Abstract:To analyze the differentially expressed proteins in bronchial epithelial carcinogenesis process and discover novel biomarkers for early detection of human lung squamous cell cancer (LSCC), iTRAQ-tagging combined with 2D LC-MS/MS analysis was used to identify differentially expressed proteins in human bronchial epithelial carcinogenic process using laser capture microdissection-purified normal bronchial epithelium (NBE), squamous metaplasia (SM), atypical hyperplasia (AH), carcinoma in situ (CIS) and invasive LSCC. As a result, 1036 proteins and 102 differentially expressed proteins were identified. Among these differentially expressed proteins, some proteins are progressively upregulation, some proteins are progressively downregulation in human bronchial epithelial carcinogenic process, and the other proteins are upregulation or downregulation in a certain stage of the process of carcinogenesis. Functional analysis and subcellular localization were performed on 102 differentially expressed proteins. Functional analysis showed that these differentially expressed proteins were associated with metabolic process, apoptosis, cell proliferation, cell differentiation, signal transduction, transcription, translation, cell adhesion, immune response, and development. Western blotting and immunohistochemistry were performed to detect the expression levels of the two proteins (S100A9 and CKB) in NBE, SM, AH, CIS and invasive LSCC, which also support the findings in MS analysis. The results suggest that these differentially expressed proteins associated with bronchial epithelial carcinogenesis, and they may be early diagnostic marker of lung squamous cell carcinoma. We further study the biological function of the differentially expressed proteins, will help to elucidate the bronchial epithelial carcinogenesis mechanism, and provide new ideas for early diagnosis and pathogenesis research of LSCC.

Abstract:Glucagon-like peptide-1(GLP-1) is an important incretin hormone that secreted from intestinal L cells. Numerous studies have proven that GLP-1 increases β-cell mass through promoting β-cell proliferation and inhibiting β-cell apoptosis, besides stimulating insulin secretion. This review focuses on the molecular mechanism of GLP-1 induced β-cell protection.

Abstract:Insulin resistance is a pathological basis for the development of type 2 diabetes mellitus (T2DM), and has been a key and hot topic for T2DM research recently. Furthermore, an increasing number of reports demonstrate that some sphingolipids play a critical role in regulating insulin signal. For instance, both ceramide and ganglioside monosialo 3 (GM3) can reduce insulin signal sensitivity and lead to insulin resistance. Their mechanisms may be associated with caveolin localization and formation of caveolae on membrane. On the contrary, another remarkable sphingolidid, sphingosine-1-phosphate (S1P), dramatically enhances insulin signal by redox signaling pathway. Both negative and positive effects of sphingolipids on insulin signal imply the involvement of calcium signal. Thus it can be seen that detecting both extracellular calcium influx and intracellular calcium transient in real time may satisfy the speculation of insulin signal regulation with sphingolipids at the ion channel level. Therefore, we will review the mechanisms of modulation of some sphingolipids, including ceramide, GM3 and S1P, on insulin signal. The potential strategies of sphingolipids activity alteration and insulin resistance improvement will be involved in this review as well, which will make great strides in T2DM research.

Abstract:ATP-binding cassette transporter 1 (ABCA1) mediated lipid efflux and evolved to maintain cholesterol homeostasis. Newborn ABCA1 should through intracellular transport and chemical modification, ultimately being a mature functional transporter with its function of lipid transport. The intracellular transport and correct localization of ABCA1 plays an important role in its function. ABCA1 researches focused on the lipid transport, and proposed the model of the various cholesterol efflux mechanisms, such as channel transport model, mushroom-like protuberances model and retroendocytosis model. Recent studies have shown, ABCA1 can regulate the plasma membrane lipid rafts, and involve in immune and inflammatory regulation. This review focuses on the current views on intracellular transport and various functions of ABCA1, in order to provide the new therapeutic targets for atherosclerosis-related diseases.

Abstract:The cyclic dinucleotides such as cyclic-di–guanosine monophosphate (c-di-GMP), c-di-AMP and cGAMP were discovered as second messengers in many species of bacteria, but weren’t identified in mammal.Recently, cGAMP was demonstrated to play critical regulatory roles as endogenous second messenger in mammal innate immune signaling. cGAMP can be catalyzed by cytoplasmic cGAMP synthase (cGAS) after DNA binding, and then bind to STING(stimulator of IFN genes) which triggers type I interferon-dependent innate immunity. These researches provide new perspective for signaling pathway in innate immunity and are beneficial for development of therapeutic agents in immune therapy.

Abstract:Resistin was a cytokine associated with insulin resistance. The objective of the study is to investigate the impact of fibroblast growth factor (FGF)-21 on glucose metabolism in resistin-induced insulin resistant liver cells. A stable HepG2 cell line expressing human resistin was developed. The cell line was treated with different concentrations of insulin or FGF-21. The glucose uptake of the treated cells was measured by the method of GOD-POD, and the mRNA expression of GLUT1 and PPAR-γ of the FGF-21-treated cells were examined by real-time PCR. We successfully developed the HepG2 line which stably expressed human resistin. The results of glucose uptake showed that the sensitivity of the cell line to the insulin treatment significantly decreased, but FGF-21 could effectively stimulate the glucose uptake by the same cell line. The mRNA expression of GLUT1 was elevated in the cell line, and the GLUT1 mRNA level of the cells significantly increased after treatment by FGF-21. The expression of PPAR-γ had no significant change. The cell line in which resistin was highly expressed has a reduced susceptibility to insulin. Although the high level of resistin makes cells insulin resistance, FGF-21 can regulate the glycometabolism effectively in the cells.

Abstract:In order to investigate the inhibitory effect of cariporide, a specific Na⁺/H⁺ exchanger 1 (NHE1) blocker, on neointimal proliferation induced by AGEs in a carotid artery balloon injury model, the rats carotid artery was balloon injured. The exemplars were collected and stained by HE. The morphology changes were observed. The intima area, media area and the ratio of area between intima and media were calculated using image analysis system. In order to explore the precise mechanism, the experiments were done on the isolated rat vascular smooth muscle cells(VSMC). Cell proliferation was assessed by [³H] thymidine incorporation. RT-PCR and Real-time RT-PCR were used to assay the cyclooxygenase-2 (COX-2) matrix metalloproteinases-2 (MMP-2) and matrix metalloproteinases-9 (MMP-9) expression; Western blot was used to assay NF-κB protein and the degradation of the inhibitor I-κBα. The in vivo results shows that neointima hyperplasia is significantly suppressed treated with cariporide in balloon-injured rats compared with AGEs treatment lonely. The in vitro study shows that cariporide dose-dependently inhibited AGEs-induced upregulation of COX-2, MMP-2 and MMP-9 expression. We also found that cariporide blocked AGEs-induced activation of nuclear factor-κB(NF-κB)and pointed out that inhibition of NF-κB was achieved by inhibiting the degradation of the inhibitor I-κBα. The results identified that NHE1 inhibitor cariporide inhibited AGEs-induced neointimal hyperplasia in rats balloon-injured model by suppressing the proliferation of VSMC and the upregulation of COX-2, MMP-2 and MMP-9 via inhibiting NF-κB activation in VSMC. These results indicated that NHE1 might be a considerable ingredient of the signal pathway in which AGEs played a key role in the processes of vascular damage.

Abstract:Vasculogenesis and angiogenesis are fundamental processes for the formation and restoration of circulatory system, which plays important role in embryonic organ development, adult wound healing and reproduction. Vascular endothelial cells are the foundation of cardiovascular systems. Although it has been proved that several kinds of stem cells could be induced to generate endothelial cells ex vivo, there are still some shortages in these cell sources. Depending on the multipotency and infinitive reproductive capability, human embryonic stem cells (hESCs) become the new sources. However, the efficiency for deriving endothelial progenitor/ endothelial cells (EPCs/ECs) from hESCs is low. To increase endothelial induction efficiency, we used a two-stage induction protocol. By the treatment of different cytokine cocktails at different stages and the using of Matrigel matrix, we obtained about 16% CD31⁺KDR⁺ cells from hESCs. The following endothelial induction stage with EGM2 medium showed approximate 32% subgroup of EPCs/ECs. The derived cells had the capability for tube-like structure formation on Matrigel and the ability to cohere with fluorescein griffonia (bandeiraea) simplicifolia lectinⅡ. Q-PCR indicated that the induced cells express endothelial specific genes. Immunofluorescence detection indicated the expression of endothelial specific marker-CD31. Taken together, we have successfully derived EPCs/ECs from human embryonic stem cells with a modified method.

Abstract:The study was aimed to investigate the effects and mechanism of manganese superoxide dismutase (MnSOD) on tert-butyl hydroperoxide(t-BHP) induced apoptosis in human bone marrow mesenchymal stem cells (MSCs). The MnSOD gene was cloned from human fetal liver by RT-PCR. The MnSOD recombinant plasmid was transfected into MSCs stably by lentiviral system. The efficiency of virus transfection was identified by expression of enhanced green fluorescence protein (EGFP) analyzed by fluorescence microscope, then MSCs were sorted by fluorescence-activated cell sorting (FACS) according to strong EGFP expression. MnSOD expression was detected by RT-PCR and Western blot. Transfected cells were then treated with different concentration of t-BHP, and the features including cell viability, senescence-associated β-gal activity, and apoptosis were evaluated. Our data demonstrated that overexpression of MnSOD could promote MSCs viability by inhibiting apoptosis or cellular senescence. Furthermore, apoptosis related genes p53 and PUMA were down-regulated. Therefore, these results indicated that overexpression of MnSOD in MSCs could protect against t-BHP induced apoptosis.

Abstract:Amylosucrase (AS) is a kind of glucosyltransferase (E.C. 2.4.1.4) belonging to the glycoside hydrolase (GH) family 13. In the presence of an activator polymer, in vitro, AS is able to catalyze the synthesis of an amylose-like polysaccharide composed of only α-1,4-linkages using sucrose as the only energy source. Unlike AS, other enzymes responsible for the synthesis of such amylose-like polymers require the addition of expensive nucleotide-activated sugars. These properties make AS an interesting enzyme for industrial applications. In spite of the great application potential, industrial applications of AS were largely hampered by its low stability. In this work, the structure of Arthrobacter chlorophenolicus amylosucrase (AcAS) was modeled according to crystal structures of Neisseria polysaccharea amylosucrase (NpAS) and Deinococcus geothermalis amylosucrase (DgAS). The structural difference of the three AS were investigated for the sake of exploring the working mechanism of AcAS. The dynamics and functional motions of AcAS were also investigated by the Gaussian network model (GNM). Based on GNM, it was found that AcAS can be divided into two regions with different moving directions. Finally, the unfolding property of AcAS was studied by the iterative GNM, and several kinetically weak regions were identified on the basis of the predicted unfolding pathway. These discoveries may be helpful to the industrial development of AS.

Abstract:We used exercise training rat as animal model, and investigated the effect of exercise training on the property of large-conductance Ca²⁺ voltage-activated K+ (BK) channels in VSMCs using patch clamp technique. The results showed that the open probability of channels increased significantly during the second and third week of exercise training, then became stable in the fourth week. After the exercise training, the sensitivity of channels to voltage and intracellular Ca²⁺ was enhanced, and its mean opening time got longer while the mean closed time got shorter. Our results also showed that the exercise training had no effect on channel conductance. These changes in biophysical properties of BK channels after exercise training could be considered to prevent membrane depolarization and vasoconstriction.