In Vitro Ballooned Hepatocytes Can Be Produced By Primary Human Hepatocytes and Hepatic Stellate Cell Sheets

Background Despite the increasing prevalence of Nonalcoholic steatohepatitis (NASH) worldwide, there is no effective treatment available for this disease. “Ballooned hepatocyte” is a characteristic finding in NASH and is correlated with disease prognosis, but their mechanisms of action are poorly understood; furthermore, neither animal nor in vitro models of NASH have been able to adequately represent ballooned hepatocytes. Herein, we engineered cell sheets to develop a new in vitro model of ballooned hepatocytes. Methods Primary human hepatocytes (PHH) and Hepatic stellate cells (HSC) were co-cultured to produce cell sheets, which were cultured in glucose and lipid containing medium, following which histological and functional analyses were performed. Results Histological findings showed hepatocyte ballooning, accumulation of fat droplets, abnormal cytokeratin arrangement, and the presence of Mallory-Denk bodies and abnormal organelles. These findings are similar to those of ballooned hepatocytes in human NASH. Functional analysis showed elevated levels of TGFβ-1, SHH, and p62, but not TNF-α, IL-8. Conclusions Exposure of PHH/HSC sheets to a glucolipotoxicity environment induces ballooned hepatocyte without inflammation. Moreover, fibrosis is an important mechanism underlying ballooned hepatocytes and could be the basis for the development of a new in vitro NASH model with ballooned hepatocytes.

Dental Pulp Mesenchymal Stem Cells Promote Polarization of M2 Macrophages and Accelerate Wound Healing by Secreting CCL2

Background Mesenchymal stem cells (MSCs) are widely used in tissue engineering owing to their regenerative potential and immunomodulatory capacity. The crosstalk between MSCs and the host immune function plays a key role in the efficiency of tissue regeneration. However, the difference in immunological modulation and tissue regeneration function between MSCs from different sources remains unclear. Methods Human mesenchymal stem cells derived from bone marrow (BMMSCs), periodontal ligament (PDLSCs), adipose (ADSCs), and dental pulp (DPSCs) were obtained and induced to form cell sheets under the condition of 20 ug/ml vitamin C. The MSC cell sheets carried by hydroxyapatite/tricalciumphosphate (HA/TCP) particles were transplanted subcutaneously into C57BL6 mice for 8 weeks. Histological analyses were performed to detect the tissue regeneration potential and macrophages polarization in vivo. Then, THP-1 macrophages were co-cultured with MSCs and quantitative real-time polymerase chain reaction, immunofluorescent staining, western blotting, and enzyme-linked immunosorbent assay were used to investigate the function and mechanism of MSCs on macrophages in vitro. Finally, a wound healing model of the palatal mucosa was performed to confirm the effect of MSCs on macrophages and tissue healing efficiency. Results Compared to PDLSCs, BMMSCs, and ADSCs, DPSCs exhibited greater tissue regeneration potential, with greater tissue volume, higher Ki67 expression, and less apoptosis in the regenerated tissue of wild-type C57BL6 mice. In addition, DSPCs triggered more M2 macrophages in the regenerated tissue than other MSCs. Our data showed that DPSCs exhibited higher expression levels of C-C Motif Chemokine Ligand 2 (CCL2), and specific blocking of CCL2 by neutralising antibodies can significantly inhibit the DPSCs-induced polarization of M2 macrophages. Finally, DPSCs transplantation promoted wound healing of the palatal mucosa and M2 macrophages polarization in vivo, which could be significantly impaired by CCL2 neutralising antibody. Conclusions Our data indicate that DPSCs exert better tissue regeneration potential and immunoregulatory function by secreting CCL2. These results suggest that CCL2 application can enhance MSC-mediated tissue regeneration or wound healing.

Strategies Using Gelatin Microparticles for Regenerative Therapy and Drug Screening Applications

Gelatin, a denatured form of collagen, is an attractive biomaterial for biotechnology. In particular, gelatin particles have been noted due to their attractive properties as drug carriers. The drug release from gelatin particles can be easily controlled by the crosslinking degree of gelatin molecule, responding to the purpose of the research. The gelatin particles capable of drug release are effective in wound healing, drug screening models. For example, a sustained release of growth factors for tissue regeneration at the injured sites can heal a wound. In the case of the drug screening model, a tissue-like model composed of cells with high activity by the sustained release of drug or growth factor provides reliable results of drug effects. Gelatin particles are effective in drug delivery and the culture of spheroids or cell sheets because the particles prevent hypoxia-derived cell death. This review introduces recent research on gelatin microparticles-based strategies for regenerative therapy and drug screening models.

Role of FBXW2 in explant cultures of bovine periosteum-derived cells

Objective Bone regeneration is a potential technique for treating osteoporosis. A previous study reported that F-box and WD-40 domain-containing protein 2 (FBXW2) localized with osteocalcin in bovine periosteum after 5 weeks of explant culture. However, the osteoblastic functions of FBXW2 remain unclear. In this study, double-fluorescent immunostaining was used to investigate the potential role of FBXW2 and its relationship with osteocalcin. Results At day 0, FBXW2 was expressed in the cambium layer between the bone and periosteum, while osteocalcin was expressed in bone. After explant culture, changes in the periosteum were observed from weeks 1 to 7. At week 1, partial FBXW2 expression was seen with a small amount of osteocalcin. At week 2, a layer of FBXW2 was observed. From weeks 3 to 7, tube-like structures of FBXW and osteocalcin were observed, and periosteum-derived cells were released from the periosteum in areas where no FBXW2 was observed. Bovine periosteum-derived cells can form a three-dimensional cell pellet, because multilayered cell sheets are formed inside of the periosteum in vitro. It is shown that in results FBXW2 is produced in periosteal explants near sites where initial osteogenic activity is observed, suggesting that it may be involved in periosteal osteogenesis.