Engineering the Vasculature of Stem-Cell-Derived Liver Organoids

The vasculature of stem-cell-derived liver organoids can be engineered using methods that recapitulate embryonic liver development. Hepatic organoids with a vascular network offer great application prospects for drug screening, disease modeling, and therapeutics. However, the application of stem cell-derived organoids is hindered by insufficient vascularization and maturation. Here, we review different theories about the origin of hepatic cells and the morphogenesis of hepatic vessels to provide potential approaches for organoid generation. We also review the main protocols for generating vascularized liver organoids from stem cells and consider their potential and limitations in the generation of vascularized liver organoids.

FIRST REPORT OF THE ANTICANCER EFFICIENCY OF AGARICUS BRASILIENSIS MUSHROOM ON HUMAN EMBRYONIC LIVER WRL68 AND HUMAN PANCREATIC ASPC-1 CANCER CELLS THERAPY

Mushrooms are food traditionally consumed in Asia, Europe and America. They are being studied for medicinal benefits. Extensive studies have shown that Agaricus brasiliensis mushroom used as a medical productto combat cancers. Our data reveal that the determined inhibitory concentration fifty(IC50) values were observed maximum dose responses (IC50) of WRL68 andAsPC-1 cancer cells reported of 172.6 µg/ml and 158.2 µg/ml respectively at 2.23 and 2.1 µg/ml ethanolic mushroom concentrations. The highly cytotoxic activity of the extract on growth inhibition AsPC-1 and WRL68were generally observed 97.9% and 95% at extract concentrations of 25 µg/ml and 50µg/ml respectively. Finally, Phytochemical profile of Agaricus brasiliensis mushroom extract found to beflavonoids, glycosides, saponins, phenols,alkaloids, tannins.The extracts of Agaricus brasiliensiswas tested throughGas Chromatography-Mass(GC-MS). There were five different compounds analyzed from the extracts of Agaricus brasiliensis. The compounds in the ethanolic extract of A. brasiliensis mushroom were comprised mostly of Acetic acid ethyl ester(38.39%),followed by(3-Methyl-oxiran-2-yl)-methanol (34.71%), Chlorbromuron (22.86%), n-Hexadecanoic acid (3.85%) and Heptane, 1-(1-butenyloxy) (0.19%). However, no studies were done using Agaricus brasiliensismushroom extract against WRL68 and AsPC-1 human cell lines. Therefore, the anticancer efficiency of Agaricus brasiliensis mushroom on human embryonic liver WRL68 andhuman pancreatic AsPC-1 cancer cellstherapy is being reported for the first time in the current study.

Zebra Fish in Toxicology Research: Streptavidin Conjugated Peroxidase Assay in the Development Phase of Zebrafish Embryos to Study Liver Toxicities

Background: Zebrafish have similar hepatic anatomy and cellular architecture just like mammals. Therefore, a number of investigators are using zebrafish to study liver pathologies. However, the evaluation model specific to liver toxicities in zebra fish was not clearly stated earlier. Aims and Objectives: The present study was designed to develop a model of embryonic liver toxicity using dexamethasone (DEXA, 0-20 μM) as a standard hepatotoxic agent. Methods: such toxicities are easily measured by streptavidin-conjugated peroxidase assay after 48- hour post-fertilization (hpf) of DEXA treatment. Results: In addition to morphological toxicities at different hpf, DEXA showed significant (*p< 0.05 &**p<0.01) reduction of peroxidase-chromogenic dye reaction in the assay as compared to DEXA untreated embryos at 10 & 20 μM concentration that concluded the hepatocellular toxicity of dexamethasone. Conclusion: Hopefully, the developed model for hepatotoxicity evaluation will be a promising model for the evaluation of new drugs or chemicals as an easy vertebrate model before the commencement of another animal model.

Prediction and Effect of Maternal Marginal Zinc Deficiency on Development, Redox Status, and Gene Expression Related to Oxidation and Apoptosis of Embryos

BackgroundMaternal severe zinc (Zn) deficiency induced oxidative damage and apoptosis in embryos, resulting in growth retardation. Therefore, it is crucial to assess maternal marginal Zn nutritional status for poultry breeders to prevent embryos subjected to severe Zn deficiency. MethodsIn Exp. 1, twelve egg embryos were sampled at incubation day 17 (E17), E20, E23, E26, E29, and E32 (day of hatch) respectively, with 6 replicates of 2 embryo each. The developmental changes of Zn mobilization and Zn transport gene mRNA expression were determined. In Exp. 2, 324 laying duck breeders were randomly allotted into 3 dietary Zn levels (0, 60, and 120 mg Zn/kg diet) with 6 replicates of 18 ducks per replicate. Plasma Zn concentration and erythrocytic Zn metalloenzyme activities in breeders as well as the development, redox status, and gene expression related to oxidation and apoptosis in embryos were measured. Blood samples were collected at the 2th, 4th and 6th weeks of the experiment. ResultsIn Exp. 1, the overall Zn mobilization rates were increased in yolk sac and embryonic liver in response to the increased incubation period, associated with the decreased ZIP10, 13, and 14 mRNA expressions in embryonic liver (P < 0.05). In Exp. 2, with the prolonged dietary Zn depletion, maternal Zn deficiency decreased plasma Zn concentration and erythrocytic alkaline phosphatase activity at the 6th week and inhibited erythrocytic 5'-nucleotidase (5’-NT) activity at 2th week (P < 0.05). On E29 of the maximal rate of Zn mobilization, maternal marginal Zn deficiency increased middle and late embryonic mortality and contents of superoxide anion radical, MDA and PPC, as well as decreasedMT content, CuZnSOD activity, and MT1 mRNA expression in embryonic livers (P < 0.05). Additionally, maternal marginal Zn deficiency increased BCL2-associated X protein and Caspase-9 mRNA expression and decreased B-cell lymphoma-2 mRNA expression in embryonic liver (P < 0.05). ConclusionErythrocytic 5’-NT activity was more rapid and reliable to assess marginal Zn-deficient status in duck breeders. Marginal Zn deficiency impaired hatchability and antioxidant defense system and then induced the oxidative damage and apoptosis in embryonic liver, contributing to the greater loss of embryonic death.

Embryonic liver developmental trajectory revealed by single-cell RNA sequencing in the Foxa2eGFP mouse

The liver and gallbladder are among the most important internal organs derived from the endoderm, yet the development of the liver and gallbladder in the early embryonic stages is not fully understood. Using a transgenic Foxa2eGFP reporter mouse line, we performed single-cell full-length mRNA sequencing on endodermal and hepatic cells isolated from ten embryonic stages, ranging from E7.5 to E15.5. We identified the embryonic liver developmental trajectory from gut endoderm to hepatoblasts and characterized the transcriptome of the hepatic lineage. More importantly, we identified liver primordium as the nascent hepatic progenitors with both gut and liver features and documented dynamic gene expression during the epithelial-hepatic transition (EHT) at the stage of liver specification during E9.5–11.5. We found six groups of genes switched on or off in the EHT process, including diverse transcripitional regulators that had not been previously known to be expressed during EHT. Moreover, we identified and revealed transcriptional profiling of gallbladder primordium at E9.5. The present data provides a high-resolution resource and critical insights for understanding the liver and gallbladder development.

ARID1A loss in adult hepatocytes activates β-catenin-mediated erythropoietin transcription

Erythropoietin (EPO) is a key regulator of erythropoiesis. The embryonic liver is the main site of erythropoietin synthesis, after which the kidney takes over. The adult liver retains the ability to express EPO, and we discovered here new players of this transcription, distinct from the classical hypoxia-inducible factor pathway. In mice, genetically invalidated in hepatocytes for the chromatin remodeler Arid1a, and for Apc, the major silencer of Wnt pathway, chromatin was more accessible and histone marks turned into active ones at the Epo downstream enhancer. Activating β-catenin signaling increased binding of Tcf4/β-catenin complex and upregulated its enhancer function. The loss of Arid1a together with β-catenin signaling, resulted in cell-autonomous EPO transcription in mouse and human hepatocytes. In mice with Apc-Arid1a gene invalidations in single hepatocytes, Epo de novo synthesis led to its secretion, to splenic erythropoiesis and to dramatic erythrocytosis. Thus, we identified new hepatic EPO regulation mechanism stimulating erythropoiesis.

Identifying Candidate Genes for Hypoxia Adaptation of Tibet Chicken Embryos by Selection Signature Analyses and RNA Sequencing

The Tibet chicken (Gallus gallus) lives on the Qinghai–Tibet Plateau and adapts to the hypoxic environment very well. The objectives of this study was to obtain candidate genes associated with hypoxia adaptation in the Tibet chicken embryos. In the present study, we used the fixation index (Fst) and cross population extended haplotype homozygosity (XPEHH) statistical methods to detect signatures of positive selection of the Tibet chicken, and analyzed the RNA sequencing data from the embryonic liver and heart with HISAT, StringTie and Ballgown for differentially expressed genes between the Tibet chicken and White leghorn (Gallus gallus, a kind of lowland chicken) embryos hatched under hypoxia condition. Genes which were screened out by both selection signature analysis and RNA sequencing analysis could be regarded as candidate genes for hypoxia adaptation of chicken embryos. We screened out 1772 genes by XPEHH and 601 genes by Fst, and obtained 384 and 353 differentially expressed genes in embryonic liver and heart, respectively. Among these genes, 89 genes were considered as candidate genes for hypoxia adaptation in chicken embryos. ARNT, AHR, GSTK1 and FGFR1 could be considered the most important candidate genes. Our findings provide references to elucidate the molecular mechanism of hypoxia adaptation in Tibet chicken embryos.