration of human retroperitoneal liposarcoma (RPLS) cells SW872.Methods:Immunohistochemistry was used to analyze the the differential expression levels of LMNB2 in 33 RPLS tissue samples . The correlation between LMNB2 expression and clinical prognosis and clinicopathological features was analyzed. siRNA was used to lower the expression level of LMNB2 in tumor cells, and the effect of LMNB2 on the scratch healing ability and migration ability of SW872 cells was examined by using wound-healing assay and transwell migration assay. The expression levels of p-AKT and AKT in each group cells were detected by Western blot.Results:Patients with high LMNB2 expression had a lower recurrence-free survival and overall survival compared to those with low LMNB2 expression, and were more likely to experience recurrence, ( χ2=4.872, P=0.027; χ2=4.180, P=0.041; χ2=7.127, P=0.008). The migration ability of cells was significantly reduced following the silencing of LMNB2 expression ( t=11.240, P<0.01; t=7.445, P<0.01). The expression level of p-AKT in the silencing group was significantly lower than that in the control group, while there was no significant difference in the expression level of AKT between the two groups ( t=9.784, P<0.01). Conclusion:LMNB2 may promote the migration of human retroperitoneal liposarcoma cells SW872 by regulating AKT signaling pathway.

Objective:To explore the standardized performance of a FISH probe before clinical detection.Methods:The probe sensitivity and specificity of ETV6/RUNX1 were analyzed via interphase and metaphase FISH in 20 discarded healthy bone marrow samples. The threshold system of the probe was established using an inverse beta distribution, and an interpretation standard was established. Finally, a parallel-controlled polymerase chain reaction detection study was conducted on 286 bone marrow samples from patients at our hospital. The clinical sensitivity, specificity, and diagnostic coincidence rate of ETV6/RUNX1 FISH detection were analyzed, and the diagnostic consistency of the two methods was analyzed by the kappa test.Results:The probe sensitivity and specificity of the ETV6/RUNX1 probe were 98.47% and 100%, respectively. When 50, 100, and 200 cells were counted, the typical positive signal pattern cutoffs were 5.81%, 2.95%, and 1.49%, respectively, and the atypical positive signal pattern cutoffs were 13.98%, 9.75%, and 6.26%, respectively. The clinical sensitivity of FISH was 96.1%, clinical specificity was 99.6%, diagnostic coincidence rate was 99.00%, diagnostic consistency test kappa value was 0.964, and P value was <0.001.Conclusion:For FISH probes without a national medical device registration certificate, standardized performance verification and methodology performance verification can be performed using laboratory developed test verification standards to ensure a reliable and accurate reference basis for clinical diagnosis and treatment.

Objective:To establish a conditional knockout mouse model of polycystic kidney disease 1 ( Pkd1) gene based on CRISPR/Cas9 and Cre-loxP gene editing technology, and to provide an animal model for in-depth research on the role of Pkd1 gene in the development of polycystic kidney disease. Methods:In-Fusion technology was used to construct a targeting vector. Corresponding gRNAs, Cas9 mRNAs, and donor vectors carrying the loxP site were prepared based on the Pkd1 gene, and injected into the fertilized eggs of C57BL/6N mice. The fertilized eggs were transferred to the fallopian tubes of female mice with pseudopregnancy. After the newborn mice were identified by PCR and sequencing analysis, Pkd1 flox/flox F0 generation positive mice were selected. The F0 generation positive mice were bred with wild-type mice, and F1 generation heterozygous mice with Pkd1 flox/+ genotype were selected for offspring. F2 generation homozygous mice with Pkd1 flox/flox genotype were obtained through internal expansion, and then hybridized with Cre positive Ggt1/ Cre mice. F3 generation mice with Pkd1 flox/+Ggt1 Cre genotype were obtained. F4 generation mice with Pkd1 flox/flox Ggt1 Cre genotype were obtained by self crossing or backcrossing with F2 generation Pkd1 flox/flox, namely kidney-specific Pkd1 gene knockout mice ( Ggt1-cKO mice). PCR method was used to identify the genotype of mice, and then the mice were divided into wild-type control (WT) group ( n=6), Pkd1 homozygous control (PKD) group ( n=6), and Ggt1-cKO knockout validation (CKO) group ( n=6) according to the gene identification results. Real-time fluorescence quantitative PCR (RT-qPCR) was used to detect the expression of Pkd1 mRNA in the kidneys and other organs of mice in each group. HE staining was used to detect the pathological changes in renal tissues of mice in each group. The automatic biochemical detector was used to detect the blood urea nitrogen and serum creatinine levels of mice, and the kidney coefficient was calculated. Results:The PCR detection results showed that the genotype of offspring mice in CKO group was consistent with Pkd1floxflox Ggt1 Cre. Pkd1 gene was only specifically expressed in the kidney, but not in other tissues. The RT-qPCR results showed that the relative expression of Pkd1 mRNA in the renal medulla of CKO group was significantly lower than that of WT and PKD groups. The kidney volume of the CKO group had increased by about twice compared to the WT group. Under the microscope, it could be observed that there were multiple vacuoles of varying sizes and shapes in the kidneys of the CKO group, and there was a significant increase in the interstitial space of the medullary tissue. The kidney coefficient, blood urea nitrogen, and serum creatinine in the CKO group were significantly higher than those in the WT and PKD groups (all P<0.05). Conclusion:Based on CRISPR/Cas9 and Cre-loxP gene editing technology, Pkd1 gene kidney conditional knockout mice can be successfully constructed, providing an animal model for further studying the action mechanism of Pkd1 gene in polycystic kidney disease.

Objective:To investigate the perioperative alterations and management of coagulation function in patients of massive blood transfusion during retroperitoneal tumor (RT)resection.Methods:Fourty-seven RT patients at Peking University International Hospital from Jan 2016 to Dec 2021 undergoing resection with massive blood transfusion more than 20 U within 24 h were reviewed for coagulation function before and after surgery.Results:Intraoperative bleeding was 3 000-25 800 ml, 10 patients had blood loss ≥10 000 ml. During the operation, (25.3±9.9) U of red blood cells were transfused, (2 720±1 369) ml plasma transfused, and (2.4±3.3) U platelets were transfused in 6 patients. Fourty-five patients received intraoperative albumin of (79.5±46.5) g; All 47 patients received fibrinogen of (2.3±1.3) g; Prothrombin complex was given in 45 patients (1 205±807) U. Preoperative hemoglobin was statistically different compared to postoperatively and days 1, 3 and 5 ( W=1 790, P<0.001; W=1 672, P<0.001; W=1 704, P<0.001; W=1 486, P=0.004);As with platelets, the difference was also statistically significant compared to postoperative days 1, 3, and 5 ( W=2 153, P<0.001; W=2 092, P<0.001; W=1 732, P<0.001); Preoperative albumin was different compared to postoperative days 1 and 3 ( W=1 568, P<0.001; W=1 578, P<0.001,); Preoperative fibrinogen was different compared to postoperative day 1 ( W=1 964, P<0.001). PT and APTT were prolonged on postoperative days 1 and 3 ( W=628, P<0.001, W=804, P=0.023) ( W=661, P<0.001, W=796, P=0.02). Patient's preoperative fibrin degradation products and D-dimer were above the normal value and were higher on postoperative days 3 and 5 ( W=498, P<0.001, W=345, P<0.001). Conclusions:Coagulation disorders occur perioperatively in patients with massive transfusion while undergoing surgery for RT.The implementation of ratiional transfusion strategy and close postoperative survey and management of coagulation dysfunction help avoid the coagulation related morbidities.

Objective:To investigate the clinical efficacy and prognostic influencing factors of open abdomen technique for acute pancreatitis with abdominal compartment syndrome (ACS).Methods:The retrospective cohort study was conducted. The clinical data of 186 patients of acute pancreatitis with ACS who were admitted to 6 hospitals, including 65 cases in the 910th Hospital of Joint Logistics Support Force of Chinese People′s Liberation Army, 46 cases in the First Affiliated Hospital of Wenzhou Medical University, 33 cases in the Fujian Provincial Hospital, 31 cases in the Second Affiliated Hospital of Fujian Medical University, 7 cases in the People′s Hospital Affiliated to Quanzhou Medical College, 4 cases in the Shishi General Hospital, from January 2013 to December 2020 were collected. There were 142 males and 44 females, aged (43±8)years. Observation indica-tors: (1) patients conditions after being treatment with open abdomen technique; (2) analysis of clinical characteristics in patients with different treatment outcomes; (3) changing trend of the volume of urine output, levels of lactic acid, levels of enteral nutrient intake and the sequential organ failure score in patients with different treatment outcomes; (4) influencing factors for prognosis of patients. Measurement data with normal distribution were represented as Mean± SD, and compari-son between groups was analyzed using the t test. Measurement data with skewed distribution were represented as M( Q1, Q3), and comparison between groups was analyzed using the Mann-Whitney U test. Count data were described as absolute numbers, and comparison between groups was analyzed using the chi-square test or the continuity correction chi-square test. Repeated measurement data were analyzed using the repeated ANOVA. Spearman correlation analysis was used for correlation analyses. The COX regression model was used for univariate analysis and COX regression model with forward regression was used for multivariate analysis. Results:(1) Patients conditions after being treatment with open abdomen technique. Intra-abdominal pressure, oxygena-tion index, levels of lactic acid and sequential organ failure score of the 186 patients were (23.3±1.9)mmHg (1 mmHg=0.133 kPa), (121±24)mmHg, (5.0±3.4)mmol/L and 10.4±3.6 before the treatment with open abdomen technique and (11.2±2.9)mmHg, (222±38)mmHg, (3.2±2.1)mmol/L and 4.4±2.3 at postoperative 168 hours, showing significant differences in time effects before and after the treatment ( Ftime=855.26,208.50, 174.91,208.47, P<0.05). (2) Analysis of clinical characteristics in patients with different treatment outcomes. Of the 186 patients, 166 cases survived and were discharged, and 20 cases died during hospitalization. Age, sequential organ failure score, duration of ACS and levels of lactic acid during hospitalization before the treatment with open abdomen technique were (41±7)years, 9.4±3.4, 13(10,21)hours and (4.2±0.6)mmol/L in surviving patients, versus (45±6)years, 11.5±2.4, 65(39,84)hours and (5.2±0.5)mmol/L in dead patients, respectively, showing significant differences between them ( t=-2.10, -2.71, Z=-5.36, t=-7.16, P<0.05). Duration of postoperative acute gastro-intestinal injury, duration of continuous renal replacement therapy, time to liberation from mech-anical ventilation, duration of vasoactive drugs therapy, cases undergoing early abdominal closure, cases without intestinal fistula or with postoperative high-order intestinal fistula and low-order intestinal fistula during hospitalization after the treatment with open abdomen technique were 4(2,6)days, 4(3,7)days, 34(21,41)days, 3(2,6)days, 126, 131, 23, 12 in surviving patients, versus 13(10,17)days, 10(8,18)days, 0(0,3)days, 8(6,12)days, 1, 2, 15, 3 in dead patients, respectively, showing significant differences between them ( Z=-5.60, -3.75, -3.64, -3.06, χ2=41.43, 45.86, P<0.05). (3) Changing trend of the volume of urine output, levels of lactic acid, levels of enteral nutrient intake and the sequential organ failure score in patients with different treatment outcomes. The volume of urine output, levels of lactic acid, levels of enteral nutrient intake and the sequential organ failure score in surviving patients during hospitalization were (0.29±0.10)mL/(kg·h), (4.2±0.6)mmol/L, 0.0 kcal/(kg·d) and 9.4±3.4 before the treatment with open abdomen technique and (2.22±0.15)mL/(kg·h), (1.9±0.7)mmol/L, (20.7±2.9)kcal/(kg·d) and 3.7±2.2 at postoperative 168 hours. The above indicators in dead patients during hospitalization were (0.28±0.08)mL/(kg·h), (5.2±0.5)mmol/L, 0.0kcal/(kg·d) and 11.5±2.4 before the treatment with open abdomen technique and (0.28±0.09)mL/(kg·h), (7.7±0.8)mmol/L, (4.6±1.8)kcal/(kg·d) and 12.4±2.1 at postoperative 168 hours. There were significant differences in time effects in the above indicators in surviving patients and dead patients before and after the treatment with open abdomen technique ( Ftime=425.57, 188.59, 394.84, 37.52, P<0.05). There were interactive effects between the above indicators and the treatment outcome at different time points ( Finteraction=383.14, 233.04, 169.83, 36.61, P<0.05). There were signifi-cant differences in the change trends of the above indicators between the surviving patients and the dead patients during hospitalization ( Fgouprs=2 739.56, 877.98, 542.05, 240.85, P<0.05). (4) Influen-cing factors for prognosis of patients. Results of univariate analysis showed that age, sequential organ failure score, duration of ACS before surgery, procalcitonin, lactic acid, postoperative high-order intestinal fistula, abdominal hemorrhage, duration of postoperative acute gastrointestinal injury, duration of continuous renal replacement therapy, duration of vasoactive drugs therapy, early abdominal closure were related factors influencing prognosis of patients under-going treatment with open abdomen technique ( hazard ratio=1.07, 1.18, 1.39, 1.16, 8.25, 12.26, 2.83, 1.29, 1.56, 1.41, 0.02, 95% confidence interval as 1.00-1.15, 1.45-2.27, 1.22-1.57, 1.02-1.32, 1.75-38.90, 7.37-41.23, 1.16-6.93, 1.22-1.37, 1.23-1.99, 1.08-1.84, 0.00-0.16, P<0.05). Results of multivariate analysis showed that extended duration of ACS before surgery, postoperative high-order intestinal fistula and extended duration of postoperative acute gastrointestinal injury were independent risk factors influencing prognosis of patients undergoing treatment with open abdomen technique ( hazard ratio=1.05, 7.95, 1.17, 95% confidence interval as 1.01-1.32, 2.05-30.87, 1.13-1.95, P<0.05) and early abdominal closure was an independent protective factor ( hazard ratio=0.10, 95% confidence interval as 0.01-0.89, P<0.05). Results of Spearman correlation analysis showed that duration of ACS was positively correlated with sequential organ failure score before surgery ( r=0.71, P<0.05). Conclusions:Open abdomen technique is effective for acute pancreatitis with ACS. Extended duration of ACS before surgery, postoperative high-order intestinal fistula and extended duration of postoperative acute gastrointestinal injury are independent risk factors for prognosis of patients during hospitalization and early abdominal closure is an independent protective factor.

Through the analysis of patent jurisprudence, it is proved that the Classical Prescription of Traditional Chinese Medicine (CPTCM) belongs to the scope of "existing technology" in the Law of Patent, and has lost the foundation of obtaining patent rights. Taking Japan's CPTCM as an example, based on the analysis of the administration of CPTCM and patent applications related to CPTCM in Japan, it is proved that CPTCM can not obtain patent authorization in Japan. Through the comparison of patent in China, the United States, Europe and worldwide region, it is proved that China is still the main source of patent applications for Traditional Chinese Medicine. At the end of the article, the author puts forward the suggestion of "active protection". It is believed that we should abandon the concept of patent-only protection and improve the influence of Traditional Chinese Medicine in worldwide from the aspects of industrial development, promotion ofits application, and intellectual property protection.

Objective To explore the formation process of vacuum pressure differential of micro power negative pressure technology to facilitate its clinical application.MethodsThe formation of negative pressure differential and changes of negative pressure values in enclosure space were studied which was formed by liquid aspiration after the compressive polyvinyl alcohol foam material was enclosed.Micro power negative pressure technology was applied to the treatment of the right ear necrosis patient after focal cleaning,and the efficacy was observed.Results Medical adhesive film was raised gradually during the expansion of polyvinyl alcohol foam due to liquid aspiration,and then the enclosure space extended increasingly to form local vacuum negative pressure.The range of maximal negative pressure was between-110 and-200 Pa,and the mean maximal negative pressure value was-132.7 Pa.Micro power negative pressure technology behaved well in healing the wound.Conclusion Micro power negative pressure technology gains advantages in reliability and easy operation over other technologies and products,and may be a new choice for healing of refractory wounds.

Objective To clone and express the alkaline protease AprA , one important virulence factor secreted by Pseudomonas aeruginosa(PAE)in Escherichia coli, to clone and express the inhibitor of AprA (AprI) and its substrate flagellin , and to detect the function of AprA and the inhibitory function of AprI .Methods The genes encoding AprA ,AprI and flagellin gene were amplified respectively by PCR using PAE PAO 1 genome DNA as the template .The expression vec-tors (pET-28a-AprA, pET-28a-AprI and pET-28a-Flagellin) were constructed and transformed into E.coli BL21(DE3) respectively.The recombinant AprA protein was expressed by IPTG induction and purified via denaturing and renaturation. The recombinant AprI and flagellin were expressed and purified by Ni 2+affinity chromatography .The cleavage activities of AprA on flagellin were detected by SDS-PAGE.Results Recombinant AprA , AprI and flagellin protein were expressed and purified .It was demonstrated that AprA cleaved flagellin , which was blocked by AprI .Conclusion Recombinant AprA could cleave its substrates as an alkaline protease , and its inhibitor AprI inhibits the activities of AprA .This study will contribute to further investigations on the role of AprA in the pathogenesis of PAE .

Objective To investigate the risk factors of severe hand,foot and mouth disease (HFMD).Methods 175 severe cases of HFMD and 183 mild cases of HFMD in the same period were randomly selected.Single factor analysis was first performed between severe and mild cases on age,gender,residence,symptoms,signs and laboratory examinations,etc,to screen out the related risk factors which P value ＜ 0.05.Then,binary logistic regression analysis was carried out to determine risk factors most related to severe HFMD.Finally,receiver operating characteristic curve (ROC) analysis was performed on severe HFMD related risk factors.Results Single factor analysis showed that there were obvious differences between children with mild HFMD and those with severe HFMD in the factors like difficulty in breathing,walking instability,vomiting,limb shaking,disturbance of consciousness,convulsions,cold sweat and weakness,thermal process,the degree of fever,pulmonary rales,heart rate,serum EV71 antibody,circulatory failure,leukocyte count,platelet count,neutrophil ratio,CRP,blood glucose,etc (x2 =15.236,19.819,33.823,52.670,12.984,10.180,29.318,52.932,34.544,14.615,46.633,31.407 and 5.303,t =3.184,3.144,2.256,2.244 and 2.828,,all P ＜0.05).Binary logistic regression analysis showed that the thermal process,startle tremor or limb jitter,serum EV71 antibody,vomiting,fever,neutrophil ratio were the related risk factors of severe HFMD (B value =2.605,2.129,1.409,1.185,0.841 and 0.103,all P ＜ 0.05).ROC analysis showed that the areas under the curve of the predicted probability and thermal process were larger than any other risk factors [(95％ CI (0.888 ～ 0.961) and (0.818 ～ 0.920)],and thus had better diagnostic values.Conclusion Children under 3 years old were the high risk population of HFMD.Such clinical symptoms as persistent high fever,vomiting,startle tremor orlimb jitter,EV71 antibody in serum and increasing neutrophil ratio were risk factors for severe HFMD.The predicted probability had more diagnostic value than any other risk factors.

Objective To determine the median effective target effect-site concentration (EC50) of sufentanil when intraoperative wake-up test was successful in patients undergoing scoliosis surgery.Methods Ninety ASA Ⅰ or Ⅱ patients,aged 18-64 yr,with body mass index ＜ 30 kg/m2,scheduled for scoliosis surgery under sevoflurane and sufentanil anesthesia,were randomly divided into Ⅰ-Ⅵ groups (n =15 each).Anesthesia was induced with sufentanil,etomidate and cisatracurium,and maintained with sevoflurane,sufentanil and cisatracurium.The patients were tracheal intubated and mechanically ventilated.Before the wake-up test,infusion of cisatracurium was stopped and the end-tidal concentration of sevoflurane was adjusted to 0.The EC50 was determined by the K(a)rber method.The target effect-site concentration of sufentanil was set at 0.19 ng/ml in group Ⅰ,0.18 ng/ml in group Ⅱ and gradually decreased in decrements of 0.01 ng/ml.The wake-up test was performed 5 min later.The EC50 and 95 ％ confidence interval of sufentanil were calculated by the K(a)rber method.Results EC50 of sufentanil obtained was 0.164 ng/ml and 95％ confidence interval of sufentanil obtained was 0.157-0.172 ng/ml when the wake-up test was successful.Conclusion The EC50 of sufentanil is 0.164 ng/ml when the intraoperative wake-up test is successful in the patients undergoing scoliosis surgery.