Citoquinas pro y anti-inflamatorias en la infección por SARS-CoV-2, en población de Colombia / Pro- and anti-inflammatory cytokines associated to SARS-COV-2 infection in the Colombian population

Resumen La infección por SARS-CoV-2 es hoy el principal problema de salud pública en el mundo. No es claro el papel de las citoquinas en la fisiopatología del COVID-19,que en algunos individuos presenta una progresión rápida, severa y mortal asociada con proinflamación sistémicos relacionada con coagulopatías y fallas multiorgánicas. En este estudio, evaluamos los niveles séricos de citoquinas y su correlación con IgM, IgG e IgA, en 24 muestras de individuos positivos y 8 muestras de individuos negativos, para SARS-CoV-2. Hallamos concentraciones significativamente menores de IFN-g, TNF, IL-2 e IL-4 y un aumento significativo de IL-6 en el grupo de infectados hospitalizados respecto a los no infectados, así como una tendencia significativa al aumento, para IgG e IgA en el mismo grupo de individuos, respecto a infectados asintomáticos. Nuestros datos soportan el papel de la IL-6 en la severidad de la enfermedad destacando su potencial papel como biomarcador en la prognosis de esta patología. También, soportan la hipótesis sobre la función de los anticuerpos en el control efectivo del patógeno; se observa una respuesta inmune humoral más débil, frente a la proteína de la nucleocápside viral, en individuos con un mejor curso de la enfermedad.

Abstract The emergency caused by the infection in humans of SARS-COV-2 and the clinical syndrome resulting from the infection (COVID-19) is a major public health crisis with global repercussions. Currently, the role of different cytokine profiles in the infection pathophysiology and its outcome remains unclear despite the coordina ted efforts of the scientific community. COVID-19 shows a rapid progression where the disease severity and mortality are linked to systemic pro-inflammatory pro cesses associated to a dysregulation in the cytokine production balance, resulting in blood clothing disorders and multiorgan failure. Here we evaluate the serum concentration for a cytokine panel as well as the antibody titers of IgM, IgG and IgA from 24 individuals who tested positive for SARS-CoV-2 by RT-PCR (divided into three separate groups according to disease severity) and eight RT-PCR-negative controls. Significantly lower concentrations of IFN-g, TNF, IL-2 and IL-4, and a higher production of IL-6 were observed in hospitalized COVID-19 patients when compared to SARS-CoV-2-negative individuals. Furthermore, a significant and sustained increase in the levels of IgG and IgA was found for the group of hospitalized patients compared to asymptomatic SARS-CoV-2-positive individuals. Our data support previous findings on the role of cytokines like IL-6 in the severity of the disease and highlight their potential use as biomarkers for the prognosis of COVID-19. Finally, we provide evidence supporting the potential function of the antibody response in the effective control of the virus, showing that a somehow weaker humoral immune response can be associated to milder forms of COVID-19.

Antileishmanial activity of synthetic analogs of the naturally occurring quinolone alkaloid N-methyl-8-methoxyflindersin.

Leishmaniasis is a neglected, parasitic tropical disease caused by an intracellular protozoan from the genus Leishmania. Quinoline alkaloids, secondary metabolites found in plants from the Rutaceae family, have antiparasitic activity against Leishmania sp. N-methyl-8-methoxyflindersin (1), isolated from the leaves of Raputia heptaphylla and also known as 7-methoxy-2,2-dimethyl-2H,5H,6H-pyran[3,2-c]quinolin-5-one, shows antiparasitic activity against Leishmania promastigotes and amastigotes. This study used in silico tools to identify synthetic quinoline alkaloids having structure similar to that of compound 1 and then tested these quinoline alkaloids for their in vitro antiparasitic activity against Leishmania (Viannia) panamensis, in vivo therapeutic response in hamsters suffering from experimental cutaneous leishmaniasis (CL), and ex vivo immunomodulatory potential in healthy donors' human peripheral blood (monocyte)-derived macrophages (hMDMs). Compounds 1 (natural), 2 (synthetic), and 8 (synthetic) were effective against intracellular promastigotes (9.9, 3.4, and 1.6 µg/mL medial effective concentration [EC50], respectively) and amastigotes (5.07, 7.94, and 1.91 µg/mL EC50, respectively). Compound 1 increased nitric oxide production in infected hMDMs and triggered necrosis-related ultrastructural alterations in intracellular amastigotes, while compound 2 stimulated oxidative breakdown in hMDMs and caused ultrastructural alterations in the parasite 4 h posttreatment, and compound 8 failed to induce macrophage modulation but selectively induced apoptosis of infected hMDMs and alterations in the intracellular parasite ultrastructure. In addition, synthetic compounds 2 and 8 improved the health of hamsters suffering from experimental CL, without evidence of treatment-associated adverse toxic effects. Therefore, synthetic compounds 2 and 8 are potential therapeutic candidates for topical treatment of CL.

Seco-limonoid derived from Raputia heptaphylla promotes the control of cutaneous leishmaniasis in hamsters (Mesocricetus auratus).

The rational search of novel bioactive molecules against pathogens with immunomodulatory activity is presently one of the most significant approaches to discover and design new therapeutic agents for effective control of infectious diseases, such as the infection caused by Leishmania parasites. In the present study, we evaluated the therapeutic efficacy of the recently characterized immunomodulatory compound 11α,19ß-dihydroxy-7-acetoxy-7-deoxoichangin, a seco-limonoid derived from the bark of Raputia heptaphylla (Pittier) using: (1) peritoneal macrophages and (2) Mesocricetus auratus hamsters infected with Leishmania (V.) panamensis and Leishmania (L.) amazonensis. We observed the ability of this seco-limonoid to induce the effective control of the parasite either in vitro [determining an effective concentration 50 (EC50) of 59 µ m at the infection model] and in vivo (inducing clinical improvement or even cure in infected animals treated compared with the groups of animals treated with vehicle solution or meglumine antimoniate).

seco-limonoids and quinoline alkaloids from Raputia heptaphylla and their antileishmanial activity.

A novel seco-limonoid, rel-(1S,5R,9S,7R,8S,9R,10S,11R,13S,14R,15R,17R)-11,19-dihydroxy-7-acetoxy-7-deoxoichangin (raputiolide) (1), and two novel quinolone alkaloids N-methyl-2-phenoxyquinolin-4(1H)-one (heptaphyllone A) (2) and 6-methylbenzofuro[2,3-b]quinolin-4(1H)-one (heptaphyllone B) (3), along with the known seco-limonoid ichangin (4), were isolated from Raputia heptaphylla PITTIER (Rutaceae) stem bark. Five known alkaloids, N-methyl-8-methoxyflindersine (5), skimmianine (6), kokusaginine (7), dictamnine (8) and flindersiamine (9), were also isolated from R. heptaphylla leaves. Their structures were established on the basis of full spectroscopic data interpretation supported by data from the pertinent literature. seco-Limonoid 1 configuration was determined by enhanced nuclear Overhauser effect spectroscopy (NOESY) experiments and density functional theory (DFT) molecular modeling. The antileishmanial effect of the isolated compounds was evaluated on Leishmania Viannia panamensis (promastigotes and amastigotes). Whereas alkaloids 2-3, 6-8 and limonoid 4 exhibited no significant parasitocide activity against internalized L. (V.) panamensis amastigotes, limonoid 1 and alkaloid 5 had leishmanicidal activity on intracellular amastigotes (EC50: 8.7 µg/ml) and promastigotes (EC(50): 14.3 µg/ml), respectively.

Pharmacogenetic impact of VKORC1 and CYP2C9 allelic variants on warfarin dose requirements in a hispanic population isolate.

Warfarin is the most prescribed oral anticoagulant worldwide. Because of the complexity of warfarin therapy, we attempted to dissect genetic from bioenvironmental factors influencing warfarin dose responses in individuals of a genetic isolate of Hispanic ancestry. A total of 191 patients with standard values of international normalized ratio were recruited. Three groups with a significantly different warfarin dose response were identified, that is, sensitive (2.28 +/- 0.50 mg/d), intermediate (4.2 +/- 0.76 mg/d), and resistant (7.40 +/- 1.54 mg/d; Tukey test, P < .001). Age had a significant inverse correlation with warfarin dose (P < .001; effective dose diminished 0.56 mg/d/decade). Required doses were higher for individuals with CYP2C9 variants containing the allele *1 compared to those individuals with variants composed of other alleles (P = .006). Similarly, individuals with VKORC1-1639GG and VKORC1-1639GA genotypes also required higher doses compared to the AA genotype (P < .001). Evaluation of potential gene-gene interactions between CYP2C9 and VKORC1 polymorphisms showed significant differences in dosing for CYP2C9 genotypes within the VKORC1-1639G/A subgroup (P = .013). A stepwise multivariate linear regression analysis showed that 38.2% of the warfarin dose response variance was accounted for by a model involving age (20.9%), VKORC1-1639G/A (11.3%), and CYP2C9*1, *2, and *3 variants (7.1%). These results corroborate previous findings on warfarin pharmacogenetics and define a contrastable gene-bioenvironment interaction model suited to be used in Hispanic populations.

Evaluación in vitro de la actividad de aceites esenciales de plantas colombianas sobre Leishmania braziliensis / In vitro evaluation of essentials oils activity from Colombians plants on Leishmania braziliensis

Las leishmaniosis son zoonosis que en el hospedero humano pueden afectar la piel, las mucosas o las vísceras, resultado del parasitismo de los macrófagos por un protozoario flagelado del género Leishmania, introducido al organismo por la picadura de un insecto flebótomo. En Colombia, la leishmaniosis cutánea es causada por parásitos del subgénero Viannia, al cual pertenecen las especies Leishmania (L) panamensis, L. braziliensis y L. guyanensis. El tratamiento clínico requiere la administración de medicamentos que, si bien son efectivos, generan efectos adversos en el individuo. Además, se ha detectado una resistencia del parásito, lo que ha favorecido la disminución de la eficacia de los tratamientos usados convencionalmente para controlar la enfermedad. En la búsqueda de nuevas y más seguras alternativas terapéuticas, los productos naturales son una fuente importante de agentes con potencial actividad antileishmanial. Éste es el caso de los aceites esenciales que por sus características físico-químicas, principalmente por su untuosidad, pueden constituirse en una alternativa tópica para el control de la leishmaniosis cutánea (lc). En este estudio, la actividad antileishmanial de trece aceites esenciales de diferentes familias de plantas colombianas (Lauraceae, Rutaceae, Verbenaceae, Lamiaceae, Zingiberaceae, Myristicaceae, Cardiopteridaceae y Pinaceae) se evaluó sobre los promastigotes de L. braziliensis. Si bien, es necesaria la valoración del efecto citotóxico que sobre los macrófagos, como células blanco de la infección por Leishmania, puedan generar estos aceites; resulta promisoria la actividad antileishmanial directa sobre las formas promastigotes, la cual puede ser comparable con la observada para pentamidina, fármaco que a diferencia de las sales antimoniales pentavalentes, no es una prodroga que requiera ser descompuesta en metabolitos activos una vez es incorporada al macrófago.

Leishmaniasis is zoonosis that in human host can affect skin, mucosa and viscera, when a flagelar protozoo is phagocyted by macrophages after bit of Phlebotomineo insect. In Colombia, cutaneous leishmaniasis is produced by subgenus Viannia parasites, genus that includes species like Leishmania panamensis (L), L. braziliensis y L. guyanensis. Clinical treatment requires the administration of effective drugs that induce severe adverse effects in the patient treated. Additionally, the parasite had generated a drug-resistance, which promotes a reduction in this kind of therapeutic schedule. In the searching of new and safer therapeutics alternatives to leishmaniasis control, natural products are a important source of active molecules with pharmacological activity. In this study we reported the antileishmanial activity of 13 essentials oils of different Colombians plants (Lauraceae, Rutaceae, Verbenaceae, Lamiaceae, Zingiberaceae, Myristicaceae, Cardiopteridaceae y Pinaceae) using L. braziliensis promastigotes. The effective concentration 50 (EC50) was 87,8 +/- 55,51 μg/mL, 265,5 +/- 7,5 μg/mL and 17,4 +/- 0,43 μg/mL for essentials oils of Persea caerulea fruit, Lippia alba leaves and Rosmarinus officinalis leaves, respectively, which permit suggest us, a parasitocide property. Although, it is necessary the cytotoxic effect study on macrophages as infection target cells, result promissory the antileishmanial effect direct to promastigote form, which can be comparable with the effect observed to pentamidine, drugs to a difference of antimonial drugs, is not a prodrug that require to be metabolized into macrophage. In other words, essentials oils would contain active principles lethal against free forms and maybe, against intracellular Leishmania spp forms that need further studies to be considered as therapeutic alternative to leishmaniasis cutaneous control.