With the rapidly development of the biotechnology industry,large quantities of recombinant proteins are needed for specific therapeutic and diagnostic applications.Bacterial cells are most often used for the production of recombinant proteins.However,recombinant proteins expressed in the cytoplasm of bacteria are often misfolded as insoluble inclusion bodies and therefore inactive.To circumvent this problem,several eukaryotic expression systems have also been developed over the years,ranging from yeast to mammalian cell-based technologies.For many mammalian proteins,especially those secreted and modified posttranslationally,a more compatible expression system is highly desirable because proper folding or modification can only be provided with closely related cells,i.e.,mammalian cells.Large scale transient transfection of mammalian cells is a recent and powerful technology for the fast production of milligram amounts of recombinant proteins.Transient expression by means of extrachromosomal replication in COS cells is frequently used to check the functional integrity of genes/plasmids and to produce small quantities of cell supernatants containing the protein of interest.As it is allowed for easy and efficient purification,many recombinant proteins used for therapeutic and structural studies are naturally secreted or engineered to be secreted.The use of a proper signal peptide is one of the major determinants for the efficient secretion of heterologous proteins from mammalian cells.The noncatalytic C-terminal hemopexin-like domain of MMP-2,PEX,can block angiogenesis and tumor growth in vivo.Large quantities of biochemically active recombinant PEX are required for the study of their functions and biochemical properties,as well as for their industrial applications.For this purpose,the rat growth hormone,mouse IgG? chain and MMP-9 signal peptides were used for expression of PEX in COS7 cells,and their secretion efficiencies were compared by Western blotting and ELISA.Western-blotting of PEX protein from culture media,resulted in detection of proteins with the predicted molecular mass,which indicate that all of the signal sequences could direct PEX secretion successfully.The MMP-9 signal peptide seems to be superior to the signal peptides from IgG and rGH both in terms of extracellular yield and in terms of secretion efficiency.Thus,expression of pM9PEX construct resulted in higher yields of extracellular PEX and the majority of the produced PEX was secreted and not trapped intracellularly.To examine whether the observed difference in secretion yields is promoted at the transcriptional level,a RT-PCR analysis was performed at 6 h after transfection.The presence of mRNA transcripts of PEX was observed in all the DNA constructs.Moreover,semiquantitative reverse transcription(RT-PCR)results show that there were no significant differences in the expression levels of PEX among the constructs at 6 h after transfection.Though there was no difference in the expression levels of PEX at an early time point after transfection,the presence of an ER-targeting signal peptide sequence in the expression vector affected the trafficking of expressed proteins in the cells.Hence,the described difference in exported yields is probably promoted at the secretion level,rather than at the transcriptional level.Chick chorioallantoic membrane(CAM)bioassay show that the PEX protein purified from cell culture had biological activity to inhibit the angiogenesis.The MMP-9's signal peptide is used for the first time as leader sequence for secretion of foreign proteins.The results revealed that higher amounts of secreted PEX were obtained when vectors containing MMP-9 signal peptide were used and it is also indicated that MMP-9 signal sequence could be effective on promoting the secretion of other heterologous proteins in eukaryotic cells.

Objective To study the better way of avoidance of hypothalamus injury in surgical management of craniopharyngioma.Methods 40 cases with craniopharyngioma accepted surgery treatment were analyzed respectively.Results The pterional approach was performed in 30 cases, interfornical approach in 8 cases,subfrontal approach in 2 cases.Total removal of tumor was achieved in 35 cases,subtotal removal in 5 cases.One died from extradural hematoma after operation. No one died from hypothalamus injury.Conclusion Some way was key to avoid hypothalamus injury and to achieve a better surgical result, such as proper approach, separating strictly along tumor, identification of pituitary stalk and hypothalamus structure,preservation of perforating arteries to hypothalamus.

Objective To evaluate the methods and consequences of surgical technique in the treatment of Stanford A aortic dissection.Methods 108 patients with type Standford A aortic dissection underwent surgery in our study,including urgent surgery in 53 and selective surgery in 55.The operation was performed under deep hypothennic circulatory arrest (DHCA) in 85 cases.Surgical procedures included ascending and semi arch replacement or total arch replacement (some cases combined with stented graft implanted into the descending aorta),"elephant trunk" procedure.Concomitant procedures included repair of intimal tear in arch or descending aorta,Bentall procedure,aortic valve replacement,Cabrol or modified Cabrol procedure,aortic valvuloplasty,mitral valvuloplasty or mitral valve replacement,tricuspid valvuloplasty and CABG.Results In-hospital mortality was 6.5% (7 of 108 patients).The mor- tality was 7.5% (4 of 53 patients) in urgent surgery group and in elective surgery group was 5.4% (3 of 55 patients).Ninety six percent survived patients were followed up for 1 month to 13.3 years [mean (3.2?1.3) years] and 2 deaths occurred during the fel- low-up period.3 patients underwent re-operatian.Conclusion The choice of surgical procedures depend on the location of intimal tear for Stanford A aortic dissection.The better operative effects can be expected with proper surgical indication,perfecting surgical technique,and enhancing postoperative treatment.

Tripterygium glycosides preparation which extracted from the traditional Chinese herb Tripterygium wilfordii (TWHY), was widely used to treat the autoimmune diseases. Previous works demonstrated that TWHF had potent anti-inflammatory and immunosuppressive properties. But the different quality and high incident rate of side effects of different manufactures inhibited its clinical application. Since TWHF had been generally known to play a therapeutical effect by synergism of multiple constituents, it was necessary to build the relationship between the HPLC fingerprint and bioactivity so as to ensure the quality safety and efficacy. The HPLC fingerprint showed that description and content of peaks from different manufactures were diverse. Only 11 common peaks were found. In this study, mice spleen cells stimulated by Con A were used to test the proliferation inhibition bioactivity of TWHF preparations, which were incubated with 30, 15, 7.5, 3.75, 1.88 and 0.94 mg x L(-1) TWHF preparations for 48 h. The results showed that mice spleen cells proliferation was inhibited by all TWHF preparations significantly compared with the control group, which suggested the TWHF preparations showed immune suppress activity. The TWHF preparations from 7 manufacture showed different IC50 value, which might belong to different contents which showed in the HPLC fingerprint. Moreover, a relationship between the HPLC fingerprint and the bioactivity were established to identify important constituents by grey relational analysis (GRA). The result showed that all the contents were relative with the IC50, especially No. 5 and 10 peaks, but No. 1 peak, which was proved to be triptolide, had few contribute to the inhibition of mice spleen cells proliferation. The study of relationship between the HPLC fingerprint and the IC50 by GRA could help to investigate mechanism of bioactive and provide an evidence for the quantification of multi-constituents.
Animals ; Anti-Inflammatory Agents ; analysis ; pharmacology ; Cell Proliferation ; drug effects ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; analysis ; pharmacology ; Glycosides ; analysis ; pharmacology ; Male ; Mice ; Mice, Inbred BALB C ; Spleen ; cytology ; drug effects ; Tripterygium ; chemistry

Aged, 80 and over ; Antibodies, Monoclonal ; metabolism ; Carcinoma, Squamous Cell ; metabolism ; pathology ; Diagnosis, Differential ; Granuloma ; metabolism ; pathology ; Humans ; Keratins ; immunology ; Male ; Mucin-1 ; metabolism ; Skin Neoplasms ; metabolism ; pathology

Adenocarcinoma ; metabolism ; pathology ; Chromogranin A ; metabolism ; Colonic Neoplasms ; metabolism ; pathology ; Humans ; Immunohistochemistry ; methods ; Proliferating Cell Nuclear Antigen ; metabolism ; Staining and Labeling ; methods ; Vimentin ; metabolism

Abstract: Objective　To investigate the distribution and drug resistance characteristics of pathogenic bacteria in patients with neutropenic acute leukemia (AL) and bloodstream infections (BSI). Methods　The clinical data of 258 neutropenic acute leukemia patients with bloodstream infections, who admitted to Shengjing Hospital of China Medical University from January 2016 to December 2021, were collected and analyzed for pathogenic bacteria and drug resistance. Results A total of 268 strains of pathogenic bacteria were isolated from 258 patients, including 180 strains of gram-negative bacteria (67.16%), 61 strains of gram-positive bacteria (22.76%), and 27 strains of fungi (10.07%). Gram-negative bacteria were mainly Klebsiella pneumoniae (53/268, 19.78%), Escherichia coli (49/268, 18.28%) and Pseudomonas aeruginosa (41/268, 15.30%). Gram-positive bacteria were mainly coagulase negative Staphylococcus (31/268, 11.57%) and Staphylococcus aureus(17/268, 6.34%). The main fungi were Candida tropicalis (25/268, 9.33%). Escherichia coli (33/268, 12.31%) was the most common pathogen isolated from acute myeloid leukemia (AML), followed by Pseudomonas aeruginosa (25/268, 9.33%), coagulase-negative Staphylococcus (18/268, 6.72%) and Candida tropicalis (18/268, 6.72%). Klebsiella pneumoniae (35/268, 13.06%) was the most common pathogen isolated from acute lymphoblastic leukemia (ALL),followed by Pseudomonas aeruginosa (15/268, 5.60%) and Escherichia coli (14/268, 5.22%). The resistance of Gram-negative bacteria to piperacillin/tazobactam, cefoperazone/sulbactam, imipenem, meropenem, ertapenem, amikacin, cefoxitin, amoxicillin/clavulanic acid was low. Gram-positive bacteria were sensitive to linezolid and vancomycin. Candida was sensitive to flucytosine, amphotericin B and itraconazole. Conclusions　In patients with granulosa after AL chemotherapy combined with BSI, the pathogenic bacteria isolated from AML are diverse, and the pathogenic bacteria isolated from ALL are mainly gram-negative bacteria. Pathogenic bacteria have different degrees of drug resistance to commonly used antibacterial drugs, so it is important to strengthen the monitoring of the distribution of pathogenic bacteria and the change of drug resistance and rational use of antibacterial drugs to minimize the death of patients.

Objective:To compare the therapeutic efficacy and safety of Hyper-CVAD/MA regimen and CHOP/CHOP-like regimen in the treatment of peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS). Methods:The 78 primary PTCL-NOS patients who were initially diagnosed and treated in Tianjin Medical University Cancer Institute and Hospital and Tianjin Union Medical Center from June 2004 to June 2012 were retrospectively analyzed. The patients were then divided into two groups:Hyper-CVAD/MA group (n=21) and CHOP/CHOP-like group (n=57). Curative efficacies and toxicities were analyzed by Chi-square test, and survival was estimated by Ka-plan-Meier method. Results: In the Hyper-CVAD/MA group, complete response (CR) was 42.9%, overall response rate (ORR) was 85.7%, median progression-free survival (PFS) was 20 months, and the three-year overall survival (OS) was 56.9%. In the CHOP/CHOP-like group, the CR, ORR, and three-year OS were 28.1%, 59.6%, and 49.6%, respectively, and the median PFS was 13 months. Compara-tive analysis showed that the ORR and three-year OS were statistically significant (P<0.05), but the relapse rates (57.1%versus 77.2%) and three-year OS were similar (P>0.05). The incidence rates ofⅢ/Ⅳneutrocytopenia and thrombocytopenia in Hyper-CVAD/MA group (66.7%and 61.9%, respectively) were significantly higher than those of the CHOP/CHOP-like group (22.8%and 14.0%, respec-tively) (P<0.05). Conclusion:Hyper-CVAD/MA regimen can achieve satisfactory efficacy in parents with PTCL-NOS, and toxicity can be controlled with granulocyte colony stimulating factor (G-CSF).

In order to authenticate the components of antler powder in the market, DNA barcoding technology coupled with cloning method were used. Cytochrome c oxidase subunit I (COI) sequences were obtained according to the DNA barcoding standard operation procedure (SOP). For antler powder with possible mixed components, the cloning method was used to get each COI sequence. 65 COI sequences were successfully obtained from commercial antler powders via sequencing PCR products. The results indicates that only 38% of these samples were derived from Cervus nippon Temminck or Cervus elaphus Linnaeus which is recorded in the 2010 edition of "Chinese Pharmacopoeia", while 62% of them were derived from other species. Rangifer tarandus Linnaeus was the most frequent species among the adulterants. Further analysis showed that some samples collected from different regions, companies and prices, contained adulterants. Analysis of 36 COI sequences obtained by the cloning method showed that C. elaphus and C. nippon were main components. In addition, some samples were marked clearly as antler powder on the label, however, C. elaphus or R. tarandus were their main components. In summary, DNA barcoding can accurately and efficiently distinguish the exact content in the commercial antler powder, which provides a new technique to ensure clinical safety and improve quality control of Chinese traditional medicine

In order to authenticate the components of antler powder in the market, DNA barcoding technology coupled with cloning method were used. Cytochrome c oxidase subunit I (COI) sequences were obtained according to the DNA barcoding standard operation procedure (SOP). For antler powder with possible mixed components, the cloning method was used to get each COI sequence. 65 COI sequences were successfully obtained from commercial antler powders via sequencing PCR products. The results indicates that only 38% of these samples were derived from Cervus nippon Temminck or Cervus elaphus Linnaeus which is recorded in the 2010 edition of "Chinese Pharmacopoeia", while 62% of them were derived from other species. Rangifer tarandus Linnaeus was the most frequent species among the adulterants. Further analysis showed that some samples collected from different regions, companies and prices, contained adulterants. Analysis of 36 COI sequences obtained by the cloning method showed that C. elaphus and C. nippon were main components. In addition, some samples were marked clearly as antler powder on the label, however, C. elaphus or R. tarandus were their main components. In summary, DNA barcoding can accurately and efficiently distinguish the exact content in the commercial antler powder, which provides a new technique to ensure clinical safety and improve quality control of Chinese traditional medicine
Animals ; Antlers ; DNA Barcoding, Taxonomic ; Deer ; Medicine, Chinese Traditional ; Polymerase Chain Reaction ; Powders ; Quality Control