Explore the vast range of titles available.

A protocol for generating biliary epithelial cells from human pluripotent stem cells facilitates disease modeling and drug screening. The study of biliary disease has been constrained by a lack of primary human cholangiocytes. Here we present an efficient, serum-free protocol for directed differentiation of human induced pluripotent stem cells into cholangiocyte-like cells (CLCs). CLCs show functional characteristics of cholangiocytes, including bile acids transfer, alkaline phosphatase activity, γ-glutamyl-transpeptidase activity and physiological responses to secretin, somatostatin and vascular endothelial growth factor. We use CLCs to model in vitro key features of Alagille syndrome, polycystic liver disease and cystic fibrosis (CF)-associated cholangiopathy. Furthermore, we use CLCs generated from healthy individuals and patients with polycystic liver disease to reproduce the effects of the drugs verapamil and octreotide, and we show that the experimental CF drug VX809 rescues the disease phenotype of CF cholangiopathy in vitro . Our differentiation protocol will facilitate the study of biological mechanisms controlling biliary development, as well as disease modeling and drug screening.

Human iPS cells have been generated using a diverse range of tissues from a variety of donors using different reprogramming vectors. However, these cell lines are heterogeneous, which presents a limitation for their use in disease modeling and personalized medicine. To explore the basis of this heterogeneity we generated 25 iPS cell lines under normalised conditions from the same set of somatic tissues across a number of donors. RNA-seq data sets from each cell line were compared to identify the majority contributors to transcriptional heterogeneity. We found that genetic differences between individual donors were the major cause of transcriptional variation between lines. In contrast, residual signatures from the somatic cell of origin, so called epigenetic memory, contributed relatively little to transcriptional variation. Thus, underlying genetic background variation is responsible for most heterogeneity between human iPS cell lines. We conclude that epigenetic effects in hIPSCs are minimal, and that hIPSCs are a stable, robust and powerful platform for large-scale studies of the function of genetic differences between individuals. Our data also suggest that future studies using hIPSCs as a model system should focus most effort on collection of large numbers of donors, rather than generating large numbers of lines from the same donor.

A fully iterative ‘two-way’ fluid-structure interaction (FSI) tool of a commercially available composite windsurfer fin was developed, which was then used to investigate the normally hidden fin behaviour for a range of typical sailing conditions. The ‘two-way’ FSI analysis gave significantly better insights into the fin behaviour than the simpler ‘one-way’ non-iterative analysis. The tool also indicated that hydro-elastic tailoring, via simple reinforcement ply rotations, can produce large changes in tip twist. This gives an opportunity for both improved passive control and higher speeds, without deviating from a hydrodynamically optimal plan form. Inexpensive cantilever tests appear to be sufficient to make qualitative comparisons between the sailing responses of fins with different layups.

This protocol describes how to recapitulate biliary development by differentiation of hPSCs into endoderm, foregut progenitor cells, hepatoblasts, cholangiocyte progenitors and mature 3D cholangiocyte-like cell organoids. The difficulty in isolating and propagating functional primary cholangiocytes is a major limitation in the study of biliary disorders and the testing of novel therapeutic agents. To overcome this problem, we have developed a platform for the differentiation of human pluripotent stem cells (hPSCs) into functional cholangiocyte-like cells (CLCs). We have previously reported that our 26-d protocol closely recapitulates key stages of biliary development, starting with the differentiation of hPSCs into endoderm and subsequently into foregut progenitor (FP) cells, followed by the generation of hepatoblasts (HBs), cholangiocyte progenitors (CPs) expressing early biliary markers and mature CLCs displaying cholangiocyte functionality. Compared with alternative protocols for biliary differentiation of hPSCs, our system does not require coculture with other cell types and relies on chemically defined conditions up to and including the generation of CPs. A complex extracellular matrix is used for the maturation of CLCs; therefore, experience in hPSC culture and 3D organoid systems may be necessary for optimal results. Finally, the capacity of our platform for generating large amounts of disease-specific functional cholangiocytes will have broad applications for cholangiopathies, in disease modeling and for screening of therapeutic compounds.

Repair of defects in the common bile duct is hampered by a lack of healthy donor tissue. Developing human extrahepatic cholangiocyte organoids and testing them in mouse models may provide a way to overcome this limitation. The treatment of common bile duct (CBD) disorders, such as biliary atresia or ischemic strictures, is restricted by the lack of biliary tissue from healthy donors suitable for surgical reconstruction. Here we report a new method for the isolation and propagation of human cholangiocytes from the extrahepatic biliary tree in the form of extrahepatic cholangiocyte organoids (ECOs) for regenerative medicine applications. The resulting ECOs closely resemble primary cholangiocytes in terms of their transcriptomic profile and functional properties. We explore the regenerative potential of these organoids in vivo and demonstrate that ECOs self-organize into bile duct–like tubes expressing biliary markers following transplantation under the kidney capsule of immunocompromised mice. In addition, when seeded on biodegradable scaffolds, ECOs form tissue-like structures retaining biliary characteristics. The resulting bioengineered tissue can reconstruct the gallbladder wall and repair the biliary epithelium following transplantation into a mouse model of injury. Furthermore, bioengineered artificial ducts can replace the native CBD, with no evidence of cholestasis or occlusion of the lumen. In conclusion, ECOs can successfully reconstruct the biliary tree, providing proof of principle for organ regeneration using human primary cholangiocytes expanded in vitro .

Production of large quantities of hepatocytes remains a major challenge for a number of clinical applications in the biomedical field. Directed differentiation of human pluripotent stem cells (hPSC) into hepatocyte-like cells (HLCs) provides an advantageous solution and a number of protocols have been developed for this purpose. However, these methods usually follow different steps of liver development in vitro which is time consuming and requires complex culture conditions. In addition, HLCs lack the full repertoire of functionalities characterising primary hepatocytes. Here, we explore the interest of forward programming to generate hepatocytes from hPSCs and to bypass these limitations. This approach relies on the overexpression of 3 hepatocyte nuclear factors (HNF1A, HNF6 and FOXA3) in combination with different nuclear receptors expressed in the adult liver using the OPTi-OX platform. Forward programming allows for the rapid production of hepatocytes (FoP-Heps) with functional characteristics using a simplified process. We also uncovered that the overexpression of nuclear receptors such as RORc can enhance specific functionalities of FoP-Heps thereby validating its role in lipid/glucose metabolism. Together, our results show that forward programming could offer a versatile alternative to direct differentiation for generating hepatocytes in vitro. Competing Interest Statement Fabian Bachinger is a PhD student sponsored by bit.bio. Ludovic Vallier is a founder and shareholder of DefiniGEN, Aculive Therapeutics and Billitech.

Despite promising preclinical results, average response rates to anti-VEGF therapies, such as bevacizumab, are reduced for most cancers, while incurring in remarkable costs and side effects. Currently, there are no biomarkers available to select patients that can benefit from this therapy. Depending on the individual tumor, anti-VEGF therapies can either block or promote metastasis. In this context, an assay able to predict individual responses prior to treatment, including the impact on metastasis would prove of great value to guide treatment options. Here we show that zebrafish xenografts are able to reveal different responses to bevacizumab in just 4 days, evaluating not only individual tumor responses but also the impact on angiogenesis and micrometastasis. Importantly, we perform proof-of-concept experiments where clinical responses in patients were compared with their matching zebrafish Patient-Derived Xenografts - zAvatars, opening the possibility of using the zebrafish model to screen bevacizumab therapy in a personalized manner. Rebelo de Almeida et al. describe the application of zebrafish tumor xenografts (zAvatars) to analyse the efficacy of bevacizumab (FDA approved drug) to treat cancer patients with highly variable outcomes. The authors suggest that these zAvatars could be used as a predictive model to determine whether or not bevacizumab treatment would be efficient for individual patients.

The Tocantins-Araguaia Basin is one of the largest river systems in South America, located entirely within Brazilian territory. In the last decades, capital-concentrating activities such as agribusiness, mining, and hydropower promoted extensive changes in land cover, hydrology, and environmental conditions. These changes are jeopardizing the basin’s biodiversity and ecosystem services. Threats are escalating as poor environmental policies continue to be formulated, such as environmentally unsustainable hydropower plants, large-scale agriculture for commodity production, and aquaculture with non-native fish. If the current model persists, it will deepen the environmental crisis in the basin, compromising broad conservation goals and social development in the long term. Better policies will require thought and planning to minimize growing threats and ensure the basin’s sustainability for future generations.

Chemotherapy remains the mainstay in most high-risk breast cancer (BC) settings, with several equivalent options of treatment. However, the efficacy of each treatment varies between patients and there is currently no test to determine which option will be the most effective for each individual patient. Here, we developed a fast in-vivo test for BC therapy screening: the zebrafish patient-derived-xenograft model (zAvatars), where in-vivo results can be obtained in just 10 days. To determine the predictive value of the BC zAvatars we performed a preclinical study, where zAvatars were treated with the same therapy as the donor-patient and their response to therapy was compared. Our data show a 100% concordance (18 out of 18) between the zAvatar-test and the corresponding patient’s clinical response to treatment. Altogether, our results suggest that the zAvatar model constitutes a promising in-vivo assay to optimize cancer treatments in a truly personalized manner.

Antibodies are critical components of the adaptive immune system, whose therapeutic applications have been growing exponentially in the last years. Discovery and development of therapeutic antibodies encompasses in vivo immunization, synthetic libraries, and surface display methodologies. To overcome some of their limitations, several platforms in higher eukaryotic cells have been developed. Moreover, these platforms aim to replicate in the bench both primary and secondary antibody diversification mechanisms that occur in vivo. Here, we describe the latest strategies that have been used to mirror both naïve and affinity-maturated antibody repertoire.Key points• Therapeutic antibodies are one of the most promising classes of drugs to fight diseases.• Antibodies discovered through hybridoma or display technologies require further engineering.• Innovative antibody discovery platforms in higher eukaryotic cells have been developed.