Specific scAbs could be obtained through biopanning from phage antibody libraries by use of antigens as target molecules. scAbs specific to thrombin were separated from mouse antibody library by the panning strategy of alternating liquid-solid phase in this paper. Thrombin was biotinylated by photobiotin at first, then avidin-coated magnetic beads were utilized to isolate specific scAbs. The eluted phages were amplified and subject to the second round panning in microtiter plate to remove the unspecific reombinant phages. 4 specific scAbs were separated from 23 phage clones after four rounds of alternative panning.

Adult ; Chemical and Drug Induced Liver Injury ; Dimethylformamide ; poisoning ; Humans ; Male

BackgroundThe effects of extremely low frequency electromagnetic fields (ELF-EMFs) on public health have attracted wide attentions.The association of the thermal effect of ELF-EMFs with cancer and ocular tissue damage has been of concern.However,the pathological changes of scleral tissue after exposure to ELF-EMFs as well as the relationship between these changes and myopia are still poorly understood.ObjectiveThe present study was to investigate the molecular pathological changes of human fetal scleral fibroblasts (HFSFs) after exposure to ELF-EMFs in vitro and to explore the possible mechanism in the occurrence and development of myopia.MethodsHFSFs were cultured and passaged and then exposed to 50 Hz electromagnetic fields,and HFSFs that did not receive the irradiation of ELF-EMFs were used as the control group.The expression of collagen type Ⅰ (COL1A1 ) mRNA and matrix metalloproteinase-2 (MMP-2) mRNA in cultured HFSFs were detected by real-time qualitative polymerase chain reaction (real-time PCR) under different magnetic field intensites (0,0.1,0.2,0.5,1.0 mT) and different exposure time (0,6,12,24,36,48 hours).Cell proliferation assay of HFSFs was detected by the cell counting kit 8 ( CCK8 ) assay.The expression levels of COL1 A1 and MMP-2 proteins in HFSFs were further confirmed by immunofluorescence staining.Results The expression of COL1A1 mRNA was significantly down-regulated under the exposure of 0.2 mT ELF-EMFs for 6 hours,in comparison with the control group;moreover,it decreased in parallel with the increased of flux density (0.099±0.008 vs.0.050±0.004) (P =0.009 ).The expression of MMP-2mRNA was up-regulated conspicuously after exposure to 0.1 mT ELF-EMFs for 24 hours,and it increased with exposure time in comparison with the control group ( 0.009 ±0.001 vs.0.018±0.003 ) ( P =0.038 ).Proliferation of HFSFs (A450) was inhibited following the exposure to 0.2 mT ELF-EMFs for 24 hours in comparison with the control group (P =0.009 ).The expression of COL1 A1 in the experimental group was decreased,compared with the control group,but the expression of MMP-2 was increased.ConclusionsELF-EMFs inhibit the proliferation of HFSFs and expression of COL1 A1 in HFSFs,which might be one of the reasons for the development of myopia.

This study is to evaluate the therapeutic effect of fibroblast growth factor 21 (FGF21) on NAFLD in MSG-IR mice and to provide mechanism insights into its therapeutic effect. The MSG-IR mice with insulin resistance were treated with high dose (0.1 micromol.kg-1d-1) and low dose (0.025 micromol.kg-1d-1) of FGF21 once a day for 5 weeks. Body weight was measured weekly. At the end of the experiment, serum lipids, insulin and aminotransferases were measured. Hepatic steatosis was observed. The expression of key genes regulating energy metabolism were detected by real-time PCR. The results showed that after 5 weeks treatment, both doses of FGF21 reduced body weight (P<0.01), corrected dyslipidemia (P<0.01), reversed steatosis and restored the liver morphology in the MSG model mice and significantly ameliorated insulin resistance. Additionally, real-time PCR showed that FGF21 significantly reduced transcription levels of fat synthetic genes, decreased fat synthesis and promoted lipolysis and energy metabolism by up-regulating key genes of lipolysis, thereby liver fat accumulation was reduced and liver function was restored to normal levels. In conclusion, FGF21 significantly reduces body weight of the MSG-IR mice, ameliorates insulin resistance, reverses hepatic steatosis. These findings provide a theoretical support for clinical application of FGF21 as a novel therapeutics for treatment of NAFLD.
Animals ; Body Weight ; drug effects ; Dose-Response Relationship, Drug ; Dyslipidemias ; metabolism ; Energy Metabolism ; drug effects ; Fatty Liver ; chemically induced ; complications ; Female ; Fibroblast Growth Factors ; administration & dosage ; pharmacology ; therapeutic use ; Insulin Resistance ; Lipolysis ; drug effects ; Liver ; metabolism ; pathology ; Male ; Mice ; Non-alcoholic Fatty Liver Disease ; drug therapy ; Sodium Glutamate

Antineoplastic Agents ; therapeutic use ; Carcinoma, Non-Small-Cell Lung ; drug therapy ; genetics ; Drug Delivery Systems ; Erlotinib Hydrochloride ; Female ; Genes, erbB-1 ; Humans ; Lung Neoplasms ; drug therapy ; genetics ; Male ; Mutation ; Protein Kinase Inhibitors ; therapeutic use ; Quinazolines ; therapeutic use ; Receptor, Epidermal Growth Factor ; antagonists & inhibitors ; genetics

OBJECTIVETo analyze the differential protein expression of left-sided colon cancer and right-sided colon cancer.

METHODSTissue samples of left-sided colon cancer (n=7) and right-sided colon cancer (n=7) were collected. Tissue protein was abstracted and two-dimensional gel electrophoresis (2-DE) was used to examine the gel images. Peptide mass fingerprintings (PMF) was acquired by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and the proteins were identified by data searching with bioinformatics. Immunohistochemical SP method was used for the detection of glucose regulated protein 78 kD (GRP78) and heat shock protein 20 (HSP20) in left-sided colon cancer (n=50) and right-sided colon cancer (n=50) tissues.

RESULTSSixteen differentiating protein spots were identified. Compared with right-sided colon cancer, 10 proteins including GRP78 up-regulated and 6 proteins including HSP20 down-regulated in left-sided colon cancer. Immunohistochemical detection showed that in left and right sided colon cancer, the positive expression rate of GRP78 was 78% (39/50) and 56% (28/50) and the positive expression rate of HSP20 was 34% (17/50) and 72% (36/50), respectively, and the differences were statistically significant (both P<0.05). The positive rate of GRP78 was associated with tumor differentiation, infiltration layer, TNM staging, lymph node metastasis, and liver metastasis, while the positive rate of HSP20 was associated with tumor gross morphology, TNM staging, and lymph node metastasis (P<0.05).

CONCLUSIONSThere are differentially expressed proteins between left-sided colon cancer and right-sided colon cancer, especially for GRP78 and HSP20, which may be the cause leading to the biological differences between left-sided and right-sided colon cancer.

Adult ; Aged ; Colonic Neoplasms ; metabolism ; Female ; Gene Expression Regulation, Neoplastic ; HSP20 Heat-Shock Proteins ; metabolism ; Heat-Shock Proteins ; metabolism ; Humans ; Male ; Middle Aged

OBJECTIVETo observe dynamically the development of fetal long bone and detect the expression and distribution of HIF-1alpha,to investigate the expression pattern and possible effects of hypoxia inducible factor-1alpha (HIF-1alpha) in fetal long bone development of mouse.

METHODSE12.5, E13.5, E14.5, E15.5, E16.5 and E17.5 pregnant C57BL6 mice were sacrificed. After sacrifice, the embryos were delivered by caesarean section. The development of fetal long bone was dynamically observed by stereoscopic microscope, and the distributional expression of HIF-1alpha protein was detected by using method of immunohistochemistry. The expression of HIF-1alpha mRNA and osteoblast marker gene at various stage were also detected by using methods of reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSThe cartilaginous long bone began to form and joints outline arised at E13.5, then the primary ossification center was observed at E14.5, showing opaque ossification under stereoscopic microscope,and then the osteogenesis expanded and extended to both sides. Immunohistochemistry demonstrated lots of HIF-1alpha protein positive chondrcytes in the center of primary ossification at E14.5, then they decreased dramatically. HIF-1alpha mRNA expressed at high level from E13.5 to E15.5, and then decreased to low level.

CONCLUSIONFetal long bone development pattern appeared to be endochondral osteogenisis process, existing hypoxia microenviroment may increase HIF-1alpha mRNA expression and thus initiate the cascade of endochondral osteogenisis.

Animals ; Bone Development ; Female ; Hypoxia-Inducible Factor 1, alpha Subunit ; analysis ; genetics ; physiology ; Immunohistochemistry ; Male ; Mice ; RNA, Messenger ; analysis

Nomination, apparatus, manipulating techniques, indications and theoretical basis of 14 single moxibustion styles including Chuijiu (insufflating moxibustion), Dianjiubi Jiu (pecking moxibustion with a pen-like stool), Jiujia Xunjiu (moxibustion with frame), Tongmai Wenyang Jiu (moxibustion for removing meridian obstructions and warming up yang), QifuJiu (moxibustion on umbilicus and abdomen), Xiongyang Jiu (moxibustion on the chest for reinforcing yang qi), Toujing Jiu (moxibustion on head and neck), Anmo Jiu (moxibustion with massage), Zhiti Jiu (moxibustion on extremities), Guan Jiu (moxibustion with a tube), Zu Jiu (moxibustion on foot), Wenzhenjiu (warm needling), Huanong Jiu (festering moxibustion) and Gewu Jiu (indirect moxibustion) are expounded in this article. And 10 compound moxibustion with the combination of 2 or more than 2 above mentioned single moxibustion style under the instruction of combination of local and distal points, combination of upper and lower points as well as combination of frontal and back points are also stated. It suggests to classify moxibustion into categories of festering moxibustion and mild moxibustion, indirect moxibustion and direct moxibustion, and to classify moxibustion apparatus into the categories of treating tools and assisting tools.
Acupuncture ; education ; history ; Acupuncture Therapy ; history ; methods ; China ; History, 19th Century ; History, 20th Century ; History, 21st Century ; Humans ; Moxibustion ; history ; methods ; Schools

OBJECTIVETo investigate the interventional effect of comprehensive behaviour psychotherapy on the ejaculatory latency of premature ejaculation (PE) patients, sexual satisfaction of sexual partners, as well as its influence on the results of clinical treatment.

METHODSNinety PE patients were randomly divided into a psychological intervention group (n = 45) and a control group (n = 45). Both groups were given medicine therapy, and the former also received comprehensive behaviour psychotherapy for 6 weeks. All the patients were assessed with the Chinese index of sexual function for PE and ejaculatory latency in the vagina, and the clinical efficacy was compared between the two groups.

RESULTSBefore treatment, the ejaculatory latency in the vagina was (0.69 +/- 0.25) min and (0.71 +/- 0.19) min respectively in the intervention and the control groups, as compared with (5.87 +/- 0.59) min and (4.76 +/- 0.54) min before treatment, with significant difference (P < 0.01). In the intervention group, the scores in the control of ejaculatory reflex, the sexual satisfaction of the patients and their sexual partners and anxiety or depress in sexual activity in CIPE were higher than in the control group, with significant difference between the two groups (t = 2.12, 2.31, 2.01, 2.24, P < 0.05). The difference in the SAS score after therapy was of significance (P < 0.01). A month after treatment, the effectivity rates of the two groups were 82.9% and 30% respectively, and the difference was significant (P < .01).

CONCLUSIONComprehensive behaviour psychotherapy obviously adds to the clinical efficacy of drugs in the treatment of PE.

Ejaculation ; Humans ; Male ; Psychotherapy ; Sexual Dysfunctions, Psychological ; therapy ; Treatment Outcome

OBJECTIVETo isolate and identify the cancer stem cells from primary human ovarian cancer tissues.

METHODSFresh tumor tissues from five cases of pathologically diagnosed ovarian cancers were taken, minced and then digested with collagenase and hyaluronidase to obtain single cell suspension. The erythrocytes were removed with ACK Lysis buffer. The suspensions were sorted by magnetic activated cell sorting (MACS) using CD133-binding microbeads. Then the sorted CD133(+) cells were verified by flow cytometry. The cells were cultured in serum-free medium supplemented with EGF, bFGF, insulin and BSA, and grew into spheroids. Immunofluorescence, differentiation and tumor formation tests of the cells were performed to characterize the properties of cancer stem cells.

RESULTSThe ovarian cancer stem cells were successfully isolated from primary human ovarian tumors, which formed typical spheroids in serum-free medium and had stronger ability of tumorigenesis. The results of related experiments verified that CD133 positive cells owned the properties of cancer stem cells.

CONCLUSIONSThe ovarian cancer stem cells presenting the characteristics of stemness in vitro and in vivo, have been successfully isolated from primary human ovarian tumor tissues by MACS. The isolated ovarian cancer stem cells could be used in future researches of their biological functions.

AC133 Antigen ; Animals ; Antigens, CD ; metabolism ; Cell Differentiation ; Cell Separation ; methods ; Female ; Flow Cytometry ; methods ; Glycoproteins ; metabolism ; Humans ; Immunomagnetic Separation ; methods ; Mice ; Mice, Inbred NOD ; Mice, SCID ; Neoplasm Transplantation ; Neoplastic Stem Cells ; metabolism ; pathology ; Ovarian Neoplasms ; metabolism ; pathology ; Peptides ; metabolism