Assessment of the corneal collagen organization after chemical burn using second harmonic generation microscopy

The organization of the corneal stoma is modified due to different factors, including pathology, surgery or external damage. Here the changes in the organization of the corneal collagen fibers during natural healing after chemical burn are investigated using second harmonic generation (SHG) imaging. Moreover, the structure tensor (ST) was used as an objective tool for morphological analyses at different time points after burn (up to 6 months). Unlike control corneas that showed a regular distribution, the collagen pattern at 1 month of burn presented a non-organized arrangement. SHG signal levels noticeably decreased and individual fibers were hardly visible. Over time, the healing process led to a progressive re-organization of the fibers that could be quantified through the ST. At 6 months, the stroma distribution reached values similar to those of control eyes and a dominant direction of the fibers re-appeared. The present results show that SHG microscopy imaging combined with the ST method is able to objectively monitor the temporal regeneration of the corneal organization after chemical burn. Future implementations of this approach into clinically adapted devices would help to diagnose and quantify corneal changes, not only due to chemical damages, but also as a result of disease or surgical procedures

HIV infection and aging: enhanced Interferon- and Tumor Necrosis Factor-alpha production by the CD8(+) CD28(-) T subset

BACKGROUND: T cells from HIV(+) and aged individuals show parallels in terms of suppressed proliferative activity and interleukin-2 (I1-2) production and an increased number of CD8(+) CD28(-) T cells. In order to compare cytokine production from T cells from these two states, CD4(+) and CD8(+) T cells from HIV(+) aged, and normal young donors (controls) were monitored for cytokine production by flow cytometry, quantitative PCR and ELISA upon activation by PMA and anti-CD3. In addition, the CD8(+) T cell subsets CD28(+) and CD28(-) from the HIV(+) and the aged groups were evaluated for cytokine production by flow cytometry, and compared with those from young controls. RESULTS: Flow cytometric analysis indicated that CD8(+) T cells from both HIV(+) and aged donors showed an increase of approximately 2–3 fold over controls in percentage of cells producing inflammatory cytokines IFN-γ and TNF-α. Similar analysis also revealed that the production of interleukins-4,6 and 10, production was very low (1–2% of cells) and unchanged in these cells. Quantitative PCR also showed a substantial increase (4–5 fold) in IFN-γ and TNF-α mRNA from HIV(+) and aged CD8(+) T cells, as did ELISA for secreted IFN-γ and TNF-α (2.3–4 fold). Flow cytometric analysis showed that the CD8(+) CD28(-) T cell subset accounts for approximately 80–86% of the IFN-γ and TNF-α production from the CD8(+) subset in the aged and HIV(+) states. The CD4(+) T cell, while not significantly changed in the HIV(+) or aged states in terms of IFN-γ production, showed a small but significant increase in TNF-α production in both states. CONCLUSIONS: Our data appear compatible with physiologic conditions existing in HIV(+) and aged individuals, i.e. elevated serum levels and elevated CD8(+) T cell production of IFN-γ and TNF-α. Thus, the capacity for increased production of cytokines IFN-γ and TNF-α in the aged individual by the dominant CD8(+) CD28(-) subset may have a profound influence on the clinical state by aggravating inflammatory pathologies such as rheumatoid arthritis, and possibly Alzheimer's disease and Crohn's disease. In AIDS, these cytokines may contribute to wasting and cachexia. We theorize that the predominant phenotypic change to the cytotoxic CD8(+) CD28(-) T cell subsets in both the HIV(+) and the aged states may reflect a natural "endpoint" in CD8(+) T cell differentiation induced after a lifetime of immune activity (toward viruses, etc) in the aged, and after a massive accelerated response to HIV in the HIV-positive individual

Drug delivery nanosystems for the localized treatment of glioblastoma multiforme

[EN] Glioblastoma multiforme is one of the most prevalent and malignant forms of central nervous system tumors. The treatment of glioblastoma remains a great challenge due to its location in the intracranial space and the presence of the blood-brain tumor barrier. There is an urgent need to develop novel therapy approaches for this tumor, to improve the clinical outcomes, and to reduce the rate of recurrence and adverse effects associated with present options. The formulation of therapeutic agents in nanostructures is one of the most promising approaches to treat glioblastoma due to the increased availability at the target site, and the possibility to co-deliver a range of drugs and diagnostic agents. Moreover, the local administration of nanostructures presents significant additional advantages, since it overcomes blood-brain barrier penetration issues to reach higher concentrations of therapeutic agents in the tumor area with minimal side effects. 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Ecosystem-bedrock interaction changes nutrient compartmentalization during early oxidative weathering

Ecosystem-bedrock interactions power the biogeochemical cycles of Earth's shallow crust, supporting life, stimulating substrate transformation, and spurring evolutionary innovation. While oxidative processes have dominated half of terrestrial history, the relative contribution of the biosphere and its chemical fingerprints on Earth's developing regolith are still poorly constrained. Here, we report results from a two-year incipient weathering experiment. We found that the mass release and compartmentalization of major elements during weathering of granite, rhyolite, schist and basalt was rock-specific and regulated by ecosystem components. A tight interplay between physiological needs of different biota, mineral dissolution rates, and substrate nutrient availability resulted in intricate elemental distribution patterns. Biota accelerated CO2 mineralization over abiotic controls as ecosystem complexity increased, and significantly modified stoichiometry of mobilized elements. Microbial and fungal components inhibited element leaching (23.4% and 7%), while plants increased leaching and biomass retention by 63.4%. All biota left comparable biosignatures in the dissolved weathering products. Nevertheless, the magnitude and allocation of weathered fractions under abiotic and biotic treatments provide quantitative evidence for the role of major biosphere components in the evolution of upper continental crust, presenting critical information for large-scale biogeochemical models and for the search for stable in situ biosignatures beyond Earth.Comment: 41 pages (MS, SI and Data), 16 figures (MS and SI), 6 tables (SI and Data). Journal article manuscrip

Biofilm formation as a novel phenotypic feature of adherent-invasive Escherichia coli (AIEC)

<p>Abstract</p> <p>Background</p> <p>Crohn's disease (CD) is a high morbidity chronic inflammatory disorder of unknown aetiology. Adherent-invasive <it>Escherichia coli </it>(AIEC) has been recently implicated in the origin and perpetuation of CD. Because bacterial biofilms in the gut mucosa are suspected to play a role in CD and biofilm formation is a feature of certain pathogenic <it>E. coli </it>strains, we compared the biofilm formation capacity of 27 AIEC and 38 non-AIEC strains isolated from the intestinal mucosa. Biofilm formation capacity was then contrasted with the AIEC phenotype, the serotype, the phylotype, and the presence of virulence genes.</p> <p>Results</p> <p>Specific biofilm formation (SBF) indices were higher amongst AIEC than non-AIEC strains (P = 0.012). In addition, 65.4% of moderate to strong biofilms producers were AIEC, whereas 74.4% of weak biofilm producers were non-AIEC (P = 0.002). These data indicate that AIEC strains were more efficient biofilm producers than non-AIEC strains. Moreover, adhesion (P = 0.009) and invasion (P = 0.003) indices correlated positively with higher SBF indices. Additionally, motility (100%, P < 0.001), H1 type flagellin (53.8%, P < 0.001), serogroups O83 (19.2%, P = 0.008) and O22 (26.9%, P = 0.001), the presence of virulence genes such as <it>sfa/focDE </it>(38.5%, P = 0.003) and <it>ibeA </it>(26.9%, P = 0.017), and B2 phylotype (80.8%, P < 0.001) were frequent characteristics amongst biofilm producers.</p> <p>Conclusion</p> <p>The principal contribution of the present work is the finding that biofilm formation capacity is a novel, complementary pathogenic feature of the recently described AIEC pathovar. Characterization of AIEC specific genetic determinants, and the regulatory pathways, involved in biofilm formation will likely bring new insights into AIEC pathogenesis.</p

Cardiotrophin-1 defends the liver against ischemia-reperfusion injury and mediates the protective effect of ischemic preconditioning

Ischemia-reperfusion (I/R) liver injury occurs when blood flow is restored after prolonged ischemia. A short interruption of blood flow (ischemic preconditioning [IP]) induces tolerance to subsequent prolonged ischemia through ill-defined mechanisms. Cardiotrophin (CT)-1, a cytokine of the interleukin-6 family, exerts hepatoprotective effects and activates key survival pathways like JAK/STAT3. Here we show that administration of CT-1 to rats or mice protects against I/R liver injury and that CT-1-deficient mice are exceedingly sensitive to this type of damage. IP markedly reduced transaminase levels and abrogated caspase-3 and c-Jun-NH2-terminal kinase activation after I/R in normal mice but not in CT-1-null mice. Moreover, the protective effect afforded by IP was reduced by previous administration of neutralizing anti-CT-1 antibody. Prominent STAT3 phosphorylation in liver tissue was observed after IP plus I/R in normal mice but not in CT-1-null mice. Oxidative stress, a process involved in IP-induced hepatoprotection, was found to stimulate CT-1 release from isolated hepatocytes. Interestingly, brief ischemia followed by short reperfusion caused mild serum transaminase elevation and strong STAT3 activation in normal and IL-6-deficient mice, but failed to activate STAT3 and provoked marked hypertransaminasemia in CT-1-null animals. In conclusion, CT-1 is an essential endogenous defense of the liver against I/R and is a key mediator of the protective effect induced by IP

RAF1 kinase activity is dispensable for KRAS/p53 mutant lung tumor progression.

We thank Dr. Shiva Malek and her colleagues (Genentech Inc.) for sharing their results with us before publication. We also thank M. San Roman, R. Villar, M.C. Gonzalez, A. Lopez, N. Cabrera, P. Villanueva, J. Condo, O. Dominguez, and S. Ortega for excellent technical support. This work was supported by grants from the European Research Council (ERC-2015-AdG/695566, THERACAN); the Spanish Ministry of Science, Innovation, and Universities (RTC-2017-6576-1 and RTI2018094664-B-I00) and the Autonomous Community of Madrid (B2017/BMD-3884 iLUNG-CM) to M.B., as well as by a grant from the Spanish Ministry of Science, Innovation and Universities (RTI2018-094664-B-I00) to M.B. and M.M. M.B. is a recipient of an Endowed Chair from the AXA Research Fund. M.S., P.N., and F.F.-G. were supported by FPU fellowships from the Spanish Ministry of Education. L.E.-B. was a recipient of an FPI fellowship from the Spanish Ministry of Economy and Competitiveness. S.G.-A. is a recipient of a postdoctoral fellowship from the Asociacion Espanola Contra el Cancer (AECC).S

Egg consumption and dyslipidemia in a Mediterranean cohort

Introduction and objectives: Our aim was to prospectively evaluate the association between egg consumption and dyslipidemia in a Mediterranean cohort. Methods: We followed-up 13,104 Spanish university graduates for a mean period of 8 years. Dietary habits at baseline were assessed using a validated semi-quantitative 136-item food-frequency questionnaire. Self-reported blood concentrations of total cholesterol, high-density lipoproteins cholesterol (HDL-c) and triglycerides were evaluated according to categories of egg consumption after 6 and 8 years of follow-up. We also assessed the association between baseline egg consumption and the incidence of hypercholesterolemia, low HDL-c concentrations and hypertriglyceridemia during follow-up. Results: We observed a significant inverse association for intermediate levels of egg consumption (2 to 4 eggs/week vs. less than 1 egg/week) and hypertriglyceridemia with OR = 0.71 (95% confidence interval [CI]: 0.54 to 0.93, p < 0.05) in the multivariable-adjusted model. Using HDL-c values after 8-year follow-up, we found an association between higher egg consumption and lower HDL-c levels (p for trend = 0.02) with an adjusted difference of –4.01 mg/dl (-7.42 to -0.61) for > 4 vs. < 1 egg/week. Lower means of triglycerides were found in each of the three upper categories of egg consumption compared to the lowest category (< 1 egg/week) with significant results for some of these categories both after 6 and 8 year follow-up. Conclusions: Our data do not support that higher egg consumption was associated with abnormal blood levels of total cholesterol or triglycerides; an inverse association with HDL-c as a quantitative variable was found only in one of our analyses.Introducción y objetivos: evaluar prospectivamente la asociación entre el consumo de huevo y el riesgo de dislipidemia en una cohorte mediterránea. Métodos: se siguieron 13.104 graduados universitarios españoles durante un periodo medio de 8 años. La dieta se evaluó al inicio utilizando un cuestionario semicuantitativo de frecuencia de consumo de alimentos repetidamente validado. Las concentraciones sanguíneas de colesterol total, lipoproteínas de alta densidad (HDL-c) y triglicéridos autorreferidas fueron evaluadas según categorías de consumo de huevo tras 6 y 8 años de seguimiento. También se evaluó la asociación entre el consumo basal de huevo y la incidencia de hipercolesterolemia, concentraciones bajas de HDL-c e hipertrigliceridemia durante el seguimiento. Resultados: se observó una asociación entre los niveles intermedios de consumo de huevo (2-4 unidades/semana frente a < 1 unidad/semana) y menor riesgo de hipertrigliceridemia con OR = 0,71 (intervalo de confianza del 95% [IC]: 0,54 a 0,93, p < 0,05) en el modelo más ajustado. Tras 8 años de seguimiento, encontramos una asociación entre un mayor consumo de huevo y menores niveles de HDL-c (p tendencia lineal = 0,02) con una diferencia ajustada de -4,01 mg/dl (-7,42 a -0,61) para > 4 vs. < 1 unidad/semana. Se encontraron menores concentraciones de triglicéridos en las tres categorías superiores de consumo de huevo en comparación con la inferior con resultados significativos para algunas de estas categorías después de 6 y 8 años de seguimiento. Conclusiones: un mayor consumo de huevo no se asoció con niveles anormales de colesterol total o triglicéridos; se encontró una asociación inversa con HDL-c como variable cuantitativa solo en uno de nuestros análisis

Tumor regression and resistance mechanisms upon CDK4 and RAF1 inactivation in KRAS/P53 mutant lung adenocarcinomas.

KRAS mutant lung adenocarcinomas remain intractable for targeted therapies. Genetic interrogation of KRAS downstream effectors, including the MAPK pathway and the interphase CDKs, identified CDK4 and RAF1 as the only targets whose genetic inactivation induces therapeutic responses without causing unacceptable toxicities. Concomitant CDK4 inactivation and RAF1 ablation prevented tumor progression and induced complete regression in 25% of KRAS/p53-driven advanced lung tumors, yet a significant percentage of those tumors that underwent partial regression retained a population of CDK4/RAF1-resistant cells. Characterization of these cells revealed two independent resistance mechanisms implicating hypermethylation of several tumor suppressors and increased PI3K activity. Importantly, these CDK4/RAF1-resistant cells can be pharmacologically controlled. These studies open the door to new therapeutic strategies to treat KRAS mutant lung cancer, including resistant tumors.We thank S. Ortega for the generation of the Cdk4FxKD mouse model; and M. San Roman, R. Villar, M. C. Gonzalez, A. Lopez, N. Cabrera, P. Villanueva, J. Condo, J. Klett, A. Cebria, A. Otero, O. Dominguez, G. Luengo, G. Garaulet, F. Mulero, and D. Megias for excellent technical support. This work was supported by European Research Council Grant ERC-2015-AdG/695566, THERACAN, Spanish Ministry of Science, Innovation, and Universities Grant RTC-2017-6576-1, and the Autonomous Community of Madrid Grant B2017/BMD-3884 iLUNG-CM (to M.B.); Spanish Ministry of Science, Innovation, and Universities Grant RTI2018-094664B-I00 (to M.B. and M.M.); and National Natural Science Foundation of China Grant 31771469 (to H.W.). M.B. is a recipient of an Endowed Chair from the AXA Research Fund. L.E.-B. is the recipient of an FPI fellowship from the Spanish Ministry of Economy and Competitiveness. F.F.-G., M.S., and P.N. were supported by an FPU fellowships from the Spanish Ministry of Education.S

Molecular features in a biphenotypic small cell sarcoma with neuroectodermal and muscle differentiation

We report a case of a 13-year-old girl with soft tissue sarcoma of the hand, which showed muscle and neuroectodermal immunophenotypes. Molecular studies were performed on RNA collected from fine-needle aspiration (FNA) cytology and peripheral blood samples by nested reverse transcriptase-polymerase chain reaction (RT-PCR) and Southern blot analysis. This biphenotypic tumor showed simultaneous expression of EWS-FLI1 and PAX3-FKHR transcripts, specific of Ewing family tumors and alveolar rhabdomyosarcoma, respectively. Although childhood sarcomas with simultaneous muscle and neural differentiation have been described to have EWS-FLI1 transcripts, there are no reports of tumors with both transcripts. Cytological specimens are a good source of RNA for molecular studie