ABSTRACT
The L1 cell adhesion molecule (L1CAM) has been implicated in tumor progression of many types of cancers, but its role in prostate cancer and its application in targeted gene therapy have not been investigated. Herein, we demonstrated that the L1CAM was expressed in androgen-insensitive and highly metastatic human prostate cancer cell lines. The correlation between L1CAM expression and prostate cancer metastasis was also validated in serum samples of prostate cancer patients. Knockdown of L1CAM expression in prostate cancer cells by RNA interference significantly decreased their aggressive behaviors, including colony formation, migration and invasion in vitro, and tumor formation in a metastatic murine model. These anti-malignant phenotypes of L1CAM-knockdown cancer cells were accompanied by G0/G1 cell cycle arrest and suppression of matrix metalloproteinase (MMP)-2 and MMP-9 expression and nuclear factor NF-κB activation. In vivo targeting of L1CAM expression using liposome-encapsulated L1CAM siRNAs effectively inhibited prostate cancer growth in mouse bone, which was associated with decreased L1CAM expression and cell proliferation by tumor cells. These results provide the first evidence for L1CAM being a major contributor to prostate cancer metastasis and translational application of siRNA-based L1CAM-targeted therapy.
Subject(s)
Bone Neoplasms/secondary , Bone Neoplasms/therapy , Neural Cell Adhesion Molecule L1/biosynthesis , Prostatic Neoplasms/pathology , Prostatic Neoplasms/therapy , RNA, Small Interfering/administration & dosage , Animals , Apoptosis/genetics , Bone Neoplasms/genetics , Bone Neoplasms/metabolism , Cell Line, Tumor , Humans , Liposomes/administration & dosage , Male , Mice , Mice, Nude , Molecular Targeted Therapy , Neural Cell Adhesion Molecule L1/genetics , Neural Cell Adhesion Molecule L1/metabolism , Prostatic Neoplasms/genetics , Prostatic Neoplasms/metabolism , RNA, Small Interfering/genetics , Signal Transduction , Transfection/methodsABSTRACT
The DOC-2/DAB2 interactive protein (DAB2IP) is a new member of the Ras GTPase-activating protein family. Recent studies have shown that, in addition to its tumor suppressive role in various tumors, DAB2IP also plays an important role in regulating neuronal migration and positioning during brain development. In this study, we determined the roles of DAB2IP in the neuronal differentiation of human mesenchymal stem cells (hMSCs). We found that lentiviral short hairpin RNA (shRNA)-mediated knockdown of DAB2IP promoted the mesenchymal-to-neuroepithelial stem cell transition (MtNeST) and neuronal differentiation, which were accompanied by a reduction of cell proliferation but not apoptosis or cellular senescence. This suggests that DAB2IP plays an important role in the neuronal induction of hMSCs. Moreover, our finding that reduction of glycogen synthase kinase 3 beta (GSK3ß) activity upon LiCl pretreatment inhibited both the MtNeST and production of MAP2-positive cells upon DAB2IP knockdown suggests that this transition is most likely mediated by regulation of the GSK3ß signaling pathway. Our study demonstrates that DAB2IP participates in the first step of neuron induction of hMSCs, which implies a potentially important role for DAB2IP in the MtNeST during neurogenesis.
Subject(s)
Cell Differentiation , Mesenchymal Stem Cells/cytology , Neuroepithelial Cells/cytology , Neurons/cytology , ras GTPase-Activating Proteins/metabolism , Apoptosis , Blotting, Western , Cell Proliferation , Cells, Cultured , Down-Regulation , Flow Cytometry , Fluorescent Antibody Technique , Humans , Immunoenzyme Techniques , Lentivirus/genetics , Mesenchymal Stem Cells/metabolism , Microscopy, Fluorescence , Neuroepithelial Cells/metabolism , Neurons/metabolism , RNA, Messenger/genetics , RNA, Small Interfering/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , ras GTPase-Activating Proteins/antagonists & inhibitors , ras GTPase-Activating Proteins/geneticsABSTRACT
Enhancer of zeste homolog 2 (EZH2) regulates stem cells renewal, maintenance, and differentiation into different cell lineages including neuron. Changes in intracellular Ca(2+) concentration play a critical role in the differentiation of neurons. However, whether EZH2 modulates intracellular Ca(2+) signaling in regulating neuronal differentiation from human mesenchymal stem cells (hMSCs) still remains unclear. When hMSCs were treated with a Ca(2+) chelator or a PLC inhibitor to block IP(3)-mediated Ca(2+) signaling, neuronal differentiation was disrupted. EZH2 bound to the promoter region of PIP5K1C to suppress its transcription in proliferating hMSCs. Interestingly, knockdown of EZH2 enhanced the expression of PIP5K1C, which in turn increased the amount of PI(4,5)P(2), a precursor of IP(3), and resulted in increasing the intracellular Ca(2+) level, suggesting that EZH2 negatively regulates intracellular Ca(2+) through suppression of PIP5K1C. Knockdown of EZH2 also enhanced hMSCs differentiation into functional neuron both in vitro and in vivo. In contrast, knockdown of PIP5K1C significantly reduced PI(4,5)P(2) contents and intracellular Ca(2+) release in EZH2-silenced cells and resulted in the disruption of neuronal differentiation from hMSCs. Here, we provide the first evidence to demonstrate that after induction to neuronal differentiation, decreased EZH2 activates the expression of PIP5K1C to evoke intracellular Ca(2+) signaling, which leads hMSCs to differentiate into functional neuron lineage. Activation of intracellular Ca(2+) signaling by repressing or knocking down EZH2 might be a potential strategy to promote neuronal differentiation from hMSCs for application to neurological dysfunction diseases.