Inhibition of mTORC1 inhibits lytic replication of Epstein-Barr virus in a cell-type specific manner

BACKGROUND: Epstein-Barr virus is a human herpesvirus that infects a majority of the human population. Primary infection of Epstein-Barr virus (EBV) causes the syndrome infectious mononucleosis. This virus is also associated with several cancers, including Burkitt’s lymphoma, post-transplant lymphoproliferative disorder and nasopharyngeal carcinoma. As all herpesvirus family members, EBV initially replicates lytically to produce abundant virus particles, then enters a latent state to remain within the host indefinitely. METHODS: Through a genetic screen in Drosophila, we determined that reduction of Drosophila Tor activity altered EBV immediate-early protein function. To further investigate this finding, we inhibited mTOR in EBV-positive cells and investigated subsequent changes to lytic replication via Western blotting, flow cytometry, and quantitative PCR. The student T-test was used to evaluate significance. RESULTS: mTOR, the human homolog of Drosophila Tor, is an important protein at the center of a major signaling pathway that controls many aspects of cell biology. As the EBV immediate-early genes are responsible for EBV lytic replication, we examined the effect of inhibition of mTORC1 on EBV lytic replication in human EBV-positive cell lines. We determined that treatment of cells with rapamycin, which is an inhibitor of mTORC1 activity, led to a reduction in the ability of B cell lines to undergo lytic replication. In contrast, EBV-positive epithelial cell lines underwent higher levels of lytic replication when treated with rapamycin. CONCLUSIONS: Overall, the responses of EBV-positive cell lines vary when treated with mTOR inhibitors, and this may be important when considering such inhibitors as anti-cancer therapeutic agents

Comparison of Regional Body Composition Estimates Obtained from Dual-energy X-ray Absorptiometry and Single-frequency Bioelectrical Impedance Analysis

The anatomical distribution of fat mass (FM) and lean mass (LM) is significant for health and athletic performance. Dual-energy x-ray absorptiometry (DXA) is often used for regional body composition analysis but is not portable, often inaccessible, and costly, while single-frequency bioelectrical impedance analysis (SFBIA) is a more affordable and accessible alternative. PURPOSE: The purpose of this analysis was to compare regional body composition estimates obtained via DXA and SFBIA. METHODS: After an overnight food and fluid fast, 102 adults (64 F, 38 M; age: 29.2 ± 13.4 y; BMI: 24.3 ± 3.9 kg/m2; BF%: 24.6 ± 8.3%) underwent assessments via DXA and SBFIA, each of which provided estimates of FM and LM for the whole body, torso, legs, and arms. DXA scans were performed using custom-made foam blocks to enhance accuracy of regional body composition estimates. SFBIA was performed using an 8-lead device with a 12-channel multiplexer. Both DXA and SFBIA were performed in the supine position. DXA was designated as the criterion method, and body composition estimates were compared using paired-samples t-tests using a Bonferroni-corrected significance level of p ≤ 0.00625. Additional evaluations were conducted using the correlation coefficient (r), constant error (CE), standard error of the estimate (SEE), and total error (TE). RESULTS: Correlations between DXA and SFBIA were high, and the magnitude of errors was generally small: LMTOTAL (r: 0.97; CE: 1.4 kg; SEE: 2.7 kg; TE: 2.9 kg), LMLEGS (r: 0.85; CE: -0.3 kg; SEE: 2.0 kg; TE: 2.1 kg), LMTORSO (r: 0.92; CE: 1.0 kg; SEE: 2.2 kg; TE: 2.5 kg), LMARMS (r: 0.96; CE: 0.6 kg; SEE: 0.6 kg; TE: 0.8 kg), FMTOTAL (r: 0.95; CE: -2.3 kg; SEE: 2.6 kg; TE: 3.5 kg), FMLEGS (r: 0.83; CE: -1.0 kg; SEE: 1.2 kg; TE: 2.0 kg), FMTORSO (r: 0.90; CE: -1.3 kg; SEE: 2.2 kg; TE: 2.6 kg), and FMARMS (r: 0.89; CE: -0.1 kg; SEE: 0.5 kg; TE: 0.5 kg). Despite the relatively small magnitude of differences in FM and LM estimates between DXA and SFBIA, results of paired-samples t-tests indicated that all differences were statistically significant (p \u3c 0.0001), with the exception of LMLEGS (p=0.13) and FMARMS (p=0.11). CONCLUSION: Despite the fact that body composition estimates for most regions exhibited statistically significant differences between DXA and SFBIA, the strong correlations (r: 0.83 to 0.97) and relatively low magnitude of error (CE: -2.3 to 1.4 kg; TE: 0.8 to 3.5 kg) indicate that SFBIA may be an acceptable alternative to DXA when regional body composition is being evaluated and DXA is unavailable. However, additional research is needed to determine the ability of SFBIA to accurately track changes in regional body composition over time. Due to its low cost, portability, and ease of use, the presently examined SFBIA device may represent a useful tool for the evaluation of regional body composition when more advanced methods are unavailable

Validity of Infrared 3-dimensional Scanning for Estimation of Body Composition: A 4-Compartment Model Comparison

Multiple infrared 3-dimensional (3D) scanning technologies exist, including time of flight (ToF) scanners and structured light scanners with static (SL-S) and dynamic (SL-D) configurations. ToF scanners measure depth by using the round-trip time of reflected photons, whereas SL scanners measure deformations in light patterns and allow for creation of a depth image using geometric triangulation. Recently, 3D scanning technologies have been proposed as novel methods of body composition assessment. PURPOSE: The purpose of this analysis was to examine the validity of four different commercially-available 3D scanners for estimation of body fat percentage (BF%) as compared to a 4-compartment (4C) model criterion. METHODS: After an overnight fast, 101 adults (63 F, 38 M; age: 29.3 ± 13.5 y; BMI: 24.3 ± 3.9 kg/m2; BF%: 24.6 ± 8.3%) completed assessments via dual-energy x-ray absorptiometry (DXA), air displacement plethysmography (ADP), bioimpedance spectroscopy (BIS), a standard body mass scale, and four infrared 3D scanners. Two scanners (3DSSL-D1; 3DSSL-D2) utilized structured light scanning with a dynamic configuration, one utilized structured light scanning with a static configuration (3DSSL-S), and one utilized time-of-flight technology (3DSToF). Using the equation of Wang et al. (2002), a criterion 4C estimate of BF% was obtained using DXA for bone mineral, ADP for body volume, scale for body mass, and BIS for total body water. BF% estimates were compared using one-way ANOVA with Bonferroni adjustment for multiple comparisons, and additional evaluations were conducted using the correlation coefficient (r), constant error (CE), standard error of the estimate (SEE), total error (TE), and 95% limits of agreement (LOA). RESULTS: Estimates of BF% did not significantly differ between 4C and any of the 3D scanners. However, metrics of group, individual, and prediction errors varied between scanners: 3DSSL-D1: p=1.0; CE: 0.4%; r: 0.91; SEE: 2.5%; TE: 3.6%; LOA: ±7.0%; 3DSSL-D2: p= 1.0; CE: 0.8%; r: 0.86; SEE: 4.2%; TE: 4.7%; LOA: ±9.2%; 3DSSL-S: p= 1.0; CE: 1.0%; r: 0.81; SEE: 4.0%; TE: 5.0%; LOA: ±9.7%; 3DSToF: p=0.08; CE: -2.9%; r: 0.86, SEE: 2.5%; TE: 5.2%; LOA: ±8.6%. CONCLUSION: All three structured light scanners exhibited low magnitudes of group error (CE ≤ 1%) and may be valid assessment methods when analyzing the body composition of groups. 3DSSL-D1 exhibited the lowest group-level error (i.e. CE), prediction errors (i.e. SEE; TE), and individual error (i.e. LOA) of all scanners. Therefore, this device was deemed the most valid 3D scanner for body composition assessment. 3DSSL-D2, 3DSSL-S, and 3DSToF exhibited comparable TE, although group-level error was lower in 3DSSL-D2 and 3DSSL-S, while the SEE and individual-level error was lower for 3DSToF. However, individual-level errors were relatively high with all scanners (LOA ≥ 7%), which calls into question the utility of these methods for assessing the body composition of individuals. Nonetheless, additional research is needed regarding the ability of 3DS to successfully detect changes in body composition over time

Validity of Four-Compartment Model Body Fat Using Single- or Multi-frequency Bioelectrical Impedance Analysis to Estimate Body Water

Most common body composition assessment techniques make assumptions about the body, including the density and hydration of fat-free mass (FFM). An advantage of the four-compartment (4C) model is the ability to take these FFM characteristics into account when assessing body composition, thus reducing potential error. The total body water (TBW) estimate utilized in 4C models is particularly important due to the large contribution of water to an adult human’s total body mass (~40 - 70%) and FFM (~68 - 81%); however, the impact of utilizing different estimates of TBW within 4C model has not been fully explored. PURPOSE: The purpose of this investigation was to examine the validity of body fat percentage (BF%) estimates produced by 4C models utilizing single- or multi-frequency bioelectrical impedance analysis (BIA) TBW estimates as compared to a criterion 4C with TBW from bioimpedance spectroscopy (BIS). METHODS: After an overnight food and fluid fast, a sample of 101 adults (63 F, 38 M; age: 29.3 ± 13.5 y; BMI: 24.3 ± 4.0 kg/m2; BF%: 24.5 ± 8.3%) completed assessments via dual-energy x-ray absorptiometry (DXA), air displacement plethysmography (ADP), BIS, single-frequency BIA (SFBIA), multi-frequency BIA (MFBIA) and a body mass scale. A criterion 4C model (4CBIS) estimate of BF% was obtained using DXA for bone mineral, ADP for body volume, scale for body mass, and BIS for TBW. BIS was used as the reference TBW method due to its more direct estimation of TBW via mathematical procedures (i.e. Cole modeling and mixture theories) as compared to the prediction equations used by BIA. Alternate 4C estimates of BF% were produced using TBW values from MFBIA (4CMFBIA) and SFBIA (4CSFBIA). BF% estimates were compared using one-way ANOVA, and additional evaluations were conducted using the coefficient of determination (R2), constant error (CE), total error (TE), and 95% limits of agreement (LOA). RESULTS: BF% did not differ between 4CBIS (24.5 ± 8.3%), 4CMFBIA (24.4 ± 8.9%), and 4CSFBIA (25.7 ± 8.3%; p=0.52). 4CMFBIA exhibited negligible CE (-0.1 ± 2.3%), R2 of 0.97, TE of 2.3%, and LOA of 4.4%. 4CSFBIA exhibited a small CE (1.2 ± 1.2%), R2 of 0.98, TE of 1.6%, and LOA of 2.3%. CONCLUSION: At the group level, BF% estimates did not differ between any 4C model, indicating that both SFBIA and MFBIA can serve as viable alternatives to BIS for TBW estimation. Although the magnitude of group error (i.e. CE) was slightly smaller in 4CMFBIA, the individual error (i.e. LOA) and total error were smaller in 4CSFBIA,indicating that SFBIA TBW estimates may be more appropriate when tracking body composition changes within individuals using a 4C model. While the MFBIA and SFBIA technologies employed in the present study exhibited good validity, these results may not be attributable to all BIA analyzers. The quality of assessment device, affordability, portability and ease of use should be considered when utilizing an impedance-based technology for TBW estimation in a 4C model

Needle Electrode-Based Electromechanical Reshaping of Cartilage

Electromechanical reshaping (EMR) of cartilage provides an alternative to the classic surgical techniques of modifying the shape of facial cartilages. The original embodiment of EMR required surface electrodes to be in direct contact with the entire cartilage region being reshaped. This study evaluates the feasibility of using needle electrode systems for EMR of facial cartilage and evaluates the relationships between electrode configuration, voltage, and application time in effecting shape change. Flat rabbit nasal septal cartilage specimens were deformed by a jig into a 90° bend, while a constant electric voltage was applied to needle electrodes that were inserted into the cartilage. The electrode configuration, voltage (0–7.5 V), and application time (1–9 min) were varied systematically to create the most effective shape change. Electric current and temperature were measured during voltage application, and the resulting specimen shape was assessed in terms of retained bend angle. In order to demonstrate the clinical feasibility of EMR, the most effective and practical settings from the septal cartilage experimentation were used to reshape intact rabbit and pig ears ex vivo. Cell viability of the cartilage after EMR was determined using confocal microscopy in conjunction with a live/dead assay. Overall, cartilage reshaping increased with increased voltage and increased application time. For all electrode configurations and application times tested, heat generation was negligible (<1 °C) up to 6 V. At 6 V, with the most effective electrode configuration, the bend angle began to significantly increase after 2 min of application time and began to plateau above 5 min. As a function of voltage at 2 min of application time, significant reshaping occurred at and above 5 V, with no significant increase in the bend angle between 6 and 7.5 V. In conclusion, electromechanical reshaping of cartilage grafts and intact ears can be effectively performed with negligible temperature elevation and spatially limited cell injury using needle electrodes

A bovine lymphosarcoma cell line infected with theileria annulata exhibits an irreversible reconfiguration of host cell gene expression

Theileria annulata, an intracellular parasite of bovine lymphoid cells, induces substantial phenotypic alterations to its host cell including continuous proliferation, cytoskeletal changes and resistance to apoptosis. While parasite induced modulation of host cell signal transduction pathways and NFκB activation are established, there remains considerable speculation on the complexities of the parasite directed control mechanisms that govern these radical changes to the host cell. Our objectives in this study were to provide a comprehensive analysis of the global changes to host cell gene expression with emphasis on those that result from direct intervention by the parasite. By using comparative microarray analysis of an uninfected bovine cell line and its Theileria infected counterpart, in conjunction with use of the specific parasitacidal agent, buparvaquone, we have identified a large number of host cell gene expression changes that result from parasite infection. Our results indicate that the viable parasite can irreversibly modify the transformed phenotype of a bovine cell line. Fifty percent of genes with altered expression failed to show a reversible response to parasite death, a possible contributing factor to initiation of host cell apoptosis. The genes that did show an early predicted response to loss of parasite viability highlighted a sub-group of genes that are likely to be under direct control by parasite infection. Network and pathway analysis demonstrated that this sub-group is significantly enriched for genes involved in regulation of chromatin modification and gene expression. The results provide evidence that the Theileria parasite has the regulatory capacity to generate widespread change to host cell gene expression in a complex and largely irreversible manner

An ion mobility mass spectrometer coupled with a cryogenic ion trap for recording electronic spectra of charged, isomer-selected clusters

Infrared and electronic spectra are indispensable for understanding the structural and energetic properties of charged molecules and clusters in the gas phase. However, the presence of isomers can potentially complicate the interpretation of spectra, even if the target molecules or clusters are mass-selected beforehand. Here, we describe an instrument for spectroscopically characterizing charged molecular clusters that have been selected according to both their isomeric form and their mass-to-charge ratio. Cluster ions generated by laser ablation of a solid sample are selected according to their collision cross sections with helium buffer gas using a drift tube ion mobility spectrometer and their mass-to-charge ratio using a quadrupole mass filter. The mobility- and mass-selected target ions are introduced into a cryogenically cooled, three-dimensional quadrupole ion trap where they are thermalized through inelastic collisions with an inert buffer gas (He or He/N2 mixture). Spectra of the molecular ions are obtained by tagging them with inert atoms or molecules (Ne and N2), which are dislodged following resonant excitation of an electronic transition, or by photodissociating the cluster itself following absorption of one or more photons. An electronic spectrum is generated by monitoring the charged photofragment yield as a function of wavelength. The capacity of the instrument is illustrated with the resonance-enhanced photodissociation action spectra of carbon clusters (Cn+) and polyacetylene cations (HC2nH+) that have been selected according to the mass-to-charge ratio and collision cross section with He buffer gas and of mass-selected Au2+ and Au2Ag+ clusters