In Vitro Liver Toxicity Testing of Chemicals: A Pragmatic Approach.

The liver is among the most frequently targeted organs by noxious chemicals of diverse nature. Liver toxicity testing using laboratory animals not only raises serious ethical questions, but is also rather poorly predictive of human safety towards chemicals. Increasing attention is, therefore, being paid to the development of non-animal and human-based testing schemes, which rely to a great extent on in vitro methodology. The present paper proposes a rationalized tiered in vitro testing strategy to detect liver toxicity triggered by chemicals, in which the first tier is focused on assessing general cytotoxicity, while the second tier is aimed at identifying liver-specific toxicity as such. A state-of-the-art overview is provided of the most commonly used in vitro assays that can be used in both tiers. Advantages and disadvantages of each assay as well as overall practical considerations are discussed.

Advances in microfluidic in vitro systems for neurological disease modeling.

Neurological disorders are the leading cause of disability and the second largest cause of death worldwide. Despite significant research efforts, neurology remains one of the most failure-prone areas of drug development. The complexity of the human brain, boundaries to examining the brain directly in vivo, and the significant evolutionary gap between animal models and humans, all serve to hamper translational success. Recent advances in microfluidic in vitro models have provided new opportunities to study human cells with enhanced physiological relevance. The ability to precisely micro-engineer cell-scale architecture, tailoring form and function, has allowed for detailed dissection of cell biology using microphysiological systems (MPS) of varying complexities from single cell systems to "Organ-on-chip" models. Simplified neuronal networks have allowed for unique insights into neuronal transport and neurogenesis, while more complex 3D heterotypic cellular models such as neurovascular unit mimetics and "Organ-on-chip" systems have enabled new understanding of metabolic coupling and blood-brain barrier transport. These systems are now being developed beyond MPS toward disease specific micro-pathophysiological systems, moving from "Organ-on-chip" to "Disease-on-chip." This review gives an outline of current state of the art in microfluidic technologies for neurological disease research, discussing the challenges and limitations while highlighting the benefits and potential of integrating technologies. We provide examples of where such toolsets have enabled novel insights and how these technologies may empower future investigation into neurological diseases.

In-vitro blood-brain barrier modeling: A review of modern and fast-advancing technologies.

The development of realistic in vitro blood-brain barrier (BBB) models that recapitulate the physiological parameters and molecular aspect of the neurovascular unit (NVU) is of fundamental importance not only in CNS drug discovery but also in translational research. Successful modeling of the NVU would provide an invaluable tool to aid in dissecting out the pathological factors, mechanism of action (and corresponding targets) prodromal to the onset of CNS disorders. The field of BBB in vitro modeling has seen many radical changes in the last few years with the introduction on novel technologies and methods to improve over existing models and develop new ones. Therefore, the goal of this review is to provide the readers with updated technical and operational details concerning current BBB platforms with special focus on stem cell technology used to establish a functional BBB model in vitro. Furthermore, we provide a detailed update on rapidly advancing 3D printing technologies used for engineering BBB models which use is now fast expanding among researchers.

In vitro and in silico liver models: Current trends, challenges and opportunities.

Most common drug development failures originate from either bioavailability problems, or unexpected toxic effects. The culprit is often the liver, which is responsible for biotransformation of a majority of xenobiotics. Liver may be modeled using "liver on a chip" devices, which may include established cell lines, primary human cells, and stem cell-derived hepatocyte-like cells. The choice of biological material along with its processing and maintenance greatly influence both the device performance and the resultant toxicity predictions. Impediments to the development of "liver on a chip" technology include the problems with standardization of cells, limitations imposed by culturing and the necessity to develop more complicated fluidic contours. Fortunately, recent breakthroughs in the development of cell-based reporters, including ones with fluorescent label, permits monitoring of the behavior of the cells embed into the "liver on a chip" devices. Finally, a set of computational approaches has been developed to model both particular toxic response and the homeostasis of human liver as a whole; these approaches pave a way to enhance the in silico stage of assessment for a potential toxicity.

Advances in predictive in vitro models of drug-induced nephrotoxicity.

In vitro screens for nephrotoxicity are currently poorly predictive of toxicity in humans. Although the functional proteins that are expressed by nephron tubules and mediate drug susceptibility are well known, current in vitro cellular models poorly replicate both the morphology and the function of kidney tubules and therefore fail to demonstrate injury responses to drugs that would be nephrotoxic in vivo. Advances in protocols to enable the directed differentiation of pluripotent stem cells into multiple renal cell types and the development of microfluidic and 3D culture systems have opened a range of potential new platforms for evaluating drug nephrotoxicity. Many of the new in vitro culture systems have been characterized by the expression and function of transporters, enzymes, and other functional proteins that are expressed by the kidney and have been implicated in drug-induced renal injury. In vitro platforms that express these proteins and exhibit molecular biomarkers that have been used as readouts of injury demonstrate improved functional maturity compared with static 2D cultures and represent an opportunity to model injury to renal cell types that have hitherto received little attention. As nephrotoxicity screening platforms become more physiologically relevant, they will facilitate the development of safer drugs and improved clinical management of nephrotoxicants.

Una nueva generación de triacetato de celulosa adecuado para hemodiafiltración on-line / A new generation of cellulose triacetate suitable for online haemodiafiltration

Antecedentes: La hemodiafiltración on-line (HDF-OL) es actualmente la técnica de hemodiálisis (HD) más efectiva y aumenta la supervivencia. Hasta el momento presente las membranas de alta permeabilidad con baja pérdida de albúmina como las de polisulfona, poliamida y poliacrilonitrilo son las más utilizadas. Las membranas de triacetato de celulosa (TAC), limitadas inicialmente para su uso en HDF-OL, han evolucionado. El objetivo del estudio fue determinar si las membranas de nueva generación de TAC asimétrico (TACA) son más adecuadas para realizar alto transporte convectivo. Pacientes y métodos: Se estudiaron 16 pacientes, 10 hombres y 6 mujeres, en programa de HDF-OL. A cada paciente se le realizaron 4 sesiones diferentes, con HD o HDF-OL, o con filtros de TAC o TACA de 1,9 m2, aleatorizando el orden. En cada sesión se determinaron concentración de urea, creatinina, β2-microglobulina, mioglobina, prolactina, α1-microglobulina, α1-glicoproteína ácida y albúmina en suero al inicio y al final de cada sesión, para calcular el porcentaje de reducción. Así mismo, se cuantificó la pérdida de solutos y albúmina en el líquido de diálisis. Resultados: Con las membranas de TACA se consiguió un volumen de sustitución en HDF-OL significativamente superior a las membranas de TAC clásicas (32,1±3,1 vs. 19,7±4,5L; p<0,001). En términos de depuración, la eliminación de moléculas pequeñas fue similar con ambas membranas, pero, en moléculas grandes, con HDF-OL la depuración fue mayor con TACA. En HDF-OL, el porcentaje de reducción de la β2-microglobulina se incrementó un 29%, un 27,7% la mioglobina, un 19,5% la prolactina, un 49% la α1-microglobulina, y se duplicó la α1-glicoproteína ácida (p<0,01 en todas las situaciones). La pérdida de albúmina fue inferior a 2 g en todas las situaciones de estudio. Conclusión: Las membranas de TAC de nueva generación han demostrado ser eficaces para alcanzar los objetivos de HDF-OL, sin que haya una mayor pérdida de albúmina (AU)

Background: Online haemodiafiltration (OL-HDF) is currently the most effective dialysis technique that also improves survival. To date, high permeability membranes with low albumin loss, such as polysulfone, polyamide and polyacrylonitrile membranes have been the most widely used. However, the initially restricted use of cellulose triacetate (CTA) membranes in OL-HDF has expanded. The aim of the study was to ascertain whether the latest generation asymmetric CTA membranes are more effective in obtaining high convective transport. Patients and methods: A total of 16 patients (10 males and 6 females) undergoing OL-HDF were studied. Each patient underwent 4 different sessions, with haemodialysis or OL-HDF, and/or with CTA or asymmetric CTA 1.9 m2 membranes. Each session was assigned in a randomised order. Serum levels of urea, creatinine, β2-microglobulin, myoglobin, prolactin, α1-microglobulin, α1-acid glycoprotein and albumin where measured at the beginning and end of each session to obtain the reduction rate. The loss of solutes and albumin was quantified from the dialysate. Results: A significantly greater replacement volume in OL-HDF (32.1±3.1 vs. 19.7±4.5 l, P<.001) was obtained by using asymmetrical CTA membranes compared to conventional CTA membranes. Regarding uraemic toxin removal, both membranes obtained similar results for small molecules, whereas asymmetric CTA membranes achieved better results for large molecules, increasing the reduction ratio by 29% for β2-microglobulin, 27.7% for myoglobin, 19.5% for prolactin, 49% for α1-microglobulin and double for α1-acid glycoprotein (P<0.001 in all situations). The loss of albumin was less than 2g for all treatment sessions. Conclusion: Latest-generation asymmetric CTA have proven to be effective in attaining OL-HDF objectives without increased albumin loss (AU)

The Next Frontier: Quantitative Biochemistry in Living Cells.

Researchers striving to convert biology into an exact science foremost rely on structural biology and biochemical reconstitution approaches to obtain quantitative data. However, cell biological research is moving at an ever-accelerating speed into areas where these approaches lose much of their edge. Intrinsically unstructured proteins and biochemical interaction networks composed of interchangeable, multivalent, and unspecific interactions pose unique challenges to quantitative biology, as do processes that occur in discrete cellular microenvironments. Here we argue that a conceptual change in our way of conducting biochemical experiments is required to take on these new challenges. We propose that reconstitution of cellular processes in vitro should be much more focused on mimicking the cellular environment in vivo, an approach that requires detailed knowledge of the material properties of cellular compartments, essentially requiring a material science of the cell. In a similar vein, we suggest that quantitative biochemical experiments in vitro should be accompanied by corresponding experiments in vivo, as many newly relevant cellular processes are highly context-dependent. In essence, this constitutes a call for chemical biologists to convert their discipline from a proof-of-principle science to an area that could rightfully be called quantitative biochemistry in living cells. In this essay, we discuss novel techniques and experimental strategies with regard to their potential to fulfill such ambitious aims.

In Vitro Models to Study Human Lung Development, Disease and Homeostasis.

The main function of the lung is to support gas exchange, and defects in lung development or diseases affecting the structure and function of the lung can have fatal consequences. Most of what we currently understand about human lung development and disease has come from animal models. However, animal models are not always fully able to recapitulate human lung development and disease, highlighting an area where in vitro models of the human lung can compliment animal models to further understanding of critical developmental and pathological mechanisms. This review will discuss current advances in generating in vitro human lung models using primary human tissue, cell lines, and human pluripotent stem cell derived lung tissue, and will discuss crucial next steps in the field.

In vitro bioavailability and cellular bioactivity studies of flavonoids and flavonoid-rich plant extracts: questions, considerations and future perspectives.

In vitro techniques are essential in elucidating biochemical mechanisms and for screening a wide range of possible bioactive candidates. The number of papers published reporting in vitro bioavailability and bioactivity of flavonoids and flavonoid-rich plant extracts is numerous and still increasing. However, even with the present knowledge on the bioavailability and metabolism of flavonoids after oral ingestion, certain inaccuracies still persist in the literature, such as the use of plant extracts to study bioactivity towards vascular cells. There is therefore a need to revisit, even question, these approaches in terms of their biological relevance. In this review, the bioavailability of flavonoid glycosides, the use of cell models for intestinal absorption and the use of flavonoid aglycones and flavonoid-rich plant extracts in in vitro bioactivity studies will be discussed. Here, we focus on the limitations of current in vitro systems and revisit the validity of some in vitro approaches, and not on the detailed mechanism of flavonoid absorption and bioactivity. Based on the results in the review, there is an apparent need for stricter guidelines on publishing data on in vitro data relating to the bioavailability and bioactivity of flavonoids and flavonoid-rich plant extracts.