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In the field of precision medicine, therapy is optimized individually for each patient, enhancing efficacy while reducing side effects. This involves the identification of promising drug candidates through high-throughput screening on human derived cells in culture. However, screening of drugs which have poor solubility or permeability remains challenging, especially when targeting intracellular components. Therefore, encapsulation of drugs into advanced delivery systems such as nanostructured lipid carries (NLC) becomes necessary. Here we show that the cellular uptake of NLC with different matrix compositions can be assessed in a high-throughput screening system based on acoustic droplet ejection (ADE) technology (Echo liquid handler). Our findings indicate that surface tension and viscosity of the NLC dispersions need to be tailored to enable a reliable ADE transfer. The automated NLC uptake studies indicated that the composition of the matrix, more specifically the amount of oleic acid, significantly influenced cellular uptake. The data obtained were corroborated by imaging based and spectral flow cytometry cellular uptake studies. These findings thus not only provide the basis for a screening tool to rapidly identify the efficacy of NLC uptake but also enable a next step toward precision high-throughput drug screening under consideration of an optimized drug delivery system., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

A variety of strategies for producing high-quality nanoparticles have been reported in recent years. Batch-based bottom-up and top-down technologies are generally the most efficient methods, but present a number of challenges, particularly in terms of variability, safety, sustainability and large-scale production. In this study, a scalable, semi-continuous production line was built by connecting individual processing units, including a high shear mixing device, the Microfluidizer® technology and a cooling system. Each unit was equipped with an adequate temperature control to allow solvent-free production of solid lipid nanoparticles (consisting of Precirol® ATO 5 or Gelucire® 43/01) and nanostructured lipid carriers (additionally comprising Labrafac™ lipophile WL 1349). Subsequently, critical formulation parameters and critical process parameters (CPPs) of the individual processing units and their effects on particle size (i.e., critical quality attribute (CQA)) were investigated to identify appropriate input parameters for the subsequent Design of Experiment (DoE) studies conducted after linking the process units to a semi-continuous production line. For particle size monitoring, spatially resolved dynamic light scattering (SR-DLS) measurements were conducted and compared to standard DLS measurements to evaluate the applicability of SR-DLS as an inline monitoring tool. It was found that matrix composition, emulsifier concentration, pressure and number of cycles when processing through Microfluidizer® processor were the most influencing parameters. By optimizing these parameters, five-times higher throughputs could be achieved by the semi-continuous manufacturing line. In addition, the particle size measurements with SR-DLS confirmed the feasibility of implementing this technology for real-time particle size monitoring as an important safety factor in quality control., Competing Interests: Declaration of competing interest The authors confirm that they do not have any relevant conflicts of interest to declare., (Copyright © 2024. Published by Elsevier B.V.)

The efficacy of nanostructured lipid carriers (NLC) for drug delivery strongly depends on their stability and cell uptake. Both properties are governed by their compositions and internal structure. To test the effect of the lipid composition of NLC on cell uptake and stability, three kinds of liquid lipids with different degrees of unsaturation are employed. After ensuring homogeneous size distributions, the thermodynamic characteristics, stability, and mixing properties of NLC are characterized. Then the rates and predominant pathways of cell uptake are determined. Although the same surfactant is used in all cases, different uptake rates are observed. This finding contradicts the view that the surface properties of NLC are dominated by the surfactant. Instead, the uptake rates are explained by the structure of the nanocarrier. Depending on the mixing properties, some liquid lipids remain inside the nanocarrier, while other liquid lipids are present on the surface. Nanocarriers with liquid lipids on the surface are taken up more readily by the cells. This shows that the engineering of efficient lipid nanocarriers requires a delicate balance of interactions between all components of the nanocarrier on the molecular level.

In vitro intestinal models are used to study biological processes, drug and food absorption, or cytotoxicity, minimizing the use of animals in the laboratory. They usually consist of enterocytes and mucus-producing cells cultured for 3 weeks, e.g., on Transwells, to obtain a fully differentiated cell layer simulating the human epithelium. Other important components are the extracellular matrix (ECM) and strong vascularization. The former serves as structural support for cells and promotes cellular processes such as differentiation, migration, and growth. The latter includes endothelial cells, which coordinate vascularization and immune cell migration and facilitate the transport of ingested substances or drugs to the liver. In most cases, animal-derived hydrogels such as Matrigel or collagen are used as ECM in in vitro intestinal models, and endothelial cells are only partially considered, if at all. However, it is well-known that animal-derived products can lead to altered cell behavior and incorrect results. To circumvent these limitations, synthetic and modifiable hydrogels (Peptigel and Vitrogel) were studied here to mimic xenofree ECM, and the data were compared with Matrigel. Careful rheological characterization was performed, and the effect on cell proliferation was investigated. The results showed that Vitrogel exhibited shear-thinning behavior with an internal structure recovery of 78.9 ± 11.2%, providing the best properties among the gels investigated. Therefore, a coculture of Caco-2 and HT29-MTX cells (ratio 7:3) was grown on Vitrogel, while simultaneously endothelial cells were cultured on the basolateral side by inverse cultivation. The model was characterized in terms of cell proliferation, differentiation, and drug permeability. It was found that the cells cultured on Vitrogel induced a 1.7-fold increase in cell proliferation and facilitated the formation of microvilli and tight junctions after 2 weeks of cultivation. At the same time, the coculture showed full differentiation indicated by high alkaline phosphatase release of Caco-2 cells (95.0 ± 15.9%) and a mucus layer produced by HT29-MTX cells. Drug tests led to ex vivo comparable permeability coefficients ( P app ) (i.e., P app ; antipyrine = (33.64 ± 5.13) × 10 -6 cm/s, P app ; atenolol = (0.59 ± 0.16) × 10 -6 cm/s). These results indicate that the newly developed intestinal model can be used for rapid and efficient assessment of drug permeability, excluding unexpected results due to animal-derived materials.

Background and Purpose: Leptin, an important hormone for body weight regulation, may be involved in the pathogenesis of cardiovascular manifestations of obesity. We tested whether leptin may be an independent risk marker for stroke in a case-referent study., Methods: Definitive acute stroke events, defined by MONICA criteria, were identified from October 1, 1995 to April 30, 1999. Referents without known cardiovascular disease were randomly selected from a population census. Patient characteristics were taken from hospital files and leptin was analyzed in stored samples. Logistic regression analysis was used to determine possible differences in leptin levels between groups., Results: One hundred and thirty-seven cases with ischemic stroke and 69 cases with hemorrhagic stroke were identified. In comparison with referents, male patients with stroke had significantly higher leptin levels. Both male and female stroke patients had increased blood pressure compared with the referents. In multivariate analyses, high leptin levels were associated with both ischemic (OR = 4.89; 95% CI: 1.89-12.62) and hemorrhagic (OR = 3.86; 95% CI: 1.13-13.16) stroke in men, and with ischemic stroke in women (OR = 4.10; 95% CI: 1.45-11.62). The combination of high leptin levels and increased blood pressure (systolic or diastolic) was associated with a strong positive interaction in males with hemorrhagic stroke., Conclusion: Leptin may be an important link for the development of cerebrovascular disease in the insulin resistance syndrome in men., (Copyright 2003 S. Karger AG, Basel)