Since the discovery of stem cells and multipotency characteristics of mesenchymal stem cells (MSCs), there has been tremendous development in regenerative medicine. MSCs derived from bone marrow have been widely used in various research applications, yet there are limitations such as invasiveness of obtaining samples, low yield and proliferation rate, and questions regarding their practicality in clinical applications. Some have suggested that MSCs from other sources, specifically those derived from palatine tonsil tissues, that is, tonsil-derived MSCs (TMSCs), could be considered as a new potential therapeutic tool in regenerative medicine due to their superior proliferation rate and differentiation capabilities with low immunogenicity and ease of obtaining. Several studies have determined that TMSCs have differentiation potential not only into the mesodermal lineage but also into the endodermal as well as ectodermal lineages, expanding their potential usage and placing them as an appealing option to consider for future studies in regenerative medicine. In this review, the differentiation capacities of TMSCs and their therapeutic competencies from past studies are addressed. Stem Cells 2019;37:1252–1260

Zearalenone (ZEN) is a mycotoxin that has been reported to damage various types of cells/tissues, yet its effects on endothelial cells (ECs) have never been investigated. Therefore, this study investigates the potential effects of ZEN using bovine aortic ECs (BAECs). In this study, we found that ZEN induced apoptosis of BAECs through increased cleavage of caspase 3 and poly ADP-ribose polymerase (PARP). ZEN also increased phosphorylation of ERK1/2 and p53, and treatment with the ERK1/2 or p53 inhibitor reversed ZEN-induced EC apoptosis. Transfection of BAECs with small interfering RNA against ERK1/2 or p53 revealed ERK1/2 as an upstream target of p53 in ZEN-stimulated apoptosis. ZEN increased the production of reactive oxygen species (ROS), yet treatment with the antioxidant did not prevent EC apoptosis. Similarly, blocking of estrogen receptors by specific inhibitors also did not prevent ZEN-induced apoptosis. Finally, chelation of cytosolic calcium (Ca²⁺) using BAPTA-AM or inhibition of endoplasmic reticulum (ER) Ca²⁺ channel using 2-APB reversed ZEN-induced EC apoptosis, but not by inhibiting ER stress using 4-PBA. Together, our findings demonstrate that ZEN induces EC apoptosis through an ERK1/2/p53/caspase 3 signaling pathway activated by Ca²⁺ release from the ER, and this pathway is independent of ROS production and estrogen receptor activation.

Our previous study found that the level of CCN1 increases as osteogenic differentiation progresses in tonsil-derived mesenchymal stem cells (TMSCs). This study investigated how CCN1 is regulated through HDAC inhibition in TMSCs and their relationship with osteogenesis. Valproic acid (VPA) (1–5 mM), a well-known histone deacetylase (HDAC) inhibitor, strongly inhibited TMSC proliferation without altering MSC-specific surface markers, CD14, 34, 45, 73, 90 and 105. However, CD146 expression increased at 5 mM VPA. VPA increased osteogenic differentiation of TMSCs but decreased adipogenesis and chondrogenesis, as evidenced by the cell-specific staining of differentiation. The former was validated by the increased osteocalcin (OCN). The changes in CCN1 by VPA was biphasic; it increased until 48 h and decreased thereafter. Knockdown of CCN1 by using siRNA inhibited the osteogenic effect of VPA. VPA had no effect on CCN1 mRNA expression, but inhibition of protein synthesis by cycloheximide showed that VPA slowed down the CCN1 protein degradation. Moreover, overexpression of HDAC1 completely inhibited VPA-induced CCN1. Our results indicate that VPA inhibits the HDAC1, inducing CCN1 protein stability rather than gene expression, thereby promoting osteogenic differentiation of TMSCs. These findings present the noble implication of VPA as an inhibitor of HDAC1 to facilitate CCN1-induced osteogenic differentiation of MSCs.

Zearalenone (ZEN) is a non-steroidal mycotoxin that has various toxicological impacts on mammalian health. Here, we found that ZEN significantly affected the production of nitric oxide (NO) and the expression of endothelial NO synthase (eNOS) of bovine aortic endothelial cells (BAECs). A promoter analysis using 5′-serially deleted human eNOS promoter revealed that the proximal region (−135 to +22) was responsible for ZEN-mediated reduction of the human eNOS promoter activity. This effect was reversed by mutation of two specificity protein 1 (Sp1) binding elements in the human eNOS promoter. A chromatin immunoprecipitation assay revealed that ZEN increased Sp1 binding to the bovine eNOS promoter region (−113 to −12), which is homologous to −135 to +22 of the human eNOS promoter region. We also found that ZEN promoted the binding of the pregnane X receptor (PXR) to Sp1 of the bovine eNOS, consequently decreasing eNOS expression. This reduction of eNOS could have contributed to the decreased acetylcholine-induced vessel relaxation upon ZEN treatment in our ex vivo study using mouse aortas. In conclusion, our data demonstrate that ZEN decreases eNOS expression by enhancing the binding of PXR-Sp1 to the eNOS promoter, thereby decreasing NO production and potentially causing vessel dysfunction.

Background: The lack of appropriate mesenchymal stem cells (MSCs) selection methods has given the challenges for standardized harvesting, processing, and phenotyping procedures of MSCs. Genetic engineering coupled with high-throughput proteomic studies of MSC surface markers arises as a promising strategy to identify stem cell-specific markers. However, the technical limitations are the key factors making it less suitable to provide an appropriate starting material for the screening platform. A more accurate, easily accessible approach is required to solve the issues. Methods: This study established a high-throughput screening strategy with forward versus side scatter gating to identify the adipogenesis-associated markers of bone marrow-derived MSCs (BMSCs) and tonsil-derived MSCs (TMSCs). We classified the MSC-derived adipogenic differentiated cells into two clusters: lipid-rich cells as side scatter (SSC)-high population and lipid-poor cells as SSC-low population. By screening the expression of 242 cell surface proteins, we identified the surface markers which exclusively found in lipid-rich subpopulation as the specific markers for BMSCs and TMSCs. Results: High-throughput screening of the expression of 242 cell surface proteins indicated that CD49f and CD146 were specific for BMSCs and TMSCs. Subsequent immunostaining confirmed the consistent specific expression of CD49f and CD146 and in BMSCs and TMSCs. Enrichment of MSCs by CD49f and CD146 surface markers demonstrated that the simultaneous expression of CD49f and CD146 is required for adipogenesis and osteogenesis of mesenchymal stem cells. Furthermore, the fate decision of MSCs from different sources is regulated by distinct responses of cells to differentiation stimulations despite sharing a common CD49f⁺CD146⁺ immunophenotype. Conclusions: We established an accurate, robust, transgene-free method for screening adipogenesis associated cell surface proteins. This provided a valuable tool to investigate MSC-specific markers. Additionally, we showed a possible crosstalk between CD49f and CD146 modulates the adipogenesis of MSCs.