N-glycosylation modifies prostaglandin E2 uptake by reducing cell surface expression of SLCO2A1.

SLCO2A1 functions as a prostaglandin (PG) influx transporter to facilitate intracellular oxidation of PGs and its defect causes dysregulation of PG signaling and metabolism. This study aimed to clarify effects of N-glycosylation on functional SLCO2A1 expression. Putative N-glycosylation site(s) (N134, N478, and/or N491) of human SLCO2A1 were mutated to Q and wild-type (WT) and mutant forms were expressed in HEK293 and human epithelial cells. Molecular weight of WT decreased to nearly 55 kDa by PNGase F treatment and was identical to that of triple mutant (TM, i.e., N134Q/N478Q/N491Q). Transport affinity of TM for PGE2 (Km of 392.7 nM) was comparable to that of WT (Km of 328.5 nM); however, immunoassays showed that TM cell surface expression remained at 24% of WT in HEK293 cells, resulting in a reduced cellular PGE2 uptake. These results suggest N-glycosylation modifies cellular PGE2 uptake by decreasing SLCO2A1 localization to the plasma membrane.

Regulation of IGF-I by IGFBP3 and IGFBP5 during odontoblast differentiation in mice.

OBJECTIVES: Although intracellular signaling pathways of insulin-like growth factor I (IGF-I) related to the proliferation of dental pulp cells have been investigated, the switching mechanism from cell proliferation to differentiation during odontogenesis remains elusive. This study aimed to elucidate the role of IGF binding protein (IGFBP) 3 and 5 in regulation of IGF-I during odontoblast differentiation in mouse incisors. METHODS: The detailed expression patterns of IGF-I, IGF-I receptor (IGF-IR), IGFBP3, and IGFBP5 together with that of an odontoblast differentiation marker, nestin, were examined by immunohistochemistry and/or in situ hybridization using paraffinized sections of TetOP-H2B-GFP mouse incisors at postnatal 4 weeks. RESULTS: Undifferentiated dental papilla cells and preodontoblasts (preOB) showed intense IGF-I- and IGF-IRα-positive reactions, and the expression was observed in differentiated odontoblasts, such as immature odontoblasts (iOB) and mature odontoblasts (mOB). IGFBP3/Igfbp3 was transiently expressed in preOB and early iOB, and the intensity of expression gradually reduced with the progression of odontoblast differentiation. In contrast, immunohistochemical analysis for IGFBP5 identified a positive reaction in the undifferentiated dental papilla cells and differentiated odontoblasts, and the expression of Igfbp5 was reduced in the differentiated odontoblasts. CONCLUSION: The present study demonstrated the expression patterns of IGF-I, IGF-IR, IGFBP3, and IGFBP5 during odontoblast differentiation in mouse incisors. These results suggested that IGFBP3 regulates the transition from the proliferative to differentiation stage by inhibiting the action of IGF-I on the proliferation of dental papilla cells, and that IGFBP5 plays an important role in the maintenance of the differentiated odontoblasts during tooth development.