My Brother’s Keeper How American Lutherans Fought to Preserve the Evangelical Lutheran Church of Russia in the USSR, 1921–1939

The Gates of Hell Shall Not Prevail The years between 1939 and the fall of the Soviet Union in 1991 were bleak for Christians. Surrounded by the daily drumbeat of atheist propaganda, many kept their faith underground, not just figuratively but literally by hiding Bibles under the floorboards. World War II enacted a severe toll upon the nation as well as Christians, whether underground or open about their faith. Friedrich Wacker had survived the Great Terror and the closing of the churches, but when Nazi Germany invaded the USSR on June 22, 1941, he knew his family would suffer as a result and he deemed it best that they suffer together as a family. Previously he would illegally take the local night train to visit his wife and children in Leningrad, assuming that under the cover of dark he could slip in and out of their apartment more easily. But with the coming of war to Russia, he decided to risk it and take the day train. The caretaker at the apartment recognized him and in-formed the NKVD. He was arrested on June 28, 1941. When his wife heard about his arrest, she went looking for him. She, too, was arrested. Wacker was shot on July 10th, the last professor of the Leningrad Lutheran seminary who had been left alive. His wife suffered the same fate as well

SARS coronavirus and innate immunity

The emergence of the highly pathogenic SARS coronavirus (SARS-CoV) has reignited interest in coronavirus biology and pathogenesis. An emerging theme in coronavirus pathogenesis is that the interaction between specific viral genes and the host immune system, specifically the innate immune system, functions as a key determinant in regulating virulence and disease outcomes. Using SARS-CoV as a model, we will review the current knowledge of the interplay between coronavirus infection and the host innate immune system in vivo, and then discuss the mechanisms by which specific gene products antagonize the host innate immune response in cell culture models. Our data suggests that the SARS-CoV uses specific strategies to evade and antagonize the sensing and signaling arms of the interferon pathway. We summarize by identifying future points of consideration that will contribute greatly to our understanding of the molecular mechanisms governing coronavirus pathogenesis and virulence, and the development of severe disease in humans and animals

A Dynamic Culture System to Support the In Vitro Growth and Maturation of Ovarian Follicles

Ovarian follicle growth is a prolonged process that involves progressive development of the follicle unit through specific histologically defined stages of development. Groups around the world have begun ovarian cryopreservation programs for young girls and women undergoing potentially sterilizing surgery or chemotherapy with the hope that follicles can be isolated from these tissues and grown in vitro at a later date. Though follicles derived from mice can be grown up to maturity using conventional culture techniques, scientists have been unsuccessful with the in vitro development of follicles from species that have larger follicles. The objective of this study was to develop a culture system that could better support the growth and maturation of these larger follicles. The aims of the study focused on maintaining structural integrity through a suspension culture technique, providing three-dimensional support by utilizing an alginate microencapsulation technique, and creating a unique oxygen environment that more closely mimicked the oxygen levels of the native ovary. The suspension culture technique was found to eliminate follicle flattening that occurred with larger follicles on flat surfaces in a static culture. The alginate microencapsulation technique was shown to improve the support of three-dimensional growth of preantral follicles; but requires the inclusion of FSH in the scaffold in order to maintain the growth rate of unencapsulated follicles. Finally, by implementing a dynamic oxygen protocol based on the unique oxygen environment of the ovary, both the yield and quality of the oocytes derived from in vitro cultured preantral follicles were significantly improved when compared to oocytes from follicles cultured at the traditional ambient oxygen levels. In addition, these oocytes were not only able to undergo parthenogenetic activation, but were also fertilized through intracytoplasmic sperm injection. A subsequent gene expression analysis uncovered that follicles grown in a high oxygen environment possessed more differentially expressed genes compared to an in vivo control than did follicles cultured in a low oxygen environment. Furthermore, these differentially expressed genes were found to regulate several key processes that contribute to proper follicular development. These findings have contributed to the development of a novel culture system that has enhanced the in vitro support of follicle and oocyte maturation

Calcium alginate microencapsulation of ovarian follicles impacts FSH delivery and follicle morphology

BACKGROUND: We have previously shown that suspension culture prevents follicle flattening and maintains three-dimensional follicle architecture better than culture on flat plates. However, many of the follicles cultured in suspension do eventually rupture, as basement membrane integrity is lost and the three-dimensional structure of the follicle is altered. Therefore, the objective of this study is to support three-dimensional follicle architecture during in vitro growth of ovarian follicles through encapsulation in calcium alginate, while maintaining responsiveness to FSH stimulation. METHODS: Preantral follicles (150 – 160 micrometers in diameter) were isolated from the ovaries of juvenile rats and grown in culture tubes or encapsulated in calcium alginate and grown in culture tubes. Previous studies revealed that follicles maintained structural integrity but did not grow as well when encapsulated in calcium alginate. In these studies, we evaluated the effect of calcium alginate on FSH-stimulated follicle growth, survival, and morphology in suspension culture. Follicles were grown under 5 culture conditions: 1) not encapsulated; with FSH in the medium, 2) encapsulated in the absence of FSH, grown in medium without FSH, 3) encapsulated with calcium alginate containing FSH but grown in medium without FSH, 4) encapsulated without FSH but grown in medium containing FSH and 5) encapsulated with calcium alginate containing FSH and in medium containing FSH. To assess growth rates, follicles were cultured for 72 hours and analyzed for follicle size increase and DNA content. Survival analysis for encapsulated and unencapsulated follicles was performed by constructing a Kaplan Meier survival curve of daily observations of intact follicle survival. Three-dimensional architecture was assessed histologically and by analysis of the pattern of connexin 43 expression in the cultured follicles. RESULTS: In the absence of FSH, follicle diameter increased by only 6.4%. When FSH was included in the alginate bead alone or the media alone, the follicle diameter increased by 13.5% and 19.9% respectively. This was greater than follicles cultured in the absence of FSH (p < 0.05), but less than that of the FSH-treated unencapsulated follicles (p < 0.05). However, when follicles were cultured with FSH included in both the media and the bead, a 32.6% increase in follicle diameter was observed, statistically no different than the growth rate of the unencapsulated follicles grown with FSH. CONCLUSION: Microencapsulation supports three-dimensional follicle growth, but may limit access to hormones in the medium resulting in altered development compared to unencapsulated follicles. Inclusion of FSH in the alginate bead restores the follicle growth response to FSH, while also providing a scaffold of support for three-dimensional growth. The application of tissue engineering principles to the problems of follicle culture in vitro may provide advances applicable to fertility preservation in women and endangered species

Surface mannosylation of dispersion polymerisation derived nanoparticles by copper mediated click chemistry

The synthesis of spherical polymeric nanoparticles containing alkyne surface functionalities for post poly- merisation glycosylation is described. The nanoparticles were obtained by a polymerisation induced self- assembly (PISA) inspired methodology in dispersed media by Cu(0) mediated polymerisation. A water soluble poly(ethylene glycol methacrylate-stat-propargyl methacrylate), poly(PEGMA18-stat-PgMA5), macroinitiator was first synthesised and chain extended with 2-hydroxypropyl methacrylate (HPMA) in water using a copper wire catalyst. It was found that irrespective of the macroinitiator to HPMA ratio and the reaction time the desired spherical morphologies (&lt;100 nm) were obtained while the absence other morphologies suggest a deviation from the classical PISA process due to chain termination in the nano- particle’s core. The obtained nanoparticles contained alkyne functionalities in the shell, which were suc- cessfully reacted by copper mediated click chemistry with fluoresceine azide and mannosides with hydro- phobic and hydrophilic spacers of different lengths. The obtained mannosylated nanoparticles displayed no significant cytotoxicity against human alveolar basal epithelial adenocarcinomic (A549) cells at any dose &lt;0.5 mg mL−1. Preliminary binding studies confirm the ability of the mannosylated nanoparticles to bind to human lectin dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN). The methodology reported here is a convenient route to well-defined spherical and shell- functionalisable nanoparticles to create libraries of bio-active nanomaterials

High molecular weight polyproline as a potential biosourced ice growth inhibitor : synthesis, ice recrystallization inhibition, and specific ice face binding

Ice-binding proteins (IBPs) from extremophile organisms can modulate ice formation and growth. There are many (bio)technological applications of IBPs, from cryopreservation to mitigating freeze–thaw damage in concrete to frozen food texture modifiers. Extraction or expression of IBPs can be challenging to scale up, and hence polymeric biomimetics have emerged. It is, however, desirable to use biosourced monomers and heteroatom-containing backbones in polymers for in vivo or environmental applications to allow degradation. Here we investigate high molecular weight polyproline as an ice recrystallization inhibitor (IRI). Low molecular weight polyproline is known to be a weak IRI. Its activity is hypothesized to be due to the unique PPI helix it adopts, but it has not been thoroughly investigated. Here an open-to-air aqueous N-carboxyanhydride polymerization is employed to obtain polyproline with molecular weights of up to 50000 g mol–1. These polymers were found to have IRI activity down to 5 mg mL–1, unlike a control peptide of polysarcosine, which did not inhibit all ice growth at up to 40 mg mL–1. The polyprolines exhibited lower critical solution temperature behavior and assembly/aggregation observed at room temperature, which may contribute to its activity. Single ice crystal assays with polyproline led to faceting, consistent with specific ice-face binding. This work shows that non-vinyl-based polymers can be designed to inhibit ice recrystallization and may offer a more sustainable or environmentally acceptable, while synthetically scalable, route to large-scale applications

A Comparison of Copper Abundances in Globular Cluster and Halo Field Giant Stars

We derive [Cu/Fe] for 117 giant stars in ten globular clusters (M3, M4, M5, M10, M13, M15, M71, NGC 7006, NCG 288, and NGC 362) and find that globular cluster Cu abundances appear to follow [Cu/Fe] trends found in the field. This result is interesting in light of recent work which indicates that the globular cluster Omega Centauri shows no trend in [Cu/Fe] with [Fe/H] over the abundance range -2.0 <[Fe/H]< -0.8. Of particular interest are the two clusters M4 and M5. While at a similar metallicity ([Fe/H] ~- 1.2), they differ greatly in some elemental abundances: M4 is largely overabundant in Si, Ba, and La compared to M5. We find that it is also overabundant in Cu with respect to M5, though this overabundance is in accord with [Cu/Fe] ratios found in the field.Comment: 39 pages, 7 figures, to appear in April 2003 A

Deep Underground Science and Engineering Laboratory - Preliminary Design Report

The DUSEL Project has produced the Preliminary Design of the Deep Underground Science and Engineering Laboratory (DUSEL) at the rehabilitated former Homestake mine in South Dakota. The Facility design calls for, on the surface, two new buildings - one a visitor and education center, the other an experiment assembly hall - and multiple repurposed existing buildings. To support underground research activities, the design includes two laboratory modules and additional spaces at a level 4,850 feet underground for physics, biology, engineering, and Earth science experiments. On the same level, the design includes a Department of Energy-shepherded Large Cavity supporting the Long Baseline Neutrino Experiment. At the 7,400-feet level, the design incorporates one laboratory module and additional spaces for physics and Earth science efforts. With input from some 25 science and engineering collaborations, the Project has designed critical experimental space and infrastructure needs, including space for a suite of multidisciplinary experiments in a laboratory whose projected life span is at least 30 years. From these experiments, a critical suite of experiments is outlined, whose construction will be funded along with the facility. The Facility design permits expansion and evolution, as may be driven by future science requirements, and enables participation by other agencies. The design leverages South Dakota's substantial investment in facility infrastructure, risk retirement, and operation of its Sanford Laboratory at Homestake. The Project is planning education and outreach programs, and has initiated efforts to establish regional partnerships with underserved populations - regional American Indian and rural populations

Identification and characterization of a novel non-structural protein of bluetongue virus

Bluetongue virus (BTV) is the causative agent of a major disease of livestock (bluetongue). For over two decades, it has been widely accepted that the 10 segments of the dsRNA genome of BTV encode for 7 structural and 3 non-structural proteins. The non-structural proteins (NS1, NS2, NS3/NS3a) play different key roles during the viral replication cycle. In this study we show that BTV expresses a fourth non-structural protein (that we designated NS4) encoded by an open reading frame in segment 9 overlapping the open reading frame encoding VP6. NS4 is 77–79 amino acid residues in length and highly conserved among several BTV serotypes/strains. NS4 was expressed early post-infection and localized in the nucleoli of BTV infected cells. By reverse genetics, we showed that NS4 is dispensable for BTV replication in vitro, both in mammalian and insect cells, and does not affect viral virulence in murine models of bluetongue infection. Interestingly, NS4 conferred a replication advantage to BTV-8, but not to BTV-1, in cells in an interferon (IFN)-induced antiviral state. However, the BTV-1 NS4 conferred a replication advantage both to a BTV-8 reassortant containing the entire segment 9 of BTV-1 and to a BTV-8 mutant with the NS4 identical to the homologous BTV-1 protein. Collectively, this study suggests that NS4 plays an important role in virus-host interaction and is one of the mechanisms played, at least by BTV-8, to counteract the antiviral response of the host. In addition, the distinct nucleolar localization of NS4, being expressed by a virus that replicates exclusively in the cytoplasm, offers new avenues to investigate the multiple roles played by the nucleolus in the biology of the cell

A Universal Next-Generation Sequencing Protocol To Generate Noninfectious Barcoded cDNA Libraries from High-Containment RNA Viruses

ABSTRACT Several biosafety level 3 and/or 4 (BSL-3/4) pathogens are high-consequence, single-stranded RNA viruses, and their genomes, when introduced into permissive cells, are infectious. Moreover, many of these viruses are select agents (SAs), and their genomes are also considered SAs. For this reason, cDNAs and/or their derivatives must be tested to ensure the absence of infectious virus and/or viral RNA before transfer out of the BSL-3/4 and/or SA laboratory. This tremendously limits the capacity to conduct viral genomic research, particularly the application of next-generation sequencing (NGS). Here, we present a sequence-independent method to rapidly amplify viral genomic RNA while simultaneously abolishing both viral and genomic RNA infectivity across multiple single-stranded positive-sense RNA (ssRNA+) virus families. The process generates barcoded DNA amplicons that range in length from 300 to 1,000 bp, which cannot be used to rescue a virus and are stable to transport at room temperature. Our barcoding approach allows for up to 288 barcoded samples to be pooled into a single library and run across various NGS platforms without potential reconstitution of the viral genome. Our data demonstrate that this approach provides full-length genomic sequence information not only from high-titer virion preparations but it can also recover specific viral sequence from samples with limited starting material in the background of cellular RNA, and it can be used to identify pathogens from unknown samples. In summary, we describe a rapid, universal standard operating procedure that generates high-quality NGS libraries free of infectious virus and infectious viral RNA. IMPORTANCE This report establishes and validates a standard operating procedure (SOP) for select agents (SAs) and other biosafety level 3 and/or 4 (BSL-3/4) RNA viruses to rapidly generate noninfectious, barcoded cDNA amenable for next-generation sequencing (NGS). This eliminates the burden of testing all processed samples derived from high-consequence pathogens prior to transfer from high-containment laboratories to lower-containment facilities for sequencing. Our established protocol can be scaled up for high-throughput sequencing of hundreds of samples simultaneously, which can dramatically reduce the cost and effort required for NGS library construction. NGS data from this SOP can provide complete genome coverage from viral stocks and can also detect virus-specific reads from limited starting material. Our data suggest that the procedure can be implemented and easily validated by institutional biosafety committees across research laboratories