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Ursolic acid (UA) and oleanolic acid (OA) are hydrophobic triterpenoid isomers with demonstrated anti-mycobacterial (Mtb) and immune-regulatory properties, although their poor solubility limits clinical use. We report the development of solid lipid microparticles (SLMs) as delivery vehicles for UA and OA and evaluate their anti-Mtb efficacy in vitro and in vivo, as well as their acute toxicity. SLMs measured 0.7–0.89 µM in size, with complete in vitro release of OA and UA at 40 and 32 h, respectively. The minimum inhibitory concentration (MIC) of SLMs loaded with OA and UA was 40 µg/mL SLMs + 20 µg/mL OA + 20 µg/mL UA for drug-sensitive Mtb and 80 µg/mL SLMs + 40 µg/mL OA + 40 µg/mL UA for multidrug-resistant (MDR) Mtb. These SLMs showed an efficient reduction in Mtb burden in infected alveolar macrophages. In a murine model of late-stage progressive MDR-TB, aerosolized delivery of SLMs containing OA and UA via a metered-dose inhaler significantly reduced pulmonary bacterial loads and extended survival. In vivo, acute toxicity studies revealed no mortality or signs of toxicity. These findings demonstrate that SLMs are an optimal delivery system for terpenoids, providing potent in vitro and in vivo anti-TB activity with an excellent safety profile.

OBJECTIVES/GOALS: Single nucleotide polymorphisms in the transcription factor 7-like 2 (TCF7L2) gene are associated with Type 2 Diabetes (T2D) and nonalcoholic fatty liver disease (NAFLD). The metabolic function of TCF7L2 in the liver remains to be fully elucidated, but we hypothesized that TCF7L2 contributes to NAFLD through regulation of zonal metabolic pathways. METHODS/STUDY POPULATION: Using single nuclei RNA sequencing, we examined Tcf7l2 expression in periportal (PP) hepatocytes around the portal triad and pericentral (PC) hepatocytes surrounding the central vein of the liver. To visualize TCF7L2 transcriptional activity we used a TCF reporter mice, which expresses an H2B-eGFP fusion protein downstream of the conserved TCF DNA binding site. We disrupted Tcf7l2 transcriptional activity in mouse liver by breeding mice with a floxed Tcf7l2 exon 11, which encodes part of the DNA binding domain (DBD), to albumin-Cre mice (Hep-TCF7L2ΔDBD). Eight-week-old mice were fed a choline-deficient amino acid-defined high fat (CDAHFD) diet for 8 weeks. In liver samples harvested from these mice, we examined disruption to several key zonated metabolic pathways, and quantified the development of fibrosis. RESULTS/ANTICIPATED RESULTS: Single nuclei analysis revealed that Tcf7l2 mRNA was expressed primarily in parenchymal cells of the liver but was ubiquitous across the liver lobule. However, in immunofluorescence analysis of TCF reporter mice, the transcriptional activity of TCF7L2 was highly restricted to PC hepatocytes. Classic PC hepatocyte markers, including glutamine synthetase (Glul), were absent in Hep-TCF7L2ΔDBD mice. Following the CDAHFD, Hep-TCF7L2ΔDBD mice developed more severe fibrosis in histological analysis, and expressed elevated levels of genes involved in fibrogenesis, collagen synthesis and TGFβ signaling. Hep-TCF7L2ΔDBD mice also displayed hepatic cholesterol accumulation following the CDAHFD, which was likely the result of impaired pericentral bile acid synthesis. DISCUSSION/SIGNIFICANCE: Our results suggest that TCF7L2 plays an important role in the regulation of zonated metabolic pathways, which may contribute to the development of fibrosis. Ongoing analyses are exploring the mechanisms regulating the zonal transcriptional activity of TCF7L2.

OBJECTIVES/GOALS: Single nucleotide polymorphisms in the transcription factor 7-like 2 (TCF7L2) gene are associated with Type 2 Diabetes (T2D) and nonalcoholic fatty liver disease (NAFLD). The metabolic function of TCF7L2 in the liver remains to be fully elucidated, but we hypothesized that TCF7L2 contributes to NAFLD through regulation of zonal metabolic pathways. METHODS/STUDY POPULATION: Using single nuclei RNA sequencing, we examined Tcf7l2 expression in periportal (PP) hepatocytes around the portal triad and pericentral (PC) hepatocytes surrounding the central vein of the liver. To visualize TCF7L2 transcriptional activity we used a TCF reporter mice, which expresses an H2B-eGFP fusion protein downstream of the conserved TCF DNA binding site. We disrupted Tcf7l2 transcriptional activity in mouse liver by breeding mice with a floxed Tcf7l2 exon 11, which encodes part of the DNA binding domain (DBD), to albumin-Cre mice (Hep-TCF7L2ΔDBD). Eight-week-old mice were fed a choline-deficient amino acid-defined high fat (CDAHFD) diet for 8 weeks. In liver samples harvested from these mice, we examined disruption to several key zonated metabolic pathways, and quantified the development of fibrosis. RESULTS/ANTICIPATED RESULTS: Single nuclei analysis revealed that Tcf7l2 mRNA was expressed primarily in parenchymal cells of the liver but was ubiquitous across the liver lobule. However, in immunofluorescence analysis of TCF reporter mice, the transcriptional activity of TCF7L2 was highly restricted to PC hepatocytes. Classic PC hepatocyte markers, including glutamine synthetase (Glul), were absent in Hep-TCF7L2ΔDBD mice. Following the CDAHFD, Hep-TCF7L2ΔDBD mice developed more severe fibrosis in histological analysis, and expressed elevated levels of genes involved in fibrogenesis, collagen synthesis and TGFβsignaling. Hep-TCF7L2ΔDBD mice also displayed hepatic cholesterol accumulation following the CDAHFD, which was likely the result of impaired pericentral bile acid synthesis. DISCUSSION/SIGNIFICANCE: Our results suggest that TCF7L2 plays an important role in the regulation of zonated metabolic pathways, which may contribute to the development of fibrosis. Ongoing analyses are exploring the mechanisms regulating the zonal transcriptional activity of TCF7L2.

El presente estudio tiene el propósito de analizar la política industrial desarrollada por la monarquía hispánica para la producción de buques de guerra durante un largo siglo XVIII comprendido entre 1670 y 1834. Su hipótesis fundamental es que una de las claves para completar nuestro conocimiento sobre la naturaleza de los sistemas administrativos de la construcción naval es la relación del Estado y la iniciativa empresarial con la mano de obra especializada, encuadrada en el marco normativo de la Matrícula de Mar. Esta línea interpretativa intersecta con el debate historiográfico dedicado a la alternativa asiento/administración directa como clave para comprender la importancia de la construcción naval en el proceso de construcción del Estado moderno. La conexión se encuentra en que la tendencia legislativa de la corona consistió, primero, en militarizar la mano de obra especializada por medio de la inclusión en la matrícula y, posteriormente, en concentrar los contratos de construcción naval exclusivamente en individuos matriculados. Esto representa una diferencia estructural entre el contratista de principios del siglo XVIII, encargado de una multiplicidad de procesos productivos, y aquél de finales de la centuria, responsable únicamente de la movilización y administración de una mano de obra previamente militarizada. De manera paralela, este estudio busca integrar la dimensión asiática de la construcción naval española, no de manera tangencial, sino como un componente fundamental y prioritario del reformismo borbónico en las industrias estratégicas. Abstract The purpose of the present article is to outline the industrial policies developed by the Spanish Monarchy for the production of warships during the long Eighteenth century (1670-1834), in Spain, America and the Philippines. Its main hypothesis is that an important element to complete our understanding of the administrative systems of Spanish naval shipbuilding is the relationship between the State, the entrepreneurs dedicated to this aspect of the armaments industry, and the specialised workforce recruited through the registry of maritime professions known as Matrícula de Mar. This line of argumentation intersects with the historiographical debate dedicated to understand the State-building process of imperial Spain through the two main administrative methods employed in its armaments industries (private contracting and direct state administration). The Matrícula de Mar allowed the Spanish crown to create new militarised corporations of shipbuilding workers and, from the last third of the Eighteenth century it followed a consistent policy of concentrating the shipbuilding contracts on small-scale entrepreneurs belonging to these corporations. This had important implications for the meaning of the alternative between private contracting and direct state administration. The article also details the peculiarities of shipbuilding administration in Spanish America and Asia, through the expansion of the Matrícula de Mar.

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This contribution analyses the role played by the mechanical properties of a decollément shale layer in the evolution of the Mexican Fold and Thrust Belt (MFTB). The mobility of overpressured shales can accommodate large strains by grain-scale plastic mechanisms, and affect the folding and thrusting styles of the overburden. Research on shale deformation mechanisms is necessary to improve the knowledge of these processes and their influence on the structural style of fold and thrust belts. The ductile behavior of rocks involving grain-scale plasticity was documented in the Jurassic Santiago shale sequence using geological mapping, microstructural observations on thin-oriented sections, and scanning electron microscopy (SEM) imaging. Structural styles such as detachment folding, fault-bend folding, and shale-cored fold-thrusts were observed at the regional scale. At the outcrop scale, the shale developed strong foliation and pencil cleavage, with immersed packstone boudins. Observed structures include thrusting, soft and open folds, and buckle folding. In thin section, the ductile textures include a strong penetrative foliation with lenticular and wavy-parallel laminae composed of carbonates, ribbons of reoriented clays and organic matter (clay+OM), s-c structures, porphyroblasts microtextures, development of oblique cleavage concerning folded foliation (crenulation cleavage), and carbonates dissolution. The Santiago shale shows also evidence of brittle deformation including calcite-filled fractures and cataclastic gouges. X-ray diffraction (XRD) analysis of the clay size fraction suggests that the authigenic calcareous shale was deformed in conditions of the deep diagenetic zone (between 100 and 200°C) and fluid overpressure (>70 MPa). The results help to improve the understanding of ductile microstructure and its role in shale deformation cretaceous cover, promoting the formation of localized fault propagation folds in the overburden. This study aims to open new perspectives in the kinematics and rheology interpretations for this sector of the MFTB, highlighting the role of the décollement layers during the progression of the orogen.