Effect of Valproic Acid on Promoting the Differentiation of Human Embryonic Stem Cells Into Cholangiocyte-Like Cells.

Cholangiocytes form a complex 3D network of bile ducts in the liver and contribute to liver function. The damage or destruction of cholangiocytes can lead to biliary diseases, and the shortage of cholangiocytes remains an obstacle for drug development targeting biliary diseases. Valproic acid (VPA) is a potent activator of Notch signaling pathway that is essential for cholangiocyte differentiation. Here, we report a VPA-based approach for cholangiocyte differentiation of human pluripotent stem cells. VPA activated Notch2 expression and upregulated HES-1, HEY-1, and Sox9 gene expression in hESC-derived hepatoblast. After 7 days treatment, VPA promoted successful differentiation of hepatoblast into cholangiocytes expressing cholangiocyte marker genes (AE2, AQP1, CFTR) and proteins (CK19 and CK7). In addition, the differentiated cholangiocytes formed bile duct-like structures after implantation into the spleen of NOD/SCID mice. Our results suggested that VPA can promote hESC differentiation to cholangiocyte-like cells. The induced cholangiocytes may serve as a potential cell source for both in vitro modeling and regenerative therapy of cholangiopathies. The findings can also support further development of small-molecule based differentiation protocols for cholangiocyte production.

Oral Delivery of Bioactive Glass-Loaded Core-Shell Hydrogel Microspheres for Effective Treatment of Inflammatory Bowel Disease.

Resolving inflammation and promoting intestinal tissue regeneration are critical for inflammatory bowel disease (IBD) treatment. Bioactive glass (BG) is a clinically approved bone graft material and has been shown to modulate inflammatory response, but it is unknown whether BG can be applied to treat IBD. Here, it is reported that BG attenuates pro-inflammatory response of lipopolysaccharide (LPS)-stimulated macrophages and hence reduces inflammatory damage to intestinal organoids in vitro. In addition, zein/sodium alginate-based core-shell microspheres (Zein/SA/BG) are developed for oral delivery of BG, which helps prevent premature dissolution of BG in the stomach. The results show that Zein/SA/BG protects BG from a gastric-simulated environment while dissolved in an intestinal-simulated environment. When administered to acute and chronic colitis mice model, Zein/SA/BG significantly reduces intestinal inflammation, promotes epithelial tissue regeneration, and partially restores microbiota homeostasis. These findings are the first to reveal the therapeutic efficacy of BG against IBD, which may provide a new therapeutic approach at low cost for effective IBD treatment.

Bioactive glass-elicited stem cell-derived extracellular vesicles regulate M2 macrophage polarization and angiogenesis to improve tendon regeneration and functional recovery.

Effective countermeasures for tendon injury remains unsatisfactory. Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs)-based therapy via regulation of Mφ-mediated angiogenesis has emerged as a promising strategy for tissue regeneration. Still, approaches to tailor the functions of EVs to treat tendon injuries have been limited. We reported a novel strategy by applying MSC-EVs boosted with bioactive glasses (BG). BG-elicited EVs (EVB) showed up-regulation of medicinal miRNAs, including miR-199b-3p and miR-125a-5p, which play a pivotal role in M2 Mφ-mediated angiogenesis. EVB accelerated angiogenesis via the reprogrammed anti-inflammatory M2 Mφs compared with naïve MSC-EVs (EVN). In rodent Achilles tendon rupture model, EVB local administration activated anti-inflammatory responses via M2 polarization and led to a spatial correlation between M2 Mφs and newly formed blood vessels. Our results showed that EVB outperformed EVN in promoting tenogenesis and in reducing detrimental morphological changes without causing heterotopic ossification. Biomechanical test revealed that EVB significantly improved ultimate load, stiffness, and tensile modulus of the repaired tendon, along with a positive correlation between M2/M1 ratio and biomechanical properties. On the basis of the boosted nature to reprogram regenerative microenvironment, EVB holds considerable potential to be developed as a next-generation therapeutic modality for enhancing functional regeneration to achieve satisfying tendon regeneration.

Efficient hepatic differentiation of hydrogel microsphere-encapsulated human pluripotent stem cells for engineering prevascularized liver tissue.

Liver tissue engineering is promising as an alternative strategy to treat liver failure. However, generating functional hepatocytes from stem cells is conventionally restricted by the immature status of differentiated cells. Besides, embedding hepatocytes in bulk scaffold is limited by a lack of vascularity and low cell-packing density. Here, we fabricate collagen type I (COL1) microspheres for efficient hepatic differentiation of pluripotent stem cells and subsequent assembly of prevascularized liver tissue (PLT). Using a microfluidic platform, we demonstrate that hydrogel COL1 microspheres (mCOL1) encapsulating human embryonic stem cells (hESCs) can be reproducibly generated and efficiently differentiated into hepatocyte-like cells (HLCs) microspheres for the first time. Compared with other culture configurations such as encapsulation of hESC in a bulk COL1 hydrogel and 2D monolayer culture, mCOL1 with high uniformity produce HLC microspheres of improved maturity based on comprehensive analyses of cell morphology, transcriptome profile, hepatic marker expression and hepatic functions. In addition, these HLC microspheres can be applied as building blocks to self-assemble with endothelial cells to construct a dense PLT. The PLT resembles native liver tissue with high cell-packing density, shows successful engraftment in mice liver following implantation, and exhibits improved hepatic functionin vivo. Overall, it is believed that this multiscale technology will advance the fabrication of stem cell-based liver tissue for regenerative medicine, drug screening, andin vitroliver modeling.

Dissolved oxygen from microalgae-gel patch promotes chronic wound healing in diabetes.

Chronic wounds in diabetes undergo a lifetime risk of developing into diabetic foot ulcers. Oxygen is crucial to wound healing by regulating cell proliferation, migration, and neovascularization. However, current oxygen therapies, including hyperbaric oxygen (HBO) and topical gaseous oxygen (TGO), mainly employ gaseous oxygen delivery, which is much less effective in penetrating the skin. Here, we introduce an oxygen-producing patch, made of living microalgae hydrogel, which can produce dissolved oxygen. The superior performance of the patch that results from its dissolved oxygen delivery is >100-fold much more efficient than TGO penetrating the skin. Further experiments indicate that the patch could promote cell proliferation, migration, and tube formation in vitro, and improve chronic wound healing and the survival of skin grafts in diabetic mice. We believe that the microalgae-gel patch can provide continuous dissolved oxygen to improve chronic wound healing.