Objective To compare the antineoplastic immunological effects of nanoemulsion-encapsulated MAGE1-HSP70 and SEA(the ratio of MAGE1-HSP70 fusion protein to SEA superantigen was 100∶1)as NE(MHS)vaccine as given in different routes,and try to look for a novel and effective immune route.Methods NE(MHS)was prepared using magnetic ultrasound methods,and the size,the encapsulation rate and the stability of the nanoemulsion vaccine were evaluated.C57BL/6 mice were immunized via p.o.,subcut.,i.v.,or i.p.route.The cellular immunocompetence was detected by ELISpot assay and LDH release assay.The therapeutic and tumor challenge assay were examined too.Results When the vaccine was given orally,the tumor masses formed 28 days after B16-MAGE1 inoculation in mice were markedly bigger than that formed in the mice of the other groups(P

Adult ; Faculty ; Female ; Humans ; Lipoproteins ; blood ; Lipoproteins, HDL ; blood ; Lipoproteins, LDL ; blood ; Male ; Middle Aged ; Multivariate Analysis ; Regression Analysis ; Sampling Studies ; Surveys and Questionnaires ; Universities

Objective:To observe the expression of B7-H4 in experimental autoimmune myocarditis (EAM).Methods:BALB/c mice were randomly divided into 2 groups:the control group and the experimental group.The mice of experimental group were injected with myosin to establish EAM models , while the mice of control group were injected with complete Freund 's adjuvant and normal saline.All the mice were killed separately at the 14th,21st,30th and 45th day for lymphocyte proliferation assay ,hematoxylin-eosin staining,immunohistochemical staining and real-time PCR.Results:The inflammation infiltration of heart was most serious at the 14th and 21st day,then it was gradually relieved with time;the results of lymphocyte proliferation assay and real-time PCR were similar to that of the inflammation infiltration of heart ,which were in high level at the 14th and 21st day,and they were both higher than that of the control group ( P<0.05 );B7-H4 protein were only detected in the experimental group ,and it was constantly expressed during the whole experiment on the endothelium of heart with myocarditis.Conclusion:B7-H4 participates in the progress of EAM ,and it may be a new way of studying the mechanism of myocarditis.

Objective To explore the expression and significance of Annexin Ⅱ protein in ovarian adenocar-einoma.Methods The expressions of Annexin Ⅱ protein were detected in 48 cases of ovarian adenocarcinoma and 10 cases of ovarian adenoma by SP immunohistochemistry.Results The positive rates of expression of Annexin Ⅱprotein in ovarian adenoma were 50.00% (5/10), and were 81.25% (39/48) in ovarian adenocareinoma (χ2 =4.414, P <0.05), in well and moderately-differentiated, poorly differentiated ovarian serous adenocarcinoma were 93.33% (14/15) and 78.95% (15/19) respectively (χ2 = 1.383 ,P0.05 ), and on the Ⅰ and Ⅱ stage, Ⅲ and Ⅳ stage of ovarian serous adenocarcinoma were 60.00% (6/10) and 95.83% ( 23/24 ) respectively (χ2=7.226, P <0.05).The positive rate in the group of ovarian serous adenocarcinoma with lymph node metastasis was 90.00% (27/30), while 50.00% (2/4) in the group without lymph node metastasis (χ2=4.502, P<0.05).Conclusion The positive expression of Annexin Ⅱ protein may be correlated with the carcinogenesis, development and lymph node metastasis of ovarian serous adenocarcinoma.

To explore the anti-tumor proliferation activity of human interleukin-29 (hIL-29) variant and based on bioinformatics analyzed data of hIL-29, a mutant gene hIL-29(mut33,35) was amplified by site-directed mutagenesis and megaprimer PCR. The hIL-29(mut33,35) was inserted into an eukaryotic expression plasmid pPIC9K and successfully expressed in Pichia pastoris GS115. A recombinant variant protein (rhIL-29(mut33,35)) was purified from the ferment supernatant of the engineering GS115. To observe the antineoplastic activity of the variant rhIL-29(mut33,35), a CCK-8 reagent was used to detect the anti-proliferation effect. Results show that it has strong anti-proliferation effect when acted on liver cancer cell BEL7402, colon cancer cell HCT8 and gastric cancer cell SGC7901. The inhibition ratios of the three tumor cells were (30.99 ± 1.58)%, (22.47 ± 1.37)% and (32.05 ± 2.02)%, respectively. In high dose group, the anti-proliferation effect of the rhIL-29(mut33,35) was stronger than that of wild type rhIL-29 (P < 0.01). This indicates the variant rhIL-29(mut33,35) has potential development value for medicine.
Antineoplastic Agents ; pharmacology ; Carcinoma, Hepatocellular ; pathology ; Cell Line, Tumor ; drug effects ; Humans ; Interleukins ; biosynthesis ; pharmacology ; Liver Neoplasms ; pathology ; Mutagenesis, Site-Directed ; Pichia ; Plasmids ; Polymerase Chain Reaction ; Recombinant Proteins ; biosynthesis ; pharmacology

9α-hydroxy-4-androstene-3,17-dione (9-OH-AD) is an important intermediate in the steroidal drugs production. 3-ketosteroid-9α-hydroxylase (KSH), a two protein system of KshA and KshB, is a key-enzyme in the microbial steroid ring B-opening pathway. KSH catalyzes the transformation of 4-androstene-3,17-dione (AD) into 9-OH-AD specifically. In the present study, the putative KshA and KshB genes were cloned from Mycobacterium smegmatis mc(2)155 and Gordonia neofelifaecis NRRL B-59395 respectively, and were inserted into the expression vector pNIT, the co-expression plasmids of kshA-kshB were obtained and electroporated into Mycobacterium sp. NRRL B-3805 cells. The recombinants were used to transform steroids, the main product was characterized as 9α-hydroxy-4-androstene-3,17-dione (9-OH-AD), showing that kshA and kshB were expressed successfully. Different from the original strain Mycobacterium sp. NRRL B-3805 that accumulates 4-androstene-3,17-dione, the recombinants accumulates 9α-hydroxy-4-androstene-3,17-dione as the main product. This results indicates that the putative genes kshA, kshB encode active KshA and KshB, respectively. The process of biotransformation was investigated and the results show that phytosterol is the most suitable substrate for biotransformation, kshA and kshB from M. smegmatis mc(2)155 seemed to exhibit high activity, because the resultant recombinant of them catalyzed the biotransformation of phytosterol to 9-OH-AD in a percent conversion of 90%, which was much higher than that of G. neofelifaecis NRRL B-59395. This study on the manipulation of the ksh genes in Mycobacterium sp. NRRL B-3805 provides a new pathway for producing steroid medicines.
Androstenedione ; analogs & derivatives ; biosynthesis ; Bacterial Proteins ; genetics ; metabolism ; Biotransformation ; Ketosteroids ; Mixed Function Oxygenases ; genetics ; metabolism ; Mycobacterium ; metabolism ; Mycobacterium smegmatis ; enzymology ; Plasmids

Objective To investigate arsenic trioxide (As2O3)-mediated apoptosis of synovlal cells in pa-tients with rheumatoid arthritis (RA) through culturing the synoviocytes in vitro. Methods Primary synovial cells were cultured by means of two-enzymatic digestion and the third cells were adopted in this test. The cultured cells were defined by MTT and flow cytometry (FCM). Results Certain concentration of As2O3 could inhibit the viability of synoviocytes at 48 h by means of MTT, which was dose-dependent. Certain concentration of As2O3 could induce the apoptosis of synoviocytos pro rata at 48h by means of FCM ,which was dose-dependent within range of 10-80 μmol/L concentration. Conclusion Certain concentration of As2O3 following 48 h effect could induce the apoptosis of syno-viocytes of RA,which is dose-dependent.

Background Researches demonstrated that CpG ODN,a immunostimulatory sequences,has preventing and treating effect on allergic conjunctivitis caused by protein allergen.However,its effect on allergic conjunctivitis caused by fungal allergen is unclear. Objective This study aimed to investigate into whether the Th1-Th2 switching immunostimulatory CpG ODN could reverse the response in the murine allergic conjunctivitis model caused by aspergillus fumigatus. Methods A mixture of spores and hyphae of aspergillus fumigatus strain was used to induce allergic conjunctivitis in male BALB/C mice aged 6-8 weeks.This experiment was designed into preventive or therapeutical treatment program.Under both settings,allergic conjunctivitis of the animals were treated with CpG ODN,nonstimulatory GpC ODN or PBS.After the last challenge with the allergen,the clinical symptoms of the animals were scored based on the criteria of Magone.The animals were sacrificed and the histopathological examination of conjunctiva was performed.Expression of TLR4 mRNA in conjunctiva was analyzed by real-time PCR assay.The responsiveness and populations of lymphocytes in spleen and draining lymph nodes were analyzed by flow cytometry.The use complied with the Standard of Association for Research in Vision and Ophthalmology. Results In the prevention mode.CpG ODN decreased subconjunctival infiltration compared with GpC ODN and PBS groups with the average neutrophil count index(21.25 ±11.59/section,30.75 ±11.44 section and 69.00±9.90/section,respectively).Expression of TLR4 mRNA was up-regulated significantly by CpG ODN.The clinical scores for CpG ODN group were insignificantly lower than those in GpC ODN group and PBS group(P＞0.05).In the therapeutic mode,compared with GpC ODN and PBS groups,the allergic symptom score in CpG ODN group manifested significantly lower(t=4.000.t=2.750,P＜0.01)and showed fewer cellular infiltration(t=4.870,t=3.829,P＜0.01)and higher expression of TLR4 mRNA(P＜0.01).In cultured splenic and draining lymph node cells,increased percentages of CD4+ CD25+ and CD4+ CD25+ CD69+ in CpG ODN group were observed compared with control groups(|P＜0.05). Conclusion CpG ODN can relieve aspergillus fumigatus-induced allergic conjunctivitis via either subconjunctival injection or topical application by upregulating expression of TLR4 and activating Treg lymphocytes.

Aim To study inhibitory effect of 5-fluorouracil encapsulated by galactosylceramide liposomes (5-Fu-GCL)on 5-Fu-resistent HepG_2 cells and its mechanisms. Methods Inhibitory effect of 5-Fu-GCL on established model of 5-Fu-resistant HepG_2 cells was assessed with MTT assay in vitro. The concentration-time course of 5-Fu-GCL in intracellular fluid was detected with high performance liquid chromatography (HPLC). Thymidylic acid synthase (TS) expression was observed with immunohistochemical method,and NO content was determined with chemical method. Results Obvious inhibitory effects of 5-Fu-GCL (75,150,300,600,1200?mol?L-1) on 5-Fu-resistant HepG_2 cells were observed with IC_ 50 of 158.6 ?mol?L-1,far lower than that of free 5-Fu (400.9 ?mol?L-1). 5-Fu-GCL (300 ?mol?L-1) inhibited 5-Fu-resistant HepG_2 cells in a time-dependent manner,and the inhibitory effect of 5-Fu-GCL was stronger than that of free 5-Fu during 12～48 h. Compared with free 5-Fu,5-Fu-GCL (300 ?mol?L-1) increased the content of intracellular fluid in 5-Fu-resistant HepG_2 cells. 5-Fu-GCL(62.5,300,1200 ?mol?L-1) not only inhibited the expression of TS,but also increased the production of NO in 5-Fu-resistant HepG_2 cells,and these effects of 5-Fu-GCL(300,1200 ?mol?L-1) were stronger than those of free 5-Fu. Conclusion 5-Fu-GCL has inhibitory effect on 5-Fu-resistant HepG_2 cells. The effect may be related to the increased concentration of 5-Fu-GCL in intracellular fluid,inhibited expression of TS and increased production of NO.

To determine the role of allogeneil graft of mesenchymal stem cells in mammalian longevity, mesenchymal stem cells were isolated from BALB/c mouse uterine-incision delivery fetus by two successive density gradient centrifugations, and then were purified and amplified by adherent culture. Identified P1 mesenchymal stem cells were injected (i.v.) through vena caudalis into the 15-month-old female BALB/c mice three times. The mice were evaluated with ultrasoundcardiogram, autopsy, score of cardiac, skin, lung, kidney, colon histopathology and serum total superoxide dismutase activity, maleic dialdehyde content, glutathione peroxidase activity. The results showed that after transplantation, the long-term surviving stem cells were found to be located in many organ tissues with in situ Y chromosomal hybridization dyeing. Median life span was increased in these animals after transplantation. Skin, cardiac, lung, kidney and colon pathology development were delayed. The retrogradation of heart function was attenuated, the increase of heart mass index (the mass of heart/the mass of the body), and serum maleic dialdehyde content, the decrease of spleen mass index (the mass of spleen/the mass of the body), serum total superoxide dismutase activity and glutathione peroxidase activity were reduced three months after transplantation (all P<0.05). These results support the idea that longevity can be enhanced by transplantation of mesenchymal stem cells and reinforce the hypothesis of mesenchymal stem cell as antiager.
Aging ; physiology ; Animals ; Female ; Fetal Stem Cells ; transplantation ; Longevity ; physiology ; Mesenchymal Stem Cell Transplantation ; Mice ; Mice, Inbred BALB C ; Random Allocation