Objective To investigate the expression of urokinase-type plasminogen activator (uPA) mRNA in gastric cancer tissues and cancer-adjacent tissues and the relationship between its expression and biologic behavior of tumor. Methods Fourty-eight cases with gastric cancer were detected for the expression of uPA mRNA by fluorogenic probe quantitative reverse transcription polymerase chain reaction (RT-PCR). Results The positive expression rate of uPA mRNA was 83.3%, 25.0%, 93.8% and 62.5% in gastric cancer tissues,cancer-adjacent tissues, gastric cancer tissues with lymph node metastasis and with non-lymph node metastasis respectively. Expression of uPA mRNA was positively related with the invasion depth of gastric cancer.Conclusion Expression of uPA mRNA is significantly increased in gastric cancer and it can be used as an indicator to judge the metastasis and prognosis of tumor.

AIM:To investigate the depressant effect and mechanism of atorvastatin on the chronic rejection of aortic allograft in rats.METHODS:The models of abdominal aorta transplantation were made with micro-surgery in rats.The recipients were divided into three groups:allograft control group,atorvastatin-treated group and isograft control group.Vascular intimal thickness in all of the groups was observed by histological examination.The expression of proliferation cell nuclear antigenl(PCNA)and ?-smooth muscle actin(?-SMA)were determined by immunohistochemistry.The content of nitric oxide was measured by nitrate reductase chromatometry.RESULTS:The vascular intimal thickness in atorvastatin-treated group(11.60%?2.40%)was lower than that in allograft control group(34.60%?6.40%,P

Objective: The impact of parenteral fish oil lipid emulsion on liver function and nutritional status of malignant tumors of the liver and gallbladder patients. Methods: From December 2007 to A-pril 2008, 32 post-operative hepatobiliary cancer patients were randomly divided into control and study groups. Two groups were treated with isocaloric, isonitrogenic parenteral nutrition and the study group was added fish oil lipid emulsion. Comparison of plasma protein, glucose, jaundice index, transaminase, ALP and the rate of infection complications was made betweent the two groups. Results: The blood glucose, transaminase and ALP levels were not significantly different between the two groups. But the plasma proteins and bilirubin levels were improved significantly (P < 0.05) with reduced infection complication in the study group. Conclusion : Fish oil lipid emulsion is conducive to the recovery of post-operative liver and gallbladder cancer patients in live function and nutritional status.

The eukaryotic expression of human arresten gene and its effect on the proliferation of in vitro cultured vascular smooth cells (VSMCs) in vitro were investigated. COS-7 cells were transfected with recombinant eukaryotic expression plasmid pSecTag2-AT or control plasmid pSecTag2 mediated by liposome. Forty-eight h after transfection, reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expression of arresten mRNA in the cells, while Western blot assay was applied to detect the expression of arresten protein in concentrated supernatant. Primary VSMCs from thoracic aorta of male Sprague-Dawley rats were cultured using the tissue explant method, and identified by immunohistochemical staining with a smooth muscle-specific anti-alpha-actin monoclonal antibody before serial subcultivation. VSMCs were then co-cultured with the concentrated supernatant and their proliferation was detected using Cell Counting Kit-8 (CCK-8) in vitro. The results showed that RT-PCR revealed that the genome of arresten-transfected cells contained a 449 bp specific fragment of arresten gene, suggesting the successful transfection. Successful protein expression in supernatants was confirmed by Western blot. CCK-8 assay showed that the proliferation of VSMCs were inhibited significantly by arresten protein as compared with control cells (F=40.154, P<0.01). It was concluded that arresten protein expressed in eukaryotic cells can inhibit proliferation of VSMCs effectively in vitro, which would provide possibility to the animal experiments.

In order to investigate the origin of neointimal smooth muscle cells in transplant arteriosclerosis in rat aortic allograft, sex-mismatched bone marrow transplantation was performed from male Wistar rats to female Wistar rats. Four weeks after transplantation, the aortic transplant model was established by means of micro-surgery in rats. The recipients were divided into 4 groups: female Wistar-female Wistar aortic isografts, female SD female Wistar aortic allografts, male SD-male Wistar aortic allografts, female SD-chimera Wistar aortic allografts. Eight weeks after transplantation, aortic grafts were removed at autopsy and processed for histological evaluation and immunohistochemistry. The results indicated that excessive accumulation of alpha-SMA-positive smooth muscle cells resulted in significant neointima formation and vascular lumen stricture in rat aortic allografts. Neointima assay revealed that the neointimal area and NIA/MA ratio of transplanted artery were significantly increased in all of aortic allograft groups as compared with those in aortic isograft group (P<0.01). Neointimal smooth muscle cells were harvested from cryostat sections of aortic allograft by microdissection method. The Sry gene-specific PCR was performed, and the result showed that a distinct DNA band of 225 bp emerged in the male-male aortic allograft group and chimera aortic allograft group respectively, but not in the female-female aortic allograft group. It was suggested that recipient bone-marrow cells, as the origin of neointimal smooth muscle cells, contributed to the pathological neointimal hyperplasia of aortic allograft and transplant arteriosclerosis.

AIM: To express human Arresten gene in eukaryotic cell,and to investigate its effect on the proliferation and migration in vitro of rat primary cultured thoracic aortic vascular smooth cells (VSMCs).METHODS: COS-7 cells were transfected with recombinant eukaryotic expression plasmid pSecTag2-AT or control plasmid pSecTag2 mediated by liposome.48 hours after transfection,polymerase chain reaction(RT-PCR) was used to detect the expression of Arresten mRNA in the cells,while Western blotting assay was applied to detect expressed Arresten protein in concentrated supernatants.VSMCs were then co-cultured with the concentrated supernatants;and its proliferation was detected using cell counting kit-8(CCK-8) in vitro.Migration of VSMCs was assayed by a microchemotaxis chamber and a polycarbonate filter (Transwell's chamber) with pores of 8 ?m in diameter.RESULTS: RT-PCR revealed that the genome of Arresten-transferred cells contained a 449bp specific fragment of Arresten gene.Successful protein expression in supernatants was confirmed by Western blotting.CCK-8 assay showed that the proliferation of VSMCs was inhibited significantly by Arresten protein as compared with control group(P

AIM: To construct prokaryotic expression vector of human angiogenesis inhibitor arresten gene and express recombinant arresten in Escherichia coli. METHODS: Human arresten gene was amplified from recombinant plasmid pGEM-Arr with polymerase chain reaction (PCR), and then cloned into prokaryotic expression vector pRSET by means of recombinant gene technology. The recombinant plasmid pRSET-Arr was transformed into E.coli BL21(DE3), and recombinant arresten was expressed in the bacteria under induction of IPTG. The expressed products were detected by SDS-PAGE analysis. RESULTS: Restriction analysis indicated that the arresten gene was successfully inserted into the expression vector, and DNA sequencing verified that the reading frame of the recombinant vector was correct. Recombinant arresten was successfully expressed in Escherichia coli; its molecular weight was about 26 kD and its amount was approximately 30% of total bacterial proteins.CONCLUSION: The successful construction of prokaryotic expression vector containing human arresten gene and the effective expression of recombinant arresten in Escherichia coli laid the foundation for further study on its biological functions.

Objective To study the depressant effect of atorvastatin on arteriosclerosis of aortic allograft in rats.Methods The models of abdominal aorta transplantation were established in rats with the use of(micro-surgery).The recipients were divided into three groups:allograft control group,allograft experimental group and isograft control group.After 60 days of transplant,vascular intimal thickness(VIT) in all of the groups was observed by histological examination.The expression of PCNA and ?-SMA was determined by(immunohistochemistry).Results The degree of VIT in rats of the allograft experimental group was lower than that in the allograft control group;the VIT area ratio in the allograft control group,allograft experimental group and isograft control group was(12.40?2.65)%,(5.20?6.35)%,and(1.2?1.10)%,respectively,A statistical difference between these groups was observed(P

Objective To investigate the value of resting energy expenditure (REE) monitoring in nutritional support therapy of critical patients on mechanical ventilation.Methods A prospective randomized controlled trial was conducted.Sixty critical patients [acute physiology and chronic health evaluation Ⅱ score (APACHE Ⅱ) ＞ 15] on ventilation admitted to intensive care unit (ICU) of Dalian Friendship Hospital from September 2016 to October 2018 were enrolled.The enrolled patients were randomly divided into Harris-Benedict formula (HB formula) group and indirect energy measurement (metabolic vehicle) group with 30 patients in each group.The HB formula group was used traditional HB formula to determine the energy supply and ratio of nutritional support therapy,and the metabolic vehicle group was regularly measured the energy supply and proportion of nutritional support therapy.Serum albumin (ALB),total protein (TP),lymphocyte ratio,blood glucose,blood gas analysis parameters and REE value were determined at 3,5,7,9,and 11 days of nutritional support therapy.Results The value of REE at 3 days of nutritional support therapy in metabolic vehicle group was significantly higher than that in HB formula group (kJ/d:7 850.4±947.3 vs.6 915.3±875.7,P ＜ 0.05).With the time of nutritional support treatment prolonged,the REE value of metabolic vehicle group was decreased gradually,and after 7 days,the patient's condition was stable and improved,and the REE value tended to be stable gradually,it was significantly lower than that of HB formula group at 11 days (kJ/d:5 046.3 ± 493.3 vs.6 915.3 ± 875.7,P ＜ 0.05).There was no significant difference in blood gas analysis or plasma protein before nutritional support therapy between the two groups.After 5 days of nutritional support therapy,the respiratory function of critical patients in both groups was improved,and the lymphocyte ratio and plasma protein parameters were alleviated.After 11 days of nutrition support therapy,the respiratory function of critical patients in both groups was further improved,the ventilator model was adjusted to continuous positive airway pressure (CPAP) mode,the lymphocyte ratio and plasma protein parameters were improved,and the skin color and elasticity were improved,the granulation of the wound was fresh and healed well,and the plasma protein level was increased obviously,ALB level in metabolic vehicle group was significantly higher than that in HB formula group (g/L:31.8 ± 2.5 vs.26.7 ± 2.3,P ＜ 0.05).In the metabolic vehicle group,REE value was decreased from the maximum level on the 3rd day (k J/d:7 850.4 ± 947.3) to a stable level after 11 days (k J/d:5 046.3 ± 493.3),and its energy ratio changed significantly,from carbohydrate:fat of 77％ ∶ 21％ with protein consumption gradually transition in the early (3 days) to carbohydrates:fat of 56％ ∶ 44％ without protein consumption in the later stage (11 days),which showed the tendency of energy consumption was reasonable.Conclusion The energy metabolism rule of critical patients on ventilation could be determined by using the accurate metabolic vehicle and dynamic monitoring of REE value,which could be used for the implementation of nutritional support therapy.

To express recombinant arresten in Escherichia coli (E. Coli) and investigate its biological activity, prokaryotic expression vector of human arresten gene was constructed by gene engineering. Human arresten gene was amplified from recombinant plasmid pGEMArr by polymerase chain reaction (PCR), and inserted into prokaryotic expression vector pRSET containing T7 promoter. Restriction analysis and DNA sequencing verified that the arresten gene was correctly cloned into the expression vector. The recombinant plasmid pRSETAt was subsequently transformed into E. Coli BL21 (DE3), and the target gene was expressed under induction of IPTG. SDS-PAGE analysis revealed that the recombinant protein with a molecular weight of 29 kD (1 kD=0. 992 1 ku) amounted to 29 % of the total bacterial proteins. After purification and renaturation, the recombinant protein could significantly suppress the proliferation of human umbilical vein endothelial cells (HUVECs). These results suggested that the expression of a biologically active form of human arresten in the pRSET expression system laid a foundation for further study on the mechanistic insight into arresten action on angiogenesis and the development of powerful anti-cancer drugs.