[ ABSTRACT] AIM:To construct a lentiviral vector for stable delivery of the ER-α36 gene and to detect its effect on SGC7901 cell growth.METHODS: The efficient RNAi targeting sequences identified for the ER-α36 gene were screened.The Oligo DNA was synthesized with target sequences and annealed to form double-stranded DNA.Then it was digested by Xho I and EcoR I and connected with GV307 vector to produce LV-ER-α36-RNAi lentiviral vector.PCR was used to screen the positive clones and sequence.The LV-ER-α36-RNAi, pHelper 1.0 and pHelper 2.0 plasmids were co-transfected into 293T cells for producing lentiviral vector and infecting SGC7901 cell line.Fluorescence microscopy, real-time PCR and Western blotting were used to detect the transfection efficiency and gene silencing effect.17β-estrodial at concentration of 1 ×10 -10 mol/L was used to stimulate the recombinant cell line, and the action on the growth of gastric cancer cells and the expression of Src, ERK1/2 and cyclin D1 were determined.RESULTS: DNA sequencing analysis confirmed the identity of recombinant shRNA expression vectors.Immunofluorescence assay demonstrated that transfection efficiency was above 80%.Transfection of LV-ER-α36-RNAi significantly knocked down the expression of ER-α36 at mR-NA and protein levels with tetracycline ( TeT) simulating as revealed by real-time PCR and Western blotting.Compared with control group, the growth of the recombinant cell line declined and the expression of Src, ERK1/2 and cyclin D1 and the activation of Src decreased (P<0.05).CONCLUSION: Lentiviral vectors that silence ER-α36 expression are con-structed successfully and can be used to study the role of ER-α36 in gastric cancer.The ER-α36 is related with many kinds of cancer cell growth, including gastric cancer cells.

Objective To determine the relationship between early postoperative delirium (EPD) and prognosis in patients undergoing non-cardiac surgery. Methods This was a prospective cohort study consisted of 698 patients admitted to postanesthesia care unit, undergoing non-cardiac surgery under general anesthesia, between June and December 2009. The risk factors affecting prognosis were collected. All the patients were assessed for the development of delirium by experienced research staff using Confusion Assessment Method for Intensive Care Unit. The patients were divided into 2 groups according to the occurrence of EPD: EPD group and no EPD (NEPD) group. The postoperative hospital length of stay was made as a major prognostic indicator. Cox proportional hazard regression model was used to analyze the risk factors affecting prognosis. Results Of the 698 patients, 197 (28.2%) developed EPD. The postoperative hospital length of stay was prolonged in group EPD compared with group NEPD. The Cox proportional hazard regression model analysis indicated that EPD was an independent risk factor affecting prognosis. Conclusion EPD is an independent risk factor affecting prognosis in patients undergoing non-cardiac surgery.

Objective To investigate the preemptive analgesic effect and safety of paracoxib sodium in patients under-going endoscopic submucosal dissection(ESD). Methods A total of 80 ASA I or II patients aged 35-65 years undergoing ESD under general anesthesia were randomized into two groups(n=40 each):parecoxib sodium group(group B) was received intrave-nous parecoxib sodium 40 mg (in 5 mL 0.9% sodium chloride solution) 10 min before anesthesia induction and control group (group A)was received 0.9% sodium chloride solution 5 mL instead of parecoxib sodium.At the end of operation,patients in both groups were received 5 mg of dezocine.Blood samples were analyzed for PT,TT,APTT,Fib,PLT and PAgT before induction of an-esthesia,at 30 min and 120 min after operation.Patients'Visual analogue scale(VAS),Numeric sedation scale(NSS),and ad-verse reactions were recorded at the end of the operation,2,4 and 6 h after operation. Results Compared with those before parecoxib sodium administration,the fibrinogen concentration and PAgT were significantly higher in group B at 30 min after the intravenous injection of parecoxib sodium(P<0.05),while there was no significant difference in PT,TT,APTT and platelet count between group B and group A(P>0.05).VAS at the end of operation,2,4 and 6 h after operation were lower in group B(P<0.05),and the patients were more satisfied in group B(P<0.05). Conclusion Parecoxib could temporarily enhance blood co-agulation in patients undergoing ESD and could offer safe and effective analgesia.

Objective To evaluate the effect of acupuncture(manual acupuncture,electroacu-puncture)on prevention and treatment of nausea and vomiting after general anesthesia.Methods By searching CNKI,VIP,China Biomedical Literature Database,Wanfang Medical,Embase,PubMed,Co-chrane Clinical Trial Registry,randomized controlled trial(RCT)on acupuncture combined with antiemetics for the prevention and treatment of nausea and vomiting after general anesthesia were performed.The retrieval period was from the establishment of the database to September 2022.RevMan 5.3 was used for statistical analysis.Results Fifteen studies involving 1 493 patients were included.There were 741 patients in the antiemetic group and 752 patients in the acupuncture combined antiemetic group.The incidence of postoperative nausea in the acupuncture combined with antiemetics group was significantly lower than that in the simple antiemetics group(OR=0.43,95％CI 0.34-0.54,P＜0.001).The incidence of postopera-tive vomiting in the acupuncture combined with antiemetics group was significantly lower than that in the simple antiemetics group(OR=0.55,95％CI 0.45-0.66,P＜0.001).Conclusion The incidence of postoperative nausea and vomiting was lower in patients treated with acupuncture(manual acupuncture,electroacupuncture)combined with antiemetics than with antiemetics alone.

Objective To construct engineered corneal epithelium from embryonic stem cells (ESCs) using Rock inhibitor combined with hypoxia-normoxia culture condition. Methods Human ESC line H1 was induced to differentiate into epithelial-like cells by addition of retinoic acid (RA) and bone morphogenetic protein 4(BMP4) in the differentiation medium under the adherent culture condition. The ESCs derived epithelial-like cells were expanded in the mixed medium of SHEM and KSFM with the mixture ratio of 1 : 2 with or without Rock inhibitor Y27632. The H1 derived epithelial-like cells were seeded on the denuded ammonic membrane to construct engineered corneal epithelium under hypoxia,normoxia and hypoxia-normoxia culture conditions,respectively. The inducted effect of ESCs into epithelial-like cells,the expansion ability of the epithelial-like cells and the characteristics of the constructed engineered corneal epithelium were evaluated by morphological observation, real-time fluorescent quantitative polymerase chain reaction (RTFQ-PCR) and immunofluorescence technology. Results Compared with the control group,the relative expressions of ESCs marker Oct4 mRNA, Notch signaling pathway related factors Notch1 and Jagged1 mRNA,and Wnt signaling pathways related factors c-myc and Cyclin D1 mRNA were significantly reduced, and the relative expressions of cutaneous ectoderm markers p63 and K18 mRNA were significantly increased at day 8 after induction in the induced group,with significant differences between them (t =14.63,20.15,93.50,11.60, 19.30,18.44,22.63;all at P<0.05). Compared with the without Y27632 group,the relative expressions of p63 and K14 mRNA,Notch signal pathway receptor Notch1 and Jagged1 mRNA were significantly increased,and Wnt signaling pathways downstream targeted gene c-myc and CylinD1 mRNA were significantly decreased at day 8 after induction in the Y27632 group,with significant differences between them (t =20.29,59.22,2.90,39.59,5.32,10.14;all at P<0.05),and the relative expression of K18 mRNA in the two groups was not significantly changed(t=1.38,P>0.05). The ESCs derived epithelium and constructed under hypoxia-normoxia culture condition showed more obvious stratification and tighter cell arrangement in comparison with those cells cultured in consistent hypoxia culture condition or normoxia culture condition. Epithelial markers Pan-CK and K18 as well as epithelial progenitor cell markers p63 and K14 expressed in the whole cell layers of the ESCs derived epithelium constructed under hypoxia-normoxia culture condition. Conclusions The addition of Y27632 enhances the proliferation ability of H1 derived epithelial cells and actives Notch signaling pathway and inhibits Wnt signaling pathway. The culture and construction in the expansion medium with Y27632 under the hypoxia-normoxia culture condition can promote the stratification of H1 derived engineered corneal epithelium.

Objective To observe the effect of Shenfu injection on lung injury caused by hemor-rhagic shock(HS)in rats and explore the related potential mechanism.Methods Thirty-six SPF healthy male SD rats,aged 16-17 weeks,weighing 400-600 g,were randomly divided into three groups:sham op-eration group(group SH),HS group(group HS),and Shenfu injection group(group SF),12 rats in each group.In group SH,only the right femoral vein and femoral artery were separated after anesthesia,and ve-nous catheterization was not performed.HS model was established in groups SF and HS.In group HS,liquid resuscitation was performed through an intravenous catheter,and the resuscitation fluid consisted of the auto-blood lost and the compound sodium chloride injection equivalent to 1.5 times the blood loss and 10 ml/kg normal saline.In group SF,the resuscitation fluid consisted of the lost autoblood and the compound sodium chloride injection equivalent to 1.5 times the blood loss and Shenfu injection 10 ml/kg.The whole perfusion time was about 60 minutes.Six rats in the three groups were randomly anesthetized 24 and 48 hours after op-eration.The wet/dry weight ratio(W/D)of lung tissues was detected.The concentrations of interleukin-6(IL-6),IL-17,IL-10,and transforming growth factor-β(TGF-β)were detected by ELISA,the mRNA ex-pression of retinoic acid-related orphan nuclear receptor γt(RORγt),transcription factor forkhead box pro-tein 3(Foxp3),and hypoxia-inducible factor-1α(HIF-1α)in lung tissues were detected by PCR.The pro-tein contents of RORγt,Foxp3,HIF-1α,aquaporin 1(AQP1),and AQP5 in lung tissue were detected by Western blot.Pathological changesunder HE staining light microscope and lung injury scores were observed.Results Compared with 24 hours after operation,W/D,the concentrations of IL-6 and IL-17,mRNA ex-pression and protein content of RORγt and HIF-1α,and lung injury score were significantly decreased(P＜0.05),the concentrations of IL-10,and TGF-β,Foxp3 mRNA expression and protein content,and AQP1 protein content were significantly increased in group SF 48 hours after operation(P＜0.05).Compared with group SH,W/D,the concentrations of IL-6,IL-17,IL-10,and TGF-β,mRNA expression and protein content of RORγt,Foxp 3,and HIF-1α,and lung injury score were significantly increased(P＜0.05),AQP1 and AQP5 protein contents were significantly decreased in groups HS and SF 24 and 48 hours after operation(P＜0.05),and alveolar structure was damaged under light microscope and alveolar interstitium was filled with a large amount of edematous fluid,during which a large number of inflammatory cells infiltra-ted.Compared with group HS,W/D,the concentrations of IL-6 and IL-17,mRNA expression and protein content of RORγt and HIF-1α,and lung injury score were significantly decreased(P＜0.05),the concen-trations of IL-10 and TGF-β,Foxp3 mRNA expression and protein content,AQP1 and AQP5 protein con-tents were significantly increased in group SF 24 and 48 hours after surgery(P＜0.05),and the alveolar structure was improved under light microscope,and edema was reduced,and the number of inflammatory cells was reduced.Conclusion Shenfu injection can regulate the balance between pro-inflammatory factors IL-6 and IL-17,and anti-inflammatory factors IL-10 and TGF-β,increase the protein content of AQP1 and AQP5 in lung tissue,and decrease the W/D and injury score in lung tissue,thus alleviating lung injury in HS rats.The mechanism may be related to the regulation of HIF-1α-RORγt/Foxp3 balance.