Histone deacetylase (HDAC) inhibitors- based drugs are effective to control Mycobacterium tuberculosis infection and promote the sensibility for rifampicin in MDR strain

BACKGROUND Tuberculosis (TB) is a major public health problem, which has been aggravated by the alarming growth of drug-resistant tuberculosis. Therefore, the development of a safer and more effective treatment is needed. OBJECTIVES The aim of this work was repositioning and evaluate histone deacetylases (HDAC) inhibitors- based drugs with potential antimycobacterial activity. METHODS Using an in silico pharmacological repositioning strategy, three molecules that bind to the catalytic site of histone deacetylase were selected. Pneumocytes type II and macrophages were infected with Mycobacterium tuberculosis and treated with pre-selected HDAC inhibitors (HDACi). Subsequently, the ability of each of these molecules to directly promote the elimination of M. tuberculosis was evaluated by colony-forming unit (CFU)/mL. We assessed the expression of antimicrobial peptides and respiratory burst using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) FINDINGS Aminoacetanilide (ACE), N-Boc-1,2-phenylenediamine (N-BOC), 1,3-Diphenylurea (DFU), reduce bacillary loads in macrophages and increase the production of β-defensin-2, LL-37, superoxide dismutase (SOD) 3 and inducible nitric oxide synthase (iNOS). While only the use of ACE in type II pneumocytes decreases the bacterial load through increasing LL-37 expression. Furthermore, the use of ACE and rifampicin inhibited the survival of intracellular multi-drug resistance M. tuberculosis. MAIN CONCLUSIONS Our data support the usefulness of in silico approaches for drug repositioning to provide a potential adjunctive therapy for TB.

La contaminación del aire atmosférico en la respuesta inmune innata antimicobacteriana / Atmospheric air pollution and innate antimycobacterial immunity

Indoor and outdoor air pollution has been considered a serious public health problem worldwide, and is associated annually with around 7 million deaths (4.8 million associated with outdoor air and 2.2 million indoor air). The main reasons for these deaths include: chronic obstructive pulmonary diseases, pneumonia, ischemic cardiopathy and lung cancer. In addition, epidemiological studies have associated exposure to this type of pollutants with a greater susceptibility to the development of infectious and non-infectious diseases. One of the most important infectious diseases is tuberculosis, which over the years has worsened with the emergence of resistant multi-drug strains, as well as with association with other diseases such as diabetes mellitus type 2 and the acquired immune deficiency syndrome (AIDS). In fact, despite efforts made by the World Health Organization to stop the epidemic, a large number of deaths (about 1.7 million worldwide) are still caused by this disease. In this review, a brief summary will be made of the effects of exposure to indoor and outdoor air pollution on the innate immune response against tuberculosis, and how these alterations could be linked to the development of pulmonary tuberculosis.

La contaminación del aire de interiores y de exteriores ha sido considerada un serio problema de salud pública a nivel mundial, el cual se asocia anualmente con alrededor de 7 millones de muertes (4.8 millones asociadas con el aire de exteriores y 2.2 millones con el aire de interiores). Entre las principales razones ligadas a estas muertes se encuentran: enfermedad pulmonar obstructiva crónica, neumonía, cardiopatía isquémica y cáncer de pulmón. Además, estudios epidemiológicos han asociado la exposición a este tipo de contaminantes con mayor susceptibilidad para la aparición de enfermedades infecciosas y no infecciosas. Entre las enfermedades infecciosas, una de las más importantes es la tuberculosis, la cual, durante el transcurso de los años, ha empeorado con el surgimiento de cepas resistentes a múltiples fármacos, así como por la asociación con otras afecciones como la diabetes mellitus tipo 2 y el síndrome de inmunodeficiencia adquirida (sida). A pesar del esfuerzo realizado por la Organización Mundial de la Salud (OMS) por detener esta epidemia, aún existen anualmente un gran número de muertes causadas por esta enfermedad: alrededor de 1.7 millones a nivel mundial. En esta revisión se hará un breve resumen de los efectos que tiene la exposición a la contaminación del aire de interiores y de exteriores en la respuesta inmune innata en contra de la tuberculosis, y cómo estas alteraciones pueden estar ligadas a la aparición de tuberculosis pulmonar.

Evaluation of the transcriptional immune biomarkers in peripheral blood from warao indigenous associate with the infection by mycobacterium tuberculosis

Abstract INTRODUCTION: Biomarkers are critical tools for finding new approaches for controlling the spread of tuberculosis (TB), including for predicting the development of TB therapeutics, vaccines, and diagnostic tools. METHODS: Expression of immune biomarkers was analyzed in peripheral blood cells stimulated and non-stimulated with M. tuberculosis antigens ESAT-6, CFP10 and TB7.7. in Warao indigenous individuals. These biomarkers may be able to differentiate TB states, such as active tuberculosis (ATB) cases and latent tuberculosis infection (LTBI) from non-infected controls (NIC). A real-time reverse transcription polymerase chain reaction (RT-qPCR) assay was performed on 100 blood samples under non-stimulation or direct ex vivo conditions (NS=50) and stimulation conditions (S=50). RESULTS: The findings are shown as the median and interquartile range (IQR) of relative gene expression levels of IFN-γ, CD14, MMP9, CCR5, CCL11, CXCL9/MIG, and uPAR/PLAUR immune biomarkers. MMP9 levels were significantly higher in the LTBI-NS and LTBI-S groups compared with the NIC-NS and NIC-S groups. However, CCR5 levels were significantly lower in the LTBI-S group compared with both NIC-NS and NIC-S groups. CCL11 levels were significantly lower in the LTBI-S group compared with the NIC-NS group. CONCLUSIONS: Preliminary findings showed that MMP9 immune biomarkers separated LTBI indigenous individuals from NIC indigenous individuals, while CCR5, CCL11, CD14, and IFN-γ did not differentiate TB states from NIC. MMP9 may be useful as a potential biomarker for LTBI and new infected case detection among Warao indigenous individuals at high risk of developing the disease. It may also be used to halt the epidemic, which will require further validation in larger studies.

Variability in the virulence of specific Mycobacterium tuberculosis clinical isolates alters the capacity of human dendritic cells to signal for T cells

BACKGROUND Once in the pulmonary alveoli, Mycobacterium tuberculosis (Mtb) enters into contact with alveolar macrophages and dendritic cells (DCs). DCs represent the link between the innate and adaptive immune system owing to their capacity to be both a sentinel and an orchestrator of the antigen-specific immune responses against Mtb. The effect that the virulence of Mtb has on the interaction between the bacilli and human DCs has not been fully explored. OBJECTIVE To evaluate the effect of Mtb virulence on human monocyte-derived DCs. METHODS We exposed human monocyte-derived DCs to Mtb clinical strains (isolated from an epidemiological Mtb diversity study in Mexico) bearing different degrees of virulence and evaluated the capacity of DCs to internalise the bacilli, control intracellular growth, engage cell death pathways, express markers for activation and antigen presentation, and expand to stimulate autologous CD4+ T cells proliferation. FINDINGS In the case of the hypervirulent Mtb strain (Phenotype 1, strain 9005186, lineage 3), we report that DCs internalise and neutralise intracellular growth of the bacilli, undergo low rates of apoptosis, and contribute poorly to T-cell expansion, as compared to the H37Rv reference strain. In the case of the hypovirulent Mtb strain (Phenotype 4, strain 9985449, lineage 4), although DCs internalise and preclude proliferation of the bacilli, the DCs also display a high level of apoptosis, massive levels of apoptosis that prevent them from maintaining autologous CD4+ T cells in a co-culture system, as compared to H37Rv. MAIN CONCLUSIONS Our findings suggest that variability in virulence among Mtb clinical strains affects the capacity of DCs to respond to pathogenic challenge and mount an immune response against it, highlighting important parallels to studies previously done in mouse models.

Los péptidos antimicrobianos y sus beneficios en el tratamiento de las úlceras de pie diabético / The benefits of antimicrobial peptides in treating diabetic foot ulcers

Diabetic foot ulcers are one of the most important complications of diabetes mellitus type 2. In fact, up to 30% of diabetic patients develop this complication. In recent years new therapies have been implemented to try to promote rapid healing in this type of ulcers, focusing on antibiotic therapy, restoration of vascularization, induction of healing and counteracting neuropathy. Antimicrobial peptides are versatile molecules that induce wound healing, have antimicrobial activity and also promote angiogenesis. Because of these properties, they have been proposed as adjuvants in the therapy for diabetic foot ulcers. In order to avoid excessive costs in the treatment of diabetic ulcers with the use of synthetic AMP, some research groups have chosen to induce peptides with exogenous molecules. Many molecules are capable of inducing antimicrobial peptides both in blood cells and in endothelial cells. Research into the use of AMP as adjuvants in the treatment of diabetic foot ulcers continues in several lines of research, such as synthesis of peptides with specific angiogenic activities, use of peptides from insects or amphibians, as well as the use of synthetic peptides. In the present review, we analyze the possible use of AMP and their inducers in the treatment of diabetic foot ulcers based on the reported studies either in vivo or in vitro.

La úlcera de pie diabético es una de las complicaciones más importantes de la diabetes mellitus tipo 2; de hecho, hasta un 30% de los pacientes diabéticos presentan esta complicación. En los últimos años se han implementado nuevas terapias para tratar de promover la rápida cicatrización de este tipo de úlceras, enfocándose en la terapia antibiótica, el restablecimiento de la vascularización, la inducción de la cicatrización y contrarrestar la neuropatía. Los péptidos antimicrobianos son moléculas versátiles que inducen cicatrización, tienen actividad antimicrobiana y además promueven angiogénesis; dadas estas propiedades se ha propuesto su uso como adyuvantes en la terapia de las úlceras de pie diabético. Con el fin de evitar costos excesivos en el tratamiento de las úlceras diabéticas con el uso AMP sintéticos, algunos grupos de investigación han optado por inducir péptidos con moléculas exógenas. Existen muchas moléculas capaces de inducir péptidos antimicrobianos tanto en células sanguíneas como en células epiteliales. La investigación en el uso de AMP como coadyuvantes en el tratamiento de las úlceras de pie diabético sigue avanzando en varias líneas de investigación: síntesis de péptidos con actividades angiogénicas específicas, uso de péptidos de insectos o anfibios, así como el uso de péptidos sintéticos. En la presente revisión se analiza, sobre la base de los estudios realizados tanto in vivo como in vitro el posible uso de AMP y sus inductores en el tratamiento de úlceras de pie diabético.

Type-2 diabetes alters the basal phenotype of human macrophages and diminishes their capacity to respond, internalise, and control Mycobacterium tuberculosis

BACKGROUND Type 2 diabetes (T2D) is a risk factor for the development of tuberculosis (TB), although the associated mechanisms are not known. OBJECTIVES To study the association between T2D and the basal phenotype of macrophages, and their immune response to Mycobacterium tuberculosis (Mtb) infection. METHODS We evaluated the influence of T2D on the response of monocyte-derived macrophages (MDM) to Mtb in patients with T2D (n = 10) compared to healthy subjects (n = 9), before and after infection with Mtb clinical isolates bearing different degrees of virulence. The levels of cell surface markers for activation secreted cytokines and chemokines, bacterial association, and intracellular bacterial growth were evaluated. FINDINGS The expression levels of HLA-DR, CD80, and CD86 were low while those of of PD-L1 were high in uninfected MDMs derived from patients with diabetes; as a result of Mtb infection, changes were only observed in the expression levels of PD-L1. The levels of cytokines (e.g., IL-6, IL-1β, IL-10, and IL-12) and chemokines (e.g., MCP-1, MIG, and RANTES) are perturbed in MDMs derived from patients with diabetes, both before infection and in response to Mtb infection. In response to the more virulent Mtb strains, the levels of association and bacterial clearance were diminished in MDMs derived from patients with diabetes. CONCLUSIONS T2D affects the basal activation state of the macrophages and its capacity to respond and control Mtb infection.

Concordance between IFNγ gene +874 A/T polymorphism and interferon-γ expression in a TB-endemic indigenous setting

Abstract INTRODUCTION: Interferon-γ (IFN-γ) plays a crucial role in resistance to mycobacterial diseases; accordingly, variants of the gene encoding this cytokine may be associated with elevated risk of contracting pulmonary tuberculosis (TB). METHODS: Blood samples were collected from 135 Warao indigenous individuals with newly diagnosed sputum culture-positive TB. Of these, 24 were diagnosed with active tuberculosis (ATB). The study comprised 111 participants, who were grouped as follows: 1) 14 tuberculin skin test (TST)-positive Warao indigenous individuals and 4 that were QuantiFERON-TB?Gold In-Tube (QFT-IT) test-positive, collectively comprising the latent TB infection group (LTBI), n = 18), and 2) healthy controls who were QFT-IT- and TST-negative, comprising the control group (CTRL, n = 93). Detection of the IFN γ gene (IFNG) +874A/T polymorphism was performed via PCR and quantification of IFNG expression via qPCR. RESULTS: Relative to indigenous and white Americans, ATB and CTRL groups had a higher frequency of the IFNG SNP (+874A): 23 (95.8%) and 108 (97.3%), respectively. Indigenous Warao individuals homozygous for the IFNG (+874) A allele exhibited 3.59-fold increased risk of developing TB (95% confidence interval, 2.60-4.96, p =0.0001). A decreased frequency of the AT genotype was observed in individuals with pulmonary TB (4.16%) and controls (0.90%). The frequency of the TT genotype was decreased among controls (1.80%); none of the patients with TB were found to have this genotype. The differences in IFNG expression between the groups, under unstimulated and stimulated conditions, were not statistically significant. CONCLUSIONS: Preliminary results demonstrate concordance between IFNG +874 A/A genotype and low expression of IFNG.

Seropositivity for ascariosis and toxocariosis nd cytokine expression among the indigenous people in the Venezuelan delta region / Seropositividad para ascariosis y toxocariosis y expresión de citocinas entre la población indígena de la región del delta Venezolano

The present study aimed at measuring seropositivities for infection by Ascaris suum and Toxocara canis using the excretory/secretory (E/S) antigens from Ascaris suum (AES) and Toxocara canis (TES) within an indigenous population. In addition, quantification of cytokine expressions in peripheral blood cells was determined. A total of 50 Warao indigenous were included; of which 43 were adults and seven children. In adults, 44.1% were seropositive for both parasites; whereas children had only seropositivity to one or the other helminth. For ascariosis, the percentage of AES seropositivity in adults and children was high; 23.3% and 57.1%, respectively. While that for toxocariosis, the percentage of TES seropositivity in adults and children was low; 9.3% and 14.3%, respectively. The percentage of seronegativity was comparable for AES and TES antigens in adults (27.9%) and children (28.6%). When positive sera were analyzed by Western blotting technique using AES antigens; three bands of 97.2, 193.6 and 200.2 kDas were mostly recognized. When the TES antigens were used, nine major bands were mostly identified; 47.4, 52.2, 84.9, 98.2, 119.1, 131.3, 175.6, 184.4 and 193.6 kDas. Stool examinations showed that Blastocystis hominis, Hymenolepis nana and Entamoeba coli were the most commonly observed intestinal parasites. Quantification of cytokines IFN-γ, IL-2, IL-6, TGF-β, TNF-α, IL-10 and IL-4 expressions showed that there was only a significant increased expression of IL-4 in indigenous with TES seropositivity (p < 0.002). Ascaris and Toxocara seropositivity was prevalent among Warao indigenous.

El objetivo del presente estudio fue determinar la seropositividad de infección por Ascaris suum y Toxocara canis, utilizando antígenos de excreción/secreción (E/S) de Ascaris suum (AES) y Toxocara canis (TES) en una población indígena. Adicionalmente, se cuantificó la expresión de citocinas a partir de células de sangre periférica. Un total de 50 indígenas Warao se incluyeron en el estudio; 43 fueron adultos y 7 niños. Entre los adultos, 44,1% fueron seropositivos para ambos parásitos; mientras que los niños sólo mostraron seropositividad a uno u otro de los helmintos. Para ascariosis, el porcentaje de seropositividad para los antígenos AES fue alto tanto en adultos como en niños; 23,3% y 57,1%, respectivamente. Para toxocariosis, el porcentaje de seropositividad para los antígenos TES fue bajo en adultos así como en niños; 9,3% y 14,3%, respectivamente. El porcentaje de seronegatividad fue similar tanto para los antígenos AES como para TES en adultos (27,9%) y niños (28,6%). Cuando la seropositividad fue analizada a través de la técnica de Western blotting utilizando los antígenos AES; 3 bandas de 97,2, 193,6 y 200,2 kDas fueron principalmente reconocidas. Para los antígenos TES, 9 bandas fueron mayormente identificadas; 47,4, 52,2, 84,9, 98,2, 119,1, 131,3, 175,6, 184,4 y 193,6 kDas. Los análisis coproparasitológicos mostraron que los parásitos Blastocystis hominis, Hymenolepis nana y Entamoeba coli fueron los parásitos intestinales más comúnmente observados. La cuantificación de la expresión de las citocinas IFN-γ, IL-2, IL-6, TGF-β, TNF-α, IL-10 e IL-4 mostró que hubo un significante incremento de la expresión de IL-4 entre los indígenas con seropositividad para los antígenos TES (p < 0.002). La seropositividad para Ascaris y Toxocara fue prevalente entre los indígenas Warao.

Tuberculin skin test and interferon-gamma release assay values are associated with antimicrobial peptides expression in polymorphonuclear cells during latent tuberculous infection

It has been reported that patients with progressive tuberculosis (TB) express abundant amounts of the antimicrobial peptides (AMPs) cathelicidin (LL-37) and human neutrophil peptide-1 (HNP-1) in circulating cells, whereas latent TB infected donors showed no differences when compared with purified protein derivative (PPD) and QuantiFERON®-TB Gold (QFT)-healthy individuals. The aim of this study was to determine whether LL-37 and HNP-1 production correlates with higher tuberculin skin test (TST) and QFT values in TB household contacts. Twenty-six TB household contact individuals between 26-58 years old TST and QFT positive with at last two years of latent TB infection were recruited. AMPs production by polymorphonuclear cells was determined by flow cytometry and correlation between TST and QFT values was analysed. Our results showed that there is a positive correlation between levels of HNP-1 and LL-37 production with reactivity to TST and/or QFT levels. This preliminary study suggests the potential use of the expression levels of these peptides as biomarkers for progression in latent infected individuals.

Péptidos antimicrobianos: un probable arsenal contra la infección por VIH / Antimicrobial peptides: a potential arsenal against HIV infection

La infección por VIH (virus de la inmunodeficiencia humana) en la actualidad es un grave problema de salud pública a nivel mundial, que requiere de nuevas estrategias vacunales para detener su propagación así como para su efectivo tratamiento. Algunos estudios relacionados con la inmunidad innata en contra de VIH, han demostrado que los péptidos antimicrobianos (AMP´s) pueden generar resistencia a las infecciones virales. En la presente revisión, se describen a los péptidos antimicrobianos de humano y su actividad en contra de VIH así como péptidos de otras especies como plantas, anfibios, insectos y varias especies de animales que poseen un potencial terapéutico o profiláctico en la infección por VIH. Se describen brevemente algunos mecanismos mediante los cuales estos péptidos pueden bloquear la replicación e infección por el VIH.

HIV (human immunodeficiency virus) infection is today a very important health issue worldwide, which demands new ways and strategies for its prevention and treatment. Several studies on the innate immunity against HIV infection have shown that antimicrobial peptides are associated with increased resistance to infection. In the present review, we briefly summarize the major characteristics of antimicrobial peptides from human and several species of plants, amphibians, insects and other animal species that have significant potential to be used as therapeutic or prophylactic agents. The mechanisms of infection inhibition and viral replication blockade are also described in the context of the biology of infection.

Células epiteliales en la inmunidad del pulmón / Immunological role of lung epithelial cells

El sistema respiratorio se encuentra en contacto con patógenos, sin embargo, gracias a la respuesta inmune innata de este, solo en raras ocasiones se establece la enfermedad. Las células epiteliales del tracto respiratorio juegan un papel importante para evitar la colonización del pulmón por agentes infecciosos, identificando a los microorganismos a través de receptores especializados, como los Toll-like,. Las células epiteliales son capaces de secretar citocinas, péptidos antimicrobianos y otras moléculas proinflamatorias las cuales evitan el establecimiento de patógenos

Péptidos antimicrobianos en la inmunidad innata de enfermedades infecciosas / Antimicrobial peptides in the innate immunity of infectious diseases

Los péptidos antimicrobianos son moléculas efectoras clave en la inmunidad innata. Generalmente contienen de 15 a 45 residuos de aminoácidos y en su gran mayoría poseen carga positiva, adem s de que tienen la propiedad de ser anfipáticos. Estos péptidos son secretados por células epiteliales y leucocitos, como es el caso de los macráfagos y neutrófilos. En la actualidad, con base en la traducción de secuencias en programas computacionales se han descrito m s de 800 tipos de péptidos antimicrobianos distribuidos en los reinos animal y vegetal. Estos péptidos pueden ser clasificados de acuerdo con su conformación estructural y la ubicación de sus puentes disulfuro. Las defensinas constituyen uno de los tipos de péptidos antimicrobianos más estudiados, y se dividen en dos familias de acuerdo con la ubicación de sus puentes disulfuro: alfa-defensinas y beta-defensinas. Algunas de estas defensinas pueden ser inducidas por citocinas proinflamatorias, así como por moléculas propias de patógenos, y se ha observado que est n relacionadas con la inmunopatogenia de varias enfermedades. El papel principal de los péptidos antimicrobianos es la lisis directa de microorganismos; sin embargo, a la fecha también se han descrito propiedades quimiotácticas, que le permiten modular el sistema inmune y de esta forma constituir un puente entre la inmunidad innata y la inmunidad adaptativa. Actualmente se han iniciado estudios con la posibilidad de utilizar esta clase de moléculas como nuevos fármacos en diferentes tipos de enfermedades infecciosas.

Papel de las células epiteliales en la respuesta inmune del pulmón / Immunological role of epithelial cells of the lung

El sistema respiratorio se encuentra en contacto con agentes patógenos; sin embargo, gracias a la respuesta inmune innata de éste, sólo en raras ocasiones se produce la enfermedad. Las células epiteliales del tracto respiratorio desempeñan un papel importante para evitar la colonización del pulmón por agentes infecciosos, identificando a los microorganismos a través de receptores especializados como los toll-like. Asimismo, son capaces de secretar citocinas, péptidos antimicrobianos y otras moléculas proinflamatorias, las cuales evitan el establecimiento de patógenos.

The respiratory tract is one of the main systems which is in perennial contact with a wide variety of pathogenic microorganisms; however, infection is seldom produced due to its innate immune response. Respiratory tract epithelial cells play a very important role to avoid colonization of the lung by infectious agents, because they recognize microbial molecules through very specialized receptors, such as toll-like receptors; moreover, these cells posses a broad variety of molecules which are related to local immunity. Respiratory tract epithelial cells produce chemokines, antimicrobial peptides and other proinflammatory molecules that prevent the establishment of pathogenic microorganisms.

Respuesta de inmunidad celular en la tuberculosis pulmonar: revisión / Cell immunity response in human pulmonary tuberculosis: review

La tuberculosis (TBC) pulmonar humana, un problema de salud pública en el mundo, es una enfermedad infecciosa causada por Mycobacterium tuberculosis. Se conoce que una minoría de las personas que son infectadas por M. tuberculosis son capaces de progresar a enfermedad clínica. Se puede decir, en términos generales, que el 90 por ciento de las personas tendrán controlados los bacilos en estado latente para toda la vida, por medio de su sistema inmunológico. Un 5 por ciento presentará TBC primaria progresiva y el otro 5 por ciento presentará la enfermedad en estados tardíos de la vida, lo que se denomina TBC de reactivación o post-primaria. En los individuos resistentes, el control de la infección o de los bacilos tuberculosos que se encuentran en la región alveolar requiere principalmente del desarrollo de una respuesta de inmunidad celular (RIC) del tipo Th1. Este tipo de respuesta incluye la participación de los macrófagos alveolares, los linfocitos T CD4+ y CD8+, principalmente los linfocitos T gd y la producción de citocinas como: IL-2, IFN-g, IL-12, IL-18 y TNF-a. Aunado están las quimiocinas como RANTES, MCP-1, MIP-1a e IL-8 que juegan un papel muy importante en la migración de las diferentes subpoblaciones celulares al sitio de infección para la formación del granuloma. Además, es primordial el papel de las células "natural killer" (NK), y de las células epiteliales como parte de la respuesta de inmunidad innata