Development of the technique of isoelectricfocusing and immunodetection of immunoglobulin A in the healthcare context as support for the diagnosis of Multiple Sclerosis

Motivation: Multiple sclerosis (MS) is characterized by demyelination and subsequent degeneration of myelin sheaths that line the axons of neurons, producing multifocal and temporarily dispersed damage to the central nervous system (CNS) [1] that leads to neuronal damage and axonal loss [2]. MS is a complex disease in which the exact etiology remains unknown, although our current understanding of the natural history of MS and its immunopathogenesis points to an immune deregulation resulting from an interaction between genetic predispositions and environmental factors [2]. Given this uncertainty, there is a need for the identification of biomarkers in both cerebrospinal fluid and peripheral blood that help us to have a more complete view of the disease, and at the same time could contribute to a faster and more accurate diagnosis. This project arises from the importance and need to study immunoglobulin A as a biomarker for MS. Methods: Isoelectric focusing is one of the most powerful techniques used to solve complex mixtures of proteins [2]. The main objective of this project is the development of the isoelectroenfocusing and immunodetection technique for oligoclonal bands of immunoglobulin A and the IgA reduction protocol [3][4] to establish a subsequent relationship between patients diagnosed with MS and intrathecal secretion of immunoglobulin A. Results: For the implementation of the technique, a set of cerebrospinal fluid (CSF) samples previously extracted by lumbar puncture were selected, presenting the patient with a diagnostic suspicion of demyelinating disease, and presenting a high quantification of intrathecal secretion of IgA, routinely carried out in the Clinical Biochemistry Unit, immunology laboratory. A total of 23 samples were selected based on IgA concentrations in both CSF and serum. Conclusions: It should be noted that today the CSF remains the most interesting fluid for the study of degenerative neurological diseases. The CSF is the best representative of the clinical manifestations that are confined to the central nervous system, hence its characterization is, increasingly, of high interest for a more accurate diagnosis. With this set-up it is intended to give new information to the clinician to help diagnose the disease. Its value as a marker of the disease will be derived from subsequent observations and studies that clinicians perform in their cohort

Evaluation of the antimicrobial activity and cytotoxicity of different components of natural origin present in essential oils

Even though essential oils (EOs) have been used for therapeutic purposes, there is now a renewed interest in the antimicrobial properties of phytochemicals and EOs in particular. Their demonstrated low levels of induction of antimicrobial resistance make them interesting for bactericidal applications, though their complex composition makes it necessary to focus on the study of their main components to identify the most effective ones. Herein, the evaluation of the antimicrobial action of different molecules present in EOs against planktonic and biofilm-forming Gram-positive (Staphylococcus aureus) and Gram-negative (Escherichia coli) bacteria was assessed. The bactericidal mechanisms of the different molecules, as well as their cytocompatibility, were also studied. Carvacrol, cinnamaldehyde, and thymol exhibit the highest in vitro antimicrobial activities against E. coli and S. aureus, with membrane disruption the bactericidal mechanism identified. The addition of those compounds (=0.5 mg/mL) hampers S. aureus biofilm formation and partially eliminates preformed biofilms. The subcytotoxic values of the tested EO molecules (0.015–0.090 mg/mL) are lower than the minimum inhibitory and bactericidal concentrations obtained for bacteria (0.2–0.5 mg/mL) but are higher than that obtained for chlorhexidine (0.004 mg/mL), indicating the reduced cytotoxicity of EOs. Therefore, carvacrol, cinnamaldehyde, and thymol are molecules contained in EOs that could be used against E. coli– and S. aureus–mediated infections without a potential induction of bactericidal resistance and with lower cell toxicity than the conventional widely used chlorhexidine

Efficiency of Antimicrobial Electrospun Thymol-Loaded Polycaprolactone Mats in Vivo

Due to the prevalence of antimicrobial resistant pathogens, natural products with long-term antimicrobial activities are considered as potential alternatives. In this work, polycaprolactone (PCL) electrospun fibers with mean diameters around 299 nm and loaded with 14.92 ± 1.31% w/w thymol (THY) were synthesized. The mats had appropriate elongation at break (74.4 ± 9.5%) and tensile strength (3.0 ± 0.5 MPa) to be potentially used as wound dressing materials. In vivo studies were performed using eight to ten week-old male SKH1 hairless mice. The infection progression was evaluated through a semiquantitative method and quantitative polymerase chain reaction. The analyses of post-mortem samples indicated that THY-loaded PCL fibers acted as inhibitors of Staphylococcus aureus ATCC 25923 strain growth being as efficient as chlorhexidine (CLXD). Histopathological and immunohistochemical studies showed that the PCL-THY-treated wounds were almost free of an inflammatory reaction. Therefore, wound dressings containing natural compounds can prevent infection and promote wound healing and prompt regeneration. Copyrigh

Drug-eluting wound dressings having sustained release of antimicrobial compounds

Wound healing is a complex and costly public health problem that should be timely addressed to achieve a rapid and adequate tissue repair avoiding or even eliminating potential pathogenic infection. Chronic infected non-healing wounds represent a serious concern for health care systems. Efficient wound dressings with tailored therapy having the best response and highest safety margin for the management of chronic non-healing wounds are still needed. The use of novel wound dressing materials has emerged as a promising tool to fulfil these requirements. In this work, asymmetric electrospun polycaprolactone (PCL)-based nanofibers (NFs) were decorated with electrosprayed poly(lactic-co-glycolic acid) microparticles (PLGA MPs) containing the natural antibacterial compound thymol (THY) in order to obtain drug eluting antimicrobial dressings having sustained release. The synthesized dressings successfully inhibited the in vitro growth of Staphylococcus aureus ATCC 25923, showing also at the same doses cytocompatibility on human dermal fibroblasts and keratinocyte cultures after treatment for 24 h, which was not observed when using free thymol. An in vivo murine excisional wound splinting model, followed by the experimental infection of the wounds with S. aureus and their treatment with the synthesized dressings, pointed to the reduction of the bacterial load in wounds after 7 days, though the total elimination of the infection was not reached. The findings indicated the relevance of the direct contact between the dressings and the bacteria, highlighting the need to tune their design considering the wound surface and the nature of the antimicrobial cargo contained

Decreased pain in split-thickness skin graft donor sites with the use of a non-adherent polyurethane dressing

Donor sites of split-thickness skin grafts (STSGs) are painful and limit patient rehabilitation. We conducted this study to assess the efficacy of a non-adherent polyurethane dressing in reducing pain and its effect on the epithelialization rate of donor sites of STSGs. Methods: Fifteen patients requiring an STSG were included. In 10 patients the donor sites were randomly divided into two halves and covered with either a non-adherent polyurethane dressing or a standard non-adherent gauze. In five patients with bilateral donor sites, one side was covered with the non-adherent polyurethane dressing and the other with non-adherent gauze. The pain was assessed with a visual analog scale and epithelialization was also assessed, calculating non-epithelialized areas with image software by a blinded surgeon. Epithelialization of the wounds covered with the non-adherent polyurethane dressing was assessed at day 8 and 10 and those with non-adherent gauze at day 10. Results: Postoperative pain significantly decreased with the non-adherent polyurethane dressing during the length of the study (6.07 ± 1.46 vs. 1.72 ± 1.6) and at each time point (p < 0.001). Epithelialization was not affected with the polyurethane dressing, compared to the standard method

Implication of VHL, ERK5, and HIF-1alpha in clear cell renal cell carcinoma: Molecular basis

Objectives: To determine the expression status of several proteins related to VHL gene function and its relationship with common clinicopathological parameters. Material and methods: Observational, analytical, cross-sectional study with 50 patients diagnosed with clear cell renal cell carcinoma. The study analyzed VHL mutations and hypermethylation as well as protein expression of VHL, CA-IX, HIF-1alpha, VEGF, ERK1/2, and ERK5, relating them to clinical variables. A bivariate and multivariate descriptive logistical regression analysis was performed, using the presence of metastasis at diagnosis as dependent variable. Results: The study identified 13 (26%) VHL mutations related to nuclear grade (P = 0.036). VHL hypermethylation was found in 20% of cases. VHL expression was associated with the presence of mutations (P = 0.013), and the absence of expression was associated with nuclear grade and the presence of metastasis (P<0.05). HIF-1alpha was negative in only 5 cases. Vascular endothelial growth factor (VEGF) was positive in 31 of 47 cases and was associated with Fuhrman nuclear grade, presence of metastasis, and stage (P<0.05). ERK5 expression was increased in 58% of cases and associated with the presence of metastasis and more advanced stages (P<0.05). In the logistic regression analysis, the only variable remaining in the model was VEGF expression (P = 0.014). Conclusions: VEGF has prognostic value in clear cell renal cell carcinoma, and ERK5 may be a new prognostic marker in this type of tumor owing to its relationship with metastasis and more advanced stages

ANAIS-112 status: Two years results on annual modulation

ANAIS (Annual modulation with Nal Scintillators) is a dark matter direct detection experiment located at the Canfranc Underground Laboratory (LSC), in Spain. The goal is to confirm or refute in a model independent way the DAMA/LIBRA positive result: an annual modulation in the low-energy detection rate compatible with the expected signal induced by dark matter particles in the galactic halo. This signal, observed for about 20 years, is in strong tension with the negative results of other very sensitive experiments, but a direct comparison using the same target material, NaI(Tl), was still lacking. ANAIS-112, consisting of 112.5 kg of NaI(Tl) scintillators, is taking data at the LSC since August 2017. Here we present the preliminary annual modulation analysis corresponding to two years of data (exposure of 220.69 kgy) and the ANAIS-112 projected sensitivity for the scheduled 5 y of operation

Extracellular vesicles from Mycobacterium tuberculosis-infected neutrophils induce maturation of monocyte-derived dendritic cells and activation of antigen-specific Th1 cells

Tuberculosis remains one of the leading public health problems in the world. The mechanisms that lead to the activation of the immune response against Mycobacterium tuberculosis have been extensively studied, with a focus on the role of cytokines as the main signals for immune cell communication. However, less is known about the role of other signals, such as extracellular vesicles, in the communication between immune cells, particularly during the activation of the adaptive immune response. In this study, we determined that extracellular vesicles released by human neutrophils infected with M. tuberculosis contained several host proteins that are ectosome markers. In addition, we demonstrated that extracellular vesicles released by human neutrophils infected with M. tuberculosis released after only 30 min of infection carried mycobacterial antigens and pathogen-associated molecular patterns, and we identified 15 mycobacterial proteins that were consistently found in high concentrations in extracellular vesicles released by human neutrophils infected with M. tuberculosis; these proteins contain epitopes for CD4 T-cell activation. We found that extracellular vesicles released by human neutrophils infected with M. tuberculosis increased the expression of the costimulatory molecule CD80 and of the coinhibitory molecule PD-L1 on immature monocyte-derived dendritic cells. We also found that immature and mature dendritic cells treated with extracellular vesicles released by human neutrophils infected with M. tuberculosis were able to induce IFN-γ production by autologous M. tuberculosis antigen-specific CD4 T cells, indicating that these extracellular vesicles acted as antigen carriers and transferred mycobacterial proteins to the antigen-presenting cells. Our results provide evidence that extracellular vesicles released by human neutrophils infected with M. tuberculosis participate in the activation of the adaptive immune response against M. tuberculosis.This work was supported by Consejo Nacional de Ciencia y Tecnología (CONACYT grant A1-S-16113 to IEG) and by Secretaría de Investigación y Posgrado, Instituto Politécnico Nacional (IPN). L.V.-F., E.S.P., M.G.-M., and D.B. were recipients of CONACYT fellowships. J.S.-L., R.C.-S., S.E.-P., and I.E.-G. are fellows of Comisión de Operación y Fomento de Actividades Académicas (COFAA)–IPN. J.S.-L., R.C.-S., S.E.-P., I.E.-G., and I.W.-B. are fellows of Estímulos al Desempeño de los Investigadores (EDI)–IPN