The Chromatin Reader ZMYND8 Regulates Igh Enhancers to Promote Immunoglobulin Class Switch Recombination.

Class switch recombination (CSR) is a DNA recombination reaction that diversifies the effector component of antibody responses. CSR is initiated by activation-induced cytidine deaminase (AID), which targets transcriptionally active immunoglobulin heavy chain (Igh) switch donor and acceptor DNA. The 3' Igh super-enhancer, 3' regulatory region (3'RR), is essential for acceptor region transcription, but how this function is regulated is unknown. Here, we identify the chromatin reader ZMYND8 as an essential regulator of the 3'RR. In B cells, ZMYND8 binds promoters and super-enhancers, including the Igh enhancers. ZMYND8 controls the 3'RR activity by modulating the enhancer transcriptional status. In its absence, there is increased 3'RR polymerase loading and decreased acceptor region transcription and CSR. In addition to CSR, ZMYND8 deficiency impairs somatic hypermutation (SHM) of Igh, which is also dependent on the 3'RR. Thus, ZMYND8 controls Igh diversification in mature B lymphocytes by regulating the activity of the 3' Igh super-enhancer.

Engineering the cellulolytic bacterium, Clostridium thermocellum, to co-utilize hemicellulose.

Consolidated bioprocessing (CBP) of lignocellulosic biomass holds promise to realize economic production of second-generation biofuels/chemicals, and Clostridium thermocellum is a leading candidate for CBP due to it being one of the fastest degraders of crystalline cellulose and lignocellulosic biomass. However, CBP by C. thermocellum is approached with co-cultures, because C. thermocellum does not utilize hemicellulose. When compared with a single-species fermentation, the co-culture system introduces unnecessary process complexity that may compromise process robustness. In this study, we engineered C. thermocellum to co-utilize hemicellulose without the need for co-culture. By evolving our previously engineered xylose-utilizing strain in xylose, an evolved clonal isolate (KJC19-9) was obtained and showed improved specific growth rate on xylose by ∼3-fold and displayed comparable growth to a minimally engineered strain grown on the bacteria's naturally preferred substrate, cellobiose. To enable full xylan deconstruction to xylose, we recombinantly expressed three different ß-xylosidase enzymes originating from Thermoanaerobacterium saccharolyticum into KJC19-9 and demonstrated growth on xylan with one of the enzymes. This recombinant strain was capable of co-utilizing cellulose and xylan simultaneously, and we integrated the ß-xylosidase gene into the KJC19-9 genome, creating the KJCBXint strain. The strain, KJC19-9, consumed monomeric xylose but accumulated xylobiose when grown on pretreated corn stover, whereas the final KJCBXint strain showed significantly greater deconstruction of xylan and xylobiose. This is the first reported C. thermocellum strain capable of degrading and assimilating hemicellulose polysaccharide while retaining its cellulolytic capabilities, unlocking significant potential for CBP in advancing the bioeconomy.

Ecological Dichotomies Arise in Microbial Communities Due to Mixing of Deep Hydrothermal Waters and Atmospheric Gas in a Circumneutral Hot Spring.

Little is known of how the confluence of subsurface and surface processes influences the assembly and habitability of hydrothermal ecosystems. To address this knowledge gap, the geochemical and microbial composition of a high-temperature, circumneutral hot spring in Yellowstone National Park was examined to identify the sources of solutes and their effect on the ecology of microbial inhabitants. Metagenomic analysis showed that populations comprising planktonic and sediment communities are archaeal dominated, are dependent on chemical energy (chemosynthetic), share little overlap in their taxonomic composition, and are differentiated by their inferred use of/tolerance to oxygen and mode of carbon metabolism. The planktonic community is dominated by putative aerobic/aerotolerant autotrophs, while the taxonomic composition of the sediment community is more evenly distributed and comprised of anaerobic heterotrophs. These observations are interpreted to reflect sourcing of the spring by anoxic, organic carbon-limited subsurface hydrothermal fluids and ingassing of atmospheric oxygen that selects for aerobic/aerotolerant organisms that have autotrophic capabilities in the water column. Autotrophy and consumption of oxygen by the planktonic community may influence the assembly of the anaerobic and heterotrophic sediment community. Support for this inference comes from higher estimated rates of genome replication in planktonic populations than sediment populations, indicating faster growth in planktonic populations. Collectively, these observations provide new insight into how mixing of subsurface waters and atmospheric oxygen create dichotomy in the ecology of hot spring communities and suggest that planktonic and sediment communities may have been less differentiated taxonomically and functionally prior to the rise of oxygen at ∼2.4 billion years ago (Gya). IMPORTANCE Understanding the source and availability of energy capable of supporting life in hydrothermal environments is central to predicting the ecology of microbial life on early Earth when volcanic activity was more widespread. Little is known of the substrates supporting microbial life in circumneutral to alkaline springs, despite their relevance to early Earth habitats. Using metagenomic and informatics approaches, water column and sediment habitats in a representative circumneutral hot spring in Yellowstone were shown to be dichotomous, with the former largely hosting aerobic/aerotolerant autotrophs and the latter primarily hosting anaerobic heterotrophs. This dichotomy is attributed to influx of atmospheric oxygen into anoxic deep hydrothermal spring waters. These results indicate that the ecology of microorganisms in circumneutral alkaline springs sourced by deep hydrothermal fluids was different prior to the rise of atmospheric oxygen ∼2.4 Gya, with planktonic and sediment communities likely to be less differentiated than contemporary circumneutral hot springs.

Unexpected Abundance and Diversity of Phototrophs in Mats from Morphologically Variable Microbialites in Great Salt Lake, Utah.

Microbial mat communities are associated with extensive (∼700 km2) and morphologically variable carbonate structures, termed microbialites, in the hypersaline Great Salt Lake (GSL), Utah. However, whether the composition of GSL mat communities covaries with microbialite morphology and lake environment is unknown. Moreover, the potential adaptations that allow the establishment of these extensive mat communities at high salinity (14% to 17% total salts) are poorly understood. To address these questions, microbial mats were sampled from seven locations in the south arm of GSL representing different lake environments and microbialite morphologies. Despite the morphological differences, microbialite-associated mats were taxonomically similar and were dominated by the cyanobacterium Euhalothece and several heterotrophic bacteria. Metagenomic sequencing of a representative mat revealed Euhalothece and subdominant Thiohalocapsa populations that harbor the Calvin cycle and nitrogenase, suggesting they supply fixed carbon and nitrogen to heterotrophic bacteria. Fifteen of the next sixteen most abundant taxa are inferred to be aerobic heterotrophs and, surprisingly, harbor reaction center, rhodopsin, and/or bacteriochlorophyll biosynthesis proteins, suggesting aerobic photoheterotrophic (APH) capabilities. Importantly, proteins involved in APH are enriched in the GSL community relative to that in microbialite mat communities from lower salinity environments. These findings indicate that the ability to integrate light into energy metabolism is a key adaptation allowing for robust mat development in the hypersaline GSL.IMPORTANCE The earliest evidence of life on Earth is from organosedimentary structures, termed microbialites, preserved in 3.481-billion-year-old (Ga) rocks. Phototrophic microbial mats form in association with an ∼700-km2 expanse of morphologically diverse microbialites in the hypersaline Great Salt Lake (GSL), Utah. Here, we show taxonomically similar microbial mat communities are associated with morphologically diverse microbialites across the lake. Metagenomic sequencing reveals an abundance and diversity of autotrophic and heterotrophic taxa capable of harvesting light energy to drive metabolism. The unexpected abundance of and diversity in the mechanisms of harvesting light energy observed in GSL mat populations likely function to minimize niche overlap among coinhabiting taxa, provide a mechanism(s) to increase energy yield and osmotic balance during salt stress, and enhance fitness. Together, these physiological benefits promote the formation of robust mats that, in turn, influence the formation of morphologically diverse microbialite structures that can be imprinted in the rock record.

Metabolically engineered Caldicellulosiruptor bescii as a platform for producing acetone and hydrogen from lignocellulose.

The production of volatile industrial chemicals utilizing metabolically engineered extreme thermophiles offers the potential for processes with simultaneous fermentation and product separation. An excellent target chemical for such a process is acetone (Tb = 56°C), ideally produced from lignocellulosic biomass. Caldicellulosiruptor bescii (Topt 78°C), an extremely thermophilic fermentative bacterium naturally capable of deconstructing and fermenting lignocellulose, was metabolically engineered to produce acetone. When the acetone pathway construct was integrated into a parent strain containing the bifunctional alcohol dehydrogenase from Clostridium thermocellum, acetone was produced at 9.1 mM (0.53 g/L), in addition to minimal ethanol 3.3 mM (0.15 g/L), along with net acetate consumption. This demonstrates that C. bescii can be engineered with balanced pathways in which renewable carbohydrate sources are converted to useful metabolites, primarily acetone and H2 , without net production of its native fermentation products, acetate and lactate.

Modification of the glycolytic pathway in Pyrococcus furiosus and the implications for metabolic engineering.

The key difference in the modified Embden-Meyerhof glycolytic pathway in hyperthermophilic Archaea, such as Pyrococcus furiosus, occurs at the conversion from glyceraldehyde-3-phosphate (GAP) to 3-phosphoglycerate (3-PG) where the typical intermediate 1,3-bisphosphoglycerate (1,3-BPG) is not present. The absence of the ATP-yielding step catalyzed by phosphoglycerate kinase (PGK) alters energy yield, redox energetics, and kinetics of carbohydrate metabolism. Either of the two enzymes, ferredoxin-dependent glyceraldehyde-3-phosphate ferredoxin oxidoreductase (GAPOR) or NADP+-dependent non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase (GAPN), responsible for this "bypass" reaction, could be deleted individually without impacting viability, albeit with differences in native fermentation product profiles. Furthermore, P. furiosus was viable in the gluconeogenic direction (growth on pyruvate or peptides plus elemental sulfur) in a ΔgapnΔgapor strain. Ethanol was utilized as a proxy for potential heterologous products (e.g., isopropanol, butanol, fatty acids) that require reducing equivalents (e.g., NAD(P)H, reduced ferredoxin) generated from glycolysis. Insertion of a single gene encoding the thermostable NADPH-dependent primary alcohol dehydrogenase (adhA) (Tte_0696) from Caldanaerobacter subterraneus, resulted in a strain producing ethanol via the previously established aldehyde oxidoreductase (AOR) pathway. This strain demonstrated a high ratio of ethanol over acetate (> 8:1) at 80 °C and enabled ethanol production up to 85 °C, the highest temperature for bio-ethanol production reported to date.

Determinants of sulphur chemolithoautotrophy in the extremely thermoacidophilic Sulfolobales.

Species in the archaeal order Sulfolobales thrive in hot acid and exhibit remarkable metabolic diversity. Some species are chemolithoautotrophic, obtaining energy through the oxidation of inorganic substrates, sulphur in particular, and acquiring carbon through the 3-hydroxypropionate/4-hydroxybutyrate (3-HP/4-HB) CO2 -fixation cycle. The current model for sulphur oxidation in the Sulfolobales is based on the biochemical analysis of specific proteins from Acidianus ambivalens, including sulphur oxygenase reductase (SOR) that disproportionates S° into H2 S and sulphite (SO3 2- ). Initial studies indicated SOR catalyses the essential first step in oxidation of elemental sulphur, but an ancillary role for SOR as a 'recycle' enzyme has also been proposed. Here, heterologous expression of both SOR and membrane-bound thiosulphate-quinone oxidoreductase (TQO) from Sulfolobus tokodaii 'restored' sulphur oxidation capacity in Sulfolobus acidocaldarius DSM639, but not autotrophy, although earlier reports indicate this strain was once capable of chemolithoautotrophy. Comparative transcriptomic analyses of Acidianus brierleyi, a chemolithoautotrophic sulphur oxidizer, and S. acidocaldarius DSM639 showed that while both share a strong transcriptional response to elemental sulphur, S. acidocaldarius DSM639 failed to upregulate key 3-HP/4-HB cycle genes used by A. brierleyi to drive chemolithoautotrophy. Thus, the inability for S. acidocaldarius DSM639 to grow chemolithoautotrophically may be rooted more in gene regulation than the biochemical capacity.

Selective Synthesis of Perfluoroalkylated Corannulenes and Investigation of their Structural, Dynamic and Electrochemical Behavior.

Herein we report general methods allowing the synthesis of various perfluoroalkylated corannulenes with a specific substitution pattern. Variable temperature NMR spectroscopic investigations revealed dynamic behavior which was analyzed by line shape analysis. The activation parameters of these dynamic processes were determined. For a tetrasubstituted compound it was possible to observe through space scalar coupling. The packing motifs were elucidated by X-ray crystallography, showing that the substitution pattern as well as the size of substituents strongly influence intermolecular π-stacking. The reduction potentials of the perfluoroalkylated compounds were determined by cyclic voltammetry.

Alternative sources of molybdenum for Methanococcus maripaludis and their implication for the evolution of molybdoenzymes.

Molybdoenzymes are essential in global nitrogen, carbon, and sulfur cycling. To date, the only known bioavailable source of molybdenum (Mo) is molybdate. However, in the sulfidic and anoxic (euxinic) habitats that predominate in modern subsurface environments and that were pervasive prior to Earth's widespread oxygenation, Mo occurs as soluble tetrathiomolybdate ion and molybdenite mineral that is not known to be bioavailable. This presents a paradox for how organisms obtain Mo to support molybdoenzymes in these environments. Here, we show that tetrathiomolybdate and molybdenite sustain the high Mo demand of a model anaerobic methanogen, Methanococcus maripaludis, grown via Mo-dependent formate dehydrogenase, formylmethanofuran dehydrogenase, and nitrogenase. Cells grown with tetrathiomolybdate and molybdenite have similar growth kinetics, Mo content, and transcript levels of proteins involved in Mo transport and cofactor biosynthesis when compared to those grown with molybdate, implying similar mechanisms of transport and cofactor biosynthesis. These results help to reconcile the paradox of how Mo is acquired in modern and ancient anaerobes and provide new insight into how molybdoenzymes could have evolved prior to Earth's oxygenation.

Covariation of hot spring geochemistry with microbial genomic diversity, function, and evolution.

The geosphere and the microbial biosphere have co-evolved for ~3.8 Ga, with many lines of evidence suggesting a hydrothermal habitat for life's origin. However, the extent that contemporary thermophiles and their hydrothermal habitats reflect those that likely existed on early Earth remains unknown. To address this knowledge gap, 64 geochemical analytes were measured and 1022 metagenome-assembled-genomes (MAGs) were generated from 34 chemosynthetic high-temperature springs in Yellowstone National Park and analysed alongside 444 MAGs from 35 published metagenomes. We used these data to evaluate co-variation in MAG taxonomy, metabolism, and phylogeny as a function of hot spring geochemistry. We found that cohorts of MAGs and their functions are discretely distributed across pH gradients that reflect different geochemical provinces. Acidic or circumneutral/alkaline springs harbor MAGs that branched later and are enriched in sulfur- and arsenic-based O2-dependent metabolic pathways that are inconsistent with early Earth conditions. In contrast, moderately acidic springs sourced by volcanic gas harbor earlier-branching MAGs that are enriched in anaerobic, gas-dependent metabolisms (e.g. H2, CO2, CH4 metabolism) that have been hypothesized to support early microbial life. Our results provide insight into the influence of redox state in the eco-evolutionary feedbacks between thermophiles and their habitats and suggest moderately acidic springs as early Earth analogs.

An active microbiome in Old Faithful geyser.

Natural thermal geysers are hot springs that periodically erupt liquid water, steam, and gas. They are found in only a few locations worldwide, with nearly half located in Yellowstone National Park (YNP). Old Faithful geyser (OFG) is the most iconic in YNP and attracts millions of visitors annually. Despite extensive geophysical and hydrological study of geysers, including OFG, far less is known of the microbiology of geysed waters. Here, we report geochemical and microbiological data from geysed vent water and vent water that collects in a splash pool adjacent to OFG during eruptions. Both waters contained microbial cells, and radiotracer studies showed that they fixed carbon dioxide (CO2) when incubated at 70°C and 90°C. Shorter lag times in CO2 fixation activity were observed in vent and splash pool waters incubated at 90°C than 70°C, suggesting cells are better adapted or acclimated to temperatures like those in the OFG vent (∼92-93°C). 16S rDNA and metagenomic sequence data indicated that both communities are dominated by the autotroph Thermocrinis, which likely fuels productivity through the aerobic oxidation of sulfide/thiosulfate in erupted waters or steam. Dominant OFG populations, including Thermocrinis and subdominant Thermus and Pyrobaculum strains, exhibited high-strain level genomic diversity (putative ecotypes) relative to populations from nongeysing YNP hot springs that is attributed to the temporal chemical and temperature dynamics caused by eruptions. These findings show that OFG is habitable and that its eruption dynamics promote genomic diversity, while highlighting the need to further research the extent of life in geyser systems such as OFG.

Iron Minerals Influence the Assembly of Microbial Communities in a Basaltic Glacial Catchment.

The influence of mineralogy on the assembly of microbial communities in glacial environments has been difficult to assess due to complications in isolating mineralogy from other variables. Here we assess the abundance and composition of microbial communities that colonized defined minerals incubated for 12 months in two meltwater streams (N and S) emanating from Kaldalónsjökull (Kal), a basalt-hosted glacier in Iceland. The two streams shared similar meltwater geochemistry as well as bedrock and proglacial sediment elemental compositions. Yet genomic DNA and PCR-amplifiable 16S rRNA genes were detected only in Kal S. The amount of recoverable DNA was highest for hematite incubated in Kal S and the composition of 16S rRNA genes recovered from Kal S sediments was most like those recovered from hematite and magnetite, an effect driven largely by similarities in the relative abundance of the putative hydrogenotrophic iron reducer Rhodoferax. We suggest this is attributable to comminution and weathering reactions involving exposed iron silicate minerals that generate and release hydrogen and Fe(III) that can be coupled to support microbial metabolism in Kaldalónsjökull, and possibly other basaltic habitats. The low abundance of cells in Kal N could be due to low availability of Fe(III) or another substrate.

An essential role for tungsten in the ecology and evolution of a previously uncultivated lineage of anaerobic, thermophilic Archaea.

Trace metals have been an important ingredient for life throughout Earth's history. Here, we describe the genome-guided cultivation of a member of the elusive archaeal lineage Caldarchaeales (syn. Aigarchaeota), Wolframiiraptor gerlachensis, and its growth dependence on tungsten. A metagenome-assembled genome (MAG) of W. gerlachensis encodes putative tungsten membrane transport systems, as well as pathways for anaerobic oxidation of sugars probably mediated by tungsten-dependent ferredoxin oxidoreductases that are expressed during growth. Catalyzed reporter deposition-fluorescence in-situ hybridization (CARD-FISH) and nanoscale secondary ion mass spectrometry (nanoSIMS) show that W. gerlachensis preferentially assimilates xylose. Phylogenetic analyses of 78 high-quality Wolframiiraptoraceae MAGs from terrestrial and marine hydrothermal systems suggest that tungsten-associated enzymes were present in the last common ancestor of extant Wolframiiraptoraceae. Our observations imply a crucial role for tungsten-dependent metabolism in the origin and evolution of this lineage, and hint at a relic metabolic dependence on this trace metal in early anaerobic thermophiles.

Suspension medium influences interaction of mesenchymal stromal cells with endothelium and pulmonary toxicity after transplantation in mice.

Intravenous (i.v.) transplantation and subsequent homing of Mesenchymal Stromal Cells (MSC) may be adversely influenced by their relatively high adhesion capacity and their tendency to aggregate, leading to clogging of capillaries especially in the lungs. We evaluated the ability of murine MSC suspended in EDTA or heparin in buffered saline solution on their spontaneous adhesion to endothelial cells in vitro, under shear stress and their in vivo tolerability after i.v. injection. We show that suspension of MSC in heparin was highly beneficial, avoiding clinical symptoms in 95% of mice, whereas application of MSC suspended in PBS/EDTA or control buffer caused severe pulmonary reactions and partly, death. In vitro studies using parallel plate flow chambers revealed increased adhesion of MSC suspended in PBS/EDTA to endothelial cells compared with MSC in PBS/heparin. These data provide a means to predict and to interfere with toxicity of i.v. transplanted MSC.

Using multivariate generalizability theory to assess the effect of content stratification on the reliability of a performance assessment.

In recent years, demand for performance assessments has continued to grow. However, performance assessments are notorious for lower reliability, and in particular, low reliability resulting from task specificity. Since reliability analyses typically treat the performance tasks as randomly sampled from an infinite universe of tasks, these estimates of reliability may not be accurate. For tests built according to a table of specifications, tasks are randomly sampled from different strata (content domains, skill areas, etc.). If these strata remain fixed in the test construction process, ignoring this stratification in the reliability analysis results in an underestimate of "parallel forms" reliability, and an overestimate of the person-by-task component. This research explores the effect of representing and misrepresenting the stratification appropriately in estimation of reliability and the standard error of measurement. Both multivariate and univariate generalizability studies are reported. Results indicate that the proper specification of the analytic design is essential in yielding the proper information both about the generalizability of the assessment and the standard error of measurement. Further, illustrative D studies present the effect under a variety of situations and test designs. Additional benefits of multivariate generalizability theory in test design and evaluation are also discussed.

Using standardized patients to assess professionalism: a generalizability study.

BACKGROUND: Assessment of professionalism in undergraduate medical education is challenging. One approach that has not been well studied in this context is performance-based examinations. PURPOSE: This study sought to investigate the reliability of standardized patients' scores of students' professionalism in performance-based examinations. METHODS: Twenty students were observed on 4 simulated cases involving professional challenges; 9 raters evaluated each encounter on 21 professionalism items. Correlational and multivariate generalizability (G) analyses were conducted. RESULTS: G coefficients were .75, .53, and .68 for physicians, standardized patients (SPs), and lay raters, respectively. Composite G coefficient for all raters reached acceptable level of .86. Results indicated SP raters were more variable than other rater types in severity with which they rated students, although rank ordering of students was consistent among SPs. CONCLUSIONS: SPs' ratings were less reliable and consistent than physician or lay ratings, although the SPs rank ordered students more consistently than the other rater types.

Community Service as Patients' Tutor to Avoid Delirium. / Ehrenamtliche Patientenbegleiter zur Delirvermeidung.

BACKGROUND: Today, nearly two thirds of inpatients in trauma surgery hospitals are over 70 years old and at significant risk for comorbidities. These patients frequently suffer delirium. Delirium occurs in 15 - 30% of these trauma patients and increases the risk of mortality in up to 15 - 25% of cases. Conversation, attention, and activity significantly reduce this risk. OBJECTIVES: Thus, the Centre for Geriatric Traumatology and the District Seniors Council have initiated a new project of a visiting service, to prevent delirium and anxiety conditions in elderly inpatients. Volunteers from the welfare program offer daily care for selected patients. METHODS: They care for and interact with the patients for approximately one hour per day. They accompany the patients in the hospital and during mobilization, explain, read aloud, converse, or simply listen, and thus provide calm, support, and company. The attendants are volunteers who have been previously trained by the clinic, who regularly attend team meetings and draw up reports from each admission. All patients and relatives are informed prior to these visits and are notified with an accompanying letter. There are structured visiting plans for each week. RESULTS: From July 17 to May 19, 4031 patients in our clinic and in three other hospitals have been accompanied in the program. None of the patients suffered delirium. The patients and attendants found the care very good and personally satisfying. Evaluations of the volunteers for the success of their work, using the German school-grading system, was an average 1.3. Noticeable was the markedly reduced frequency of patient calls to nursing, who were thus relieved. Travel expenses and insurance costs for the volunteers were reimbursed. CONCLUSION: In times of skilled labour shortages, patient support provided by volunteer patient attendants enables innovative care for patients at risk for delirium. The assistance of the attendants alone reduced the workload on the nursing staff.

Efficacy of adding ramipril (VAsotop) to the combination of furosemide (Lasix) and pimobendan (VEtmedin) in dogs with mitral valve degeneration: The VALVE trial.

BACKGROUND: Triple therapy (TT) consisting of furosemide, pimobendan, and an angiotensin-converting enzyme inhibitor (ACEI) frequently is recommended for the treatment of congestive heart failure (CHF) attributable to myxomatous mitral valve disease (MMVD). However, the effect of adding an ACEI to the combination of pimobendan and furosemide (dual therapy [DT]) so far has not been evaluated prospectively. HYPOTHESIS: Triple therapy will extend survival time compared to DT in dogs with CHF secondary to MMVD. ANIMALS: Client-owned dogs presented with the first episode of CHF caused by MMVD. METHODS: Prospective, single-blinded, randomized multicenter study. One-hundred and fifty-eight dogs were recruited and prospectively randomized to receive either DT (furosemide and pimobendan) or TT (furosemide, pimobendan, and ramipril). The primary endpoint was a composite of cardiac death, euthanasia for heart failure, or treatment failure. RESULTS: Seventy-seven dogs were randomized to receive DT and 79 to receive TT. Two dogs were excluded from analysis. The primary endpoint was reached by 136 dogs (87%; 66 dogs, DT; 70 dogs, TT). Median time to reach the primary endpoint for all dogs in the study was 214 days (95% confidence interval [CI], 168-259 days). Median time to reach the primary endpoint was not significantly different between the DT group (227 days; interquartile range [IQR], 103-636 days) compared with TT group (186 days; IQR, 72-453 days; P = .42). CONCLUSIONS AND CLINICAL IMPORTANCE: Addition of the ACEI ramipril to pimobendan and furosemide did not have any beneficial effect on survival time in dogs with CHF secondary to MMVD.

PDGFA-associated protein 1 protects mature B lymphocytes from stress-induced cell death and promotes antibody gene diversification.

The establishment of protective humoral immunity is dependent on the ability of mature B cells to undergo antibody gene diversification while adjusting to the physiological stressors induced by activation with the antigen. Mature B cells diversify their antibody genes by class switch recombination (CSR) and somatic hypermutation (SHM), which are both dependent on efficient induction of activation-induced cytidine deaminase (AID). Here, we identified PDGFA-associated protein 1 (Pdap1) as an essential regulator of cellular homeostasis in mature B cells. Pdap1 deficiency leads to sustained expression of the integrated stress response (ISR) effector activating transcription factor 4 (Atf4) and induction of the ISR transcriptional program, increased cell death, and defective AID expression. As a consequence, loss of Pdap1 reduces germinal center B cell formation and impairs CSR and SHM. Thus, Pdap1 protects mature B cells against chronic ISR activation and ensures efficient antibody diversification by promoting their survival and optimal function.

Influence of physical activity on the immune system in breast cancer patients during chemotherapy.

PURPOSE: Physical activity can impact the immune system in different ways, e.g. by alteration of the humoral and cellular immune response. Physical activity at medium intensity enhances numbers of cytotoxic T cells, NK cells and macrophages in healthy people. The aim of this study was to compare the effects of endurance and resistance training on the immune system in breast cancer patients during adjuvant chemotherapy. METHODS: In a prospective, controlled and randomized intervention exploratory trial, 12-week supervised endurance or resistance training were compared with usual care twice a week. Endpoints were the absolute numbers of the immune cells such as CD3+ T lymphocytes including CD4+- and CD8+, αß T cells, γδT cells, CD3-/CD16+/56+ NK cells and CD19+ B cells, before and after 12 weeks of treatment. Cell numbers were analyzed using fluorescence-activated cell sorting. RESULTS: Despite different physical interventions in all groups immune cell count decreased in CD3 T cells including TCR αß and CD4 T cells, NK cells and CD19 B cells 12 weeks after initiation of chemotherapy and start of the physical intervention program, while the reduction of γδ T cells and CD8 T cells is less prominent in the RT and UC group. CONCLUSION: Chemotherapy led to a decrease in nearly all measured immune cells. In this study, physical intervention with endurance or resistance training did not suppress cellular immunity any further. Larger multicenter trials are needed to evaluate the exact impact of sports intervention on immune cell subpopulations.