ABSTRACT
Lysophosphatidic acid (LPA) signaling via LPA receptors (LPA1 to LPA6) exhibits a variety of malignant properties in cancer cells. Intracellular ATP depletion leads to the development of necrosis and apoptosis. The present study aimed to evaluate the effects of LPA receptor-mediated signaling on the regulation of cancer cell functions associated with ATP reduction. Long-term ethidium bromide (EtBr) treated (MG63-EtBr) cells were established from osteosarcoma MG-63 cells. The intracellular ATP levels of MG63-EtBr cells were significantly lower than that of MG-63 cells. LPAR2, LPAR3, LPAR4 and LPAR6 gene expressions were elevated in MG63-EtBr cells. The cell motile and invasive activities of MG63-EtBr cells were markedly higher than those of MG-63 cells. The cell motile activity of MG-63 cells was increased by LPA4 and LPA6 knockdowns. In cell survival assay, cells were treated with cisplatin (CDDP) every 24 h for 3 days. The cell survival to CDDP of MG63-EtBr cells was lower than that of MG-63 cells. LPA2 knockdown decreased the cell survival to CDDP of MG-63 cells. The cell survival to CDDP of MG-63 cells was inhibited by (2 S)-OMPT (LPA3 agonist). Moreover, the cell survival to CDDP of MG-63 cells was enhanced by LPA4 and LPA6 knockdowns. These results indicate that LPA signaling via LPA receptors is involved in the regulation of cellular functions associated with ATP reduction in MG-63 cells treated with EtBr.
Subject(s)
Bone Neoplasms , Osteosarcoma , Adenosine Triphosphate/pharmacology , Bone Neoplasms/drug therapy , Bone Neoplasms/genetics , Cell Movement , Ethidium/pharmacology , Gene Expression Regulation, Neoplastic , Humans , Lysophospholipids/metabolism , Osteosarcoma/drug therapy , Osteosarcoma/genetics , Receptors, Lysophosphatidic Acid/genetics , Receptors, Lysophosphatidic Acid/metabolismABSTRACT
Lysophosphatidic acid (LPA) signaling via LPA receptors (LPA1 to LPA6) exhibits a variety of biological responses. In tumor microenvironment, endothelial cells promote cancer cell functions. In this study, we investigated the roles of endothelial cells in the regulation of cell motile activity via LPA2 and LPA3 in human osteosarcoma MG-63 cells. In cell motility assay, the cell motile activity of MG-63 cells was markedly increased by the supernatants of endothelial F2 cells. MG-63 cell motility elevated by the supernatants was enhanced by GRI-977143 (LPA2 agonist) and reduced by (2S)-OMPT (LPA3 agonist). LPAR2 and LPAR3 expressions were increased in highly migratory MG63-CR7(F2) cells, which were generated from MG-63 cells by co-culture with F2 cell supernatants. MG63-CR7(F2) cell motility was stimulated by LPA treatment. In the presence of F2 cell supernatants, MG63-CR7(F2) cell motility was markedly enhanced by GRI-977143 and suppressed by (2S)-OMPT. Autotaxin (ATX) enzymatically converts lysophosphatidylcholine (LPC) to LPA. ATX expression was higher in MG63-CR(F2) cells than in MG-63 cells. MG63-CR7(F2) cell motility was markedly increased by LPC in comparison with MG-63 cells. In addition, MG63-CR(F2) cell motility was significantly stimulated by the supernatants of LPC treated F2 cells. The present results suggest that the activation of LPA signaling via LPA2 and LPA3 by endothelial cells is involved in the modulation of cell motile activity of MG-63 cells.
Subject(s)
Bone Neoplasms/pathology , Cell Movement , Endothelial Cells/pathology , Gene Expression Regulation, Neoplastic , Osteosarcoma/pathology , Receptors, Lysophosphatidic Acid/metabolism , Apoptosis , Bone Neoplasms/genetics , Bone Neoplasms/metabolism , Cell Proliferation , Endothelial Cells/metabolism , Humans , Lysophospholipids/metabolism , Osteosarcoma/genetics , Osteosarcoma/metabolism , Receptors, Lysophosphatidic Acid/genetics , Signal Transduction , Tumor Cells, CulturedABSTRACT
Lysophosphatidic acid (LPA) is a simple physiological lipid and exhibits several biological functions by binding to G-protein-coupled LPA receptors (LPA receptor-1 (LPA1) to LPA6). The present study aimed to evaluate whether LPA signaling via LPA2 and LPA5 is involved in the chemoresistance to anticancer drugs in colon cancer DLD1 cells. In cell survival assay, cells were treated with fluorouracil (5-FU) every 24 h for 2 days. The cell survival rate to 5-FU of DLD1 cells was significantly decreased by LPA treatment. In the presence of LPA, the cell survival rate to 5-FU was significantly elevated by LPA5 knockdown. Before initiation of the cell survival assay, cells were pretreated with an LPA2 agonist, GRI-977143. The cell survival rate to 5-FU was markedly increased in DLD1 cells treated with GRI-977143. In the presence of GRI-977143, the elevated cell survival rate of DLD1 cells was reduced by LPA2 knockdown. To assess the effects of LPA2 and LPA5 on the enhancement of chemoresistance, long-term 5-FU treated (DLD-5FU) cells were generated from DLD1 cells. The cell survival rate to 5-FU of DLD-5FU cells were significantly elevated by LPA5 knockdown. GRI-977143 treatment increased the cell survival rate to 5-FU of DLD-5FU cells. These results suggest that LPA2 promotes and LPA5 suppresses the acquisition of chemoresistance in colon cancer cells treated with anticancer drugs.
Subject(s)
Colonic Neoplasms/drug therapy , Fluorouracil/administration & dosage , Receptors, Lysophosphatidic Acid/genetics , Cell Line, Tumor , Cell Survival/drug effects , Colonic Neoplasms/genetics , Colonic Neoplasms/pathology , Drug Resistance, Neoplasm/genetics , Gene Knockdown Techniques , Humans , Receptors, Lysophosphatidic Acid/agonists , Receptors, Lysophosphatidic Acid/antagonists & inhibitorsABSTRACT
Lysophosphatidic acid (LPA) through six subtypes of G protein-coupled LPA receptors (LPA1 to LPA6) mediates a variety of cancer cell functions. The aim of this study was to evaluate the cooperative effects of G12/13 and Gi proteins through LPA2 on cancer cell survival to cisplatin (CDDP). In cell survival assay, cells were treated with CDDP every 24 h for 2 days. The long-term CDDP treated (HT-CDDP) cells established from fibrosarcoma HT1080 cells were pretreated with an LPA2 agonist, GRI-977143. The cell survival rate to CDDP of HT-CDDP cells was significantly increased by GRI-977143. The elevated cell survival to CDDP was suppressed by LPA2 knockdown. Since G12/13 protein stimulates Rho-mediated signaling, RhoA and RhoC knockdown cells were generated from HT1080 cells (HT1080-RhoA and HT1080-RhoC cells, respectively). In the presence of GRI-977143, HT1080-RhoA and HT1080-RhoC cells showed the low cell survival rates to CDDP. On the other hand, Gi protein inhibits adenylyl cyclase (AC) activity. Before cell survival assay, cells were treated with a Gi protein inhibitor, pertussis toxin (PTX) for 24 h. The cell survival rate to CDDP of HT1080 cells was significantly reduced by PTX. Furthermore, when HT1080-RhoA and HT1080-RhoC cells were pretreated with PTX, the cell survival rates to CDDP of both cells were markedly inhibited by PTX. The present results suggest that cooperation of G12/13 and Gi proteins activated by LPA2 enhances the cell survival of HT1080 cells treated with CDDP.
Subject(s)
Cell Survival/drug effects , Cisplatin/pharmacology , Fibrosarcoma/drug therapy , GTP-Binding Protein alpha Subunits, G12-G13/metabolism , GTP-Binding Protein alpha Subunits, Gi-Go/metabolism , Receptors, Lysophosphatidic Acid/metabolism , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cell Survival/physiology , Fibrosarcoma/metabolism , Fibrosarcoma/pathology , Gene Expression Regulation, Neoplastic/drug effects , Gene Knockdown Techniques , Humans , Models, Biological , Receptors, Lysophosphatidic Acid/antagonists & inhibitors , Receptors, Lysophosphatidic Acid/genetics , Signal Transduction/drug effects , rhoA GTP-Binding Protein/metabolism , rhoC GTP-Binding Protein/metabolismABSTRACT
Lysophosphatidic acid (LPA) signaling through LPA receptors (LPA1 to LPA6) regulates a variety of malignant properties in cancer cells. Recently, we show that LPA2 expression is elevated by long-term cisplatin (CDDP) treatment in melanoma A375 cells. In the present study, we investigated whether LPA2-mediated signaling is involved in the modulation of chemoresistance in A375 cells. In cell survival assay, cells were treated with CDDP and dacarbazine (DTIC) every 24 h for 2 days. The cell survival rates to CDDP and DTIC were markedly increased by an LPA2 agonist, GRI-977143. To validate the effects of LPA2 on cell survival, LPA2 knockdown cells were generated from A375 cells. The cell survival rates elevated by GRI-977143 were suppressed by LPA2 knockdown. To evaluate the roles of LPA2-mediated signaling in cell survival, cells were pretreated with a Gi protein inhibitor, pertussis toxin (PTX). In the presence of GRI-977143, the cell survival rates to CDDP and DTIC were significantly lower in PTX-treated cells than in untreated cells. In addition, pretreatment of an adenylyl cyclase inhibitor, SQ22536, increased the cell survival of A375 cells treated with CDDP and DTIC. These results suggest that LPA2-mediated signaling plays an important role in the enhancement of chemoresistance of A375 cells treated with anticancer drugs.
Subject(s)
Antineoplastic Agents/pharmacology , Cisplatin/pharmacology , Dacarbazine/pharmacology , Drug Resistance, Neoplasm/genetics , Lysophospholipids/metabolism , Melanoma/metabolism , Receptors, Lysophosphatidic Acid/metabolism , Cell Line, Tumor , Cell Survival/drug effects , Gene Knockdown Techniques , Humans , Lysophospholipids/agonists , Lysophospholipids/genetics , Melanoma/genetics , Pertussis Toxin/toxicity , Receptors, Lysophosphatidic Acid/genetics , Signal Transduction/drug effects , Signal Transduction/geneticsABSTRACT
Lysophosphatidic acid (LPA) mediates a variety of biological functions via the binding of G protein-coupled LPA receptors (LPA receptor-1 (LPA1) to LPA6). This study aimed to investigate the roles of LPA2 and LPA3 in the modulation of chemoresistance to anticancer drug in lung cancer A549 cells. In cell survival assay, cells were treated with cisplatin (CDDP) every 24 h for 2 days. The cell survival rate to CDDP of A549 cells was significantly elevated by an LPA2 agonist, GRI-977143. To evaluate the roles of LPA2-mediated signaling in cell survival during tumor progression, highly migratory (A549-R10) cells were generated from A549 cells. In the presence of GRI-977143, the cell survival rate to CDDP of A549-R10 cells were markedly higher than that of A549 cells, correlating with LPAR2 expression level. Moreover, to assess the effects of long-term anticancer drug treatment on cell survival, the long-term CDDP treated (A549-CDDP) cells were established from A549 cells. The cell survival rate to CDDP of A549-CDDP cells was elevated by GRI-977143. Since LPAR3 expression level was significantly higher in A549-CDDP cells than in A549 cells, we investigated the roles of LPA3 in the cell survival to CDDP of A549 cells, using an LPA3 agonist, 1-oleoyl-2-methyl-sn-glycero-3-phosphothionate ((2S)-OMPT). The cell survival rate to CDDP of A549 cells was significantly reduced by (2S)-OMPT treatment. In the presence of (2S)-OMPT, the cell survival rate to CDDP of A549 cells was elevated by LPA3 knockdown. These results suggest that LPA signaling via LPA2 and LPA3 is involved in the regulation of chemoresistance in A549 cells treated with CDDP.
Subject(s)
Antineoplastic Agents/therapeutic use , Cisplatin/therapeutic use , Drug Resistance, Neoplasm , Lung Neoplasms/drug therapy , Receptors, Lysophosphatidic Acid/physiology , A549 Cells , Cell Survival , Gene Expression Regulation, Neoplastic , HumansABSTRACT
Free fatty acid receptor 1 (FFA1) and FFA4 belong to a family of free fatty acid (FFA) receptors. FFA1- and FFA4-mediated signaling regulates a variety of malignant properties in cancer cells. It is known that stromal cells in the tumor microenvironment promote tumor progression. In the present study, to assess the roles of FFA1 and FFA4 in cellular functions modulated by endothelial cells, highly migratory MG63-CR7(F2) cells were generated from osteosarcoma MG-63 cells, using endothelial F2 cell supernatants. Expression levels of FFAR1 and FFAR4 genes in MG63-CR7(F2) cells were significantly higher than those of MG-63 cells. In cell survival assay, cells were treated with cisplatin (CDDP) every 24 h for 2 days. The cell survival rate of MG-63 cells was significantly elevated by an FFA1 agonist TUG-770 as well as an FFA4 agonist TUG-891. Moreover, the cell survival rate of MG63-CR7(F2) cells was higher than that of MG-63 cells in the presence of TUG-770 or TUG-891, correlating with FFAR1 and FFAR4 expression levels. To validate the effects of FFA1 and FFA4 on cell survival to CDDP, FFA1 and FFA4 knockdown cell were generated from MG-63 cells. The cell survival rate of MG-63 cells was markedly inhibited by FFA1 or FFA4 knockdown. These results suggest that FFA1 and FFA4 may play an important role in the modulation of cellular functions by endothelial cells in osteosarcoma cells.
Subject(s)
Carcinogenesis/genetics , Osteosarcoma/drug therapy , Receptors, G-Protein-Coupled/genetics , Cell Line, Tumor , Cell Survival/drug effects , Cisplatin/pharmacology , Endothelial Cells/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Humans , Osteosarcoma/genetics , Osteosarcoma/pathology , Tumor Microenvironment/drug effectsABSTRACT
Lysophosphatidic acid (LPA) signaling via LPA receptors (LPA1 to LPA6) regulates a variety of malignant properties of cancer cells. It is known that endothelial cells promote tumor progression and chemoresistance. The present study aimed to investigate the roles of LPA5 in cellular functions modulated by endothelial cells and anticancer drug in osteosarcoma cells. Human osteosarcoma MG-63 cells were maintained in endothelial F2 cell supernatants. After culturing for 3 months, MG63-F2 cells were established. LPAR5 expression level in MG63-F2 cells was significantly elevated, compared with MG-63 cells. The cell motile activity of MG63-F2 cells was markedly higher than that of MG-63 cells. To validate the effects of LPA5 on cell motile activity, LPA5 knockdown cells were generated from MG-63 cells. The cell motile activity of MG-63 cells was inhibited by LPA5 knockdown. The cell survival to cisplatin (CDDP) was reduced in MG-63 cells treated with LPA. In the presence of LPA, the cell survival rate was significantly lower in MG63-F2 cells than MG-63 cells, correlating with LPAR5 expression. LPA5 knockdown cells indicated the high cell survival rate to CDDP. Moreover, LPAR5 expression level was increased in the long-term CDDP treated MG63-C cells. The cell survival to CDDP of MG63-C cells was enhanced by LPA5 knockdown. These results suggest that cellular functions are regulated through LPA5-mediatd signaling induced by endothelial cells and CDDP in MG-63 cells.
Subject(s)
Endothelial Cells/metabolism , Osteosarcoma/metabolism , Receptors, Lysophosphatidic Acid/metabolism , Antineoplastic Agents/pharmacology , Cell Death/drug effects , Cell Movement/drug effects , Cisplatin/analogs & derivatives , Cisplatin/pharmacology , Culture Media, Conditioned/pharmacology , Humans , Lysophospholipids/metabolism , Receptors, Lysophosphatidic Acid/genetics , Signal Transduction , Tumor Cells, CulturedABSTRACT
Cell migration is closely involved in cancer cell invasion into surrounding tissue and metastasis to the distant organs. It is crucial for understanding the molecular mechanisms that regulate cell migration in cancer cells. The aim of this study is to establish a rapid induction method of highly migratory cells from cancer cells. Osteosarcoma MG-63 and colon cancer DLD1 cells were seeded at 1 × 105 cells in 6-well plates. After 10 min, unattached cells were washed off three times with PBS. The cells which remained attached on the bottom of plates were cultured in DMEM containing 10% FBS. When the cells reached approximately 80% confluence, cells were harvested using trypsin/EDTA. The harvested cells were seeded in other 6-well plates and incubated for 10 min. The unattached cells were washed off and attached cells were further cultured. By repeating this procedure 11-12 times for 2 months, highly migratory MG63-A12 and DLD-A11 cells were obtained from MG-63 and DLD1 cells, respectively. In cell motility assay, the cell motile activities of MG63-A12 and DLD-A11 cells was 10.3 and 13.7 times higher than those of the parental cells, respectively. This procedure is useful to generate highly migratory cells for investigating cellular functions during tumor progression in cancer cells.
Subject(s)
Cell Movement , Osteosarcoma/pathology , Cell Line, Tumor , Cell Proliferation , Humans , Receptors, Cell Surface/metabolismABSTRACT
Lysophosphatidic acid (LPA) signaling via LPA receptors (LPA1 to LPA6) contributes to the promotion of malignant potency in cancer cells. The cell motile activity are stimulated through the induction of LPA5 in melanoma cells treated with anticancer drugs. The present study aimed to investigate whether LPA signaling via LPA5 regulates chemoresistance in melanoma A375â¯cells. Cells were treated with cisplatin (CDDP) or dacarbazine (DTIC) every 24â¯h for 2 days. CDDP and DTIC treatment increased LPAR5 expressions. The cell survival rates of A375â¯cells treated with CDDP and DTIC were significantly decreased by LPA. In addition, LPAR5 expression was markedly elevated in long-term CDDP treated (A375-CDDP) cells. LPA decreased the cell survival rate of A375-CDDP cells treated with CDDP. To evaluate the roles of LPA5 in chemoresistance during tumor progression, highly migratory (A375-R11) cells were established from A375â¯cells. LPAR5 expression level was significantly lower in A375-R11â¯cells than in A375â¯cells. The cell survival rates of A375-R11â¯cells treated with CDDP and DTIC were increased, compared with A375â¯cells. Moreover, we generated LPA5 knockdown cells from A375â¯cells. The cell survival rates of A375â¯cells treated with CDDP and DTIC were significantly elevated by LPA5 knockdown. These results suggest that LPA signaling via LPA5 is involved in the modulation of chemoresistance in melanoma A375â¯cells.
Subject(s)
Antineoplastic Agents/pharmacology , Cisplatin/pharmacology , Dacarbazine/pharmacology , Lysophospholipids/metabolism , Melanoma/drug therapy , Receptors, Lysophosphatidic Acid/metabolism , Cell Line, Tumor , Cell Movement/drug effects , Cell Survival/drug effects , Drug Resistance, Neoplasm , Gene Expression Regulation, Neoplastic/drug effects , Gene Knockdown Techniques , Humans , Melanoma/genetics , Melanoma/metabolism , Receptors, Lysophosphatidic Acid/genetics , Signal Transduction/drug effectsABSTRACT
Lysophosphatidic acid (LPA) is a simple biological lipid and mediates several biological functions with LPA receptors (LPA1 to LPA6). In the present study, to assess whether LPA receptors promote cell-invasive activity of pancreatic cancer cells, highly invasion PANC-R9 cells were established from PANC-1 cells, using Matrigel-coated Cell Culture Insert. The cell-invasive activity of PANC-R9 cells was shown to be approximately 15 times higher than that of PANC-1 cells. LPAR1 expression level was markedly elevated in PANC-R9 cells in comparison with PANC-1 cells, while LPAR3 expression level was reduced. The cell-invasive activity of PANC-R9 cells was enhanced by LPA, but LPA had no impact on PANC-1 cell invasion. Before initiation of the cell invasion assay, PANC-R9 cells were pretreated with dioctanoylglycerol pyrophosphate (DGPP), an antagonist of LPA1/LPA3. The invasive activity of PANC-R9 cells was markedly suppressed by DGPP. Autotaxin (ATX) is a key enzyme that catalyzes the conversion of lysophosphatidylcholine (LPC) to LPA. ATX expression level was elevated in PANC-R9 cells compared with PANC-1 cells. In the presence of LPC, the cell motile activity of PANC-R9 cells was markedly stimulated. In contrast, LPC did not affect the cell motile activity of PANC-1 cells. PANC-R9 cell motility was inhibited by an ATX inhibitor, PF-8380. These results suggest that LPA signaling via LPA1 is a potent molecular target for the regulation of tumor progression in PANC-1 cells.
Subject(s)
Lysophosphatidylcholines/metabolism , Pancreatic Neoplasms/drug therapy , Phosphoric Diester Hydrolases/genetics , Receptors, Lysophosphatidic Acid/genetics , Benzoxazoles/pharmacology , Cell Line, Tumor , Cell Movement/drug effects , Cell Proliferation/drug effects , Gene Expression Regulation, Neoplastic , Humans , Lysophosphatidylcholines/genetics , Neoplasm Invasiveness/genetics , Pancreatic Neoplasms/genetics , Phosphatidic Acids/metabolism , Piperazines/pharmacology , Receptors, Lysophosphatidic Acid/metabolism , Signal Transduction/drug effectsABSTRACT
Lysophosphatidic acid (LPA) receptors (LPA1 to LPA6) regulate a variety of malignant properties in cancer cells. In the present study, we investigated the roles of LPA receptors in the promotion of cellular functions during tumor progression in fibrosarcoma cells. To obtain long-term anticancer drug treated cells, human fibrosarcoma HT1080â¯cells were treated with methotrexate (MTX) and cisplatin (CDDP) for 6 months. LPAR2 and LPAR5 expressions were significantly higher in MTX-treated (HT-MTX) cells than in HT1080â¯cells. The cell motile and invasive activities of HT-MTX cells were significantly elevated compared with HT1080â¯cells. Although LPAR5 expression was increased in MTX and CDDP treated (HT-M-C) cells, no change of LPAR2 expression was observed. The cell motile and invasive activities of HT-M-C cells were lower than those of HT1080â¯cells. Moreover, to evaluate whether LPA receptors promote cell invasive activity, highly invasion (HT1080-M6) cells were established from HT1080â¯cells. The cell invasive activity of HT1080-M6 cells was approximately 4.5 times higher than HT1080â¯cell invasion. LPAR2 expression was markedly elevated in HT1080-M6 cells compared with HT1080â¯cells. The high cell invasion activity of HT1080-M6 cells was significantly suppressed by an antagonist of LPA2, H2L5186303. These results suggest that LPA2 acts as a key regulator of malignant properties in HT1080â¯cells.
Subject(s)
Fibroblasts/metabolism , Gene Expression Regulation, Neoplastic , Receptors, Lysophosphatidic Acid/genetics , Antineoplastic Agents/pharmacology , Benzene Derivatives/pharmacology , Cell Line, Tumor , Cell Movement/drug effects , Cell Proliferation/drug effects , Cisplatin/pharmacology , Drug Resistance, Neoplasm/genetics , Fibroblasts/drug effects , Fibroblasts/pathology , Humans , Methotrexate/pharmacology , Receptors, Lysophosphatidic Acid/antagonists & inhibitors , Receptors, Lysophosphatidic Acid/metabolism , Signal TransductionABSTRACT
Free fatty acid (FFA) receptors belong to a member of G-protein-coupled receptors. GPCR 120 (GPR120) and GPR40 are identified as FFA receptors and activated via the binding of long- and medium-chain FFAs. The aim of this study was to assess the effects of GPR120 and GPR40 on cell motility and growth in breast cancer cells treated with tamoxifen (TAM). MCF-7 cells were continuously treated with TAM for approximately 6 months. The expression level of GPR40 gene was markedly higher in the long-term TAM treated (MCF-TAM) cells than in MCF-7 cells. In cell motility assay, MCF-TAM cells indicated the high cell motile activity, compared with MCF-7 cells. The cell motile activity of MCF-TAM cells was suppressed by a selective GPR40 antagonist, GW1100. To evaluate the effects of GPR40 on cell growth activity under estrogen-free conditions, cells were maintained in serum-free DMEM without phenol red for 2 days. In estrogen-free conditioned medium, the cell growth rate of MCF-TAM cells was significantly higher than that of MCF-7 cells. In addition, treatment of GW1100 reduced the cell growth rate of MCF-TAM cells. These results suggest that the cell motile and growth activities may be positively regulated through the induction of GPR40 by the long-term TAM treatment in MCF-7 cells.
Subject(s)
Breast Neoplasms/drug therapy , Cell Proliferation/drug effects , Receptors, G-Protein-Coupled/genetics , Tamoxifen/pharmacology , Benzoates/pharmacology , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Cell Movement/drug effects , Female , Gene Expression Regulation, Neoplastic/drug effects , Humans , MCF-7 Cells , Pyrimidines/pharmacologyABSTRACT
Lysophosphatidic acid (LPA) signaling via G protein-coupled LPA receptors mediates various biological effects in cancer cells. This study aimed to investigate the roles of LPA receptors in the regulation of cellular functions during tumor progression in osteosarcoma cells. Long-term cisplatin (CDDP)-treated MG63-C and MG63-R7-C cells were generated from osteosarcoma MG-63 and highly-migratory MG63-R7 cells, respectively. LPAR2 and LPAR3 expression levels were significantly higher in MG63-C cells than in MG-63 cells, while LPAR1 expression was reduced. MG63-C cells were highly motile, compared with MG-63 cells. MG63-C cell motility was suppressed by LPA2 knockdown and enhanced by the LPA1/LPA3 antagonist, dioctanoylglycerol pyrophosphate. LPAR2 and LPAR3 expression levels were significantly elevated in MG63-R7-C cells in comparison with MG63-R7 cells. MG63-R7-C cells were found to be highly invasive, correlating with metalloproteinase-2 activation. MG63-R7-C cells formed large colonies, whereas colony formation was absent from MG63-R7 cells. Notably, MG63-R7-C cell activities were inhibited by LPA2 knockdown. These results suggest that LPA signaling via LPA2 plays an important role in the acquisition of malignant properties during tumor progression in MG-63 cells.
Subject(s)
Lysophospholipids/metabolism , Osteosarcoma/etiology , Osteosarcoma/metabolism , Receptors, Lysophosphatidic Acid/metabolism , Cell Line, Tumor , Cell Movement , Cisplatin/pharmacology , Disease Progression , Gene Expression Regulation, Neoplastic/drug effects , Gene Knockdown Techniques , Humans , Neoplasm Invasiveness , Osteosarcoma/genetics , Phosphoric Diester Hydrolases/metabolism , Receptors, Lysophosphatidic Acid/antagonists & inhibitors , Receptors, Lysophosphatidic Acid/genetics , Signal Transduction , Tumor Stem Cell AssayABSTRACT
Free fatty acid receptor 1 (FFA1) and FFA4 mediate a variety of biological responses through binding of medium- and long-chain free fatty acids. The aim of this study was to investigate an involvement of FFA1 and FFA4 in the regulation of cellular functions during tumor progression in colon cancer cells. The long-term fluorouracil (5-FU) and cisplatin (CDDP) treated cells were generated from DLD1 cells (DLD-5FU and DLD-CDDP cells, respectively). FFAR1 expressions were lower in DLD-5FU and DLD-CDDP cells than in DLD1 cells. In contrast, DLD-5FU and DLD-CDDP cells showed the high FFAR4 expressions, compared with DLD1 cells. The cell motile activities of DLD-5FU and DLD-CDDP cells were reduced by GW9508 which is an agonist of FFA1 and FFA4. Moreover, GW1100, an antagonist of FFA1, inhibited the cell motile activities of DLD-5FU and DLD-CDDP cells. To evaluate whether FFA1 and FFA4 regulate the enhancement of cell motility, invasion and colony formation, highly migratory (hmDLD1) cells were established from DLD1 cells. FFAR1 expression was significantly higher in hmDLD1 cells than in DLD1 cells, but no change of FFAR4 expression was observed. The elevated cell motile and invasive activities and colony formation of hmDLD1 cells were suppressed by FFA1 inhibition. These results suggest that FFA1 and FFA4 are involved in the regulation of cellular functions during tumor progression in colon cancer DLD1 cells.
Subject(s)
Cell Movement/genetics , Colon/pathology , Colonic Neoplasms/pathology , Epithelial Cells/physiology , Intestinal Mucosa/pathology , Receptors, G-Protein-Coupled/physiology , Antineoplastic Agents/pharmacology , Benzoates/pharmacology , Cell Line, Tumor , Cell Movement/drug effects , Cisplatin/pharmacology , Colon/drug effects , Colon/metabolism , Colonic Neoplasms/genetics , Colonic Neoplasms/physiopathology , Disease Progression , Drug Resistance, Neoplasm/genetics , Epithelial Cells/drug effects , Epithelial Cells/pathology , Fluorouracil/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , Humans , Intestinal Mucosa/drug effects , Intestinal Mucosa/metabolism , Methylamines/pharmacology , Propionates/pharmacology , Pyrimidines/pharmacology , Receptors, G-Protein-Coupled/antagonists & inhibitorsABSTRACT
Lysophosphatidic acid (LPA) signaling through six subtypes of LPA receptors (LPA1 to LPA6) regulates a variety of biological responses in cancer cells. The aim of our study was to evaluate an involvement of LPA receptors in the activation of cell motility by phorbol ester and anticancer drug treatments in melanoma A375â¯cells. Cells were treated with 12-O-tetradecanoylphorbol- 13-acetate (TPA) and phorbol-12,13-dibutyrate (PDBu) for 3 days. The cell motile activity of TPA treated cells was significantly higher than that of PDBu treated cells, correlating with LPAR5 expression levels. LPA5 knockdown suppressed the high cell motile activity induced by TPA. To assess whether the cell motile activity of A375â¯cells is stimulated through LPA5 induced by anticancer drugs, the long-term cisplatin (CDDP) and dacarbazine (DTIC) treated cells were generated from A375â¯cells (A375-CDDP and A375-DTIC cells, respectively). The expression levels of LPA receptor genes were changed in A375-CDDP and A375-DTIC cells. In particular, CDDP and DTIC treatment markedly elevated LPAR5 expressions. The cell motile activities of A375-CDDP and A375-DTIC cells were significantly higher than that of untreated cells. These results suggest that the cell motile activity is regulated through the induction of LPA5 by phorbol ester and anticancer drug treatments in A375â¯cells.
Subject(s)
Antineoplastic Agents/administration & dosage , Cell Movement/drug effects , Melanoma/drug therapy , Melanoma/metabolism , Phorbol Esters/administration & dosage , Receptors, Lysophosphatidic Acid/metabolism , Skin Neoplasms/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Humans , Skin Neoplasms/drug therapyABSTRACT
Lysophosphatidic acid (LPA) is a simple physiological lipid and exhibits a variety of cellular responses via the activation of G protein-coupled transmembrane LPA receptors (LPA receptor-1 (LPA1) to LPA6). The aim of our study was to investigate effects of LPA receptors on soft agar colony formation in colon cancer cells treated with anticancer drugs. DLD1 cells were treated with fluorouracil (5-FU) or cisplatin (CDDP) for at least six months (DLD-5FU and DLD-CDDP cells, respectively). LPAR1 gene expression was markedly elevated in DLD-5FU cells. In contrast, DLD-CDDP cells showed the high expression of LPAR6 gene. In colony formation assay, DLD-5FU cells formed markedly large-sized colonies, while no colony formation was observed in DLD1 and DLD-CDDP cells. The large-sized colonies formed in DLD-5FU cells were suppressed by LPA1 knockdown. In contrast, LPA6 knockdown increased the size of colonies. In addition, DLD-5FU cells were further treated with CDDP for three months (DLD-C-F cells). DLD-CDDP cells were also treated with 5-FU (DLD-F-C cells). DLD-C-F cells formed large-sized colonies, but not DLD-F-C cells, correlating with LPAR1 and LPAR6 gene expression levels. These results suggest that LPA1 and LPA6 may regulate the colony formation activity in DLD1 cells treated with anticancer drugs.
