Medicine and Health Sciences Commons^™

Imaging Activity Of Neuronal Populations With New Long-Wavelength Voltage-Sensitive Dyes, Joseph P. Wuskell, Leslie M. Loew

UCHC Articles - Research

We have assessed the utility of five new long-wavelength fluorescent voltage-sensitive dyes (VSD) for imaging the activity of populations of neurons in mouse brain slices. Although all the five were capable of detecting activity resulting from activation of the Schaffer collateral-CA1 pyramidal cell synapse, they differed significantly in their properties, most notably in the signal-to-noise ratio of the changes in dye fluorescence associated with neuronal activity. Two of these dyes, Di-2-ANBDQPQ and Di-1-APEFEQPQ, should prove particularly useful for imaging activity in brain tissue and for combining VSD imaging with the control of neuronal activity via light-activated proteins such as channelrhodopsin-2 …

Dna Polymorphisms And Response To Treatment In Patients With Chronic Hepatitis C: Results From The Halt-C Trial, Richard W. Lambrecht, Herbert L. Bonkovsky

UCHC Articles - Research

Background/Aim

Certain host genetic polymorphisms reportedly affect the likelihood of a sustained virological response (SVR) to interferon treatment in subjects infected with hepatitis C virus (HCV). As part of the HALT-C trial we evaluated genetic associations among patients infected with HCV genotype 1 who had failed previous interferon treatment.

Methods

SVR was determined 24 weeks after completing treatment with pegylated interferon alfa-2a and ribavirin. Eight single nucleotide polymorphisms (SNPs) were selected on the basis of previously reported associations with treatment response. Genotypes were assessed by polymerase chain reaction-based assays. The percentage of patients who achieved SVR was determined for each …

Use Of An Alpha-Smooth Muscle Actin (Smaa) Gfp Reporter To Identify An Osteoprogenitor Population, Zana Kalajzic, Haitao Li, Li-Ping Wang, Xi Jiang, Katie B. Lamothe, Douglas J. Adams, Hector L. Aguila, David W. Rowe, Ivo Kalajzic

UCHC Articles - Research

Identification of a reliable marker of skeletal precursor cells within calcified and soft tissues remains a major challenge for the field. To address this, we used a transgenic model in which osteoblasts can be eliminated by pharmacological treatment. Following osteoblast ablation a dramatic increase in a population of α-smooth muscle actin (α-SMA) positive cells was observed. During early recovery phase from ablation we have detected cells with the simultaneous expression of SMAA and a preosteoblastic 3.6GFP marker, indicating the potential for transition of α-SMA⁺ cells towards osteoprogenitor lineage. Utilizing α-SMAGFP transgene, α-SMAGFP⁺ positive cells were detected in the …

Teriparatide (1-34 Human Pth) Regulation Of Osterix During Fracture Repair, Do Y. Soung, Graziello Geneau, Hicham Drissi

UCHC Articles - Research

Based on remarkable success of PTH as an anabolic drug for osteoporosis, case reports of off-label use of teriparatide (1-34 PTH) in patients with complicated fractures and non-unions are emerging. We investigated the mechanisms underlying PTH accelerated fracture repair. Bone marrow cells from 7 days 40cg/kg of teriparatide treated or saline control mice were cultured and Osx and osteoblast phenotypic gene expression assessed by real time RT-PCR in these cells. Fractured animals injected daily with either saline or 40cg/kg of teriparatide for up to 21 days were X-rayed and histological assessment performed, as well as immunohistochemical analyses of the Osx …

Amino(Oligo)Thiophene-Based Environmentally Sensitive Biomembrane Chromophores, Ping Yan, Aifang Xie, Meide Wei, Leslie M. Loew

UCHC Articles - Research

There is a growing need for cellular imaging with fluorescent probes that emit at longer wavelengths to minimize the effects of absorption, autofluorescence and scattering from biological tissue. In this paper a series of new environmentally-sensitive hemicyanine dyes featuring amino(oligo)thiophene donors have been synthesized via aldol condensation between a 4-methylpyridinium salt and various amino(oligo)thiophene carboxaldehydes, which were, in turn, obtained from amination of bromo (oligo)thiophene carboxaldehyde. Side chains on these fluorophores impart a strong affinity for biological membranes. Compared with benzene analogues, these thiophene fluorophores show significant red shift in the absorption and emission spectra, offering compact red and near-infrared …

A Crucial Role In Cell Spreading For The Interaction Of Abl Pxxp Motifs With Crk And Nck Adaptors, Susumu Antoku, Gonzalo M. Rivera, Bruce J. Mayer

UCHC Articles - Research

The dynamic reorganization of actin structures helps to mediate the interaction of cells with their environment. The Abl non-receptor tyrosine kinase can modulate actin rearrangement during cell attachment. Here we report that the Abl PxxP motifs, which bind Src homology 3 (SH3) domains, are indispensable for the coordinated regulation of filopodium and focal adhesion formation and cell-spreading dynamics during attachment. Candidate Abl PxxP-motif-binding partners were identified by screening a comprehensive SH3-domain phage-display library. A combination of protein overexpression, silencing, pharmacological manipulation and mutational analysis demonstrated that the PxxP motifs of Abl exert their effects on actin organization by two distinct …

Microsome Biocolloids For Rapid Drug Metabolism And Inhibition Assessment By Lc-Ms, Besnik Bajrami, Sadagopan Krishnan, James F. Rusling

UCHC Articles - Research

Rat liver microsomes attached to nanoparticles were used for LC-MS studies of CYP3A and 2E1 enzymes in metabolism of N-nitroso compounds. Using these biocolloids, turnover rates were measured within 2 min. Inhibitor IC₅₀ values for ketoconazole (KET) and 4-methylpyrazole (4-MEP) were estimated.

Osteopenia In Transgenic Mice With Osteoblast-Targeted Expression Of The Inducible Camp Early Repressor, Taranpreet K. Chandhoke, Yu-Feng Huang, Fei Liu, Gloria A. Gronowicz, Douglas J. Adams, John R. Harrison, Barbara E. Kream

UCHC Articles - Research

ICER is a member of the CREM family of basic leucine zipper transcription factors that acts as a dominant negative regulator of gene transcription. Four different isoforms of ICER (I, Iγ, II and IIγ) are transcribed from the P2 promoter of the Crem gene. We previously found that each of the ICER isoforms is induced by parathyroid hormone in osteoblasts. The goal of the present study was to assess the function of ICER in bone by overexpressing ICER in osteoblasts of transgenic mice. ICER I and ICER II cDNAs, each containing an N-terminal FLAG epitope tag, were cloned downstream of …

Understanding How Lipopolysaccharide Impacts Cd4 T Cell Immunity, Jeremy P. Mcaleer, Anthony T. Vella

UCHC Articles - Research

Lipopolysaccharide (LPS) is a natural adjuvant synthesized by gram-negative bacteria that has profound effects on CD4 T cell responses. LPS stimulates cells through Toll-like receptor 4 (TLR4), causing the release of inflammatory cytokines and upregulation of costimulatory molecules on antigen presenting cells. The combination of signals from antigen, costimulation, and cytokines allow CD4 T cells to overcome suppressive barriers and accumulate in large numbers. T cells that are primed in an LPS-stimulated environment are programmed for long-term survival following clonal expansion. LPS is well-known for generating Th1 responses, however, under appropriate conditions it can also support differentiation into other T …

Synergistic Metabolic Toxicity Screening Using Microsome/Dna Electrochemiluminescent Arrays And Nanoreactors, Sadagopan Krishnan, Eli G. Hvastkovs, Besnik Bajrami, Dharamainder Choudhary, John B. Schenkman, James F. Rusling

UCHC Articles - Research

Platforms based on thin enzyme/DNA films were used in two-tier screening of chemicals for reactive metabolites capable of producing toxicity. Microsomes were used for the first time as sources of cytochrome (cyt) P450 enzymes in these devices. Initial rapid screening involved electrochemiluminescent (ECL) arrays featuring spots containing ruthenium poly(vinylpyridine), DNA, and rat liver microsomes or bicistronically expressed human cyt P450 2E1 (h2E1). Cyt P450 enzymes were activated via the NADPH/reductase cycle. When bioactivation of substrates in the films gives reactive metabolites, they are trapped by covalent attachment to DNA bases. The rate of increase in ECL with enzyme reaction time …

Prevalence And Correlates Of Shoplifting In The United States: Results From The National Epidemiologic Survey On Alcohol And Related Conditions (Nesarc), Nancy M. Petry

UCHC Articles - Research

Objective

This study presented nationally representative data on the lifetime prevalence, correlates, and comorbidity of shoplifting among adults in the United States.

Method

Data were derived from a large national sample of the United States population. Face-to-face surveys of more than 43,000 adults ages 18 years and older residing in households were conducted during the 2001–2002 period. Diagnoses of mood, anxiety, and drug disorders as well as personality disorders were based on the Alcohol Use Disorder and Associated Disabilities Interview Schedule—DSM-IV Version.

Results

The prevalence of lifetime shoplifting in the U.S. population was 11.3%. Associations between shoplifting and all antisocial …

Subcutaneous Late Phase Responses Are Augmented During Local Inhalational Tolerance In A Murine Asthma Model, Anurag Singh, Roger S. Thrall, Linda A. Guernsey, William F. Carson Iv, Eric R. Secor Jr, Robert E. Cone, Thiruchandurai V. Rajan, Craig M. Schramm

UCHC Articles - Research

Acute exposure of sensitized mice to antigen elicits allergic airway disease (AAD) characterized by Th2 cytokine-dependent pulmonary eosinophilia, methacholine hyperresponsiveness and antigen-specific IgE elevation. However, chronic exposure induces a local inhalational tolerance (LIT), with resolution of the airway responses but persistent systemic IgE production. To further determine if systemic immunologic responses were maintained during LIT, we assessed subcutaneous late phase responses to ovalbumin in this model. Sensitized and AAD mice developed small subcutaneous responses to ovalbumin, with footpad thickness increasing to 113.7 and 113.6% of baseline, respectively. In comparison, LIT mice developed marked foot swelling (141.6%). Histologic examination confirmed increased …

Molecular Biology: Power Sequencing, Brenton R. Graveley

UCHC Articles - Research

Advances in DNA-sequencing technology provide unprecedented insight into the entire collection of four genomes' transcribed sequences; they herald a new era in the study of gene regulation and genome function.

Social-Emotional Screening Status In Early Childhood Predicts Elementary School Outcomes, Margaret J. Briggs-Gowan

UCHC Articles - Research

OBJECTIVE—The goal was to examine whether children who screen positive for socialemotional/ behavioral problems at 12 to 36 months of age are at elevated risk for social-emotional/ behavioral problems in early elementary school.
METHODS—The sample studied (N = 1004) comprised an ethnically (33.3% minority) and socioeconomically (17.8% living in poverty and 11.3% living in borderline poverty) diverse, healthy, birth cohort from a metropolitan region of the northeastern United States. When children were 12 to 36 months of age (mean age: 23.8 months; SD: 7.1 months), parents completed the Brief Infant-Toddler Social and Emotional Assessment and questions concerning their level of …

Expression And Function Of Dlx Genes In The Osteoblast Lineage, Haitao Li, Inga Marijanovic, Mark S. Kronenberg, Ivana Erceg, Mary Louise Stover, Dimitrios Velonis, Mina Mina, William B. Upholt, Ivo Kalajzic, Alexander C. Lichtler

UCHC Articles - Research

Our laboratory and others have shown that overexpression of Dlx5 stimulates osteoblast differentiation. Dlx5^−/−/Dlx6^−/− mice have more severe craniofacial and limb defects than Dlx5^−/−, some of which are potentially due to defects in osteoblast maturation. We wished to investigate the degree to which other Dlx genes compensate for the lack of Dlx5, thus allowing normal development of the majority of skeletal elements in Dlx5^−/− mice. Dlx gene expression in cells from different stages of the osteoblast lineage isolated by FACS sorting showed that Dlx2, Dlx5 and Dlx6 are expressed most strongly in less mature …

Roles Of Fgfr3 During Morphogenesis Of Meckel's Cartilage And Mandibular Bones, Bruce A. Havens, Dimitris Velonis, Mark S. Kronenberg, Alex C. Lichtler, Bonnie Oliver, Mina Mina

UCHC Articles - Research

To address the functions of FGFR2 and FGFR3 signaling during mandibular skeletogenesis, we over-expressed in the developing chick mandible, replication-competent retroviruses carrying truncated FGFR2c or FGFR3c that function as dominant negative receptors (RCAS-dnFGFR2 and RCAS-dnFGFR3). Injection of RCAS-dnFGFR3 between HH15−20 led to reduced proliferation, increased apoptosis, and decreased differentiation of chondroblasts in Meckel's cartilage. These changes resulted in the formation of a hypoplastic mandibular process and truncated Meckel's cartilage. This treatment also affected the proliferation and survival of osteoprogenitor cells in osteogenic condensations, leading to the absence of five mandibular bones on the injected side. Injection of RCAS-dnFGFR2 between HH15−20 …

The Effects Of Hiv On P300 Are Moderated By Familial Risk For Substance Dependence: Implications For A Theory Of Brain Reserve, Lance O. Bauer

UCHC Articles - Research

Abstract

Background

The goal of the study was to test the validity of additive versus synergistic versus underadditive versions of brain reserve theory within the context of HIV/AIDS. In addition, it tested the convergent validity of 2 operational definitions of premorbid reserve: verbal IQ (VIQ) and a family history (FH) of substance abuse or dependence.

Methods

Seventy HIV-1 seronegative and 115 HIV-1 seropositive male and female volunteers were assigned to 4 subgroups defined by the crossing of a VIQ score < versus ≥ 90 with the presence versus absence of a paternal history of alcohol, cocaine, or opiate abuse or dependence. The principal dependent measure was the P300 event related brain potential elicited during the Stroop color-word interference task.

Results

The principal finding was an underadditive effect of FH plus HIV/AIDS on P300 area over the frontal region: FH …

Accurate Dna Fragment Sizing By Capillary Electrophoresis With Laser-Induced Fluorescence Array For Detection Of Sequence Specificity Of Dna Damage, Erwin V. Fundador, Dharamainder Choudhary, John B. Schenkman, James F. Rusling

UCHC Articles - Research

Cancer has been linked to mutations within specific codons in genes that code for critical biomolecules such as tumor suppressor proteins (e.g., p53). Activated metabolites like benzo[a]pyrenediol epoxide act on preferred nucleotide sequences of DNA, and such mutations have been identified in cancers. DNA reaction site identification depends on accurate analysis of oligonucleotide fragment sizes produced by strand breakage at the damaged sites. Herein, we report a new method for DNA fragment sizing using capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). Absolute sizing accuracy and speed are achieved by utilizing a CE-LIF array with two-color fluorescence detection. Accuracy …

Heat-Shock Protein 90 Associates With N-Terminal Extended Peptides And Is Required For Direct And Indirect Antigen Presentation, Margaret K. Callahan, Manish Garg, Pramod K. Srivastava

UCHC Articles - Research

CD8⁺ T cells recognize peptide fragments of endogenously synthesized antigens of cancers or viruses, presented by MHC I molecules. Such antigen presentation requires the generation of peptides in the cytosol, their passage to the endoplasmic reticulum, loading of MHC I with peptides, and transport of MHC I–peptide complexes to the cell surface. Heat-shock protein (hsp) 90 is a cytosolic chaperone known to associate with peptide and peptide precursors of MHC I epitopes. We report here that treatment of cells with hsp90 inhibitors leads to generation of “empty” MHC I caused by inhibited loading of MHC I with peptides. Inhibition …

Fas And Cfl Forms Of Verbal Fluency Differ In Difficulty: A Meta-Analytic Study, Danielle Barry

UCHC Articles - Research

The Controlled Oral Word Association (COWA) Test is a brief and sensitive measure of executive cognitive dysfunction. There are two commonly used forms of the test, one using the letters F, A, and S, and the other using C, F, and L. This study examines the relative difficulty of the two forms using a meta-analytic approach that includes multiple samples of normal individuals. The effects of age, education, gender composition, exclusion criteria, and age of study are also examined. Results indicate that the CFL form of the test is more difficult and that age, education, and the use of strict …

Biochemical Applications Of Ultrathin Films Of Enzymes, Polyions And Dna, James F. Rusling, Dominic O. Hull, Eli G. Hvastkovs, John B. Schenkman

UCHC Articles - Research

This feature article summarizes recent applications of ultrathin films of enzymes and DNA assembled layer-by-layer (LbL). Using examples mainly from our own research, we focus on systems developed for biocatalysis and biosensors for toxicity screening. Enzyme–poly(l-lysine) (PLL) films, especially when stabilized by crosslinking, can be used for biocatalysis at unprecedented high temperatures or in acidic or basic solutions on electrodes or sub-micron sized beads. Such films have bright prospects for chiral synthesis and biofuel cells. Excellent bioactivity and retention of enzyme structure in these films facilitates their use in detailed kinetic studies. Biosensors and arrays employing DNA–enzyme films show great …

Management Of Oral Mucositis In Patients With Cancer, Rajesh V. Lalla, Douglas E. Peterson

UCHC Articles - Research

Oral mucositis refers to erythematous and ulcerative lesions of the oral mucosa observed in patients with cancer being treated with chemotherapy, and/or with radiation therapy to fields involving the oral cavity. Lesions of oral mucositis are often very painful and compromise nutrition and oral hygiene as well as increase risk for local and systemic infection. Mucositis can also involve other areas of the alimentary tract; for example, gastrointestinal (GI) mucositis can manifest as diarrhea. Thus, mucositis is a highly significant and sometimes dose-limiting complication of cancer therapy.

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The Validity Of Cocaine Dependence Subtypes, Henry R. Kranzler, Victor M. Hesselbrock

UCHC Articles - Research

Cocaine dependence (CD) is a multifactorial disorder, variable in its manifestations, and heritable. We examined the concurrent validity of homogeneous subgroups of CD as phenotypes for genetic analysis. We applied data reduction methods and an empirical cluster-analytic approach to measures of cocaine use, cocaine-related effects, and cocaine treatment history in 1393 subjects, from 660 small nuclear families. Four of the six clusters that were derived yielded heritability estimates in excess of 0.3. Linkage analysis showed genomewide significant results for two of the clusters. Here we examine the concurrent validity of the six clusters using a variety of demographic and substance-related …

Phagocytosis Of Borrelia Burgdorferi, The Lyme Disease Spirochete, Potentiates Innate Immune Activation And Induces Apoptosis In Human Monocytes, Adriana R. Cruz, Meagan W. Moore, Carson J. La Vake, Christian H. Eggers, Juan C. Salazar, Justin D. Radolf

UCHC Articles - Research

We have previously demonstrated that phagocytosed Borrelia burgdorferi induces activation programs in human peripheral blood mononuclear cells that differ qualitatively and quantitatively from those evoked by equivalent lipoprotein-rich lysates. Here we report that ingested B. burgdorferi induces significantly greater transcription of proinflammatory cytokine genes than do lysates and that live B. burgdorferi, but not B. burgdorferi lysate, is avidly internalized by monocytes, where the bacteria are completely degraded within phagolysosomes. In the course of these experiments, we discovered that live B. burgdorferi also induced a dose-dependent decrease in monocytes but not a decrease in dendritic cells or T cells …