Application of human liver organoids as a patient-derived primary model for HBV infection and related hepatocellular carcinoma.

The molecular events that drive hepatitis B virus (HBV)-mediated transformation and tumorigenesis have remained largely unclear, due to the absence of a relevant primary model system. Here we propose the use of human liver organoids as a platform for modeling HBV infection and related tumorigenesis. We first describe a primary ex vivo HBV-infection model derived from healthy donor liver organoids after challenge with recombinant virus or HBV-infected patient serum. HBV-infected organoids produced covalently closed circular DNA (cccDNA) and HBV early antigen (HBeAg), expressed intracellular HBV RNA and proteins, and produced infectious HBV. This ex vivo HBV-infected primary differentiated hepatocyte organoid platform was amenable to drug screening for both anti-HBV activity and drug-induced toxicity. We also studied HBV replication in transgenically modified organoids; liver organoids exogenously overexpressing the HBV receptor sodium taurocholate co-transporting polypeptide (NTCP) after lentiviral transduction were not more susceptible to HBV, suggesting the necessity for additional host factors for efficient infection. We also generated transgenic organoids harboring integrated HBV, representing a long-term culture system also suitable for viral production and the study of HBV transcription. Finally, we generated HBV-infected patient-derived liver organoids from non-tumor cirrhotic tissue of explants from liver transplant patients. Interestingly, transcriptomic analysis of patient-derived liver organoids indicated the presence of an aberrant early cancer gene signature, which clustered with the hepatocellular carcinoma (HCC) cohort on The Cancer Genome Atlas Liver Hepatocellular Carcinoma dataset and away from healthy liver tissue, and may provide invaluable novel biomarkers for the development of HCC and surveillance in HBV-infected patients.

Lectin Profile Variation in Mesenchymal Stem Cells Derived from Different Sources.

Human mesenchymal stem cells (MSCs), a promising source of stem cells for regenerative medicine, have different morphological and functional characteristics. Carbohydrate moieties on the cell surface play an important role, including cell-cell interaction and cell recognition. The objective of this study was to determine possible differences in glycoconjugate distribution patterns of MSCs derived from various sources. MSCs were isolated from adipose tissue, bone marrow, Wharton's jelly, and cord blood. Then, they were stained with FITC-conjugated wheat germ agglutinin (WGA), peanut agglutinin (PNA), concanavalin A (ConA), Ulex europaeus (UEA), Dolichos biflorus (DBA), and Atto-488 conjugated Phytolacca americana (PWM) lectins. The intensity of the reactions was scored using ImageJ software. Flow cytometry was performed to detect the expression of the endothelial marker CD144. The obtained data were analyzed by ANOVA and LSD. Cord blood-derived MSCs showed the most significant staining intensities with all lectins. All MSCs were also moderately stained with PNA. Bone marrow-derived MSCs failed to react with UEA, DBA, and ConA. Wharton's jelly-derived MSCs could also not be stained with ConA. Cord blood-derived MSCs contained 2 subpopulations: osteoclast- and fibroblast-like cells. Both lectin staining intensity and distribution pattern were different in these 2 cell types; therefore, the central part of osteoclast-like cells stained more intensive with PNA and PWM, while that part in fibroblast-like cells stained more intensive with ConA. None of them expressed CD144. The glycoconjugate content of MSCs derived from various sources is different.

Activation of mitosis and angiogenesis in diabetes-impaired wound healing by processed human amniotic fluid.

BACKGROUND: Functional characterization of human amniotic fluid (AF) proteome, 845 proteins, has revealed that top three functions are cell proliferation, movement and differentiation, events fundamental to development, and tissue repair. Although these findings fortify the idea that AF components play roles in regeneration-like fetal wound healing, it is not known whether the components endure processing. Therefore, we processed AF and tested its effects on diabetes-impaired wound healing in an animal model. MATERIALS AND METHODS: Through a germfree procedure, mature and premature AF samples were collected, respectively, from the mothers of full-term and preterm infants. Excisional wounds were generated on the dorsum of diabetic rats. Wounds were treated on day 3 and harvested on day 7 postwounding. Proliferating cell nuclear antigen and alpha-smooth muscles actin, markers for mitosis and angiogenesis, respectively, were assessed by in situ immunodetection method. RESULTS: Significant increases in the rate of wound closure and proliferating cell nuclear antigen-expressing cells were observed in AF-treated wounds when compared with that of sham and control wounds. Likewise, the number of large vessels was significantly increased in the wounds treated with the AF. However, population of myofibroblasts was not affected by the treatment. The mature and premature AF were almost equally effective. CONCLUSIONS: Our data, for the first time, show that processed AF accelerates diabetes-impaired wound healing by activating mitosis and angiogenesis, indicating that bioactive molecules in AF may endure processing. We believe that processed forms of this naturally designed "Cocktail" of bioactive molecules may have multiple clinical applications.