Comparison of Inflammatory Cytokine Levels in Hepatic and Jugular Veins of Patients with Cirrhosis.

Background: Systemic inflammation with elevated inflammatory cytokines is a hallmark in patients with cirrhosis and the main driver of decompensation. There is insufficient data on whether inflammatory cytokine levels differ between hepatic and jugular veins, which may have implications for further immunological studies. Methods: Blood from the hepatic and jugular veins of 40 patients with cirrhosis was collected during hepatic venous pressure gradient (HVPG) measurements. Serum levels of 13 inflammatory cytokines (IL-1ß, Int-α2, Int-γ, TNF-α, MCP-1, IL-6, IL-8, IL-10, IL-12p70, IL-17A, IL-18, IL-23, and IL-33) were quantified by cytometric bead array. Results: Cytokine levels of IFN-α2, IFN-γ, TNF-α, IL-6, IL-8, IL-10, IL-17A, IL-18, IL-23, and IL-33 were significantly elevated in patients with decompensated cirrhosis compared to patients with compensated cirrhosis. When comparing patients with clinically significant portal hypertension (CSPH, HVPG ≥ 10 mmHg) to patients without CSPH, there were significantly enhanced serum levels of IL-6 and IL-18 in the former group. There was no significant difference between cytokine serum levels between blood obtained from the jugular versus hepatic veins. Even in subgroup analyses stratified for an early cirrhosis stage (Child-Pugh (CP) A) or more decompensated stages (CP B/C), cytokine levels were similar. Conclusion: Cytokine levels increase with decompensation and increasing portal hypertension in patients with cirrhosis. There is no relevant difference in cytokine levels between hepatic and jugular blood in patients with cirrhosis.

Transcriptional Targeting of Dendritic Cells Using an Optimized Human Fascin1 Gene Promoter.

Deeper knowledge about the role of the tumor microenvironment (TME) in cancer development and progression has resulted in new strategies such as gene-based cancer immunotherapy. Whereas some approaches focus on the expression of tumoricidal genes within the TME, DNA-based vaccines are intended to be expressed in antigen-presenting cells (e.g., dendritic cells, DCs) in secondary lymphoid organs, which in turn induce anti-tumor T cell responses. Besides effective delivery systems and the requirement of appropriate adjuvants, DNA vaccines themselves need to be optimized regarding efficacy and selectivity. In this work, the concept of DC-focused transcriptional targeting was tested by applying a plasmid encoding for the luciferase reporter gene under the control of a derivative of the human fascin1 gene promoter (pFscnLuc), comprising the proximal core promoter fused to the normally more distantly located DC enhancer region. DC-focused activity of this reporter construct was confirmed in cell culture in comparison to a standard reporter vector encoding for luciferase under the control of the strong ubiquitously active cytomegalovirus promoter and enhancer (pCMVLuc). Both plasmids were also compared upon intravenous administration in mice. The organ- and cell type-specific expression profile of pFscnLuc versus pCMVLuc demonstrated favorable activity especially in the spleen as a central immune organ and within the spleen in DCs.

Peptide-Decorated Degradable Polycarbonate Nanogels for Eliciting Antigen-Specific Immune Responses.

For successful therapeutic interventions in cancer immunotherapy, strong antigen-specific immune responses are required. To this end, immunostimulating cues must be combined with antigens to simultaneously arrive at antigen-presenting cells and initiate cellular immune responses. Recently, imidazoquinolines have shown their vast potential as small molecular Toll-like receptor 7/8 (TLR7/8) agonists for immunostimulation when delivered by nanocarriers. At the same time, peptide antigens are promising antigen candidates but require combination with immune-stimulating adjuvants to boost their immunogenicity and exploit their full potential. Consequently, we herein present biodegradable polycarbonate nanogels as versatile delivery system for adjuvants within the particles' core as well as for peptide antigens by surface decoration. For that purpose, orthogonally addressable multifunctional polycarbonate block copolymers were synthesized, enabling adjuvant conjugation through reactive ester chemistry and peptide decoration by strain-promoted alkyne-azide cycloaddition (SPAAC). In preparation for SPAAC, CD4+-specific peptide sequences of the model protein antigen ovalbumin were equipped with DBCO-moieties by site-selective modification at their N-terminal cysteine. With their azide groups exposed on their surface, the adjuvant-loaded nanogels were then efficiently decorated with DBCO-functional CD4+-peptides by SPAAC. In vitro evaluation of the adjuvant-loaded peptide-decorated gels then confirmed their strong immunostimulating properties as well as their high biocompatibility. Despite their covalent conjugation, the CD4+-peptide-decorated nanogels led to maturation of primary antigen-presenting cells and the downstream priming of CD4+-T cells. Subsequently, the peptide-decorated nanogels loaded with TLR7/8 agonist were successfully processed by antigen-presenting cells, enabling potent immune responses for future application in antigen-specific cancer immunotherapy.

Genetic testing in the evaluation of individuals with clinical diagnosis of atypical Sturge-Weber syndrome.

Sturge-Weber Syndrome (SWS) is a rare vascular malformation disorder characterized by abnormal blood vessels in the brain, skin, and eye. SWS is most commonly caused by a somatic mosaic GNAQ-p.Arg183Gln variant. In this series, 12 patients presented for clinical evaluation of SWS but were noted to have atypical features, and therefore germline and/or somatic genetic testing was performed. Atypical features included extensive capillary malformation on the body as well as the face, frontal bossing, macrocephaly, telangiectasia, overgrowth of extremities, absence of neurologic signs and symptoms, and family history of vascular malformations. Five patients had a somatic GNAQ or GNA11 pathogenic variant, one patient had a somatic mosaic likely-pathogenic variant in PIK3CA, and another one had a somatic mosaic deletion that disrupted PTPRD. The other five patients had germline variants in RASA1, EPHB4, or KIT. Our findings suggest that patients presenting for SWS evaluation who have atypical clinical characteristics may have pathogenic germline or somatic variants in genes other than GNAQ or GNA11. Broad germline and somatic genetic testing in these patients with atypical findings may have implications for medical care, prognosis, and trial eligibility.

Nanostructured Lipid Carriers Loaded with Dexamethasone Prevent Inflammatory Responses in Primary Non-Parenchymal Liver Cells.

Liver inflammation represents a major clinical problem in a wide range of pathologies. Among the strategies to prevent liver failure, dexamethasone (DXM) has been widely used to suppress inflammatory responses. The use of nanocarriers for encapsulation and sustained release of glucocorticoids to liver cells could provide a solution to prevent severe side effects associated with systemic delivery as the conventional treatment regime. Here we describe a nanostructured lipid carrier developed to efficiently encapsulate and release DXM. This nano-formulation proved to be stable over time, did not interact in vitro with plasma opsonins, and was well tolerated by primary non-parenchymal liver cells (NPCs). Released DXM preserved its pharmacological activity, as evidenced by inducing robust anti-inflammatory responses in NPCs. Taken together, nanostructured lipid carriers may constitute a reliable platform for the delivery of DXM to treat pathologies associated with chronic liver inflammation.

Enrichment Methods for Murine Liver Non-Parenchymal Cells Differentially Affect Their Immunophenotype and Responsiveness towards Stimulation.

Hepatocytes comprise the majority of the liver and largely exert metabolic functions, whereas non-parenchymal cells (NPCs)-comprising Kupffer cells, dendritic cells and liver sinusoidal endothelial cells-control the immunological state within this organ. Here, we compared the suitability of two isolation methods for murine liver NPCs. Liver perfusion (LP) with collagenase/DNase I applied via the portal vein leads to efficient liver digestion, whereas the modified liver dissociation (LD) method combines mechanical dissociation of the retrieved organ with enzymatic degradation of the extracellular matrix. In cases of both LP and LD, NPCs were enriched by subsequent gradient density centrifugation. Our results indicate that LP and LD are largely comparable with regards to the yield, purity, and composition of liver NPCs. However, LD-enriched liver NPCs displayed a higher degree of activation after overnight cultivation, and accordingly were less responsive towards stimulation with toll-like receptor ligands that are frequently used as adjuvants, e.g., in nano-vaccines. We conclude that LP is more suitable for obtaining liver NPCs for subsequent in vitro studies, whereas LD as the less laborious method, is more convenient for parallel isolation of larger numbers of samples for ex vivo analysis.

Influence of Advanced Organ Support (ADVOS) on Cytokine Levels in Patients with Acute-on-Chronic Liver Failure (ACLF).

Background: ADVanced Organ Support (ADVOS) is a novel type of extracorporeal albumin dialysis that supports multiorgan function in patients with acute-on-chronic liver failure (ACLF). No data exist on whether ADVOS affects inflammatory cytokine levels, which play a relevant role in ACLF. Aim: Our aim was to quantify cytokine levels both before and after a single ADVOS treatment in patients with ACLF at a regular dialysis ward. Methods and results: In this prospective study, 15 patients (60% men) with ACLF and an indication for renal replacement therapy were included. Patient liver function was severely compromised, reflected by a median CLIF-consortium ACLF score of 38 (IQR 35; 40). Blood samples were directly taken before and after ADVOS dialysis. The concentration of cytokines for IL-1ß, IFN-α2, IFN-γ, TNF-α, MCP-1, IL-6, IL-8, IL-10, IL-12p70, IL-17A, IL-18, IL-23, IL-33 were quantified via a cytometric bead array. We found no significant (p > 0.05) change in cytokine levels, even when patients were stratified for dialysis time (<480 min versus ≥480 min). The relevance of the assessed cytokines in contributing to systemic inflammation in ACLF was demonstrated by Ingenuity pathway analysis®. Conclusion: Concentrations of pathomechanistically relevant cytokines remained unchanged both before and after ADVOS treatment in patients with ACLF.

Diagnosis and discovery: Insights from the NIH Undiagnosed Diseases Program.

Living with an undiagnosed medical condition places a tremendous burden on patients, their families, and their healthcare providers. The Undiagnosed Diseases Program (UDP) was established at the National Institutes of Health (NIH) in 2008 with the primary goals of providing a diagnosis for patients with mysterious conditions and advancing medical knowledge about rare and common diseases. The program reviews applications from referring clinicians for cases that are considered undiagnosed despite a thorough evaluation. Those that are accepted receive clinical evaluations involving deep phenotyping and genetic testing that includes exome and genomic sequencing. Selected candidate gene variants are evaluated by collaborators using functional assays. Since its inception, the UDP has received more than 4500 applications and has completed evaluations on nearly 1300 individuals. Here we present six cases that exemplify the discovery of novel disease mechanisms, the importance of deep phenotyping for rare diseases, and how genetic diagnoses have led to appropriate treatment. The creation of the Undiagnosed Diseases Network (UDN) in 2014 has substantially increased the number of patients evaluated and allowed for greater opportunities for data sharing. Expansion to the Undiagnosed Diseases Network International (UDNI) has the possibility to extend this reach even farther. Together, networks of undiagnosed diseases programs are powerful tools to advance our knowledge of pathophysiology, accelerate accurate diagnoses, and improve patient care for patients with rare conditions.

Genome-wide DNA methylation profiling confirms a case of low-level mosaic Kabuki syndrome 1.

Kabuki syndrome is a Mendelian disorder of the epigenetic machinery characterized by typical dysmorphic features, intellectual disability, and postnatal growth deficiency. Pathogenic variants in the genes encoding the chromatin modifiers KMT2D and KDM6A are responsible for Kabuki syndrome 1 (KS1) and Kabuki syndrome 2 (KS2), respectively. In addition, 11 cases of KS1 caused by mosaic variants in KMT2D have been reported in the literature. Some of these individuals display milder craniofacial and growth phenotypes, and most do not have congenital heart defects. We report the case of an infant with severe hypoplastic left heart syndrome with mitral atresia and aortic atresia (HLHS MA-AA), pulmonary vein stenosis, and atypical facies with a somatic mosaic de novo nonsense variant in KMT2D (c.8200C>T, p.R2734*) identified on trio exome sequencing of peripheral blood and present in 11.2% of sequencing reads. KS was confirmed with EpiSign, a diagnostic genome-wide DNA methylation platform used to identify epigenetic signatures. This case suggests that use of this newly available clinical test can guide the interpretation of low-level mosaic variants identified through sequencing and suggests a new lower limit of mosaicism in whole blood required for a diagnosis of KS.

End Group Dye-Labeled Polycarbonate Block Copolymers for Micellar (Immuno-)Drug Delivery.

Defined conjugation of functional molecules to block copolymer end groups is a powerful strategy to enhance the scope of micellar carriers for drug delivery. In this study, an approach to access well-defined polycarbonate-based block copolymers by labeling their end groups with single fluorescent dye molecules is established. Following controlled polymerization conditions, the block copolymers' primary hydroxy end group can be converted into activated pentafluorophenyl ester carbonates and subsequently aminolyzed with fluorescent dyes that are equipped with primary amines. During a solvent-evaporation process, the resulting end group dye-labeled block copolymers self-assemble into narrowly dispersed ∼25 nm-sized micelles and simultaneously encapsulate hydrophobic (immuno-)drugs. The covalently attached fluorescent tracer can be used to monitor both uptake into cells and stability under biologically relevant conditions, including incubation with blood plasma or during blood circulation in zebrafish embryos. By encapsulation of the toll-like receptor 7/8 (TLR7/8) agonist CL075, immune stimulatory polymeric micelles are generated that get internalized by various antigen-presenting dendritic cells and promote their maturation. Generally, such end group dye-labeled polycarbonate block copolymers display ideal features to permit targeted delivery of hydrophobic drugs to key immune cells for vaccination and cancer immunotherapy.

Systemically Administered TLR7/8 Agonist and Antigen-Conjugated Nanogels Govern Immune Responses against Tumors.

The generation of specific humoral and cellular immune responses plays a pivotal role in the development of effective vaccines against tumors. Especially the presence of antigen-specific, cytotoxic T cells influences the outcome of therapeutic cancer vaccinations. Different strategies, ranging from delivering antigen-encoding mRNAs to peptides or full antigens, are accessible but often suffer from insufficient immunogenicity and require immune-boosting adjuvants as well as carrier platforms to ensure stability and adequate retention. Here, we introduce a pH-responsive nanogel platform as a two-component antitumor vaccine that is safe for intravenous application and elicits robust immune responses in vitro and in vivo. The underlying chemical design allows for straightforward covalent attachment of a model antigen (ovalbumin) and an immune adjuvant (imidazoquinoline-type TLR7/8 agonist) onto the same nanocarrier system. In addition to eliciting antigen-specific T and B cell responses that outperform mixtures of individual components, our two-component nanovaccine leads in prophylactic and therapeutic studies to an antigen-specific growth reduction of different tumors expressing ovalbumin intracellularly or on their surface. Regarding the versatile opportunities for functionalization, our nanogels are promising for the development of highly customized and potent nanovaccines.

Density of Conjugated Antibody Determines the Extent of Fc Receptor Dependent Capture of Nanoparticles by Liver Sinusoidal Endothelial Cells.

Despite considerable progress in the design of multifunctionalized nanoparticles (NPs) that selectively target specific cell types, their systemic application often results in unwanted liver accumulation. The exact mechanisms for this general observation are still unclear. Here we asked whether the number of cell-targeting antibodies per NP determines the extent of NP liver accumulation and also addressed the mechanisms by which antibody-coated NPs are retained in the liver. We used polysarcosine-based peptobrushes (PBs), which in an unmodified form remain in the circulation for >24 h due to the absence of a protein corona formation and low unspecific cell binding, and conjugated them with specific average numbers (2, 6, and 12) of antibodies specific for the dendritic cell (DC) surface receptor, DEC205. We assessed the time-dependent biodistribution of PB-antibody conjugates by in vivo imaging and flow cytometry. We observed that PB-antibody conjugates were trapped in the liver and that the extent of liver accumulation strongly increased with the number of attached antibodies. PB-antibody conjugates were selectively captured in the liver via Fc receptors (FcR) on liver sinusoidal endothelial cells, since systemic administration of FcR-blocking agents or the use of F(ab')2 fragments prevented liver accumulation. Cumulatively, our study demonstrates that liver endothelial cells play a yet scarcely acknowledged role in liver entrapment of antibody-coated NPs and that low antibody numbers on NPs and the use of F(ab')2 antibody fragments are both sufficient for cell type-specific targeting of secondary lymphoid organs and necessary to minimize unwanted liver accumulation.

Squaric Ester-Based, pH-Degradable Nanogels: Modular Nanocarriers for Safe, Systemic Administration of Toll-like Receptor 7/8 Agonistic Immune Modulators.

Small-molecular Toll-like receptor 7/8 (TLR7/8) agonists hold promise as immune modulators for a variety of immune therapeutic purposes including cancer therapy or vaccination. However, due to their rapid systemic distribution causing difficult-to-control inflammatory off-target effects, their application is still problematic, in particular systemically. To address this problem, we designed and robustly fabricated pH-responsive nanogels serving as versatile immunodrug nanocarriers for safe delivery of TLR7/8-stimulating imidazoquinolines after intravenous administration. To this aim, a primary amine-reactive methacrylamide monomer bearing a pendant squaric ester amide is introduced, which is polymerized under controlled RAFT polymerization conditions. Corresponding PEG-derived squaric ester amide block copolymers self-assemble into precursor micelles in polar protic solvents. Their cores are amine-reactive and can sequentially be transformed by acid-sensitive cross-linkers, dyes, and imidazoquinolines. Remaining squaric ester amides are hydrophilized affording fully hydrophilic nanogels with profound stability in human plasma but stimuli-responsive degradation upon exposure to endolysosomal pH conditions. The immunomodulatory behavior of the imidazoquinolines alone or conjugated to the nanogels was demonstrated by macrophages in vitro. In vivo, however, we observed a remarkable impact of the nanogel: After intravenous injection, a spatially controlled immunostimulatory activity was evident in the spleen, whereas systemic off-target inflammatory responses triggered by the small-molecular imidazoquinoline analogue were absent. These findings underline the potential of squaric ester-based, pH-degradable nanogels as a promising platform to permit intravenous administration routes of small-molecular TLR7/8 agonists and, thus, the opportunity to explore their adjuvant potency for systemic vaccination or cancer immunotherapy purposes.

Complement-Opsonized Nano-Carriers Are Bound by Dendritic Cells (DC) via Complement Receptor (CR)3, and by B Cell Subpopulations via CR-1/2, and Affect the Activation of DC and B-1 Cells.

The development of nanocarriers (NC) for biomedical applications has gained large interest due to their potential to co-deliver drugs in a cell-type-targeting manner. However, depending on their surface characteristics, NC accumulate serum factors, termed protein corona, which may affect their cellular binding. We have previously shown that NC coated with carbohydrates to enable biocompatibility triggered the lectin-dependent complement pathway, resulting in enhanced binding to B cells via complement receptor (CR)1/2. Here we show that such NC also engaged all types of splenic leukocytes known to express CR3 at a high rate when NC were pre-incubated with native mouse serum resulting in complement opsonization. By focusing on dendritic cells (DC) as an important antigen-presenting cell type, we show that CR3 was essential for binding/uptake of complement-opsonized NC, whereas CR4, which in mouse is specifically expressed by DC, played no role. Further, a minor B cell subpopulation (B-1), which is important for first-line pathogen responses, and co-expressed CR1/2 and CR3, in general, engaged NC to a much higher extent than normal B cells. Here, we identified CR-1/2 as necessary for binding of complement-opsonized NC, whereas CR3 was dispensable. Interestingly, the binding of complement-opsonized NC to both DC and B-1 cells affected the expression of activation markers. Our findings may have important implications for the design of nano-vaccines against infectious diseases, which codeliver pathogen-specific protein antigen and adjuvant, aimed to induce a broad adaptive cellular and humoral immune response by inducing cytotoxic T lymphocytes that kill infected cells and pathogen-neutralizing antibodies, respectively. Decoration of nano-vaccines either with carbohydrates to trigger complement activation in vivo or with active complement may result in concomitant targeting of DC and B cells and thereby may strongly enhance the extent of dual cellular/humoral immune responses.

Role of Liver-Mediated Tolerance in Nanoparticle-Based Tumor Therapy.

In the last decades, the use of nanocarriers for immunotherapeutic purposes has gained a lot of attention, especially in the field of tumor therapy. However, most types of nanocarriers accumulate strongly in the liver after systemic application. Due to the default tolerance-promoting role of liver non-parenchymal cells (NPCs), Kupffer cells (KCs), liver sinusoidal endothelial cells (LSECs), and hepatic stellate cells (HSCs), their potential role on the immunological outcome of systemic nano-vaccination approaches for therapy of tumors in the liver and in other organs needs to be considered. Concerning immunological functions, KCs have been the focus until now, but recent studies have elucidated an important role of LSECs and HSCs as well. Therefore, this review aims to summarize current knowledge on the employment of nanocarriers for immunotherapeutic therapy of liver diseases and the overall role of liver NPCs in the context of nano-vaccination approaches. With regard to the latter, we discuss strategies on how to address liver NPCs, aiming to exploit and modulate their immunological properties, and alternatively how to avoid unwanted engagement of nano-vaccines by liver NPCs for tumor therapy.

Independent Methylome-Wide Association Studies of Schizophrenia Detect Consistent Case-Control Differences.

Methylome-wide association studies (MWASs) are promising complements to sequence variation studies. We used existing sequencing-based methylation data, which assayed the majority of all 28 million CpGs in the human genome, to perform an MWAS for schizophrenia in blood, while controlling for cell-type heterogeneity with a recently generated platform-specific reference panel. Next, we compared the MWAS results with findings from 3 existing large-scale array-based schizophrenia methylation studies in blood that assayed up to ~450 000 CpGs. Our MWAS identified 22 highly significant loci (P < 5 × 10-8) and 852 suggestively significant loci (P < 1 × 10-5). The top finding (P = 5.62 × 10-11, q = 0.001) was located in MFN2, which encodes mitofusin-2 that regulates Ca2+ transfer from the endoplasmic reticulum to mitochondria in cooperation with DISC1. The second-most significant site (P = 1.38 × 10-9, q = 0.013) was located in ALDH1A2, which encodes an enzyme for astrocyte-derived retinoic acid-a key neuronal morphogen with relevance for schizophrenia. Although the most significant MWAS findings were not assayed on the arrays, we observed significant enrichment of overlapping findings with 2 of the 3 array datasets (P = 0.0315, 0.0045, 0.1946). Overrepresentation analysis of Gene Ontology terms for the genes in the significant overlaps suggested high similarity in the biological functions detected by the different datasets. Top terms were related to immune and/or stress responses, cell adhesion and motility, and a broad range of processes essential for neurodevelopment.

Efecto de borde en la distribución de líquenes y el contenido de clorofilas en fragmentos de Polylepis quadrijuga (Rosaceae) en el páramo de La Rusia (Boyacá-Colombia) / Edge effect on lichen's distribution and chlorophyll content, in fragments of Polylepis quadrijuga (Rosaceae) in Páramo de la Rusia (Boyacá-Colombia)

Resumen:La fragmentación de los ecosistemas es uno de los fenómenos antrópicos de mayor impacto global, y el efecto borde provoca que solo el interior de los fragmentos conserve sus características bióticas y abióticas originales. Los líquenes son organismos especialmente susceptibles a la variabilidad ambiental, lo que podría ser útil para la bio-indicación del efecto borde. En este trabajo se evaluó el efecto borde en dos fragmentos de Polylepis quadrijuga en el Páramo de la Rusia (Boyacá-Colombia), para determinar si existe un efecto borde sobre la distribución de los líquenes asociados a P. quadrijuga y su contenido de clorofilas. Se utilizaron tres transectos de 70 m a través del gradiente matriz-borde-interior en cada fragmento. Se escogieron nueve forofitos por transecto para medir variables ambientales: radiación fotosintéticamente activa, humedad relativa y temperatura del aire, y variables biológicas: riqueza y cobertura de cada especie. Además, fueron elegidas las especies que estaban presentes en las tres zonas del gradiente para cuantificar el contenido de clorofilas a y b, y determinar si hay cambios en la relación de clorofilas a/b que podrían sugerir plasticidad fisiológica como respuesta al efecto borde. Nuestros resultados mostraron que el fragmento 2 tenía una mayor exposición al borde debido a su alta relación perímetro/área, lo que causa una homogenización ambiental y pérdida de biodiversidad con relación con el fragmento 1. En general, se encontraron 55 especies que se distribuyen diferencialmente en relación con los fragmentos y el gradiente matriz-borde-interior. El interior del fragmento 1 fue la zona más conservada, que alberga una composición diferente en más de un 40 % a la composición de cualquier otra zona. Se clasificaron los líquenes de acuerdo con sus hábitos: gelatinosos, fruticosos, costrosos o foliosos, pero no se encontró ninguna relación entre la distribución por hábito y el efecto borde. Seis especies de amplia distribución mostraron cambios en el contenido de clorofila a lo largo del gradiente matriz-borde-interior, lo que es una evidencia de la plasticidad fisiológica al efecto borde. También fue posible diferenciar entre especies con preferencia por ambientes de alta temperatura y especies con preferencia a lugares más húmedos y suficientemente irradiados. Se concluye que algunas especies podrían tener un importante potencial como bio-indicadores de la fragmentación en ambientes de páramo.

Abstract:The ecosystems fragmentation is one of the anthropic phenomena with highest impact at global level and the edge effect causes that only the interior of fragments conserves their original biotic and abiotic characteristics. Lichens are organisms especially susceptible to environmental variability, what could be useful for bio-indication of edge effect. In this work, we evaluated the edge effect in two fragments of Polylepis quadrijuga in the Páramo de la Rusia (Boyacá-Colombia) to determine if there is an edge effect on distribution of lichens associated to P. quadrijuga and their chlorophyll content. We used three transects of 70 m across the matrix-edge-interior gradient in each fragment. We chose nine phorophytes per transect to measure the environmental variables: photosynthetically active radiation, relative humidity and air temperature, and the biological variables: richness and cover per species. Besides, we employed the species that were present in all the three zones of the gradient to quantify the content of chlorophylls a and b, and determine if there are changes in the ratio of chlorophylls a/b that could suggest physiological plasticity as a response to the edge effect. Our results showed that fragment 2 had a higher edge exposition because of its high relation perimeter/area, allowing to an environmental homogenization and lose of biodiversity in relation with fragment 1. Overall, we found 55 differentially distributed species in relation with the fragments and the matrix-edge-interior gradient. The interior of fragment 1 was the most conserved zone, harboring a composition different in more than 40 % to the composition of any other zone. We classified the lichens according with their habits: gelatinous, fruticose, crusty or foliose, but we did not find any relationship between the habit distribution and the edge effect. Six species of wide distribution showed changes in the chlorophyll content along the matrix- edge-interior gradient, what is an evidence of physiological plasticity to edge effect. It was also possible to distinguish between species with preference to warmer environment and species with preference to more humid and sufficiently irradiated places. We concluded that some species of lichens could have an important potential as bio-indicators of fragmentation in the páramo. Rev. Biol. Trop. 64 (4): 1683-1697. Epub 2016 December 01.

High-specificity bioinformatics framework for epigenomic profiling of discordant twins reveals specific and shared markers for ACPA and ACPA-positive rheumatoid arthritis.

BACKGROUND: Twin studies are powerful models to elucidate epigenetic modifications resulting from gene-environment interactions. Yet, commonly a limited number of clinical twin samples are available, leading to an underpowered situation afflicted with false positives and hampered by low sensitivity. We investigated genome-wide DNA methylation data from two small sets of monozygotic twins representing different phases during the progression of rheumatoid arthritis (RA) to find novel genes for further research. METHODS: We implemented a robust statistical methodology aimed at investigating a small number of samples to identify differential methylation utilizing the comprehensive CHARM platform with whole blood cell DNA from two sets of twin pairs discordant either for ACPA (antibodies to citrullinated protein antigens)-positive RA versus ACPA-negative healthy or for ACPA-positive healthy (a pre-RA stage) versus ACPA-negative healthy. To deconvolute cell type-dependent differential methylation, we assayed the methylation patterns of sorted cells and used computational algorithms to resolve the relative contributions of different cell types and used them as covariates. RESULTS: To identify methylation biomarkers, five healthy twin pairs discordant for ACPAs were profiled, revealing a single differentially methylated region (DMR). Seven twin pairs discordant for ACPA-positive RA revealed six significant DMRs. After deconvolution of cell type proportions, profiling of the healthy ACPA discordant twin-set revealed 17 genome-wide significant DMRs. When methylation profiles of ACPA-positive RA twin pairs were adjusted for cell type, the analysis disclosed one significant DMR, associated with the EXOSC1 gene. Additionally, the results from our methodology suggest a temporal connection of the protocadherine beta-14 gene to ACPA-positivity with clinical RA. CONCLUSIONS: Our biostatistical methodology, optimized for a low-sample twin design, revealed non-genetically linked genes associated with two distinct phases of RA. Functional evidence is still lacking but the results reinforce further study of epigenetic modifications influencing the progression of RA. Our study design and methodology may prove generally useful in twin studies.

Association of DNA Methylation Differences With Schizophrenia in an Epigenome-Wide Association Study.

IMPORTANCE: DNA methylation may play an important role in schizophrenia (SZ), either directly as a mechanism of pathogenesis or as a biomarker of risk. OBJECTIVE: To scan genome-wide DNA methylation data to identify differentially methylated CpGs between SZ cases and controls. DESIGN, SETTING, AND PARTICIPANTS: Epigenome-wide association study begun in 2008 using DNA methylation levels of 456 513 CpG loci measured on the Infinium HumanMethylation450 array (Illumina) in a consortium of case-control studies for initial discovery and in an independent replication set. Primary analyses used general linear regression, adjusting for age, sex, race/ethnicity, smoking, batch, and cell type heterogeneity. The discovery set contained 689 SZ cases and 645 controls (n = 1334), from 3 multisite consortia: the Consortium on the Genetics of Endophenotypes in Schizophrenia, the Project among African-Americans To Explore Risks for Schizophrenia, and the Multiplex Multigenerational Family Study of Schizophrenia. The replication set contained 247 SZ cases and 250 controls (n = 497) from the Genomic Psychiatry Cohort. MAIN OUTCOMES AND MEASURES: Identification of differentially methylated positions across the genome in SZ cases compared with controls. RESULTS: Of the 689 case participants in the discovery set, 477 (69%) were men and 258 (37%) were non-African American; of the 645 controls, 273 (42%) were men and 419 (65%) were non-African American. In our replication set, cases/controls were 76% male and 100% non-African American. We identified SZ-associated methylation differences at 923 CpGs in the discovery set (false discovery rate, <0.2). Of these, 625 showed changes in the same direction including 172 with P < .05 in the replication set. Some replicated differentially methylated positions are located in a top-ranked SZ region from genome-wide association study analyses. CONCLUSIONS AND RELEVANCE: This analysis identified 172 replicated new associations with SZ after careful correction for cell type heterogeneity and other potential confounders. The overlap with previous genome-wide association study data can provide potential insights into the functional relevance of genetic signals for SZ.