Increasing the production and secretion of endogenous opioid peptide by immune cell can significantly induce myocardial protective effects against ischemia-reperfusion injury. Gene therapy is promising to increase endogenous enkephalin (ENK). However, classical viral and plasmid vectors for gene delivery are hampered by immunogenicity, gene recombination, oncogene activation, the production of antibacterial antibody and changes in physiological gene expression. Minimalistic immunologically defined gene expression (MIDGE) can overcome all the deficients of viral and plasmid vectors. The exon of rat's preproenkephalin (PPENK) gene was amplified by PCR and the fragments were cloned into pEGFP-N1 plasmids. The recombined plasmids were digested with enzymes to obtain a linear vector contained promoter, preproenkephalin gene, RNA stable sequences and oligodesoxy nucleotides (ODNs) added to both ends of the gene vector to protect gene vector from exonuclease degradation. A nuclear localization sequence (NLS) was attached to an ODN to ensure the effective transport to the nucleus and transgene expression. Flow cytometry, laser confocal microscopy and Western blotting demonstrated that PPENK-MIDGE-NLS can transfect leukocyte of rat in vivo, increase the expression of proenkephalin (PENK) in tissue, and the transfection efficiency depends on gene vector's dosage. These results indicate that PPENK-MIDGE-NLS could be an innovative method to protect and treatment of myocardial ischemia-reperfusion injury.
Animals ; Cloning, Molecular ; Enkephalins ; genetics ; Gene Expression ; Genetic Therapy ; Genetic Vectors ; Leukocytes ; Plasmids ; Promoter Regions, Genetic ; Protein Precursors ; genetics ; Rats ; Transfection ; Transgenes

Objective To observe the efficacy of Zhuang’s medicated thread moxibustion plus acupoint thread embedding in treating allergic rhinitis.Method By using random number table, 103 patients with allergic rhinitis were randomized into a treatment group (n=53) and a control group (n=50). The treatment group was intervened by Zhuang’s medicated thread moxibustion and acupoint thread embedding, while the control group was by acupoint thread embedding alone. The subjects were treated for 6 courses with a one-month follow-up. The therapeutic efficacies were compared between the two groups.Result The total effective rates were respectively 94.3% and 80.0% in the treatment group and control group.Conclusion Zhuang’s medicated thread moxibustion plus acupoint thread embedding has significant advantage in improving the symptoms of allergic rhinitis.

Objective:To explore the effect of instant blood glucose monitoring system (FGM) in perioperative blood glucose management in patients with hip fracture complicated with diabetes mellitus, and to provide guidance for perioperative blood glucose management.Methods:A total of 100 patients with hip fracture complicated with diabetes mellitus treated in the Department of Orthopaedics, Gongli Hospital, Pudong New Area, Shanghai from July 2018 to July 2020 were selected as subjects.The patients were divided into experimental group and control group by random digits table method with 50 cases in each group. The patients in both groups adopted the mode of multidisciplinary cooperation.The experimental group was used to monitor blood glucose based on FGM, and traditional blood glucose monitoring was used in the control group. The waiting time before operation, the pain caused by blood glucose acupuncture, the satisfaction of patients and medical staff, the blood glucose level at different time points and the incidence of adverse reactions were compared between the two groups.Results:The waiting time before operation, the proportion of patients with moderate and severe pain after acupuncture and the incidence of hyperglycemia in the experimental group were (2.58 ± 1.30) d, 2% (1/50) and 30% (15/50), respectively, the control group were (3.67 ± 1.59) d, 22% (11/50) and 50% (25/50), respectively. The differences were statistically significant( t value was -2.087, χ2 values were 9.470, 4.170, P<0.05). The satisfaction of patients and medical staff in the experimental group were (46.43 ± 1.87), (46.58 ± 2.10) points, respectively, the control group were (40.67 ± 3.24), (43.84 ± 2.92) points, respectively.The differences were statistically significant( t values were 8.441, 8.087, P<0.05). There was no significant difference in fasting blood glucose, 2-hour postprandial blood glucose and pre-bedtime blood glucose between the two groups( P>0.05). Conclusions:The application of FGM can realize the continuous blood glucose monitoring and management of patients,making the blood glucose reach the standard more quickly and smoothly, meanwhile it may effectively shorten the waiting time before operation and reduce the acupuncture pain of blood glucose monitoring and recognize patients with concealed abnormal blood glucose. The application can improve the satisfaction of patients and medical staff, and promote the rapid recovery of patients as well.

Objective To construct human surfactant protein B (SP-B) gene promoter luciferase reporter plasmids and detect their transcriptional activities in H441 cells.Methods (1)The fragment of SP-B promoter (-218/+ 435 bp) was acquired from human genome DNA by polymerase chain reaction (PCR) amplification and then was inserted into pGM-T vector by the T4 DNA ligase.The vector was transfected into TOP10 E.coli.The positive clone was identified by DNA sequencing.The identified target SP-B promoter sequence was cloned into pGL3-basic vector to construct the recombinant vector pGL3-basic-SP-B-promoter and was identified by enzyme digestion and sequencing; (2)The pGL3-basic-SP-B-promoter vector was converted into pGL4.17-SP-B-promoter vector through enzyme digestion.The identified recombinant vectors and control plasmid pRL-TK were transfected into H441 cells by lipofectamine 2000,and luciferase assays was performed using the dual-luciferase reporter assay system.Results The sequences of SP-B promoter in the recombinant luciferase reporter plasmids were consistent with the one published on Genebank.The firefly/renilla luciferase activity ratio of pGL3-basic/pGL4.17-SP-B-promoter vector (2.8 ± 1.1,66.5±3.8) was significantly higher than pGL3-Basic,pGL4.17 control vector (0.2 ±0.1,4.3 ±0.4) with statistical significance (t =4.182,27.419,P =0.000),respectively.The SP-B promoter activity of pGL4.17-SP-B-promoter vector was significantly higher than pGL3-basic-SP-B-promoter vector (t =27.712,P =0.000).Conclusions The pGL3-basic/pGL4.17-SP-B-promoter vectors are successfully constructed with SP-B promoter activity in H441 cells and pGL4.17-SP-B-promoter vector is the better choice for further study with higher luciferase activity.

Objective To construct two kinds of eukaryotic ccll expression vcctors pIRES2-EGFP-SP-B-C/T 1580 and evaluate their expressions in 293T cells,for the further study of relationship between polymorphism of surfactant protein B (SP-B) gene and bronchopulmonary dysplasia (BPD).Methods The eukaryotic pIRES2-EGFP-SP-B-C/T 1580 expression vectors were constructed by gene recombination,and identified by gene sequencing.The recombinant expression vectors were transfected into 293T cells by lipofectamine2000.The expression of green fluorescence protein in 293T cells was observed by fluorescence microscopy.The mRNAs and proteins of SP-B-C/T 1580 were tested and identified by reverse transcriptionpolymerase chain reaction-restriction fragment length polymorphism(RT-PCR-RFLP) and western blot.Results Two recombinant plasmids contained the complete cDNA of SP-B with the same sequence as in gene bank.The base of SP-B 1580 gene of pIRES2-EGFP-SP-B-C 1580 was C,that of pIRES2-EGFP-SP-B-T 1580 was T.After being transfected into 293T cells,highly efficient expression of SP-B-C/T 1580 gene was detected at mRNA and protein levels.Conclusions The pIRES2-EGFP-SP-B-C/T 1580 eukaryotic cell expression vectors were successfully constructed.

Objective:To understand the continuing care needs of patients based on the App and "hospital-community-home" linkage.Methods:With descriptive qualitative research, a semi-structured in-depth interview was conducted in 15 patients with type 2 diabetes in Gongli Hospital, Pudong New Area, Shanghai City, and community hospital from January to February 2020. Thematic analysis and the software NVivo 10.0 were used for data analysis.Results:Five themes were extracted: desire a "hospital-led,community-implemented, family-supported" model of continuing care; desire an appropriate, convenient and personalized access to information and follow-up; expect continuous, comprehensive, professional health guidance; expect to receive continuing care service led by specialized nurses and managed by multidisciplinary team; expect diabetes App to be simple and practical, satisfying various needs such as post-hospitalization condition monitoring, assessment feedback, health education and health intervention,ensure information security and free use.Conclusions:Patients have a strong willingness to receive continuing care based on the App and "hospital-community-family" linkage. A mechanism of "hospital-community-family" linkage continuing care service with clear division of labor should be established, multidisciplinary cooperation should be strengthened, and team advantages should be utilized. Providing comprehensive, professional and evidence-based continuing care services for patients, while further optimizing software functions and focusing on information security construction and following evidence-based guidelines to standardize App content in order to adapt to more patient characteristics and needs.