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1.  An SPM8-based Approach for Attenuation Correction Combining Segmentation and Non-rigid Template Formation: Application to Simultaneous PET/MR Brain Imaging 
We present an approach for head MR-based attenuation correction (MR-AC) based on the Statistical Parametric Mapping (SPM8) software that combines segmentation- and atlas-based features to provide a robust technique to generate attenuation maps (µ-maps) from MR data in integrated PET/MR scanners.
Methods
Coregistered anatomical MR and CT images acquired in 15 glioblastoma subjects were used to generate the templates. The MR images from these subjects were first segmented into 6 tissue classes (gray and white matter, cerebro-spinal fluid, bone and soft tissue, and air), which were then non-rigidly coregistered using a diffeomorphic approach. A similar procedure was used to coregister the anatomical MR data for a new subject to the template. Finally, the CT-like images obtained by applying the inverse transformations were converted to linear attenuation coefficients (LACs) to be used for AC of PET data. The method was validated on sixteen new subjects with brain tumors (N=12) or mild cognitive impairment (N=4) who underwent CT and PET/MR scans. The µ-maps and corresponding reconstructed PET images were compared to those obtained using the gold standard CT-based approach and the Dixon-based method available on the Siemens Biograph mMR scanner. Relative change (RC) images were generated in each case and voxel- and region of interest (ROI)-based analyses were performed.
Results
The leave-one-out cross-validation analysis of the data from the 15 atlas-generation subjects showed small errors in brain LACs (RC=1.38%±4.52%) compared to the gold standard. Similar results (RC=1.86±4.06%) were obtained from the analysis of the atlas-validation datasets. The voxel- and ROI-based analysis of the corresponding reconstructed PET images revealed quantification errors of 3.87±5.0% and 2.74±2.28%, respectively. The Dixon-based method performed substantially worse (the mean RC values were 13.0±10.25% and 9.38±4.97%, respectively). Areas closer to skull showed the largest improvement.
Conclusion
We have presented an SPM8-based approach for deriving the head µ-map from MR data to be used for PET AC in integrated PET/MR scanners. Its implementation is straightforward and only requires the morphological data acquired with a single MR sequence. The method is very accurate and robust, combining the strengths of both segmentation- and atlas-based approaches while minimizing their drawbacks.
doi:10.2967/jnumed.113.136341
PMCID: PMC4246705  PMID: 25278515
integrated PET/MRI; attenuation correction; segmentation; template
2.  Fully printed flexible and disposable wireless cyclic voltammetry tag 
Scientific Reports  2015;5:8105.
A disposable cyclic voltammetry (CV) tag is printed on a plastic film by integrating wireless power transmitter, polarized triangle wave generator, electrochemical cell and signage through a scalable gravure printing method. By proximity of 13.56 MHz RF reader, the printed CV tag generates 320 mHz of triangular sweep wave from +500 mV to −500 mV which enable to scan a printed electrochemical cell in the CV tag. By simply dropping any specimen solution on the electrochemical cell in the CV tag, the presence of solutes in the solution can be detected and shown on the signage of the CV tag in five sec. 10 mM of N,N,N′,N′-tetramethyl-p-phenylenediamine (TMPD) was used as a standard solute to prove the working concept of fully printed disposable wireless CV tag. Within five seconds, we can wirelessly diagnose the presence of TMPD in the solution using the CV tag in the proximity of the 13.56 MHz RF reader. This fully printed and wirelessly operated flexible CV tag is the first of its kind and marks the path for the utilization of inexpensive and disposable wireless electrochemical sensor systems for initial diagnose hazardous chemicals and biological molecules to improve public hygiene and health.
doi:10.1038/srep08105
PMCID: PMC4309959  PMID: 25630250
3.  Monoamine Oxidase B Prompts Mitochondrial and Cardiac Dysfunction in Pressure Overloaded Hearts 
Antioxidants & Redox Signaling  2014;20(2):267-280.
Abstract
Aims: Monoamine oxidases (MAOs) are mitochondrial flavoenzymes responsible for neurotransmitter and biogenic amines catabolism. MAO-A contributes to heart failure progression via enhanced norepinephrine catabolism and oxidative stress. The potential pathogenetic role of the isoenzyme MAO-B in cardiac diseases is currently unknown. Moreover, it is has not been determined yet whether MAO activation can directly affect mitochondrial function. Results: In wild type mice, pressure overload induced by transverse aortic constriction (TAC) resulted in enhanced dopamine catabolism, left ventricular (LV) remodeling, and dysfunction. Conversely, mice lacking MAO-B (MAO-B−/−) subjected to TAC maintained concentric hypertrophy accompanied by extracellular signal regulated kinase (ERK)1/2 activation, and preserved LV function, both at early (3 weeks) and late stages (9 weeks). Enhanced MAO activation triggered oxidative stress, and dropped mitochondrial membrane potential in the presence of ATP synthase inhibitor oligomycin both in neonatal and adult cardiomyocytes. The MAO-B inhibitor pargyline completely offset this change, suggesting that MAO activation induces a latent mitochondrial dysfunction, causing these organelles to hydrolyze ATP. Moreover, MAO-dependent aldehyde formation due to inhibition of aldehyde dehydrogenase 2 activity also contributed to alter mitochondrial bioenergetics. Innovation: Our study unravels a novel role for MAO-B in the pathogenesis of heart failure, showing that both MAO-driven reactive oxygen species production and impaired aldehyde metabolism affect mitochondrial function. Conclusion: Under conditions of chronic hemodynamic stress, enhanced MAO-B activity is a major determinant of cardiac structural and functional disarrangement. Both increased oxidative stress and the accumulation of aldehyde intermediates are likely liable for these adverse morphological and mechanical changes by directly targeting mitochondria. Antioxid. Redox Signal. 20, 267–280.
doi:10.1089/ars.2012.4616
PMCID: PMC3887464  PMID: 23581564
4.  Human Pluripotent Stem Cell Culture: Considerations for Maintenance, Expansion, and Therapeutics 
Cell stem cell  2014;14(1):13-26.
Summary
Human pluripotent stem cells (hPSCs) provide powerful resources for application in regenerative medicine and pharmaceutical development. In the past decade, various methods have been developed for large-scale hPSC culture that rely on combined use of multiple growth components, including media containing various growth factors, extracellular matrices, three-dimensional environmental (3D) cues and modes of multicellular association. In this review, we dissect these growth components by comparing cell culture methods and identifying the benefits and pitfalls associated with each one. We further provide criteria, considerations, and suggestions to achieve optimal cell growth for hPSC expansion, differentiation, and use in future therapeutic applications.
doi:10.1016/j.stem.2013.12.005
PMCID: PMC3915741  PMID: 24388173
Human embryonic stem cells; induced pluripotent stem cells; cell culture; expansion; differentiation; regenerative medicine
5.  Directed Differentiation of Human Induced Pluripotent Stem Cells Toward Bone and Cartilage: In Vitro Versus In Vivo Assays 
The ability to generate large numbers of bone- and cartilage-forming cells from induced pluripotent stem cells (iPSCs) would mark a major advance in tissue engineering. A number of protocols exist, but the overall quality and consistency of this type of differentiation are under-reported. In this study, the authors analyzed differentiated iPSCs in vitro and in vivo by stringent criteria, and found that in vitro analysis does not predict in vivo differentiation.
The ability to differentiate induced pluripotent stem cells (iPSCs) into committed skeletal progenitors could allow for an unlimited autologous supply of such cells for therapeutic uses; therefore, we attempted to create novel bone-forming cells from human iPSCs using lines from two distinct tissue sources and methods of differentiation that we previously devised for osteogenic differentiation of human embryonic stem cells, and as suggested by other publications. The resulting cells were assayed using in vitro methods, and the results were compared with those obtained from in vivo transplantation assays. Our results show that true bone was formed in vivo by derivatives of several iPSC lines, but that the successful cell lines and differentiation methodologies were not predicted by the results of the in vitro assays. In addition, bone was formed equally well from iPSCs originating from skin or bone marrow stromal cells (also known as bone marrow-derived mesenchymal stem cells), suggesting that the iPSCs did not retain a “memory” of their previous life. Furthermore, one of the iPSC-derived cell lines formed verifiable cartilage in vivo, which likewise was not predicted by in vitro assays.
doi:10.5966/sctm.2013-0154
PMCID: PMC4073820  PMID: 24855277
Induced pluripotent stem cells; Bone; Osteoblast; Chondrogenesis; Transplantation
6.  Infusion of imaging and therapeutic molecules into the plant virus-based carrier cowpea mosaic virus: cargo-loading and delivery 
This work is focused on the development of a plant virus-based carrier system for cargo delivery, specifically 30 nm-sized cowpea mosaic virus (CPMV). Whereas previous reports described the engineering of CPMV through genetic or chemical modification, we report a non-covalent infusion technique that facilitates efficient cargo loading. Infusion and retention of 130–155 fluorescent dye molecules per CPMV using DAPI (4’,6-diamidino-2-phenylindole dihydrochloride), propidium iodide (3,8-diamino-5-[3-(diethylmethylammonio)propyl]-6-phenylphenanthridinium diiodide), and acridine orange (3,6-bis(dimethylamino)acridinium chloride), as well as 140 copies of therapeutic payload proflavine (PF, acridine-3,6-diamine hydrochloride), is reported. Loading is achieved through interaction of the cargo with the CPMV’s encapsidated RNA molecules. The loading mechanism is specific; empty RNA-free eCPMV nanoparticles could not be loaded. Cargo-infused CPMV nanoparticles remain chemically active, and surface lysine residues were covalent modified with dyes leading to the development of dual-functional CPMV carrier systems. We demonstrate cargo-delivery to a panel of cancer cells (cervical, breast, and colon): CPMV nanoparticles enter cells via the surface marker vimentin, the nanoparticles target the endolysosome, where the carrier is degraded and the cargo released allowing imaging and/or cell killing. In conclusion, we demonstrate cargo-infusion and delivery to cells; the methods discussed provide a useful means for functionalization of CPMV toward its application as drug and/or contrast agent delivery vehicle.
doi:10.1016/j.jconrel.2013.04.023
PMCID: PMC3815978  PMID: 23665254
7.  Early postnatal inhibition of serotonin synthesis results in long-term reductions of perseverative behaviors, but not aggression, in MAO A-deficient mice 
Neuropharmacology  2013;0:10.1016/j.neuropharm.2013.07.003.
Monoamine oxidase (MAO) A, the major enzyme catalyzing the oxidative degradation of serotonin (5-hydroxytryptamine, 5-HT), plays a key role in emotional regulation. In humans and mice, MAO-A deficiency results in high 5-HT levels, antisocial, aggressive, and perseverative behaviors. We previously showed that the elevation in brain 5-HT levels in MAO-A knockout (KO) mice is particularly marked during the first two weeks of postnatal life. Building on this finding, we hypothesized that the reduction of 5-HT levels during these early stages may lead to enduring attenuations of the aggression and other behavioral aberrances observed in MAO-A KO mice. To test this possibility, MAO-A KO mice were treated with daily injections of a 5-HT synthesis blocker, the tryptophan hydroxylase inhibitor p-chloro-phenylalanine (pCPA, 300 mg/kg/day, IP), from postnatal day 1 through 7. As expected, this regimen significantly reduced 5-HT forebrain levels in MAO-A KO pups. These neurochemical changes persisted throughout adulthood, and resulted in significant reductions in marble-burying behavior, as well as increases in spontaneous alternations within a T-maze. Conversely, pCPA-treated MAO-A KO mice did not exhibit significant changes in anxiety-like behaviors in a novel open-field and elevated plus-maze; furthermore, this regimen did not modify their social deficits, aggressive behaviors and impairments in tactile sensitivity. Treatment with pCPA from postnatal day 8 through 14 elicited similar, yet milder, behavioral effects on marble-burying behavior. These results suggest that early developmental enhancements in 5-HT levels have long-term effects on the modulation of behavioral flexibility associated with MAO-A deficiency.
doi:10.1016/j.neuropharm.2013.07.003
PMCID: PMC3849223  PMID: 23871843
8.  Variation in piRNA and Transposable Element Content in Strains of Drosophila melanogaster 
Genome Biology and Evolution  2014;6(10):2786-2798.
Transposable elements (TEs) are one of the most important features of genome architecture, so their evolution and relationship with host defense mechanisms have been topics of intense study, especially in model systems such as Drosophila melanogaster. Recently, a novel small RNA-based defense mechanism in animals called the Piwi-interacting RNA (piRNA) pathway was discovered to form an adaptive defense mechanism against TEs. To investigate the relationship between piRNA and TE content between strains of a species, we sequenced piRNAs from 16 inbred lines of D. melanogaster from the Drosophila Genetic Reference Panel. Instead of a global correlation of piRNA expression and TE content, we found evidence for a host response through de novo piRNA production from novel TE insertions. Although approximately 20% of novel TE insertions induced de novo piRNA production, the abundance of de novo piRNAs was low and did not markedly affect the global pool of ovarian piRNAs. Our results provide new insights into the evolution of TEs and the piRNA system in an important model organism.
doi:10.1093/gbe/evu217
PMCID: PMC4224344  PMID: 25267446
piRNA; transposable elements; Drosophila melanogaster; de novo piRNA production
9.  MixMir: microRNA motif discovery from gene expression data using mixed linear models 
Nucleic Acids Research  2014;42(17):e135.
microRNAs (miRNAs) are a class of ∼22nt non-coding RNAs that potentially regulate over 60% of human protein-coding genes. miRNA activity is highly specific, differing between cell types, developmental stages and environmental conditions, so the identification of active miRNAs in a given sample is of great interest. Here we present a novel computational approach for analyzing both mRNA sequence and gene expression data, called MixMir. Our method corrects for 3’ UTR background sequence similarity between transcripts, which is known to correlate with mRNA transcript abundance. We demonstrate that after accounting for kmer sequence similarities in 3’ UTRs, a statistical linear model based on motif presence/absence can effectively discover active miRNAs in a sample. MixMir utilizes fast software implementations for solving mixed linear models, which are widely used in genome-wide association studies (GWASs). Essentially we use 3’ UTR sequence similarity in place of population cryptic relatedness in the GWAS problem. Compared to similar methods such as miReduce, Sylamer and cWords, we found that MixMir performed better at discovering true miRNA motifs in three mouse Dicer-knockout experiments from different tissues, two of which were collected by our group. We confirmed these results on protein and mRNA expression data obtained from miRNA transfection experiments in human cell lines. MixMir can be freely downloaded from https://github.com/ldiao/MixMir.
doi:10.1093/nar/gku672
PMCID: PMC4176157  PMID: 25081207
10.  A comprehensive analysis of piRNAs from adult human testis and their relationship with genes and mobile elements 
BMC Genomics  2014;15(1):545.
Background
Piwi-interacting RNAs (piRNAs) are a recently discovered class of small non-coding RNAs whose best-understood function is to repress mobile element (ME) activity in animal germline. To date, nearly all piRNA studies have been conducted in model organisms and little is known about piRNA diversity, target specificity and biological function in human.
Results
Here we performed high-throughput sequencing of piRNAs from three human adult testis samples. We found that more than 81% of the ~17 million putative piRNAs mapped to ~6,000 piRNA-producing genomic clusters using a relaxed definition of clusters. A set of human protein-coding genes produces a relatively large amount of putative piRNAs from their 3’UTRs, and are significantly enriched for certain biological processes, suggestive of non-random sampling by the piRNA biogenesis machinery. Up to 16% of putative piRNAs mapped to a few hundred annotated long non-coding RNA (lncRNA) genes, suggesting that some lncRNA genes can act as piRNA precursors. Among major ME families, young families of LTR and endogenous retroviruses have a greater association with putative piRNAs than other MEs. In addition, piRNAs preferentially mapped to specific regions in the consensus sequences of several ME (sub)families and some piRNA mapping peaks showed patterns consistent with the “ping-pong” cycle of piRNA targeting and amplification.
Conclusions
Overall our data provide a comprehensive analysis and improved annotation of human piRNAs in adult human testes and shed new light into the relationship of piRNAs with protein-coding genes, lncRNAs, and mobile genetic elements in human.
Electronic supplementary material
The online version of this article (doi:10.1186/1471-2164-15-545) contains supplementary material, which is available to authorized users.
doi:10.1186/1471-2164-15-545
PMCID: PMC4094622  PMID: 24981367
Human piRNA; piRNA cluster; Protein coding gene; Mobile element; High-throughput sequencing
11.  Cerebral cortical blood flow maps are reorganized in MAOB-deficient mice 
Brain research  1999;824(1):36-44.
Cerebral cortical blood flow (CBF) was measured autoradiographically in conscious mice without the monoamine oxidase B (MAOB) gene (KO, n = 11) and the corresponding wild-type animals (WILD, n = 11). Subgroups of animals of each genotype received a continuous intravenous infusion over 30 min of phenylethylamine (PEA), an endogenous substrate of MAOB, (8 nmol g−1 min−1 in normal saline at a volume rate of 0.11 μl g−1 min−1) or saline at the same volume rate. Maps of relative CBF distribution showed predominance of midline motor and sensory area CBF in KO mice over WILD mice that received saline. PEA enhanced CBF in lateral frontal and piriform cortex in both KO and WILD mice. These changes may reflect a differential activation due to chronic and acute PEA elevations on motor and olfactory function, as well as on the anxiogenic effects of this amine. In addition to its effects on regional CBF distribution, PEA decreased CBF globally in KO mice (range −31% to −41% decrease from control levels) with a lesser effect in WILD mice. It is concluded that MAOB may normally regulate CBF distribution and its response to blood PEA.
PMCID: PMC4075434  PMID: 10095040
Cerebral blood flow; Monoamine oxidase; Phenylethylamine; Autoradiography; Iodo-antipyrine; Mouse
12.  High-Throughput Screening of Effective siRNAs Using Luciferase-Linked Chimeric mRNA 
PLoS ONE  2014;9(5):e96445.
The use of siRNAs to knock down gene expression can potentially be an approach to treat various diseases. To avoid siRNA toxicity the less transcriptionally active H1 pol III promoter, rather than the U6 promoter, was proposed for siRNA expression. To identify highly efficacious siRNA sequences, extensive screening is required, since current computer programs may not render ideal results. Here, we used CCR5 gene silencing as a model to investigate a rapid and efficient screening approach. We constructed a chimeric luciferase-CCR5 gene for high-throughput screening of siRNA libraries. After screening approximately 900 shRNA clones, 12 siRNA sequences were identified. Sequence analysis demonstrated that most (11 of the 12 sequences) of these siRNAs did not match those identified by available siRNA prediction algorithms. Significant inhibition of CCR5 in a T-lymphocyte cell line and primary T cells by these identified siRNAs was confirmed using the siRNA lentiviral vectors to infect these cells. The inhibition of CCR5 expression significantly protected cells from R5 HIV-1JRCSF infection. These results indicated that the high-throughput screening method allows efficient identification of siRNA sequences to inhibit the target genes at low levels of expression.
doi:10.1371/journal.pone.0096445
PMCID: PMC4022502  PMID: 24831610
13.  ‘Marker-of-self’ functionalization of nanoscale particles through a top-down cellular membrane coating approach 
Nanoscale  2013;5(7):2664-2668.
We investigate the ‘marker-of-self’ functionalization of nanoparticles through coating of natural RBC membranes. The membrane translocation approach is shown to be highly efficient and bestows nanoparticles with correctly oriented and functional immunomodulatory proteins such as CD47 at equivalent density to natural RBCs.
doi:10.1039/c3nr00015j
PMCID: PMC3667603  PMID: 23462967
14.  Evidence for the biogenesis of more than 1,000 novel human microRNAs 
Genome Biology  2014;15(4):R57.
Background
MicroRNAs (miRNAs) are established regulators of development, cell identity and disease. Although nearly two thousand human miRNA genes are known and new ones are continuously discovered, no attempt has been made to gauge the total miRNA content of the human genome.
Results
Employing an innovative computational method on massively pooled small RNA sequencing data, we report 2,469 novel human miRNA candidates of which 1,098 are validated by in-house and published experiments. Almost 300 candidates are robustly expressed in a neuronal cell system and are regulated during differentiation or when biogenesis factors Dicer, Drosha, DGCR8 or Ago2 are silenced. To improve expression profiling, we devised a quantitative miRNA capture system. In a kidney cell system, 400 candidates interact with DGCR8 at transcript positions that suggest miRNA hairpin recognition, and 1,000 of the new miRNA candidates interact with Ago1 or Ago2, indicating that they are directly bound by miRNA effector proteins. From kidney cell CLASH experiments, in which miRNA-target pairs are ligated and sequenced, we observe hundreds of interactions between novel miRNAs and mRNA targets. The novel miRNA candidates are specifically but lowly expressed, raising the possibility that not all may be functional. Interestingly, the majority are evolutionarily young and overrepresented in the human brain.
Conclusions
In summary, we present evidence that the complement of human miRNA genes is substantially larger than anticipated, and that more are likely to be discovered in the future as more tissues and experimental conditions are sequenced to greater depth.
doi:10.1186/gb-2014-15-4-r57
PMCID: PMC4054668  PMID: 24708865
15.  Clinical Characteristics as a Function of Referral Status among Substance Users in Residential Treatment 
Addictive behaviors  2012;38(4):1924-1930.
In the United States, substance users who voluntarily (VO) elect to receive treatment and substance users who are court-mandated (CM) to receive treatment typically obtain care within the same facilities. Little is known about the clinical characteristics that differentiate these individuals. The current study provides rates of specific DSM-IV Axis I and II psychiatric and substance use disorders, comorbidities, childhood trauma, motivation, and other clinical and demographic characteristics as a function of referral status, among individuals in residential substance use treatment (463 participants, M age = 43.3; 69.7% male; 88.4% African American). Participants were interviewed and diagnosed using the Structure Clinical Interview for DSM-IV and the Diagnostic Interview for Personality Disorders. Within our sample, VO individuals, as compared to CM individuals had significantly higher rates of psychiatric disorders (68.7% versus 55.2%, respectively), including mood disorders, major depressive disorder, generalized anxiety disorder, and borderline personality disorder. Additionally, they were significantly more likely to have alcohol dependence (43.0% versus 20.8%) and cocaine dependence (66.5% versus 48.9%). Elevated rates of comorbidities and childhood abuse were also observed among VO individuals, while motivation did not differ as a function of referral status. Overall, VO individuals appeared to have more severe problems than their CM counterparts which may suggest that they require more intensive or different types of treatment.
doi:10.1016/j.addbeh.2012.12.015
PMCID: PMC3578165  PMID: 23380487
court-mandated; residential substance use treatment; comorbidities; substance dependence; psychiatric disorders
16.  Probabilistic atlas-based segmentation of combined T1-weighted and DUTE MRI for calculation of head attenuation maps in integrated PET/MRI scanners 
We present a new MRI-based attenuation correction (AC) approach for integrated PET/MRI systems that combines both segmentation- and atlas-based methods by incorporating dual-echo ultra-short echo-time (DUTE) and T1-weighted (T1w) MRI data and a probabilistic atlas. Segmented atlases were constructed from CT training data using a leave-one-out framework and combined with T1w, DUTE, and CT data to train a classifier that computes the probability of air/soft tissue/bone at each voxel. This classifier was applied to segment the MRI of the subject of interest and attenuation maps (μ-maps) were generated by assigning specific linear attenuation coefficients (LACs) to each tissue class. The μ-maps generated with this “Atlas-T1w-DUTE” approach were compared to those obtained from DUTE data using a previously proposed method. For validation of the segmentation results, segmented CT μ-maps were considered to the “silver standard”; the segmentation accuracy was assessed qualitatively and quantitatively through calculation of the Dice similarity coefficient (DSC). Relative change (RC) maps between the CT and MRI-based attenuation corrected PET volumes were also calculated for a global voxel-wise assessment of the reconstruction results. The μ-maps obtained using the Atlas-T1w-DUTE classifier agreed well with those derived from CT; the mean DSCs for the Atlas-T1w-DUTE-based μ-maps across all subjects were higher than those for DUTE-based μ-maps; the atlas-based μ-maps also showed a lower percentage of misclassified voxels across all subjects. RC maps from the atlas-based technique also demonstrated improvement in the PET data compared to the DUTE method, both globally as well as regionally.
PMCID: PMC3992209  PMID: 24753982
PET; MRI; attenuation correction; segmentation; atlas
17.  Identification of Genes with Consistent Methylation Levels across Different Human Tissues 
Scientific Reports  2014;4:4351.
DNA methylation plays an important role in regulating cell growth and disease development. Methylation profiles are examined by bisulfite conversion; however, the lack of markers for bisulfite conversion efficiency and appropriate internal control genes remains a major challenge. To address these issues, we utilized two bioinformatics approaches, coefficients of variances and resampling tests, to identify probes showing stable methylation levels from several independent microarray datasets. Mass spectrometry validated the consistently high methylation levels of the five probes (N4BP2, EGFL8, CTRB1, TSPAN3, and ZNF690) in 13 human tissue types from 24 cell lines. Linear associations between detected methylation levels and methyl concentrations of DNA samples were further demonstrated in three genes (N4BP2, EGFL8, and CTRB1). To summarize, we identified five genes which may serve as internal controls for methylation studies by analyzing large-scale microarray data, and three of them can be used as markers for evaluating the efficiency of bisulfite conversion.
doi:10.1038/srep04351
PMCID: PMC3950633  PMID: 24619003
19.  Prevalence and clinical correlates of explosive outbursts in Tourette Syndrome 
Psychiatry research  2012;205(3):269-275.
The aim of this study was to examine the prevalence and clinical correlates of explosive outbursts in two large samples of individuals with TS, including one collected primarily from non-clinical sources. Participants included 218 TS-affected individuals who were part of a genetic study (N=104 from Costa Rica (CR) and N=114 from the US). The relationship between explosive outbursts and comorbid attention deficit hyperactivity disorder (ADHD), obsessive compulsive disorder (OCD), tic severity, and prenatal and perinatal complications were examined using regression analyses. Twenty percent of participants had explosive outbursts, with no significant differences in prevalence between the CR (non-clinical) and the US (primarily clinical) samples. In the overall sample, ADHD, greater tic severity, and lower age of tic onset were strongly associated with explosive outbursts. ADHD, prenatal exposure to tobacco, and male gender were significantly associated with explosive outbursts in the US sample. Lower age of onset and greater severity of tics were significantly associated with explosive outbursts in the CR sample. This study confirms previous studies that suggest that clinically significant explosive outbursts are common in TS and associated with ADHD and tic severity. An additional potential risk factor, prenatal exposure to tobacco, was also identified.
doi:10.1016/j.psychres.2012.09.029
PMCID: PMC3543492  PMID: 23040794
impulse control; tic disorders; prenatal maternal smoking; rage; co-morbidity
20.  Principal expression of two mRNA isoforms (ABCB5α and ABCB5β) of the ATP-binding cassette transporter gene ABCB5 in melanoma cells and melanocytes 
Summary
ATP-binding cassette (ABC) transporters play a pivotal role in physiology and pathology. We identified and cloned two novel mRNA isoforms (ABCB5α and ABCB5β) of the ABC transporter ABCB5 in human melanoma cells. The deduced ABCB5α protein appears to be an altered splice variant containing only a putative ABC, whereas the ABCB5β isoform shares approximately 70% similarity with ABCB1 (MDR1) and has a deduced topological arrangement similar to that of the whole carboxyl terminal half of the ABCB1 gene product, P-glycoprotein, including an intact ABC. Northern blot, real-time PCR, and conventional RT-PCR were used to verify the expression profiles of ABCB5α/β. We found that the melanomas included among the NCI-60 panel of cell lines preferentially expressed both ABCB5α and ABCB5β. However, ABCB5α/β expression was undetectable in two amelanotic melanomas (M14 and LOX-IMVI). The expression profile of ABCB5α/β in all of the other melanomas of the panel was confirmed both by RT-PCR and by sequencing. Neither ABCB5α nor ABCB5β expression was found in normal tissues such as liver, spleen, thymus, kidney, lung, colon, small intestines or placenta. ABCB5α/β mRNAs were also expressed in normal melanocytes and in retinal pigment epithelial cells, suggesting that ABCB5α/β expression is pigment cell-specific and might be involved in melanogenesis. Our findings indicate that expression of ABCB5α/β might possibly provide two novel molecular markers for differential diagnosis of melanomas and constitute potential molecular targets for therapy of melanomas.
doi:10.1111/j.1600-0749.2005.00214.x
PMCID: PMC3915408  PMID: 15760339
ATP-binding cassette transporter; ABCB5; melanoma/melanocytes
21.  A Pilot Study of Qigong for Reducing Cocaine Craving Early in Recovery 
Abstract
Objectives
This pilot study examined the feasibility, preliminary efficacy, and determined the effect sizes of external qigong therapy (EQT) in reducing cue-elicited cocaine craving and associated symptoms among recently abstinent cocaine-dependent (CD) individuals.
Methods
This study randomized 101 CD subjects to either a real EQT (n=51) or sham EQT control (n=50) group. Subjects underwent a baseline assessment and a weekly cue-exposure session for 2 weeks. Total EQT or sham treatments ranged from 4 to 6 sessions in 2 weeks.
Results
EQT-treated subjects displayed a greater reduction in cue-elicited craving (p=0.06) and symptoms of depression (p<0.05) with medium effect sizes.
Conclusions
This study demonstrated the feasibility of delivering EQT among CD individuals early in residential treatment. Future research should include a larger sample and examine the mechanisms and potential longitudinal benefits of EQT.
doi:10.1089/acm.2012.0052
PMCID: PMC3576894  PMID: 22757968
22.  StemCellDB: The Human Pluripotent Stem Cell Database at the National Institutes of Health 
Stem cell research  2012;10(1):57-66.
Much of the excitement generated by induced pluripotent stem cell technology is concerned with the possibility of disease modeling as well as the potential for personalized cell therapy. However, to pursue this it is important to understand the ‘normal’ pluripotent state including its inherent variability. We have performed various molecular profiling assays for 21 hESC lines and 8 hiPSC lines to generate a comprehensive snapshot of the undifferentiated state of pluripotent stem cells. Analysis of the gene expression data revealed no iPSC-specific gene expression pattern in accordance with previous reports. We further compared cells, differentiated as embryoid bodies in 2 media proposed to initiate differentiation towards separate cell fates, as well as 20 adult tissues. From this analysis we have generated a gene list which defines pluripotency and establishes a baseline for the pluripotent state. Finally, we provide lists of genes enriched under both differentiation conditions which show the proposed bias toward independent cell fates.
doi:10.1016/j.scr.2012.09.002
PMCID: PMC3590800  PMID: 23117585
23.  Human Neural Stem Cells Induce Functional Myelination in Mice with Severe Dysmyelination 
Science translational medicine  2012;4(155):10.1126/scitranslmed.3004371.
Shiverer-immunodeficient (Shi-id) mice demonstrate defective myelination in the central nervous system (CNS) and significant ataxia by 2 to 3 weeks of life. Expanded, banked human neural stem cells (HuCNS-SCs) were transplanted into three sites in the brains of neonatal or juvenile Shi-id mice, which were asymptomatic or showed advanced hypomyelination, respectively. In both groups of mice, HuCNS-SCs engrafted and underwent preferential differentiation into oligodendrocytes. These oligodendrocytes generated compact myelin with normalized nodal organization, ultrastructure, and axon conduction velocities. Myelination was equivalent in neonatal and juvenile mice by quantitative histopathology and high-field ex vivo magnetic resonance imaging, which, through fractional anisotropy, revealed CNS myelination 5 to 7 weeks after HuCNS-SC transplantation. Transplanted HuCNS-SCs generated functional myelin in the CNS, even in animals with severe symptomatic hypomyelination, suggesting that this strategy may be useful for treating dysmyelinating diseases.
doi:10.1126/scitranslmed.3004371
PMCID: PMC3864816  PMID: 23052293
24.  Prenatal Cerebral Ischemia Disrupts MRI-Defined Cortical Microstructure Through Disturbances in Neuronal Arborization 
Science translational medicine  2013;5(168):10.1126/scitranslmed.3004669.
Children who survive preterm birth exhibit persistent unexplained disturbances in cerebral cortical growth with associated cognitive and learning disabilities. The mechanisms underlying these deficits remain elusive. We used ex vivo diffusion magnetic resonance imaging to demonstrate in a preterm large-animal model that cerebral ischemia impairs cortical growth and the normal maturational decline in cortical fractional anisotropy (FA). Analysis of pyramidal neurons revealed that cortical deficits were associated with impaired expansion of the dendritic arbor and reduced synaptic density. Together, these findings suggest a link between abnormal cortical FA and disturbances of neuronal morphological development. To experimentally investigate this possibility, we measured the orientation distribution of dendritic branches and observed that it corresponds with the theoretically predicted pattern of increased anisotropy within cases that exhibited elevated cortical FA after ischemia. We conclude that cortical growth impairments are associated with diffuse disturbances in the dendritic arbor and synapse formation of cortical neurons, which may underlie the cognitive and learning disabilities in survivors of preterm birth. Further, measurement of cortical FA may be useful for noninvasively detecting neurological disorders affecting cortical development.
doi:10.1126/scitranslmed.3004669
PMCID: PMC3857141  PMID: 23325800
25.  Lipid-insertion enables targeting functionalization of erythrocyte membrane-cloaked nanoparticles† 
Nanoscale  2013;5(19):10.1039/c3nr03064d.
RBC membrane-cloaked polymeric nanoparticles represent an emerging nanocarrier platform with extended circulation in vivo. A lipid-insertion method is employed to functionalize these nanoparticles without the need for direct chemical conjugation. Insertion of both folate and the nucleolin-targeting aptamer AS1411 shows receptor-specific targeting against model cancer cell lines.
doi:10.1039/c3nr03064d
PMCID: PMC3831007  PMID: 23907698

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