Glucosylceramide is the primary molecule of glycosphingolipids, and its metabolic regulation is crucial for life. Defects in the catabolizing enzyme, glucocerebrosidase (GCase), cause a lysosomal storage disorder known as Gaucher disease. However, the genetic regulation of GCase has not been fully understood. Here we show the redefined structure of the GCase coding gene (GBA), and clarify the regulatory mechanisms of its transcription and translation. First, alternative uses of the two GBA gene promoters were identified in fibroblasts and HL60-derived macrophages. Intriguingly, both GBA transcripts and GCase activities were induced in macrophages but not in neutrophils. Second, we observed cap-independent translation occurs via unique internal ribosome entry site activities in first promoter-driven GBA transcripts. Third, the reciprocal expression was observed in GBA and miR22-3p versus GBAP1 transcripts before and after HL60-induced macrophage differentiation. Nevertheless, these findings clearly demonstrate novel cell-type-specific GBA gene expression regulatory mechanisms, providing new insights into GCase biology.

Dental pulp cells have a capacity to differentiate into mineralization-inducing cells. To clarify the molecular mechanism, we established an in vitro mineralization-inducing system by rat clonal dental pulp cell line, RPC-C2A, and tried to purify a mineralization-inducing factor in conditioned medium (CM) from preosteoblastic MC3T3-E1 cells. The active factor was impermeable to an ultrafiltrating membrane, and sedimented by ultracentrifugation. The sedimented factor was found as a needle-like structure about 1.3 µm in average length as observed by transmission electron microscopy. The factor contained type I collagen, suggesting not a matrix vesicle, but a soluble matrix. The mineralization-inducing activity was also detected in CM from primary culture of rat calvaria (RC) cells. These results suggested that the soluble matrices from osteoblastic cells serve, at least in part, as differentiation-inducing agents. J. Med. Invest. 53: 297-302, August, 2006

Considering that every tissue/organ has the most suitable microenvironment for its functional cells, controlling induced pluripotent stem cell (iPSC) differentiation by culture on frozen sections having a suitable microenvironment is possible. Induced PSCs were cultured on frozen sections of the liver, the brain, the spinal cord, and cover glasses (control) for 9 days. The iPSCs cultured on the sections of the liver resembled hepatocytes, whereas those on sections of the brain and the spinal cord resembled neuronal cells. The percentage of hepatocytic marker-positive cells in the iPSCs cultured on the sections of the liver was statistically higher than that of those in the iPSCs cultured on the sections of the brain and the spinal cord or on cover glasses. In contrast, the iPSCs cultured on the sections of the brain and the spinal cord revealed a high percentage of neural marker-positive cells. Thus, iPSCs can be differentiated into a specific cell lineage in response to specific factors within frozen sections of tissues/organs. Differentiation efficacy of the frozen sections markedly differed between the iPSC clones. Therefore, our induction method could be simple and effective for evaluating the iPSC quality.