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1.  Development and Characterization of Lecithin-based Self-assembling Mixed Polymeric Micellar (saMPMs) Drug Delivery Systems for Curcumin 
Scientific Reports  2016;6:37122.
Self-assembling mixed polymeric micelles (saMPMs) were developed for overcoming major obstacles of poor bioavailability (BA) associated with curcumin delivery. Lecithin added was functioned to enlarge the hydrophobic core of MPMs providing greater solubilization capacity. Amphiphilic polymers (sodium deoxycholate [NaDOC], TPGS, CREMOPHOR, or a PLURONIC series) were examined for potentially self-assembling to form MPMs (saMPMs) with the addition of lecithin. Particle size, size distribution, encapsulation efficacy (E.E.), and drug loading (D.L.) of the mixed micelles were optimally studied for their influences on the physical stability and release of encapsulated drugs. Overall, curcumin:lecithin:NaDOC and curcumin:lecithin:PLURONIC P123 in ratios of 2:1:5 and 5:2:20, respectively, were optimally obtained with a particle size of < 200 nm, an E.E. of >80%, and a D.L. of >10%. The formulated system efficiently stabilized curcumin in phosphate-buffered saline (PBS) at room temperature or 4 °C and in fetal bovine serum or PBS at 37 °C and delayed the in vitro curcumin release. In vivo results further demonstrated that the slow release of curcumin from micelles and prolonged duration increased the curcumin BA followed oral and intravenous administrations in rats. Thus, lecithin-based saMPMs represent an effective curcumin delivery system, and enhancing BA of curcumin can enable its wide applications for treating human disorders.
PMCID: PMC5111064  PMID: 27848996
2.  1α,25-dihydroxyvitamin D3 acts via transforming growth factor-β to up-regulate expression of immunosuppressive CD73 on human CD4+ Foxp3– T cells 
Immunology  2015;146(3):423-431.
Vitamin D deficiency is associated with increased incidence and severity of various immune-mediated diseases. Active vitamin D (1α,25-dihydroxyvitamin D3; 1,25(OH)2D3) up-regulates CD4+ T-cell expression of the purine ectonucleotidase CD39, a molecule that is associated with the generation of anti-inflammatory adenosine. Here we aimed to investigate the direct impact of 1,25(OH)2D3 on expression of the downstream ecto-5′-nucleotidase CD73 by human CD4 T cells, and components of the transforming growth factor-β (TGF-β) pathway, which have been implicated in the modulation of CD73 by murine T cells. At 10−8 to 10−7 m, 1,25(OH)2D3 significantly increased expression of CD73 on peripheral human CD4+ T cells. Although 1,25(OH)2D3 did not affect the mRNA expression of latent TGF-β1, 1,25(OH)2D3 did up-regulate expression of TGF-β-associated molecules [latency-associated peptide (LAP), glycophorin A repetitions predominant (GARP), GP96, neuropilin-1, thrombospondin-1 and αv integrin] which is likely to have contributed to the observed enhancement in TGF-β bioactivity. CD73 was highly co-expressed with LAP and GARP following 1,25(OH)2D3 treatment, but unexpectedly, each of these cell surface molecules was expressed primarily on CD4+ Foxp3– T cells, rather than CD4+ Foxp3+ T cells. Notably, neutralization of TGF-β significantly impaired 1,25(OH)2D3-mediated induction of CD73. Collectively, we show that 1,25(OH)2D3 enhances expression of CD73 on CD4+ Foxp3– T cells in a process that is at least partially TGF-β-dependent. These data reveal an additional contributing mechanism by which vitamin D may be protective in immune-mediated disease.
PMCID: PMC4610631  PMID: 26251265
1α,25-dihydroxyvitamin D3; CD73; Foxp3; transforming growth factor-β
3.  A Novel Nuclear Function for the Interleukin-17 Signaling Adaptor Protein Act1 
PLoS ONE  2016;11(10):e0163323.
In the context of the human airway, interleukin-17A (IL-17A) signaling is associated with severe inflammation, as well as protection against pathogenic infection, particularly at mucosal surfaces such as the airway. The intracellular molecule Act1 has been demonstrated to be an essential mediator of IL-17A signaling. In the cytoplasm, it serves as an adaptor protein, binding to both the intracellular domain of the IL-17 receptor as well as members of the canonical nuclear factor kappa B (NF-κB) pathway. It also has enzymatic activity, and serves as an E3 ubiquitin ligase. In the context of airway epithelial cells, we demonstrate for the first time that Act1 is also present in the nucleus, especially after IL-17A stimulation. Ectopic Act1 expression can also increase the nuclear localization of Act1. Act1 can up-regulate the expression and promoter activity of a subset of IL-17A target genes in the absence of IL-17A signaling in a manner that is dependent on its N- and C-terminal domains, but is NF-κB independent. Finally, we show that nuclear Act1 can bind to both distal and proximal promoter regions of DEFB4, one of the IL-17A responsive genes. This transcriptional regulatory activity represents a novel function for Act1. Taken together, this is the first report to describe a non-adaptor function of Act1 by directly binding to the promoter region of IL-17A responsive genes and directly regulate their transcription.
PMCID: PMC5056742  PMID: 27723765
4.  Cisplatin-induced regulation of signal transduction pathways and transcription factors in p53-mutated subclone variants of hepatoma cells: Potential application for therapeutic targeting 
Oncology Letters  2016;12(5):3723-3730.
Cisplatin is commonly recognized as a DNA-damaging drug; however, its versatile antitumor effects have been demonstrated to extend beyond this narrow functional attribute. The present study determined how cisplatin regulates alternative pathways and transcription factors to exert its additional antitumor actions. Cisplatin was observed to be able to trigger an endoplasmic reticulum stress response through aggravated nitrosative stress coupled to perturbed mitochondrial calcium (Ca2+) homeostasis, which substantially downregulated glucose-regulated protein (GRP) 78 expression by suppressing the cleavage of activating transcription factor (ATF) 6α (90 kDa) to its active 50 kDa subunit. Concomitantly, the ATF4-ATF3-C/emopamil binding protein homologous protein axis was activated by cisplatin, which triggered cellular glutathione (GSH) depletion by strongly inhibiting γ-glutamylcysteine synthetase heavy chain (γ-GCSh), a key enzyme in GSH biosynthesis. The present study also demonstrated that cisplatin substantially inhibited β-catenin, causing a marked downregulation of survivin and B-cell lymphoma (Bcl)-2. Taken together, the present results uncovered a novel mechanism of cisplatin that could simultaneously trigger the inhibition of three prominent antiapoptotic effector molecules (Bcl-2, survivin and GRP78) and effectively promote GSH depletion by inhibiting γ-GCSh. These newly discovered functional attributes of cisplatin can provide an avenue for novel combined therapeutic strategies to kill hepatocellular carcinoma cells effectively.
PMCID: PMC5104157  PMID: 27895723
cisplatin; GSH depletion; ER stress; GRP78; survivin; Bcl-2
5.  Anaerobic choline metabolism in microcompartments promotes growth and swarming of P roteus mirabilis  
Environmental Microbiology  2015;18(9):2886-2898.
Gammaproteobacteria are important gut microbes but only persist at low levels in the healthy gut. The ecology of G ammaproteobacteria in the gut environment is poorly understood. Here, we demonstrate that choline is an important growth substrate for representatives of G ammaproteobacteria. Using P roteus mirabilis as a model, we investigate the role of choline metabolism and demonstrate that the cut C gene, encoding a choline‐trimethylamine lyase, is essential for choline degradation to trimethylamine by targeted mutagenesis of cut C and subsequent complementation experiments. P roteus mirabilis can rapidly utilize choline to enhance growth rate and cell yield in broth culture. Importantly, choline also enhances swarming‐associated colony expansion of P . mirabilis under anaerobic conditions on a solid surface. Comparative transcriptomics demonstrated that choline not only induces choline‐trimethylamine lyase but also genes encoding shell proteins for the formation of bacterial microcompartments. Subsequent analyses by transmission electron microscopy confirmed the presence of such novel microcompartments in cells cultivated in liquid broth and hyper‐flagellated swarmer cells from solid medium. Together, our study reveals choline metabolism as an adaptation strategy for P . mirabilis and contributes to better understand the ecology of this bacterium in health and disease.
PMCID: PMC5026066  PMID: 26404097
6.  1α,25‐dihydroxyvitamin D3 acts via transforming growth factor‐β to up‐regulate expression of immunosuppressive CD73 on human CD4+ Foxp3– T cells 
Immunology  2015;146(3):423-431.
Vitamin D deficiency is associated with increased incidence and severity of various immune‐mediated diseases. Active vitamin D (1α,25‐dihydroxyvitamin D3; 1,25(OH)2D3) up‐regulates CD4+ T‐cell expression of the purine ectonucleotidase CD39, a molecule that is associated with the generation of anti‐inflammatory adenosine. Here we aimed to investigate the direct impact of 1,25(OH)2D3 on expression of the downstream ecto‐5′‐nucleotidase CD73 by human CD4 T cells, and components of the transforming growth factor‐β (TGF‐β) pathway, which have been implicated in the modulation of CD73 by murine T cells. At 10−8 to 10−7 m, 1,25(OH)2D3 significantly increased expression of CD73 on peripheral human CD4+ T cells. Although 1,25(OH)2D3 did not affect the mRNA expression of latent TGF‐β 1, 1,25(OH)2D3 did up‐regulate expression of TGF‐β‐associated molecules [latency‐associated peptide (LAP), glycophorin A repetitions predominant (GARP), GP96, neuropilin‐1, thrombospondin‐1 and αv integrin] which is likely to have contributed to the observed enhancement in TGF‐β bioactivity. CD73 was highly co‐expressed with LAP and GARP following 1,25(OH)2D3 treatment, but unexpectedly, each of these cell surface molecules was expressed primarily on CD4+ Foxp3– T cells, rather than CD4+ Foxp3+ T cells. Notably, neutralization of TGF‐β significantly impaired 1,25(OH)2D3‐mediated induction of CD73. Collectively, we show that 1,25(OH)2D3 enhances expression of CD73 on CD4+ Foxp3– T cells in a process that is at least partially TGF‐β‐dependent. These data reveal an additional contributing mechanism by which vitamin D may be protective in immune‐mediated disease.
PMCID: PMC4610631  PMID: 26251265
1α,25‐dihydroxyvitamin D3; CD73; Foxp3; transforming growth factor‐β
7.  Bio- chemical and physical characterizations of mesenchymal stromal cells along the time course of directed differentiation 
Scientific Reports  2016;6:31547.
Cellular biophysical properties are novel biomarkers of cell phenotypes which may reflect the status of differentiating stem cells. Accurate characterizations of cellular biophysical properties, in conjunction with the corresponding biochemical properties could help to distinguish stem cells from primary cells, cancer cells, and differentiated cells. However, the correlated evolution of these properties in the course of directed stem cells differentiation has not been well characterized. In this study, we applied video particle tracking microrheology (VPTM) to measure intracellular viscoelasticity of differentiating human mesenchymal stromal/stem cells (hMSCs). Our results showed that osteogenesis not only increased both elastic and viscous moduli, but also converted the intracellular viscoelasticity of differentiating hMSCs from viscous-like to elastic-like. In contrast, adipogenesis decreased both elastic and viscous moduli while hMSCs remained viscous-like during the differentiation. In conjunction with bio- chemical and physical parameters, such as gene expression profiles, cell morphology, and cytoskeleton arrangement, we demonstrated that VPTM is a unique approach to quantify, with high data throughput, the maturation level of differentiating hMSCs and to anticipate their fate decisions. This approach is well suited for time-lapsed study of the mechanobiology of differentiating stem cells especially in three dimensional physico-chemical biomimetic environments including porous scaffolds.
PMCID: PMC4985743  PMID: 27526936
8.  Quantitative Determination of Flexible Pharmacological Mechanisms Based On Topological Variation in Mice Anti-Ischemic Modular Networks 
PLoS ONE  2016;11(7):e0158379.
Targeting modules or signalings may open a new path to understanding the complex pharmacological mechanisms of reversing disease processes. However, determining how to quantify the structural alteration of these signalings or modules in pharmacological networks poses a great challenge towards realizing rational drug use in clinical medicine. Here, we explore a novel approach for dynamic comparative and quantitative analysis of the topological structural variation of modules in molecular networks, proposing the concept of allosteric modules (AMs). Based on the ischemic brain of mice, we optimize module distribution in different compound-dependent modular networks by using the minimum entropy criterion and then calculate the variation in similarity values of AMs under various conditions using a novel method of SimiNEF. The diverse pharmacological dynamic stereo-scrolls of AMs with functional gradient alteration, which consist of five types of AMs, may robustly deconstruct modular networks under the same ischemic conditions. The concept of AMs can not only integrate the responsive mechanisms of different compounds based on topological cascading variation but also obtain valuable structural information about disease and pharmacological networks beyond pathway analysis. We thereby provide a new systemic quantitative strategy for rationally determining pharmacological mechanisms of altered modular networks based on topological variation.
PMCID: PMC4934924  PMID: 27383195
9.  Induction of potent NK cell-dependent anti-myeloma cytotoxic T cells in response to combined mapatumumab and bortezomib 
Oncoimmunology  2015;4(9):e1038011.
There is increasing evidence that some cancer therapies can promote tumor immunogenicity to boost the endogenous antitumor immune response. In this study, we used the novel combination of agonistic anti-TRAIL-R1 antibody (mapatumumab, Mapa) with low dose bortezomib (LDB) for this purpose. The combination induced profound myeloma cell apoptosis, greatly enhanced the uptake of myeloma cell apoptotic bodies by dendritic cell (DC) and induced anti-myeloma cytotoxicity by both CD8+ T cells and NK cells. Cytotoxic lymphocyte expansion was detected within 24 h of commencing therapy and was maximized when myeloma-pulsed DC were co-treated with low dose bortezomib and mapatumumab (LDB+Mapa) in the presence of NK cells. This study shows that Mapa has two distinct but connected modes of action against multiple myeloma (MM). First, when combined with LDB, Mapa produced powerful myeloma cell apoptosis; secondly, it promoted DC priming and an NK cell-mediated expansion of anti-myeloma cytotoxic lymphocyte (CTL). Overall, this study indicates that Mapa can be used to drive potent anti-MM immune responses.
PMCID: PMC4570118  PMID: 26405606
dendritic cell; multiple myeloma; natural killer cell; proteasome inhibitor; T cell; tumor necrosis factor apoptosis inducing ligand and receptor
10.  LncRNA MALAT1 exerts oncogenic functions in lung adenocarcinoma by targeting miR-204 
American Journal of Cancer Research  2016;6(5):1099-1107.
Accumulating evidence indicates that the lncRNAs play a critical role in cancer progression and metastasis. In this study, we found that MALAT1 upregulation was associated with larger tumor size and lymph-node metastasis, and also correlated with shorter overall survival of lung adenocarcinoma patients. Furthermore, MALAT1 promotes EMT and metastasis of lung adenocarcinoma cells in vitro and in vivo. In particular, MALAT1 upregulated the expression of miR-204 target gene SLUG through competitively ‘spongeing’ miR-204. In summary we unveil a branch of the MALAT1/miR-204/SLUG pathway that regulates the progression of lung adenocarcinoma.
PMCID: PMC4889723  PMID: 27294002
Lung adenocarcinoma; MALAT1; miR-204; SLUG
11.  Sleep deprivation disturbed regional brain activity in healthy subjects: evidence from a functional magnetic resonance-imaging study 
The aim of this study was to use amplitude of low-frequency fluctuation (ALFF) to explore regional brain activities in healthy subjects after sleep deprivation (SD).
Materials and methods
A total of 16 healthy subjects (eight females, eight males) underwent the session twice: once was after normal sleep (NS), and the other was after SD. ALFF was used to assess local brain features. The mean ALFF-signal values of the different brain areas were evaluated to investigate relationships with clinical features and were analyzed with a receiver-operating characteristic curve.
Compared with NS subjects, SD subjects showed a lower response-accuracy rate, longer response time, and higher lapse rate. Compared with NS subjects, SD subjects showed higher ALFF area in the right cuneus and lower ALFF area in the right lentiform nucleus, right claustrum, left dorsolateral prefrontal cortex, and left inferior parietal cortex. ALFF differences in regional brain areas showed high sensitivity and specificity. In the SD group, mean ALFF of the right claustrum showed a significant positive correlation with accuracy rate (r=0.687, P=0.013) and a negative correlation with lapse rate (r=−0.706, P=0.01). Mean ALFF of the dorsolateral prefrontal cortex showed a significant positive correlation with response time (r=0.675, P=0.016).
SD disturbed the regional brain activity of the default-mode network, its anticorrelated “task-positive” network, and the advanced cognitive function brain areas.
PMCID: PMC4835129  PMID: 27110113
sleep deprivation; amplitude of low-frequency fluctuation; default-mode network; functional magnetic resonance imaging
12.  Lipid remodelling is a widespread strategy in marine heterotrophic bacteria upon phosphorus deficiency 
The ISME Journal  2015;10(4):968-978.
Upon phosphorus (P) deficiency, marine phytoplankton reduce their requirements for P by replacing membrane phospholipids with alternative non-phosphorus lipids. It was very recently demonstrated that a SAR11 isolate also shares this capability when phosphate starved in culture. Yet, the extent to which this process occurs in other marine heterotrophic bacteria and in the natural environment is unknown. Here, we demonstrate that the substitution of membrane phospholipids for a variety of non-phosphorus lipids is a conserved response to P deficiency among phylogenetically diverse marine heterotrophic bacteria, including members of the Alphaproteobacteria and Flavobacteria. By deletion mutagenesis and complementation in the model marine bacterium Phaeobacter sp. MED193 and heterologous expression in recombinant Escherichia coli, we confirm the roles of a phospholipase C (PlcP) and a glycosyltransferase in lipid remodelling. Analyses of the Global Ocean Sampling and Tara Oceans metagenome data sets demonstrate that PlcP is particularly abundant in areas characterized by low phosphate concentrations. Furthermore, we show that lipid remodelling occurs seasonally and responds to changing nutrient conditions in natural microbial communities from the Mediterranean Sea. Together, our results point to the key role of lipid substitution as an adaptive strategy enabling heterotrophic bacteria to thrive in the vast P-depleted areas of the ocean.
PMCID: PMC4796936  PMID: 26565724
13.  Trimethylamine and trimethylamine N-oxide are supplementary energy sources for a marine heterotrophic bacterium: implications for marine carbon and nitrogen cycling 
The ISME Journal  2014;9(3):760-769.
Bacteria of the marine Roseobacter clade are characterised by their ability to utilise a wide range of organic and inorganic compounds to support growth. Trimethylamine (TMA) and trimethylamine N-oxide (TMAO) are methylated amines (MA) and form part of the dissolved organic nitrogen pool, the second largest source of nitrogen after N2 gas, in the oceans. We investigated if the marine heterotrophic bacterium, Ruegeria pomeroyi DSS-3, could utilise TMA and TMAO as a supplementary energy source and whether this trait had any beneficial effect on growth. In R. pomeroyi, catabolism of TMA and TMAO resulted in the production of intracellular ATP which in turn helped to enhance growth rate and growth yield as well as enhancing cell survival during prolonged energy starvation. Furthermore, the simultaneous use of two different exogenous energy sources led to a greater enhancement of chemoorganoheterotrophic growth. The use of TMA and TMAO primarily as an energy source resulted in the remineralisation of nitrogen in the form of ammonium, which could cross feed into another bacterium. This study provides greater insight into the microbial metabolism of MAs in the marine environment and how it may affect both nutrient flow within marine surface waters and the flux of these climatically important compounds into the atmosphere.
PMCID: PMC4331573  PMID: 25148480
14.  Magnetic nanoparticle-mediated isolation of functional bacteria in a complex microbial community 
The ISME Journal  2014;9(3):603-614.
Although uncultured microorganisms have important roles in ecosystems, their ecophysiology in situ remains elusive owing to the difficulty of obtaining live cells from their natural habitats. In this study, we employed a novel magnetic nanoparticle-mediated isolation (MMI) method to recover metabolically active cells of a group of previously uncultured phenol degraders, Burkholderiales spp., from coking plant wastewater biosludge; five other culturable phenol degraders—Rhodococcus sp., Chryseobacterium sp. and three different Pseudomonas spp.—were also isolated from the same biosludge using traditional methods. The kinetics of phenol degradation by MMI-recovered cells (MRCs) was similar to that of the original sludge. Stable isotope probing (SIP) and pyrosequencing of the 16S rRNA from the ‘heavy' DNA (13C-DNA) fractions indicated that Burkholderiales spp. were the key phenol degraders in situ in the biosludge, consistent with the results of MRCs. Single-cell Raman micro-spectroscopy was applied to probe individual bacteria in the MRCs obtained from the SIP experiment and showed that 79% of them were fully 13C-labelled. Biolog assays on the MRCs revealed the impact of various carbon and nitrogen substrates on the efficiency of phenol degradation in the wastewater treatment plant biosludge. Specifically, hydroxylamine, a metabolite of ammonia oxidisation, but not nitrite, nitrate or ammonia, inhibited phenol degradation in the biosludge. Our results provided a novel insight into the occasional abrupt failure events that occur in the wastewater treatment plant. This study demonstrated that MMI is a powerful tool to recover live and functional cells in situ from a complex microbial community to enable further characterisation of their physiology.
PMCID: PMC4331582  PMID: 25191996
15.  Comparative genomics and mutagenesis analyses of choline metabolism in the marine R oseobacter clade 
Environmental Microbiology  2015;17(12):5048-5062.
Choline is ubiquitous in marine eukaryotes and appears to be widely distributed in surface marine waters; however, its metabolism by marine bacteria is poorly understood. Here, using comparative genomics and molecular genetic approaches, we reveal that the capacity for choline catabolism is widespread in marine heterotrophs of the marine Roseobacter clade (MRC). Using the model bacterium R uegeria pomeroyi, we confirm that the bet A, bet B and bet C genes, encoding choline dehydrogenase, betaine aldehyde dehydrogenase and choline sulfatase, respectively, are involved in choline metabolism. The bet T gene, encoding an organic solute transporter, was essential for the rapid uptake of choline but not glycine betaine (GBT). Growth of choline and GBT as a sole carbon source resulted in the re‐mineralization of these nitrogen‐rich compounds into ammonium. Oxidation of the methyl groups from choline requires formyltetrahydrofolate synthetase encoded by fhs in R . pomeroyi, deletion of which resulted in incomplete degradation of GBT. We demonstrate that this was due to an imbalance in the supply of reducing equivalents required for choline catabolism, which can be alleviated by the addition of formate. Together, our results demonstrate that choline metabolism is ubiquitous in the MRC and reveal the role of Fhs in methyl group oxidation in R . pomeroyi.
PMCID: PMC4744692  PMID: 26058574
16.  Repeated measures of urinary oxidative stress biomarkers during pregnancy and preterm birth 
To investigate oxidative stress as a mechanism of preterm birth in human subjects, we examined associations between urinary biomarkers of oxidative stress measured at multiple time points during pregnancy and preterm birth.
Study Design
This nested case-control study included 130 mothers who delivered preterm and 352 who delivered term who were originally recruited as part of an ongoing prospective birth cohort at Brigham and Women’s Hospital. Two biomarkers, including 8-hydroxydeoxyguanosine (8-OHdG) and 8-isoprostane were measured in urine samples collected at up to four time points (median 10, 18, 26, and 35 weeks) during gestation.
Urinary concentrations of 8-isoprostane and 8-OHdG decreased and increased, respectively, as pregnancy progressed. Average levels of 8-isoprostane across pregnancy were associated with increased odds of spontaneous preterm birth (adjusted odds ratio [aOR]=6.25, 95% confidence interval [CI]=2.86, 13.7), and associations were strongest with levels measured later in pregnancy. Average levels of 8-OHdG were protective against overall preterm birth (aOR=0.19, 95%CI=0.10, 0.34), and there were no apparent differences in the protective effect in cases of spontaneous preterm birth compared to cases of placental origin. Odds ratios for overall preterm birth were more protective in association with urinary 8-OHdG concentrations measured early in pregnancy.
Maternal oxidative stress may be an important contributor to preterm birth, regardless of subtype and timing of exposure during pregnancy. The two biomarkers measured in the present study had opposite associations with preterm birth; an improved understanding of what each represents may help to more precisely identify important mechanisms in the pathway to preterm birth.
PMCID: PMC4312513  PMID: 25111586
Epidemiology; longitudinal; oxidative stress; preterm birth; repeated measures
17.  Relative 11C-PiB Delivery as a Proxy of Relative CBF: Quantitative Evaluation Using Single-Session 15O-Water and 11C-PiB PET 
The primary goal of this study was to assess the suitability of 11C-Pittsburgh compound B (11C-PiB) blood–brain barrier delivery (K1) and relative delivery (R1) parameters as surrogate indices of cerebral blood flow (CBF), with a secondary goal of directly examining the extent to which simplified uptake measures of 11C-PiB retention (amyloid-β load) may be influenced by CBF, in a cohort of controls and patients with mild cognitive impairment (MCI) and Alzheimer disease (AD).
Nineteen participants (6 controls, 5 AD, 8 MCI) underwent MR imaging, 15O-water PET, and 11C-PiB PET in a single session. Fourteen regions of interest (including cerebellar reference region) were defined on MR imaging and applied to dynamic coregistered PET to generate time–activity curves. Multiple analysis approaches provided regional 15O-water and 11C-PiB measures of delivery and 11C-PiB retention that included compartmental modeling distribution volume ratio (DVR), arterial- and reference-based Logan DVR, simplified reference tissue modeling 2 (SRTM2) DVR, and standardized uptake value ratios. Spearman correlation was performed among delivery measures (i.e., 15O-water K1 and 11C-PiB K1, relative K1 normalized to cerebellum [Rel-K1-Water and Rel-K1-PiB], and 11C-PiB SRTM2-R1) and between delivery measures and 11C-PiB retention, using the Bonferroni method for multiple-comparison correction.
Primary analysis showed positive correlations (ρ ≈0.2–0.5) between 15O-water K1 and 11C-PiB K1 that did not survive Bonferroni adjustment. Significant positive correlations were found between Rel-K1-Water and Rel-K1-PiB and between Rel-K1-Water and 11C-PiB SRTM2-R1 (ρ ≈0.5–0.8, P < 0.0036) across primary cortical regions. Secondary analysis showed few significant correlations between 11C-PiB retention and relative 11C-PiB delivery measures (but not 15O-water delivery measures) in primary cortical areas that arose only after accounting for cerebrospinal fluid dilution.
11C-PiB SRTM2-R1 is highly correlated with regional relative CBF, as measured by 15O-water K1 normalized to cerebellum, and cross-sectional 11C-PiB retention did not strongly depend on CBF across primary cortical regions. These results provide further support for potential dual-imaging assessments of regional brain status (i.e., amyloid-β load and relative CBF) through dynamic 11C-PiB imaging.
PMCID: PMC4730871  PMID: 26045309
amyloid; blood flow; PiB PET; Alzheimer’s disease; kinetic modeling
18.  Cardiopulmonary Function, Exercise Capacity, and Echocardiography Finding of Pediatric Patients With Kawasaki Disease 
Medicine  2016;95(2):e2444.
Coronary artery (CA) abnormalities influence exercise capacity (EC) of patients with Kawasaki disease (KD), and Z-score of CA is a well established method for detecting CA aneurysm. We studied the influence of KD on cardiopulmonary function and EC; meanwhile we analyzed echocardiographic findings of KD patients. We also assessed the correlation between CA Z-score and EC of KD patients to see if CA Z-score of KD patients could reflect EC during exercise.
Sixty-three KD patients were recruited as KD group 1 from children (aged 5–18 y) who received transthoracic echocardiographic examinations and symptom-limited treadmill exercise test for regular follow-up of KD from January 2010 to October 2014 in 1 medical center. We then divided KD group 1 into KD group 2 (<5 y, n = 12) and KD group 3 (≥5 y, n = 51) according to time interval between KD onset to when patients received test. Control groups were matched by age, sex, and body mass index. Max-Z of CA was defined as the maximal Z-score of the proximal LCA or RCA by Dalliarre equation or Fuse calculator.
All routine parameters measured during standard exercise test were similar between KD and control groups, except that peak rate pressure products (PRPPs) in KD group 1 to 3 were all lower than corresponding control groups significantly (P = 0.010, 0.020, and 0.049, respectively). PRPPs correlated with Max-Z of CA by both equations modest inversely (by Dallaire, P = 0.017, Spearman rho = −0.301; by Fuse, P = 0.014, Spearman rho = −0.309).
Our study recruited larger number of KD patients and provided a newer data of EC of KD patients. Our finding suggests that after acute stage of KD, patients could maintain normal cardiorespiratory fitness. Therefore, we believe that it is important to promote cardiovascular health to KD patients and KD patients should exercise as normal peers. However, since KD patients might still have compromised coronary perfusion during exercise, it remains crucial to assess and monitor cardiovascular risk of KD patients. Max-Z of CA correlates with PRPP modest inversely and might be used as a follow-up indicator of CA reserve during exercise after acute stage of KD.
PMCID: PMC4718257  PMID: 26765431
19.  Utilizing Longitudinal Measures of Fetal Growth to Create a Standard Method to Assess the Impacts of Maternal Disease and Environmental Exposure 
PLoS ONE  2016;11(1):e0146532.
Impaired or suboptimal fetal growth is associated with an increased risk of perinatal morbidity and mortality. By utilizing readily available clinical data on the relative size of the fetus at multiple points in pregnancy, including delivery, future epidemiological research can improve our understanding of the impacts of maternal, fetal, and environmental factors on fetal growth at different windows during pregnancy. This study presents mean and standard deviation ultrasound measurements from a clinically representative US population that can be utilized for creating Z-scores to this end. Between 2006 and 2012, 18, 904 non-anomalous pregnancies that received prenatal care, first and second trimester ultrasound evaluations, and ultimately delivered singleton newborns at Brigham and Women’s hospital in Boston were used to create the standard population. To illustrate the utility of this standard, we created Z-scores for ultrasound and delivery measurements for a cohort study population and examined associations with factors known to be associated with fetal growth. In addition to cross-sectional regression models, we created linear mixed models and generalized additive mixed models to illustrate how these scores can be utilized longitudinally and for the identification of windows of susceptibility. After adjustment for a priori confounders, maternal BMI was positively associated with increased fetal size beginning in the second trimester in cross-sectional models. Female infants and maternal smoking were associated with consistently reduced fetal size in the longitudinal models. Maternal age had a non-significant association with increased size in the first trimester that was attenuated as gestation progressed. As the growth measurements examined here are widely available in contemporary obstetrical practice, these data may be abstracted from medical records by investigators and standardized with the population means presented here. This will enable easy extension of clinical data to epidemiologic studies investigating novel maternal, fetal, and environmental factors that may impact fetal growth.
PMCID: PMC4701464  PMID: 26731406
20.  Multiple viral introductions: molecular characterization of influenza B virus in Wenzhou, Zhejiang, China, from 2011 to 2014 based on hemagglutinin and neuraminidase genes 
Archives of Virology  2016;161:1005-1013.
Influenza B virus is a major causative agent of respiratory disease in humans. Our study of an outbreak of influenza B virus in Wenzhou from 2011 to 2014 revealed that 163 (5.58 %) of 2921 samples were influenza B positive. Sequencing of the hemagglutinin and neuraminidase genes showed substitutions at the amino acid level. Phylogenetic analysis revealed co-circulation of the B/Victoria and B/Yamagata lineages in the Wenzhou area from 2011 to 2014. Multiple viral introductions from both Chinese and international sources played important roles in endemic co-circulation and transmission in coastal southeastern China.
Electronic supplementary material
The online version of this article (doi:10.1007/s00705-015-2721-7) contains supplementary material, which is available to authorized users.
PMCID: PMC4819752  PMID: 26724821
Influenza B virus; Molecular epidemiology; Phylogenetic; Victoria; Yamagata
21.  Bacterial metabolism of methylated amines and identification of novel methylotrophs in Movile Cave 
The ISME Journal  2014;9(1):195-206.
Movile Cave, Romania, is an unusual underground ecosystem that has been sealed off from the outside world for several million years and is sustained by non-phototrophic carbon fixation. Methane and sulfur-oxidising bacteria are the main primary producers, supporting a complex food web that includes bacteria, fungi and cave-adapted invertebrates. A range of methylotrophic bacteria in Movile Cave grow on one-carbon compounds including methylated amines, which are produced via decomposition of organic-rich microbial mats. The role of methylated amines as a carbon and nitrogen source for bacteria in Movile Cave was investigated using a combination of cultivation studies and DNA stable isotope probing (DNA-SIP) using 13C-monomethylamine (MMA). Two newly developed primer sets targeting the gene for gamma-glutamylmethylamide synthetase (gmaS), the first enzyme of the recently-discovered indirect MMA-oxidation pathway, were applied in functional gene probing. SIP experiments revealed that the obligate methylotroph Methylotenera mobilis is one of the dominant MMA utilisers in the cave. DNA-SIP experiments also showed that a new facultative methylotroph isolated in this study, Catellibacterium sp. LW-1 is probably one of the most active MMA utilisers in Movile Cave. Methylated amines were also used as a nitrogen source by a wide range of non-methylotrophic bacteria in Movile Cave. PCR-based screening of bacterial isolates suggested that the indirect MMA-oxidation pathway involving GMA and N-methylglutamate is widespread among both methylotrophic and non-methylotrophic MMA utilisers from the cave.
PMCID: PMC4274414  PMID: 25050523
22.  Activation of GLP-1 Receptor Enhances Neuronal Base Excision Repair via PI3K-AKT-Induced Expression of Apurinic/Apyrimidinic Endonuclease 1 
Theranostics  2016;6(12):2015-2027.
Glucagon-like peptide-1 (GLP-1) is an intestinal-secreted incretin that increases cellular glucose up-take to decrease blood sugar. Recent studies, however, suggest that the function of GLP-1 is not only to decrease blood sugar, but also acts as a neurotrophic factor that plays a role in neuronal survival, neurite outgrowth, and protects synaptic plasticity and memory formation from effects of β-amyloid. Oxidative DNA damage occurs during normal neuron-activity and in many neurological diseases. Our study describes how GLP-1 affected the ability of neurons to ameliorate oxidative DNA damage. We show that activation of GLP-1 receptor (GLP-1R) protect cortical neurons from menadione induced oxidative DNA damage via a signaling pathway involving enhanced DNA repair. GLP-1 stimulates DNA repair by activating the cyclic AMP response element binding protein (CREB) which, consequently, induces the expression of apurinic/apyrimidinic endonuclease 1 (APE1), a key enzyme in the base excision DNA repair (BER) pathway. In this study, APE1 expression was down-regulated as a consequence phosphatidylinositol-3 kinase (PI3K) suppression by the inhibitor LY294002, but not by the suppression of MEK activity. Ischemic stroke is typically caused by overwhelming oxidative-stress in brain cells. Administration of exentin-4, an analogue of GLP-1, efficiently enhanced DNA repair in brain cells of ischemic stroke rats. Our study suggests that a new function of GLP-1 is to elevate DNA repair by inducing the expression of the DNA repair protein APE1.
PMCID: PMC5039677  PMID: 27698937
Glucagon-like peptide-1(GLP-1); Exendin-4; Oxidative DNA damage; Base excision repair (BER); Apurinic/apyrimidinic endonuclease 1 (APE1)
23.  Loss of CPEB3 Upregulates MEGF10 to Impair Mosaic Development of ON Starburst Amacrine Cells 
Cytoplasmic polyadenylation element binding protein 3 (CPEB3) regulates target RNA translation in neurons. Here, we examined CPEB3 distribution and function in the mouse retina. CPEB3 is expressed in retinal neurons, including those located in the inner nuclear layer (INL) and ganglion cell layer (GCL) but not in cone and rod photoreceptors in the outer nuclear layer (ONL). A previous study found CPEB3 expressed in cholinergic starburst amacrine cells (SACs). We first examined these cells and observed aberrant SAC mosaicism in CPEB3-knockout (KO) retinas. Retinal neurons showed orderly spatial arrangements. Many individual subtypes are organized non-randomly in patterns called mosaics. Despite CPEB3 being expressed in both populations of SACs, OFF SACs in the INL and ON SACs in the GCL, aberrant mosaic regularity was observed in only ON SACs of CPEB3-KO retinas. Molecular characterization revealed that translation of multiple epidermal growth factor 10 (Megf10) RNA is suppressed by CPEB3 during the first week of postnatal development, when MEGF10 is primarily expressed in SACs and mediates homotypic repulsive interactions to define intercellular spacing of SACs. Thus, elevated MEGF10 expression in the absence of the translational repressor CPEB3 may account for the defective spatial organization of ON SACs. Our findings uncover for the first time that translational control plays a role in shaping retinal mosaic arrangement.
PMCID: PMC5075539  PMID: 27822178
CPEB; MEGF10; retinal mosaic; starburst amacrine cells; translational control
24.  Focal adhesion kinase-dependent focal adhesion recruitment of SH2 domains directs SRC into focal adhesions to regulate cell adhesion and migration 
Scientific Reports  2015;5:18476.
Directed cell migration requires dynamical control of the protein complex within focal adhesions (FAs) and this control is regulated by signaling events involving tyrosine phosphorylation. We screened the SH2 domains present in tyrosine-specific kinases and phosphatases found within FAs, including SRC, SHP1 and SHP2, and examined whether these enzymes transiently target FAs via their SH2 domains. We found that the SRC_SH2 domain and the SHP2_N-SH2 domain are associated with FAs, but only the SRC_SH2 domain is able to be regulated by focal adhesion kinase (FAK). The FAK-dependent association of the SRC_SH2 domain is necessary and sufficient for SRC FA targeting. When the targeting of SRC into FAs is inhibited, there is significant suppression of SRC-mediated phosphorylation of paxillin and FAK; this results in an inhibition of FA formation and maturation and a reduction in cell migration. This study reveals an association between FAs and the SRC_SH2 domain as well as between FAs and the SHP2_N-SH2 domains. This supports the hypothesis that the FAK-regulated SRC_SH2 domain plays an important role in directing SRC into FAs and that this SRC-mediated FA signaling drives cell migration.
PMCID: PMC4683442  PMID: 26681405
25.  Zebrafish Egg Infection Model for Studying Candida albicans Adhesion Factors 
PLoS ONE  2015;10(11):e0143048.
Disseminated candidiasis is associated with 30–40% mortality in severely immunocompromised patients. Among the causal agents, Candida albicans is the dominant one. Various animal models have been developed for investigating gene functions in C. albicans. Zebrafish injection models have increasingly been applied in elucidating C. albicans pathogenesis because of the conserved immunity, prolific fecundity of the zebrafish and the low costs of care systems. In this study, we established a simple, noninvasive zebrafish egg bath infection model, defined its optimal conditions, and evaluated the model with various C. albicans mutant strains. The deletion of SAP6 did not have significant effect on the virulence. By contrast, the deletion of BCR1, CPH1, EFG1, or TEC1 significantly reduced the virulence under current conditions. Furthermore, all embryos survived when co-incubated with bcr1/bcr1, cph1/cph1 efg1/efg1, efg1/efg1, or tec1/tec1 mutant cells. The results indicated that our novel zebrafish model is time-saving and cost effective.
PMCID: PMC4646526  PMID: 26569623

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