In JoVE (1)

Other Publications (35)

Articles by Jingwen Yang in JoVE

Other articles by Jingwen Yang on PubMed

Preparation and Characterization of Glycerol Plasticized-pea Starch/ZnO-carboxymethylcellulose Sodium Nanocomposites

Bioresource Technology. Jun, 2009  |  Pubmed ID: 19217775

Among natural polymers, starch is one of the most promising biodegradable materials because it is a renewable bioresource that is universally available and of low cost. However, the properties of starch-based materials are not satisfactory. One approach is the use of nano-filler as reinforcement for starch-based materials. In this paper, a nanocomposite is prepared using ZnO nanoparticles stabilized by carboxymethylcellulose sodium (CMC) as the filler in glycerol plasticized-pea starch (GPS) matrix by the casting process. According to the characterization of ZnO-CMC particles with Fourier transform infrared (FTIR), Ultraviolet-visible (UV-vis), X-ray diffraction (XRD), transmission electron microscope (TEM) and thermogravimetric analysis (TG), ZnO (about 60 wt%) is encapsulated with CMC (about 40 wt%) in ZnO-CMC particles with the size of about 30-40 nm. A low loading of ZnO-CMC particles can obviously improve the pasting viscosity, storage modulus, the glass transition temperature and UV absorbance of GPS/ZnO-CMC nanocomposites. When the ZnO-CMC contents vary from 0 to 5 wt%, the tensile yield strength increase from 3.94 MPa to 9.81 MPa, while the elongation at break reduce from 42.2% to 25.8%. The water vapor permeability decrease from 4.76 x 10(-10) to 1.65 x 10(-10) g m(-1) s(-1) Pa(-1).

Identification and Functional Characterization of Two Orphan G-protein-coupled Receptors for Adipokinetic Hormones from Silkworm Bombyx Mori

The Journal of Biological Chemistry. Dec, 2011  |  Pubmed ID: 22009754

Adipokinetic hormones (AKHs) are the best studied insect neuropeptides with the function of mobilizing lipids and carbohydrates during energy-expensive activities and modulating fundamental physiological processes, such as sugar homeostasis, lipid metabolism, and reproduction. Three distinct cDNAs encoding the prepro-Bombyx AKH1-3 have been cloned and confirmed by mass spectrometric methods. Our previous research suggested the Bombyx AKH receptor is activated by AKH1 and AKH2 with high affinity but by AKH3 with quite low affinity. In this study, using stable functional expression of the receptors in HEK293 cells, we have now identified AKH3 as a specific ligand for two orphan G-protein-coupled receptors, and we therefore named them AKHR2a and AKHR2b, respectively. We demonstrated that both AKHR2a and AKHR2b were activated by AKH3 at high affinity and by AKH1 and AKH2 at low affinity, leading to an increase of intracellular cAMP levels and activation of ERK1/2 and receptor internalization, but they were not activated by Bombyx corazonin. Conversely, the Bombyx corazonin receptor was activated by corazonin but not by AKH1-3. Quantitative RT-PCR revealed that AKHR2a and AKHR2b were both highly expressed in the testis but were also detected at low levels in other tissues. These results will lead to a better understanding of the AKH/AKHR system in the regulation of fundamental physiological processes.

Relative Expression of Type II MAGE Genes During Retinoic Acid-induced Neural Differentiation of Mouse Embryonic Carcinoma P19 Cells: a Comparative Real-time PCR Analysis

Cellular and Molecular Neurobiology. Aug, 2012  |  Pubmed ID: 22410673

In mammals, the type II melanoma antigen (MAGE) protein family is constituted by at least ten closely related members, but our understanding of their function in the developing nervous system remains poor. To systematically study the expression pattern of type II MAGE genes during neurogenesis, we employed mouse embryonic carcinoma P19 cells as an in vitro model for neural differentiation by retinoic acid (RA) induction. The expression of type II MAGE genes was investigated under distinct steps of differentiation by a comparative ΔΔC (T) paradigm of real-time quantitative reverse-transcription PCR (qRT-PCR). The relative levels of each gene expression at various steps of differentiation were expressed as a fold change compared with that in RA-untreated P19 cells. The results revealed that: (1) the expression of MAGE-E1, E2, and Necdin transcripts was steadily increased, and the relative levels of MAGE-D1, D2, D3, F1, G1, and H1 mRNA were fluctuantly elevated after the RA-treatment at embryoid body and neural stages; (2) during RA-treatment and subsequent differentiation, the expression of MAGE-L2 mRNA was decreased. Therefore, our results suggested that MAGE-D1, D2, D3, E1, E2, F1, G1, H1, and Necdin might be involved in the early process of neurogenesis, and MAGE-L2 connected with maintenance of pluripotency of stem cells. These studies may present some clues for a better understanding of the fundamental aspects of type II MAGE genes during neurogenesis.

Mutations in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) in Chinese Patients with Congenital Bilateral Absence of Vas Deferens

Journal of Cystic Fibrosis : Official Journal of the European Cystic Fibrosis Society. Jul, 2012  |  Pubmed ID: 22483971

Genetic testing of the cystic fibrosis transmembrane conductance (CFTR) gene is currently performed in patients with congenital bilateral absence of vas deferens (CBAVD). This study was conducted to investigate the role of mutations in the CFTR gene in CBAVD-dependent male infertility.

Specific Activation of the G Protein-coupled Receptor BNGR-A21 by the Neuropeptide Corazonin from the Silkworm, Bombyx Mori, Dually Couples to the G(q) and G(s) Signaling Cascades

The Journal of Biological Chemistry. Apr, 2013  |  Pubmed ID: 23457297

Corazonin, an undecapeptide neurohormone sharing a highly conserved amino acid sequence across Insecta, plays different physiological roles in the regulation of heart contraction rates, silk spinning rates, the induction of dark color and morphometric phase changes, and ecdysis. Corazonin receptors have been identified in Drosophila melanogaster, Manduca sexta, and Musca domestica. However, detailed information on the signaling and major physiological functions of corazonin and its receptor is largely unknown. In the current study, using both the mammalian cell line HEK293 and insect cell lines BmN and Sf21, we paired the Bombyx corazonin neuropeptide as a specific endogenous ligand for the Bombyx neuropeptide G protein-coupled receptor A21 (BNGR-A21), and we therefore designated this receptor as BmCrzR. Further characterization indicated that synthetic BmCrz demonstrated a high affinity for and activated BmCrzR, resulting in intracellular cAMP accumulation, Ca(2+) mobilization, and ERK1/2 phosphorylation via the Gq- and Gs-coupled signaling pathways. The direct interaction of BmCrzR with BmCrz was confirmed by a rhodamine-labeled BmCrz peptide. Moreover, experiments with double-stranded RNA and synthetic peptide injection suggested a possible role of BmCrz/BmCrzR in the regulation of larval growth and spinning rate. Our present results provide the first in-depth information on BmCrzR-mediated signaling for further elucidation of the BmCrz/BmCrzR system in the regulation of fundamental physiological processes.

The Second Intracellular Loop of the Human Cannabinoid CB2 Receptor Governs G Protein Coupling in Coordination with the Carboxyl Terminal Domain

PloS One. 2013  |  Pubmed ID: 23667597

The major effects of cannabinoids and endocannabinoids are mediated via two G protein-coupled receptors, CB1 and CB2, elucidation of the mechanism and structural determinants of the CB2 receptor coupling with G proteins will have a significant impact on drug discovery. In the present study, we systematically investigated the role of the intracellular loops in the interaction of the CB2 receptor with G proteins using chimeric receptors alongside the characterization of cAMP accumulation and ERK1/2 phosphorylation. We provided evidence that ICL2 was significantly involved in G protein coupling in coordination with the C-terminal end. Moreover, a single alanine substitution of the Pro-139 in the CB2 receptor that corresponds to Leu-222 in the CB1 receptor resulted in a moderate impairment in the inhibition of cAMP accumulation, whereas mutants P139F, P139M and P139L were able to couple to the Gs protein in a CRE-driven luciferase assay. With the ERK activation experiments, we further found that P139L has the ability to activate ERK through both Gi- and Gs-mediated pathways. Our findings defined an essential role of the second intracellular loop of the CB2 receptor in coordination with the C-terminal tail in G protein coupling and receptor activation.

Localization of Beclin1 in Mouse Developing Tooth Germs: Possible Implication of the Interrelation Between Autophagy and Apoptosis

Journal of Molecular Histology. Dec, 2013  |  Pubmed ID: 23793984

Our previous study identified the appearance of autophagy in developing tooth germs, and suggested its possible association with apoptosis in odontogenesis. Beclin1 was recently indicated to play a central role in bridging autophagy and apoptosis, and occupied a key position in the process of development. This study hypothesized that Beclin1 may be involved, and act as the molecular basis of the connection between autophagy and apoptosis in odontogenesis. Immunohistochemical analysis showed the spatiotemporal expression pattern of Beclin1 in odontogenesis from embryonic (E) day 13.5 to postnatal (P) day 5.5. At E stages, Beclin1 was mainly immunolocalized in the cytoplasm of the cells in the enamel organ. Meanwhile, the nucleus localization of Beclin1 was detected in part of the stellate reticulum, outer and inner enamel epithelium, especially at E16.5 and E18.5. At P stages, Beclin1 was detected in the cytoplasm of the odontoblasts, besides the dental epithelium cells. Triple immunofluorescence analysis showed the partial colocalization of Beclin1, autophagic marker LC3, or activated caspase-3 in the E14.5 tooth germs, especially the Beclin1(+)LC3(+)Caspase-3(+) cells in the PEK. Furthermore, western blot analysis revealed that the full-length (60 kDa) and/or cleaved (50, 37, and 35 kDa) Beclin1 in the developing tooth germs. Taken together, our findings indicate that Beclin1 is involved, and might be responsible for the crosstalk between autophagy and apoptosis in mouse odontogenesis.

Transcription Factor SCIRR69 Involved in the Activation of Brain-derived Neurotrophic Factor Gene Promoter II in Mechanically Injured Neurons

Neuromolecular Medicine. Sep, 2013  |  Pubmed ID: 23842743

The spinal cord injury and regeneration-related gene #69 (SCIRR69), which was identified in our screen for genes upregulated after spinal cord injury, encode a protein belonging to the cAMP response element-binding protein (CREB)/ATF family of transcription factors. Our previous study showed that SCIRR69 functions as a transcriptional activator. However, the target gene regulated by SCIRR69 and its roles in injured neurons remain unknown. In this study, we showed that SCIRR69 is widely distributed in the central nervous system. Full-length SCIRR69 is an endoplasmic reticulum-bound protein. Following mechanical injury to neurons, SCIRR69 was induced and proteolytically cleaved by site-1 and site-2 proteases, and the proteolytically cleaved SCIRR69 (p60-SCIRR69) was translocated to the nucleus where it bound to brain-derived neurotrophic factor (BDNF) gene promoter II. In addition, loss- and gain-of-function studies confirmed that SCIRR69 is involved in the regulation of BDNF expression in injured neurons. As expected, the culture supernatants of PC12 cells stably expressing p60-SCIRR69 contained higher levels of BDNF, and more remarkably promoted neurite outgrowth in a spinal cord slice culture model in vitro than the supernatants of control cells. These results suggest that SCIRR69 is a novel regulator of the BDNF gene and may play an important role in the repair and/or regeneration of damaged neural tissues by specifically activating BDNF promoter II.

Activation of CAMP-response Element-binding Protein is Positively Regulated by PKA and Calcium-sensitive Calcineurin and Negatively by PKC in Insect

Insect Biochemistry and Molecular Biology. Nov, 2013  |  Pubmed ID: 24018109

The cAMP response element binding protein, CREB, is a G protein-coupled receptor (GPCR) signal-activated transcription factor implicated in the control of many biological processes. In the current study, we constructed a cAMP response element (CRE)-driven luciferase assay system for GPCR characterization in insect cells. Our results indicated that Gs-coupled Bombyx adipokinetic hormone receptor (AKHR) and corazonin receptor could effectively initiate CRE-driven luciferase transcription, but forskolin, a reagent widely used to activate adenylyl cyclase in mammalian systems, failed to induce luciferase activity in insect cells co-transfected with a CRE-driven reporter construct upon agonist treatment. Further investigation revealed that the specific protein kinase C (PKC) inhibitors exhibited stimulatory effects on CRE-driven reporter transcription, and blockage of Ca(2+) signals and inhibition of Ca(2+)-dependent calcineurin resulted in a significant decrease in the luciferase activity. Taken together, these results suggest that PKC likely acts as a negative regulator to modulate CREB activation; in contrast, Ca(2+) signals and Ca(2+)-dependent calcineurin, in addition to PKA, essentially contribute to the positive regulation of CREB activity. This study presents evidence to elucidate the underlying molecular mechanism by which CREB activation is regulated in insects.

The Presence of Autophagy in Human Periapical Lesions

Journal of Endodontics. Nov, 2013  |  Pubmed ID: 24139258

Autophagy, a lysosome- or endosome-mediated self-degradation process, participates in diverse neurodegenerative diseases, cancer, and inflammatory diseases associated with apoptosis. This study aims to identify the presence of autophagy in human periapical lesions and its possible colocalization with apoptosis.

The Distribution and Ultrastructure of the Forming Blood Capillaries and the Effect of Apoptosis on Vascularization in Mouse Embryonic Molar Mesenchyme

Cell and Tissue Research. Apr, 2014  |  Pubmed ID: 24477797

Vascularization is essential for organ and tissue development. Teeth develop through interactions between epithelium and mesenchyme. The developing capillaries in the enamel organ, the dental epithelial structure, occur simultaneously by mechanisms of vasculogenesis and angiogenesis at the onset of dentinogenesis. The vascular neoformation in the dental mesenchyme has been reported to start from the cap stage. However, the mechanisms of vascularization in the dental mesenchyme remain unknown. In the hope of understanding the mechanisms of the formation of dental mesenchymal vasculature, mouse lower molar germs from embryonic day (E) 13.5 to E16.5 were processed for immunostaining of CD31 and CD34, terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) and transmission electron microscopy (TEM). In addition, the role of apoptosis for the vascularization in dental mesenchyme was examined by in vitro culture of E14.0 lower molars in the presence of the apoptosis inhibitor (z-VAD-fmk) and a subsequent subrenal culture. Our results showed that CD31- and CD34-positive cells progressively entered the central part of the dental papilla from the peridental mesenchyme. For TEM, angioblasts, young capillaries with thick endothelium and endothelial cells containing vacuoles were observed in peripheral dental mesenchyme, suggesting vasculogenesis was taking place. The presence of lateral sprouting, cytoplasmic filopodia and transluminal bridges in the dental papilla suggested angiogenesis was also occurring. Inhibition of apoptosis delayed the angiogenic vascularization of the dental papilla. Therefore, these data demonstrated that molar mesenchyme is progressively vascularized by mechanisms of both vasculogenesis and angiogenesis and apoptosis partially contributes to the vascularization of the dental papilla.

A Novel Mutation of Hyaluronan Synthase 2 Gene in Chinese Children with Ventricular Septal Defect

PloS One. 2014  |  Pubmed ID: 24558368

As a major product of extracellular matrix (ECM), Hyaluronic acid (HA) is involved in early cardiac development and mainly synthesized by Hyaluronan synthase 2 (HAS2) during embryogenesis. Targeted deletion of HAS2 gene in mice led to obvious cardiac and vascular defects. To clarify the potential association of the mutation in HAS2 with the development of congenital heart disease (CHD), in this study, we sequenced the coding region of HAS2 and identified a novel non-synonymous variant c.A1496T (p.Glu499Val) in one of 100 non-syndromic Ventricular Septal Defect (VSD) patients. The variant was not observed in 250 controls. In addition, to determine the contribution of HAS2 variant in VSD, we compared HA content in supernatant using HA quantitative analysis and found that the mutation obviously affected the HA synthetic activity of HAS2. To our knowledge, this is the first time that the mutation in HAS2 was found in Chinese VSD patients, which suggested that HAS2 may be involved in the etiology of non-syndromic VSD and have the vital function in the development of heart septum.

MMP9 Deficiency Increased the Size of Experimentally Induced Apical Periodontitis

Journal of Endodontics. May, 2014  |  Pubmed ID: 24767560

Apical periodontitis is an inflammation and destruction of periapical tissues. Matrix metalloproteinase-9 (MMP-9) is thought to be involved in periapical lesion formation and progression. The aim of this study was to evaluate the lesion progression in MMP-9 knockout (KO) mice compared with that in control mice (wild type [WT]).

LPS-induced Dental Pulp Inflammation Increases Expression of Ionotropic Purinergic Receptors in Rat Trigeminal Ganglion

Neuroreport. Sep, 2014  |  Pubmed ID: 25055139

Severe toothache can be caused by dental pulp inflammation. The ionotropic purinergic receptor family (P2X) is reported to mediate nociception in primary afferent neurons. This study aims to investigate the involvement of P2X receptors in the sensitization of the trigeminal ganglion (TG) caused by dental pulp inflammation. Lipopolysaccharides were unilaterally applied to the pulp of the upper molar of the rat to induce dental pulp inflammation. Increased expression of c-fos, a marker of neuronal activity, was induced in V1-V2 division, indicating the activation of TG neurons. The expressions of P2X2, P2X3, and P2X5 were also increased in the V1-V2 division of TG, primarily in small-sized and medium-sized neurons. Markers of glutamatergic afferents, VGluT1, and GABAergic afferents, GAD67, were induced by lipopolysaccharides and coexpressed with P2X in small-sized TG neurons. The present findings suggest that the P2X2, P2X3, and P2X5 receptors are upregulated as part of the sensitization produced by dental pulp inflammation.

A Comparative Study of BioAggregate and ProRoot MTA on Adhesion, Migration, and Attachment of Human Dental Pulp Cells

Journal of Endodontics. Aug, 2014  |  Pubmed ID: 25069918

The aim of the present study was to evaluate the effects of a novel bioceramic nanoparticular cement, BioAggregate (Innovative Bioceramix, Vancouver, BC, Canada), on the adhesion, migration, and attachment of human dental pulp cells (HDPCs) and to compare its performance with that of ProRoot mineral trioxide aggregate (MTA) (Dentsply, Tulsa, OK).

In Vitro and in Vivo Evaluation of a Nanoparticulate Bioceramic Paste for Dental Pulp Repair

Acta Biomaterialia. Dec, 2014  |  Pubmed ID: 25182220

Bioactive materials play an important role in facilitating dental pulp repair when living dental pulp is exposed after injuries. Mineral trioxide aggregate is the currently recommended material of choice for pulp repair procedures though has several disadvantages, especially the inconvenience of handling. Little information is yet available about the early events and molecular mechanisms involved in bioceramic-mediated dental pulp repair. We aimed to characterize and determine the apatite-forming ability of the novel ready-to-use nanoparticulate bioceramic iRoot BP Plus, and investigate its effects on the in vitro recruitment of human dental pulp stem cells (DPSCs), as well as its capacity to induce dentin bridge formation in an in vivo model of pulp repair. It was found that iRoot BP Plus was nanosized and had excellent apatite-forming ability in vitro. Treatment with iRoot BP Plus extracts promoted the adhesion, migration and attachment of DPSCs, and optimized focal adhesion formation (Vinculin, p-Paxillin and p-Focal adhesion kinase) and stress fibre assembly. Consistent with the in vitro results, we observed the formation of a homogeneous dentin bridge and the expression of odontogenic (dentin sialoprotein, dentin matrix protein 1) and focal adhesion molecules (Vinculin, p-Paxillin) at the injury site of pulp repair model by iRoot BP Plus. Our findings provide valuable insights into the mechanism of bioceramic-mediated dental pulp repair, and the novel revolutionary ready-to-use nanoparticulate bioceramic paste shows promising therapeutic potential in dental pulp repair application.

Activation of BNGR-A24 by Direct Interaction with Tachykinin-related Peptides from the Silkworm Bombyx Mori Leads to the Gq- and Gs-coupled Signaling Cascades

Biochemistry. Oct, 2014  |  Pubmed ID: 25275886

Tachykinins constitute one of the largest peptide families in the animal kingdom and exert their diverse actions via G protein-coupled receptors (GPCRs). In this study, the Bombyx tachykinin-related peptides (TKRPs) were identified as specific endogenous ligands for the Bombyx neuropeptide GPCR A24 (BNGR-A24) and thus designated BNGR-A24 as BmTKRPR. Using both mammalian cell line HEK293 and insect cell line Sf21, further characterization demonstrated that BmTKRPR was activated, thus resulting in intracellular accumulation of cAMP, Ca(2+) mobilization, and ERK1/2 phosphorylation in a Gs and Gq inhibitor-sensitive manner. Moreover, quantitative reverse transcriptase polymerase chain reaction analysis and dsRNA-mediated knockdown experiments suggested a possible role for BmTKRPR in the regulation of feeding and growth. Our findings enhance the understanding of the Bombyx TKRP system in the regulation of fundamental physiological processes.

The Induction of Prolonged Myelopoietic Effects in Monkeys by GW003, a Recombinant Human Granulocyte Colony-stimulating Factor Genetically Fused to Recombinant Human Albumin

Journal of Pharmaceutical Sciences. Feb, 2015  |  Pubmed ID: 25174614

GW003, a genetic fusion protein of human serum albumin and granulocyte colony-stimulating factor (G-CSF), was developed based on a novel strategy for producing long-acting proteins. The purpose of this study was to evaluate the hematologic, pharmacokinetic, and toxicokinetic effects of GW003 on cynomolgus monkeys. We show that following a single subcutaneous administration of GW003, the absolute neutrophil count increased significantly compared with monkeys that received only the vehicle, and the magnitude of the neutrophilic response to GW003 was dose dependent. After an injection at equal molar dose, the clearance of GW003 in the monkeys was approximately fourfold slower, and the terminal half-life (T1/2 ) was fivefold longer than the corresponding values for recombinant methionyl human G-CSF. Interestingly, both the clearance and T1/2 decreased with increasing doses of GW003, and much faster elimination was observed after multidose exposure. In toxicokinetic studies, the serum concentration of GW003 after the eighth injection was much lower than it was after the first injection, and a neutralizing antibody against G-CSF was found to have a dose-dependent effect upon the treatment groups. Overall, the favorable pharmacokinetic and pharmacodynamic properties supported the selection and development of GW003 as a promising candidate for neutropenia therapy.

Phosphorus-containing Flame Retardant Modified Layered Double Hydroxides and Their Applications on Polylactide Film with Good Transparency

Journal of Colloid and Interface Science. Feb, 2015  |  Pubmed ID: 25460688

Polylactide (PLA)/layered double hydroxide (LDH) films with good flame retardant property and transparency were synthesized by solution exfoliation and film casting method. The organic-inorganic interfacial interaction between PLA and NiAl-LDH was carefully modified by 2-carboxylethyl-phenyl-phosphinic acid (CEPPA) to well solve the dispersion problem of NiAl-LDH nanolayers and get enhanced flame retardancy of PLA composites. The results showed the NiAl-LDH/CEPPA (LC) nanolayers had exfoliated structures and were homogenously dispersed in PLA matrixes. All the PLA/LDH films had good transparency even LC content was up to 10 wt%. The PLA/LDH films absorbed the ultraviolet light, which alleviates the embrittlement of PLA films in the using procedure. The flame retardant effect characterized by microscale combustion calorimeter was observed when LC contents increased. The total heat release value of the sample with 10 wt% LC decreased to 9.7 kJ/g from 12.0 kJ/g of virgin PLA.

GC-MS-based Plasma Metabolomic Investigations of Morphine Dependent Rats at Different States of Euphoria, Tolerance and Naloxone-precipitated Withdrawal

Metabolic Brain Disease. Jun, 2015  |  Pubmed ID: 25472920

Long-term or excessive application of morphine leads to tolerance and addiction, which hindered its conventional applications as a drug. Although tremendous progress has been made on the mechanisms of morphine, crucial evidence elaborating the neurobiological basis of tolerance and dependence is still lacking. To further explore the physiological adaptions during morphine's application, a systematic screening of small molecules in blood has been carried out. The plasma of morphine dependent rats was collected at different time points with or without naloxone treatment, and was analyzed by gas chromatography-mass spectrometry (GC-MS). Partial least squares discriminate analysis (PLS-DA) and the Student's t Tests with the false discovery rate (FDR) correction were conducted on the normalized data for the distinction of groups and the identification of the most contributed metabolites. Clear separation is observed between different treatments, and 29 out of 41 metabolites changes significantly compared with the corresponding controls. The concentration of threonine, glycine, serine, beta-d-glucose and oxalic acid are consistently changed in all morphine treated groups compared with controls. Through this experiment we find characteristic metabolites in different dependent states and discuss the possible compensation effects. The interpretation of these metabolites would throw light on the biological effects of morphine and reveal the possibilities to become marker of morphine addiction.

Graded Porous β-tricalcium Phosphate Scaffolds Enhance Bone Regeneration in Mandible Augmentation

The Journal of Craniofacial Surgery. Mar, 2015  |  Pubmed ID: 25675019

Bone augmentation requires scaffold to promote forming of natural bone structure. Currently, most of the reported bone scaffolds are porous solids with uniform pores. The aim of the current study is to evaluate the effect of a graded porous β-tricalcium phosphate scaffolds on alveolar bone augmentation. Three groups of scaffolds were fabricated by a template-casting method: (1) graded porous scaffolds with large pores in the center and small pores at the periphery, (2) scaffolds with large uniform pores, and (3) scaffolds with small uniform pores. Bone augmentation on rabbit mandible was investigated by microcomputed tomography, sequential fluorescent labeling, and histologic examination 3 months after implantation.The result presents that all the scaffold groups maintain their augmented bone height after 3-month observation, whereas the autografting group presents an obvious bone resorption. Microcomputed tomography reveals that the graded porous group has significantly greater volume of new bone (P < 0.05) and similar bone density compared with the uniform pores groups. Bone substance distributes unevenly in all the 3 experimental groups. Greater bone volume can be observed in the area closer to the bone bed. The sequential fluorescent labeling observation reveals robust bone regeneration in the first month and faster bone growth in the graded porous scaffold group than that in the large porous scaffold group. Histologic examinations confirm bone structure in the aspect of distribution, activity, and maturity. We conclude that graded porous designed biodegradable β-tricalcium phosphate scaffolds are beneficial to promote bone augmentation in the aspect of bone volume.

[The Research of SurgiCase CMF Software in Surgical Simulation and Prediction for Mandibular Asymmetry]

Zhonghua Zheng Xing Wai Ke Za Zhi = Zhonghua Zhengxing Waike Zazhi = Chinese Journal of Plastic Surgery. Mar, 2015  |  Pubmed ID: 26211186

To evaluate the predictive accuracy of the SurgiCase CMF software in surgical simulation and prediction for mandibular asymmetry with 3-dimensional simulation and measurement.

Evolution and Coevolution of PRC2 Genes in Vertebrates and Mammals

Advances in Protein Chemistry and Structural Biology. 2015  |  Pubmed ID: 26572978

Recruited by noncoding RNAs (ncRNAs) to specific genomic sites, polycomb repressive complexes 2 (PRC2) modify chromatin states in nearly all eukaryotes. The limited ncRNAs in Drosophila but abundant in mammals should have made PRC2 genes evolved significantly in Deuterostomia to adapt to the much increased ncRNAs. This study analyzes the evolution and coevolution of seven PRC2 genes in 29 Deuterostomia. These genes, previously assumed highly conserved, are found to have obtained multiple insertions in vertebrates and mammals and undergone significant positive selections in marsupials and prosimians, indicating adaptions to substantially increased lncRNAs (long noncoding RNAs) in mammals and in primates. Some insertions occur notably in homologous sequences of human nonsense-mediated decay (NMD) transcripts. Moreover, positive selections and signals of convergent evolution imply the independent increase of lncRNAs in mammals and in primates. Coevolutionary analysis reveals that patterns of interaction between PRC2 proteins have also much evolved from vertebrates to mammals, indicating adaptation at the protein complex level. The potential functions of mammalian-specific insertions and NMD transcripts deserve further experimental examination.

Decreased Expression of Rab27A and Rab27B Correlates with Metastasis and Poor Prognosis in Colorectal Cancer

Discovery Medicine. Dec, 2015  |  Pubmed ID: 26760980

The Rab27 subfamily of secretory small GTPase plays a vital role in vesicle trafficking and regulates tumor growth and metastasis in several cancer types. Thus, this research was designed to explore the clinical and prognostic significance of Rab27A and Rab27B in colorectal cancer (CRC) patients. Reverse transcription-polymerase chain reaction (RT-PCR), western blot, and immunohistochemistry (IHC) analysis were used to examine Rab27A and Rab27B expression in human CRC cell lines and primary tumors. The correlation of gene expression with clinicopathological features and prognosis was also evaluated. Our results indicated that Rab27A expression was down-regulated in primary tumors compared with matched adjacent tissues (100%, 8/8), as detected by western blot. IHC analysis revealed that the positive expression rate of Rab27A in primary CRC tissues was lower than in adjacent normal tissues (P = 0.005). Negative expression of Rab27A and Rab27B significantly correlated with poor tumor differentiation (both P < 0.001) and positive vascular invasion (P = 0.005, P = 0.021, respectively). Moreover, absence of Rab27A was associated with advanced tumor node metastasis (TNM) stage (P = 0.006), distant metastasis (P = 0.002), and local recurrence (P = 0.038). Survival analysis also showed a significant correlation between unfavorable survival and negative expression of Rab27A (P = 0.002). In addition, positive expression of both Rab27A and Rab27B was a protective factor in CRC. In conclusion, decreased expression of Rab27A and Rab27B, especially Rab27A, closely correlated with tumor progression and are valuable prognostic indicators in CRC patients.

Cathepsin L of the Sea Cucumber Apostichopus Japonicus-molecular Characterization and Transcriptional Response to Vibrio Splendidus Infection

Fish & Shellfish Immunology. Feb, 2016  |  Pubmed ID: 26777896

Cathepsin L, a lysosomal endopeptidase, has been noted for its involvement in the innate immune response in invertebrates. Here, the cathepsin L cDNA of the sea cucumber Apostichopus japonicus (AjCatL) is identified from an EST library and then cloned by the rapid amplification of the cDNA ends (RACE) PCR. The full-length cDNA is 1678 bp long containing an open reading frame (ORF) of 1002 bp, an 80 bp 5' UTR and a 599 bp 3' UTR. The cDNA encodes 333 amino acid residues with a predicted molecular mass of 37.07 kDa and a theoretical isoelectric point (pI) of 5.01. The full-length AjCatL contains three active sites of eukaryotic thiol (cysteine) protease at positions 133-144, 278-288 and 295-314. Analysis of the predicted tertiary structure of prepro-CatL (17-333 aa) and mature-CatL (116-333 aa) reveals that the propeptide region (17-115 aa) blocks access to the substrate-binding cleft. Phylogenetic analysis shows that the AjCatL is clustered together with two other CatLs from Strongylocentrotus purpuratus. The enzymatic activity of AjCatL was verified using a substrate hydrolyzing assay with recombinant mAjCatL. Further analysis of real time-PCR demonstrates that the expression of AjCatL mRNA is significantly up-regulated in the coelomocytes in cases of infection with the common bacterial pathogen, Vibrio splendidus. This suggests that the AjCatL is likely to be involved in the immune response.

Abundant Members of Scavenger Receptors Family and Their Identification, Characterization and Expression Against Vibrio Alginolyticus Infection in Juvenile Larimichthys Crocea

Fish & Shellfish Immunology. Mar, 2016  |  Pubmed ID: 26876357

Scavenger receptors (SRs) are crucial pattern recognition receptors (PRRs) to defense pathogen infection in fish innate immunity. In this paper, some members in SRs family of Larimichthys crocea were identified, including eight genes in the class A, B, D and F families. (G + C) % of all SRs members held 51% ∼ 59%, and these genes were no obvious codon bias by analyzing the distribution of A-, T-, G- and C-ended codons. The order of Enc for all SRs members by sequencing was LycCD68 > LycSCARA5 > LycSCARB1 > LycCD163 > LycMARCO > LycSREC1 > LycSCARA3 > LycSREC2. Moreover, different lengths and numbers of exons and introns led to the diverse mRNAs and respective functional domains or motifs, for example, an optional cysteine-rich (SRCR) domain in LycMARCO and LycSCARA5, an epidermal growth factor (EGF) and EGF-like domain in LycSREC1 and LycSREC2. The sub-cellular localization demonstrated SRs members mainly located in plasma membrane or extracellular matrix. Further, all of the SRs members in L. crocea were almost low expressed in heart, gill and intestine, whereas high in spleen and liver. After stimulation by Vibrio alginolyticus, the class A and F families were induced significantly, but the class B and D families expressed less even none after pathogenic infection. All the findings would pave the way to understand not only the evolution of the SR-mediated immune response, but also the complexity of fish immunity.

Pulp Regeneration: Current Approaches and Future Challenges

Frontiers in Physiology. 2016  |  Pubmed ID: 27014076

Regenerative endodontics aims to replace inflamed/necrotic pulp tissues with regenerated pulp-like tissues to revitalize teeth and improve life quality. Pulp revascularization case reports, which showed successful clinical and radiographic outcomes, indicated the possible clinical application of pulp regeneration via cell homing strategy. From a clinical point of view, functional pulp-like tissues should be regenerated with the characterization of vascularization, re-innervation, and dentin deposition with a regulated rate similar to that of normal pulp. Efficient root canal disinfection and proper size of the apical foramen are the two requisite preconditions for pulp regeneration. Progress has been made on pulp regeneration via cell homing strategies. This review focused on the requisite preconditions and cell homing strategies for pulp regeneration. In addition to the traditionally used mechanical preparation and irrigation, antibiotics, irrigation assisted with EndoVac apical negative-pressure system, and ultrasonic and laser irradiation are now being used in root canal disinfection. In addition, pulp-like tissues could be formed with the apical foramen less than 1 mm, although more studies are needed to determine the appropriate size. Moreover, signaling molecules including stromal cell derived factor (SDF-1α), basic Fibroblast Growth Factor (bFGF), Platelet Derived Growth Factor (PDGF), stem cell factor (SCF), and Granulocyte Colony-Stimulating Factor (G-CSF) were used to achieve pulp-like tissue formation via a cell homing strategy. Studies on the cell sources of pulp regeneration might give some indications on the signaling molecular selection. The active recruitment of endogenous cells into root canals to regenerate pulp-like tissues is a novel concept that may offer an unprecedented opportunity for the near-term clinical translation of current biology-based therapies for dental pulp regeneration.

Exome Sequencing and Gene Prioritization Correct Misdiagnosis in a Chinese Kindred with Familial Amyloid Polyneuropathy

Scientific Reports. May, 2016  |  Pubmed ID: 27212199

Inherited neuropathies show considerable heterogeneity in clinical manifestations and genetic etiologies, and are therefore often difficult to diagnose. Whole-exome sequencing (WES) has been widely adopted to make definite diagnosis of unclear conditions, with proven efficacy in optimizing patients' management. In this study, a large Chinese kindred segregating autosomal dominant polyneuropathy with incomplete penetrance was ascertained through a patient who was initially diagnosed as Charcot-Marie-Tooth disease. To investigate the genetic cause, forty-six living family members were genotyped by SNP microarrays, and one confirmed patient was subject to WES. Through systematic computational prioritization, we identified a missense mutation c.G148T in TTR gene which results in a p.V50L substitution known to cause transthyretin-related familial amyloid polyneuropathy. Co-segregation analysis and clinical follow-up confirmed the new diagnosis, which suggested new therapeutic options to the patients and informed high risk family members. This study confirms WES as a powerful tool in translational medicine, and further demostrates the practical utility of gene prioritization in narrowing the scope of causative mutation.

Agonist-mediated Activation of Bombyx Mori Diapause Hormone Receptor Signals to Extracellular Signal-regulated Kinases 1 and 2 Through Gq-PLC-PKC-dependent Cascade

Insect Biochemistry and Molecular Biology. Aug, 2016  |  Pubmed ID: 27318251

Diapause is a developmental strategy adopted by insects to survive in challenging environments such as the low temperatures of a winter. This unique process is regulated by diapause hormone (DH), which is a neuropeptide hormone that induces egg diapause in Bombyx mori and is involved in terminating pupal diapause in heliothis moths. An G protein-coupled receptor from the silkworm, B. mori, has been identified as a specific cell surface receptor for DH. However, the detailed information on the DH-DHR system and its mechanism(s) involved in the induction of embryonic diapause remains unknown. Here, we combined functional assays with various specific inhibitors to elucidate the DHR-mediated signaling pathways. Upon activation by DH, B. mori DHR is coupled to the Gq protein, leading to a significant increase of intracellular Ca(2+) and cAMP response element-driven luciferase activity in an UBO-QIC, a specific Gq inhibitor, sensitive manner. B. mori DHR elicited ERK1/2 phosphorylation in a dose- and time-dependent manner in response to DH. This effect was almost completely inhibited by co-incubation with UBO-QIC and was also significantly suppressed by PLC inhibitor U73122, PKC inhibitors Gö6983 and the Ca(2+) chelator EGTA. Moreover, DHR-induced activation of ERK1/2 was significantly attenuated by treatment with the Gβγ specific inhibitors gallein and M119K and the PI3K specific inhibitor Wortmannin, but not by the Src specific inhibitor PP2. Our data also demonstrates that the EGFR-transactivation pathway is not involved in the DHR-mediated ERK1/2 phosphorylation. Future efforts are needed to clarify the role of the ERK1/2 signaling pathway in the DH-mediated induction of B. mori embryonic diapause.

Agonist-Activated Bombyx Corazonin Receptor Is Internalized Via an Arrestin-Dependent and Clathrin-Independent Pathway

Biochemistry. Jul, 2016  |  Pubmed ID: 27348044

Agonist-induced internalization plays a key role in the tight regulation of the extent and duration of G protein-coupled receptor signaling. Previously, we have shown that the Bombyx corazonin receptor (BmCrzR) activates both Gαq- and Gαs-dependent signaling cascades. However, the molecular mechanisms involved in the regulation of the internalization and desensitization of BmCrzR remain to be elucidated. Here, vectors for expressing BmCrzR fused with enhanced green fluorescent protein (EGFP) at the C-terminal end were used to further characterize BmCrzR internalization. We found that the BmCrzR heterologously expressed in HEK-293 and BmN cells was rapidly internalized from the plasma membrane into the cytoplasm in a concentration- and time-dependent manner via a β-arrestin (Kurtz)-dependent and clathrin-independent pathway in response to agonist challenge. While most of the internalized receptors were recycled to the cell surface via early endosomes, some others were transported to lysosomes for degradation. Assays using RNA interference revealed that both GRK2 and GRK5 were essentially involved in the regulation of BmCrzR phosphorylation and internalization. Further investigations indicated that the identified cluster of Ser/Thr residues ((411)TSS(413)) was responsible for GRK-mediated phosphorylation and internalization. This is the first detailed investigation of the internalization and trafficking of Bombyx corazonin receptors.

Double-layer Versus Single-layer Bone-patellar Tendon-bone Anterior Cruciate Ligament Reconstruction: a Prospective Randomized Study with 3-year Follow-up

Archives of Orthopaedic and Trauma Surgery. Dec, 2016  |  Pubmed ID: 27568219

To evaluate the clinical results of a randomized controlled trial of single-layer versus double-layer bone-patellar tendon-bone (BPTB) anterior cruciate ligament (ACL) reconstruction.

Low Temperature and Daphnia-associated Infochemicals Promote Colony Formation of Scenedesmus Obliquus and Its Harvesting

Biotechnology Letters. Jan, 2017  |  Pubmed ID: 27654822

To explore the combined effects of temperature and Daphnia-associated infochemicals on colony formation of Scenedesmus obliquus to faciliate harvesting the algal biomass.

Preparation of Bioactive β-tricalcium Phosphate Microspheres As Bone Graft Substitute Materials

Materials Science & Engineering. C, Materials for Biological Applications. Jan, 2017  |  Pubmed ID: 27772722

In this study, β-tricalcium phosphate (Ca3PO4, β-TCP) microspheres with different diameters were fabricated via a solid-in-oil-in-water (S/O/W) emulsion method. After soaking in simulated body fluid (SBF), the fabricated β-TCP microspheres were fully covered with a new bone-like apatite layer; subsequent analysis suggested that the microspheres have excellent bioactivity properties, specifically in inducing apatite deposition. The calcium release profiles of the microspheres were tested in pH7.4 Tris-HCl buffer, and results demonstrated that the Ca(2+) continually released from microspheres during the two-week test period. We then co-cultured bone marrow stem cells (BMSCs) in vitro with β-TCP microspheres, and performed SEM and confocal microscope analyses to find that β-TCP microspheres efficiently promoted BMSC attachment and bone-related gene expression. The co-cultured BMSCs and microspheres were successfully implanted subcutaneously into nude mice for 8weeks. The H&E neo-tissue staining results showed that abundant new bone-like structures had formed between the β-TCP microspheres, implying that β-TCP microspheres used as a cell carrier and bone graft substitute material show highly promising potential application for irregular-shaped bone defect regeneration.

A Transcriptome Profile in Hepatocellular Carcinomas Based on Integrated Analysis of Microarray Studies

Diagnostic Pathology. Jan, 2017  |  Pubmed ID: 28086821

Despite new treatment options for hepatocellular carcinomas (HCC) recently, 5-year survival remains poor, ranging from 50 to 70%, which may attribute to the lack of early diagnostic biomarkers. Thus, developing new biomarkers for early diagnosis of HCC, is extremely urgent, aiming to decrease HCC-related deaths.

Specific and Spatial Labeling of P0-Cre Versus Wnt1-Cre in Cranial Neural Crest in Early Mouse Embryos

Genesis (New York, N.Y. : 2000). Mar, 2017  |  Pubmed ID: 28371069

P0-Cre and Wnt1-Cre mouse lines have been widely used in combination with loxP-flanked mice to label and genetically modify neural crest (NC) cells and their derivatives. Wnt1-Cre has been regarded as the gold standard and there have been concerns about the specificity of P0-Cre because it is not clear about the timing and spatial distribution of the P0-Cre transgene in labeling NC cells at early embryonic stages. We re-visited P0-Cre and Wnt1-Cre models in the labeling of NC cells in early mouse embryos with a focus on cranial NC. We found that R26-lacZ Cre reporter responded to Cre activity more reliably than CAAG-lacZ Cre reporter during early embryogenesis. Cre immunosignals in P0-Cre and reporter (lacZ and RFP) activity in P0-Cre/R26-lacZ and P0-Cre/R26-RFP embryos were detected in the cranial NC and notochord regions in E8.0-9.5 (4-19 somite) embryos. P0-Cre transgene expression was observed in migrating NC cells and was more extensive in the forebrain and hindbrain but not apparent in the midbrain. Differences in the Cre distribution patterns of P0-Cre and Wnt1-Cre were profound in the midbrain and hindbrain regions, i.e., extensive in the midbrain of Wnt1-Cre and in the hindbrain of P0-Cre embryos. The difference between P0-Cre and Wnt1-Cre in labeling cranial NC may provide a better explanation of the differential distributions of their NC derivatives and of the phenotypes caused by Cre-driven genetic modifications. This article is protected by copyright. All rights reserved.

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