In recent years , the emergence and progress of cancer genomics and targeted therapies have remarkably expanded the application fields of protein-protein interaction in cancer research .Fluorescence resonance energy transfer ( FRET ) is a kind of nonradiation energy transfer technique , which can timing , quantitative , positioning , and dynamically monitor the interaction between protein and protein in living cells .With the discovery of novel fluorescent proteins and the development of FRET -based biosensors , FRET has became an important method for visualizing spatial and temporal dynamics of interactions among biological macromolecules in native environments .This review summarises the recent studies and technological advances that have enhanced the use of FRET tech -nology in cancer basic research , early diagnosis , prognosis evaluation , drug development and other areas .

Objective To study the effect of static magnetic field (SMF) on levels of lipid peroxidation in liver kidney and brain tissues in mice. MethodsThirty mice were randomly assigned to groups A,B,C and D, and exposed to static magnetic fields with four different intensities of(24.6?4.2)mT, (42.0?2.1)mT, (63.5?3.0)mT, (85.1?2.9)mT, respectively, for an average of 4 hours daily for 15 days. Then the mice were sacrificed and the amount of MDA in liver, kidney and brain tissues in mice were measured. ResultsThe amount of MDA were significantly decreased in the liver and kidney in rat exposed to (24.6?4.2)mT, (42.0?2.1)mT MSF as compared with that in the control group( P

BACKGROUND: As the end product of lipid peroxidation, malondialdehyde (MDA) content can be used for assessment lipid peroxidation injury.Glutathione peroxidase (GSH-Px) acts as a free radical scavenger. Currently the effect of static magnetic field on the organism, whether positive or negative, has not been elucidated.OBJECTIVE: To study the effect of static magnetic field on anti-oxidation capacity of mouse hepatic tissues and its intensity dependence for producing such effects.DESIGN: A controlled comparative experiment.SETTING: Laboratories of Medical Physics and Biochemistry of Jiangsu University.MATERIALS: The experiment was conducted in the Laboratories of Medical Physics and Biochemistry of Jiangsu University from January to December 2003. Totally 30 mice of either sex weighing 18-20 g were selected and subjected to magnetic filed exposure using a self-designed ferrite magnet apparatus.METHODS: The mice were equally randomized into normal control group and 4 exposure groups exposed to magnetic field of (24.6±4.2) mT,(42.0±2.1) mT, (63.5±3.0) mT, and (85.1±2.9) mT, respectively. The mice in the 4 exposure groups were exposed to static magnetic field of the specified intensity for 2 hours twice a day, while those in the normal control group were subjected to the sham exposure apparatus without magnetic field at scheduled time points every day. After 15 days of exposure, the mice were sacrificed and the GSH-Px activity and the MDA content in the hepatic tissue were assayed.MAIN OUTCOME MEASURES: GSH-Px activity and MDA content in hepatic tissue of the mice.RESULTS: Thirty mice entered the final analysis without losses. MDA content in (24.6±4.2) mT and (42.0±2.1) mT groups were obviously lower than that in the normal control group [(12.70±0.53), (12.96±0.72), and (17.62±0.91) μmol/g, respectively, F=10.4, 9.89, P ＜ 0.01]. The GSH-Px activity in the hepatic tissue in (24.6±4.2) mT and (42.0±2.1) mT groups were obviously higher than that in the normal control group [(143.36±8.34),(150.69±12.00), (87.51±11.34) μkat/g, respectively, F=10.0, 11.3, P ＜ 0.01].CONCLUSION: Static magnetic field of appropriate intensity can lower MDA content and enhance the GSH-Px activity in the hepatic tissue of mice, and may also improve the activity of antioxidase and reduce the production of lipid peroxidation to diminish the consequent injuries and delay the aging process.

Objective:To evaluate the effect of cryopreservation on clonogenic ability and apoptosis rate of mono-nuclear cells and CD34+cells in umbilical blood (UB), and to choose the index to present the freezing injury and optimize the cryopreservation of UB. Methods:hTe mono-nuclear cells (MNC) and CD34+cells were separated from UB and frozen.Atfer 30 days, they were thawed in warm water. Clonogenic capacity and clonogenic recovery before and atfer the cryopreservation was compared. We also used Annexin V-FITC-PI to investigate the apoptosis rate of the cells before and atfer the cryopreservation of these 2 types of cells. Results:hTe number of colony forming unit-granulocyte/monocyte (CFU-GMs) was not changed atfer freezing and thawing in both MNCs and CD34+cells, while the number of colony forming unit-granulocyte, erythrocyte, monocyte and megakaryocyte (CFU-GEMM) was obviously reduced after freezing in CD34+cells. The 2 types of cryopreserved cells had certain degree of apoptosis before the cryopreservation. MNC-type cryopreservation increased the cells apoptosis a little, while CD34+-type cryopreservation increased more. Conclusion:hTe cells have certain degree of apoptosis before the cryopreservation. hTe freezing and thawing procedure does affect the early stage progenitor cells-CFU-GEMM in the CD34+-type cryopreserved cells in UB. hTe damage may be induced by the cell apoptosis.

【Objective】To observe the dynamic interference of cellular endotoxin specific receptors of Toll-like receptor 4(TLR4) mRNA and nuclear factor ?B p65(NF-?B p65) in rats with damp-heat syndrome of seasonal febrile disease by Ganlu Xiaodu Dan(GXD),and to explore the therapeutic mechanism of prescriptions with the actions of clearing heat and resolving dampness.【Methods】Wistar rats were randomized into 3 groups: A(normal control),B(models established by feeding of high-fat diet and colon bacillus under the high-temperature and high-humidity condition),and C(rat models treated with GXD).Reverse transcription-polymerase chain reaction(RT-PCR) analysis was used to detect the TLR4 mRNA level in hepatic macrophages and immunohistochemical method was used to detect the activation of NF-?B p65.【Results】The expression of TLR4 mRNA and NF-?B p65 was increased gradually in group B 6,12 and 24 hours after infection with colon bacillus,and the differences were significant,indicating that the expression of TLR4 mRNA and NF-?B p65 increased with the prolongation of disease course.In group C,TLR4 mRNA expression was decreased in different time points after infection,the difference of TLR4 mRNA expression 12 and 24 hours after infection was significant as compared with group B and group A(P

A low calcium medium developed for epidermal keratinocytes were prepared according to the MCDB 153 modified formula and used in human epidermal keratinocyte culture compared with DMEM culture system. The observation by contrast microscopy and electron microscopy showed that in the low calcium medium keratinocytes grew as a monolayer of high proliferation and had many characteristics of basal cells, with a more rounded shape and large intercellular spaces. Increasing the calcium ion concentration in the medium or changing the other culture conditions the cells in these cultures could be induced stratification and terminal differentiation. The results suggest that the growth, proliferation and differentiation of cultured human epidermal keratinocytes can be controlled and regulated someway.

OBJECTIVE: To observe the effects of Guben Huatan Tongmai Recipe (GBHTTMR), a compound Chinese herbal recipe, on expressions of macrophages and cell adhesion molecules (CAMs) of aortic endothelia in rats with syndrome of phlegm blocking blood vessel, and to explore the pathogenesis of the phlegm-pathogen. METHODS: Fifty normal male Wistar rats, 7-week in age, were randomly divided into five groups: normal control group, untreated group, high-dose GBHTTMR-treated group, low-dose GBHTTMR-treated group and simvastatin-treated group, with 10 rats in each group. Syndrome of phlegm blocking blood vessel was induced in rats of the latter 4 groups by feeding the rats with high lipid diet. Levels of blood lipid were compared among the 5 groups. The expressions of macrophages and CAMs in aortic endothelia were tested by immunohistochemical staining method. RESULTS: The level of blood lipid, and the expressions of macrophages and CAMs showed statistical differences between the normal control group and the untreated group (P<0.01), and between the untreated group and the low-, high-dose GBHTTMR-treated and simvastatin-treated groups as well (P<0.05). CONCLUSIONS: GBHTTMR can decrease the level of serum cholesterol and triglycerides, and increase the level of high density lipoprotein. It also can inhibit the expressions of macrophages, intercellular adhesion molecule-1, vascular cell adhesion molecule-1, P-selectin, and E-selectin.

Objective To determine the relation between Marburg I polymorphism of FactorⅦ-activating protease (FSAP) and cerebral infarction,and to analyze whether it is one of the risk factors of cerebral infarction.Methods Single strand conformation polymorphism-polymerase chain reaction (SSCP-PCR) was applied for the polymorphism analysis of FSAP in 159 patients with cerebral infarction and 179 non-cerebral infarction subjects.Results The phenotypes of FSAP in both the patients and the control subjects were wild type GG;no mutant of Marburg I was found. But a new gene mutation was tested, which had not been reported, requiring further investigation. Conclusion Marburg I polymorphism of FSAP may not be associated with cerebral infarction.

BACKGROUND: There still was not any report about inducing stem cells into matured cells to form products in vitro.OBJECTIVE: To induce CD34+ cells of umbilical cord blood to differentiate into mature megakaryocytes, and to investigate the mechanism of production of platelets.DESIGN, TIME AND SETTING: This cytology in vitro study was conducted at the Central Laboratory of Xiangya Hospital and Xiangya Third Hospital from 2004 to 2006. MATERIALS: Umbilical cord was collected from healthy full-term pregnant puerperants at the Xiangya Hospital.METHODS: The CD34+ cells were isolated from umbilical cord blood by magnetic activated cell sorting (MACS) and then cultured in 24-well culture plate at 5x107/L in StemPro-34 serum-free medium, supplemented with L-glutamine, saturated human transferrin, CaCl2, insulin, deionized bovine serum albumin and recombinant human thrombopoietin at 37℃, under 0.05 volume fraction CO2 saturated humidity to be differentiated into megakaryocytes for 14-21 days. Cell medium was absorbed, and centrifuged to obtain supernatant. Samples were centrifuged again, and then supernatant was removed. The remaining was platelet-like particles in cell culture plate. Platelet was isolated from normal platelet-rich plasma.MAIN OUTCOME MEASURES: The following parameters were measured: morphological changes in cultured cells and platelet-like particles in supematant; results of immunohistochemistry; observation results under a microscope; platelet aggregation; CD41 expression.RESULTS: At day 10, silk-like substances were found in megakaryocyte culture medium, with the presence of platelet-sized particles. The production of platelet-sized particles reached a peal at day 16. Cultured cells were strongly positively for platelet-specific antigen GP Ⅱb Ⅲa. Under the optical microscope, mature megakaryocytes were detected, with the presence of some immature megakaryocytes, and platelet-sized particles were found surrounding megakaryocytes. Under the electron microscope, a majority of mature megakaryocytes and a few apoptotic megakaryocytes were detected, and platelet-sized particles in the supernatant had the same size and structure with the platelet in the platelet-rich plasma. Some platelet surfaces were smooth or irregular. Platelet-sized particles in the supematant aggregated in response to thrombin as platelets in normal platelet-rich plasma. Flow cytometry demonstrated that the cultured platelets had the same high expression rate of CD41 as the platelets from platelet rich plasma.CONCLUSION: Umbilical cord blood CD34+ cells can be induced to differentiate into pudfied and mature megakaryocytes and platelets in vitro.

BACKGROUND:Adiponectin possess functions of lowering blood glucose and blood lipids, and improve insulin sensitivity. But, controversy results about the effect of adiponectin on skeletal muscle have been reported.OBJECTIVE:To study the effects of eukaryon expressed adiponectin on the glycogen synthesis and glucose oxidation in skeletal muscle cell strain C2C12 myotubes by transfecting plasmids carrying mouse adiponectin.DESIGN: A controlled experiment.SETTING: The Second Affiliated Hospital of Sun Yat-sen University.MATERIALS: PcDNA3.0 plasmid with mouse adiponectin cDNA, pcDNA3.0-mad (generously presented from Dr. Gong,University of Maryland), C2C12 cell strain (purchased from ATCC, GRL-1722), DMEM high glucose (Gibco), MEM (Hyclone), fetal bovine serum (Hanagzhou Sijiqing), equine serum (Hyclone), lipofectamine 2000 (Invitrogene), G418 (Gibco), rabbit anti-mouse adiponectin IgG (ACRP303-A, Alpha Diagnostic International), chemiluminescence kit (ECL+PLUS,Amersham), SABC instant immunohistochemistry kit (Boster), D-[U-14C] glucose (specific activity 9.25-13.32 GBq/mmol,NEC), scintillation fluid POP, POPOP (SIGMA), liquid scintillation counter (LS3801, Beckman, USA).METHODS:This study was carried out in the Central Laboratory of the Second Affiliated Hospital of Sun Yat-sen University from March to August, 2003. ① After extraction of plasmid, double digest with Xho Ⅰ and Xba Ⅰ and identification with HindⅢ digest were carried out. ② Plasmid pcDNA3.0-mad and pcDNA3.0 blank vector were transfected using liposome to C2C12 cells, and the stably transfected cells were screened by 500 mg/L G418 for 3 weeks, G418 resistant C2C12 cells were thereafter harvested, therefore stable transfected pcDNA3.0-mad and pcDNA 3.0 C2C12 cell strains were established.③ Adiponectin protein expression was determined by Western blot analysis and immunohistochemistry. ④ Glucose oxidation and glycogen synthesis detections were divided into control, vector and pcDNA3.0-mad (mad)group. Each group was further divided into 4 subgroups with 0, 0.5, 5 and 100 nmol/L insulin (n =6), respectively. Detection of glucose oxidation and glycogen synthesis was carried out with 14C-labeled glucose by counting radioactivity of 14CO2 or 14C labeled glycogen with scintillation, respectively.MAIN OUTCOME MEASURES:Changes of glycogen synthesis and glucose oxidation in skeletal muscle cell strain C2C12myotubes.RESULTS: ① Results of plasmid transfection and restrict digest: After plasmid extraction, double digest with Xba Ⅰ and Xho Ⅰ was carried out along with HindⅢ digest identification.Digest fragments were in accordance with expectation.Length of adiponectin cDNA fragment was 781 bp, plasmid fragment was 5 446 bp, adiponectin cDNA was inserted between digest sites (Xho Ⅰ and Xba Ⅰ ) of eukaryotic expression vector pcDNA3.0. ② Plasmid transfection of C2C12 cell and positive clone screening: On the 10th day of G418 media culture screening after transfection, most C2C12 cells died.Positive clone appeared at the 2nd week. G418 resistant C2C12 colonies were harvested at the 3rd week. ③ Western blot and immunohistochemical identifications: Both confirmed that adipoenctin gene was stably transfected into cells in the Mad group, with successful adipoenctin expression. ④ Effect of stably transfected adiponectin gene to myocyte glucose metabolism:The myocyte glycogen synthesis and glucose oxidation increased along with the increasing of insulin concentration. The linear regression analyses of measured myocyte glucose oxidation amount showed that the regression coefficients of the control group, blank vector group and mad group were 23.34, 2;3.23 and 26.06 respectively. This result indicated that in C2C12 cell stably transfected with adiponectin gene, when insulin concentration increased, the acceleration rate of glucose oxidation increasing was higher than other 2 groups. However, no significant difference could be observed in glycogen synthesis and glucose oxidation of C2C12 cells under basic status without insulin stimulus and treatment status with different insulin concentrations between control group, blank vector group and mad group (P＞ 0.05).CONCLUSION: ① We have successfully established stably adiponectin gene transfected C2C12 cell strain with adiponectin protein expression ability. ② Transfection with adiponectin cDNA had no significant effect on the glucose oxidation and glycogen synthesis of C2C12 myotubes.③ The glucose oxidation and glycogen synthesis of C2C12 myotubes increased with the increasing of insulin concentration. ④ Adipoenctin may coordinate with insulin in improving myocyte glucose oxidation and increasing myocyte glucose uptake.