Dong-Soo Suh

Cancer‐Derived Lysophosphatidic Acid Stimulates Differentiation of Human Mesenchymal Stem Cells to Myofibroblast‐Like Cells

Lysophosphatidic acid (LPA) is enriched in ascites of ovarian cancer patients and is involved in growth and invasion of ovarian cancer cells. Accumulating evidence suggests cancer‐associated myofibroblasts play a pivotal role in tumorigenesis through secreting stromal cell‐derived factor‐1 (SDF‐1). In the present study, we demonstrate that LPA induces expression of α‐smooth muscle actin (α‐SMA), a marker for myofibroblasts, in human adipose tissue‐derived mesenchymal stem cells (hADSCs). The LPA‐induced expression of α‐SMA was completely abrogated by pretreatment of the cells with Ki16425, an antagonist of LPA receptors, or by silencing LPA1 or LPA2 isoform expression with small interference RNA (siRNA). LPA elicited phosphorylation of Smad2/3, and siRNA‐mediated depletion of endogenous Smad2/3 or adenoviral expression of Smad7, an inhibitory Smad, abrogated the LPA induced expression of α‐SMA and phosphorylation of Smad2/3. LPA‐induced secretion of transforming growth factor (TGF)‐β1 in hADSCs, and pretreatment of the cells with SB431542, a TGF‐β type I receptor kinase inhibitor, or anti‐TGF‐β1 neutralizing antibody inhibited the LPA‐induced expression of α‐SMA and phosphorylation of Smad2. Furthermore, ascites from ovarian cancer patients or conditioned medium from ovarian cancer cells induced expression of α‐SMA and phosphorylation of Smad2, and pretreatment of the cells with Ki16425 or SB431542 abrogated the expression of α‐SMA and phosphorylation of Smad2. In addition, LPA increased the expression of SDF‐1 in hADSCs, and pretreatment of the cells with Ki16425 or SB431562 attenuated the LPA‐stimulated expression of SDF‐1. These results suggest that cancer‐derived LPA stimulates differentiation of hADSCs to myofibroblast‐like cells and increases SDF‐1 expression through activating autocrine TGF‐β1‐Smad signaling pathway.

Lysophosphatidic acid in malignant ascites stimulates migration of human mesenchymal stem cells

Lysophosphatidic acid (LPA) is elevated in ascites of ovarian cancer patients and is involved in growth and invasion of ovarian cancer cells. Accumulating evidence suggests a pivotal role of mesenchymal stem cells (MSCs) or stromal cells in tumorigenesis. In the present study, we demonstrated that ascites from ovarian cancer patients and LPA increased migration of human MSCs. The migration of MSCs induced by LPA and malignant ascites was completely abrogated by pretreatment with Ki16425, an antagonist of LPA receptors, and by silencing of endogenous LPA1, but not LPA2, with small interference RNA, suggesting a key role of LPA played in the malignant ascites‐induced migration. LPA induced activation of ERK through pertussis toxin‐sensitive manner, and pretreatment of MSCs with U0126, a MEK inhibitor, or pertussis toxin attenuated the LPA‐induced migration. Moreover, LPA induced activation of RhoA in MSCs, and pretreatment of the cells with Y27632, a Rho kinase inhibitor, markedly inhibited the LPA‐induced migration. In addition, LPA and malignant ascites increased intracellular concentration of calcium in MSCs, and Ki16425 completely inhibited the elevation of intracellular calcium. These results suggest that LPA is a crucial component of the malignant ascites which induce the migration of MSCs and elevation of intracellular calcium. J. Cell. Biochem. 104: 499–510, 2008.

Lysophosphatidic acid induces cell migration through the selective activation of Akt1

Akt plays pivotal roles in many physiological responses including growth, proliferation, survival, metabolism, and migration. In the current studies, we have evaluated the isoform-specific role of akt in lysophosphatidic acid (LPA)-induced cell migration. Ascites from ovarian cancer patients (AOCP) induced mouse embryo fibroblast (MEF) cell migration in a dose-dependent manner. On the other hand, ascites from liver cirrhosis patients (ALCP) did not induce MEF cell migration. AOCP-induced MEF cell migration was completely blocked by pre-treatment of cells with LPA receptor antagonist, Ki16425. Both LPA- and AOCP-induced MEF cell migration was completely attenuated by PI3K inhibitor, LY294002. Furthermore, cells lacking Akt1 displayed defect in LPA-induced cell migration. Re-expression of Akt1 in DKO (Akt1-/-Akt2-/-) cells restored LPA-induced cell migration, whereas re-expression of Akt2 in DKO cells could not restore the LPA-induced cell migration. Finally, Akt1 was selectively phosphorylated by LPA and AOCP stimulation. These results suggest that LPA is a major factor responsible for AOCP-induced cell migration and signaling specificity of Akt1 may dictate LPA-induced cell migration.

Silibinin polarizes Th1/Th2 immune responses through the inhibition of immunostimulatory function of dendritic cells.

Silibinin is the primary active compound in silymarin. It has been demonstrated to exert anti‐carcinogenic effects and hepato‐protective effects. However, the effects of silibinin on the maturation and immunostimulatory activities exhibited by dendritic cells (DCs) remain, for the most part, unknown. In this study, we have attempted to determine whether silibinin can influence surface molecule expression, dextran uptake, cytokine production, capacity to induce T‐cell differentiation, and the signaling pathways underlying these phenomena in murine bone marrow‐derived DCs. Silibinin was shown to significantly suppress the expression of CD80, CD86, MHC class I, and MHC class II in the DCs, and was also associated with impairments of LPS‐induced IL‐12 expression in the DCs. Silibinin‐treated DCs proved highly efficient with regard to Ag capture via mannose receptor‐mediated endocytosis. Silibinin also inhibited the LPS‐induced activation of MAPKs and the nuclear translocation of the NF‐κB p65 subunit. Additionally, silibinin‐treated DCs evidenced an impaired induction of Th1 response, and a normal cell‐mediated immune response. These findings provide new insight into the immunopharmacological functions of silibinin, especially with regard to their impact on the DCs. These findings expand our current understanding of the immunopharmacological functions of silibinin, and may prove useful in the development of therapeutic adjuvants for acute and chronic DC‐associated diseases. J. Cell. Physiol. 210: 385–397, 2007.

Lysophosphatidic acid-induced expression of periostin in stromal cells: prognoistic relevance of periostin expression in epithelial ovarian cancer

Lysophosphatidic acid (LPA) is a bioactive lipid crucial for the initiation and progression of ovarian cancer. Identification of LPA‐induced biomarkers is necessary for predicting prognosis of ovarian cancer patients. Here we report periostin, an extracellular matrix protein, as an LPA‐induced protein in stromal cells and as a prognostic marker in patients with epithelial ovarian cancer (EOC). In human EOC tissues, periostin was mainly expressed in cancer‐associated stromal fibroblasts, but not in cancer cells. The expression levels of periostin highly correlated with poor survival and tumor recurrence of ovarian cancer patients. Treatment of human adipose tissue‐derived stromal cells with LPA or conditioned media from human ovarian adenocarcinoma cell lines, such as SK‐OV‐3 and OVCAR‐3, induced expression of periostin. The periostin expression induced by cancer‐conditioned media was abrogated by silencing of the LPA receptor 1 expression using small hairpin RNA lentivirus. Recombinant periostin stimulated adhesion and invasion of SK‐OV‐3 human ovarian adenocarcinoma cells and induced expression of matrix metalloprotease‐2 in the cancer cells. These results suggest that LPA is associated with the expression of periostin in cancer‐associated fibroblasts of EOC.

Significance of E2F-1 overexpression in epithelial ovarian cancer

E2F-1 is a downstream regulator of the Rb pathway and is a transcription factor that plays a key role in the control of cell cycle progression. Deregulation of E2F-1 expression and Rb pathway is involved in carcinogenesis. The aim of this study was to evaluate E2F-1 expression and Rb pathway alteration and to elucidate their correlation with clinical and pathologic parameters in epithelial ovarian cancer (EOC). We investigated overexpression of E2F-1 and alterations of p16INK4a, cyclin D1, CDK4, and pRb using immunohistochemistry and tissue microarray methods in 72 EOC patients. Overexpression of E2F-1 was detected in 45.8% of samples. The overall abnormal expression frequencies of p16INK4a, cyclin D1, CDK4, and pRb were 33.3%, 11.1%, 12.5%, and 38.9%, respectively. E2F-1 overexpression was not associated with alteration of the Rb pathway. E2F-1 overexpression was correlated with FIGO stage, histologic grade, and mitotic index; it was a valuable prognostic variable along with FIGO stage in the multivariated analysis. The results suggest that E2F-1 has a growth-promoting effect in EOC and that E2F-1 overexpression may provide a useful prognostic indicator for EOC.

TAZ Mediates Lysophosphatidic Acid-Induced Migration and Proliferation of Epithelial Ovarian Cancer Cells

Background: Transcriptional co-activator with PDZ-binding motif (TAZ), a downstream effector of the Hippo pathway, has been reported to regulate organ size, tissue homeostasis, and tumorigenesis by acting as a transcriptional co-activator. Lysophosphatidic acid (LPA) is a bioactive lipid implicated in tumorigenesis and metastasis of ovarian cancer through activation of G protein-coupled receptors. However, the involvement of TAZ in LPA-induced tumorigenesis of ovarian cancer has not been elucidated. Methods: In order to demonstrate the role of TAZ in LPA-stimulated tumorigenesis, the effects of LPA on TAZ expression and cell migration were determined by Western blotting and chemotaxis analyses in R182 human epithelial ovarian cancer cells. Results and Conclusion: Treatment of R182 cells with the LPA receptor inhibitor Ki16425 blocked LPA-induced cell migration. In addition, transfection of R182 cells with small interfering RNA specific for LPA receptor 1 resulted in abrogation of LPA-stimulated cell migration. LPA induced phosphorylation of ERK and p38 MAP kinase in R182 cells and pretreatment of cells with the MEK-ERK pathway inhibitor U0126, but not the p38 MAPK inhibitor SB202190, resulted in abrogation of LPA-induced cell migration. Pretreatment of R182 cells with U0126 attenuated LPA-induced mRNA levels of TAZ and its transcriptional target genes, such as CTGF and CYR61, without affecting phosphorylation level of YAP. These results suggest that MEK-ERK pathway plays a key role in LPA-induced cell migration and mRNA expression of TAZ in R182 cells, without affecting stability of TAZ protein. In addition, small interfering RNA-mediated silencing of TAZ expression attenuated LPA-stimulated migration of R182 cells. These results suggest that TAZ plays a key role in LPA-stimulated migration of epithelial ovarian cancer cells.

FOXP1 functions as an oncogene in promoting cancer stem cell-like characteristics in ovarian cancer cells

Ovarian cancer has the highest mortality rate of all gynecological cancers with a high recurrence rate. It is important to understand the nature of recurring cancer cells to terminally eliminate ovarian cancer. The winged helix transcription factor Forkhead box P1 (FOXP1) has been reported to function as either oncogene or tumor-suppressor in various cancers. In the current study, we show that FOXP1 promotes cancer stem cell-like characteristics in ovarian cancer cells. Knockdown of FOXP1 expression in A2780 or SKOV3 ovarian cancer cells decreased spheroid formation, expression of stemness-related genes and epithelial to mesenchymal transition-related genes, cell migration, and resistance to Paclitaxel or Cisplatin treatment, whereas overexpression of FOXP1 in A2780 or SKOV3 ovarian cancer cells increased spheroid formation, expression of stemness-related genes and epithelial to mesenchymal transition-related genes, cell migration, and resistance to Paclitaxel or Cisplatin treatment. In addition, overexpression of FOXP1 increased promoter activity of ABCG2, OCT4, NANOG, and SOX2, among which the increases in ABCG2, OCT4, and SOX2 promoter activity were dependent on the presence of FOXP1-binding site. In xenotransplantation of A2780 ovarian cancer cells into nude mice, knockdown of FOXP1 expression significantly decreased tumor size. These results strongly suggest FOXP1 functions as an oncogene by promoting cancer stem cell-like characteristics in ovarian cancer cells. Targeting FOXP1 may provide a novel therapeutic opportunity for developing a relapse-free treatment for ovarian cancer patients.

Proteomic Identification of ADAM12 as a Regulator for TGF-β1-Induced Differentiation of Human Mesenchymal Stem Cells to Smooth Muscle Cells

Background Transforming growth factor-β1 (TGF-β1) induces the differentiation of human adipose tissue-derived mesenchymal stem cells (hASCs) into smooth muscle cells. Lipid rafts are cholesterol-rich microdomains in cell membranes that reportedly play a key role in receptor-mediated signal transduction and cellular responses. In order to clarify whether lipid rafts are involved in TGF-β1-induced differentiation of hASCs into smooth muscle cells, we analyzed the lipid raft proteome of hASCs. Methods and Results Pretreatment of hASCs with the lipid raft disruptor methyl-β-cyclodextrin abrogated TGF-β1-induced expression of α-smooth muscle actin, a smooth muscle cell marker, suggesting a pivotal role of lipid rafts in TGF-β1-induced differentiation of hASCs to smooth muscle cells. Sucrose density gradient centrifugation along with a shotgun proteomic strategy using liquid chromatography-tandem mass spectrometry identified 1002 individual proteins as the lipid raft proteome, and 242 of these were induced by TGF-β1 treatment. ADAM12, a disintegrin and metalloproteases family member, was identified as the most highly up-regulated protein in response to TGF-β1 treatment. TGF-β1 treatment of hASCs stimulated the production of both ADAM12 protein and mRNA. Silencing of endogenous ADAM12 expression using lentiviral small hairpin RNA or small interfering RNA abrogated the TGF-β1-induced differentiation of hASCs into smooth muscle cells. Conclusions These results suggest a pivotal role for lipid raft-associated ADAM12 in the TGF-β1-induced differentiation of hASCs into smooth muscle cells.

Decreased expressions of vascular endothelial growth factor and visfatin in the placental bed of pregnancies complicated by preeclampsia

Aim:  The aim of the present study was to examine the expression of vascular endothelial growth factor (VEGF) and visfatin in the third trimester placental bed of pregnancies with and without preeclampsia (PE).