Abstract:Neurofibrillary tangle (NFT), paired helical filament (PHF) as the major component, is an important pathological character of Alzheimer's disease. Further evidence suggests that PHF plays an important role in neural cell death, including necrotic and apoptotic death. Induction of reactive oxygen species (ROS) by PHF production may be an important mechanism of neural cell death.

Abstract:As an important kinase in the organism, adenylate kinase (AK) not only plays a crucial role in keeping the energy balance, but also takes part in the procedure of apoptosis. In apoptotic cells the release of AK2 from mitochondrial intermembrane to the cytosol is very common, but its function is still not clear. The latest research of AK in apoptosis and the possible role of AK during apoptosis are reviewed.

Abstract:The development of conventional BLMs (planar lipid bilayers) and later s-BLMs and sb-BLMs, have made it possible for the first time to study, directly, electrical properties and transport phenomena across a 5 nm ultrathin film separating two phases, in particular as models of biomembranes. As a result of extensive studies over the past 4 decades, biomembranes have now been recognized as the basic structure of Nature's sensors and molecular devices. To impart relevant functions in BLMs, a variety of compounds such as ionophores, enzymes, receptors, pigments, tissues, etc. have been embedded. Some of these incorporated compounds cause the BLMs to exhibit non-linear phenomena and photoelectric effects. The self-assembled lipid bilayer, the most crucial component of all biomembranes, is in a liquid-crystalline and dynamic state. Such a system, as we know intuitively, must act as some sort of a transducer capable of gathering information, processing it, and then delivering a response based on this information. Today, planar lipid bilayer research is a matured field of endeavor, as a result of applications of many disciplines and techniques including interfacial chemistry, electrochemistry, patch-clamp techniques, spectroscopy, microelectronics, and others. In membrane reconstitution experiments, for example, the evidence is that intracellular signal transduction begins at membrane receptors. The research area covered in this paper is highly interdisciplinary. Emphasis has been placed on basic research. The past work has been benefited by a cross-fertilization of ideas among various branches of sciences. The biomimetic approach to practical applications is unique and full of exciting possibilities. We can glean the design principles from Nature's successful products and apply them to our research and development from which molecular medicine and advanced biosensors may ultimately depend.

Abstract:During the course of NF-κB dimer activation, IκB kinase(IKK) play a crucial role by phosphorylation of inhibitory κB(IκBs). There are lots of existing forms in cytoplasm about IKK complex, which activate IκBs through different ways. Generally, IKK has two catalytic subunits, IKK-α、IKK-β, which have 52% amino acids identity and similar construction, one regulatory subunit, IKK-γ. Both NF-κB-inducing kinase(NIK) and mitogen -activated protein kinase kinase kinase-1(MEKK₁) are upstream kinases of IKK. MEKK₁ preferentially activates IKK-β,whereas NIK efficiently phosphorylates both IKK-α Ser176 and IKK-β. Through cascade reaction, IκBs are phosphorylated by IKK and dissociated from IκB- NF-κB complex, NF-κB dimer enter the nucleus and activate a series of genes.

Abstract:Keratinocyte growth factor(KGF),a member of fibroblast growth factor(FGF),exerts proliferative and differenting effects on a variety of epithelial cells via binding to a specific FGF receptor. KGF gene expression is subject to positive and negative regulation. A fine balance of the regulation is important for normal function of KGF. Results have suggested that KGF play important roles in several aspects: the development of tissues and organs; prevent wound and facilitate wound healing; involvement in tissues malignant transformation.

Abstract:Structure and function of gap junction as well as the role of pH_i, voltage and cytokines on the modulating of gap junction are introduced. Gap junction is an information channel between the adjacent cells, with the function that delivering message and coordinating the actions of cell groups, but its formation and its function on the physical events need more research. The lower pH_i closes the gap junctional channel and it is explained by “particle-receptor” model. The increase of voltage downregulates the conductance of the channel. Cytokines regulate the permeability of gap junction by changing the synthesis and degradation of connexin, enhancing the phosphatizing the connexin.

Abstract:Cellular senescence contributes greatly to organism aging, organism self-protection, cell carcinogenesis and many significant physiological or pathological processes. The mechanism of cellular senescence could be applied to cancer therapy. The way in these years to seek for the mammalian cellular senescence associated genes is described.

Abstract:There exists a special kind protein, lacking intrinsic enzymatic activity, in signal transduction system, termed scaffold protein. It has ability of binding two or more proteins simultaneously. It permits signal transduction in a specific and efficient manner by binding functionally related proteins into a signalosome.

Abstract:The causes of the proton leak, its contribution to basal metabolic rate (BMR), influencing effect factors, and its physiological significance are reviewed. The mitochondrial proton leak occurs in intact cells and tissues, and divides energy into heat production and ATP turnover. The heat production can account for 20%～30% of BMR. However the ability to allow rapid switching of proton flux from leak to ATP turnover may have an even more important significance than that of heat production. The physiological functions of proton leak are including producing heat, increasing the potential for regulation of metabolism, reducing harmful free radical production and regulating carbon fluxes.

Abstract:The peroxisome proliferator-activated receptors (PPARs), new members of nuclear hormone receptors were discovered in 1990. Recent developments regarding these receptors were reviewed. The specificity of ligands，the differential tissue distribution as well as target genes are described for each of the three isotypes α、β、γ. Their implication in the control of inflammtory response, lipid metabolism, cell proliferation and differentiation, and the possible role played in chronic diseases such as cancer and atherosclerosis are discussed.

Abstract:Strategies for decreasing the formation of inclusion bodies, isolation and resolution of inclusion bodies, and refolding of inclusion body proteins were included. The advances in the principle of refolding, assisting cosolvent and refolding processing were reviewed in detail.

Abstract:The strategic and tactical advantages, principle, screening methods, achievements of RNA as a small molecule drug target were reviewed. The process of RNA maturation, transport, intracellular localization and translation are rich in RNA recognition sites that provide good opportunities for drug binding. Both RNAs and proteins are potential drug-binding sites.

Abstract:TALL-1(tumor necrosis factor and apoptosis ligand-related leukocyte-expressed ligand 1), a member of the TNF family, is a novel cytokine identified recently. The human and mice TALL-1 consist of 285 and 309 amino acids, respectively. It was shown that TALL-1 is a type Ⅱ transmembrane protein which is produced in monocytes and macrophages. A soluble form of human TALL-1(sTALL-1) has a extracellular domain composed of 152 amino acids which corresponds to C-terminal 134～285 site of amino acids. The recombinant human sTALL-1 is functionally involved in stimulating B cell growth, activating NF-κB(nuclear factor-kappa B) and JNK(c-Jun NH₂-terminal kinase), and inhibiting tumor cell growth. Moreover, the overexpression of TALL-1 in transgenic mice can result in severe B cell hyperplasia and autoimmune lupus-like disease. Therefore, TALL-1 functions as a potent regulatory factor in vivo and in vitro.

Abstract:Plant expressed sequence tags (EST) project is a new research area in plant genome. By a single pass large scale sequencing of cDNA libraries, ESTs can be acquired and used to analyze gene expression, organization, construction. Some kinds of plant genome projects, the major research in plant EST project, the fuction of bioinformatics in EST analysis, dbEST and inquiry service, some problems in EST research are discussed.

Abstract:Eph receptors are the largest known family of receptor tyrosine kinases. The Eph receptors and their membrane-attached ligands,the ephrins,show diverse expression patterns during development.Functional studies have demonstrated that Eph receptors and ephrins play important roles in many developmental processes including formation of neuronal network,patterning of neural tube and paraxial mesoderm, guidance of cell migration and axon pathfinding,and vascular formation.Recent studies have also suggested that Eph receptors and ephrins may be involved in carcinogenesis.It is therefore of clinical importance to further analyze the function of these molecules,particularly their function in tumor cell growth,since manipulation of the activities of these molecules may have therapeutic applications.

Abstract:The cDNA encoding glial cell-derived neurotrophic factor (GDNF) was isolated from the human astrocytoma cell BT-325. GDNF was efficiently expressed in E.coli. The recombinant protein was purified. The renaturation occurred in the presence of sodium tetrathionate system. In order to study the biological activity of the recombinant protein, the effects of rhGDNF on the dorsal root ganglion (DRG) of chicken embryo (8 d), motor neurons of rat embryo (14 d) spinal cord and glial cell were explored. The results show that rhGDNF promotes the growth of neurite of DRG, increases the number of survival motor neurons cultured for 3 d, 7 d, 14 d and 21 d, respectively, and promotes glial cell proliferation.

Abstract:ARF GAP is a kind of important regulator of introcellular transport. Recently, a novel human gene has been found from a cDNA library of second trimester human fetal liver. The amino acid sequence encoded by the novel gene has 32% similarity to rat ARF1 GAP, was thus termed as ARFGAP3. Functional studies of the new gene were performed. The full-length cDNA of ARFGAP3 was amplified from the human total placenta RNA by RT-PCR technique, then subcloned into pGEM-T vector and sequenced. The RNA Master blot and multiple tissue Northern blot analysis were used to define the expression profile and the transcript size of ARFGAP3 in human tissues. It was shown that ARFGAP3 was strongly expressed in glands and testis and that ARFGAP3 mRNA existed as only one kind of transcript of 2.7 kb in various human tissues. Then, the expression and purification of the recombinant human ARFGAP3 (rhARFGAP3) were performed. It was demonstrated that rhARFGAP3 exhibited strong GTPase-activating protein (GAP) activity towards the recombinant ARF1 in vitro by an assay of a single round of GTP hydrolysis on recombinant ARF1, and that GAP activity of ARFGAP3 was stimulated by PIP2 and inhibited by PC.

Abstract:Two toxin polypeptides (designated RSAPⅠ and RSAPⅡ) have been isolated from Sea anemone Actinia cari. Toxins isolation was accomplished by freezing-thawing extraction, acetone fractional precipitation, cation exchange chromagraphy on SP Sephadex C 25 and reverse phase HPLC on ODS C₁₈.They were found to be homogenous by SDS-PAGE and MALDI-TOF-MS, and their relative molecular masses are 5 008 and 4 992, respectively. They are found to be active on sodium channel in Guinea pig cardiac cell, RSAPI, increasing inward Na(+) currents, opens the sodium channel; RSAPⅡ, decreasing inward Na(+) currents, inhibits the opening of the sodium channel. The circular dichroic spectra of these two polypeptides are 100% β turn.

Abstract:To Investigate the mechanism of postmenopausal osteoporosis in the metabolism of bone organic matrix，the levels of type Ⅰ collagen mRNA in ovariectomized rats bone tissue were detected by RT-PCR. The activities of bone metalloproteinases(MMPs) were determined by Gel-SDS-PAGE.The amounts of type Ⅰ collagen in bone tissues were estimated by immuno-histochemistry. The expression levels of type Ⅰ collagen mRNA decreased about 26.3%，and the activities of pro-MMP-9 increased markedly in ovariectomized group compared with Sham group. Estrogen increased the levels of type Ⅰ collagen mRNA about 34.1% and obviously reduced the pro-MMP-9 activities compared with OVX group. Ovariectomizing in rats resulted in the decrease of type Ⅰ collagen mRNA expression and the increase of pro-MMP-9 activity，both contributed to the decrease of bone organic matrix and ovariectomized osteoprosis.

Abstract:Polyadenylation of 3′-forming in eukaryote concerns three elements located in precursor mRNA downstream region: efficiency element (EE), position element (PE) and the actual site for cleavage and polyadenylation. Several base sequences of EE and PE have been proposed by many experiments. The secondary structures of 180 yeast genomic downstream regions (200 bases downstream the stop codon) have been analysed in detail. It is showed that about 86% of EE sites and 89% of PE sites are related to the regions of non-paired loop (hairpin loop, bulge loop, interior loop or multi-branched loop) or to connecting single strand. This result suggests that the identifications and actions of EE and PE by trans-factors, to certain extent, are dependent on the structural features of EE and PE in the secondary structure. According to the secondary structure of mRNA, the prediction accuracy of EE and PE sites may be improved.

Abstract:Using hepatopioetin (HPO) as “bait” in an yeast two-hybrid screen resulted in the identification of JAB1(a protein initially as a co-activator of c-Jun) as a putative HPO binding partner.Then the full length cDNA of JAB1 was obtained from human fetal liver cDNA library by PCR for GST-JAB1 expression.The binding assay showed that JAB1 binded to rHPO and HPO expressed in COS7 cells in vitro.

Abstract:79 EST fragments which represents corresponding novel genes were obtained by sequencing and bioinformatics analysis of human fetal kidney cDNA library. Microarray was prepared by using these novel EST fragments by automatic spotting. Expression patters of 79 ESTs of novel genes from human fetal kidney were analyzed in fetal brain and fetal heart tissues of 20-week-age and 26-week-age fetus by performing of cDNA chip hybridization. This provides clues for studying exact functions of the novel genes. 8 genes were obtained which were expressed differentially in the fetal brain and heart of 20-week- and 26-week-age respectively. Then differentially expressed genes were identified by Northern analysis. The more exact function of the novel genes are under study.

Abstract:To study the possibility of oral gene therapy using live attenuated Salmonella. A live attenuated AraA^- autotrophic mutant of Salmonella typhimurium (SL3261) was used as carrier for eukaryotic expression vectors EGFPN1, pCMVmIL-12, pCMVmGM-CSF and was administered orally to BALB/c and C57BL/6 mice. After 6 weeks, these mice were challenged with 4T₁ or Lewis tumor cells respectively. Flow cytometer and laser scanning confocal microscopy were used to detect the expression of GFP in murine tissues. PCR and ELISA were used to detect the integration and expression of mIL-12, mGM-CSF gene. The survival time of mice was also investigated. GFP expression and mIL-12, mGM-CSF gene integration could be detected in murine liver, spleen, intestine, kidney and tumor. The serum level of mIFN-γ，mIL-12 increased significantly in the mIL-12 orally treated mice (P＜0.05); The serum level of mGM-CSF increased also in mGM-CSF orally treated mice (P＜0.05); which resulted in the prolongation of the survival time of those mice, compared with the control (P＜0.05). Oral gene therapy using live attenuated Salmonella has the potential to be a simple, effective and above all, safe way against tumor.

Abstract:Asymmetric synthesis of (R)-cyanohydrins from aldehydes and hydrogen cyanide using (R)-oxynitrilase from almond in micro-aqueous phase was studied by GC chiral analysis. Reaction time, content of acetic acid, reaction medium, water activity, reaction temperature and substrate structure all had remarkable effects on the reaction. (R)-oxynitrilase from almond accommodates a wide variety of aromatic, heterocyclic and aliphatic aldehydes. Benzaldehyde was found to be the best substrate for the (R)-oxynitrilase from almond. At low temperature 0～5℃,both conversion rate and enantiomeric excess of the product were above 99%.

Abstract:The effect of oxyphenamone (Oxy, a new inodilator) on high energy phosphate metabolism of isolated Langendorff perfused rat hearts exposed to 60 minutes of ischemia and 60 minutes of reperfusion injury was determined using phosphorus nuclear magnetic resonance (³¹P NMR) spectroscopy. The results led to a paradox of rapid intracellular pH recovery and a low level of high energy phosphate pools reserve during reperfusion compared to the control. It can be stated that Oxy improves energy consumption and, in turn, improves pH recovery, but also causes a deletion of high energy phosphate pools during reperfusion. The relationship between ions regulation, including H^＋ regulation, and myocardial functions recovery may display a key factor for cell survival during postischemic reperfusion.

Abstract:A novel method was developed to prepare for PCR- mediated DNA footprinting walking map. This method is based on the amplification of the cleaved strands using multi-specific primers to walk along the whole DNA template. At first, long enough target DNA fragment without being labeled is incubated in the presence or absence of nuclear proteins and cleaved randomly on average once by either a chemical reagent or an enzyme such as DNaseⅠ. Then the labeled multi-specific primers (sense or antisense) are utilized to amplify the cleaved DNA template to produce labeled single strands which can walk along the whole template. At last, the single strand DNA is separated by electrophoresis in a denaturing polyacrylamide gel, followed by autoradiography analysis. The result is the appearance of a “gap” in the sequence ladder of protein-combined DNA compared to the pattern of unbound or unprotected DNA. This method can footprint walking along any length of DNA template in one time theoretically if enough specific primers are utilized. This method was applied to investigate the partial footprinting map of 5′ flanking region from －1190 to －273 of human stem cell factor gene.

Abstract:To evaluate reasonably the neurotrophic action of small molecule. MTT, differentiation quantity and image analysis methods were used, with the FK506 and GPI1046 as positive control chemicals. A PC12 cell survival and differentiation based screening system were established, in which FK506 and GPI1046 sharply enhanced the neurotrophic action of NGF both in survival and differentiation model just as expected. The system will be helpful in screening neurotrophic novel compounds from chemical library produced by combinatorial chemistry.

Abstract:The RRTR can linearly increase cDNA products via repeated reverse transcription reactions controlled by temperatures and catalyzed by FD-TRT (FD thermostable reverse transcriptase). The stability of RNA in RRTR is proved via the Northern blot, and the increase in cDNA products along with that in reacting cycles is shown via the Dot hybridization. After components of the reaction system are optimized, the RRTR combined with PCR is applied in quantification of HCV RNA through changing the reacting cycles of the RRTR. It can be concluded that due to the linearity of RRTR, the RRTR is a convenient method to enhance the sensitivity and efficacy of RNA related study. It can be used in some fields which full-length cDNA is not required, such as quantification of RNA or cDNA, RACE （rapid amplification of cDNA end), and different display.

Abstract:The significant results are gene expression when cytokine acts on receptors. To study genes related with IL-6 effect, here a simple and rapid PCR-based protocol is utilized to detect and isolate cDNA fragments of the differential expressed genes between the IL-6 treated and untreated Sko007 cells. To generate cDNAs from most internal regions of mRNAs by the reverse transcription using three fully degenerated 6-mer oligonucleotide as primers. And then PCR amplification of the cDNAs with two or three longer primers that are arbitrary but defined sequences is carried on, The PCR amplification was repeated on the same cDNA templates (first step) with different sets of primers. PCR products were electrophoresed in the 2% agarose gel and then the recovered different fragments were used directly in cloning and sequencing. 3 different ESTs were obtained, one of which is a novel gene fragment, and its were proved to be IL-6 related genes by reverse Northern blot analysis. Further bioinformatics analysis indicated that new STRF17 fragments are expressed in many tissues and cells.

Abstract:In order to evaluate the role of inherited variation in the BRD7 gene in human nasoparyngeal carcinoma. RT-PCR, SSCP and sequencing were used to identified multiple coding-region single nucleotide polymorphisms (cSNPs), three cSNPs in BRD7 were found in fifty-seven sporadic biopsy specimens and corresponding blood samples from the same NPC patients and two pedigrees of NPC family. One of those cSNPs is A538C transition, which creates a missense change of Asp162Ala in three of NPC patients analyzed and three of fifty healthy individuals. Adjacent to the bromodomain (504～609 nt), another SNP (C450T) behind the start condon (484nt) results in a G133Ter substitution in 87.7% NPC biopsies and corresponding blood samples, all affected family members in the pedigrees and eight of nine susceptible members, the third SNP is synonymous polymorphism (A737G) which is coupled interestingly with the C450T alteration. The latter two forms of SNPs are also observed in eleven of fifty control normal blood samples. This findings suggest that the presence of the two coupled SNPs in BRD7 might be one of important risk factors for the development and/or progression of NPC (P＜0.01), and the BRD7 gene might have two modes of translation and the terminational SNP at 450 nt in BRD7 gene results in the second alternatively truncated isoform.

Abstract:The current study was designed to reveal the preliminary function of NGX6 to NPC cells. In search of mechanisms of NGX6 2-D PAGE was used to identify proteins that were overexpressed in HNE cells that were transfected with NGX6. After staining and image analysis, spots of interest were isolated and subjected to mass spectrometry. Seven proteins such as Fas,ZNF and MHC-ⅡAg were identified. The possible significance of these findings is discussed. NGX6 may exert its effect on NPC cells by many ways.

Abstract:A new assay for indirectly detecting nitric oxide in serum was established. Nitrate was reduced to nitrite by activated spongy cadmium with copper ion on a microscale chromatographic column, then the nitrite was measured by the Griess reaction. The cadmium column has good vitality, high speed and low interference for the reducing reaction at pH 9.7. The reducing rates are 96.4%～100.0%. The linear range of the assay is 0～400 μmol·L^-1, the limit is 1.85 μmol·L^-1. The RSDs are 2.56%～3.46%, and the recoveries are 96.4%～102.2%, 95.2%～101.3% and 98.7%～104.4% for NO^-₃, NO^-₂ and NO^-₃/NO^-₂ (95∶5) respectively. This method is easy, rapid and precise, needing only a small amount of sample and reagents, and can be used in clinical assay.

Abstract:Based on the decline of A₆₃₀ with the clot lyses, the －A₆₃₀ of many samples was detected at the same time with EL311 Auto Reader. It was found that the A₆₃₀ declined linearly with time, and k under different dillutions could reflect the fibrinolytic activity of enzyme. This method costs less time, less reagent and correlated with CLT very well, indicating it was a good and useful method for determination of fibrinolytic enzymes.

Abstract:Two modified one-step acid guanidinium thiocyanate-phenol methods were reported for the isolation of total cellular RNA from mineralized skeletal tissues. The measurement of optical density of RNA extracts showed that the ratios of A₂₆₀ to A₂₈₀ were greater than 1.85, the values of A₂₃₀ decreased to an acceptable level which meant the contaminated carbohydrate was removed mostly from the preparations of total RNA. Bands of 28S RNA and 18S RNA could be seen apparently on the 1 % formaldehyde-agarose gel. Total RNA could be reverse transcripted into cDNA and β-actin and BMP-2 gene were successfully amplified by PCR. The results showed that the total bone tissue RNA was the qualified contemplate for RT-PCR.

Abstract:DNaseⅠ footprinting is labor intensive, time consuming and the target protein should be purified to some extent.Solid-phase DNaseⅠ footprinting can enrich a sequence specific protein by target DNA immobilized onto paramagnetic beads. After DNaseⅠ digestion, separate the products by sequencing PAGE. This new method is quick ,simple and repeatable,which can minimize the exposure of the researcher to radiation. The solid-phase approach may also facilitate the analysis of factors using crude protein mixtures.

Abstract:Polymorphisms in genomic fingerprints generated by arbitrarily primed PCR(AP-PCR) can distinguish between slightly divergent strains of any organism. Sequence polymorphisms detected by genomic fingerprinting can be mapped genetically or used in phylogenetic and population studies, but too many factors affect the AP-PCR result. Sometimes, it is difficult to obtain a perfect genomic fingerprints are difficult. So an optimization project with minimum times of test is needed. The optimization of the amplified conditions included concentration of Mg²⁺，concentration of DNA, concentration of primers were reported. The results show that the optimum amplified condition of AP-PCR are 2 mmol/L MgCl₂, 50 ng of DNA and 0.3 μmol/L of primers. Under these conditions the AP-PCR productions are richness and the most clear.

Abstract:There are two major hurdles on the way of xenotransplantation entering the clinic. The first is the technical issue of being able to overcome the human immune response that leads to rejection of transplanted organs from other species. The second concerns the potential risk of inadvertent transfer of animal viruses such as the much discussed porcine endogenous retroviruses (PERV) which might cross the species barrier and cause disease in human hosts. Many closely monitored clinical trials are needed as an approach to assess the safety and efficacy of using porcine cells, tissues, or organs therapeutically in humans.