Nowadays,small peptides are always expressed in the form of fusion protein.The expression product contains many superfluous amino acids which can affect the biological functions of small peptides even expressed by GST fusion protein expression system.SUMO protease can cut SUMO fusion protein expressed by fusion expression system without any amino acid residues left on target protein thus become a hot topic in this field.Recombinant His-UlP1/pET3c/BL21(DE3)engineering strain was constructed by genetic engineering technology and the expression conditions were optimized in shake flaks.The process of high density fermentation was explored and different purification conditions were detected by chromatography.The results showed that SUMO protease could be expressed well after inducing the engineering strain by IPTG of 1.0mmol/L at 30℃ for 6 hours.The expression level of the strain in fermentation pots could reach 24.3% analyzed by SDS-PAGE.The purity of SUMO protease was more than 98% after further purification by cation exchange chromatography.The yield was 355mg SUMO protease per liter fermentation liquid.Western blot analysis demonstrated that there were immune reactions between IlP1 and 6?His antibodies,so it has established a good foundation for large-scale industralazation in the future.

Enterotoxins ; analysis ; Humans ; Microbial Sensitivity Tests ; Staphylococcal Food Poisoning ; microbiology ; Staphylococcal Protein A ; analysis ; Staphylococcus aureus ; isolation & purification

OBJECTIVETo review the recent research progress in lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) including its protein, ligands, expression and pathophysiological significance. Data sources Information included in this article was identified by searching of PUBMED (1997 - 2006) online resources using the key term LOX-1.

STUDY SELECTIONMainly original milestone articles and critical reviews written by major pioneer investigators of the field were selected.

RESULTSThe key issues related to the LOX-1 protein as well as ligands for LOX-1. Factors regulating the expression of LOX-1 were summarized. The pathophysiological functions of LOX-1 in several diseases were discussed.

CONCLUSIONSIdentification of LOX-1 and a definition of its biological role in pathophysiologic states provide deeper insight into the pathogenesis of some cardiovascular diseases especially in atherosclerosis and provide a potential selective therapeutic approach. LOX-1 is unlocking and drugs targeting LOX-1 might be a promising direction to explore.

Animals ; Arthritis, Rheumatoid ; etiology ; Atherosclerosis ; etiology ; Humans ; Ligands ; Myocardial Infarction ; etiology ; Osteoarthritis ; etiology ; Scavenger Receptors, Class E ; chemistry ; genetics ; physiology

Objective To observe the sequence of fat deposit and its relationship with insulin resistance in SD rats fed by high fat diet.Methods Normal 8-week-old male SD rats were randomly divided into normal chow (NC,n=40)and high fat diet(HF,n=40)groups.Triglyceride(TG)in serum,liver and muscle were measured;glucose infusion rate(GIR)and the mRNA level of genes related to lipid metabolism in liver and muscle were determined in different stages.GIR was detected by eugiyeemic-hyperinsulinemic clamp for evaluating the insulin sensitivity.Gene expression was determined by real-time PCR.Results(1)As compared with NC group,serum TG was not increased after high fat feeding for4 and 8 weeks,it began to increase after 12 weeks [0.52(0.15-1.00) mmol/L vs O.31(0.09-0.53)retool/L, P0.05)in skeletal muscle.After 8 weeks,the expression of ACC1 in liver in HF group was increased by 20.6%,CPT-1 was decreased by 27.1%(P

OBJECTIVETo evaluate the role of human leukocyte antigen G (HLA-G) in the better effect of allogenetic bone marrow transplantation than that of peripheral blood stem cell transplantation.

METHODSFlow cytometry was used to detect the expression of membrane-bound HLA-G (mHLA-G) on donor peripheral blood (PBC) or bone marrow (BM) mononuclear cells. The levels of soluble HLA-G (sHLA-G) in the plasma and bone marrow fluid were determined using enzyme-linked immunosorbent assay (ELISA) before and after granulocyte colony-stimulating factor (G-CSF) mobilization.

RESULTSThe mean levels of mHLA-G after G-CSF mobilization in the PBC and BM were significantly higher than that before G-CSF mobilization (P=0.001 and 0.000), but the plasma levels of sHLA-G showed no significant changes after the mobilization (P=0.279). The mean levels of sHLA-G in the BM fluid significantly increased (P=0.002) to a level higher than that in the PBC after G-CSF mobilization (P=0.004).

CONCLUSIONHLA-G plays an important role in immune tolerance after hematopoietic stem cell transplantation with G-CSF mobilization.

Adult ; Bone Marrow Transplantation ; immunology ; Female ; Granulocyte Colony-Stimulating Factor ; pharmacology ; HLA Antigens ; immunology ; metabolism ; HLA-G Antigens ; Hematopoietic Stem Cell Mobilization ; methods ; Hematopoietic Stem Cell Transplantation ; Histocompatibility Antigens Class I ; immunology ; metabolism ; Humans ; Immune Tolerance ; Male ; Middle Aged

OBJECTIVETo establish strategies for preventing graft versus host disease (GVHD) and reducing treatment associated morbidity while preserving graft versus leukemia (GVL) effect in nonmyeloablative allogeneic bone marrow transplantation (allo-BMT), with or without donor lymphocyte infusion (DLI) after BMT.

METHODS3 x 10(7) bone marrow cells mixed with 1 x 10(7) spleen cells from the same BALB/c mouse were transplanted into the nonablative irradiated inbred 615 mouse which received a single subcutaneous injection of 1 x 10(6) L615 leukemia cells three days before. The experiments were designed as follows (ten mice in each group): myeloablative BMT control group (group A), nonmyeloablative conditioning without BMT group (group B), nonmyeloablative BMT group (group C), and nonmyeloablative BMT + DLI group (group D). GVL effects were assessed by survival time, white blood cell count and L615 cells in peripheral blood and histologic changes. GVHD was assessed by signs of weight loss, ruffled fur, diarrhea and histologic changes of skin, liver and small intestines. Chimerism was detected by cytogenetic analysis and PCR technique.

RESULTSThe survival time of group A, B, C and D was (20.3 +/- 13.4), (15.9 +/- 1.1), (21.6 +/- 1.7) and (37.8 +/- 2.0) days, respectively, being no significant difference between group A and group C (P > 0.05). The survival time of group C was longer than that of group B (P < 0.01). And among group B, C and D, group D had the longest survival time (P < 0.01). GVHD signs and histologic changes were observed in 60% of control group mice at + 14 day, but none of group C and group D. 40% of mice in group A died of treatment associated morbidity within two weeks, but none in group C and group D. Allogeneic chimerism was kept in group A, but excluded gradually in group C.

CONCLUSIONGVL effect seems preserved in nonmyeloablative BMT mice, but weaker than that in myeloablative BMT mice. GVL effect seems to be enhanced by DLI after nonmyeloablative BMT. GVHD and transplantation associated morbidity seems to be reduced in nonmyeloablative BMT.

Animals ; Bone Marrow Transplantation ; immunology ; methods ; Combined Modality Therapy ; Female ; Graft vs Host Disease ; prevention & control ; Graft vs Leukemia Effect ; Leukemia, Experimental ; therapy ; Leukemia, Lymphoid ; therapy ; Lymphocyte Transfusion ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Inbred Strains ; Transplantation Conditioning ; methods ; Transplantation, Heterologous

This study was aimed to explore the role of human fetal bone marrow stromal cells (FBMSC) in combination with exogenous cytokines in supporting the in vitro expansion of cord blood mononuclear cells and the expression of CXCR4(+) and CD49d(+) in CD34(+) cells. Mononuclear cells (MNC) separated from cord blood (CB) were cultured in a serum-free support culture system with FBMSC or exogenous cytokines or both of them. On day 0, 6, 10 and 14, total cells were counted, CD34(+), CD34(+)CXCR4(+) and CD34(+)CD49d(+) cells were quantitated by FACS, and hematopoietic progenitor cells were assessed by semisolid culture assay. The results showed that after culturing for 14 days, CD34(+) cells, CD34(+)CXCR4(+) cells, CD34(+) CD49d(+) cells and colony forming unit (CFU) were significantly increased (P < 0.05). Compared with other groups, expansion multiple of CD34(+), CD34(+)CXCR4(+), CD34(+)CD49d(+) cells and CFU were higher than that in FBMSC and cytokine group (P < 0.05). It is concluded that the culture system used in this study can not only support the expansion of CB MNCs but also increase the number of hematopoietic stem and progenitor cells which has chemokine and adhesion capacity. This culture system may be a feasible way for in vitro culture of cord blood cells.
Antigens, CD34 ; blood ; Bone Marrow Cells ; cytology ; immunology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Coculture Techniques ; Cytokines ; pharmacology ; Fetal Blood ; cytology ; Fetus ; Flow Cytometry ; Granulocyte-Macrophage Colony-Stimulating Factor ; pharmacology ; Humans ; Integrin alpha4 ; blood ; Interleukin-3 ; pharmacology ; Leukocytes, Mononuclear ; cytology ; immunology ; Receptors, CXCR4 ; blood ; Stromal Cells ; cytology ; immunology ; Time Factors

OBJECTIVETo establish a restriction endonuclease pattern which could detect and differentiate four major human herpesviruses by polymerase chain reaction (PCR), restriction fragment length polymorphism (RFLP), DNA cloning and sequence analysis.

METHODSA pair of primer, which was designed according to sequences in well-conserved regions of the DNA polymerase gene in human herpesviruses, was designed to amplify herpes simplex virus type 1 and 2 (HSVI/II), Epstein-Barr virus (EBV) and cytomegalovirus (CMV). Sequences of the primers are as follows: P(1) (5'-CGACTTTGCCAGCCTGACC-3') and P(2) (5'-AGTCCGTGTCCCCGTAGATG-3'). DNA of four strains of standard herpesviruses were amplified by PCR, and further studied by DNA cloning, sequence analysis and RFLP. At last, the authors established the PCR-RFLP technique to differentiate the four different herpesviruses. Meanwhile, 75 clinical blood specimens from infants with suspected viral infection and 38 blood specimens from healthy children were evaluated for herpesviruses DNA or virus-specific IgM antibody by PCR-RFLP or by ELISA.

RESULTSThe PCR amplified products of four human herpesviruses were from 510 bp to 592 bp in length and were analyzed for herpesvirus types with restriction endonuclease technique. The specificity and sensitivity of this PCR-RFLP were examined. There was no cross-reaction with Escherichia coli, Staphylococcus aureus, hepatitis B virus (HBV), Clostridium neoformans and human-genomic DNA and the lowest detection level was 0.1 fg DNA. Among 75 specimens, 23 were positive by PCR and the positive rate was 30.7%, including 13 for CMV, four for EBV, five for HSVII and one for HSVI after restriction enzyme digestion with BamHI and BstUI, while 10 were positive by ELISA and positive rate was 13.3%. All ELISA-positive specimens were likewise positive by PCR. Thirteen of 65 specimens that were ELISA-negative were tested positive by PCR. An infant with CMV infection was determined with viral DNA and virus-specific IgM antibody in blood at 3, 4 and 6 months after birth, respectively. The result showed that she was still CMV DNA-positive in blood whereas IgM antibody was positive only at month 3 after birth. None of the 38 control blood specimens was positive for herpesvirus by this PCR-RFLP or by ELISA.

CONCLUSIONSThis PCR-RFLP technique was specific, sensitive, rapid and accurate in diagnosing herpesviruses infection in infants, and it could detect herpesviruses DNA in specimens which were negative for IgM antibody by ELISA.

Antibodies, Viral ; blood ; Cytomegalovirus ; genetics ; Enzyme-Linked Immunosorbent Assay ; Herpesviridae ; genetics ; Herpesvirus 4, Human ; genetics ; Humans ; Immunoglobulin M ; blood ; Infant ; Infant, Newborn ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Sequence Analysis, DNA

BACKGROUNDMenopausal symptoms and sleep difficulty were physiological processes that were affected by genetic and other factors. This study was to investigate the prevalence of menopausal symptoms and sleep quality in menopausal transition (MT) and postmenopause (PM) women in Taiyuan, Shanxi.

METHODSA community-based survey of women's menopausal symptoms and sleep quality was conducted between July 2012 and May 2013 at six municipal districts of Taiyuan, Shanxi. A sample of 2429 women aged 40-59 years was divided into four groups: early MT, late MT, early PM, and late PM. Sleep quality in the past 2 weeks before the interview was recorded. The data were analyzed using SPSS 16.0.

RESULTSThe prevalence of menopausal symptoms was 49.8%. Mild, moderate, and severe symptoms were observed in 28.9%, 18.5%, and 2.5% of participants, respectively. The highest prevalence of menopausal symptoms occurred in the early postmenopausal stage; the subsequences were the late postmenopausal stage and the early MT stage. Interestingly, among the 13 items of modified Kupperman index, the five most common symptoms were fatigue, arthralgia and myalgia, decreased libido, insomnia, and nervousness. Meanwhile, 55% perimenopausal women had poor sleep.

CONCLUSIONSMenopausal symptoms are common but mild among women in Taiyuan, Shanxi during MT and PM. In these stages, the prevalence of poor sleep is high.

Adult ; Cross-Sectional Studies ; Female ; Humans ; Logistic Models ; Menopause ; physiology ; Middle Aged ; Postmenopause ; physiology ; Sleep

Adjuvants, Immunologic ; therapeutic use ; Animals ; Antiviral Agents ; pharmacology ; therapeutic use ; Drug Design ; Drugs, Chinese Herbal ; therapeutic use ; Humans ; SARS Virus ; chemistry ; drug effects ; Severe Acute Respiratory Syndrome ; drug therapy ; rehabilitation