Resuscitation of Newborn Piglets. Short-Term Influence of FiO2 on Matrix Metalloproteinases, Caspase-3 and BDNF

Perinatal hypoxia-ischemia is a major cause of mortality and cerebral morbidity, and using oxygen during newborn resuscitation may further harm the brain. The aim was to examine how supplementary oxygen used for newborn resuscitation would influence early brain tissue injury, cell death and repair processes and the regulation of genes related to apoptosis, neurodegeneration and neuroprotection.Anesthetized newborn piglets were subjected to global hypoxia and then randomly assigned to resuscitation with 21%, 40% or 100% O(2) for 30 min and followed for 9 h. An additional group received 100% O(2) for 30 min without preceding hypoxia. The left hemisphere was used for histopathology and immunohistochemistry and the right hemisphere was used for in situ zymography in the corpus striatum; gene expression and the activity of various relevant biofactors were measured in the frontal cortex. There was an increase in the net matrix metalloproteinase gelatinolytic activity in the corpus striatum from piglets resuscitated with 100% oxygen vs. 21%. Hematoxylin-eosin (HE) staining revealed no significant changes. Nine hours after oxygen-assisted resuscitation, caspase-3 expression and activity was increased by 30-40% in the 100% O(2) group (n = 9/10) vs. the 21% O(2) group (n = 10; p<0.04), whereas brain-derived neurotrophic factor (BDNF) activity was decreased by 65% p<0.03.The use of 100% oxygen for resuscitation resulted in increased potentially harmful proteolytic activities and attenuated BDNF activity when compared with 21%. Although there were no significant changes in short term cell loss, hyperoxia seems to cause an early imbalance between neuroprotective and neurotoxic mechanisms that might compromise the final pathological outcome

Local Induction of Immunosuppressive CD8+ T Cells in the Gut-Associated Lymphoid Tissues

Background: In contrast to intestinal CD4 + regulatory T cells (Tregs), the generation and function of immunomodulatory intestinal CD8 + T cells is less well defined. To dissect the immunologic mechanisms of CD8 + T cell function in the mucosa, reactivity against hemagglutinin (HA) expressed in intestinal epithelial cells of mice bearing a MHC class-I-restricted T-cellreceptor specific for HA was studied. Methodology and Principal Findings: HA-specific CD8 + T cells were isolated from gut-associated tissues and phenotypically and functionally characterized for the expression of Foxp3 + and their suppressive capacity. We demonstrate that intestinal HA expression led to peripheral induction of HA-specific CD8 + Foxp3 + T cells. Antigen-experienced CD8 + T cells in this transgenic mouse model suppressed the proliferation of CD8 + and CD4 + T cells in vitro. Gene expression analysis of suppressive HA-specific CD8 + T cells revealed a specific up-regulation of CD103, Nrp1, Tnfrsf9 and Pdcd1, molecules also expressed on CD4 + T reg subsets. Finally, gut-associated dendritic cells were able to induce HA-specific CD8 + Foxp3 + T cells. Conclusion and Significance: We demonstrate that gut specific antigen presentation is sufficient to induce CD8 + T regs in vivo which may maintain intestinal homeostasis by down-modulating effector functions of T cells

Baicalin Improves Survival in a Murine Model of Polymicrobial Sepsis via Suppressing Inflammatory Response and Lymphocyte Apoptosis

BACKGROUND: An imbalance between overwhelming inflammation and lymphocyte apoptosis is the main cause of high mortality in patients with sepsis. Baicalin, the main active ingredient of the Scutellaria root, exerts anti-inflammatory, anti-apoptotic, and even antibacterial properties in inflammatory and infectious diseases. However, the therapeutic effect of baicalin on polymicrobial sepsis remains unknown. METHODOLOGY/PRINCIPAL FINDINGS: Polymicrobial sepsis was induced by cecal ligation and puncture (CLP) in C57BL/6 mice. Mice were infused with baicalin intraperitoneally at 1 h, 6 h and 12 h after CLP. Survival rates were assessed over the subsequent 8 days. Bacterial burdens in blood and peritoneal cavity were calculated to assess the bacterial clearance. Neutrophil count in peritoneal lavage fluid was also calculated. Injuries to the lung and liver were detected by hematoxylin and eosin staining. Levels of cytokines, including tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, IL-10 and IL-17, in blood and peritoneum were measured by enzyme-linked immunosorbent assay. Adaptive immune function was assessed by apoptosis of lymphocytes in the thymus and counts of different cell types in the spleen. Baicalin significantly enhanced bacterial clearance and improved survival of septic mice. The number of neutrophils in peritoneal lavage fluid was reduced by baicalin. Less neutrophil infiltration of the lung and liver in baicalin-treated mice was associated with attenuated injuries to these organs. Baicalin significantly reduced the levels of proinflammatory cytokines but increased the level of anti-inflammatory cytokine in blood and peritoneum. Apoptosis of CD3(+) T cell was inhibited in the thymus. The numbers of CD4(+), CD8(+) T lymphocytes and dendritic cells (DCs) were higher, while the number of CD4(+)CD25(+) regulatory T cells was lower in the baicalin group compared with the CLP group. CONCLUSIONS/SIGNIFICANCE: Baicalin improves survival of mice with polymicrobial sepsis, and this may be attributed to its antibacterial property as well as its anti-inflammatory and anti-apoptotic effects

Factor structure of the Hospital Anxiety and Depression Scale in Japanese psychiatric outpatient and student populations

<p>Abstract</p> <p>Background</p> <p>The Hospital Anxiety and Depression Scale (HADS) is a common screening instrument excluding somatic symptoms of depression and anxiety, but previous studies have reported inconsistencies of its factor structure. The construct validity of the Japanese version of the HADS has yet to be reported. To examine the factor structure of the HADS in a Japanese population is needed.</p> <p>Methods</p> <p>Exploratory and confirmatory factor analyses were conducted in the combined data of 408 psychiatric outpatients and 1069 undergraduate students. The data pool was randomly split in half for a cross validation. An exploratory factor analysis was performed on one half of the data, and the fitness of the plausible model was examined in the other half of the data using a confirmatory factor analysis. Simultaneous multi-group analyses between the subgroups (outpatients vs. students, and men vs. women) were subsequently conducted.</p> <p>Results</p> <p>A two-factor model where items 6 and 7 had dual loadings was supported. These factors were interpreted as reflecting anxiety and depression. Item 10 showed low contributions to both of the factors. Simultaneous multi-group analyses indicated a factor pattern stability across the subgroups.</p> <p>Conclusion</p> <p>The Japanese version of HADS indicated good factorial validity in our samples. However, ambiguous wording of item 7 should be clarified in future revisions.</p

New Model of Macrophage Acquisition of the Lymphatic Endothelial Phenotype

Macrophage-derived lymphatic endothelial cell progenitors (M-LECPs) contribute to new lymphatic vessel formation, but the mechanisms regulating their differentiation, recruitment, and function are poorly understood. Detailed characterization of M-LECPs is limited by low frequency in vivo and lack of model systems allowing in-depth molecular analyses in vitro. Our goal was to establish a cell culture model to characterize inflammation-induced macrophage-to-LECP differentiation under controlled conditions.Time-course analysis of diaphragms from lipopolysaccharide (LPS)-treated mice revealed rapid mobilization of bone marrow-derived and peritoneal macrophages to the proximity of lymphatic vessels followed by widespread (∼50%) incorporation of M-LECPs into the inflamed lymphatic vasculature. A differentiation shift toward the lymphatic phenotype was found in three LPS-induced subsets of activated macrophages that were positive for VEGFR-3 and many other lymphatic-specific markers. VEGFR-3 was strongly elevated in the early stage of macrophage transition to LECPs but undetectable in M-LECPs prior to vascular integration. Similar transient pattern of VEGFR-3 expression was found in RAW264.7 macrophages activated by LPS in vitro. Activated RAW264.7 cells co-expressed VEGF-C that induced an autocrine signaling loop as indicated by VEGFR-3 phosphorylation inhibited by a soluble receptor. LPS-activated RAW264.7 macrophages also showed a 68% overlap with endogenous CD11b(+)/VEGFR-3(+) LECPs in the expression of lymphatic-specific genes. Moreover, when injected into LPS- but not saline-treated mice, GFP-tagged RAW264.7 cells massively infiltrated the inflamed diaphragm followed by integration into 18% of lymphatic vessels.We present a new model for macrophage-LECP differentiation based on LPS activation of cultured RAW264.7 cells. This system designated here as the "RAW model" mimics fundamental features of endogenous M-LECPs. Unlike native LECPs, this model is unrestricted by cell numbers, heterogeneity of population, and ability to change genetic composition for experimental purposes. As such, this model can provide a valuable tool for understanding the LECP and lymphatic biology

Characterization of Bioactive Recombinant Human Lysozyme Expressed in Milk of Cloned Transgenic Cattle

BACKGROUND: There is great potential for using transgenic technology to improve the quality of cow milk and to produce biopharmaceuticals within the mammary gland. Lysozyme, a bactericidal protein that protects human infants from microbial infections, is highly expressed in human milk but is found in only trace amounts in cow milk. METHODOLOGY/PRINCIPAL FINDINGS: We have produced 17 healthy cloned cattle expressing recombinant human lysozyme using somatic cell nuclear transfer. In this study, we just focus on four transgenic cattle which were natural lactation. The expression level of the recombinant lysozyme was up to 25.96 mg/L, as measured by radioimmunoassay. Purified recombinant human lysozyme showed the same physicochemical properties, such as molecular mass and bacterial lysis, as its natural counterpart. Moreover, both recombinant and natural lysozyme had similar conditions for reactivity as well as for pH and temperature stability during in vitro simulations. The gross composition of transgenic and non-transgenic milk, including levels of lactose, total protein, total fat, and total solids were not found significant differences. CONCLUSIONS/SIGNIFICANCE: Thus, our study not only describes transgenic cattle whose milk offers the similar nutritional benefits as human milk but also reports techniques that could be further refined for production of active human lysozyme on a large scale

Unraveling infectious structures, strain variants and species barriers for the yeast prion [PSI+]

Prions are proteins that can access multiple conformations, at least one of which is beta-sheet rich, infectious and self-perpetuating in nature. These infectious proteins show several remarkable biological activities, including the ability to form multiple infectious prion conformations, also known as strains or variants, encoding unique biological phenotypes, and to establish and overcome prion species (transmission) barriers. In this Perspective, we highlight recent studies of the yeast prion [PSI+], using various biochemical and structural methods, that have begun to illuminate the molecular mechanisms by which self-perpetuating prions encipher such biological activities. We also discuss several aspects of prion conformational change and structure that remain either unknown or controversial, and we propose approaches to accelerate the understanding of these enigmatic, infectious conformers

20-Year Risks of Breast-Cancer Recurrence after Stopping Endocrine Therapy at 5 Years

The administration of endocrine therapy for 5 years substantially reduces recurrence rates during and after treatment in women with early-stage, estrogen-receptor (ER)-positive breast cancer. Extending such therapy beyond 5 years offers further protection but has additional side effects. Obtaining data on the absolute risk of subsequent distant recurrence if therapy stops at 5 years could help determine whether to extend treatment