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1.  Bone marrow mesenchymal stem cells promote osteosarcoma cell proliferation and invasion 
Background
Bone marrow-derived stem cells (BMSCs) are locally adjacent to the tumor tissues and may interact with tumor cells directly. The purpose of this study was to explore the effects of BMSCs on the proliferation and invasion of osteosarcoma cells in vitro and the possible mechanism involved.
Methods
BMSCs were co-cultured with osteosarcoma cells, and CCK-8 assay was used to measure cell proliferation. The ELISA method was used to determine the concentration of stromal cell-derived factor-1 (SDF-1) in the supernatants. Reverse transcription polymerase chain reaction (RT-PCR) was performed to detect the expression of CXCR4 in osteosarcoma cells and BMSCs. Matrigel invasion assay was performed to measure tumor cell invasion.
Results
SDF-1 was detected in the supernatants of BMSCs, but not in osteosarcoma cells. Higher CXCR4 mRNA levels were detected in the osteosarcoma cell lines compared to BMSCs. In addition, conditioned medium from BMSCs can promote the proliferation and invasion of osteosarcoma cells, and AMD3100, an antagonist for CXCR4, can significantly downregulate these growth-promoting effects.
Conclusions
BMSCs can promote the proliferation and invasion of osteosarcoma cells, which may involve the SDF-1/CXCR4 axis.
doi:10.1186/s12957-015-0465-1
PMCID: PMC4334855
Bone marrow mesenchymal stem cells; Osteosarcoma
2.  Highly Aligned Poly(3,4-ethylene dioxythiophene) (PEDOT) Nano- and Microscale Fibers and Tubes 
Polymer  2013;54(2):702-708.
This study reports a facile method for the fabrication of aligned Poly(3,4-ethylene dioxythiophene) (PEDOT) fibers and tubes based on electrospinning and oxidative chemical polymerization. Discrete PEDOT nano- and microfibers and nano- and microtubes are difficult to fabricate quickly and reproducibly. We employed poly(lactide-co-glycolide) (PLGA) polymers that were loaded with polymerizable 3,4-ethylene dioxythiophene (EDOT) monomer to create aligned nanofiber assemblies using a rotating glass mandrel during electrospinning. The EDOT monomer/PLGA polymer blends were then polymerized by exposure to an oxidative catalyst (FeCl3). PEDOT was polymerized by continuously dripping a FeCl3 solution onto the glass rod during electrospinning. The resulting PEDOT fibers were conductive, aligned and discrete. Fiber bundles could be easily produced in lengths of several centimeters. The PEDOT sheath/PLGA core fibers were immersed in chloroform to remove the PLGA and any residual EDOT resulting in hollow PEDOT tubes. This approach made it possible to easily generate large areas of aligned PEDOT fibers/tubes. The structure and properties of the aligned assemblies were measured using optical microscopy, electron microscopy, Raman spectroscopy, thermal gravimetric analysis, and DC conductivity measurements. We also demonstrated that the aligned PEDOT sheath/PLGA core fiber assemblies could be used in supporting and directing the extension of dorsal root ganglia (DRG) neurons in vitro.
doi:10.1016/j.polymer.2012.10.057
PMCID: PMC4322418
3.  Identification of Transcription Factor AML-1 Binding Site Upstream of Human Cytomegalovirus UL111A Gene 
PLoS ONE  2015;10(2):e0117773.
Human cytomegalovirus (HCMV) interleukin-10 (hcmvIL-10), encoded by HCMV UL111A gene, is a homolog of human IL-10. It exerts immunomodulatory effects that allow HCMV to evade host defense mechanisms. However, the exact mechanism underlying the regulation of hcmvIL-10 expression is not well understood. The transcription factor acute myeloid leukemia 1 (AML-1) plays an important role in the regulation of various genes involved in the differentiation of hematopoietic lineages. A putative AML-1 binding site is present within the upstream regulatory region (URR) of UL111A gene. To provide evidence that AML-1 is involved in regulating UL111A gene expression, we examined the interaction of AML-1 with the URR of UL111A in HCMV-infected human monocytic THP-1 cells using a chromatin immunoprecipitation assay. HcmvIL-10 transcription was detected in differentiated THP-1 cells, but not in undifferentiated ones. Furthermore, the URR of UL111A showed a higher intensity of AML-1 binding, a higher level of histone H3 acetyl-K9, but a lower level of histone H3 dimethyl-K9 in differentiated THP-1 cells than undifferentiated cells. Down-regulation of AML1 by RNA interference decreased the expression of the UL111A gene. Our results suggest that AML-1 may contribute to the epigenetic regulation of UL111A gene via histone modification in HCMV-infected differentiated THP-1 cells. This finding could be useful for the development of new anti-viral therapies.
doi:10.1371/journal.pone.0117773
PMCID: PMC4320089  PMID: 25658598
4.  The glycosyltransferase involved in thurandacin biosynthesis catalyzes both O- and S-glycosylation 
The S-glycosyltransferase SunS is a recently discovered enzyme that selectively catalyzes the conjugation of carbohydrates to the cysteine thiol of proteins. This study reports the discovery of a second S-glycosyltransferase, ThuS, and shows that ThuS catalyzes both S-glycosylation of the thiol of cysteine and O-glycosylation of the hydroxyl group of serine in peptide substrates. ThuS-catalyzed S-glycosylation is more efficient than O-glycosylation and the enzyme demonstrates high tolerance with respect to both nucleotide sugars and peptide substrates. The biosynthesis of the putative products of the thuS gene cluster were reconstituted in vitro and the resulting S-glycosylated peptides thurandacin A and thurandacin B exhibit highly selective antimicrobial activity towards Bacillus thuringiensis.
doi:10.1021/ja411159k
PMCID: PMC3913795  PMID: 24325644
5.  A referencing strategy for the direct comparison of NMR and MD motional parameters in RNA 
Nuclear magnetic resonance (NMR) spectroscopy and molecular dynamics (MD) simulations are both techniques that can be used to characterize the structural dynamics of biomolecules and their underlying timescales. Comparison of relaxation parameters obtained through each methodology allows for cross validation of techniques, and for complementarity in the analysis of dynamics. Here we present a combined NMR/MD study of the dynamics of HIV-1 TAR RNA. We compute relaxation constants (R1, R2, and NOE) and model-free parameters (S2 and τ) from a 65 ns molecular dynamics (MD) trajectory and compare them with the respective parameters measured in a domain-elongation NMR experiment. Using the elongated domain as the frame of reference for all computed parameters allows for a direct comparison between experiment and simulation. We see good agreement for many parameters and gain further insight into the nature of the local and global dynamics of TAR, which are found to be quite complex, spanning multiple timescales. For the few cases where agreement is poor, comparison of the dynamical parameters provides insight into the limits of each technique. We suggest a frequency-matching procedure that yields an upper bound for the timescale of dynamics to which the NMR relaxation experiment is sensitive.
doi:10.1021/jp905286h
PMCID: PMC4287414  PMID: 20039757
6.  Efficacy and safety of gemcitabine-based chemotherapies in biliary tract cancer: A meta-analysis 
World Journal of Gastroenterology : WJG  2014;20(47):18001-18012.
AIM: To investigate the efficacy and safety of gemcitabine (Gem)-based combination chemotherapies for the treatment of advanced biliary tract cancer.
METHODS: Clinical trials were identified by searching scientific literature databases (PubMed, EMBASE and the Cochrane Library) for studies published between 1975 and 2013. Two reviewers independently evaluated the relevant studies and manually searched references from these reports to locate additional eligible studies. The disease response and control rates, progression-free and overall survivals, and the grade 3-4 toxicities were evaluated by a meta-analysis. Odds-ratios (ORs) of the disease response and control rates and grade 3-4 toxicities, and the mean difference (MD) of both progression-free and overall survivals were calculated and used for statistical analysis.
RESULTS: Seven randomized trials with a total of 858 patients were selected and included in the final analysis. The studies were divided into subgroups based on the chemotherapy regimens, including Gem-based and non-Gem-based chemotherapies. The overall analyses revealed that the patients treated with Gem-based combination chemotherapy had significantly higher disease response rates [OR = 1.69, 95% confidence interval (CI): 1.17-2.43; P = 0.01], a longer progression-free survival (MD = 1.95, 95%CI: 0.90-3.00; P = 0.00) and a longer overall survival (MD = 1.85, 95%CI: 0.26-3.44; P = 0.02). A higher incidence of grade 3-4 hematological toxicities, including leukopenia (OR = 2.98, 95%CI: 1.44-6.20; P = 0.00), anemia (OR = 2.96, 95%CI: 1.79-4.92; P = 0.00) and neutropenia (OR = 2.80, 95%CI: 1.39-5.64; P = 0.00) was found in the Gem-based combination chemotherapy group compared with the Gem monotherapy and non-Gem-based chemotherapy groups.
CONCLUSION: Gem-based combination chemotherapy is a potential first-line treatment for advanced biliary tract cancer as a result of improved survival, though with additional toxicity.
doi:10.3748/wjg.v20.i47.18001
PMCID: PMC4273152  PMID: 25548500
Biliary tract cancer; Combination chemotherapy; Gemcitabine; Meta-analysis; Randomized trial
7.  The Achyranthes bidentata polypeptide k fraction enhances neuronal growth in vitro and promotes peripheral nerve regeneration after crush injury in vivo 
Neural Regeneration Research  2014;9(24):2142-2150.
We have previously shown that Achyranthes bidentata polypeptides (ABPP), isolated from Achyranthes bidentata Blume (a medicinal herb), exhibit neurotrophic and neuroprotective effects on the nervous system. To identify the major active component of ABPP, and thus optimize the use of ABPP, we used reverse-phase high performance liquid chromatography to separate ABPP. We obtained 12 fractions, among which the fraction of ABPPk demonstrated the strongest neuroactivity. Immunocytochemistry and western blot analysis showed that ABPPk promoted neurite growth in cultured dorsal root ganglion explant and dorsal root ganglion neurons, which might be associated with activation of Erk1/2. A combination of behavioral tests, electrophysiological assessment, and histomorphometric analysis indicated that ABPPk enhanced nerve regeneration and function restoration in a mouse model of crushed sciatic nerve. All the results suggest that ABPPk, as the key component of ABPP, can be used for peripheral nerve repair to yield better outcomes than ABPP.
doi:10.4103/1673-5374.147948
PMCID: PMC4316447  PMID: 25657735
nerve regeneration; Achyranthes bidentata polypeptides; neuroactive component; dorsal root ganglion; neurite outgrowth; crush injury; sciatic nerve; peripheral nerve regeneration; neural regeneration
8.  The Effects of Prediction on the Perception for Own-Race and Other-Race Faces 
PLoS ONE  2014;9(11):e114011.
Human beings do not passively perceive important social features about others such as race and age in social interactions. Instead, it is proposed that humans might continuously generate predictions about these social features based on prior similar experiences. Pre-awareness of racial information conveyed by others' faces enables individuals to act in “culturally appropriate” ways, which is useful for interpersonal relations in different ethnicity groups. However, little is known about the effects of prediction on the perception for own-race and other-race faces. Here, we addressed this issue using high temporal resolution event-related potential techniques. In total, data from 24 participants (13 women and 11 men) were analyzed. It was found that the N170 amplitudes elicited by other-race faces, but not own-race faces, were significantly smaller in the predictable condition compared to the unpredictable condition, reflecting a switch to holistic processing of other-race faces when those faces were predictable. In this respect, top-down prediction about face race might contribute to the elimination of the other-race effect (one face recognition impairment). Furthermore, smaller P300 amplitudes were observed for the predictable than for unpredictable conditions, which suggested that the prediction of race reduced the neural responses of human brains.
doi:10.1371/journal.pone.0114011
PMCID: PMC4244206  PMID: 25422892
9.  The role of IMP dehydrogenase 2 in Inauhzin-induced ribosomal stress 
eLife  2014;3:e03077.
The ‘ribosomal stress (RS)-p53 pathway’ is triggered by any stressor or genetic alteration that disrupts ribosomal biogenesis, and mediated by several ribosomal proteins (RPs), such as RPL11 and RPL5, which inhibit MDM2 and activate p53. Inosine monophosphate (IMP) dehydrogenase 2 (IMPDH2) is a rate-limiting enzyme in de novo guanine nucleotide biosynthesis and crucial for maintaining cellular guanine deoxy- and ribonucleotide pools needed for DNA and RNA synthesis. It is highly expressed in many malignancies. We previously showed that inhibition of IMPDH2 leads to p53 activation by causing RS. Surprisingly, our current study reveals that Inauzhin (INZ), a novel non-genotoxic p53 activator by inhibiting SIRT1, can also inhibit cellular IMPDH2 activity, and reduce the levels of cellular GTP and GTP-binding nucleostemin that is essential for rRNA processing. Consequently, INZ induces RS and the RPL11/RPL5-MDM2 interaction, activating p53. These results support the new notion that INZ suppresses cancer cell growth by dually targeting SIRT1 and IMPDH2.
DOI: http://dx.doi.org/10.7554/eLife.03077.001
eLife digest
Cancer develops when cells lose the ability to control their own growth. About half of cancerous tumors carry a dysfunctional version of a protein called p53, while the other half have defects in proteins that are important for p53's production and function. When a healthy cell is exposed to damaging chemicals or agents, the p53 protein triggers responses that are aimed at repairing the damage. However, if these attempts fail, p53 causes the damaged cell to essentially destroy itself.
As defects in p53-controlled processes cause cells to grow unrestrictedly and can lead to cancer, it is a very attractive target for cancer therapies. Cancer drug developments have focused on both targeting p53 directly and targeting the proteins that work with p53. Two proteins called Mdm2 and SIRT1 are of particular interest. Mdm2 binds to, inactivates, and leads to the degradation of p53. SIRT1 can modify p53 and make it more accessible to Mdm2, and is often found in very high levels in cancer cells.
In 2012, researchers identified Inauhzin as a small molecule that could potentially be used to treat tumors that still have a functional version of the p53 protein. Inauhzin was thought to work by inhibiting SIRT1, which increases p53 levels—probably through its effects on Mdm2. This restores the cell's ability to control its growth and to die if it is irreparably damaged. However, not all of this small molecule's effects on cells can be explained by its interaction with SIRT1.
Now Zhang et al., including some of the researchers involved in the 2012 work, have investigated whether Inauhzin also interacts with other proteins in the cell; and Inauhzin was revealed to bind an enzyme called IMPDH2. This enzyme is involved in making GTP—a small molecule that is involved in many important processes in living cells. Zhang et al. demonstrated that Inauhzin's effect on the IMPDH enzyme triggered a response that did not involve the SIRT1 protein, and that ultimately led to a decrease in Mdm2 activity and restored p53 activity.
Cancer treatments often include a combination of drugs that target different proteins with the goal of reducing the likelihood of a tumor becoming resistant to the treatment. Inauhzin's effect on two different proteins that lead to p53 activation not only increases its potency, but also makes it less likely that drug resistance will develop.
DOI: http://dx.doi.org/10.7554/eLife.03077.002
doi:10.7554/eLife.03077
PMCID: PMC4209374  PMID: 25347121
Inauhzin; p53; ribosomal stress; IMPDH2; nucleostemin; MDM2; human
10.  A Homeostatic Sleep-Stabilizing Pathway in Drosophila Composed of the Sex Peptide Receptor and Its Ligand, the Myoinhibitory Peptide 
PLoS Biology  2014;12(10):e1001974.
A ligand of the sex peptide receptor maintains sleep stability and homeostasis by inhibiting wakefulness-promoting neurons in Drosophila.
Sleep, a reversible quiescent state found in both invertebrate and vertebrate animals, disconnects animals from their environment and is highly regulated for coordination with wakeful activities, such as reproduction. The fruit fly, Drosophila melanogaster, has proven to be a valuable model for studying the regulation of sleep by circadian clock and homeostatic mechanisms. Here, we demonstrate that the sex peptide receptor (SPR) of Drosophila, known for its role in female reproduction, is also important in stabilizing sleep in both males and females. Mutants lacking either the SPR or its central ligand, myoinhibitory peptide (MIP), fall asleep normally, but have difficulty in maintaining a sleep-like state. Our analyses have mapped the SPR sleep function to pigment dispersing factor (pdf) neurons, an arousal center in the insect brain. MIP downregulates intracellular cAMP levels in pdf neurons through the SPR. MIP is released centrally before and during night-time sleep, when the sleep drive is elevated. Sleep deprivation during the night facilitates MIP secretion from specific brain neurons innervating pdf neurons. Moreover, flies lacking either SPR or MIP cannot recover sleep after the night-time sleep deprivation. These results delineate a central neuropeptide circuit that stabilizes the sleep state by feeding a slow-acting inhibitory input into the arousal system and plays an important role in sleep homeostasis.
Author Summary
Sleep is a common trait in animals, from insects to mammals, and it needs to be coordinated with other critical activities such as feeding and reproduction. However, the mechanisms by which this is achieved are not fully understood. The fruit fly Drosophila melanogaster has become a key model organism for sleep research and it has been shown that reproduction is one of the factors that can modulate sleep in these animals. Researchers have observed that mating reduces the daytime sleep of female flies and shown that the seminal fluid protein Sex Peptide (SP), a ligand of the Sex Peptide Receptor (SPR) that is transferred to females during copulation, is responsible for this reduction of siesta sleep. Here, we investigated further the role of SPR in sleep regulation in Drosophila. We show that SPR is required for sleep stabilization in both sexes and that in mutant flies lacking SPR or its ligand myoinhibitory peptide (MIP) sleep is fragmented independently of reproduction. Unlike SP, MIP is expressed in the brain of both sexes and acts on SPR to silence specific neurons that keep flies awake, stabilizing sleep. Hence, our results reveal that SPR interacts with two distinct ligands to control different behaviors: SP for reproduction and MIP for sleep.
doi:10.1371/journal.pbio.1001974
PMCID: PMC4204809  PMID: 25333796
11.  Imaging cytosolic translocation of Mycobacteria with two-photon fluorescence resonance energy transfer microscopy 
Biomedical Optics Express  2014;5(11):3990-4001.
Transition from latency to active tuberculosis requires Mycobacterium tuberculosis (Mtb) to penetrate the phagosomal membrane and translocate to the cytosol of the host macrophage. Quantitative two-photon fluorescence resonance energy transfer (FRET) microscopy is developed to measure cytosolic translocation using Mycobacterium marinum (Mm) as a model organism for Mtb. Macrophages were infected with Mm or non-pathogenic Mycobacterium smegmatis (Ms) as a control, then loaded with a FRET substrate. Once translocation occurs, mycobacterium-bearing β-lactamase cleaves the substrate, resulting in decrease of FRET signal. Quantification of this FRET signal change revealed that Mm, but not Ms, is capable of translocating to the cytosol.
doi:10.1364/BOE.5.003990
PMCID: PMC4242033  PMID: 25426325
(170.2520) Fluorescence microscopy; (170.1530) Cell analysis
12.  An Active Component of Achyranthes bidentata Polypeptides Provides Neuroprotection through Inhibition of Mitochondrial-Dependent Apoptotic Pathway in Cultured Neurons and in Animal Models of Cerebral Ischemia 
PLoS ONE  2014;9(10):e109923.
An active component has been isolated by reverse-phase high performance liquid chromatography (HPLC) from Achyranthes bidentata Blume polypeptides that are extracted from Achyranthes bidentata Blume, a Chinese medicinal herb. The active component is called ABPPk based on the order of HPLC elution. In this study, we used in vitro and in vivo experimental models of cerebral ischemia to investigate the possible neuroprotective effect of ABPPk. ABPPk treatment promoted neuronal survival and inhibited neuronal apoptosis in primary cortical neurons exposed to oxygen and glucose deprivation and in rats subjected to transient middle cerebral artery occlusion. The role of ABPPk in protection against ischemia-induced neuronal damage might be mediated by mitochondrial-dependent pathways, including modulation of apoptosis-related gene expression, regulation of mitochondrial dysfunction through restoring mitochondrial membrane potential, reducing release of mitochondrial apoptogenic factors, and inhibiting intracellular ROS production. The neuroprotective effect of ABPPk may suggest the possible use of this agent in the treatment and prevention of cerebral ischemic stroke.
doi:10.1371/journal.pone.0109923
PMCID: PMC4198176  PMID: 25334016
13.  TET3 mediates the activation of human hepatic stellate cells via modulating the expression of long non-coding RNA HIF1A-AS1 
Activated Hepatic stellate cells (HSCs) play a critical role in liver fibrosis and a lot of efforts have been made to dissect the underlying mechanism involved in activation of HSCs. However, the underlying mechanism remains douteux up to now. In the present study, we found that TET3, one member of ten-eleven translocation (TET) protein family, reduced significantly in HSCs LX-2 activated by TGF-β1. To study the function of TET3 in activation of HSCs, knockdown was performed by RNA interference. Results showed that cell proliferation rise significantly and cell apoptosis reduce obviously after knockdown of TET3. Meanwhile, IHC showed that the expression of α-SMA rise significantly compared to control. These results indicated that TET3 is closely associated with the activation of HSCs. Further studies found that long non-coding RNA HIF1A-AS1 was reduced significantly in LX-2 cell after treatment with siRNA for TET3. The result hinted that TET3 activate HSCs through modulating the expression of HIF1A-AS1. To confirm this hypothesis, RNA interference was performed to silence the HIF1A-AS1. Results showed that HIF1A-AS1 silencing lead to enhancing in cell proliferation and declining apoptosis. Taken together, TET3 can mediate the activation of HSCs via modulating the expression of the long non-coding RNA HIF1A-AS1.
PMCID: PMC4270585  PMID: 25550811
Liver fibrosis; HSCs; TET3; long non-coding RNA; HIF1A-AS1
14.  Rice and cold stress: methods for its evaluation and summary of cold tolerance-related quantitative trait loci 
Rice  2014;7(1):24.
Cold stress adversely affects rice (Oryza sativa L.) growth and productivity, and has so far determined its geographical distribution. Dissecting cold stress-mediated physiological changes and understanding their genetic causes will facilitate the breeding of rice for cold tolerance. Here, we review recent progress in research on cold stress-mediated physiological traits and metabolites, and indicate their roles in the cold-response network and cold-tolerance evaluation. We also discuss criteria for evaluating cold tolerance and evaluate the scope and shortcomings of each application. Moreover, we summarize research on quantitative trait loci (QTL) related to cold stress at the germination, seedling, and reproductive stages that should provide useful information to accelerate progress in breeding cold-tolerant rice.
doi:10.1186/s12284-014-0024-3
PMCID: PMC4182278  PMID: 25279026
Cold tolerance; Physiological metabolites; Evaluation criteria; QTL; Oryza sativa
15.  Metabolomic analysis of simvastatin and fenofibrate intervention in high-lipid diet-induced hyperlipidemia rats 
Acta Pharmacologica Sinica  2014;35(10):1265-1273.
Aim:
To investigate the metabolite changes caused by simvastatin or fenofibrate intervention in diet-induced hyperlipidemia rats using a GC-MS-based metabolomic profiling approach.
Methods:
SD rats were fed with high-lipid diet for 4 weeks to induce hyperlipidemia, then the rats were fed with normal diet, and orally administered with simvastatin (10 mg·kg−1·d−1) or fenofibrate (150 mg·kg−1·d−1) for 2 weeks. Blood samples were collected once a week, and potential biomarkers were examined using commercial assay kits and a metabolomic approach. The metabolomics data were analyzed using a multivariate statistical technique and a principal component analysis (PCA).
Results:
Oral administration of simvastatin or fenofibrate significantly decreased the plasma levels of total cholesterol (TC) and low-density lipoprotein (LDL) cholesterol and increased the plasma level of high-density lipoprotein (HDL) cholesterol in the hyperlipidemia rats. Plasma samples were scattered in the PCA scores plots in response to the diet and to the drugs administered. The main metabolites changed in the hyperlipidemia rats were cholesterol, creatinine, linoleic acid, β-hydroxybutyric acid, tyrosine, isoleucine and ornithine. The plasma level of creatinine was significantly lower in the simvastatin-treated rats than in the fenofibrate-treated rats. The plasma tyrosine concentration was declined following intake of high-lipid diet, which was reversed by fenobrate, but not by simvastatin.
Conclusion:
A series of potential biomarkers including tyrosine, creatinine, linoleic acid, β-hydroxybutyric acid and ornithine have been identified by metabolomic profiling, which may be used to identify the metabolic changes during hyperlipidemia progression.
doi:10.1038/aps.2014.72
PMCID: PMC4186989  PMID: 25220639
simvastatin; fenofibrate; hyperlipidemia; metabolomics; biomarker; creatinine; LDL cholesterol; GC-MS
16.  ROCK Is Involved in Vasculogenic Mimicry Formation in Hepatocellular Carcinoma Cell Line 
PLoS ONE  2014;9(9):e107661.
Ras homolog family member A (RhoA) and Rho-associated coiled coil-containing protein kinases 1 and 2 (ROCK1 and 2) are key regulators of focal adhesion, actomyosin contraction and cell motility. RhoA/ROCK signaling has emerged as an attractive target for the development of new cancer therapeutics. Whether RhoA/ROCK is involved in regulating the formation of tumor cell vasculogenic mimicry (VM) is largely unknown. To confirm this hypothesis, we performed in vitro experiments using hepatocellular carcinoma (HCC) cell lines. Firstly, we demonstrated that HCC cells with higher active RhoA/ROCK expression were prone to form VM channels, as compared with RhoA/ROCK low-expressing cells. Furthermore, Y27632 (a specific inhibitor of ROCK) rather than exoenzyme C3 (a specific inhibitor of RhoA) effectively inhibited the formation of tubular network structures in a dose-dependent manner. To elucidate the possible mechanism of ROCK on VM formation, real-time qPCR, western blot and immunofluorescence were used to detect changes of the key VM-related factors, including VE-cadherin, erythropoietin-producing hepatocellular carcinoma-A2 (EphA2), phosphoinositide 3-kinase (PI3K), matrix metalloproteinase (MMP)14, MMP2, MMP9 and laminin 5γ2-chain (LAMC2), and epithelial-mesenchymal-transition (EMT) markers: E-cadherin and Vimentin. The results showed that all the expression profiles were attenuated by blockage of ROCK. In addition, in vitro cell migration and invasion assays showed that Y27632 inhibited the migration and invasion capacity of HCC cell lines in a dose-dependent manner markedly. These data indicate that ROCK is an important mediator in the formation of tumor cell VM, and suggest that ROCK inhibition may prove useful in the treatment of VM in HCC.
doi:10.1371/journal.pone.0107661
PMCID: PMC4169566  PMID: 25238232
17.  Optimal enamel conditioning strategy for rebonding orthodontic brackets: a laboratory study 
Objective: To compare the conventional etching and primer method (CEP) and the self-etching primer method (SEP) in rebonding brackets. Methods: Forty human maxillary second premolars extracted for orthodontic purpose were randomly divided into 4 equal groups. Group 1 and Group 2 were bonded using the CEP method; Group 3 and Group 4 using the SEP method. All the brackets were debonded and 40 new brackets were rebonded with four different protocols after surface cleaning: Group 1: CEP + adhesive; Group 2: CEP without etch step + adhesive; Group 3: SEP + adhesive; Group 4: non-acidic primer + adhesive. Then, the shear bond strength (SBS) of each group was tested and the measurements of adhesive remnant index scores (ARI) and SEM examination were performed. Results: The mean SBSs for Group 1, 2, 3 and 4 were 14.18, 6.57, 11.90, 5.91 MPa, respectively. Statistical differences of the SBS existed between Group 1 and 2 (P < 0.05) and between Group 3 and 4 (P < 0.05). No difference was found between Group 1 and 3, or Group 2 and 4. Conclusion: Omission of the acid-etching step in rebonding orthodontic brackets may be adequate for the clinical requirement. No differences in SBS and ARI of the rebonded brackets were showed between CEP and SEP methods.
PMCID: PMC4211778  PMID: 25356128
Orthodontics; dental bonding; acid-etching; bond strength
18.  New Diagnosis and Therapy Model for Ischemic-Type Biliary Lesions following Liver Transplantation—A Retrospective Cohort Study 
PLoS ONE  2014;9(9):e105795.
Ischemic-type biliary lesions (ITBLs) are a major cause of graft loss and mortality after orthotopic liver transplantation (OLT). Impaired blood supply to the bile ducts may cause focal or extensive damage, resulting in intra- or extrahepatic bile duct strictures or dilatations that can be detected by ultrasonography, computed tomography, magnetic resonance cholangiopancreatography, and cholangiography. However, the radiographic changes occur at an advanced stage, after the optimal period for therapeutic intervention. Endoscopic retrograde cholangio-pancreatography (ERCP) and percutaneous transhepatic cholangiodrainage (PTCD) are the gold standard methods of detecting ITBLs, but these procedures cannot be used for continuous monitoring. Traditional methods of follow-up and diagnosis result in delayed diagnosis and treatment of ITBLs. Our center has used the early diagnosis and intervention model (EDIM) for the diagnosis and treatment of ITBLs since February 2008. This model mainly involves preventive medication to protect the epithelial cellular membrane of the bile ducts, regular testing of liver function, and weekly monitor of contrast-enhanced ultrasonography (CEUS) to detect ischemic changes to the bile ducts. If the liver enzyme levels become abnormal or CEUS shows low or no enhancement of the wall of the hilar bile duct during the arterial phase, early ERCP and PTCD are performed to confirm the diagnosis and to maintain biliary drainage. Compared with patients treated by the traditional model used prior to February 2008, patients in the EDIM group had a lower incidence of biliary tract infection (28.6% vs. 48.6%, P = 0.04), longer survival time of liver grafts (24±9.6 months vs. 17±12.3 months, P = 0.02), and better outcomes after treatment of ITBLs.
doi:10.1371/journal.pone.0105795
PMCID: PMC4156319  PMID: 25192214
19.  Isolation of alpaca anti-idiotypic heavy chain single domain antibody for the aflatoxin immunoassay 
Analytical chemistry  2013;85(17):8298-8303.
Anti-idiotypic antibodies recognize the antigenic determinants of an antibody, thus can be used as surrogate antigens. Single domain antibodies from camlid heavy chain antibodies with the benefit features of small size, thermostability and ease in expression, are leading candidates to produce anti-idiotypic antibodies. In this work, we constructed an antibody phage library from the mRNA of an alpaca immunized with an anti-aflatoxin monoclonal antibody (MAb) 1C11. Three anti-idiotypic VHH antibodies were isolated and applied to immunoassay towards aflatoxin as a coating antigen. The best immunoassay developed with one of these VHH antibodies shows an IC50 of 0.16 ng/mL towards aflatoxin B1 and cross-reactivity towards aflatoxin B2, G1 and G2 of 90.4%, 54.4% and 37.7%, respectively. The VHH-based immunoassay was successfully applied to the analysis of peanuts, corn and rice, which are the predominant commodities regularly contaminated by aflatoxins. A good correlation (r2=0.89) was found between the data obtained from the conventional ELISA and the ELISA based on a VHH coating antigen for the analysis of aflatoxins in peanuts and feedstuff. The use of biotechnology in developing the surrogate, the absence of standard aflatoxin and organic solvents in the synthesis procedures, and the reproducibility of the VHH antibody makes it an ideal strategy for replacing conventional synthesized antigens.
doi:10.1021/ac4015885
PMCID: PMC3787825  PMID: 23965250
20.  An Ultra-Sensitive Monoclonal Antibody-Based Competitive Enzyme Immunoassay for Sterigmatocystin in Cereal and Oil Products 
PLoS ONE  2014;9(9):e106415.
Sterigmatocystin (STG), a biosynthesis precursor of aflatoxin B1, is well known for its toxic and carcinogenic effects in humans and animals. STG derivatives and protein conjugates are needed for generation of monoclonal antibodies (mAbs). This work describes a reliable and fast synthesis of novel STG derivatives, based on which novel STG bovine serum albumin conjugates were prepared. With the novel STG bovine serum albumin conjugates, three sensitive and specific mAbs against STG, named VerA 3, VerA 4, and VerA 6, were prepared by semi-solid hypoxanthine/aminopterin/thymidine (HAT) medium using a modified two-step screening procedure. They exhibited high affinity for STG and no cross-reactivity (CR) with aflatoxins B1, B2, G1, G2, and M1. Based on the most sensitive antibody VerA 3, an ultra-sensitive competitive enzyme-linked immunosorbent assay (ELISA) was developed for STG in wheat, maize, and peanuts. Assays were performed in the STG-GA-BSA-coated (0.5 µg·mL−1) ELISA format, in which the antibody was diluted to 1∶80,000. Several physicochemical factors influencing assay performance, such as pH, ionic strength, blocking solution, and diluting solution, were optimized. The final results showed that the assays had the detection limits of 0.08 ng·g−1 for wheat, 0.06 ng·g−1 for maize, and 0.1 ng·g−1 for peanuts, inter-assay and intra-assay variations of less than 10%, and recoveries ranging from 83% to 110%. These recoveries were in good agreement with those obtained by using HPLC-MS/MS method (90–104%), indicating the importance of the mAb VerA 3 in the study of STG in crude agricultural products.
doi:10.1371/journal.pone.0106415
PMCID: PMC4153633  PMID: 25184275
21.  Nucleus-targeted Dmp1 transgene fails to rescue dental defects in Dmp1 null mice 
Dentin matrix protein 1 (DMP1) is essential to odontogenesis. Its mutations in human subjects lead to dental problems such as dental deformities, hypomineralization and periodontal impairment. Primarily, DMP1 is considered as an extracellular matrix protein that promotes hydroxyapatite formation and activates intracellular signaling pathway via interacting with αvβ3 integrin. Recent in vitro studies suggested that DMP1 might also act as a transcription factor. In this study, we examined whether full-length DMP1 could function as a transcription factor in the nucleus and regulate odontogenesis in vivo. We first demonstrated that a patient with the DMP1 M1V mutation, which presumably causes a loss of the secretory DMP1 but does not affect the nuclear translocation of DMP1, shows a typical rachitic tooth defect. Furthermore, we generated transgenic mice expressing NLSDMP1, in which the endoplasmic reticulum (ER) entry signal sequence of DMP1 was replaced by a nuclear localization signal (NLS) sequence, under the control of a 3.6 kb rat type I collagen promoter plus a 1.6 kb intron 1. We then crossbred the NLSDMP1 transgenic mice with Dmp1 null mice to express the NLSDMP1 in Dmp1-deficient genetic background. Although immunohistochemistry demonstrated that NLSDMP1 was localized in the nuclei of the preodontoblasts and odontoblasts, the histological, morphological and biochemical analyses showed that it failed to rescue the dental and periodontal defects as well as the delayed tooth eruption in Dmp1 null mice. These data suggest that the full-length DMP1 plays no apparent role in the nucleus during odontogenesis.
doi:10.1038/ijos.2014.44
PMCID: PMC4170153  PMID: 25105818
autosomal recessive hypophosphatemic rickets; dentin matrix protein 1; development; odontoblast; odontogenesis
24.  High fidelity quantum state transfer in electromechanical systems with intermediate coupling 
Scientific Reports  2014;4:6237.
Hybrid quantum systems usually consist of two or more subsystems, which may take the advantages of the different systems. Recently, the hybrid system consisting of circuit electromechanical subsystems have attracted great attention due to its advanced fabrication and scalable integrated photonic circuit techniques. Here, we propose a scheme for high fidelity quantum state transfer between a superconducting qubit and a nitrogen-vacancy center in diamond, which are coupled to a superconducting transmission-line resonator with coupling strength g1 and a nanomechanical resonator with coupling strength g2, respectively. Meanwhile, the two resonators are parametrically coupled with coupling strength J. The system dynamics, including the decoherence effects, is numerical investigated. It is found that both the small () and large () coupling regimes of this hybrid system can not support high fidelity quantum state transfer before significant technique advances. However, in the intermediate coupling regime (J ~ g1 ~ g2), in contrast to a conventional wisdom, high fidelity quantum information transfer can be implemented, providing a promising route towards high fidelity quantum state transfer in similar coupled resonators systems.
doi:10.1038/srep06237
PMCID: PMC4148701  PMID: 25168206
25.  Transcriptional Regulation of Cell Cycle Genes in Response to Abiotic Stresses Correlates with Dynamic Changes in Histone Modifications in Maize 
PLoS ONE  2014;9(8):e106070.
The histone modification level has been shown to be related with gene activation and repression in stress-responsive process, but there is little information on the relationship between histone modification and cell cycle gene expression responsive to environmental cues. In this study, the function of histone modifications in mediating the transcriptional regulation of cell cycle genes under various types of stress was investigated in maize (Zea mays L.). Abiotic stresses all inhibit the growth of maize seedlings, and induce total acetylation level increase compared with the control group in maize roots. The positive and negative regulation of the expression of some cell cycle genes leads to perturbation of cell cycle progression in response to abiotic stresses. Chromatin immunoprecipitation analysis reveals that dynamic histone acetylation change in the promoter region of cell cycle genes is involved in the control of gene expression in response to external stress and different cell cycle genes have their own characteristic patterns for histone acetylation. The data also showed that the combinations of hyperacetylation and hypoacetylation states of specific lysine sites on the H3 and H4 tails on the promoter regions of cell cycle genes regulate specific cell cycle gene expression under abiotic stress conditions, thus resulting in prolonged cell cycle duration and an inhibitory effect on growth and development in maize seedlings.
doi:10.1371/journal.pone.0106070
PMCID: PMC4149478  PMID: 25171199

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