IPO7 promotes lipopolysaccharide-induced inflammatory responses in human dental pulp cells via p38 MAPK and NF-κB signaling pathways.

Pulpitis is a chronic inflammatory process that greatly affects the physical, mental health and life quality of patients. Human dental pulp cells (hDPCs) are essential components of dental pulp tissue and play a significant role in pulpitis. Lipopolysaccharide (LPS) is an initiator of pulpitis and can induce the production of inflammatory cytokines in hDPCs by activating p38 MAPK and NF-κB signaling pathways. Importin7 (IPO7), a member of the importin-ß family, is widely expressed in many tissues. Previous studies have shown that IPO7 mediated nuclear translocation of p-p38 after stimulation, and IPO7 homologous protein IPO8 participated in human dental pulp inflammation. This research aims to investigate whether IPO7 is involved in pulpitis and explore its underlying mechanisms. In the current study, we found the expression of IPO7 was increased in pulpitis tissue. In vitro, hDPCs treated with LPS to mimic the inflammatory environment, the expression of IPO7 was increased. Knockdown of IPO7 significantly inhibited the production of inflammatory cytokines and suppressed the p38 MAPK and NF-κB signaling pathways. Activating the p38 MAPK and NF-κB signaling pathways by the p38 activator and p65 activator reversed the inflammatory responses. IPO7 interacted with p-p38 under LPS stimulation in hDPCs. In addition, the increased binding between IPO7 and p-p38 is associated with the decreased binding ability of IPO7 to Sirt2. In conclusion, we found that IPO7 was highly expressed in pulpitis and played a vital role in modulating human dental pulp inflammation.

Reactive Oxygen Species-Induced Inhibition of Odontoblastic Differentiation of Mouse Dental Papilla Cells Is Mediated by Downregulation of Importin 7.

Tooth dentin is a crucial tooth structure. The biological process of odontoblast differentiation is essential for formation of normal dentin. Accumulation of reactive oxygen species (ROS) leads to oxidative stress, which can influence the differentiation of several cells. As a member of the importin-ß superfamily, importin 7 (IPO7) is essential for nucleocytoplasmic transport and plays an important role in the processes of odontoblast differentiation and oxidative stress. Nevertheless, the association between ROS, IPO7, and odontoblast differentiation in mouse dental papilla cells (mDPCs) and the underlying mechanisms remain to be elucidated. In this study, we confirmed that ROS suppressed odontoblastic differentiation of mDPCs as well as the expression and nucleocytoplasmic shuttle of IPO7 in cells, while overexpression of IPO7 can rescue these effects. ROS resulted in increased phosphorylation of p38 and cytoplasmic aggregation of phosphorylated p38 (p-p38), which was able to be reversed by overexpression of IPO7. p-p38 interacted with IPO7 in mDPCs without hydrogen peroxide (H2O2) treatment, but in the presence of H2O2, the interaction between p-p38 and IPO7 was significantly decreased. Inhibition of IPO7 increased the expression level and nuclear translocation of p53, which are mediated by cytoplasmic aggregation of p-p38. In conclusion, ROS inhibited odontoblastic differentiation of mDPCs, which is mediated by downregulation and damaged nucleocytoplasmic shuttle of IPO7.

IPO7 Promotes Odontoblastic Differentiation and Inhibits Osteoblastic Differentiation Through Regulation of RUNX2 Expression and Translocation.

RUNX2, an important transcriptional factor for both odontoblastic and osteoblastic differentiation, is upregulated during osteoblastic differentiation, but downregulated during late odontoblastic differentiation. However, the specific mechanism of the different RUNX2 expression in bone and dentin remains largely unknown. Importin 7 (IPO7), a member of the karyopherin ß-superfamily, mediates nucleocytoplasmic transport of proteins. In this study, we found that IPO7 was increasingly expressed from pre-odontoblasts to mature odontoblasts. IPO7 expression was increased with odontoblastic differentiation of mouse dental papilla cells (mDPCs) and knockdown of IPO7-inhibited cell differentiation. While in MC3T3-E1 cells, IPO7 was decreased during osteoblastic differentiation and knockdown of IPO7-promoted cell differentiation. In mPDCs, IPO7 was able to bind with some odontoblastic transcription factors, and imported them into the nucleus, but not with RUNX2. Furthermore, IPO7 inhibited the total RUNX2 expression by promoting HDAC6 nuclear localization during odontoblastic differentiation. However, in MC3T3-E1 cells, IPO7 inhibited the nuclear distribution of RUNX2 but did not affect the total protein level of RUNX2. In conclusion, we found that IPO7 promotes odontoblastic differentiation and inhibits osteoblastic differentiation through regulating RUNX2 expression and translocation differently.

The regulation of Msx1 by BMP4/pSmad1/5 signaling is mediated by importin7 in dental mesenchymal cells.

Msx1 is essential for the maintenance of the odontogenic fate of dental mesenchymal cells, and is regulated by BMP/Smad1/5 signaling in a Smad4-independent manner. However, the exact co-factors that assist pSmad1/5 entering the nucleus to regulate Msx1 in dental mesenchymal cells are still unknown. Importin7 (IPO7) is one of the important members of importin ß-superfamily, which is mainly responsible for nucleocytoplasmic shuttling of RNAs and proteins, including transcription factors. This study aims to investigate whether IPO7 participates in the nuclear translocation of pSmad1/5 activated by BMP4 to regulate Msx1 expression in mouse dental mesenchymal cells. In the current study, we found that IPO7 was strongly expressed in the mouse dental mesenchymal cells at postnatal day 1 (PN1) both in vitro and in vivo. With BMP4 stimulation, IPO7 showed a translocation from the cytoplasm to the nucleus. Knockdown of IPO7 with siRNA inhibited the nuclear accumulation of pSmad1/5 in response to BMP4 stimulation. Furthermore, the co-immunoprecipitation assay showed pSmad1/5 was a nuclear import cargo of IPO7. Next, knockdown of IPO7 abolished the upregulation of Msx1 induced by BMP4, while overexpression of Smad1 was able to rescue the Msx1 expression. Finally, ChIP and Re-ChIP assay showed IPO7 facilitated the recruitment of pSmad1/5 to the Msx1 promoter. Taken together, our data demonstrated that the regulation of Msx1 by BMP4/pSmad1/5 signaling is mediated by importin7 in mouse dental mesenchymal cells.