OBJECTIVE:To build up a method for determining the concentration of zaleplon in human plasma by HPLC. METHODS:The Kromasil-C 8 column was used and mobile phase consisted of methanol-water(65∶35);the flow rate was1.0ml/min,fluorescence excitation wave-length was233nm;fluorescence emission wave-length was460nm.RESULTS:The calibration curve revealed linearity in the concentration range of0.42～73.92ng/ml(r=0.9999)with a regressive equation of Y=1.870?147.The average recovery was99.47%.The with-day and between-day RSDs were4.9%and4.73%respec?tively.CONCLUSION:The method is accurate,reliable and highly sensitive so it can be used in study of pharmacokinetics of zaleplon.

OBJECTIVE:Dual-wavelength spectrophotometry was developed for the determination of ciprofloxacin lactate in its eye drops.METHODS:Detection was performed at wavelength 237nm and 279nm with 0.1mol/L HCl as diluent.RESU_LTS:The assay was linear for ciprofloxacin lactate in the concentration range of 3～9?g/ml,r=0.9 998;The mean recovery was 101.6%,RSD was 2.16%.CONCLUSION:This method is simple,quick,accurate and suitable for the determination of ciprofloxacin lactate in its eye drops.

AIM: To study the effect of ginsenosides on lipopolysaccharide-induced expression of tissue factor (TF) and plasminogen activator inhibitor type-1 (PAI-1) in vascular endothelial cells (EC), and to investigate the mechanism of ginsenosides in the healthy protection and treatment of cardiovascular diseases. METHODS: Human umbilical vein endothelial cells (HUVEC) were cultured by trypsin digestion method. PAI-1 was measured in the conditioned medium of HUVEC by a specific enzyme-linked immunosorbent assay (ELISA), whereas TF activity was measured in the lysates of these cells by a single step clotting assay. Specific mRNA expressions were determined by Northern blotting. RESULTS: Treatment of HUVEC with LPS resulted in a significant increase in PAI-1 antigen and TF activity. Ginsenosides inhibited this LPS-induced upregulation of PAI-1 protein and TF activity in HUVEC. These effects were also confirmed on the level of specific PAI-1 and TF mRNA expression by Northern blotting. CONCLUSION: Ginsenosides counteract activated endothelial cells by inhibiting LPS-induced PAI-1 and TF expression in these cells. This ability of ginsenosides might explain its efficacy in the healthy protection and the treatment of cardiovascular diseases.

AIM: To investigate the effect of homocysteine (HCY) on the induction of macrophage inflammatory protein-1? (MIP-1?) expression in cultured human umbilical vein endothelial cells (HUVECs). METHODS: After exposure of the cultured HUVECs to HCY at increasing concentrations (0.1, 0.5 and 1 mmol/L) for 8 h, the MIP-1? mRNA expression was determined by in situ hybridization using a MIP-1? cDNA probe, and the MIP-1? protein expression was measured by cell enzyme-linked immunosorbent assay (ELISA) using a goat anti-human MIP-1? monoclonal antibody. RESULTS: The in situ hybridization showed that cultured HUVECs were able to express MIP-1? mRNA at a low level that was purplish blue granules in cytoplasm. After exposure to HCY at the concentrations mentioned above, the expression of MIP-1? mRNA was significantly increased in a dose-dependent manner. Analysis of variance showed that there was significant difference between groups ( F= 606.38, P

Objective:To study preoperative MRI imaging and its enhanced mode on tumor features in predicting microvascular invasion (MVI) in patients with hepatocellular carcinoma (HCC).Methods:The clinical data of patients with a solitary HCC who underwent MRI examination followed by surgical resection at the General Hospital of Ningxia Medical University from January 2017 to June 2019 were studied. The patients were divided into the MVI (+ ) and MVI (-) groups according to the findings on postoperative pathological diagnosis. The relationship between the rates of MVI and MRI tumor features including diffusion weighted imaging (DWI) signal, enhancement mode, enhancement type and other imaging characteristics were analysed.Results:Of 84 patients with HCC enrolled into this study, there were 65 males and 19 females. Their age (Mean±SD) was (54.94±11.51) years. MVI (+ ) was found in 46 patients and MVI (-) in 38 patients. The maximum tumor diameters (Mean±SD) of the two groups were (7.08±3.45) cm and (4.28±2.47) cm ( P<0.01). Single-factor analysis and comparison of imaging characteristics of the two groups of patients showed tumor DWI signal, tumor encapsulation, enhancement mode, tumor edge smoothness, abnormal enhancement around tumors, and intratumoral arteries were significantly different ( P<0.05); There were no significant differences in T 1WI signals, T 2WI signals, tumor periphery, and enhancement types between groups. After inputting MVI(+ ) as a risk factor into the logistic regression model, tumor maximum diameters >6.33 cm, type 3/4 enhancement mode, and unsmoothness of tumor edge were independent risk factors (all P<0.05). Through combined diagnosis using ROC curve analysis with a cut-off value of 0.53, the area under the curve was 0.881, the sensitivity 0.870, specificity 0.789, and the Youden index 0.659. Conclusion:The multivariate logistic regression model and combined diagnosis using ROC curve analysis improved the diagnostic efficacy of MVI in its prediction of HCC on imaging studies. The risk predictors were easy to use and to promote in clinical practice.

To construct the eukaryotic expression plasmid of human PRX3 and measure its expression in the HEK-293FT cells, the full-length coding region of human PRX3 was cloned by PCR and inserted into the eukaryotic expression vector pcDNA4-Xpress (A). HEK-293FT cells were transiently transfected with the recombinant plasmid. Western blot and immuofluorescence were used to detect the expression of the fusion protein. In the experiment, restriction analysis identified the construction of the recombinant plasmid and the inserted sequence was identical with that published on GenBank. Western blot and immunofluorescence confirmed the expression of the recombinant protein in transfected HEK-293FT cells. It was concluded that the eukaryotic expression plasmid of human PRX3 was constructed successfully and the recombinant could be expressed efficiently in HEK-293FT cells, which provides a sound basis for the further study on human PRX3.
Cell Line, Transformed ; Cloning, Molecular ; Embryo ; Eukaryotic Cells/*metabolism ; Gene Expression ; Genetic Vectors ; Kidney/cytology ; Kidney/*metabolism ; Peroxidases/*biosynthesis ; Peroxidases/genetics ; Plasmids/*genetics ; Transfection

AIM:To construct recombinant adenovirus vector carrying human miR-133a and study its expression in human mesenchymal stem cells(hMSCs).METHODS:The PCR product containing miR-133a was amplified from human genomic DNA and inserted into the adenoviral shuttle vector pAdTrack-CMV.Then the recombinant shuttle plasmid linearized by pmeⅠwas cotransformed into competent E.coli.BJ5183 with the adenoviral backbone plasmid pAdEasy-1 to generate the recombinant adenovirus vector rAd-mir-133a.rAd-mir-133a was then packaged and amplified in human embryonic kidney 293(HEK293) cells.The purified rAd-miR-133a was used to infect the hMSCs and the expression of miR-133a was detected by non-quantitative RT-PCR and real-time PCR.RESULTS:The recombinant adenovirus shuttle vector pAdTrack-CMV-miR-133a was constructed and verified by restriction endonuclease analysis and DNA sequence analysis.rAd-miR-133a was successfully packaged and amplified in HEK293 cells.The transcriptions of primary miR-133a and mature miR-133a were over-expressed in the hMSCs infected with rAd-miR-133a.CONCLUSION:The recombinant adenovirus vector carrying human miR-133a is successfully constructed,which lay a foundation for miR-133a function study.

Objective:To analyze the imaging features of spontaneous rupture of primary liver cancer (PLC) and to study the high-risk factors associated with tumor rupture.Methods:From September 2016 to August 2020, 81 patients who developed spontaneous rupture of PLC at the General Hospital of Ningxia Medical University were included into this study. A control group of 81 patients with tumors located on the periphery of the liver but without rupture treated in the same period were selected by matching the two groups with age, sex and BCLC staging. The clinical data and CT imaging characteristics including tumor location, extent, size, and morphology of the two groups of patients were compared retrospectively between groups.Multivariate logistics regression was used to analyze.Results:A total of 81 patients were included in the case group, including 72 males and 9 females, aged (53.69±10.34) years. The control group included 81 patients, 64 males and 17 females, aged (54.78±9.04) years. The main risk factors for spontaneous rupture of PLC included in this study were cirrhosis, tumor close to diaphragm, biolobar distribution, portal vein obstruction, tumor diameter >10 cm, invasion of liver capsule (arc-to-chord ratio>1) and tumor protrusion ≥25% ( P<0.05). Logistic regression analysis showed that cirrhosis ( OR=2.796, 95% CI: 1.721-10.834), portal vein obstruction ( OR=3.586, 95% CI: 1.272-10.107) and tumor protrusion (≥25%) ( OR=2.831, 95% CI: 1.668-22.210) were independent predictive factors of spontaneous rupture of PLC. Conclusion:Tumor protrusion≥25%, cirrhosis and portal vein obstruction were closely related to spontaneous rupture of PLC. They were independent risk factors in predicting rupture of primary liver cancer.

Objective:To investigate the differences and changes of blood flow status of splenic volume, common hepatic artery, splenic arteriovenous, inner diameter of portal vein and hepatic in patients with hypersplenism of different degrees using multi-slice spiral CT whole-liver perfusion model.Methods:42 cases with hypersplenism of chronic hepatitis B with cirrhosis and 15 cases without hepatosplenic disease were collected as controls. All patients underwent multi-slice spiral CT whole-liver perfusion imaging. (1) The differences of spleen volume, common hepatic artery, splenic arteriovenous, and portal vein diameter between different degrees of hypersplenism and the control group were measured and compared. (2) The correlation between spleen volume and the inner diameter of each related vessels were analyzed and compared. (3) The values of perfusion parameters related to the five lobes of the liver in Couinaud segments based on hepatic artery perfusion (HAP), portal venous perfusion (PVP), total hepatic perfusion (TLP) and hepatic artery perfusion index (HPI) were measured and compared. One-way ANOVA was used to analyze the measurement data. The correlation between the spleen volume and the inner diameter of each blood vessel was analyzed by Pearson’s correlation analysis.Results:(1) spleen volume and the inner diameter of splenic artery, splenic vein and portal vein in the cirrhotic hypersplenism group were significantly larger than control group, and the difference was statistically significant ( F = 37.108, 17.484, 23.124, 13.636, P < 0.05). (2) spleen volume and the inner diameter of splenic artery, vein and portal vein in the moderate and severe hypersplenism groups were significantly larger than the mild hypersplenism group, and the difference was statistically significant ( F = 25.418, 13.293, 15.136, 7.093, P < 0.05), but there was no statistically significant difference between the moderate and severe hypersplenism groups ( P > 0.05). (3) The inner diameter of splenic vein, portal vein, and splenic artery was positively correlated with spleen volume ( r = 0.680, 0.548, and 0.726). (4) PVP and TLP of the whole liver in hypersplenism group were lower than control group ( P < 0.05), and the differences were statistically significant ( P < 0.05). HPI in the right posterior lobe of the liver in the moderate and severe hypersplenism group was higher than mild hypersplenism group ( F = 3.555, 4.570, P < 0.05), and there was no significant difference in the HAP in the whole liver among the groups ( P > 0.05), but the HAP in the whole liver in the severe hypersplenism group was lower than control, mild and moderate hypersplenism group. Conclusion:The inner diameter of the splenic arteriovenous in patients with hypersplenism of different degrees has widened to varying degrees, and is consistent with the increase in spleen volume, particularly in moderate and severe cases. Portal venous perfusion and total liver perfusion in patients with hypersplenism of different degrees have declined and the hepatic arterial perfusion in patients with severe hypersplenism is significantly reduced.

To construct the eukaryotic expression plasmid of human PRX3 and measure its expression in the HEK-293FT cells, the full-length coding region of human PRX3 was cloned by PCR and inserted into the eukaryotic expression vector pcDNA4-Xpress (A). HEK-293FT cells were transiently transfected with the recombinant plasmid. Western blot and immuofluorescence were used to detect the expression of the fusion protein. In the experiment, restriction analysis identified the construction of the recombinant plasmid and the inserted sequence was identical with that published on GenBank. Western blot and immunofluorescence confirmed the expression of the recombinant protein in transfected HEK-293FT cells. It was concluded that the eukaryotic expression plasmid of human PRX3 was constructed successfully and the recombinant could be expressed efficiently in HEK-293FT cells, which provides a sound basis for the further study on human PRX3.
Cell Line, Transformed ; Cloning, Molecular ; Embryo, Mammalian ; Eukaryotic Cells ; metabolism ; Gene Expression ; Genetic Vectors ; Humans ; Kidney ; cytology ; metabolism ; Peroxidases ; biosynthesis ; genetics ; Peroxiredoxin III ; Peroxiredoxins ; Plasmids ; genetics ; Transfection