Raman spectroscopy: the gateway into tomorrow's virology

In the molecular world, researchers act as detectives working hard to unravel the mysteries surrounding cells. One of the researchers' greatest tools in this endeavor has been Raman spectroscopy. Raman spectroscopy is a spectroscopic technique that measures the unique Raman spectra for every type of biological molecule. As such, Raman spectroscopy has the potential to provide scientists with a library of spectra that can be used to unravel the makeup of an unknown molecule. However, this technique is limited in that it is not able to manipulate particular structures without disturbing their unique environment. Recently, a novel technology that combines Raman spectroscopy with optical tweezers, termed Raman tweezers, evades this problem due to its ability to manipulate a sample without physical contact. As such, Raman tweezers has the potential to become an incredibly effective diagnostic tool for differentially distinguishing tissue, and therefore holds great promise in the field of virology for distinguishing between various virally infected cells. This review provides an introduction for a virologist into the world of spectroscopy and explores many of the potential applications of Raman tweezers in virology. Originally published in Virology Journal, 2006 Vol. 3, No. 51

Examining the reversibility of long-term behavioral disruptions in progeny of maternal SSRI exposure

Serotonergic dysregulation is implicated in numerous psychiatric disorders. Serotonin plays widespread trophic roles during neurodevelopment; thus perturbations to this system during development may increase risk for neurodevelopmental disorders. Epidemiological studies have examined association between selective serotonin reuptake inhibitor (SSRI) treatment during pregnancy and increased autism spectrum disorder (ASD) risk in offspring. It is unclear from these studies whether ASD susceptibility is purely related to maternal psychiatric diagnosis, or if treatment poses additional risk. We sought to determine whether maternal SSRI treatment alone or in combination with genetically vulnerable background was sufficient to induce offspring behavior disruptions relevant to ASD. We exposed C57BL/6J or Celf6(+/-) mouse dams to fluoxetine (FLX) during different periods of gestation and lactation and characterized offspring on tasks assessing social communicative interaction and repetitive behavior patterns including sensory sensitivities. We demonstrate robust reductions in pup ultrasonic vocalizations (USVs) and alterations in social hierarchy behaviors, as well as perseverative behaviors and tactile hypersensitivity. Celf6 mutant mice demonstrate social communicative deficits and perseverative behaviors, without further interaction with FLX. FLX re-exposure in adulthood ameliorates the tactile hypersensitivity yet exacerbates the dominance phenotype. This suggests acute deficiencies in serotonin levels likely underlie the abnormal responses to sensory stimuli, while the social alterations are instead due to altered development of social circuits. These findings indicate maternal FLX treatment, independent of maternal stress, can induce behavioral disruptions in mammalian offspring, thus contributing to our understanding of the developmental role of the serotonin system and the possible risks to offspring of SSRI treatment during pregnancy

Subcellular fractionation method to study endosomal trafficking of Kaposi’s sarcoma-associated herpesvirus

Background Virus entry involves multiple steps and is a highly orchestrated process on which successful infection collectively depends. Entry processes are commonly analyzed by monitoring internalized virus particles via Western blotting, polymerase chain reaction, and imaging techniques that allow scientist to track the intracellular location of the pathogen. Such studies have provided abundant direct evidence on how viruses interact with receptor molecules on the cell surface, induce cell signaling at the point of initial contact with the cell to facilitate internalization, and exploit existing endocytic mechanisms of the cell for their ultimate infectious agenda. However, there is dearth of knowledge in regards to trafficking of a virus via endosomes. Herein, we describe an optimized laboratory procedure to isolate individual organelles during different stages of endocytosis by performing subcellular fractionation. This methodology is established using Kaposi’s sarcoma-associated herpesvirus (KSHV) infection of human foreskin fibroblast (HFF) cells as a model. With KSHV and other herpesviruses alike, envelope glycoproteins have been widely reported to physically engage target cell surface receptors, such as integrins, in interactions leading to entry and subsequent infection. Results Subcellular fractionation was used to isolate early and late endosomes (EEs and LEs) by performing a series of centrifugations steps. Specifically, a centrifugation step post-homogenization was utilized to obtain the post-nuclear supernatant containing intact intracellular organelles in suspension. Successive fractionation via sucrose density gradient centrifugation was performed to isolate specific organelles including EEs and LEs. Intracellular KSHV trafficking was directly traced in the isolated endosomal fractions. Additionally, the subcellular fractionation approach demonstrates a key role for integrins in the endosomal trafficking of KSHV. The results obtained from fractionation studies corroborated those obtained by traditional imaging studies. Conclusions This study is the first of its kind to employ a sucrose flotation gradient assay to map intracellular KSHV trafficking in HFF cells. We are confident that such an approach will serve as a powerful tool to directly study intracellular trafficking of a virus, signaling events occurring on endosomal membranes, and dynamics of molecular events within endosomes that are crucial for uncoating and virus escape into the cytosol

Tuning supersymmetric models at the LHC: A comparative analysis at two-loop level

We provide a comparative study of the fine tuning amount (Delta) at the two-loop leading log level in supersymmetric models commonly used in SUSY searches at the LHC. These are the constrained MSSM (CMSSM), non-universal Higgs masses models (NUHM1, NUHM2), non-universal gaugino masses model (NUGM) and GUT related gaugino masses models (NUGMd). Two definitions of the fine tuning are used, the first (Delta_{max}) measures maximal fine-tuning wrt individual parameters while the second (Delta_q) adds their contribution in "quadrature". As a direct result of two theoretical constraints (the EW minimum conditions), fine tuning (Delta_q) emerges as a suppressing factor (effective prior) of the averaged likelihood (under the priors), under the integral of the global probability of measuring the data (Bayesian evidence p(D)). For each model, there is little difference between Delta_q, Delta_{max} in the region allowed by the data, with similar behaviour as functions of the Higgs, gluino, stop mass or SUSY scale (m_{susy}=(m_{\tilde t_1} m_{\tilde t_2})^{1/2}) or dark matter and g-2 constraints. The analysis has the advantage that by replacing any of these mass scales or constraints by their latest bounds one easily infers for each model the value of Delta_q, Delta_{max} or vice versa. For all models, minimal fine tuning is achieved for M_{higgs} near 115 GeV with a Delta_q\approx Delta_{max}\approx 10 to 100 depending on the model, and in the CMSSM this is actually a global minimum. Due to a strong ($\approx$ exponential) dependence of Delta on M_{higgs}, for a Higgs mass near 125 GeV, the above values of Delta_q\approx Delta_{max} increase to between 500 and 1000. Possible corrections to these values are briefly discussed.Comment: 23 pages, 46 figures; references added; some clarifications (section 2

Raman tweezers provide the fingerprint of cells supporting the late stages of KSHV reactivation

Kaposi's sarcoma-associated herpesvirus (KSHV) has both latent and lytic phases of replication. The molecular switch that triggers a reactivation is still unclear. Cells from S phase of cell cycle provide apt conditions for an active reactivation. In order to specifically delineate the Raman spectra of cells supporting KSHV reactivation, we followed a novel approach where cells were sorted based on the state of infection (latent Vs lytic) by a flow cytometer and then analyzed by the Raman tweezers. The Raman bands at 785, 813, 830, 1095, and 1128 cm-1 are specifically altered in cells supporting KSHV reactivation. These 5 peaks make up the Raman fingerprint of cells supporting KSHV reactivation. The physiological relevance of the changes in these peaks with respect to KSHV reactivation is discussed in the following report. Originally published Journal of Cellular and Molecular Medicine, Vol. 13, No. 8b, Aug 200

Properties of 125 GeV Higgs boson in non-decoupling MSSM scenarios

Tantalizing hints of the Higgs boson of mass around 125 GeV have been reported at the LHC. We explore the MSSM parameter space in which the 125 GeV state is identified as the heavier of the CP even Higgs bosons, and study two scenarios where the two photon production rate can be significantly larger than the standard model (SM). In one scenario, $\Gamma(H\to \gamma\gamma)$ is enhanced by a light stau contribution, while the $WW^{\ast}$ ($ZZ^{\ast}$) rate stays around the SM rate. In the other scenario, $\Gamma(H\to b\bar{b})$ is suppressed and not only the $\gamma\gamma$ but also the $WW^{\ast}$ ($ZZ^{\ast}$) rates should be enhanced. The $\tau\bar{\tau}$ rate can be significantly larger or smaller than the SM rate in both scenarios. Other common features of the scenarios include top quark decays into charged Higgs boson, single and pair production of all Higgs bosons in $e^+e^-$ collisions at $\sqrt{s}\lesssim 300$ GeV.Comment: 20 pages, 5 figures, accepted version for publication in JHE

BBMS + + – basic bioinformatics meta-searcher

In this paper we present a Basic Bioinformatics Meta-searcher (BBMS), a web-based service aiming to simplify and integrate biological data searching through selected biological databases. BBMS facilitates biological data searching enabling multiple sources transparently, increasing research productivity as it avoids time consuming learning and parameterization of different search engines. As a complementary service, BBMS provides insight and links to common online bioinformatics tools. Users’ feedback when evaluating BBMS in terms of usability, usefulness and efficiency was very positive

The Neurovascular Relation in Oxygen-induced Retinopathy

Purpose: Longitudinal studies in rat models of retinopathy of prematurity (ROP) have demonstrated that abnormalities of retinal vasculature and function change hand-in-hand. In the developing retina, vascular and neural structures are under cooperative molecular control. In this study of rats with oxygen-induced retinopathy (OIR) models of ROP, mRNA expression of vascular endothelial growth factor (VEGF), semaphorin (Sema), and their neuropilin receptor (NRP) were examined during the course of retinopathy to evaluate their roles in the observed neurovascular congruency. Methods: Oxygen exposures designed to induce retinopathy were delivered to Sprague-Dawley rat pups (n=36) from postnatal day (P) 0 to P14 or from P7 to P14. Room-air-reared controls (n=18) were also studied. Sensitivities of the rod photoreceptors ($S_{rod}$) and the postreceptor cells (Sm) were derived from electroretinographic (ERG) records. Arteriolar tortuosity, $T_A$, was derived from digital fundus images using Retinal Image multi-Scale Analysis (RISA) image analysis software. mRNA expression of $VEGF_{164}$, semaphorin IIIA (Sema3A), and neuropilin-1 (NRP-1) was evaluated by RT–PCR of retinal extracts. Tests were performed at P15–P16, P18–P19, and P25–P26. Relations among ERG, RISA, and PCR parameters were evaluated using linear regression on log transformed data. Results: Sm was low and $T_A$ was high at young ages, then both resolved by P25–P26. $VEGF_{164}$ and Sema3A mRNA expression were also elevated early and decreased with age. Low Sm was significantly associated with high $VEGF_{164}$ and Sema3A expression. Low Srod was also significantly associated with high VEGF164. $S_{rod}$ and Sm were both correlated with $T_A$. NRP-1 expression was little affected by OIR. Conclusions: The postreceptor retina appears to mediate the vascular abnormalities that characterize OIR. Because of the relationships revealed by these data, early treatment that targets the neural retina may mitigate the effects of ROP

Many faces of low mass neutralino dark matter in the unconstrained MSSM, LHC data and new signals

If all strongly interacting sparticles (the squarks and the gluinos) in an unconstrained minimal supersymmetric standard model (MSSM) are heavier than the corresponding mass lower limits in the minimal supergravity (mSUGRA) model, obtained by the current LHC experiments, then the existing data allow a variety of electroweak (EW) sectors with light sparticles yielding dark matter (DM) relic density allowed by the WMAP data. Some of the sparticles may lie just above the existing lower bounds from LEP and lead to many novel DM producing mechanisms not common in mSUGRA. This is illustrated by revisiting the above squark-gluino mass limits obtained by the ATLAS Collaboration, with an unconstrained EW sector with masses not correlated with the strong sector. Using their selection criteria and the corresponding cross section limits, we find at the generator level using Pythia, that the changes in the mass limits, if any, are by at most 10-12% in most scenarios. In some cases, however, the relaxation of the gluino mass limits are larger ($\approx 20$). If a subset of the strongly interacting sparticles in an unconstrained MSSM are within the reach of the LHC, then signals sensitive to the EW sector may be obtained. This is illustrated by simulating the $blj$\etslash, $l= e and \mu$, and $b\tau j$\etslash signals in i) the light stop scenario and ii) the light stop-gluino scenario with various light EW sectors allowed by the WMAP data. Some of the more general models may be realized with non-universal scalar and gaugino masses.Comment: 27 pages, 1 figure, references added, minor changes in text, to appear in JHE

Interpreting a 1 fb^-1 ATLAS Search in the Minimal Anomaly Mediated Supersymmetry Breaking Model

Recent LHC data significantly extend the exclusion limits for supersymmetric particles, particularly in the jets plus missing transverse momentum channels. The most recent such data have so far been interpreted by the experiment in only two different supersymmetry breaking models: the constrained minimal supersymmetric standard model (CMSSM) and a simplified model with only squarks and gluinos and massless neutralinos. We compare kinematical distributions of supersymmetric signal events predicted by the CMSSM and anomaly mediated supersymmetry breaking (mAMSB) before calculating exclusion limits in mAMSB. We obtain a lower limit of 900 GeV on squark and gluino masses at the 95% confidence level for the equal mass limit, tan(beta)=10 and mu>0.Comment: 18 pages, 11 figure