Ascorbic Acid/Retinol and/or Inflammatory Stimuli’s Effect on Proliferation/Differentiation Properties and Transcriptomics of Gingival Stem/Progenitor Cells

The present study explored the effects of ascorbic-acid (AA)/retinol and timed inflammation on the stemness, the regenerative potential, and the transcriptomics profile of gingival mesenchymal stem/progenitor cells’ (G-MSCs). STRO-1 (mesenchymal stem cell marker) immuno-magnetically sorted G-MSCs were cultured in basic medium (control group), in basic medium with IL-1β (1 ng/mL), TNF-α (10 ng/mL) and IFN-γ (100 ng/mL, inflammatory-medium), in basic medium with AA (250 µmol/L) and retinol (20 µmol/L) (AA/retinol group) or in inflammatory medium with AA/retinol (inflammatory/AA/retinol group; n = 5/group). The intracellular levels of phosphorylated and total β-Catenin at 1 h, the expression of stemness genes over 7 days, the number of colony-forming units (CFUs) as well as the cellular proliferation aptitude over 14 days, and the G-MSCs’ multilineage differentiation potential were assessed. Next-generation sequencing was undertaken to elaborate on up-/downregulated genes and altered intracellular pathways. G-MSCs demonstrated all mesenchymal stem/progenitor cells characteristics. Controlled inflammation with AA/retinol significantly elevated NANOG (p < 0.05). The AA/retinol-mediated reduction in intracellular phosphorylated β-Catenin was restored through the effect of controlled inflammation (p < 0.05). Cellular proliferation was highest in the AA/retinol group (p < 0.05)

Surfactant-Free Peroxidase-Mediated Enzymatic Polymerization of a Biorenewable Butyrolactone Monomer via a Green Approach:Synthesis of Sustainable Biobased Latexes

A green surfactant-free one-pot horseradish peroxidase-mediated enzymatic polymerization is successfully applied to produce a sustainable and thermally stable biobased high average molar mass poly(α-methylene-γ-butyrolactone) (PMBL) at ambient conditions in water for the first time. The initiation step required only very low concentrations of hydrogen peroxide and 2,4-pentanedione water-soluble initiator to generate the keto-enoxy radicals responsible for forming the primary latex particles. The polymer nanoparticles can be seen as monodisperse, and the biobased latexes are colloidally stable and likely stabilized by the adsorption of 2,4-pentanedione moieties on the particle surfaces. Polymerizations in air produced a 98% yield of PMBL after only 3 h, highlighting the relevance of molecular oxygen. An array of characterization techniques such as dynamic light scattering (DLS), Fourier transform infrared (FTIR), 1H, 13C, and HSQC two-dimensional (2D) nuclear magnetic resonance (NMR), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), and size-exclusion chromatography (SEC) are used to confirm the properties of the synthesized latexes. The PMBL exhibited high thermal stability, with only a 5% weight loss at 340 °C and a glass-transition temperature of 200 °C, which is double that of polymethyl methacrylate (PMMA). This research provides an interesting pathway for the synthesis of sustainable biobased latexes via enzymes in a green environment using just water at ambient conditions and the potential use of the polymer in high-temperature applications.</p

Surfactant-Free Peroxidase-Mediated Enzymatic Polymerization of a Biorenewable Butyrolactone Monomer via a Green Approach:Synthesis of Sustainable Biobased Latexes

A green surfactant-free one-pot horseradish peroxidase-mediated enzymatic polymerization is successfully applied to produce a sustainable and thermally stable biobased high average molar mass poly(α-methylene-γ-butyrolactone) (PMBL) at ambient conditions in water for the first time. The initiation step required only very low concentrations of hydrogen peroxide and 2,4-pentanedione water-soluble initiator to generate the keto-enoxy radicals responsible for forming the primary latex particles. The polymer nanoparticles can be seen as monodisperse, and the biobased latexes are colloidally stable and likely stabilized by the adsorption of 2,4-pentanedione moieties on the particle surfaces. Polymerizations in air produced a 98% yield of PMBL after only 3 h, highlighting the relevance of molecular oxygen. An array of characterization techniques such as dynamic light scattering (DLS), Fourier transform infrared (FTIR), 1H, 13C, and HSQC two-dimensional (2D) nuclear magnetic resonance (NMR), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), and size-exclusion chromatography (SEC) are used to confirm the properties of the synthesized latexes. The PMBL exhibited high thermal stability, with only a 5% weight loss at 340 °C and a glass-transition temperature of 200 °C, which is double that of polymethyl methacrylate (PMMA). This research provides an interesting pathway for the synthesis of sustainable biobased latexes via enzymes in a green environment using just water at ambient conditions and the potential use of the polymer in high-temperature applications.</p

Surfactant-Free Peroxidase-Mediated Enzymatic Polymerization of a Biorenewable Butyrolactone Monomer via a Green Approach:Synthesis of Sustainable Biobased Latexes

A green surfactant-free one-pot horseradish peroxidase-mediated enzymatic polymerization is successfully applied to produce a sustainable and thermally stable biobased high average molar mass poly(α-methylene-γ-butyrolactone) (PMBL) at ambient conditions in water for the first time. The initiation step required only very low concentrations of hydrogen peroxide and 2,4-pentanedione water-soluble initiator to generate the keto-enoxy radicals responsible for forming the primary latex particles. The polymer nanoparticles can be seen as monodisperse, and the biobased latexes are colloidally stable and likely stabilized by the adsorption of 2,4-pentanedione moieties on the particle surfaces. Polymerizations in air produced a 98% yield of PMBL after only 3 h, highlighting the relevance of molecular oxygen. An array of characterization techniques such as dynamic light scattering (DLS), Fourier transform infrared (FTIR), 1H, 13C, and HSQC two-dimensional (2D) nuclear magnetic resonance (NMR), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), and size-exclusion chromatography (SEC) are used to confirm the properties of the synthesized latexes. The PMBL exhibited high thermal stability, with only a 5% weight loss at 340 °C and a glass-transition temperature of 200 °C, which is double that of polymethyl methacrylate (PMMA). This research provides an interesting pathway for the synthesis of sustainable biobased latexes via enzymes in a green environment using just water at ambient conditions and the potential use of the polymer in high-temperature applications.</p

Diagnostic utility of snail in metaplastic breast carcinoma

Metaplastic breast carcinoma (MBC) is a rare subtype of breast cancer characterized by coexistence of carcinomatous and sarcomatous components. Snail is a nuclear transcription factor incriminated in the transition of epithelial to mesenchymal differentiation of breast cancer. Aberrant Snail expression results in lost expression of the cell adhesion molecule E-cadherin, an event associated with changes in epithelial architecture and invasive growth. We aimed to identify the utility of Snail, and of traditional immunohistochemical markers, in accurate MBC classification and to evaluate clinicopathologic characteristics and outcome

Contribution of microscopy for understanding the mechanism of action against trypanosomatids

Transmission electron microscopy (TEM) has proved to be a useful tool to study the ultrastructural alterations and the target organelles of new antitrypanosomatid drugs. Thus, it has been observed that sesquiterpene lactones induce diverse ultrastructural alterations in both T. cruzi and Leishmania spp., such as cytoplasmic vacuolization, appearance of multilamellar structures, condensation of nuclear DNA, and, in some cases, an important accumulation of lipid vacuoles. This accumulation could be related to apoptotic events. Some of the sesquiterpene lactones (e.g., psilostachyin) have also been demonstrated to cause an intense mitochondrial swelling accompanied by a visible kinetoplast deformation as well as the appearance of multivesicular bodies. This mitochondrial swelling could be related to the generation of oxidative stress and associated to alterations in the ergosterol metabolism. The appearance of multilamellar structures and multiple kinetoplasts and flagella induced by the sesquiterpene lactone psilostachyin C indicates that this compound would act at the parasite cell cycle level, in an intermediate stage between kinetoplast segregation and nuclear division. In turn, the diterpene lactone icetexane has proved to induce the external membrane budding on T. cruzi together with an apparent disorganization of the pericellar cytoskeleton. Thus, ultrastructural TEM studies allow elucidating the possible mechanisms and the subsequent identification of molecular targets for the action of natural compounds on trypanosomatids.Fil: Lozano, Esteban Sebastián. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Spina Zapata, Renata María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Barrera, Patricia Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Tonn, Carlos Eugenio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - San Luis. Instituto de Investigaciones en Tecnología Química. Universidad Nacional de San Luis. Facultad de Química, Bioquímica y Farmacia. Instituto de Investigaciones en Tecnología Química; ArgentinaFil: Sosa Escudero, Miguel Angel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentin

Neuroinflammation, Mast Cells, and Glia: Dangerous Liaisons

The perspective of neuroinflammation as an epiphenomenon following neuron damage is being replaced by the awareness of glia and their importance in neural functions and disorders. Systemic inflammation generates signals that communicate with the brain and leads to changes in metabolism and behavior, with microglia assuming a pro-inflammatory phenotype. Identification of potential peripheral-to-central cellular links is thus a critical step in designing effective therapeutics. Mast cells may fulfill such a role. These resident immune cells are found close to and within peripheral nerves and in brain parenchyma/meninges, where they exercise a key role in orchestrating the inflammatory process from initiation through chronic activation. Mast cells and glia engage in crosstalk that contributes to accelerate disease progression; such interactions become exaggerated with aging and increased cell sensitivity to stress. Emerging evidence for oligodendrocytes, independent of myelin and support of axonal integrity, points to their having strong immune functions, innate immune receptor expression, and production/response to chemokines and cytokines that modulate immune responses in the central nervous system while engaging in crosstalk with microglia and astrocytes. In this review, we summarize the findings related to our understanding of the biology and cellular signaling mechanisms of neuroinflammation, with emphasis on mast cell-glia interactions

Transcript Expression Analysis of Putative Trypanosoma brucei GPI-Anchored Surface Proteins during Development in the Tsetse and Mammalian Hosts

Human African Trypanosomiasis is a devastating disease caused by the parasite Trypanosoma brucei. Trypanosomes live extracellularly in both the tsetse fly and the mammal. Trypanosome surface proteins can directly interact with the host environment, allowing parasites to effectively establish and maintain infections. Glycosylphosphatidylinositol (GPI) anchoring is a common posttranslational modification associated with eukaryotic surface proteins. In T. brucei, three GPI-anchored major surface proteins have been identified: variant surface glycoproteins (VSGs), procyclic acidic repetitive protein (PARP or procyclins), and brucei alanine rich proteins (BARP). The objective of this study was to select genes encoding predicted GPI-anchored proteins with unknown function(s) from the T. brucei genome and characterize the expression profile of a subset during cyclical development in the tsetse and mammalian hosts. An initial in silico screen of putative T. brucei proteins by Big PI algorithm identified 163 predicted GPI-anchored proteins, 106 of which had no known functions. Application of a second GPI-anchor prediction algorithm (FragAnchor), signal peptide and trans-membrane domain prediction software resulted in the identification of 25 putative hypothetical proteins. Eighty-one gene products with hypothetical functions were analyzed for stage-regulated expression using semi-quantitative RT-PCR. The expression of most of these genes were found to be upregulated in trypanosomes infecting tsetse salivary gland and proventriculus tissues, and 38% were specifically expressed only by parasites infecting salivary gland tissues. Transcripts for all of the genes specifically expressed in salivary glands were also detected in mammalian infective metacyclic trypomastigotes, suggesting a possible role for these putative proteins in invasion and/or establishment processes in the mammalian host. These results represent the first large-scale report of the differential expression of unknown genes encoding predicted T. brucei surface proteins during the complete developmental cycle. This knowledge may form the foundation for the development of future novel transmission blocking strategies against metacyclic parasites

Poly(Glycerol Adipate-co-ω-Pentadecalactone) Spray-Dried Microparticles as Sustained Release Carriers for Pulmonary Delivery

Purpose The aim of this work was to optimize biodegradable polyester poly(glycerol adipate-co-ω-pentadecalactone), PGA-co-PDL, microparticles as sustained release (SR) carriers for pulmonary drug delivery. Methods Microparticles were produced by spray drying directly from double emulsion with and without dispersibility enhancers ( L -arginine and L -leucine) (0.5–1.5%w/w) using sodium fluorescein (SF) as a model hydrophilic drug. Results Spray-dried microparticles without dispersibility enhancers exhibited aggregated powders leading to low fine particle fraction (%FPF) (28.79 ± 3.24), fine particle dose (FPD) (14.42 ± 1.57 μg), with a mass median aerodynamic diameter (MMAD) 2.86 ± 0.24 μm. However, L -leucine was significantly superior in enhancing the aerosolization performance ( L- arginine:%FPF 27.61 ± 4.49–26.57 ± 1.85; FPD 12.40 ± 0.99–19.54 ± 0.16 μg and MMAD 2.18 ± 0.35–2.98 ± 0.25 μm, L -leucine:%FPF 36.90 ± 3.6–43.38 ± 5.6; FPD 18.66 ± 2.90–21.58 ± 2.46 μg and MMAD 2.55 ± 0.03–3.68 ± 0.12 μm). Incorporating L -leucine (1.5%w/w) reduced the burst release (24.04 ± 3.87%) of SF compared to unmodified formulations (41.87 ± 2.46%), with both undergoing a square root of time (Higuchi’s pattern) dependent release. Comparing the toxicity profiles of PGA-co-PDL with L -leucine (1.5%w/w) (5 mg/ml) and poly(lactide-co-glycolide), (5 mg/ml) spray-dried microparticles in human bronchial epithelial 16HBE14o- cell lines, resulted in cell viability of 85.57 ± 5.44 and 60.66 ± 6.75%, respectively, after 72 h treatment. Conclusion The above data suggest that PGA-co-PDL may be a useful polymer for preparing SR microparticle carriers, together with dispersibility enhancers, for pulmonary delivery

ReRep: Computational detection of repetitive sequences in genome survey sequences (GSS)

<p>Abstract</p> <p>Background</p> <p>Genome survey sequences (GSS) offer a preliminary global view of a genome since, unlike ESTs, they cover coding as well as non-coding DNA and include repetitive regions of the genome. A more precise estimation of the nature, quantity and variability of repetitive sequences very early in a genome sequencing project is of considerable importance, as such data strongly influence the estimation of genome coverage, library quality and progress in scaffold construction. Also, the elimination of repetitive sequences from the initial assembly process is important to avoid errors and unnecessary complexity. Repetitive sequences are also of interest in a variety of other studies, for instance as molecular markers.</p> <p>Results</p> <p>We designed and implemented a straightforward pipeline called ReRep, which combines bioinformatics tools for identifying repetitive structures in a GSS dataset. In a case study, we first applied the pipeline to a set of 970 GSSs, sequenced in our laboratory from the human pathogen <it>Leishmania braziliensis</it>, the causative agent of leishmaniosis, an important public health problem in Brazil. We also verified the applicability of ReRep to new sequencing technologies using a set of 454-reads of an <it>Escheria coli</it>. The behaviour of several parameters in the algorithm is evaluated and suggestions are made for tuning of the analysis.</p> <p>Conclusion</p> <p>The ReRep approach for identification of repetitive elements in GSS datasets proved to be straightforward and efficient. Several potential repetitive sequences were found in a <it>L. braziliensis </it>GSS dataset generated in our laboratory, and further validated by the analysis of a more complete genomic dataset from the EMBL and Sanger Centre databases. ReRep also identified most of the <it>E. coli </it>K12 repeats prior to assembly in an example dataset obtained by automated sequencing using 454 technology. The parameters controlling the algorithm behaved consistently and may be tuned to the properties of the dataset, in particular to the length of sequencing reads and the genome coverage. ReRep is freely available for academic use at <url>http://bioinfo.pdtis.fiocruz.br/ReRep/</url>.</p