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1.  Highly efficient generation of airway and lung epithelial cells from human pluripotent stem cells 
Nature biotechnology  2013;32(1):84-91.
The ability to generate lung and airway epithelial cells from human pluripotent stem cells (hPSCs) would have applications in regenerative medicine, drug screening and modeling of lung disease, and studies of human lung development. We established, based on developmental paradigms, a highly efficient method for directed differentiation of hPSCs into lung and airway epithelial cells. Long-term differentiation in vivo and in vitro yielded basal, goblet, Clara, ciliated, type I and type II alveolar epithelial cells. Type II alveolar epithelial cells generated were capable of surfactant protein-B uptake and stimulated surfactant release, providing evidence of specific function. Inhibiting or removing agonists to signaling pathways critical for early lung development in the mouse—retinoic acid, Wnt and BMP—recapitulated defects in corresponding genetic mouse knockouts. The capability of this protocol to generate most cell types of the respiratory system suggests its utility for deriving patient-specific therapeutic cells.
PMCID: PMC4101921  PMID: 24291815
2.  Improved Survival of Mice After Total Body Irradiation with 10 MV Photon, 2400 MU/min SRS Beam 
In vivo (Athens, Greece)  2014;28(1):1-12.
We evaluated the radiobiological effects of stereotactic radiosurgery (SRS) photon beams on survival of C57BL/6NTac mice following total body irradiation.
Materials and Methods
Survival of Lewis lung carcinoma (3LL) cells was tested after irradiation using 6 MV: 300 MU/min or 1400 MU/min; or 10 MV: 300 MU/min or 2400 MU/min. Survival of C57BL/6NTac mice after a dose which is lethal to 50% of the mice in 30 days (LD50/30) (9.25 Gy) total body irradiation (TBI) and 21 Gy to orthotopic 3LL tumors was tested. We quantitated levels of organ-specific gene transcripts by Real Time Polymerase Chain Reaction (RT-PCR).
While 3LL cell survival and inhibition of orthotopic tumor growth was uniform, 10 MV photons at 2400 MU/min TBI led to significantly greater survival (p=0.0218), with higher levels of intestinal (Sod2), (Gpx1), (Nrf2), and (NFκB) RNA transcripts.
Clinical 10 MV-2400 cGy/min SRS beams led to unexpected protection of mice on TBI and increased radioprotective gene transcripts.
PMCID: PMC4046118  PMID: 24425830
Stereotactic radiosurgery; dose rate; photon energy
3.  The neurobehavioral impact of manganese: Results and challenges obtained by a meta-analysis of individual participant data 
Neurotoxicology  2013;36:1-9.
Results from a meta-analysis of aggregated data provoked a new analysis using individual data on the neuropsychological performance of occupationally exposed workers.
Data from eight studies examining 579 exposed and 433 reference participants were included, 28 performance variables analyzed. The performance scores were adjusted for well-known individual-level covariates; the influence of possible, but unknown study-level covariates was attenuated by means of a z-normalization. Associations between performance and exposure were estimated by ANOVAs and ANCOVAs, the latter representing multi-level models.
Four cognitive and motor performance variables each indicated significantly lower performances of exposed individuals when confounding was considered; slowed motor performances and deficits in attention and short-term memory were found. Performance on a single test was significantly related to the biomarker manganese in blood. The outcomes on susceptibility were weak.
The slowing of responses was the most distinct feature of performances of exposed workers. It remains unclear, whether this result is related to the employed tests or provides important information about early stages of the neurotoxic impairment. More specific cognitive tests need to be employed to answer this question. The lack of dose–response relationships was related to features of the biomarker: it does not reflect the Mn in brain responsible for changes in performances.
PMCID: PMC4074005  PMID: 23419685
Systematic review; Neurobehavioral toxicology; Neuropsychological test; Neurobiological model; Biomarker
4.  MicroRNA-9 expression is a prognostic biomarker in patients with osteosarcoma 
The purpose of the present study was to examine the expression levels of microRNA-9 (miR-9) in osteosarcoma tissues and normal bone tissues, and investigate the relationships between miR-9 expression, clinicopathological features and the prognosis of patients with osteosarcoma.
The expression levels of miR-9 in osteosarcoma tissues and corresponding non-cancerous tissues were detected using a real-time quantitative assay. Differences in patient survival were determined using the Kaplan–Meier method and a log-rank test. A Cox proportional hazards regression analysis was used for univariate and multivariate analyses of prognostic values.
Compared to non-cancerous bone tissues, the expression levels of miR-9 in osteosarcoma tissues were significantly elevated (P < 0.001). We found that the expression level of miR-9 was significantly associated with tumor size (P = 0.011), clinical stage (P = 0.009) and distant metastasis (P < 0.001). The Kaplan–Meier curve showed that patients with low miR-9 expression survived significantly longer than patients with high miR-9 expression (P = 0.0017). Multivariate analysis suggested that miR-9 expression level (P = 0.002) is an independent prognostic factors for overall survival.
The findings of our study suggest that increased miR-9 expression has a strong correlation with the aggressive progression of osteosarcoma and its overexpression is a statistically significant risk factor affecting overall survival, suggesting that increased miR-9 expression could be a valuable marker of tumor progression and for prognosis of osteosarcoma.
PMCID: PMC4085232  PMID: 24969351
Osteosarcoma; MicroRNA-9; Prognosis
5.  DJ-1 contributes to adipogenesis and obesity-induced inflammation 
Scientific Reports  2014;4:4805.
Adipose tissue functions as an endocrine organ, and the development of systemic inflammation in adipose tissue is closely associated with metabolic diseases, such as obesity and insulin resistance. Accordingly, the fine regulation of the inflammatory response caused by obesity has therapeutic potential for the treatment of metabolic syndrome. In this study, we analyzed the role of DJ-1 (PARK7) in adipogenesis and inflammation related to obesity in vitro and in vivo. Many intracellular functions of DJ-1, including oxidative stress regulation, are known. However, the possibility of DJ-1 involvement in metabolic disease is largely unknown. Our results suggest that DJ-1 deficiency results in reduced adipogenesis and the down-regulation of pro-inflammatory cytokines in vitro. Furthermore, DJ-1-deficient mice show a low-level inflammatory response in the high-fat diet-induced obesity model. These results indicate previously unknown functions of DJ-1 in metabolism and therefore suggest that precise regulation of DJ-1 in adipose tissue might have a therapeutic advantage for metabolic disease treatment.
PMCID: PMC4055892  PMID: 24925581
6.  Characterization of the Xiamenmycin Biosynthesis Gene Cluster in Streptomyces xiamenensis 318 
PLoS ONE  2014;9(6):e99537.
Xiamenmycin (1) is a prenylated benzopyran derivative with anti-fibrotic activity. To investigate the genetic basis of xiamenmycin biosynthesis, we performed genome mining in the xiamenmycin-producing Streptomyces xiamenensis wild-type strain 318 to identify a candidate gene cluster. The complete gene cluster, consisting of five genes, was confirmed by a series of gene inactivations and heterologous expression. Based on bioinformatics analyses of each gene and feeding experiments, we found that the structure of an intermediate xiamenmycin B (3) accumulated in a ximA inactivation mutant, allowing us to propose a biosynthetic pathway. All five of the genes in the pathway were genetically and biochemically characterized. XimA was biochemically characterized as an ATP-dependent amide synthetase, catalyzing an amide bond formation in the presence of ATP as the final step in Xiamenmycin biosynthesis. The Km value of XimA was determined to be 474.38 µM for the substrate xiamenmycin B. These studies provide opportunities to use genetic and chemo-enzymatic methods to create new benzopyran derivatives as potential therapeutic agents.
PMCID: PMC4053376  PMID: 24919072
7.  Identification of an Inhibitory Mechanism of Luteolin on Insulin-like Growth Factor-1 Ligand-receptor Interaction 
Using single-molecule force measurement and single-molecule fluorescence imaging, we demonstrated that luteolin is able to perform an inhibitory effect on IGF-1 ligand-receptor binding, the initial step of IGF-1 signaling. Furthermore, this inhibition mechanism was also confirmed by flow cytometry and molecular docking. Our study demonstrates the novel molecular mechanism of luteolin as an IGF-1 signaling inhibitor in cancer therapy.
PMCID: PMC3940256  PMID: 23630137
Insulin-like growth factor 1 receptor; Luteolin; Single-molecule force measurement; Single-molecule fluorescence imaging
8.  Modification, Biological Evaluation and 3D QSAR Studies of Novel 2-(1,3-Diaryl- 4,5-Dihydro-1H-Pyrazol-5-yl)Phenol Derivatives as Inhibitors of B-Raf Kinase 
PLoS ONE  2014;9(5):e95702.
A series of novel 2-(1,3-diaryl- 4,5-dihydro-1H-pyrazol-5-yl)phenol derivatives (C1–C24) have been synthesized. The B-Raf inhibitory activity and anti-proliferation activity of these compounds have been tested. Compound C6 displayed the most potent biological activity against B-RafV600E (IC50 = 0.15 µM) and WM266.4 human melanoma cell line (GI50 = 1.75 µM), being comparable with the positive control (Vemurafenib and Erlotinib) and more potent than our previous best compounds. The docking simulation was performed to analyze the probable binding models and poses while the QSAR model was built to check the previous work as well as to introduce new directions. This work aimed at seeking more potent inhibitors as well as discussing some previous findings. As a result, the introduction of ortho-hydroxyl group on 4,5-dihydro-1H-pyrazole skeleton did reinforce the anti-tumor activity while enlarging the group on N-1 of pyrazoline was also helpful.
PMCID: PMC4020744  PMID: 24827980
9.  Extracellular Matrix Regulates UNC-6 (Netrin) Axon Guidance by Controlling the Direction of Intracellular UNC-40 (DCC) Outgrowth Activity 
PLoS ONE  2014;9(5):e97258.
How extracellular molecules influence the direction of axon guidance is poorly understood. The HSN axon of Caenorhabditis elegans is guided towards a ventral source of secreted UNC-6 (netrin). The axon’s outgrowth response to UNC-6 is mediated by the UNC-40 (DCC) receptor. We have proposed that in response to the UNC-6 molecule the direction of UNC-40-mediated axon outgrowth is stochastically determined. The direction of guidance is controlled by asymmetric cues, including the gradient of UNC-6, that regulate the probability that UNC-40-mediated axon outgrowth is directed on average, over time, in a specific direction. Here we provide genetic evidence that a specialized extracellular matrix, which lies ventral to the HSN cell body, regulates the probability that UNC-40-mediated axon outgrowth will be directed ventrally towards the matrix. We show that mutations that disrupt the function of proteins associated with this matrix, UNC-52 (perlecan), UNC-112 (kindlin), VAB-19 (Kank), and UNC-97 (PINCH), decrease the probability of UNC-40-mediated axon outgrowth in the ventral direction, while increasing the probability of outgrowth in the anterior and posterior directions. Other results suggest that INA-1 (α integrin) and MIG-15 (NIK kinase) signaling mediate the response in HSN. Although the AVM axon also migrates through this matrix, the mutations have little effect on the direction of AVM axon outgrowth, indicating that responses to the matrix are cell-specific. Together, these results suggest that an extracellular matrix can regulate the direction of UNC-6 guidance by increasing the probability that UNC-40-mediated axon outgrowth activity will be oriented in a specific direction.
PMCID: PMC4019552  PMID: 24824544
10.  Characterizations of Some Fuzzy Prefilters (Filters) in EQ-Algebras 
The Scientific World Journal  2014;2014:829527.
We introduce and study some types of fuzzy prefilters (filters) in EQ-algebras. First, we present several characterizations of fuzzy positive implicative prefilters (filters), fuzzy implicative prefilters (filters), and fuzzy fantastic prefilters (filters). Next, using their characterizations, we mainly consider the relationships among these special fuzzy filters. Particularly, we find some conditions under which a fuzzy implicative prefilter (filter) is equivalent to a fuzzy positive implicative prefilter (filter). As applications, we obtain some new results about classical filters in EQ-algebras and some related results about fuzzy filters in residuated lattices.
PMCID: PMC4032709  PMID: 24892096
11.  Radiographic Evaluation of Chronic Static Scapholunate Dissociation Post Soft Tissue Reconstruction 
Journal of Wrist Surgery  2013;2(2):155-159.
The purpose of this study was to examine the radiographic outcomes of dorsal intercarpal ligament capsulodesis (DILC), documenting the time to carpal collapse postoperatively. From January 2008 to January 2011, 12 patients were identified with chronic scapholunate (SL) dissociation. The average follow-up period was 15.8 months. Paired t-tests were used preoperatively, one month after pin removal, and at final follow-up to determine significance in radiographic outcomes. The Disabilities of the Arm, Shoulder, and Hand (DASH) survey was administered to patients before and after surgery to assess subjective levels of pain, function, and satisfaction. Intraoperatively all deformities were reduced completely. One month after pin removal, the mean SL gap was 3.3 mm, the SL angle was 74°, the radiolunate (RL) angle was 17°, and the lunatocapitate (LC) angle was 8°. Only the SL angle improved; the other measurements remained unchanged. At final follow-up, the mean SL gap was 3.6 mm, the SL angle was 78°, the RL angle was 20°, and the LC angle was 10°. SL angle worsened, but with no statistically significant difference. The other radiographic measurements remained unchanged at final follow up. Wrist flexion and extension decreased from 76% and 69% of the contralateral side to 62% and 56% of the contralateral side after surgery. Grip strength was 64% of the contralateral side before surgery and 83% after surgery. Visual Analog Scale (VAS) results improved from 6.3 to 1.7, and DASH scores improved from 39 to 8 after the surgery. DILC cannot withstand large and repetitive forces. Carpal collapse recurred within a short time after DILC. However, our small patient numbers and short term follow-up preclude any conclusions with respect to clinical efficacy of this procedure. Limitations of this study include the fact that this is a retrospective study with no control group. In addition, it represents a single-surgeon series, which introduces a source of bias and carries the risk of technical and methodological flaws, which may have contributed to the observed radiographic outcomes.
PMCID: PMC3699270  PMID: 24436809
chronic scapholunate dissociation; reconstruction of scapholunate ligament; radiographic evaluation scapholunate reconstruction
12.  Neutralizing antibody responses to enterovirus and adenovirus in healthy adults in China 
Hand, foot and mouth disease (HFMD) is an important public health problem that has emerged over the past several years. HFMD predominantly infects children under seven years old and occasionally causes severe disease in adults. Among the enteroviruses, enterovirus 71 (EV71) and coxsackievirus 16 (CA16) are the major causative agents of HFMD. In addition, adenovirus cocirculates with enterovirus and has become a possible additional pathogenic factor for HFMD in some cases. Here, we have investigated the neutralizing antibody responses to both enterovirus and adenovirus in adults, with the aim of exploring the prevalence trends of these viruses and the nature of protective immunity in humans to these viral infections. Sera from 391 healthy adults from 21 provinces and cities in China were tested for the presence of antibodies against EV71, CA16, adenovirus human serotype 5 (AdHu5) and chimpanzee adenovirus pan7 (AdC7) using neutralization tests. High seroprevalence rates of EV71, CA16 and AdHu5 were found in the population (85.7%, 58.8% and 74.2%, respectively). The coseropositivity rate of these three viruses was 39.4% (154 of 391), with median neutralizing antibody titers of 80, 40 and 640, respectively, and the neutralizing antibody titer for EV71 was found to be correlated with those of CA16 and AdHu5. AdC7 was found to be a rare adenovirus serotype in the human population, with a seropositivity rate of 11.8%, suggesting that it could be a good choice for a vaccine carrier that could be used in vaccine development.
PMCID: PMC4051363
AdC7; AdHu5; CA16; EV71; hand, foot and mouth disease; seroprevalence; neutralizing antibody
13.  Combined probiotic bacteria promotes intestinal epithelial barrier function in interleukin-10-gene-deficient mice 
AIM: To investigate the protective effects of combinations of probiotic (Bifico) on interleukin (IL)-10-gene-deficient (IL-10 KO) mice and Caco-2 cell monolayers.
METHODS: IL-10 KO mice were used to assess the benefits of Bifico in vivo. IL-10 KO and control mice received approximately 1.5 × 108 cfu/d of Bifico for 4 wk. Colons were then removed and analyzed for epithelial barrier function by Ussing Chamber, while an ELISA was used to evaluate proinflammatory cytokines. The colon epithelial cell line, Caco-2, was used to test the benefit of Bifico in vitro. Enteroinvasive Escherichia coli (EIEC) and the probiotic mixture Bifico, or single probiotic strains, were applied to cultured Caco-2 monolayers. Barrier function was determined by measuring transepithelial electrical resistance and tight junction protein expression.
RESULTS: Treatment of IL-10 KO mice with Bifico partially restored body weight, colon length, and epithelial barrier integrity to wild-type levels. In addition, IL-10 KO mice receiving Bifico treatment had reduced mucosal secretion of tumor necrosis factor-α and interferon-γ, and attenuated colonic disease. Moreover, treatment of Caco-2 monolayers with Bifico or single-strain probiotics in vitro inhibited EIEC invasion and reduced the secretion of proinflammatory cytokines.
CONCLUSION: Bifico reduced colon inflammation in IL-10 KO mice, and promoted and improved epithelial-barrier function, enhanced resistance to EIEC invasion, and decreased proinflammatory cytokine secretion.
PMCID: PMC4000500  PMID: 24782616
Probiotic bacteria; Intestinal barrier function; Tight junction proteins; Interleukin-10 gene-deficient mice; Caco-2 monolayers
14.  Targeting gallbladder cancer: oncolytic virotherapy with myxoma virus is enhanced by rapamycin in vitro and further improved by hyaluronan in vivo 
Molecular Cancer  2014;13:82.
Gallbladder carcinoma (GBC) is highly lethal, and effective treatment will require synergistic anti-tumor management. The study is aimed at investigating the oncolytic value of myxoma virus (MYXV) infection against GBC and optimizing MYXV oncolytic efficiency.
We examined the permissiveness of GBC cell lines to MYXV infection and compared the effects of MYXV on cell viability among GBC and control permissive glioma cells in vitro and in vivo after MYXV + rapamycin (Rap) treatment, which is known to enhance cell permissiveness to MYXV by upregulating p-Akt levels. We also assessed MYXV + hyaluronan (HA) therapy efficiency by examinating Akt activation status, MMP-9 expression, cell viability, and collagen distribution. We further compared hydraulic conductivity, tumor area, and survival of tumor-bearing mice between the MYXV + Rap and MYXV + HA therapeutic regimens.
MYXV + Rap treatment could considerably increase the oncolytic ability of MYXV against GBC cell lines in vitro but not against GBC xenografts in vivo. We found higher levels of collagen IV in GBC tumors than in glioma tumors. Diffusion analysis demonstrated that collagen IV could physically hinder MYXV intratumoral distribution. HA–CD44 interplay was found to activate the Akt signaling pathway, which increases oncolytic rates. HA was also found to enhance the MMP-9 secretion, which contributes to collagen IV degradation.
Unlike MYXV + Rap, MYXV + HA therapy significantly enhanced the anti-tumor effects of MYXV in vivo and prolonged survival of GBC tumor-bearing mice. HA may optimize the oncolytic effects of MYXV on GBC via the HA–CD44 interaction which can promote viral infection and diffusion.
PMCID: PMC4021541  PMID: 24725816
Gallbladder cancer; Myxoma virus; Oncolytic virotherapy; Collagen IV
15.  Prediction of antiviral efficacy in patients with chronic hepatitis C by changes in forkhead box protein 3 levels 
The aim of the present study was to investigate the distribution of CD4+CD25+ regulatory T cells (Tregs) in the peripheral blood of patients with chronic hepatitis C; in addition to identifying whether the distribution of CD4+CD25+ Tregs predicts the efficacy of antiviral therapy for HCV. The expression of CD4+CD25+ forkhead box protein (FOXP) 3+ Tregs within a CD4+ T cell population was detected in the peripheral blood obtained from patients with chronic hepatitis C and from healthy control subjects using flow cytometry. The hepatitis C virus (HCV)-RNA load was measured using quantitative-fluorescence polymerase chain reaction. CD4+CD25+FOXP3+ Tregs accounted for 14.24±1.33% of the CD4+ T cells in the peripheral blood of patients with chronic hepatitis C, which was higher than that of the healthy control subjects (5.62±1.21%; P<0.001). Furthermore, the frequency of CD4+CD25+FOXP3+ Tregs in CD4+ T cells of the peripheral blood positively correlated with the HCV-RNA load (r=0.73; P=0.032). Therefore, the results of the present study indicated that the expression of CD4+CD25+FOXP3+ Tregs increased in patients that were chronically infected with HCV and positively correlated with the HCV-RNA load.
PMCID: PMC4061195  PMID: 24944616
chronic hepatitis C; CD4+CD25+ forkhead box P3+ regulatory T cells; hepatitis C virus-RNA load
16.  Colon Cancer-Specific Antigen-2 May Be Used as a Detecting and Prognostic Marker in Colorectal Cancer: A Preliminary Observation 
PLoS ONE  2014;9(4):e94252.
A specific and sensitive serum marker for colorectal cancer (CRC) detection and surveillance is central to effective treatment. It was preliminarily reported that some nuclear matrix proteins may be served as a specific blood based marker for colon cancer. The objective of this study is to evaluate the value of serum CCSA-2 detection in diagnosis, prognostic estimation and surveillance for CRC.
Serum CCSA-2 protein was measured in 181 various patient populations and 20 healthy donors before surgery. For 106 CRC patients, it was also measured on day 7 after surgery. Among them, 49 CRC patients' CCSA-2 protein were measured during the follow-up period according to NCCN Guideline.
The serum CCSA-2 concentration in CRC patients was significantly higher than which in other patients and healthy individuals. Serum CCSA-2, at the cut-off point of 64.10 ng/mL, had a sensitivity of 98.10% and a specificity of 97.90% in separating CRC populations from all other individuals. The CCSA-2 assay was significantly more sensitive than CEA and CA19-9 assay in CRC detection. After surgery, the serum CCSA-2 level of CRC patients declined significantly, but it rebounded to a high level when recurrences occurred. The pre-operative serum CCSA-2 level in patients who had a relapse within the follow-up period was significantly higher than which in patients without relapse.
Serum CCSA-2 not only may be a potential biomarker using in screening and surveillance of CRC, but also may be an independent prognostic marker for CRC patients. Further clinical trials need to be performed in a larger population of patients to ulteriorly confirm these results.
PMCID: PMC3978011  PMID: 24710115
17.  Pollen sensitivity to ultraviolet-B (UV-B) suggests floral structure evolution in alpine plants 
Scientific Reports  2014;4:4520.
Various biotic and abiotic factors are known to exert selection pressures on floral traits, but the influence of ultraviolet-B (UV-B) light on the evolution of flower structure remains relatively unexplored. We have examined the effectiveness of flower structure in blocking radiation and the effects of UV-B on pollen viability in 42 species of alpine plants in the Hengduan Mountains, China. Floral forms were categorized as either protecting or exposing pollen grains to UV-B. The floral materials of plants with exposed and protected pollen grains were able to block UV-B at similar levels. Exposure to UV-B radiation in vitro resulted in a significantly greater loss of viability in pollen from plant species with protective floral structures. The pronounced sensitivity of protected pollen to UV-B radiation was associated with the type of flower structure. These findings demonstrate that UV-B plays an important role in the evolution of protective floral forms in alpine plants.
PMCID: PMC3970119  PMID: 24682234
18.  Expression analysis of genes and pathways associated with liver metastases of the uveal melanoma 
BMC Medical Genetics  2014;15:29.
Uveal melanoma is an aggressive cancer which has a high percentage metastasizing to the liver, with a worse prognosis. Identification of patients at high risk of metastases may provide information for early detection of metastases and treatment.
Expression profiling of ocular tumor tissues from 46 liver metastatic uveal melanoma samples and 45 non-metastatic uveal melanoma samples were got from GEO database. Bioinformatic analyses such as the Gene Oncology and Kyoto Encyclopedia of Genes and Genomes were used to identify genes and pathways specifically associated with liver metastases of the uveal melanoma.
A total of 1138 probes were differentially expressed in two group samples. All differential gene interactions in the Signal-Net were analyzed. Of them, 768 probes were up-regulated and 370 down-regulated. They mainly participated in 125 GO terms and 16 pathways. Of the genes differentially expressed between two group cancers, HTR2B, CHL1, the ZNF family, YWHAZ and FYN were the most significantly altered.
Bioinformatics may help excavate and analyze large amounts of data in microarrays by means of rigorous experimental planning, scientific statistical analysis and collection of complete data about liver metastases of uveal melanoma patients. In the present study, a novel differential gene expression pattern was constructed and advanced study will provide new targets for diagnosis and mechanism of uveal melanoma liver metastases.
PMCID: PMC4015751  PMID: 24597767
Uveal melanoma; Liver metastases; Gene expression; GO analysis; Pathway analysis
19.  Negative Regulation of RIG-I-Mediated Innate Antiviral Signaling by SEC14L1 
Journal of Virology  2013;87(18):10037-10046.
Retinoic acid-inducible gene I (RIG-I) is a key sensor for recognizing nucleic acids derived from RNA viruses and triggers beta interferon (IFN-β) production. Because of its important role in antiviral innate immunity, the activity of RIG-I must be tightly controlled. Here, we used yeast two-hybrid screening to identify a SEC14 family member, SEC14L1, as a RIG-I-associated negative regulator. Transfected SEC14L1 interacted with RIG-I, and endogenous SEC14L1 associated with RIG-I in a viral infection-inducible manner. Overexpression of SEC14L1 inhibited transcriptional activity of the IFN-β promoter induced by RIG-I but not TANK-binding kinase 1 (TBK1) and interferon regulatory factor 3 (IRF3). Knockdown of endogenous SEC14L1 in both HEK293T cells and HT1080 cells potentiated RIG-I and Sendai virus-triggered IFN-β production as well as attenuated the replication of Newcastle disease virus. SEC14L1 interacted with the N-terminal domain of RIG-I (RIG-I caspase activation and recruitment domain [RIG-I-CARD]) and competed with VISA/MAVS/IPS-1/Cardif for RIG-I-CARD binding. Domain mapping further indicated that the PRELI-MSF1 and CRAL-TRIO domains but not the GOLD domain of SEC14L1 are required for interaction and inhibitory function. These findings suggest that SEC14L1 functions as a novel negative regulator of RIG-I-mediated antiviral signaling by preventing RIG-I interaction with the downstream effector.
PMCID: PMC3754010  PMID: 23843640
20.  Effective Multifocus Image Fusion Based on HVS and BP Neural Network 
The Scientific World Journal  2014;2014:281073.
The aim of multifocus image fusion is to fuse the images taken from the same scene with different focuses to obtain a resultant image with all objects in focus. In this paper, a novel multifocus image fusion method based on human visual system (HVS) and back propagation (BP) neural network is presented. Three features which reflect the clarity of a pixel are firstly extracted and used to train a BP neural network to determine which pixel is clearer. The clearer pixels are then used to construct the initial fused image. Thirdly, the focused regions are detected by measuring the similarity between the source images and the initial fused image followed by morphological opening and closing operations. Finally, the final fused image is obtained by a fusion rule for those focused regions. Experimental results show that the proposed method can provide better performance and outperform several existing popular fusion methods in terms of both objective and subjective evaluations.
PMCID: PMC3933522  PMID: 24683327
21.  MicroRNA-362 induces cell proliferation and apoptosis resistance in gastric cancer by activation of NF-κB signaling 
According to cancer-related microRNA (miRNA) expression microarray research available in public databases, miR-362 expression is elevated in gastric cancer. However, the expression and biological role of miR-362 in gastric progression remain unclear.
miR-362 expression levels in gastric cancer tissues and cell lines were determined using real-time PCR. The roles of miR-362, in promoting gastric cancer cell proliferation and apoptosis resistance, were assessed by different biological assays, such as colony assay, flow cytometry and TUNEL assay. The effect of miR-362 on NF-κB activation was investigated using the luciferase reporter assay, fluorescent immunostaining.
MiR-362 overexpression induced cell proliferation, colony formation, and resistance to cisplatin-induced apoptosis in BGC-823 and SGC-7901 gastric cancer cells. MiR-362 increased NF-κB activity and relative mRNA expression of NF-κB–regulated genes, and induced nuclear translocation of p65. Expression of the tumor suppressor CYLD was inhibited by miR-362 in gastric cancer cells; miR-362 levels were inversely correlated with CYLD expression in gastric cancer tissue. MiR-362 downregulated CYLD expression by binding its 3′ untranslated region. NF-κB activation was mechanistically associated with siRNA-mediated downregulation of CYLD. MiR-362 inhibitor reversed all the effects of miR-362.
The results suggest that miR-362 plays an important role in repressing the tumor suppressor CYLD and present a novel mechanism of miRNA-mediated NF-κB activation in gastric cancer.
PMCID: PMC3916099  PMID: 24495516
miR-362; NF-κB; CYLD; Gastric cancer; Proliferation; Apoptosis
22.  Activated CD8 T cells acquire NK1.1 expression and preferentially locate in the liver in mice after allogeneic hematopoietic cell transplantation 
Immunology letters  2013;150(1-2):75-78.
Immune cells expressing both NK and T cell markers include CD1d-dependent NKT cells and - independent NKT-like cells. We now describe the presence of NK1.1+CD8+ T cells in the liver, but not other tissues (spleen, bone marrow, thymus or peripheral blood) in mice receiving allogeneic hematopoietic cell transplantation (allo-HCT). These cells are CD1d-independent TCRαβ+ T cells with an effector/memory CD44hiCD62L− phenotype, and do not express Ly49 receptors. Furthermore, these cells were derived from donor splenocytes, but not bone marrow cells. Depletion of CD8+, but not NK1.1+, cells from donor splenocytes prior to transplantation prevented the generation of NK1.1+CD8+ T cells, indicating that these cells arose from donor NK1.1−CD8+ splenic T cells. These results provide direct evidence that donor CD8+ T cells can acquire NK1.1 expression upon activation in allo-HCT recipients and that these NK1.1+CD8+ NKT-like cells maintain an effector/memory phenotype and persist in the recipients with preferential localization in the liver.
PMCID: PMC3602380  PMID: 23291124
CD8 T cells; NK1.1+CD8+ T cells; allogeneic hematopoietic cell transplantation; liver
23.  The Effects of Tenascin C Knockdown on Trabecular Meshwork Outflow Resistance 
Tenascin C (TNC) is a matricellular glycoprotein whose expression in adult tissue is indicative of tissue remodeling. The purpose of the current study was to determine the localization of TNC in trabecular meshwork (TM) tissue and to analyze the effects of TNC on intraocular pressure (IOP).
Human TM frontal sections were immunostained with anti-TNC and imaged by confocal microscopy. TNC mRNA and protein levels were quantitated in anterior segments perfused at physiological and elevated pressure. Short, hairpin RNA (shRNA) silencing lentivirus targeting full-length TNC (shTNC) was applied to anterior segment perfusion organ cultures. The IOPs and central corneal thickness (CCT) of wild-type, TNC−/−, and tenascin X (TNX−/−) knockout mice were measured.
TNC was distributed in the juxtacanalicular (JCT) region of adult human TM, predominantly in the basement membrane underlying the inner wall of Schlemm's canal. Application of shTNC lentivirus to human and porcine anterior segments in perfusion culture did not significantly affect outflow rate. Although TNC was upregulated in response to pressure, there was no difference in outflow rate when shTNC-silenced anterior segments were subjected to elevated pressure. Furthermore, IOPs and CCTs were not significantly different between TNC−/− or TNX−/− and wild-type mice.
TNC does not appear to contribute directly to outflow resistance. However, TNC immunolocalization in the JCT of adult human eyes suggests that certain areas of the TM are being continuously remodeled with or without an IOP increase.
Tenascin C does not appear to contribute directly to outflow resistance. However, its high expression levels and specific localization in the juxtacanalicular region of adult trabecular meshwork suggest that certain areas of this tissue are being continuously remodeled.
PMCID: PMC3747717  PMID: 23882691
tenascin C; trabecular meshwork; outflow resistance; extracellular matrix
24.  Involvement of Multiple Types of Dehydrins in the Freezing Response in Loquat (Eriobotrya japonica) 
PLoS ONE  2014;9(1):e87575.
Dehydrins (DHNs) are a family of plant proteins typically induced in response to stress conditions that cause cellular dehydration, such as low temperatures, high salinity, and drought. Loquat (Eriobotrya japonica) is a perennial fruit crop that blossoms during winter. Loquat fruitlets are frequently injured by freezing. To evaluate the role of the EjDHNs in freezing resistance in loquat fruitlets, two cultivars of loquat, the freezing-sensitive ‘Ninghaibai’ (FS-NHB) and the freezing-tolerant ‘Jiajiao’ (FT-JJ), were analyzed under induced freezing stress. Freezing stress led to obvious accumulation of reactive oxygen species and considerable lipid peroxidation in membranes during the treatment period. Both these phenomena were more pronounced in ‘FS-NHB’ than in ‘FS-JJ.’ Immunogold labeling of dehydrin protein was performed. DHN proteins were found to be concentrated mainly in the vicinity of the plasma membrane, and the density of the immunogold labeling was significantly higher after freezing treatment, especially in the more freezing-tolerant cultivar ‘FT-JJ.’ Seven DHNs, showing four different structure types, were obtained from loquat fruitlets and used to study the characteristics of different EjDHN proteins. These DHN proteins are all highly hydrophilic, but they differ significantly in size, ranging from 188 to 475 amino acids, and in biochemical properties, such as theoretical pI, aliphatic index, and instability index. Freezing treatment resulted in up-regulation of the expression levels of all seven EjDHNs, regardless of structure type. The accumulation of the transcripts of these EjDHN genes was much more pronounced in ‘FT-JJ’ than in ‘FS-NHB.’ Altogether, this study provides evidence that EjDHNs are involved in the cryoprotection of the plasma membrane during freeze-induced dehydration in loquat fruitlets.
PMCID: PMC3909202  PMID: 24498141
25.  Specific Expression of DR5 Promoter in Rice Roots Using a tCUP Derived Promoter-Reporter System 
PLoS ONE  2014;9(1):e87008.
Variation of transgene expression caused by either position effect at the insertion site or the promoter/enhancer elements employed for the expression of selectable marker genes has complicated phenotype characterization and caused misinterpretation. We have developed a reporter system in rice to analyze the influence of vector configuration, spacer and selectable marker gene promoter on the expression of the promoterless GUS reporter and DR5 promoter. Our results indicate that a spacer inserted between the reversed 35S promoter and the GUS reporter could reduce leaky expression of the reporter but was unable to block the nonspecific expression of DR5::GUS. Stacking the selectable marker unit in head to tail with the GUS reporter aided the gene specific expression of the GUS reporter under the DR5 promoter even when the 35S promoter is used for expression of the selectable marker. Compared to 35S under this configuration, a quick and distinctive expression of DR5::GUS was observed in the root cap, quiescent center and xylem cells in the root apical meristem by using the tCUP derived promoter (tCUP1) for selection, that is similar to the pattern obtained by a sensitive DR5 variant (DR5rev) in Arabidopsis. These data suggest a conserved property of the tCUP promoter in preventing enhancer-promoter interactions in rice as it does in Arabidopsis, and also demonstrate that an analogous distal auxin maximum exists in roots of rice. Therefore, the tCUP promoter based selection system provides a new strategy for specific expression of transgenes in rice.
PMCID: PMC3899362  PMID: 24466314

Results 1-25 (171)