The feasibility of non-viral gene transfer to the diaphragm in vivo

Gene transfer using electroporation is an essential method for the study of developmental biology, especially to understand the internal control of degeneration and apoptosis of the muscle cells that occurs earlier and quicker than the usual degeneration process occurring by aging. Such experimental studies may have a role in developing new strategies for treating patients suffering from inherited primary myopathies such as Duchenne muscular dystrophy (DMD). The present study was designed to evaluate the feasibility of electroporation mediated transfer of reporter genes to the diaphragm in vivo. This is the first report of gene transfer of naked plasmid DNA into the diaphragm muscle in vivo using electroporation. Our results showed that in vivo gene transfer of naked plasmid DNA into the diaphragm muscle using electroporation is feasible.

Programmed death 1 signaling on chronic myeloid leukemia-specific T cells results in T-cell exhaustion and disease progression

Chronic myeloid leukemia (CML) is a malignant myeloproliferative disease with a characteristic chronic phase (cp) of several years before progression to blast crisis (bc). The immune system may contribute to disease control in CML. We analyzed leukemia-specific immune responses in cpCML and bcCML in a retroviral-induced murine CML model. In the presence of cpCML and bcCML expressing the glycoprotein of lymphocytic choriomeningitis virus as a model leukemia antigen, leukemia-specific cytotoxic T lymphocytes (CTLs) became exhausted. They maintained only limited cytotoxic activity, and did not produce interferon-gamma or tumor necrosis factor-alpha or expand after restimulation. CML-specific CTLs were characterized by high expression of programmed death 1 (PD-1), whereas CML cells expressed PD-ligand 1 (PD-L1). Blocking the PD-1/PD-L1 interaction by generating bcCML in PD-1-deficient mice or by repetitive administration of alphaPD-L1 antibody prolonged survival. In addition, we found that PD-1 is up-regulated on CD8(+) T cells from CML patients. Taken together, our results suggest that blocking the PD-1/PD-L1 interaction may restore the function of CML-specific CTLs and may represent a novel therapeutic approach for CML.

Mammary epithelial-specific knockout of the ephrin-B2 gene leads to precocious epithelial cell death at lactation

The family of Eph receptor tyrosine kinases and their membrane bound ligands, the ephrins, are involved in a wide variety of morphogenic processes during embryonic development and adult tissue homeostasis. Receptor-ligand interaction requires direct cell-cell contact and results in forward and reverse signaling originating from the receptor and ligand, respectively. We have previously shown that EphB4 and ephrinB2 are differentially expressed during the development of the adult mammary parenchyma. Overexpression of EphB4 in the mammary epithelium of transgenic mice leads to perturbations in mammary epithelial morphology, motility and growth. To investigate the role of ephrinB2 signaling in mammary gland biology, we have established transgenic mice exhibiting conditional ephrinB2 knockout in the mammary epithelium. In homozygote double transgenic CreLox mice, specific knockout of ephrinB2 occurred in the mammary epithelium during the first pregnancy-lactating period. Abolishing ephrinB2 function led to severe interference with the architecture and functioning of the mammary gland at lactation. The morphology of the transgenic lactating glands resembled that of involuting controls, with decreased epithelial cell number and collapsed lobulo-alveolar structures. Accordingly, massive epithelial cell death and expression of involution-specific genes were observed. Interestingly, in parallel to cell death, significant cell proliferation was apparent, suggestive of tissue regeneration.

Discontinuation of enfuvirtide in heavily pretreated HIV-infected individuals

BACKGROUND: Enfuvirtide was shown to be highly effective in treatment- experienced patients. Data on discontinuation of enfuvirtide and switch to new antiretroviral drugs are scarce. We aimed to evaluate the efficacy and the impact of discontinuing and/or switching enfuvirtide on virologic and clinical parameters in clinical practice. METHODS: All HIV-infected individuals participating in the Swiss HIV Cohort Study who were treated with enfuvirtide for at least 4 weeks in combination with an optimized background antiretroviral regimen were included in this study. RESULTS: A total of 151 patients were analyzed. The median baseline CD4 cell count was 108 cells/microL (interquartile range [IQR] 50-206) and HIV RNA was 4.7 log10 copies/mL (IQR 4.1-5.2). Virologic suppression, defined as a viral load below 50 copies/mL at 12 months, was achieved by 57.6% of patients. Overall, a median CD4 cell increase of 121 cells/microL (IQR 50-189) from baseline was noted. Up to 50% of patients discontinued enfuvirtide within the first year of treatment, mainly because of the patient's choice. After discontinuation of enfuvirtide, high rates of virologic failure and clinical progression were observed, notably when CD4 cell count at stopping enfuvirtide was below 100 cells/microL and no switch to new potent antiretroviral drugs such as darunavir, maraviroc, or raltegravir was performed. CONCLUSIONS: Enfuvirtide provides high virologic and immunologic response in treatment-experienced patients in the setting of clinical practice. Enfuvirtide should not be discontinued but should be replaced by new potent antiretrovirals, particularly in case of severe immunosuppression.

Parameter-free extraction of the functional network topology from intracranial EEG recordings: a time-resolved study of graph properties in focal onset seizures

Rationale: Focal onset epileptic seizures are due to abnormal interactions between distributed brain areas. By estimating the cross-correlation matrix of multi-site intra-cerebral EEG recordings (iEEG), one can quantify these interactions. To assess the topology of the underlying functional network, the binary connectivity matrix has to be derived from the cross-correlation matrix by use of a threshold. Classically, a unique threshold is used that constrains the topology [1]. Our method aims to set the threshold in a data-driven way by separating genuine from random cross-correlation. We compare our approach to the fixed threshold method and study the dynamics of the functional topology. Methods: We investigate the iEEG of patients suffering from focal onset seizures who underwent evaluation for the possibility of surgery. The equal-time cross-correlation matrices are evaluated using a sliding time window. We then compare 3 approaches assessing the corresponding binary networks. For each time window: * Our parameter-free method derives from the cross-correlation strength matrix (CCS)[2]. It aims at disentangling genuine from random correlations (due to finite length and varying frequency content of the signals). In practice, a threshold is evaluated for each pair of channels independently, in a data-driven way. * The fixed mean degree (FMD) uses a unique threshold on the whole connectivity matrix so as to ensure a user defined mean degree. * The varying mean degree (VMD) uses the mean degree of the CCS network to set a unique threshold for the entire connectivity matrix. * Finally, the connectivity (c), connectedness (given by k, the number of disconnected sub-networks), mean global and local efficiencies (Eg, El, resp.) are computed from FMD, CCS, VMD, and their corresponding random and lattice networks. Results: Compared to FMD and VMD, CCS networks present: *topologies that are different in terms of c, k, Eg and El. *from the pre-ictal to the ictal and then post-ictal period, topological features time courses that are more stable within a period, and more contrasted from one period to the next. For CCS, pre-ictal connectivity is low, increases to a high level during the seizure, then decreases at offset. k shows a ‘‘U-curve’’ underlining the synchronization of all electrodes during the seizure. Eg and El time courses fluctuate between the corresponding random and lattice networks values in a reproducible manner. Conclusions: The definition of a data-driven threshold provides new insights into the topology of the epileptic functional networks.

Effects of thermal environment on hypothalamic-pituitary-adrenal axis hormones, oxytocin, and behavioral activity in periparturient sows

Provision of additional floor heating (33 to 34 degrees C) at birth and during the early postnatal hours is favorable for newborn piglets of domestic sows (Sus scrofa). We investigated whether this relatively high temperature influenced sow behavior and physiology around farrowing. One-half of 28 second-parity pregnant sows were randomly chosen to be exposed to floor heating 12 h after onset of nest building and until 48 h after birth of the first piglet (heat treatment), whereas the rest of the sows entered the control group (control treatment) with no floor heating. Hourly blood sampling from 8 h before and until 24 h after the birth of the first piglet was used for investigation of temporal changes in plasma concentrations of oxytocin, cortisol, and ACTH. In addition, occurrence and duration of sow postures were recorded -8 to +48 h relative to the birth of the first piglet. There was a clear temporal development in sow behavior and hormone concentrations (ACTH, cortisol, and oxytocin) across parturition (P < 0.001), independent of treatment. In general, hormone concentrations increased from the start to the end of farrowing. The observed oxytocin increase and peak late in farrowing coincided with the passive phase where sows lie laterally with an overall reduced activity. Floor heating increased the mean concentration of cortisol (P = 0.02; estimated as 29% greater than in controls) and tended to increase the mean concentration of ACTH (P = 0.08; estimated as 17% greater than in controls), but we did not find any treatment effect on mean oxytocin concentrations, the course of parturition, or the behavior of sows. Behavioral thermoregulation may, however, have lost some function for the sows because the floor was fully heated in our study. In addition, exposure to heat decreased the between-sow variation of plasma oxytocin (approximately 31% less relative to control) and ACTH (approximately 46% less relative to control). Whether this decreased variation may be indicative of acute stress or linked to other biological events is unclear. In conclusion, inescapable floor heating (around 33.5 degrees C) may be considered a stressor for sows around farrowing, giving rise to elevated plasma concentrations of cortisol, but without concurrent changes in oxytocin or behavioral activity.

A Benzaldehyde Derivative as a Chelating Ligand: Helical Manganese(II) Coordination Polymers Assembling into a Porous Solid

A molecular, porous crystalline material constructed from neutral helical coordination polymers incorporating manganese(II) ions and two types of bridging ligands, namely the deprotonated form of 2-hydroxy-5-methoxy-3-nitrobenzaldehyde (HL) and isobutyrate (iB−), has been obtained and structurally characterized. Structural analysis reveals that within the coordination polymer each benzaldehyde derivative ligates two manganese ions in 6-membered chelating rings, and the isobutyrate ligands cooperatively chelate either two or three manganese ions. The solid state assembly of the resulting polymeric chains of formula [Mn4(L)2(iB)6]n (1), described in the polar space group R3c, is associated with tubular channels occupied by MeCN solvent molecules (1·xMeCN; x ≤ 9). TGA profiles and PXRD measurements demonstrate that the crystallinity of the solid remains intact in its fully desolvated form, and its stability and crystallinity are ensured up to a temperature of 190 °C. Gas adsorption properties of desolvated crystals were probed, but no remarkable sorption capacity of N2 and only a limited one for CO2 could be observed. Magnetic susceptibility data reveal an antiferromagnetic type of coupling between adjacent manganese(II) ions along the helical chains with energy parameters J1 = −5.9(6) cm−1 and J2 = −1.8(9) cm−1.

The Usual Suspects: Jewish Magical Realism, Trauma and the Holocaust

Jewish magical realism is often associated immediately with the historical trauma of the Holocaust. This article sets out to point to some of the problems and limitations such an interpretive approach may bring. After a reconsideration of the allegedly inherent subversive qualities of the mode and a discussion of its frequent association with trauma narratives, especially in the case of Jewish texts engaging with the Holocaust, the discussion will focus on two examples, Michael Chabon’s The Amazing Adventures of Kavalier & Clay (2000) and Jonathan Safran Foer’s Everything Is Illuminated (2002). The aim throughout is to caution against rashly reproducing familiar interpretive maneuvers, which may be in need of reassessment in face of recent fictions like the ones under discussion in this article.

A Staphylococcus xylosus isolate with a new mecC allotype

Recently, a novel variant of mecA known as mecC (mecA(LGA251)) was identified in Staphylococcus aureus isolates from both humans and animals. In this study, we identified a Staphylococcus xylosus isolate that harbors a new allotype of the mecC gene, mecC1. Whole-genome sequencing revealed that mecC1 forms part of a class E mec complex (mecI-mecR1-mecC1-blaZ) located at the orfX locus as part of a likely staphylococcal cassette chromosome mec element (SCCmec) remnant, which also contains a number of other genes present on the type XI SCCmec.

Macular hole surgery in patients with end-stage choroideremia

OBJECTIVE To assess macular hole surgery in patients with end-stage choroideremia with regard to anatomic closure and visual outcome. DESIGN Retrospective, interventional case series. PARTICIPANTS Thirty adult male patients with a diagnosis of advanced choroideremia were reviewed and underwent spectral domain optical coherence tomography (OCT) as part of the screening process for a gene therapy clinical trial. From within that cohort, 3 were identified as having a full-thickness macular hole (FTMH). METHODS A 23-gauge pars plana vitrectomy was performed with peeling of the inner limiting membrane and gas tamponade. Preoperative best-corrected visual acuity ranged from perception of light to 6/24. MAIN OUTCOME MEASURES Prevalence of FTMH in advanced choroideremia, morphologic phenotype characteristics of FTMH in OCT, pre- and postoperative best-corrected visual acuity, and closure rate after surgery. RESULTS The prevalence of FTMH in advanced choroideremia in our cohort was 10%. One hole was associated with significant macular schisis, presumed to be attributable to degeneration of outer retinal layers. Anatomic closure was achieved in all 3 patients and confirmed with spectral domain OCT. Gas tamponade lasted approximately twice as long as might be expected compared with standard FTMH surgery. Objective visual acuity did not improve; however, perceived vision improved in all patients. CONCLUSIONS Although FTMH in choroideremia is a rare finding, it could potentially mask central progression of the disease. Regular screening may help to diagnose holes at an earlier stage when the visual prognosis after surgery may be better. Standard macular hole surgery seems to be effective in gaining anatomic closure, which would be significant for patients who subsequently require macula detachment for subretinal gene therapy. FINANCIAL DISCLOSURE(S) The authors have no proprietary or commercial interest in any of the materials discussed in this article.

Correlation between Accurate Electron Density and Linear Optical Properties in Amino Acid Derivatives: L-Histidinium Hydrogen Oxalate

The accurate electron density and linear optical properties of L-histidinium hydrogen oxalate are discussed. Two high-resolution single crystal X-ray diffraction experiments were performed and compared with density functional calculations in the solid state as well as in the gas phase. The crystal packing and the hydrogen bond network are accurately investigated using topological analysis based on quantum theory of atoms in molecules, Hirshfeld surface analysis, and electrostatic potential mapping. The refractive indices are computed from couple perturbed Kohn-Sham calculations and measured experimentally. Moreover, distributed atomic polarizabilities are used to analyze the origin of the linear susceptibility in the crystal, in order to separate molecular and intermolecular causes. The optical properties are also correlated with the electron density distribution. This compound also offers the possibility to test the electron density building block approach for material science and different refinement schemes for accurate positions and displacement parameters of hydrogen atoms, in the absence of neutron diffraction data.