Abstract:Developmental dyslexia (DD) is a specific learning disability. Exploring the mechanism of DD is of great assistance in developing the technique for diagnosis and therapy. There is a battery of researches about DD in west countries using alphabetic language as their mother tongue, though some opinions have not been widely accepted. The studies about Chinese DD are limited because it started relatively later. The authors review the advances of DD researches at three levels of behavior, neuron and genetics, surrounding three leading theories of DD: the phonological theory, the magnocellular (visual and auditory) theory and the cerebellar theory. At the same time, the authors compare the findings of DD between alphabetic languages and Chinese in order to uncover the differences of DD due to their mother tongue. Finally, the authors point out that Chinese dyslexics have their special characters different from those in alphabetic language, it?蒺s greatly necessary to strengthen the researches of Chinese DD, not only for establishing the theory of Chinese DD, but also for providing Chinese evidence of language specialization.

Abstract:About 70% of Parkinson's pathogenesis comes from environment factor and one important of which is oxidative stress although the genetic factor plays an important role. The antioxidant of green tea polyphenols(GTP) and they can enter into plasma even penetrate blood brain barie provides an important condition for the protective effects of GTP against Parkinson's Disease(PD). In cellular model the protective mechanisms of GTP on PC12 and SH-SY5Y cells against apoptosis induced by 6-hydroxydopamine (6-OHDA) were investigated. GTP attenuated 6-OHDA-induced early apoptosis, prevented the decrease in mitochondrial membrane potential, suppressed accumulation of reactive oxygen species (ROS) and of intracellular free Ca²⁺. GTP also counteracted 6-OHDA-induced nitric oxide increase and over-expression of nNOS and iNOS, and decreased the level of protein bound 3-Nitro-tyrosine (3-NT). Using PD rat model injected by 6-OHDA, the effect of GTP were investigated on animal model. Results showed that GTP attenuated the injury in a dose and time dependent manner. Pretreatment of the animals with GTP decreased ROS and NO production, thiobarituric acid reactive substances content, nitrite/nitrate concentration, and protein bound 3-Nitro-tyrosine (3-NT) in brain homogenate of midbrain and striatum in a concentration and time dependent manner. NOS participated in the neuron death induced by 6-OHDA and it was found that the pretreatment with GTP could decrease the protein level of nNOS and iNOS. More neurons survived and less cells suffered apoptosis in the substantia nigra pars compacta (SNc) of GTP treated animal brain. These results suggest that oral administration of GTP increases the antioxidant level in the brain and protects the brain against cell death caused by 6-OHDA. The experimental results of present study support the neuroprotection of GTP and provided new strategy of preventing and curing Parkinson's diseases by ROS-NO pathway.

Abstract:Y-box binding protein is a group of proteins that exist in a wide range, from low level to high level, of biologic species and they perform multiple biological functions. Numerous researches indicate that YB-1, as a member of the Y-box binding protein family, can interact with many important biological molecules and significantly influence the physiological function in organisms. More importantly, YB-1 also has a very profound role in relation to many diseases, especially to occurrence and development of cancer, and it can induce and maintain all the phenotypes and multiple drug resistance (MDR) of cancer cells. It is anticipated that a novel strategy of cancer treatment targeting YB-1 will effectively slow down the conditional deterioration and abrogate MDR in the cancer patients. The current progresses made in the YB-1 researches to its association with the occurrence/development of cancer were discussed and prospect on new strategies targeting YB-1 was provided.

Abstract:The investigations on the interactive relationships of DNA replication, damage repair and recombination have been locating in the frontline and becoming one of the hotspots in today's life science research. More and more studies show that the processes of DNA replication, damage repair and recombination are both independent and interdependent in the molecular level. These pathways coordinate and conform each other through interactions of many critical proteins in the pathways, by which DNA molecules, known as genetical materials, can be well maintained in cell and faithfully transferred through cellular generations. By using E. coli as a model system, the recent progresses and the possible rules underlying E. coli DNA replication, repair and recombination have been analyzed. As it is believed that the researches on E. coli DNA replication, repair and recombination may be capable of providing clues to the eukaryotic research based on the universal conservations of the critical proteins in the pathways.

Abstract:The ability to feed on vertebrate blood has evolved many times in various arthropod clades. Consequently, saliva of blood-feeding arthropods has proven to be a rich source of antihemostatic molecules. A variety of platelet aggregation inhibitors antagonize platelet responses to wound-generated signals, including ADP, thrombin, and collagen. Anticoagulants disrupt elements of both the intrinsic and extrinsic pathways. Vasodilators include nitrophorins (nitric oxide storage and transport heme proteins), a variety of peptides that mimic endogenous vasodilatory neuropeptides, and proteins that catabolize or sequester endogenous vasoconstrictors. Multiple salivary proteins may be directed against each component of hemostasis, resulting in both redundancy and in some cases cooperative interactions between antihemostatic proteins. The complexity and redundancy of saliva ensures an efficient blood meal for the arthropod, but it also provides a diverse array of novel antihemostatic molecules for the pharmacologist.

Abstract:To study the effects of connexin43 down regulation on the development of the embryonic heart and vasculature in zebrafish, two types of well designed morpholino oligonucleotide antisenses were injected into zebrafish embryos to block the translation of cx43 at one or two cells stage. After injection, the phenotypes of heart and vasculature were monitored by whole mount in situ hybridization, whole-mount immunofluorescence and microangiography. Whole-mount in situ hybridization with vmhc and amhc RNA probes showed that the vmhc expression cell domain was reduced; meanwhile, amhc expression cell domain was increased in cx43 down regulation group. Whole-mount immunofluorescence provided the evidence that down regulation of cx43 resulted in enlarged atrium and retrenched ventricle. Both in situ hybridization and microangiography indicated that vasculature pattern of cx43 morphants are almost normal compared with wildtype. Besides, the function of heart was affected obviously. Down regulation of cx43 caused the development defects of zebrafish embryonic heart, which may be involved in the wrong destination of two migratory cell populations, but it did not nearly affect vascular development.

Abstract:SR proteins play important roles in regulating alternative pre-mRNA splicing. As a newly discovered neural and reproductive tissue specific SR protein, SRp38 regulates the alternative splicing of several genes important for neural function, such as GluR-B, Trk-C and NCAML1. It also acts as a splicing inhibitor during mitosis or stress response in order to prevent wrong splicing. The expression of SRp38 in mouse retina was investigated by Western blot and immunohistochemistry (IHC) analyses. The result shows that the expression of SRp38 proteins in mouse retina is region-specific, with extensive distribution in the outer and inner plexiform layers, inner nuclear layer and ganglion cell layer, but no expression in outer nuclear layer. Double staining of isolated retina cells with anti-SRp38 and anti-Trk-C antibodies showed that SRp38 is localized in the dendrites, somata and axon terminals of rod-bipolar cells. By transient co-transmission of over-expressed SRp38 plasmid and RT-PCR analyses, the further results showed that overexpressed SRp38 could promote the splicing of the Flip isoform of GluR-B minegene in R28 cells. The result suggests that SRp38 may play important roles in the retinal function, possibly via regulating the neural-specific alternative splicing of genes as GluR-B.

Abstract:To study the regulation mechanism of SOCS-3 (Suppressor of cytokines signals-3) for erythropoietic development, small interference RNA expression vectors of SOCS-3 were constructed and transferred in to the K562 cell lines stably by lentiviral system. The efficiency of virus transfection was identified by expression of green fluorescence protein(GFP) analyzed by fluorescence microscope, then the high GFP expression K562 cells were sorted by fluorescence-activated cell sorting (FACS) according to strong GFP expression. The efficiency of RNA interferencing on SOCS-3 were detected by Real time-PCR and Western blot. The SOCS-3 gene expression at mRNA level of K562 cells transfected by lentivirus plasmids was 22.1% of the K562 cells transfected by the control lentivirus. The SOCS-3 protein expression of K562 cells transfected by lentivirus plasmids was also down regulated. Furthermore, K562 cells transfected by lentivirus plasmids were induced into the erythropoietic cells and the erythropoietic differentiation of K562 cells were examined by benzidine staining, immunocytochemistry and RT-PCR. Then it was found that the K562 cells could be induced into erythroid lineage cells more easily after silencing of SOCS-3. The experiment confirmed the important role of SOCS-3 in erythropoietic development and provided a useful new way for production of erythroid lineage cells.

Abstract:To explore whether the 3′-untranslated region (3′-UTR) of human high-affinity sodium-dependent dicarboxylate transporter (hSDCT2) plays a role in the regulation of gene expression, sequence characteristics of the 3′-UTR was analyzed using bioinformatics. The results found that within the 3′-UTR of SDCT2β mRNA there is an AU-rich region (AUR) of 585 nt, which contains three AU-rich elements (ARE). The AUR fragment was inserted into the 3′-UTR of GFP reporter gene within a expression vector pcDNA-GFP; and a pcDNA-GFP-AUR recombinant vector was constructed and transfected into HEK293, HKC and LLC-PK1 cell lines. The expression level of intracellular GFP was determined by Western blot and flow cytometry. The results showed that the AUR of SDCT2β could significantly reduce the expression level of GFP in the pcDNA-GFP-AUR- transfected cells (P < 0.01). After blockade of RNA transcription with actinomycin D, total RNA was extracted from stably transfected HEK293 cells with an interval of 2 h, and the stability of GFP-AUR and GFP mRNAs was analyzed by Northern blot. The results indicated that the stability of GFP-AUR mRNA is less than that of GFP mRNA. These results suggest that within the AU-rich region of 3′-UTR of SDCT2β mRNA there is a negative regulation region which can decrease the stability of mRNA, accelerate the degradation of GFP mRNA and may play a negative regulation role for gene expression at post-transcriptional level.

Abstract:In order to investigate the effects of hyperbaric oxygen (HBO) on the proliferation and differentiation of human osteoblasts isolated from alveolar bone under various pressure and exposure time, osteoblasts from human alveolar bone were seeded in 24 well plates at a cell density of 2 500 cells per well. There are four treatment groups which were 2.4ATA for 90 min, 2.4ATA for 30 min, 1.5ATA for 90 min and 1.5ATA for 30 min. Osteoblasts culture were treated one time everyday for up to 10 days in a temperature and humidity controlled custom-made seven-litre hyperbaric unit. Control samples were incubated in a standard humidified incubator at 37℃ containing 5% CO₂ and 95% atmospheric air. Proliferation of osteoblasts were evaluated by WST-1 assay before and 16 h after HBO on day 1,2,3,4,6,8,10. The cytotoxic effect of HBO on osteoblasts was assessed by a toxicology assay kit. For differentiation study, osteoblasts were seeded in 96 well plates at a cell density of 10 000 cells per well. After 3 days normal culture, medium was changed to osteogenic medium. Subsequently, cultures were exposed daily to HBO of 2.4ATA for 90 min and 1.5ATA for 90 min up to 13 days. Mineralization was evaluated by calcium deposition assay, alkaline phosphatase (ALP) activity and von Kossa staining. To assess the effect of pressure on cell proliferation and differentiation, hyperbaric air treatment was observed in this study. It showed that HBO treatment promotes proliferation of osteoblast in the presence of 10% foetal calf serum (FCS). No significant change in extracellular LDH activity before and after HBO treatment. The study of differentiation demonstrated that HBO enhanced differentiation associated with increased bone nodule formation, calcium deposition and alkaline phosphatase activity. These result suggests that HBO treatment significantly stimulated osteogentic differentiation, which implies a potential application of HBO in bone tissue engineering.

Abstract:Protein arginine methyltransferase 5 (PRMT5) has been implicated as an important regulator of many cellular processes and signaling pathways, including chromatin remodeling, RNA splicing, DNA transcription, and cell proliferation. Therefore, structural and functional studies on PRMT5 are quite important. The full length of PRMT5 gene was cloned into vector pGEX-4T-1, resulting in only low expression levels in Escherichia coli (E. coli). Here, it was showed that the several N-terminal amino acids deletions could result in a significant increase in the amount of soluble fraction, while one of them did not affect the protein-arginine methyltransferase activity. And it was also found that the N-terminal 15 amino acids region of PRMT5 may be important for the catalytic activity.

Abstract:To study the effect of AMD3100 on the mobilization, proliferation, migration and adhesion of endothelial progenitor cells (EPC), EPC was isolated from apoE^-/- mice bone marrow, 12 male apoE^-/- mice, with 8 weeks old,were randomly divided into two groups, AMD3100 group (2.5mg / (kg·2d)) and control group(PBS,0.1 ml/2d). After feeding western (high fat and cholesterol) for 12 weeks, the bone marrow cells were isolated and cultured by way of differential-speed-adherence and Micropore-Method. CD133⁺ VEGFR-2⁺ bone marrow cell was identified as endothelial progenitor cells by immunofluorescence. The proliferation, migration and adhesion of endothelial progenitor cells were detected by MTT chromometry, transwell and adhesion test, respectively. By counting the typical endothelial progenitor cells-colony forming units (EPC-CFUs) and observing the size and cell density of second EPC-CFUs, the clonality of endothelial progenitor cells was determined. The expression of CXCR4 mRNA and protein were measured by RT-PCR and Western blot. As a result, the proliferation, migration, adhesion and clonality of endothelial progenitor cells derived from AMD3100 group were attenuated in comparison to the control group; the expression of CXCR4 mRNA and protein of AMD3100 group were also lower to control group. It can be concluded that, lasting administration of AMD3100 inhibits the proliferation, migration, adhesion and clonality of bone marrow endothelial progenitor cells and down-regulate the expression of CXCR4 on endothelial progenitor cells.

Abstract:Lithium carbonate could be used to treat or prevent brain damage following traumatic injury and neurodegenerative diseases. It has been shown that its protective effect is related to protein kinase C (PKC) and extracellular signal-related kinase (ERK). It was demonstrated that PDBu, a PKC activator, inhibited amplitudes of delayed rectifier potassium current (I_K) and produced a hyperpolarizing shift in the activation-voltage curve. The responses to PDBu were inhibited by lithium carbonate (50 μmol/L). Further studies showed that when pretreated with MEK/ERK inhibitor U0126 (20 μmol/L), although PDBu significantly reduced I_K, lithium did not reverse the effect of PDBu. Thus, the results suggested that PKC signaling cascades, along with MAPK (mitogen-activated protein kinase) pathway, were required in the phosphorylation of potassium channel, which was presented by regulation of potassium channel characteristic. AC-cAMP and their cross-talk with GC-cGMP pathway could also modulate the effect of lithium on PKC activation, which could be one of underlying mechanisms likely related to neuroprotective effect of lithium.

Abstract:To investigate the effect of advanced glycation end products (AGE) modified protein on IL-8 secretion by human endothelial cells and the role of receptor for advanced glycation end products (RAGE) in this pathological procedure. Human umbilical vein endothelial cells (HUVEC) were cultured in vitro with different concentration AGE modified human serum albumin (AGE-HSA) which had been treated with soluble receptor for advanced glycation end products (sRAGE) or not. IL-8 levels in the supernatant were determined using Liquid Chip method. RNA was extracted from the cells and RT-PCR was performed to determine the mRNA expression levels of IL-8 in each group, and GAPDH levels were served as reference during this process. The results showed that AGE-HSA increased IL-8 secretion as a dose- and time- dependent manner. Unmodified HSA had no such effect. AGE-induced IL-8 secretion was significantly inhibited by sRAGE as a dose-dependent manner, and the decrease extent was 82.4% when the cells were incubated with 500 mg/L of sRAGE. AGE-RAGE interaction upregulated IL-8 mRNA expression in HUVEC, and this effect could be blocked by pretreatment of AGE-HSA with intact sRAGE. It was proved that AGE modified protein increases the secretion of IL-8 by endothelial cells through RAGE on protein and gene levels, which maybe provide a new aspect to study the machines and treatment methods of AGE-associated diseases.

Abstract:As₂O₃ is a Chinese traditional medicine. In previous study, 1.0 μmol/L As₂O₃ have been found that could induce MG-63 cell apoptosis in vitro. And then it was identified that IEX-1 gene expression was down regulated after the addition of 1.0 μmol/L As₂O₃ using cDNA Array, RT-PCR, Northern blot. IEX-1, a recently discovered early response gene, regulates cell growth and apoptosis. The exact transcriptional mechanism of IEX-1 was found using Dual-luciferase assay, EMSA, Western blot. Transcriptional factor p53 was up-regulated after the addition of 1.0 μmol/L As₂O₃, and the increased p53 protein bind the promoter region of IEX-1 gene, which repress the transcription and result in down-regulation of IEX-1 gene expression. It is further confirmed that IEX-1 gene is associated with osteosarcoma and the exact transcriptional mechanism of IEX-1 after the addition of As₂O₃ was found.

Abstract:miRNA was 20～25 nt endogenous non-coding RNA. miRNAs are encoded small RNAs that hybridize with messenger RNAs, resulting in degradation or translational inhibition of targeted transcripts. In order to investigate the function of miR-138 on mouse mammary epithelial cells, technique for gene silencing-miRNA inhibitor (anti-miRNA) was applied to make miR-138 silence, qRT-PCR was showed valid for inhibitor miR-138. And Western blot, CASY^®-technology was put in use to study some change of mouse mammary epithelial cells after miR138 inhibitor. It was shown that miR-138 suppresses the exepress of PRL-R(P < 0.05). It was proved that miR-138 can inhibit activity and proliferation of mouse mammary epithelial cells after miR-138 inhibitor(P < 0.01). HPLC was used to study expression of β-casin mouse mammary epithelial cells after miR-138 inhibitor, it shows down express of β-casin(P < 0.05). Results indicated that the target of miR-138 was PRL-R, miR-138 suppresses translation of PRL-R. MiR-138 suppressed the viable and proliferation of mouse mammary epithelial cells.

Abstract:Mass spectrometry has been extensively used in the identification of protein and its posttranslational modification (PTM). Mass spectrometry coupled with nano high performance liquid chromatography (HPLC) could improve its resolution and efficiency. Ubiquitination, one kind of the important protein posttranslational modifications (PTM), plays a pivotal role in dynamic balance and functional regulation of protein. A new strategy taking the advantage of immunoprecipitation, 2D nano HPLC and matrix-assisted laser desorption/ionization-time of flight mass spectrometry was established to identify the ubiquitination of endogenous protein in mammalian cells. By using this strategy, the ubiquitinated sites of c-ABL in K562 leukemia cells was successfully identified. Thus, it may provide a valuable tool to characterizing the ubiquitination of endogenous proteins under physiologic and pathologic conditions.