Hepatitis B virus (HBV) is a major cause of hepatocellular carcinoma (HCC). However, very little is known about the replication of HBV in HCC tissues. We analyzed viral and cellular parameters in HCC (T) and nontumor liver (NT) samples from 99 hepatitis B surface antigen (HBsAg)‐positive, virologically suppressed patients treated by tumor resection or liver transplantation. We examined total HBV DNA and RNA as well as covalently closed circular DNA (cccDNA) and pregenomic RNA (pgRNA), which are considered as markers of active HBV replication. Total HBV DNA and RNA were detected in both T and NT samples in a majority of cases, but only a subset of tumors harbored detectable levels of HBV cccDNA and pgRNA (39% and 67%) compared to NT livers (66% and 90%; P < 0.01). Further evidence for HBV replication in tumor tissues was provided by sequencing of the X gene derived from episomal forms, showing that HBV genotypes differed between T and matched NT samples in 11 cases. The detection of pgRNA and cccDNA in tumors was correlated to the absence of tumorous microvascular invasion and to better patient survival. Analysis of gene expression profiles by Agilent microarrays revealed that pgRNA‐positive HCCs were characterized by low levels of cell cycle and DNA repair markers and expression of the HBV receptor, sodium taurocholate cotransporting polypeptide, indicating well‐differentiated tumors. Conclusion: HCC replicating HBV represents a subtype of weakly invasive HCC with a transcriptomic signature. pgRNA originating from nonintegrated, complete HBV genomes is a sensitive marker for viral replication and prognosis. (Hepatology 2018;67:86‐96).

The founder cells of the Nucleus pulposus, the centre of the intervertebral disc, originate in the embryonic notochord. After birth, mature notochordal cells (NC) are identified as key regulators of disc homeostasis. Better understanding of their biology has great potential in delaying the onset of disc degeneration or as a regenerative-cell source for disc repair. Using human pluripotent stem cells, we developed a two-step method to generate a stable NC-like population with a distinct molecular signature. Time-course analysis of lineage-specific markers shows that WNT pathway activation and transfection of the notochord-related transcription factor NOTO are sufficient to induce high levels of mesendoderm progenitors and favour their commitment toward the notochordal lineage instead of paraxial and lateral mesodermal or endodermal lineages. This study results in the identification of NOTO-regulated genes including some that are found expressed in human healthy disc tissue and highlights NOTO function in coordinating the gene network to human notochord differentiation.

The cellular microenvironment plays a major role in the biological functions of cells. Thus, biomaterials, especially hydrogels, which can be design to mimic the cellular microenvironment, are being increasingly used for cell encapsulation, delivery, and 3D culture, with the hope of controlling cell functions. Yet, much remains to be understood about the effects of cell-material interactions, and advanced synthetic strategies need to be developed to independently control the mechanical and biochemical properties of hydrogels. To address this challenge, we designed a new hyaluronic acid (HA)-based hydrogel platform using a click and bioorthogonal strain-promoted azide-alkyne cycloaddition (SPAAC) reaction. This approach facilitates the synthesis of hydrogels that are easy to synthesize and sterilize, have minimal swelling, are stable long term, and are cytocompatible. It provides bioorthogonal HA gels over an uncommonly large range of stiffness (0.5-45 kPa), all forming within 1-15 min. More importantly, our approach offers a versatile one-pot procedure to independently tune the hydrogel composition (e.g., polymer and adhesive peptides). Using this platform, we investigate the independent effects of polymer type, stiffness, and adhesion on the secretory properties of human adipose-derived stromal cells (hASCs) and demonstrate that HA can enhance the secretion of immunomodulatory factors by hASCs.

Polysaccharides have received a lot of attention in biomedical research for their high potential as scaffolds owing to their unique biological properties. Fibrillar scaffolds made of chitosan demonstrated high promise in tissue engineering, especially for skin. As far as bone regeneration is concerned, curdlan (1,3-β-glucan) is particularly interesting as it enhances bone growth by helping mesenchymal stem cell adhesion, by favoring their differentiation into osteoblasts and by limiting the osteoclastic activity. Therefore, we aim to combine both chitosan and curdlan polysaccharides in a new scaffold for bone regeneration. For that purpose, curdlan was electrospun as a blend with chitosan into a fibrillar scaffold. We show that this novel scaffold is biodegradable (8% at two weeks), exhibits a good swelling behavior (350%) and is non-cytotoxic in vitro. In addition, the benefit of incorporating curdlan in the scaffold was demonstrated in a scratch assay that evidences the ability of curdlan to express its immunomodulatory properties by enhancing cell migration. Thus, these innovative electrospun curdlan–chitosan scaffolds show great potential for bone tissue engineering.

Articular cartilage (AC) may be affected by many injuries including traumatic lesions that predispose to osteoarthritis. Currently there is no efficient cure for cartilage lesions. In that respect, new strategies for regenerating AC are contemplated with interest. In this context, we aim to develop and characterize an injectable, self-hardening, mechanically reinforced hydrogel (Si-HPCH) composed of silanised hydroxypropymethyl cellulose (Si-HPMC) mixed with silanised chitosan. The in vitro cytocompatibility of Si-HPCH was tested using human adipose stromal cells (hASC). In vivo, we first mixed Si-HPCH with hASC to observe cell viability after implantation in nude mice subcutis. Si-HPCH associated or not with canine ASC (cASC), was then tested for the repair of osteochondral defects in canine femoral condyles. Our data demonstrated that Si-HPCH supports hASC viability in culture. Moreover, Si-HPCH allows the transplantation of hASC in the subcutis of nude mice while maintaining their viability and secretory activity. In the canine osteochondral defect model, while the empty defects were only partially filled with a fibrous tissue, defects filled with Si-HPCH with or without cASC, revealed a significant osteochondral regeneration. To conclude, Si-HPCH is an injectable, self-setting and cytocompatible hydrogel able to support the in vitro and in vivo viability and activity of hASC as well as the regeneration of osteochondral defects in dogs when implanted alone or with ASC.

Hepatitis B virus infection is a major cause of liver diseases including hepatocellular carcinoma (HCC). The viral regulatory protein HBx is essential for viral replication and has been involved in the development of HCC. Recently, we characterized a subset of HCCs that replicate HBV. Our aim was to characterize HBx encoded by the full-length HBV DNA (cccDNA) in HCC and non-HCC liver. HBx genes were amplified and sequenced from eight paired HCC and non-HCC tissues in which HBV cccDNA and pgRNA were both present. Sequence analyses identified twelve amino acid positions mutated between HCC and non-HCC liver, and detected in at least three cases. We next assessed the impact of these mutations on HBx function by testing their transcriptional activity. We examined their ability to rescue the transcription of HBV virus deficient for HBx in differentiated HepaRG cells and to induce Smc5/6 degradation, which is mandatory for viral replication. We assessed their capacity to activate a CREB-dependent reporter. Finally we analyzed their growth suppressive activity using colony formation assays. Our results showed that most HBx variants isolated from HCC retain their ability to support HBV cccDNA transcription and to degrade Smc5/6. Strikingly, HCC specific HBx variants are impaired in their antiproliferative activity, which may be detrimental for tumor growth. In conclusion, in contrast to previous observations that tumor HBx variants lack transcriptional activity, we showed here that HBx variants have retained their ability to counteract Smc5/6 and thus to activate cccDNA transcription although they tend to lose antiproliferative activity.