Development of a Resveratrol Nanoformulation for the Treatment of Diabetic Retinopathy.

Diabetic retinopathy (RD) is a microvascular disease that can cause the formation of fragile neovessels, increasing the risk of hemorrhages and leading to vision loss. Current therapies are based on the intravitreal injection of anti-VEGF (vascular endothelial growth factor), which is invasive and can cause secondary effects. The development of new treatments that complement the current therapies is necessary to improve the patient's outcomes. Nanostructured formulations offer several advantages regarding drug delivery and penetration. In this research, a resveratrol nanosuspension (RSV-NS) was prepared and characterized using dynamic light scattering, field emission scanning electron microscopy, and infrared spectroscopy. The RSV-NS had an average particle size of 304.0 ± 81.21 nm with a PDI of 0.225 ± 0.036, and a spherical-like morphology and uniform particle distribution. Cell viability, proliferation, and migration were tested on endothelial cells (HMRECs). RSV-NS in a concentration of less than 18.75 µM did not have a cytotoxic effect on HMRECs. Likewise, proliferation and migration were significantly reduced compared to the unstimulated control at 37.5 µM. The RSV-NS did not present cytotoxic effects but decreased cell proliferation and migration, indicating that it could provide an important contribution to future medical implementations and could have a high potential to treat this disease.

Mutations in SORL1 and MTHFDL1 possibly contribute to the development of Alzheimer's disease in a multigenerational Colombian Family.

Alzheimer's disease (AD) is the most common cause of dementia in the elderly, affecting over 50 million people worldwide in 2020 and this number will triple to 152 million by 2050. Much of the increase will be in developing countries like Colombia. In familial forms, highly penetrant mutations have been identified in three genes, APP, PSEN1, and PSEN2, supporting a role for amyloid-ß peptide. In sporadic forms, more than 30 risk genes involved in the lipid metabolism, the immune system, and synaptic functioning mechanisms. We used whole-exome sequencing (WES) to evaluate a family of 97 members, spanning three generations, with a familiar AD, and without mutations in APP, PSEN1, or PSEN2. We sequenced two affected and one unaffected member with the aim of identifying genetic variants that could explain the presence of the disease in the family and the candidate variants were validated in eleven members. We also built a structural model to try to determine the effect on protein function. WES analysis identified two rare variants in SORL1 and MTHFD1L genes segregating in the family with other potential risk variants in APOE, ABCA7, and CHAT, suggesting an oligogenic inheritance. Additionally, the structural 3D models of SORL1 and MTHFD1L variants shows that these variants produce polarity changes that favor hydrophobic interactions, resulting in local structural changes that could affect the protein function and may contribute to the development of the disease in this family.

A Langmuir-Blodgett Study of the Interaction between Amphotericin B and Lipids of Histoplasma capsulatum.

Histoplasma capsulatum is a dimorphic, thermal, and nutritional fungus. In the environment and at an average temperature of 28 °C, it develops as a mold that is composed of infecting particles. Once in the host or in cultures at 37 °C, it undergoes a transition into the parasitic form. In the present work, we performed chemical extraction and characterization using chromatography techniques of the associated lipid composition of the external surface of the cell wall of the mycelial phase of two isolates of the H. capsulatum: one clinical and one environmental. Several differences were evidenced in the fatty acids in the phospholipid composition. Surface pressure-area isotherms and compression module curves of the Amphotericin B and lipid extract monolayers, as well as (AmB)-lipid extract mixed monolayers were recorded. Results show a high affinity of AmB towards lipid extracts. The most stable monolayers were formed by AmB + environmental with a mass ratio of 1:3 and AmB + clinical with a mass ratio of 1:2. Knowledge of the AmB aggregation processes at a molecular level and the characterization of the lipid extracts allows the possibility to understand the interaction between the AmB and the lipid fractions of H. capsulatum.

Structural Predictive Model of Presenilin-2 Protein and Analysis of Structural Effects of Familial Alzheimer's Disease Mutations.

Alzheimer's disease manifests itself in brain tissue by neuronal death, due to aggregation of ß-amyloid, produced by senile plaques, and hyperphosphorylation of the tau protein, which produces neurofibrillary tangles. One of the genetic markers of the disease is the gene that translates the presenilin-2 protein, which has mutations that favor the appearance of the disease and has no reported crystallographic structure. In view of this, protein modeling is performed using prediction and structural refinement tools followed by an energetic and stereochemical characterization for its validation. For the simulation, four reported mutations are chosen, which are Met239Ile, Met239Val, Ser130Leu, and Thr122Arg, all associated with various functional responses. From a theoretical analysis, a preliminary bioinformatic study is made to find the phosphorylation patterns in the protein and the hydropathic index according to the polarity and chemical environment. Molecular visualization was carried out with the Chimera 1.14 software, and the theoretical calculation with the hybrid quantum mechanics/molecular mechanics system from the semi-empirical method, with Spartan18 software and an AustinModel1 basis. These relationships allow for studying the system from a structural approach with the determination of small distance changes, potential surfaces, electrostatic maps, and angle changes, which favor the comparison between wild-type and mutant systems. With the results obtained, it is expected to complement experimental data reported in the literature from models that would allow us to understand the effects of the selected mutations.

Formulation and Characterization of a New Injectable Bone Substitute Composed PVA/Borax/CaCO3 and Demineralized Bone Matrix.

The occurrence of bone-related disorders and diseases has dramatically increased in recent years around the world. Demineralized bone matrix (DBM) has been widely used as a bone implant due to its osteoinduction and bioactivity. However, the use of DBM is limited because it is a particulate material, which makes it difficult to manipulate and implant with precision. In addition, these particles are susceptible to migration to other sites. To address this situation, DBM is commonly incorporated into a variety of carriers. An injectable scaffold has advantages over bone grafts or preformed scaffolds, such as the ability to flow and fill a bone defect. The aim of this research was to develop a DBM carrier with such viscoelastic properties in order to obtain an injectable bone substitute (IBS). The developed DBM carrier consisted of a PVA/glycerol network cross-linked with borax and reinforced with CaCO3 as a pH neutralizer, porosity generator, and source of Ca. The physicochemical properties were determined by an injectability test, FTIR, SEM, and TGA. Porosity, degradation, bioactivity, possible cytotoxic effect, and proliferation in osteoblasts were also determined. The results showed that the developed material has great potential to be used in bone tissue regeneration.

Protein Predictive Modeling and Simulation of Mutations of Presenilin-1 Familial Alzheimer's Disease on the Orthosteric Site.

Alzheimer's disease pathology is characterized by ß-amyloid plaques and neurofibrillary tangles. Amyloid precursor protein is processed by ß and γ secretase, resulting in the production of ß-amyloid peptides with a length ranging from 38 to 43 amino acids. Presenilin 1 (PS1) is the catalytic unit of γ-secretase, and more than 200 PS1 pathogenic mutations have been identified as causative for Alzheimer's disease. A complete monocrystal structure of PS1 has not been determined so far due to the presence of two flexible domains. We have developed a complete structural model of PS1 using a computational approach with structure prediction software. Missing fragments Met1-Glut72 and Ser290-Glu375 were modeled and validated by their energetic and stereochemical characteristics. Then, with the complete structure of PS1, we defined that these fragments do not have a direct effect in the structure of the pore. Next, we used our hypothetical model for the analysis of the functional effects of PS1 mutations Ala246GLu, Leu248Pro, Leu248Arg, Leu250Val, Tyr256Ser, Ala260Val, and Val261Phe, localized in the catalytic pore. For this, we used a quantum mechanics/molecular mechanics (QM/MM) hybrid method, evaluating modifications in the topology, potential surface density, and electrostatic potential map of mutated PS1 proteins. We found that each mutation exerts changes resulting in structural modifications of the active site and in the shape of the pore. We suggest this as a valid approach for functional studies of PS1 in view of the possible impact in substrate processing and for the design of targeted therapeutic strategies.

Lipid Secretion by Parasitic Cells of Coccidioides Contributes to Disseminated Disease.

Coccidioides is a soil-borne fungal pathogen and causative agent of a human respiratory disease (coccidioidomycosis) endemic to semi-desert regions of southwestern United States, Mexico, Central and South America. Aerosolized arthroconidia inhaled by the mammalian host first undergo conversion to large parasitic cells (spherules, 80-100 µm diameter) followed by endosporulation, a process by which the contents of spherules give rise to multiple endospores. The latter are released upon rupture of the maternal spherules and establish new foci of lung infection. A novel feature of spherule maturation prior to endosporulation is the secretion of a lipid-rich, membranous cell surface layer shed in vivo during growth of the parasitic cells and secretion into liquid culture medium during in vitro growth. Chemical analysis of the culture derived spherule outer wall (SOW) fraction showed that it is composed largely of phospholipids and is enriched with saturated fatty acids, including myristic, palmitic, elaidic, oleic, and stearic acid. NMR revealed the presence of monosaccharide- and disaccharide-linked acylglycerols and sphingolipids. The major sphingolipid components are sphingosine and ceramide. Primary neutrophils derived from healthy C57BL/6 and DBA/2 mice incubated with SOW lipids revealed a significant reduction in fungicidal activity against viable Coccidioides arthroconidia compared to incubation of neutrophils with arthroconidia alone. Host cell exposure to SOW lipids had no effect on neutrophil viability. Furthermore, C57BL/6 mice that were challenged subcutaneously with Coccidioides arthroconidia in the presence of the isolated SOW fraction developed disseminated disease, while control mice challenged with arthroconidia alone by the same route showed no dissemination of infection. We hypothesize that SOW lipids contribute to suppression of inflammatory response to Coccidioides infection. Studies are underway to characterize the immunosuppressive mechanism(s) of SOW lipids.

Caracterización de los compuestos de aroma del aceite de sacha inchi (Plukenetia volubilis L.) por HS-SPME-GC-MS-O / Characterization of aroma compounds in sacha inchi (Plukenetia volubilis L.) by HS-SPME-GC-MS-O / Caracterização de compostos de aroma em sacha inchi (Plukenetia volubilis L.) por HS-SPME-GC-MS-O

Resumen Se identificaron los componentes aromáticos activos provenientes de muestras comerciales del aceite de sacha inchi, obtenidos de cultivos de Santa Rosa de Osos (Antioquia, Colombia), mediante la técnica de microextracción en fase sólida acoplada a cromatografía de gases, espectrometría de masas y olfatometría (HS-SPME-GC-MS-O). En la optimización de la técnica de extracción se definieron las siguientes condiciones: fibra de SPME de divinilbenceno/ carboxen/polidimetilsiloxano (DVB/CAR/PDMS), temperatura de extracción de 50 °C y tiempo de exposición de 40 min. Bajo estas condiciones, se encontraron un total de 20 compuestos aromáticos activos, donde se destacan el E-2-octenal, E-heptanal, (E,E)-3,5-octadien-2-ona, ácido hexanóico y (E,E)-2,4-heptadienal como los componentes con mayor aporte a la formación del aroma del aceite de sacha inchi.

Abstract The active aromatic components from commercial samples of sacha inchi oil obtained from Santa Rosa de Osos (Antioquia, Colombia) crops were identified by solid phase microextraction technique coupled to gas chromatography, mass spectrometry, and olfactometry (HS -SPME-GC-MS-O). In the optimization of the extraction technique, the following conditions were defined: SPME fiber of divinylbenzene/carboxen/ polydimethylsiloxane (DVB/CAR/PDMS), extraction temperature 50 °C, and exposure time 40 min. Under these conditions, a total of 20 active aromatic compounds were found, which include E-2-octenal, E-heptanal, (E,E)-3,5-octadien-2-one, hexanoic acid and (E,E)-2,4-heptadienal as the components with the greatest contribution to the aroma formation of sacha inchi oil.

Resumo Os componentes de aroma ativos de amostras comerciais de óleo de sacha inchi obtidos de Santa Rosa de Osos (Antioquia, Colômbia) foram separados pela técnica de microextração em fase sólida acoplada a cromatografia gasosa, espectrometria de massas e olfatometria para a identificação (HS-SPME-GC-MS-O). Na otimização da técnica de extração, foram definidas as seguintes condições: fibra de SPME de divinilbenzeno/carboxeno/polidimetilsiloxano (DVB/CAR/PDMS), temperatura de extração 50 °C e tempo de exposição de 40 min. Nestas condições, foi encontrado um total de 20 compostos de aroma ativos no óleo, que incluem E-2-octenal, E-heptanal, (E,E)-3,5-octadien-2-ona, ácido hexanoico e (E,E)-2,4 heptadienal como os componentes com maior contribuição para a formação do aroma.

Genetic variants, structural, and functional changes of Myelin Protein Zero and Mannose-Binding Lectin 2 protein involved in immune response and its allelic transmission in families of patients with leprosy in Colombia.

Leprosy is a chronic infectious disease caused by Mycobacterium leprae. Genetic factors associated with immune response contribute to infection development and disease. M. leprae has the capacity to invade Schwann cells in the peripheral nervous system and cause neuropathy. However, while the responsible molecular mechanisms remain to be fully unveiled, they have begun being elucidated. We studied genetic variants Myelin Protein Zero (MPZ), a major structural component of the myelin sheath, and Mannose Binding Lectin 2 (MBL2), a protein involved in immune response, in 112 family groups of 114 leprosy patients using PCR-RFLP, aiming to calculate the association and allelic transmission of variants associated in first, second and third-degree relatives. Polymorphisms found in MPZ and MBL2 showed association with leprosy. Different probabilities for allelic transmission were found for first and second-degree relatives, a fact that is important to take into account when evaluating risk in contacts of leprosy patients. Structural analysis allows the study of putative amino acids and their possible effect on protein structure and function, as well as on the assembly of a protein homotetramer. Our results suggest that the identified MPZ and MBL2 gene mutations are associated with leprosy in a Colombian population, which correlates with MPZ and MBL2 protein function, and increase the risk of M. leprae infection in leprosy-patients' family members. Additionally, structural analyses were carried out specifically for MPZ protein using information available in databases, and analyzing the substitutions in wildtype and mutant protein. The results show significant structural changes, which may be associated to infection and pathogenicity.

Actividad antihelmíntica in vitro de extractos oleosos de Azadirachta indica y extractos acuosos de Nicotiana tabacum sobre nematodos gastrointestinales de cabras / In vitro Anthelmintic Activity of Oily Extracts of Azadirachta indica and Aqueous Extracts of Nicotiana tabacum on Gastrointestinal Nematodes in Goats / Atividade anti-helmíntica in vitro de extratos oleosos de Azadirachta indica e extratos aquosos de Nicotiana tabacum sobre nematoides gastrointestinais de cabras

Los nematodos gastrointestinales son los parásitos más frecuentes en los rumiantes del mundo. Estas parasitosis causan gastroenteritis parasitaria con un impacto negativo sobre la productividad. El control de nematodos se ha basado en el uso de antihelmínticos químicos -ante los cuales los nematodos han desarrollado cierto grado de resistencia- que se han valorado como sustancias residuales en productos de la cabra que pueden desencadenar efectos nocivos en el consumidor final. Por tanto, el estudio de componentes de plantas se ha propuesto como una alternativa sostenible para el control de la nematodosis caprina. En este trabajo se evaluó in vitro el potencial antihelmíntico de extractos acuosos de Nicotiana tabacum y de extractos oleosos de Azadirachta indica sobre nematodos gastrointestinales que afectan la cadena caprina. Se realizaron ensayos de actividad nematicida sobre muestras de materia fecal de caprinos con alta carga parasitaria (trichostrongilidos), mediante la realización de curvas dosis/respuesta. El porcentaje de inhibición en la eclosión de huevos para el extracto acuoso de N. tabacum y el extracto oleoso de A. indica fue de 99% y 80%, respectivamente. Los extractos presentaron efecto sobre larva 3 (estadio infectante), con un tiempo letal medio para extractos de N. tabacum de 8 ± 1 minutos, y para el extracto A. indica de 8 ± 1 minutos. Los resultados de la actividad nematicida a nivel in vitro muestran que los extractos de N. tabacum y A. indica pueden ser una alternativa promisoria para el control de nematodos en rumiantes.

Gastrointestinal nematodes are the most common parasites found in ruminants in the world. These parasites cause parasitic gastroenteritis and have a negative effect on productivity. Nematode control has been based on the use of anthelmintic chemicals-against which nematodes have developed a certain degree of resistance-which have been rated as residual substances in goat products that may cause adverse effects on the final consumer. As a result, the study of plant components has been proposed as a sustainable alternative to control nematodosis in goats. The anthelmintic potential of aqueous extracts of Nicotiana tabacum and oily extracts of Azadirachta indica on gastrointestinal nematodes affecting the goat chain was evaluated in vitro in this study. Nematicidal activity tests were performed on stool samples from goats with a high parasite load (trichostrongyles), by performing dose/ response curves. Percent inhibition in egg hatch for the aqueous extract of N. tabacum and the oily extract of A. indica was 99% and 80%, respectively. Extracts showed an effect on larva 3 (infective stage), with a mean lethal time of 8 ± 1 minutes for extracts of N. Tabacum, and of 8 ± 1 minutes for extract of A. indica. The in vitro results of the nematicidal activity show that N. tabacum and A. indica extracts can be a promising alternative for controlling nematodes in ruminants.

Os nematoides gastrointestinais são os parasitas mais frequentes nos ruminantes do mundo. Estas parasitoses causam gastroenterite parasitária e influem negativamente na produtividade. O controle de nematoides tem se baseado no uso de anti-helmínticos químicos - perante os quais os nematoides têm desenvolvido certo grau de resistência - que têm se valorado como substâncias residuais em produtos da cabra que podem desencadear efeitos nocivos no consumidor final. Por tanto, se propôs o estudo de componentes de plantas como uma alternativa sustentável para o controle de nematódeos em caprinos. Neste trabalho se avaliou in vitro o potencial anti-helmíntico de extratos aquosos de Nicotiana tabacum e de extratos oleosos de Azadirachta indica sobre nematoides gastrointestinais que afetam a cadeia caprina. Realizaram-se ensaios de atividade nematicida sobre amostras de fezes de caprinos com alta carga parasitária (tricostrongilídeos), mediante a realização de curvas dose/resposta. A porcentagem de inibição na eclosão de ovos para o extrato aquoso de N. tabacum e o extrato oleoso de A. indica fue de 99% e 80%, respectivamente. Os extratos apresentaram efeito sobre larva 3 (estado infectante), com um tempo letal médio para extratos de N. tabacum de 8 ± 1 minutos, e para o extrato A. indica de 8 ± 1 minutos. Os resultados in vitro da atividade nematicida mostram que os extratos de N. tabacum e A. indica podem ser uma alternativa promissória para o controle de nematoides em ruminantes.

Actividad insecticida de una emulsión aceite/agua del aceite esencial de Eucalyptus tereticornis / Insecticide activity of oil/water emulsion of Eucalyptus tereticornis essential oil

Introducción: el aceite esencial extraído de especies del género Eucalyptus, entre estas Eucalyptus tereticornis, ofrece un importante potencial para el control de hongos patógenos e insectos plaga y de importancia médica. Objetivo: evaluar la actividad anti-insecto (aguda, crónica y repelente) de una emulsión aceite/agua del aceite esencial de Eucalyptus tereticornis, en función de su composición química y fraccionamiento. Métodos: se prepararon emulsiones aceite/agua con aceite esencial previamente extraído, separado y caracterizado, y 2 de sus fracciones (A y B). Se evaluó su actividad anti-insecto mediante bioensayos con Drosophila melanogaster. Se determinó la fitotoxicidad foliar sobre plántulas de Phaseolus vulgaris. Resultados: el aceite completo mostró considerable actividad repelente e insecticida aguda que guarda relación con su composición química. La actividad insecticida aguda de las fracciones disminuyó de manera notable con respecto a la mostrada por el aceite completo, sin embargo, no se observaron efectos significativos sobre la actividad repelente. En ningún caso se presentó fitotoxicidad foliar sobre frijol. Conclusión: el aceite esencial de esta especie de eucalipto posee actividad anti-insecto considerable y podría formularse directamente como insecticida para el control de insectos de interés agrícola y médico.

Introduction: the essential oil extracted from Eucalyptus species, including Eucalyptus tereticornis, offers significant potential for control of fungal pathogens and insect pests and medical importance. Objectives: to evaluate the anti-insect activity (acute, chronic and repellent) of an oil/water emulsion of Eucalyptus tereticornis essential oil in terms of their chemical composition. Methods: oil/water emulsion was prepared with essential oil previously extracted, separated and characterized, and two of its fractions (A and B). Their anti-insect activity was evaluated through bioassays on Drosophila melanogaster. Foliar phytotoxicity was evaluated on seedlings of Phaseolus vulgaris. Results: the complete oil showed considerable acute repellent and insecticide activity related to its chemical composition. The acute insecticidal activity of the fractions decreased significantly if compare with that of the whole oil , but neither significant effects on the repellent activity nor phytotoxicity on the bean leaves were observed. Conclusions: the essential oil of this eucalyptus species has considerable anti-insect properties and could be formulated as an insecticide for the control of insects of agricultural and medical significance.