Abstract:Metabonomics is the branch of science concerned with the quantitative understandings of the metabolite complement of integrated living systems and its dynamic responses to the changes of both endogenous factors (such as physiology and development) and exogenous factors (such as environmental factors and xenobiotics). As a holistic approach, metabonomics detects, quantifies and catalogues the time related metabolic processes of an integrated biological system, ultimately, relates such processes to the trajectories of the pathophysiological events. Ever since its birth in 1999, metabonomics has already been described in more than 800 scientific papers and half dozen patents, amongst which almost 700 papers were experimental articles. Now, metabonomics has been established as an extremely powerful analytical tool and hence found successful applications in many research areas including molecular pathology and physiology, drug efficacy and toxicity, gene modifications and functional genomics, and environmental sciences. This holistic approach has thus become an important part of systems biology and has now evolved to be a unique part in global systems biology. The essence of metabonomics and some of the present applications were reviewed to illustrate the rapid development of this extremely exciting new frontier.

Abstract:Mutations in human mitochondrial tRNA genes are responsible for a variety of human inherited diseases. Investigations of the molecular mechanisms of these diseases are of great interest for nowadays scientists. According to the study of post-transcriptional modification patterns of human mitochondrial tRNAs, novel taurine-containing modifications were identified at the anticodon wobble nucleotides of mitochondrial tRNA^Leu(UUR) and tRNA^Lys. Recently, it was reported that mitochondrial tRNAs harboring one of those encephalomyopathies related mutations, such as A8344G, A3243G, T3271C etc., lacked the normal taurine-containing modification at their anticodon wobble positions. Wobble modification deficiencies of mutant mitochondrial tRNAs were found from cybrid cells, as well as from patient tissues. Molecular surgery experiments showed that the wobble modification is essential for the interaction between the anticodon in tRNA and the codon in mRNA. Furthermore, the enzyme that is responsible for the formation of the modification was identified and characterized. These studies strongly suggested a key molecular factor responsible for the inherited mitochondrial encephalomyopathies and could potentially lead to the development of a gene therapy for these diseases.

Abstract:The ubiquitin-proteasome system is composed by multiple enzymes which are ubiquitously expressed in mammalian cells and plays an essential role in a variety of biological processes. As key members of the protein degradation enzymatic system, the function of E3 ubiquitin ligases has been extensively investigated. BMP and TGF-β are critical molecules that regulate proliferation, differentiation and apoptosis of osteoblasts and chondrocytes through different signaling pathways. Resent findings indicate that the ubiquitin-proteasome system functions as a key regulator in bone cells and the E3 ligase mediates the proteolytic degradation of critical molecules in BMP and TGF-β signaling pathways. Recent progress on studies of HECT domain E3 ligase, Smurf, in osteoblasts and chondrocytes were summarized. The regulatory role of Smurf1 and Smurf2 in BMP and TGF-β signaling and osteoblast and chondrocyte function has been reviewed.

Abstract:Photoacoustic tomography is a developing, promising，non-invasive imaging method in the medical clinic diagnosis. It is an ultrasound-mediated biophotonic imaging method based on the intrinsic optical absorption properties of tissue and ultrasonic detection, and combines the merits of both high contrast advantage of pure optical imaging and high resolution advantage of pure ultrasound imaging. Photoacoustic tomography can be performed by detecting photoacoustic waves instead of detecting photons. In photoacoustic tomography, imaging contrast is based primarily on the optical properties of biological tissues, and imaging resolution is based primarily on the ultrasonic waves. It can avoid the influence of optical scattering on imaging resolution in principle, and can provide tomography of tissues with high contrast and high spatial resolution at medium depths. Photoacoustic tomography can provide an effective approach to studying the structures, physiological properties, metabolisms, pathological properties of biological tissues. It has important potential clinical applications in the early non-invasive detection of cancers, structural and functional in vivo imaging. A brief introduction of photoacoustic imaging mechanisms is gives, and the imaging methods, the image reconstruction algorithm and the potential biomedical applications of photoacoustic tomography are reveiewed.

Abstract:To investigate the mechanism of the cellular repressor of E1A-stimulated genes (CREG) on the differentiation of vascular smooth muscle cells (VSMCs), the human internal thoracic artery cells (named HITASY cells) were stably infected with sense-CREG [pLNCX₂(＋)/CREG] and antisense-CREG [pLXSN(－)/CREG] retroviral vectors and positive clone was abtained by G418 selection. The cells infected with pLNCX₂(＋)/CREG take on the differentiated phenotype and overexpression of CREG can enhance SM α-actin expression accompanied with increase of the total RhoA and nuclear serum response factor (SRF) expression detected by immunofluorescence and Western blot. And treated with a selective Rho kinase inhibitor Y-27632, the expression of SM α-actin and nuclear SRF protein induced by CREG was effectively reduced. Meanwhile, the opposite results were observed in the CREG-depression HITASY cells. And immunoprecipitation assays indicate that as a secreted protein, CREG can be detected in the media of pLNCX₂(＋)/CREG cells and can interact with insulin-like growth factorⅡ/mannose 6-phosphate receptor (M6P/IGF2R). Furthermore, that using okadaic acid (an inhibitor of the protein phosphatases 2A) to decrease the number of M6P/IGF2R at the cell surface can significantly inhibit the expression of total RhoA, nuclear SRF and SM α-actin induced by CREG-overexpression. Taken together, as a secreted protein, CREG can enhance the expression of SM α-actin by interaction with M6P/IGF2R and perform a pivotal role in the process of VSMCs differentiation and phenotype modulation.

Abstract:In the post-genomics era in which gene sequences have been decoded, large-scale protein-protein interaction data are generated with the rapid development of system biology experiments. It is important in functional genomics to search for function modules and predict protein functions from the data. A new method called modularized clustering method(MCM), which are based on the direct and second-order interactions of modules, is applied to the latest high-throughput protein-protein network of yeast to predict the function of unknown proteins in the modules. P value of hypergeometric cumulative distribution of modules and the disturbance analysis on the data, including adding, removing and rewiring interactions, are employed to evaluate the prediction quality and robustness of the method. The results show that MCM has high prediction precise rate and coverage, and it is robust to high false-positive data and missing data. The predicted results of unknown proteins with high prediction precise rate can be instructive in biological analysis and the algorithm can be generalized to other networks with the similar structures.

Abstract:It is known that extracorporeal shock wave (SW) may promote healing of fracture. A previous study reported that SW promoted human bone marrow stromal cells (hMSCs) towards osteoblasts in vitro. To study the osteogenesis ability of hMSCs treated by shock wave in porous hydroxyapatite (HA) in vivo, primary hMSCs of SW group and control group were cultured in the porous HA for 2 weeks and then implanted into subcutaneous sites of nude mouse. These implants were harvested and prepared for the biochemical analysis of alkaline phosphatase activity by AKP kit, histological analysis of decalcified and undecalcified sections and morphology by scan electric microscope (SEM), as well as osteocalcin mRNA expression by RT- PCR 4 weeks and 8 weeks after implantation. It showed that cells of SW and control group almost covered the rough surface of HA before implantation and the extracelluar matrix of SW group was abundant by SEM photomicrograph . The histological analysis and SEM photomicrograph showed active bone formation 4 weeks and 8 weeks after implantation, as well as tetracycline labeling under fluoroscopy analysis in SW group. Alkaline phosphatase in supernatants of the implants detected 4 weeks and 8 weeks after implantation in SW group was higher than in control group (P < 0.001). The expression of osteocalcin mRNA was found 4 weeks and 8 weeks after implantation in SW group. The results suggest that postnatal hMSCs treated by SW could form bone tissue in vivo using tissue engineering technique, and this new method would be applied in the field of bone tissue engineering.

Abstract:Abnormally hyperphosphorylation of tau plays a critical role in the pathogenesis of Alzheimer disease (AD) and type 2 diabetes is a known risk factor of AD. Phosphorylation of tau in type 2 diabetic and obese rats was investigated by Western blots. Tau protein was found to be hyperphosphorylated at several AD-related phophorylation sites. The activity of glycogen synthase kinase 3β (GSK-3β), a key component of insulin signal transduction pathway and a known tau kinase, was also found to be increased in the brains of both diabetic and obese rats. Intrahippocampal injection of LiCl blocked activation of GSK-3β in both groups, but only blocked hyperphosphorylation of tau in the obese rats. In addition, the β-subunit of the hippocampal membrane insulin receptor was found to be reduced in the brains of obese and type 2 diabetic rats. These findings suggest that obese and type 2 diabetes increase the probability of AD by increased insulin resistance and consequent upregulation of GSK-3β, which leads to hyperphosphorylation of tau, and that impaired glucose metabolism may also contribute to tau hyperphosphorylation in type 2 diabetes.

Abstract:Triose phosphate isomerase (TIM) was prepared and purified from chicken breast muscle. The equilibrium unfolding of TIM induced by guanidine hydrochloride was investigated by following the changes in intrinsic fluorescence, circular dichroism and second-derivative spectroscopy. Results show that the unfolding of TIM is highly cooperative and no folding intermediate was detected in the experimental conditions used. The thermodynamic parameters of TIM during guanidine denaturation were calculated according to a two-state unimolecular unfolding model. Thermal denaturation of TIM monitored by CD at 222 nm shows also a single, cooperative transition with an apparent T_m of 64.6℃ and the thermal stability of TIM was decreased in the presence of low concentrations of guanidine. The possible unfolding pathway of the dimeric TIM was discussed. It shows that the secondary and tertiary structural changes of TIM occur simultaneously during guanidine denaturation and the unfolding of dimeric TIM follows an apparent two-state transition without detectable dissociation model.

Abstract:A fast, high-throughput, automated multiple antigenic peptides microarray platform was established based on multiple peptide design and synthesis technology. Human cytomegalovirus(HCMV) glycoprotein B, phosphoprotein pp150 and Helicobacter pylori (Hp) urease (Ure) beta subunit were chosen as target protein. The advantageous linear epitope of these proteins was analyzed and screened. The multiple antigenic peptides(MAPs) containing the selected sequence were synthesized by Fmoc solid phase method,purified through high performance liquid chromatography(HPLC) and printed on nitrocellulose membrane in microarrays by computer-controlled high-speed robotics. The nitrocellulose membrane was blocked with 2% bovine serum albumin solution. The MAPs microarray was finished by assembling the membrane with plastic outer shell. Some microarrays were selected at random for quality control identified by control sera and compared with ELISA method. 4 MAPs were screened out, synthesized and identified. The result of positive and negative sera of Hp and HCMV detected by MAPs microarray were consistent with the control sera. In the clinical trial of 120 random sera, the microarray performed almost equally with ELISA method using recombinant antigen and microbial lysate antigen. The sensitivity and specificity of Ure-1, Ure-2 and PP150 MAPs were higher than 90%. The CV were lower than 7% among microarrays showing a good repeatability. It can be concluded that a MAPs microarray analytical platform which has a vast application prospect in assistant design of peptide vaccine was primarily established.

Abstract:Twenty kinds of amino acids are simplified into 3 types: hydrophobic amino acids (H), hydrophilic amino acids (P) and neutral amino acids (N). Each residue is reduced to a bead which locates in the position of the C_α atom. The off-lattice model is adopted and the relative entropy is used as a minimization function to predict the tertiary structure of a protein. A new contact intensity function is given to consist with protein design research based on the relative entropy. Testing on several real proteins from Protein Data Bank (PDB) shows the good results obtained with the model and method. The root mean square deviations (RMSD) of the predicted structures relative to the native structures range from 0.30 to 0.70 nm. A foundation for studying protein design using the HNP model and the relative entropy was made.

Abstract:Two-dimensional (2D) gel electrophoresis followed by matrix-assisted laser desorption/time of flight mass spectrometry was utilized to compare the protein expression profiles between two colorectal cancer (CRC) cell lines with high and low metastatic potentials, SW620 and SW480. 10 metastasis-associated proteins were identified and further validated by Western blotting and/or semi-quantitative RT-PCR analysis. Of the identified proteins, the expressions of phosphoglycerate mutase 1, phosphatidylethanolamine binding protein and high-mobility group box 1 were elevated in SW620 cells. However, heat shock protein 27, annexin Ⅰ, methylthioadenosine phosphorylase, cofilin-1 and epidermal fatty acid binding protein were down-regulated in SW620 cells. Most of the candidate proteins have been evident to be somehow associated with various aspects of tumor metastasis such as cell growth, motility, invasion, adhesion, apoptosis and tumor immunity. These results provide the basis for searching for potential markers for CRC prognosis and give some clues to elucidate the mechanism of CRC metastasis.

Abstract:E-cadherin was one of the early expressed genes during embryo development. The intact form of E-cadherin can link to cytoskeleton via β catenin, and the complex plays essential roles in cell adhesion. Evidences indicated the essential functions of E-cadherin in processes of embryoic implantation and placentation. Immunohistochemistry and RT-PCR were used to identify the localization of E-cadherin in human placenta. It was found that E-cadherin was expressed mainly in villous cytotrophoblasts and trophoblast column, with the immunoreactivity decreased obviously in distal trophoblast column. Temporally, the mRNA level of E-cadherin in placenta was the highest at gestaional week 6, and began to be down-regulated from week 8 on, reaching a nadir at week 9. However, the mRNA expression was up-regulated in placenta at week 26 and full-term. In human normal placenta origin cytotrophoblast cell line (NPC), the mRNA and protein expressions of E-cadherin was significantly stimulated by TGFβ1 in dose- and time-dependent manners. Meanwhile, the cell-cell adhesion of NPC cells was promoted by TGFβ1. All these data indicated that there exists paracrine regulation of E-cadherin in human placenta, and E-cadherin may be involved in regulating trophoblast cell behaviors, likely inhibiting cell invasion through facilitating cell-cell adhesion.