tretinoin and Stomach-Neoplasms

Gastric cancer (GC) is the third leading cause of cancer-related death worldwide with a five-year survival rate of around 25%, and 4% when diagnosed at a metastatic stage. Cancer stem cells (CSC) have recently been characterized as being responsible for resistance to radio/chemotherapies and metastasis formation, opening up perspectives for new targeted therapies. Those CSCs express biomarkers such as cluster of differentiation 44 (CD44) and display high aldehyde dehydrogenase activity that converts vitamin A-derived retinal into retinoic acids. All-trans retinoic acid (ATRA), which has pro-differentiating properties, has revolutionized the prognosis of acute promyelotic leukemia by increasing its remission rate from 15% to 85%. Recent studies have started to show that ATRA also has an anti-tumoral role on solid cancers such as GC. The purpose of this review is therefore to summarize the work that evaluated the effects of ATRA in GC and to evaluate whether its anti-cancerous action involves gastric CSCs targeting. It has been demonstrated that ATRA can block the cell cycle, enhance apoptosis, and decrease gastric CSCs properties in GC cell lines, tumorspheres, and patient-derived xenograft mice models. Therefore, retinoids and new synthetic retinoids seem to be a promising step forward in targeted therapy of gastric CSC in combination with existing chemotherapies. Future studies should probably focus on these points.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cell Cycle; Humans; Hyaluronan Receptors; Neoplastic Stem Cells; Retinoid X Receptors; Stomach; Stomach Neoplasms; Tretinoin

To investigate the efficacy of all-trans retinoic acid (ATRA) in human gastric dysplasia.. In this double-blind study, patients with precancerous gastric dysplasia with or without intestinal metaplasia (IM) received either conventional treatment consisting of omeprazole and sucralfate (control group) or conventional treatment plus ATRA. Gastric mucosal biopsies were performed before and after drug treatment and were analysed histologically; expression of retinoblastoma (Rb) protein and HER2 protein in gastric mucosa were measured using immunohistochemistry.. A total of 122 patients were included in the study, 63 in the ATRA group and 59 in the control group. In the ATRA group, dysplasia was attenuated in 43 out of 63 patients (68%) compared with 22 out of 59 patients (37%) in the control group; however, IM was not affected by treatment in either group. ATRA treatment was associated with significantly increased Rb expression and decreased HER2 expression in gastric mucosa.. The use of conventional therapy plus ATRA for gastric dysplasia was associated with improved efficacy compared with conventional therapy alone. It was also accompanied by increased Rb expression and decreased HER2 expression in gastric mucosa. The addition of ATRA to conventional therapy for gastritis may improve the prognosis of gastric dysplasia.

Topics: Adult; Aged; Double-Blind Method; Drug Therapy, Combination; Female; Gastric Mucosa; Humans; Intestines; Male; Metaplasia; Middle Aged; Omeprazole; Precancerous Conditions; Prognosis; Retinoblastoma Protein; Stomach; Stomach Neoplasms; Sucralfate; Treatment Outcome; Tretinoin

Gastric-cancer is a heterogeneous type of neoplastic disease and it lacks appropriate therapeutic options. There is an urgent need for the development of innovative pharmacological strategies, particularly in consideration of the potential stratified/personalized treatment of this tumor. All-Trans Retinoic-acid (ATRA) is one of the active metabolites of vitamin-A. This natural compound is the first example of clinically approved cyto-differentiating agent, being used in the treatment of acute promyelocytic leukemia. ATRA may have significant therapeutic potential also in the context of solid tumors, including gastric-cancer. The present study provides pre-clinical evidence supporting the use of ATRA in the treatment of gastric-cancer using high-throughput approaches.. We evaluated the anti-proliferative action of ATRA in 27 gastric-cancer cell-lines and tissue-slice cultures from 13 gastric-cancer patients. We performed RNA-sequencing studies in 13 cell-lines exposed to ATRA. We used these and the gastric-cancer RNA-sequencing data of the TCGA/CCLE datasets to conduct multiple computational analyses.. Profiling of our large panel of gastric-cancer cell-lines for their quantitative response to the anti-proliferative effects of ATRA indicate that approximately half of the cell-lines are characterized by sensitivity to the retinoid. The constitutive transcriptomic profiles of these cell-lines permitted the construction of a model consisting of 42 genes, whose expression correlates with ATRA-sensitivity.  The model predicts that 45% of the TCGA gastric-cancers are sensitive to ATRA. RNA-sequencing studies performed in retinoid-treated gastric-cancer cell-lines provide insights into the gene-networks underlying ATRA anti-tumor activity. In addition, our data demonstrate that ATRA exerts significant immune-modulatory effects, which seem to be largely controlled by IRF1 up-regulation. Finally, we provide evidence of a feed-back loop between IRF1 and DHRS3, another gene which is up-regulated by ATRA.. ATRA is endowed with significant therapeutic potential in the stratified/personalized treatment gastric-cancer. Our data represent the fundaments for the design of clinical trials focusing on the use of ATRA in the personalized treatment of this heterogeneous tumor. Our gene-expression model will permit the development of a predictive tool for the selection of ATRA-sensitive gastric-cancer patients. The immune-regulatory responses activated by ATRA suggest that the retinoid and immune-checkpoint inhibitors constitute rational combinations for the management of gastric-cancer.

Topics: Antineoplastic Agents; Humans; Retinoids; RNA; Stomach Neoplasms; Transcriptome; Tretinoin

Long noncoding RNAs (LncRNAs) play a pivotal role in gastric tumorigenesis, while exosomes facilitate the LncRNAs transferring to recipient cells. However, the roles of exosomal LncRNAs in gastric premalignant lesions (GPL) remain unclear.. We analyzed the expression of LncHOXA10 and its role in GPL progression. The protective effect of all-trans retinoic acid (ATRA) on GPL was explored in vitro and in vivo.. Here, we found that LncHOXA10 expression was obviously increased in serum exosomes and gastric tissues from individuals with GPL, and exosomal LncHOXA10 from patients with GPL markedly promoted the malignant progression of human gastric epithelial cell line GES-1. Furthermore, RNA-pulldown assay revealed that LncHOXA10 mainly interacted with pyruvate carboxylase (PC), an essential enzyme in various cellular metabolic pathways. In gastric tissues from patients with GPL and gastric cancer (GC), PC was also upregulated and positively correlated with LncHOXA10 expression, which predicted a poor prognosis as well. Moreover, PC silencing attenuated the malignant effects of exosomal LncHOXA10 on GES-1 cells. ATRA also ameliorated the deterioration of GPL and prevented the malignant progression of GPL by reducing exosomal LncHOXA10 and PC expression.. Collectively, the LncHOXA10-PC axis participated in the early stage of GC tumorigenesis, and ATRA might be useful to prevent GPL from developing into GC because it targets this axis.

Topics: Animals; Antineoplastic Agents; Carcinogenesis; Cell Line, Tumor; Exosomes; Female; Gene Expression Regulation, Neoplastic; Homeobox A10 Proteins; Humans; Male; Middle Aged; Precancerous Conditions; Pyruvate Carboxylase; Rats; Rats, Wistar; RNA, Long Noncoding; Stomach Neoplasms; Tretinoin

The vitamin A metabolite all-trans retinoic acid (ATRA) plays a key role in immune response, but effects of ATRA on cancer-associated immunity remains unclear. Previously, we have shown that ATRA regulates the expression of PD-L1 in gastric cancer (GC) cells. We herein reported the mechanism underlying ATRA-induced PD-L1 expression in GC cells and the effects of ATRA on cancer-associated immunosuppression in vitro and in vivo. ATRA enhanced PD-L1 expression through increasing its protein stability and protein synthesis, which was suppressed by JAK pan-inhibitor ruxolitinib (RUX) but enhanced in the combination with IFN-γ. In T-cell-mediated killing assay, the upregulation of PD-L1-induced by ATRA rendered GC cells strongly resistant to activated T-cell killing, which was reversed by RUX. In vivo, PD-L1 antibody restricted tumor growth, but ATRA antagonized PD-L1 antibody efficacy. Importantly, RUX not only inhibited the expression of PD-L1 induced by ATRA, but also resensitized GC cells to PD-L1 antibody. In conclusion, our study illustrated that ATRA attenuated the effect of PD-L1 blockade through upregulating PD-L1 and blocking PD-L1 expression is an important role for the generation of effective anti-tumor immune response in the combination of immunotherapy and chemotherapy or targeted therapy.

Topics: B7-H1 Antigen; Cell Line, Tumor; Humans; Immunotherapy; Stomach Neoplasms; T-Lymphocytes; Tretinoin

The potential of all-trans retinoic acid (ATRA) in regulating some microRNAs (miRNAs) involved in multiple cancer-related pathways, including resistance to chemotherapeutics, may be a valuable idea for overcoming the CDDP resistance of GC cells.. Treatment of gastric AGS and MKN-45 cells with CDDP enriched the CDDP surviving cells (CDDP-SCs). The abilities of chemoresistance to CDDP drug, migration, either apoptosis or cell cycle distribution, spheroid body formation and changes at miRNA and protein levels were evaluated in vitro by MTT assay, colony formation assay, flow cytometry, tumor spheres culture, qRT-PCR and western blot assay in CDDP-SCs and ATRA-treated CDDP-SCs cells, respectively.. CDDP-based chemotherapy significantly reduced microRNA-30a (miR-30a) levels in GC cells. We also observed elevated autophagy activity in cancer cells that possess stem cell-like properties with overexpressed specific stem cell markers. Our extended study suggested that the reduction of miR-30a by CDDP treatment, is the possible underlying mechanism of enhanced autophagic activity, as demonstrated by enhancing autophagy-related protein beclin 1 and LC3-II/LC-I ratio. The addition of ATRA in the culture medium of GC cells increased the expression of miR-30a, and disturbed characteristic CSC-like properties. Additional studies revealed that the increased expression of miR-30a declined the expression level of its target gene, beclin 1, and beclin 1-mediated autophagy. This leads to promoted CDDP-induced GC cell apoptosis and G2/M cell cycle arrest.. Overall, miR-30a/autophagy signaling has a critical role in regulating the chemoresistance of GC cells that ATRA could modulate.

Topics: Apoptosis; Autophagy; Beclin-1; Cell Line, Tumor; Cisplatin; Humans; MicroRNAs; Stomach Neoplasms; Tretinoin; Up-Regulation

High expression of glucose transporter family members, which augment glucose uptake and glycolytic flux, has been shown to play a pivotal role in the proliferation and survival of tumor cells, contributing to the energy supply, biosynthesis and homeostasis of cancer cells. Among the many members, solute carrier family 2 member 1 (SLC2A1) encodes a glucose transporter, GLUT1, that is critical in the metabolism of glucose, which is an energy source for cell growth that contributes to cancer progression and development. The aim of this study was to analyze the survival and genetic changes/immune profiles in patients with gastric cancer with high SLC2A1 expression and to provide treatment for improving prognosis. This study investigated the clinicopathologic parameters, the proportion of immune cells and gene sets affecting SLC2A1 expression in 279 and 415 patients with gastric cancer from the Eulji Hospital cohort and The Cancer Genome Atlas, respectively. We assessed the response to conventional chemotherapy drugs, including fluorouracil, a compound of fluoropyrimidine S-1, oxaliplatin, and all-trans-retinoic acid (ATRA), in gastric cancer cell lines with high SLC2A1 expression. High SLC2A1 expression was associated with poor prognosis, cancer cell proliferation, decreased immune cells, including CD8 T cells and B cells, and a low prognostic nutrition index, representing body nutrition-related status. In pathway network analysis, SLC2A1 was indirectly linked to the retinoic signaling pathway and negatively regulated immune cells/receptors. In the drug response analysis, the drug ATRA inhibited gastric cancer cell lines with high SLC2A1 expression. Treatment involving the use of SLC2A1 could contribute to better clinical management/research for patients with gastric cancer.

Topics: Aged; Antineoplastic Agents; B-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Line, Tumor; Cell Proliferation; Disease-Free Survival; DNA Copy Number Variations; Down-Regulation; Female; Glucose Transporter Type 1; Humans; Male; Middle Aged; Mutation; Prognosis; Signal Transduction; Stomach Neoplasms; Survival Rate; Tretinoin

Gastric cancer is the third leading cause of cancer-related death worldwide. Diffuse type gastric cancer has the worst prognosis due to notorious resistance to chemotherapy and enrichment of cancer stem-like cells (CSC) associated with the epithelial-to-mesenchymal transition (EMT). The unique proline isomerase PIN1 is a common regulator of oncogenic signaling networks and is important for gastric cancer development. However, little is known about its roles in CSCs and drug resistance in gastric cancer. In this article, we demonstrate that PIN1 overexpression is closely correlated with advanced tumor stages, poor chemo-response and shorter recurrence-free survival in diffuse type gastric cancer in human patients. Furthermore, shRNA-mediated genetic or all-

Topics: Adult; Animals; Antineoplastic Agents; Apoptosis; Biomarkers, Tumor; Cell Proliferation; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplastic Stem Cells; NIMA-Interacting Peptidylprolyl Isomerase; Prognosis; RNA, Small Interfering; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Gastric cancer is the second leading cause of cancer-related mortality and the fourth most common cancer globally. High intratumor heterogeneity of advanced gastric cancer poses great challenges to targeted therapy due to simultaneous activation of many redundant cancer-driving pathways. A central common signaling mechanism in cancer is proline-directed phosphorylation, which is further regulated by the unique proline isomerase Pin1. Pin1 inhibition exerts anticancer activity by blocking multiple cancer-driving pathways in some cancers, but its role in gastric cancer is not fully understood. Here we detected Pin1 protein expression in 1065 gastric cancer patients and paired normal tissues using immunohistochemistry and Western blot, and then examined the effects of Pin1 overexpression, and genetic and chemical Pin1 inhibition using Pin1 short hairpin RNA or small molecule inhibitor all-trans retinoic acid (ATRA) on tumorigenesis of human gastric cancer in vitro and in vivo, followed by biochemical analyses to elucidate Pin1 regulated oncogenic pathways. We found that Pin1 was significantly overexpressed in primary and metastasized tumors, with Pin1 overexpression being correlated with advanced stage and poor prognosis. Furthermore, whereas Pin1 overexpression promoted the transformed phenotype in immortalized and nontransformed human gastric cells, either genetic or chemical Pin1 inhibition in multiple human gastric cancer cells potently suppressed cell growth, G1/S transition and colony formation in vitro, as well as tumor growth in xenograft tumor models in vivo, which were further supported by downregulation of multiple key oncoproteins in PI3K/AKT and Wnt/β-catenin signaling pathways. These results not only provide the first evidence for a critical role of Pin1 in the tumorigenesis of gastric cancer but also suggest that targeting Pin1 using ATRA or other inhibitors offers an effective new therapeutic approach for treating advanced gastric cancer.

Topics: Animals; beta Catenin; Carcinogenesis; Cell Line, Tumor; Female; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; NIMA-Interacting Peptidylprolyl Isomerase; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; RNA Interference; RNA, Small Interfering; Stomach Neoplasms; Tretinoin; Wnt Proteins; Wnt Signaling Pathway

In this study, miR-542-3p appended SRF/ATRA-loaded solid lipid nanoparticle was successfully prepared and demonstrated for its therapeutic efficacy against gastric cancers.. The particles were nanosized and typically spherical in shape. In vitro release study showed that release of ATRA was significantly slower compared to that of SRF from the NPs.. MTT assay showed that miR-542-3p have a strong inhibitory effect on the proliferation of MGC-803 cancer cells in a typical dose dependent manner. Nanocarrier encapsulation of SRF + ATRA induced a significantly higher cytotoxic effect compared to either individual drug or cocktail combinations indicating that the cellular uptake of different formulations was rate limiting factor in the therapeutic efficacy. Importantly, miR-542-3p-based miSRNP exhibited an extremely significant toxic effect compared to any other treated group. Importantly, miSRNP induced a significantly higher early (~55%) and late (~15%) apoptotic effect in gastric cancer cells. In vivo anticancer analysis results clearly suggest that nanoparticle encapsulation of combination of SRF and miRNA (with miRNA) will have greater antitumor efficacy in tumor mice.. Overall, unique combination of miRNA coupled with SRF + ATRA in a lipid nanocarrier could be a promising therapeutic approach in gastric cancer treatment.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Proliferation; Humans; Mice, Nude; MicroRNAs; Nanoparticles; Niacinamide; Phenylurea Compounds; Sorafenib; Stomach Neoplasms; Tretinoin

Gastric carcinoma is the third leading cause of cancer-related death worldwide. This cancer, most of the time metastatic, is essentially treated by surgery associated with conventional chemotherapy, and has a poor prognosis. The existence of cancer stem cells (CSC) expressing CD44 and a high aldehyde dehydrogenase (ALDH) activity has recently been demonstrated in gastric carcinoma and has opened new perspectives to develop targeted therapy. In this study, we evaluated the effects of all-trans-retinoic acid (ATRA) on CSCs in human gastric carcinoma. ATRA effects were evaluated on the proliferation and tumorigenic properties of gastric carcinoma cells from patient-derived tumors and cell lines in conventional 2D cultures, in 3D culture systems (tumorsphere assay) and in mouse xenograft models. ATRA inhibited both tumorspheres initiation and growth in vitro, which was associated with a cell-cycle arrest through the upregulation of cyclin-dependent kinase (CDK) inhibitors and the downregulation of cell-cycle progression activators. More importantly, ATRA downregulated the expression of the CSC markers CD44 and ALDH as well as stemness genes such as Klf4 and Sox2 and induced differentiation of tumorspheres. Finally, 2 weeks of daily ATRA treatment were sufficient to inhibit gastric tumor progression in vivo, which was associated with a decrease in CD44, ALDH1, Ki67 and PCNA expression in the remaining tumor cells. Administration of ATRA appears to be a potent strategy to efficiently inhibit tumor growth and more importantly to target gastric CSCs in both intestinal and diffuse types of gastric carcinoma.

Topics: Aldehyde Dehydrogenase; Animals; Antineoplastic Agents; Biomarkers; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Disease Models, Animal; Female; Humans; Hyaluronan Receptors; Immunophenotyping; Kruppel-Like Factor 4; Mice; Neoplastic Stem Cells; Spheroids, Cellular; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Cis-diamminedichloridoplatinum(II)(CDDP)-based combination chemotherapy is frequently used in gastrointestinal cancer. The synergistic mechanism of all-trans retinoic acid (ATRA), cisplatin (CDDP) and 5-fluorouracil (5-FU) in combination remains unclear. Despite their potent antitumor properties, resistance to CDDP and 5-FU develops frequently in tumors. To clarify this mechanism, we determined the sensitivity to each drug and their combination in two gastrointestinal cancer stem cells (CSCs) subpopulation. Here, we report the identification and separation of CD44(+) cells from human gastric carcinoma (AGS) and human esophageal squamous cell carcinoma (KYSE-30) cancer cell lines by magnetic activated cell sorting (MACS). We allowed the CD44(±) cells to grow 6 days at a subtoxic concentration of ATRA and then treated with different concentration of CDDP and 5-FU for 24h. The cytotoxicity was examined by cell proliferation MTT assay. Additionally, AO/EB staining was used for detection of apoptotic cells. In order to determine whether the growth inhibition was also associated with changes in cell cycle distribution, cell cycle analysis was performed using flow cytometry. Low concentration of ATRA (1μM, 6days) followed by 5-FU and CDDP was found to be more effective than either drugs alone, thus resulting in synergistic cytotoxicity in Kyse-30 and AGSCD44(±) cells. Furthermore, there was an indication that the combination of ATRA with 5FU and CDDP caused an increase in cell cycle arrest in G2/M and G0/G1. We conclude that low concentration of ATRA enhances the cytotoxicity of CDDP and 5FU by facilitating apoptosis and cell cycle arrest in gastrointestinal CSCs and provide a rational basis for the design of novel, well-tolerated CDDP- and 5FU-based chemotherapy in human gastrointestinal carcinoma.

Topics: Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinoma, Squamous Cell; Cell Cycle; Cell Cycle Checkpoints; Cell Line, Tumor; Cisplatin; Dose-Response Relationship, Drug; Drug Resistance, Neoplasm; Drug Synergism; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Flow Cytometry; Fluorouracil; Humans; Hyaluronan Receptors; Neoplastic Stem Cells; Stomach Neoplasms; Tretinoin

Retinoic acid (RA) analogs have been used in the treatment of a variety of cancers; however, their application is limited due to serious therapy-related sequelae. In the present study, the effects of a novel RA analog, 4-amino-2-trifluoromethyl-phenyl retinate (ATPR), on the growth of gastric cancer cells were evaluated. Three gastric cancer cell lines, AGS, MKN-74 and SC-M1, were treated with either all‑trans retinoic acid (ATRA) or ATPR, and their growth and distribution in different cell cycle phases were assessed using an MTT assay and propidium iodide (PI) staining followed by flow cytometry. The binding affinity of ATPR to the retinoic acid receptors, retinoic acid receptor-α (RAR-α) and retinoid X receptor-α (RXR-α), was determined using ligand-binding assays. Activator protein-1 (AP-1) activity was measured using a luciferase reporter assay. Western blot analysis was used to determine cyclin E, Bcl-2 and Bax protein expression. ATPR preferentially bound RXR-α (0.04 nM) as compared with RAR-α (20.96 nM). Although both ATRA and ATPR inhibited the growth of AGS, MKN-74 and SC-M1 cells in a dose-dependent manner, a significantly greater inhibitory effect was observed with treatment with 5 and 500 µM ATPR for 3 days (P<0.05). In addition, ATPR (50 µM), but not ATRA, significantly increased the population of AGS and MKN-74 cells in the subG1 phase and decreased the Bcl-2/Bax ratio (P<0.05). Furthermore, in MNK-74 and SC-M1 cells treated with 12-O-tetradecanoylphorbol-13-acetate (TPA) and 5 or 10 µM of ATPR significantly suppressed the activity of the AP-1 reporter as compared to treatment with ATRA (P<0.05). Thus, ATPR inhibits cancer cell proliferation to a greater extent compared to ATRA, possibly through the RXR-mediated inhibition of AP-1 activity.

Topics: Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Humans; Retinoids; Stomach Neoplasms; Transcription Factor AP-1; Tretinoin

In order to investigate the inhibitory effects and mechanisms of troglitazone (TGZ), a peroxisome proliferator-activated receptor γ (PPARγ) agonist, and retinoid X receptor (RXR) agonist (9-cis-retinoic acid (RA)) on gastric carcinoma cells SGC7901, SGC7901 cells were treated with TGZ and 9-cis-RA, respectively, or in combination. Then, the cell growth, apoptosis, morphological changes, and the expression of PPARγ, RXRγ, Bcl-2, and Bax were detected by MTT assay, flow cytometry, HE staining, immunocytochemistry staining, and Western blot assay, respectively. Our results showed that the growth of SGC7901 cells was inhibited and the cells got sparser at the concentrations of 50 μmol/L TGZ, 20 μmol/L 9-cis-RA, or combination of TGZ (25 μmol/L) and 9-cis-RA (10 μmol/L). Immunocytochemistry and Western blot showed that after 72 h, the expression of PPARγ, RXRγ, and Bax were upregulated; Bcl-2 was downregulated compared with the negative control group. These data indicated that PPARγ agonist and RXR agonist could inhibit the proliferation of SGC7901 cells via inducing the apoptosis, which involved the increase in the level of Bax/Bcl-2. The combination of RXR agonist and PPARγ agonist could induce the maximal inhibitory effects on tumor growth and apoptosis via promoting the formation of RXR/PPARγ heterodimer.

Topics: Alitretinoin; Antineoplastic Agents; Apoptosis; bcl-2-Associated X Protein; Cell Line, Tumor; Cell Proliferation; Chromans; Down-Regulation; Humans; Hypoglycemic Agents; Intercalating Agents; PPAR gamma; Propidium; Proto-Oncogene Proteins c-bcl-2; Retinoid X Receptors; Stomach Neoplasms; Thiazolidinediones; Tretinoin; Troglitazone; Up-Regulation

Gastric cancer is frequently lethal despite aggressive multimodal therapies, and new treatment approaches are therefore needed. Retinoids are potential candidate drugs: they prevent cell differentiation, proliferation and malignant transformation in gastric cancer cell lines. They interact with nuclear retinoid receptors (the retinoic acid receptors [RARs] and retinoid X receptors [RXRs]), which function as transcription factors, each with three subclasses, α, β and γ. At present, little is known about retinoid expression and influence on prognosis in gastric cancers.. We retrospectively analyzed the expression of the subtypes RARα, RARβ, RARγ, RXRα, RXRβ, RXRγ by immunohistochemistry in 147 gastric cancers and 51 normal gastric epithelium tissues for whom clinical follow-up data were available and correlated the results with clinical characteristics. In addition, we quantified the expression of retinoid receptor mRNA using real- time PCR (RT-PCR) in another 6 gastric adenocarcinoma and 3 normal gastric tissues. From 2008 to 2010, 80 patients with gastric cancers were enrolled onto therapy with all-trans-retinoic acid (ATRA).. RARα, RARβ, RARγ and RXRγ positively correlated with each other (p<0.001) and demonstrated significantly lower levels in the carcinoma tissue sections (p<0.01), with lower RARβ, RARγ and RXRα expression significantly related to advanced stages (p<=0.01). Tumors with poor histopathologic grade had lower levels of RARα and RARβ in different histological types of gastric carcinoma (p<0.01). Patients whose tumors exhibited low levels of RARa expression had significantly lower overall survival compared with patients who had higher expression levels of this receptor (p<0.001, HR=0.42, 95.0% CI 0.24-0.73), and patients undergoing ATRA treatment had significantly longer median survival times (p=0.007, HR=0.41, 95.0% CI 0.21-0.80).. Retinoic acid receptors are frequently expressed in epithelial gastric cancer with a decreased tendency of expression and RARa may be an indicator of a positive prognosis. This study provides a molecular basis for the therapeutic use of retinoids against gastric cancer.

Topics: Adenocarcinoma; Aged; Antineoplastic Agents; Female; Humans; Immunoenzyme Techniques; Male; Middle Aged; Neoplasm Staging; Prognosis; Receptors, Retinoic Acid; Retrospective Studies; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stomach Neoplasms; Survival Rate; Tretinoin

To treat cancer cells overexpressing P-glycoprotein (P-gp), we propose a new concept using a nanodrug. The nanodrug was prepared from polyethyleneimine (PEI)/all-trans retinoic acid (ATRA) conjugates (PRA) and covered with hyaluronic acid (HA) to control the cytotoxicity of PRA (yielding PRA-H). The size distribution of PRA-H was narrow, with an average particle size of approximately 143 nm. Its superior stability in phosphate-buffered saline (PBS) was verified by monitoring changes in particle size and zeta potential for 24 h, which were negligible. In contrast, PEI-H (not conjugated with ATRA) exhibited a significant change in particle size and zeta potential. Although PRA was highly cytotoxic against HCT-8 and SNU-484 cancer cells, both of which overexpress P-gp, the cytotoxicity was significantly reduced by shielding with HA. The cytotoxicity of PRA-H was recovered by treatment with hyaluronidase (HAase), which degrades HA and is present in tumors at high concentrations. These results were confirmed by optical microscopy, fluorescence-activated cell sorting (FACs) analysis, and confocal microscopy. The cytotoxic mechanism of PRA was revealed as a type of necrotic lysis by FACs analysis with propidium iodide (PI) staining. Furthermore, PRA increased HCT-8 cell (colon cancer) permeability to doxorubicin (DOX). Therefore, we concluded that PRA-H is a promising new candidate for the treatment of cells with multidrug resistance (MDR) induced by overexpression of P-gp and cancer stem cells.

Topics: Apoptosis; ATP Binding Cassette Transporter, Subfamily B, Member 1; Cell Membrane Permeability; Cell Proliferation; Colonic Neoplasms; Drug Resistance, Multiple; Drug Resistance, Neoplasm; Humans; Hyaluronic Acid; Nanoparticles; Polyethyleneimine; Polymers; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

To determine the inhibitory effects of all-trans-retinoic acid (ATRA) on cell growth, cell cycle and vascular endothelial growth factor (VEGF) expression in the human gastric cancer cell line BGC-823 in vitro.. Human gastric cancer BGC-823 cells were treated with various concentrations of ATRA and the cell growth was then determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide viability assay. The cell cycle distribution was analyzed using a flow cytometer. The VEGF mRNA and protein expression were analyzed by semi-quantitative RT-PCR and Western blotting, respectively.. ATRA at concentrations of 0.1-10 micromol/L inhibited the growth of BGC-823 cells grown in culture; a time- and dose-dependent inhibitory influence was found. ATRA arrested BGC-823 cells at the G0/G1 phase in a dose-dependent way. Both VEGF mRNA and protein were decreased by ATRA in a dose-dependent way.. The anti-tumor effects of ATRA on human gastric cancer cells are associated with G0/G1 phase arrest and decreased VEGF expression.

Topics: Antineoplastic Agents; Cell Cycle; Cell Proliferation; Dose-Response Relationship, Drug; Humans; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured; Vascular Endothelial Growth Factor A

Antitumor effect of 9-cis retinoic acid (9-cis RA) on gastric carcinoma is unclear yet. This study was to explore the inducement effect of 9-cis RA on cell cycle arrest and apoptosis of gastric carcinoma cell line MGC803 and its mechanism.. The expression of RXRalpha, Cyclin D1, and CDK4 in MGC803 cells was detected by reverse transcription-polymerase chain reaction (RT-PCR). Cell cycle was detected by flow cytometry. The growth inhibition was analyzed by MTT assay. The apoptosis was detected by agarose gel electrophoresis and Hoechst33342/PI staining. The expression of apoptosis-associated gene Bcl-2 was detected by SP immunocytochemistry.. When treated with 0.1-10 micromol/L 9-cis RA for 96 h, the proliferation of MGC803 cells was significantly inhibited. The proportion of MGC803 cells at G1 phase was significantly increased when treated with 10 micromol/L 9-cis RA for 48, 72, and 96 h, and showed an apparent G1 phase arrest. When treated with 9-cis RA for 72 h, typical apoptotic changes, such as chromatin condensation and DNA ladder, were observed in MGC803 cells. The expression of Bcl-2 was significantly decreased in MGC803 cells when treated with 10 micromol/L 9-cis RA for 48 h. RXRalpha expression was at a low level in MGC803 cells and up-regulated when treated with 10 micromol/L 9-cis RA for 48 h (P<0.01). The expression of Cyclin D1 and CDK4 in MGC803 cells was both significantly down-regulated when treated with 10 micromol/L 9-cis RA for 96 h (P<0.01).. 9-Cis RA could induce G1 phase arrest and apoptosis in MGC803 cells through down-regulating the expression of cell cycle factors Cyclin D1 and CDK4.

Topics: Alitretinoin; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase 4; G1 Phase; Humans; Proto-Oncogene Proteins c-bcl-2; Retinoid X Receptor alpha; RNA, Messenger; Stomach Neoplasms; Tretinoin

p62 is a cancer-associated antigen binding to mRNA encoding insulin-like growth factor II that was isolated by immunoscreening a cDNA expression library with autoantibodies from patients with hepatocellular carcinoma (HCC). In the present study, multiple methods including flow cytometry, confocal laser-scanning microscope, electron microscope were used to characterize the effect of ATRA on BGC-823 cells, which presented two phenotypes of differentiation and apoptosis in cells treated with 1.0 and 50 microM ATRA, respectively. Interestingly, we found that p62 was cytoplasmic in location, but it significantly decreased in cytoplasm and appeared in nucleus of cells when the cells were treated with 50 microM all-trans retinoic acid (ATRA) for 5 days. Furthermore, proteomics approach on differential nucleus proteins showed that the up-regulation and/or down-regulation of cell cycle proteins and IGF binding proteins were involved in the apoptosis of BGC-823 cells induced by ATRA. These results suggest that there is a significant association between expression and distribution of p62 and the growth arrest of tumor cells, in which p62 is associated with cell apoptosis induced by ATRA.

Topics: Apoptosis; Blotting, Western; Carcinoma, Hepatocellular; Cell Differentiation; Cell Line, Tumor; Dose-Response Relationship, Drug; Electrophoresis, Gel, Two-Dimensional; Flow Cytometry; Fluorescent Antibody Technique, Indirect; Humans; Image Processing, Computer-Assisted; Microscopy, Confocal; Microscopy, Electron; Neoplasm Proteins; Peptide Mapping; Proteomics; RNA-Binding Proteins; RNA, Messenger; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Stomach Neoplasms; Subcellular Fractions; Time Factors; Tretinoin

The distribution of the cancer-associated protein p62 in human gastric carcinoma (BGC-823) cells was examined by confocal laser scanning microscopy and electron microscope immunocytochemistry. In control cells p62 was cytoplasmic in location and concentrated in the cytoplasmic matrix, but when cell growth was inhibited by treatment with 50 microM all-trans-retinoic acid (ATRA) for 5 days, p62 expression decreased and the protein was translocated from cytoplasm to nucleus. Ultrastructural localization using gold particles showed that p62 was bond mainly to a linear structure in nucleus. The speculation that p62 binds Insulin-like growth factor (IGF)-II mRNA indicates its probable involvement in the posttranscriptional IGF-II mRNA processing and p62 could play a role in tumorgenesis by regulating the expression of IGF-II. Further studies will be needed to confirm this view.

Topics: Carcinoma; Cell Line, Tumor; Cell Nucleus; Humans; Microscopy, Confocal; RNA-Binding Proteins; Stomach Neoplasms; Tretinoin

The anticarcinogenic effect of vitamin A2 (dehydroretinol and 3-hydroxyretinol) compounds was studied and compared with that of vitamin A1 (retinoic acid, retinol and retinal) and carotenoids (lutein and beta-carotene) in the benzo[a]pyrene (B(a)P)-induced forestomach tumour model of female Swiss mice in vivo. Tumour growth and gross tumour incidence observed after the administration of B(a)P (eight doses of 1 mg, twice weekly for 4 weeks) and retinoids/carotenoids (2.5 and 4.7 microm per animal per d, 2 weeks before, during and 2 weeks after B(a)P) showed that the groups supplemented with lutein and 3-hydroxyretinol produced the best results in inhibiting tumour growth and had low tumour incidence compared with the control group given B(a)P only (P<0.05). Weights recorded after the different treatments showed that the beta-carotene-supplemented group exhibited maximum weight gain, followed by retinal, retinol, retinoic acid, lutein, dehydroretinol and 3-hydroxyretinol. These results indicate that the anticarcinogenicity of the compounds is not related to the vitamin A biopotencies. Vitamin A2 compounds having half the biopotency of the vitamin A1 compounds were seen to be anticarcinogenic. Again, among the carotenoids, lutein, having 50 % less biopotency, showed more significant results than beta-carotene. Thus it is imperative to conclude that the low animal growth achieved with these compounds has a correlation with the highest suppression of tumour occurrence in the present experiment. Therefore, the daily consumption of foods having high content of lutein and vitamin A2 should be given due importance and weight in further studies.

Topics: Animals; Anticarcinogenic Agents; Benzo(a)pyrene; beta Carotene; Biological Availability; Carcinogens; Carotenoids; Dietary Supplements; Female; Lutein; Mice; Retinaldehyde; Retinoids; Stomach Neoplasms; Tretinoin; Vitamin A; Vitamins

All-trans retinoic acid (ATRA) affects cell proliferation, differentiation and apoptosis through its receptors, RARs and RXRs. Besides these, other receptors such as orphan receptor TR3, are also involved in the regulatory process of ATRA. However, how different receptors function in response to ATRA is still largely unknown. In the present study, we found that formation of TR3/RXRalpha heterodimers in the nucleus and their subsequent translocation into the cytoplasm, in association with regulation of apoptosis-related proteins Bcl-2, Bcl-xl and Bax, was critical for apoptosis induction by ATRA in breast cancer cells MCF-7. When such translocation was blocked by Leptomycin B (LMB), ATRA-induced apoptosis was consequently abolished. However, in ATRA-induced gastric cancer cells MGC80-3, RXRalpha heterodimerised with RARalpha but not with TR3, and remained in the nucleus exerting its effect on cell cycle regulation. When transfected with antisense-RARalpha, MGC80-3 cells changed from ATRA-sensitive to ATRA-resistant and most cells were arrested in the S phase, implying the importance of RARalpha in cell cycle regulation. Furthermore, we demonstrated that the effects of ATRA depend on the relative levels of TR3, RARalpha and RXRalpha expression in cancer cells. In ATRA-induced MCF-7 cells, highly expressed TR3 favours the formation of TR3/RXRalpha and promotes the TR3/RXRalpha signalling pathway causing apoptosis; while in ATRA-induced MGC80-3 cells, high expression of RARalpha favours the formation of RARalpha/RXRalpha and promotes the RXRalpha/RARalpha signalling pathway in mediating cell cycle regulation. In conclusion, these results reveal the novel mechanism that cellular expression and location of protein is associated with diverse signalling transduction pathways and the resultant physiological process.

Topics: Apoptosis; Blotting, Western; Breast Neoplasms; Cell Division; Cell Line, Tumor; Female; Gene Expression Regulation, Neoplastic; Humans; Nuclear Receptor Subfamily 4, Group A, Member 1; Receptors, Retinoic Acid; Receptors, Steroid; Receptors, Thyroid Hormone; Retinoic Acid Receptor alpha; Signal Transduction; Stomach Neoplasms; Tretinoin

Retinoic acid receptor alpha (RARalpha) plays an important role in mediating all-trans retinoic acid (ATRA) signals. In this study, we found that ATRA up-regulated RARalpha mRNA and protein expression in gastric cancer BGC-823 cells. However, in breast cancer MCF-7 cells it down-regulated RARalpha protein expression with no effect on its RARalpha mRNA. Immunoprecipitation/Western blot analysis showed that, although sumoylated and ubiquitinated RARalpha existed simultaneously in both cancer cell lines, ATRA exerted different regulatory effects on sumoylation and ubiquitination of RARalpha. In MCF-7 cells, ATRA treatment enhanced the ubiquitination of RARalpha and the subsequent degradation of RARalpha through the ubiquitin/proteasome pathway. This resulted in a reduction in the DNA binding activity of RARalpha/retinoid X receptor alpha (RXRalpha) heterodimer, the separation of RXRalpha from RARalpha and the translocation of RXRalpha from the nucleus to the cytoplasm. By contrast, in BGC-823 cells, ATRA augmented sumoylation, not ubiquitination, of RARalpha. The stability of sumoylated RARalpha was significantly stronger than in non-sumoylated RARalpha. These results also showed an increase in the DNA binding activity of the RARalpha/RXRalpha heterodimer and the stability of nuclear localization of this heterodimer, which normally facilitates the ATRA signal transduction. In conclusion, our results reveal a novel mechanism for the regulation of RARalpha-dependent signal transduction through the ubiquitin/proteasome pathway in breast cancer cells and the sumoylation pathway in gastric cancer cells.

Topics: Breast Neoplasms; Cell Line, Tumor; Dimerization; Female; Humans; Polyubiquitin; Proteasome Endopeptidase Complex; Proteasome Inhibitors; Protein Processing, Post-Translational; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoid X Receptor alpha; Stomach Neoplasms; SUMO-1 Protein; Tretinoin

Retinoid X receptor (RXR) plays a crucial role in the cross talk between retinoid receptors and other hormone receptors including the orphan receptor TR3, forming different heterodimers that transduce diverse steroid/thyroid hormone signaling. Here we show that RXRalpha exhibits nucleocytoplasmic shuttling in MGC80-3 gastric cancer cells and that RXRalpha shuttling is energy-dependent through a nuclear pore complex (NPC)-mediated pathway for its import and an intact DNA binding domain-mediated pathway for its export. In the presence of its ligand 9-cis retinoic acid, RXRalpha was almost exclusively located in the cytoplasm. More importantly, we also show that RXRalpha acts as a carrier to assist translocation of TR3, which plays an important role in apoptosis. Both RXRalpha and TR3 colocalized in the nucleus; however, upon stimulation by 9-cis retinoic acid they cotranslocated to the cytoplasm and then localized in the mitochondria. TR3 export depends on RXRalpha, as in living cells GFP-TR3 alone did not result in export from the nucleus even in the presence of 9-cis retinoic acid, whereas GFP-TR3 cotransfected with RXRalpha was exported out of the nucleus in response to 9-cis retinoic acid. Moreover, specific reduction of RXRalpha levels caused by anti-sense RXRalpha abolished TR3 nuclear export. In contrast, specific knockdown of TR3 by antisense-TR3 or TR3-siRNA did not affect RXRalpha shuttling. These results indicate that RXRalpha is responsible for TR3 nucleocytoplasmic translocation, which is facilitated by the RXRalpha ligand 9-cis retinoic acid. In addition, mitochondrial TR3, but not RXRalpha, was critical for apoptosis, as TR3 mutants that were distributed in the mitochondria induced apoptosis in the presence or absence of 9-cis retinoic acid. These data reveal a novel aspect of RXRalpha function, in which it acts as a carrier for nucleocytoplasmic translocation of orphan receptors.

Topics: 3T3 Cells; Active Transport, Cell Nucleus; Alitretinoin; Animals; Apoptosis; Cell Nucleus; Cytoplasm; DNA-Binding Proteins; Down-Regulation; Green Fluorescent Proteins; HeLa Cells; Humans; Mice; Mitochondria; Nuclear Pore; Nuclear Receptor Subfamily 4, Group A, Member 1; Oligonucleotides, Antisense; Protein Transport; Receptors, Steroid; Receptors, Thyroid Hormone; Retinoid X Receptor alpha; RNA Interference; Stomach Neoplasms; Tretinoin

Many protein tyrosine kinases are key regulators involved in cellular growth, differentiation, development, apoptosis and signal transduction pathways. Obtaining a comprehensive tyrosine-kinase expression profile in tumour cells is essential to learning more about their oncogenic potentials and responses to various chemotherapeutic reagents - such as retinoic acid, which has been shown to suppress the growth of gastric cancer cells and modulate gene expression. Expression of tyrosine kinases in retionic acid-treated cancer cells was investigated by reverse trancriptase-polymerase chain reaction (RT-PCR) and a novel restriction analysis of gene expression (RAGE) display technique. We first established comprehensive tyrosine-kinase profiles in different human gastric cancer cell lines. In cells treated with 9-cis-retinoic acid or all-trans-retinoic acid, we found that two PTKs (Eph and Hek5) appeared to be upregulated. In the present study, we demonstrate an efficient and simple RAGE approach for examining tyrosine kinases' expression in tumour cells and their alterations following drug treatments.

Topics: Gene Expression Profiling; Humans; Protein-Tyrosine Kinases; Receptor, EphA3; Receptors, Eph Family; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoic Acid Receptor gamma; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

To investigate the role of retinoic acid receptor beta (RARbeta) in mediating inhibitory effect of all-trans retinoic acid (ATRA) on activator protein-1 (AP-1) activity in gastric cancer cells.. Transient transfection and chloramphenicol acetyltransferase (CAT) assay, Nort hern blot, gene transfection, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, and anchorage independent growth assay were used.. Transient transfection of RARbeta expression vector into MKN-45 cells resulted in the RARbeta concentration dependent repression of AP-1 activity induced by 12-o-tetradecanoylphorbol-13-acetate (TPA), regardless of the presence of ATRA. When the c-jun and c-fos expression vectors were cotransfected with the RARbeta expression vector into MKN-45 cells, AP-1 activity was also obviously repressed. The inhibitory effect, again, was RARbeta-concentration-dependent. The stable transfection of the RARbeta gene into MKN-45 cells led to cell growth inhibition and colony formation inhibition by ATRA. Furthermore, Cotransfection of both RARbeta/DNA binding domain (DBD) and reporter gene could not alter AP-1 activity, even in the presence of ATRA.However, when the cotransfection was substituted with the RARbeta/ligand binding domain (LBD), the inhibition was significantly enhanced by ATRA.. RARbeta might be required for anti-AP-1 activity, and contribute to growth inhibition of gastric cancer cells by ATRA.

Topics: Antineoplastic Agents; Binding Sites; Cell Division; DNA; Humans; Receptors, Retinoic Acid; Stomach Neoplasms; Transcription Factor AP-1; Tretinoin; Tumor Cells, Cultured

To investigate the effects of all-trans retinoic acid (ATRA) on metastasis and its related proteins in human gastric cancer cells in vivo and in vitro.. Gastric cancer cells, MGC80-3 and SGC-7901, were inoculated into spleen subcapsule of nude mice, respectively. Nude mice were administered with ATRA (0.7 mg/kg, ig) every other day. Six weeks later, nude mice were sacrificed. All the tumors formed in spleen and in liver were removed. Some of them were fixed, and then embedded. Others were kept in liquid nitrogen for further use. Expression level of proteins in tumor and in cell was analyzed by Western blot. Microvessel in tumor section was shown by immunohistochemistry and adhesive ability of cell to amnion was measured by adhesion assay.. When inoculated nude mice were treated with ATRA, the xenograft tumors in spleen and metastatic tumors in liver were suppressed by 50 % respectively, and inhibition of microvessel formation in xenograft and metastatic tumors was also observed obviously. Although ATRA regulated expression of nm23 and mts1/p16 proteins at different patterns in vivo and in vitro, high ratio of nm23:mts1/p16 was in association with low adhesive activity of cells. In addition, ATRA induced ICAM-1 protein expression in vivo and in vitro.. ATRA inhibits the growth of xenograft tumors and their metastasis to liver. This process may be associated with regulation of metastatic related proteins, including nm23, mts1/p16, and ICAM-1 in vivo and in vitro.

Topics: Animals; Antineoplastic Agents; Cyclin-Dependent Kinase Inhibitor p16; Disease Models, Animal; Humans; Intercellular Adhesion Molecule-1; Mice; Mice, Inbred BALB C; Mice, Nude; Monomeric GTP-Binding Proteins; Neoplasm Metastasis; Neoplasm Transplantation; Neoplasms, Experimental; NM23 Nucleoside Diphosphate Kinases; Nucleoside-Diphosphate Kinase; Stomach Neoplasms; Transcription Factors; Tretinoin; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Nur77 is an orphan receptor. Although Nur77 affects cell proliferation and apoptosis through its capability of binding to a variety of response elements and regulating their transactivation activities, the intrinsic function of Nur77 is not yet fully understood; in particular, its regulation of apoptosis and proliferation has been characterized as cell type-dependent and agent context-dependent. In this study, Nur77 can be seen to regulate apoptosis via its expression and translocation, rather than its transactivation activity in gastric cancer cells. Nur77 was constitutively expressed in BGC-823 cells. The tetradecanoylphorbol-1,3-acetate (TPA) treatment not only resulted in up-regulation of the Nur77 mRNA level, but also led to translocation of Nur77 protein from the nucleus to the mitochondria, and caused the release of cytochrome c. This TPA-induced translocation of Nur77 was in association with the initiation of apoptosis in gastric cancer cells. Although all-trans retinoic acid (ATRA) could not induce apoptosis in BGC-823 cells due to failure of stimulating Nur77 translocation, expression of Nur77 in the nucleus was required for cell growth inhibition by ATRA. Transfection of antisense Nur77 receptor into BGC-823 cells resulted in resistance of cell growth against ATRA inhibition, and the cells were still arrested in the S phase. Furthermore, the action of Nur77 in TPA-induced apoptosis was mediated through a protein kinase C signaling pathway, while mitogen-activated protein kinase and phosphatidylinositol 3-kinase signaling pathways were responsible for the regulation of Nur77 mRNA expression. Taken together, the data revealed the dual functioning mechanisms of Nur77 in gastric cancer cells in response to TPA and ATRA.

Topics: Apoptosis; Cell Cycle; Cycloheximide; DNA-Binding Proteins; Epidermal Growth Factor; Genetic Vectors; Humans; Nuclear Receptor Subfamily 4, Group A, Member 1; Receptors, Cytoplasmic and Nuclear; Receptors, Steroid; Recombinant Proteins; Stomach Neoplasms; Tetradecanoylphorbol Acetate; Transcription Factors; Transfection; Tretinoin; Tumor Cells, Cultured

To uncover the mechanisms relating to the anticancer effect of retinoic acids in gastric cancer cells, the mediation of activator protein-1 (AP-1) activity repression by retinoic acid receptors (RARs) was investigated. All-trans retinoic acid (ATRA) inhibited AP-1 activity in BGC-823 cells (RARalpha(+), RARbeta(+)), but not in MKN-45 cells (RARalpha(lo), RARbeta(-)). Transient transfection of RARbeta expression vector into MKN-45 cells significantly resulted in direct repression of AP-1 activity in a receptor concentration-dependent manner, and this could be strengthened by ATRA. Stable transfection of RARbeta into MKN-45 cells directly inhibited cell growth and colony formation, and ATRA also enhanced these effects. Transient transfection of RARalpha into MKN-45 cells however, displayed receptor concentration-dependent AP-1 activity inhibition only in the presence of ATRA. Stable transfection of RARalpha into MKN-45 cells resulted in ATRA-dependent inhibition of cell growth and colony formation. For AP-1 binding activity induced by TPA, the repressive effect of ATRA was only observed in BGC-823 and RARalpha and RARbeta stably transfected MKN-45 cells, but not in intact MKN-45 cells. This indicates the necessity for sufficient cellular RARalpha and/or RARbeta in order for AP-1 activity repression to occur. Deletion of DNA binding domain (DBD) of RARbeta, but not ligand binding domain (LBD), eliminated the anti-AP-1 function of RARbeta. It is therefore concluded that both RARalpha and RARbeta are mediators in the anticancer function of ATRA via AP-1 activity inhibition, and that RARbeta, not RARalpha, can inhibit AP-1 activity to a certain extent directly by itself. Thus DBD, not LBD, is critical for anti-AP-1 activity.

Topics: Antineoplastic Agents; Cell Division; DNA-Binding Proteins; Genes, Reporter; Humans; Protein Binding; Protein Structure, Tertiary; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Stomach Neoplasms; Tetradecanoylphorbol Acetate; Transcription Factor AP-1; Tretinoin; Tumor Cells, Cultured

To investigate the mechanism of all-trans retinoic acid (ATRA) on the regulation of the cell cycle in gastric cancer cells.. The protein level was detected by Western blot. Immunoprecipitation was used in protein kinase activity determination. Cell growth and cell cycle phase were examined by MTT assay and flow-cytometric analysis, respectively.. ATRA could effectively induce G0/G1 arrest and inhibit cell growth in certain human gastric cancer cell lines. ATRA might induce p21WAF1/CIP1 expression in ATRA-sensitive cell lines through p53-dependent and p53-independent pathways. Induction of p21WAF1/CIP1 caused decrease in CDK4 and CDK2 activities independent of CDK4 and CDK2 protein expression levels. In addition, the dephosphorylated form of Rb protein increased because of the down-regulation of CDK4 and CDK2 activities by ATRA.. Growth inhibition on gastric cancer cells by ATRA occurs through the regulation of relevant proteins leading to the arrest of cell cycle progression.

Topics: Antineoplastic Agents; Blotting, Western; Cell Cycle; Cell Division; Cyclin E; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Dose-Response Relationship, Drug; Humans; Phosphorylation; Retinoblastoma Protein; Stomach Neoplasms; Time Factors; Tretinoin; Tumor Cells, Cultured; Tumor Suppressor Protein p53

To evaluate the role of RARalpha gene in mediating the growth inhibitory effect of all-trans retinoic acid (ATRA) on gastric cancer cells.. The expression levels of retinoic acid receptors (RARs) in gastric cancer cells were detected by Northern blot. Transient transfection and chlorophenicol acetyl transferase (CAT) assay were used to show the transcriptional activity of beta retinoic acid response element (betaRARE) and AP-1 activity. Cell growth inhibition was determined by MTT assay and anchorage-independent growth assay, respectively. Stable transfection was performed by the method of Lipofectamine, and the cells were screened by G418.. ATRA could induce expression level of RARalpha in MGC80-3, BGC-823 and SGC-7901 cells obviously, resulting in growth inhibition of these cell lines. After sense RARalpha gene was transfected into MKN-45 cells that expressed rather low level of RARalpha and could not be induced by ATRA, the cell growth was inhibited by ATRA markedly. In contrast, when antisense RARalpha gene was transfected into BGC-823 cells, a little inhibitory effect by ATRA was seen, compared with the parallel BGC-823 cells. In transient transfection assay, ATRA effectively induced transcriptional activity of betaRARE in MGC80-3, BGC-823, SGC-7902 and MKN/RARalpha cell lines, but not in MKN-45 and BGC/aRARalpha cell lines. Similar results were observed in measuring-antiAP-1 activity by ATRA in these cancer cell lines.. ATRA inhibits the growth of gastric cancer cells by up-regulating the level of RARalpha RARalpha is the major mediator of ATRA action in gastric cancer cells; and adequate level of RARalpha is required for ATRA effect on gastric cancer cells.

Topics: Agar; Antineoplastic Agents; Cell Division; Clone Cells; Gene Expression Regulation, Neoplastic; Humans; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoic Acid Receptor gamma; Retinoid X Receptors; RNA, Messenger; Stomach Neoplasms; Transcription Factor AP-1; Transcription Factors; Transcriptional Activation; Transfection; Tretinoin; Tumor Cells, Cultured

The role of Bcl-2 as an anti-apoptotic protein has been well documented. In the present work, we present evidence that Bcl-2 may also be involved in cell growth regulation. SC-M1 is an unique cell line which responds to retinoic acid (RA) treatment with reversible growth arrest [Shyu, Jiang, Huang, Chang, Wu, Roffler and Yeh (1995) Eur. J. Cancer 31, 237-243]. In this study, when treated with RA, SC-M1/Bcl2 cells, which were generated by transfecting SC-M1 cells with bcl-2 DNA, were growth-arrested two days earlier than SC-M1/neo cells, which were generated by transfecting SC-M1 cells with vector DNA. This indicates that Bcl-2 accelerates RA-induced growth arrest. In addition to the accelerated growth arrest, RA-treated SC-M1/Bcl2 cells also recovered from growth arrest two days faster than SC-M1/neo cells after the removal of RA. Previously, we had identified the cyclin-dependent kinase inhibitor p21((WAF1/CIP1)) (p21) as a mediator of RA-induced growth arrest [Tsao, Li, Kuo, Liu and Chen (1996) Biochem. J. 317, 707-711]. In a search for the mechanism by which Bcl-2 affects growth regulation, we found that p21 gene expression was more prominent in SC-M1/Bcl2 cells than in SC-M1/neo cells in the presence of RA, but when RA was removed, p21 gene expression levels in SC-M1/Bcl2 cells were also reduced earlier than in SC-M1/neo cells. The present report is the first to show that Bcl-2 accelerates not only growth arrest but also recovery from growth arrest. Moreover, the close correlation between the effect of Bcl-2 on both RA-induced growth arrest and RA-induced p21 gene expression suggests the possibility that Bcl-2 affects cell growth through the mechanism of p21.

Topics: Cell Cycle; Cell Division; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; DNA, Neoplasm; Enzyme Inhibitors; Flow Cytometry; Genes, bcl-2; Humans; Kinetics; Proto-Oncogene Proteins c-bcl-2; Recombinant Proteins; Stomach Neoplasms; Transfection; Tretinoin; Tumor Cells, Cultured

Peroxisome proliferator-activated receptor (PPAR) gamma is expressed in human colon cancer, prostate cancer and breast cancer cells, and PPARgamma activation induces growth inhibition in these cells. PPARgamma expression in human gastric cancer cells, however, has not been fully investigated. We report the PPARgamma expression in human gastric cancer, and the effect of PPARgamma ligands on proliferation of gastric carcinoma cell lines. Immunohistochemistry was used to demonstrate the presence of PPARgamma protein in surgically resected specimens from well differentiated, moderately differentiated and poorly differentiated adenocarcinoma. We used reverse transcription-polymerase chain reaction and Northern and Western blot analyses to demonstrate PPARgamma expression in four human gastric cancer cell lines. PPARgamma agonists (troglitazone and 15-deoxy-Delta(12,14)-prostaglandin J2) showed dose-dependent inhibitory effects on the proliferation of the gastric cancer cells, and their effect was augmented by the simultaneous addition of 9- cis retinoic acid, a ligand of RXRalpha. Flow cytometry demonstrated G1 cell cycle arrest and a significant increase of annexin V-positive cells after treatment with troglitazone. These results suggest that induction of apoptosis together with G1 cell cycle arrest may be one of the mechanisms of the antiproliferative effect of PPARgamma activation in human gastric cancer cells.

Topics: Apoptosis; Cell Cycle; Cell Division; Flow Cytometry; Gene Expression Regulation, Neoplastic; Humans; Ligands; Receptors, Cytoplasmic and Nuclear; Stomach Neoplasms; Transcription Factors; Tretinoin; Tumor Cells, Cultured

To determine the mechanism of all-trans retinoic acid (ATRA) on growth inhibition in human gastric cancer cells.. Gastric cancer cell lines: MGC80-3, BGC-823, SGC-7901 and MKN-45. CAT assay, Northern blot, Western blot, gene transfection and MTT assay.. ATRA can inhibit the activator protein-1 (AP-1) activity in ATRA-sensitive cell lines, but not in ATRA-resistant cell line, and the anti-AP-1 activity of ATRA is mediated by its receptor, retinoic acid receptor alpha (RAR alpha). ATRA can also inhibit the expression of cJun and cFos. One of the mechanisms for ATRA to inhibit the growth of gastric cancer cells may be through its inhibitory effect on the AP-1 activity and its influence on up-regulation of RAR alpha expression. The inhibition of cJun and cFos expressions by ATRA may also contribute to the anti-AP-1 activity.. ATRA inhibits the growth of gastric cancer cells through the regulation of AP-1 activity. This action is mediated by RAR alpha.

Topics: Antineoplastic Agents; Cell Division; Humans; Stomach Neoplasms; Transcription Factor AP-1; Tretinoin; Tumor Cells, Cultured

To study the mechanism of retinoic acid receptor alpha(RAR alpha) in mediating growth inhibition of gastric cancer cells by all-trans retinoic acid(ATRA).. Expression of RAR alpha was detected by Northern blot. After anti-sense RAR alpha or sense RAR alpha had been transfected into gastric cancer cell lines BGC-823 and MKN-45 respectively, the inhibitory effect of ATRA on cell growth in stable clones was analyzed using MTT assay and colony forming assay in soft agar. The transcriptional activation of retinoic acid response element (RARE) was measured by CAT assay.. ATRA could induce expression of RAR alpha in BGC-823 cells, but not in MKN-45 cells. In stable clones, ATRA could inhibit growth of MKN-45 cells transfected with RAR alpha gene, but could not inhibit that of BGC-823 cells transfected with antisense RAR alpha. Transient transfection and CAT assay showed higher beta-RAR response element (beta RARE) transcriptional activation induced by ATRA in MKN-45 cells transfected with RAR alpha compared to parental MKN-45 cells, while lower beta RARE transcriptional activation was seen in BGC cells transfected with antisense RAR alpha gene compared to parental BGC-823 cells.. Sufficient level of RAR alpha is required for growth inhibition of gastric cancer cells by ATRA.

Topics: Antineoplastic Agents; Cell Division; DNA, Antisense; Drug Interactions; Humans; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoic Acid Receptor gamma; Stomach Neoplasms; Transfection; Tretinoin; Tumor Cells, Cultured; Tumor Stem Cell Assay

In this study, the biological effects and molecular mechanism of recombinant RA538 and antisense c-myc adenovirus on human gastric, esophageal, 2BS and high-expression bcl-2 gene cancer cell lines were studied in vitro and in vivo. The results were as follows: Ad-RA538 and Ad-ASc-myc could strongly inhibit cell growth and induce apoptosis of SGC7901 cells in vitro and in vivo, and could down-regulate expression of c-myc, bcl-2 and cyclinD1 gene, up-regulate expression of bax gene. Ad-RA538 or Ad-AS c-myc could not inhibit cell growth and induce apoptosis changes of EC109, EC8712, 2BS and high-expression bcl-2 gene cancer cell lines, and could not down-regulate expression of c-myc and bcl-2 gene. The results indicated that: Ad-RA538 or Ad-AS c-myc can inhibit growth and induce apoptosis of gastric cancer cell in vitro and in vivo. They relate to c-myc, bcl-2, cyclinD1 and bax gene closely and play a key role on biologic effects in gastric cancer cells. Ad-RA538 and Ad-AS c-myc could not produce relevant changes on esophageal cancer, 2BS and high-expression bcl-2 gene cell lines.

Topics: Adenoviruses, Human; DNA, Antisense; Esophageal Neoplasms; Humans; Proto-Oncogene Proteins c-bcl-2; Proto-Oncogene Proteins c-myc; Recombinant Proteins; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

Retinoic acid can induce growth inhibition and apoptosis, and regulate cell cycle in many types of cancer cell lines. In this study, we investigated the role of all-trans retinoic acid (ATRA) and its mechanism of action in human gastric cancer cell lines. Our results demonstrated that ATRA effectively inhibited growth in three of four gastric cancer cell lines by induction of G0/G1 arrest, and did not induce apoptosis in four gastric cancer cell lines. In RA-sensitive cell lines, ATRA-induced G0/G1 arrest is associated with down regulaton of c-myc and hyperphosphorylated Rb expression, and up regulation of p21WAF1/CIP1 and p53 expression. There were no significant changes in cyclin D1 or CDK4 expression induced by ATRA. Futhermore, expression of these genes were not regulated by ATRA in ATRA-resistant gastric cancer cell line. These results indicate that growth inhibition, rather than apoptosis, is correlated with G0/G1 arrest of these cell lines, more important molecules related cell cycle, including c-myc, p21WAF1/CIP1, p53 and Rb, are involveed in regulation of cell cycle in gastric cancer cells.

Topics: Antineoplastic Agents; Apoptosis; Cell Cycle; Cell Division; Humans; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

The effect of prolonged administration of all-trans-retinoic acid (RA) on sodium chloride-enhanced gastric carcinogenesis induced by N-methyl-N'-nitro-N-nitrosoguanidine, and the labelling and apoptotic indices and immunoreactivity of transforming growth factor (TGF) alpha in the gastric cancers was investigated in Wistar rats. After 25 weeks of carcinogen treatment, the rats were given chow pellets containing 10% sodium chloride and subcutaneous injections of RA at doses of 0.75 or 1.5 mg kg(-1) body weight every other day. In week 52, oral supplementation with sodium chloride significantly increased the incidence of gastric cancers compared with the untreated controls. Long-term administration of RA at both doses significantly reduced the incidence of gastric cancers, which was enhanced by oral administration of sodium chloride. RA at both doses significantly decreased the labelling index and TGF-alpha immunoreactivity of gastric cancers, which were enhanced by administration of sodium chloride, and significantly increased the apoptotic index of cancers, which was lowered by administration of sodium chloride. These findings suggest that RA attenuates gastric carcinogenesis, enhanced by sodium chloride, by increasing apoptosis, decreasing DNA synthesis, and reducing TGF-alpha expression in gastric cancers.

Topics: Adenocarcinoma; Animals; Antineoplastic Agents; Apoptosis; Carcinogens; Dietary Supplements; Drug Synergism; Gastric Mucosa; Male; Methylnitronitrosoguanidine; Mitotic Index; Rats; Rats, Wistar; Sodium, Dietary; Stomach Neoplasms; Transforming Growth Factor alpha; Tretinoin

In order to better understand the mechanisms that underlie the antiproliferative effect of retinoids, we have examined the response of human carcinoma cell lines to all-trans retinoic acid (RA) and N-(4-hydroxyphenyl) retinamide (4HPR) in terms of cell growth, apoptosis and regulation of retinoic acid receptors (RARs) and retinoid X receptors (RXRs) mRNA. GLC82 (lung adenocarcinoma), BGC823 (stomach adenocarcinoma) and EC109 (esophageal squamous carcinoma) cells were treated with 10 microM of RA or 4HPR for various length of time and analyzed. The results show that growth inhibition by RA and 4HPR in GLC82 and BGC823 cells correlates with the induction of RARbeta2 gene, whereas RA resistance in EC109 cells parallels loss of RARbeta2 induction. Exogenous RARbeta2 expression did not restore RA responsiveness in EC109 cells, but potentiated 4HPR-induced growth inhibition, suggesting that 4HPR acts at least in part via the RARbeta receptor. We speculate that the loss of RARbeta2 inducibility in EC109 cells may be due to an unknown repressor.

Topics: Adenocarcinoma; Antineoplastic Agents; Cell Division; Drug Resistance, Neoplasm; Esophageal Neoplasms; Fenretinide; Humans; Lung Neoplasms; Neoplasms; Receptors, Retinoic Acid; Retinoid X Receptors; Stomach Neoplasms; Transcription Factors; Transfection; Tretinoin; Tumor Cells, Cultured

Induction of apoptosis has been implicated as an anticarcinogenic mechanism of both folic acid and retinoic acid. The ability of retinoic acid or folic acid to induce gastric cancer cell apoptosis was investigated in the human gastric cancer cell lines MKN-45 and MKN-28, and DNA fragmentation was studied in situ by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling and DNA agarose gel electrophoresis. The rates of apoptosis in both the poorly differentiated MKN-45 and the well differentiated MKN-28 cell line were less than 5% after treatment with either retinoic or folic acid. Apoptosis may be induced by the administration of retinoic acid or folic acid, and the apoptosis indices of MKN-45 and MKN-28 cells were related to the doses of these drugs. The induction of gastric cancer cell apoptosis may play a role in the anticarcinogenic effect of retinoic acid and folic acid, both of which are potential agents for the treatment of human gastric cancer.

Topics: Antineoplastic Agents; Apoptosis; Electrophoresis, Agar Gel; Folic Acid; Hematinics; Humans; In Situ Nick-End Labeling; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

All-trans retinoic acid (RA) was previously shown to regulate the growth of gastric cancer cells derived from the cell line SC-M1. This study was designed to investigate the effect of RA on the sensitivity of SC-M1 cells to lymphokine-activated killer (LAK) activity. RA at the concentration range of 0.001-10 microM was shown to induce SC-M1 cells to exhibit resistance to LAK activity in a dose-dependent manner. A kinetics study indicated that a significantly increased resistance was detected after 2 days of co-culturing SC-M1 cells with RA and reached a maximum after 6 days of culture. Similar results were obtained from two other cancer cell lines: promyelocytic leukaemia HL-60 and hepatic cancer Hep 3B. A binding assay demonstrated that the binding efficacy between target SC-M1 cells and effector LAK cells was not altered by RA. Flow cytometric analyses revealed that RA exhibited no effect on the expression of cell surface molecules, including HLA class I and class II antigens, intercellular adhesion molecule-1 and -2, and lymphocyte function antigen-3. Cell cycle analysis revealed that culture of SC-M1 cells with RA resulted in an increase in G0/G1 phase and a decrease in S phase, accompanied by a decrease in cyclin A and cyclin B1 mRNA as determined by Northern blot analysis. Additionally, RA was shown to enhance the expression of retinoic acid receptor alpha (RAR alpha) in SC-M1 cells, and to have no effect on the expression of RARbeta or RARgamma. Taken together, these results indicate that RA can significantly increase gastric cancer cells SC-M1 to resist LAK cytotoxicity by means of a cytostatic effect through a mechanism relating to cell cycle regulation. The prevailing ideas, such as a decrease in effector to target cell binding, a reduced MHC class I antigen expression or an altered RARbeta expression, are not involved.

Topics: Antineoplastic Agents; Binding Sites; Blotting, Northern; Cell Adhesion Molecules; Cell Cycle; Cyclins; Cytotoxicity, Immunologic; Dose-Response Relationship, Drug; Flow Cytometry; HLA Antigens; Humans; Immunotherapy, Adoptive; Killer Cells, Lymphokine-Activated; Leukemia; Liver Neoplasms; Receptors, Retinoic Acid; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

The effect of 9-cis-retinoic acid (9-cis-RA) on the growth of eight gastric cancer cell lines was related to their transcription levels of mRNAs for retinoid receptors. Northern blot analysis showed that seven (TMK-1, MKN-1, -28, -45, -74, HSC-39, KATO-III) out of eight gastric cancer cell lines synthesized mRNAs for retinoic acid receptors (RARs) and retinoid X receptor-alpha (RXR-alpha). MKN-7 cells did not transcribe either RARs or RXR-alpha at the mRNA level although they appeared to have no alterations at the gene level. The growth of all of the cell lines except for MKN-7 cells was inhibited by 1 x 10(-6) M 9-cis-RA. Cell cycle distribution analysis revealed that G0-G1 arrest was not induced by exposure to 9-cis-RA in the sensitive TMK-1 and KATO-III cells or the resistant MKN-7 cells. Interestingly, 9-cis-RA temporarily increased the amount of the cyclin dependent kinase (cdk) inhibitor, Waf1/Cip1/Sdi1/p21 protein, and also reduced the amount of cdk-7, epidermal growth factor receptor (EGFR) and cyclin D1 proteins, followed by reduction in phosphorylation of the product of the retinoblastoma tumor suppressor gene (Rb) in the sensitive TMK-1 cells, but not in the resistant MKN-7 cells. These results suggest that 9-cis-RA has a cytostatic effect on gastric cancer cells that synthesize the receptor molecules through cell cycle regulatory machinery.

Topics: Adenocarcinoma; Alitretinoin; Antineoplastic Agents; Cell Cycle; Cell Division; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Humans; Receptors, Retinoic Acid; Retinoid X Receptors; RNA, Messenger; Signal Transduction; Stomach Neoplasms; Transcription Factors; Tretinoin; Tumor Cells, Cultured

We found that vitamin D3 up-regulates the expression of cytidine deaminase (CDD) gene in some human solid tumor cell lines as well as the monocytic leukemia cell lines. Two kinds of full length CDD cDNA were identified from human placenta: one has glutamine and the other one has lysine at codon 27. The expression was tested in various normal tissues and the cancer cell lines. Northern blot analysis demonstrated high levels of CDD mRNA in leukocytes and moderate levels in liver, kidney, placenta, spleen and lung. Expression of CDD in 20 human cancer cell lines was highly variable and not related to its expression in normal tissues. Treatment of the cell lines with 1 alpha,25-dihydroxyvitamin D3 resulted in up-regulation of CDD expression in some lines but not others. Three of five gastric carcinoma cell lines and five of eight colorectal cancer lines had increased levels of CDD mRNA following 24 h treatment with vitamin D3. Increased mRNA was detected in gastric cancer MKN 45 cells after 3 h of treatment with vitamin D3 and increased enzyme activity was measured after 24 to 48 h. But no combined effect of calcitriol with 9-cis retinoic acid was found. Our results demonstrate that CDD can be up-regulated by vitamin D3 in some solid tumor cell lines.

Topics: Base Sequence; Blotting, Northern; Calcitriol; Cell Line; Cloning, Molecular; Cytidine Deaminase; DNA Primers; Escherichia coli; Gene Expression Regulation, Neoplastic; Humans; Molecular Sequence Data; Recombinant Fusion Proteins; RNA, Messenger; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured; Up-Regulation

Vitamin A and its analogus, the retinoids, are agents that are known to induce differentiation and inhibit growth on cell in vitro and in vivo. These agents show promise as prophylactic and therapeutic ones in human cancer and were noted extensively by scholars abroad, but the research has just been developed in recent years in our country. Retinamide (named briefly R II), a kind of new retinoids made in China, can induce differentiation and inhibit growth of cell, but its toxic effect is smaller than its maternal chemical compound (retinoic acid). Using automatic image analysis technology, polyacrylamide gel LDH isoenzyme electrophoresis, radioimmunoassay and condensing reaction of Con A, we have observed induction of differentiation of the R II on SGC-7901 cells. Our experiments indicate that R II can inhibit cell proliferation, decrease obviously average DNA content and nuclear area, reduce CEA secretion and condensing of the cells, and change LDH isozymoraphy of SGC-7901 cell with promoting H-type LDH and reducing M-type LDH. The change of morphology under light microscope by R II showed that the cells spread, flattened and partially lined up, and the alkalophilic quality of cytoplasm became weak. These results suggest that SGC-7901 cell under the action of R II should undergo the changes of reverse differentiation in morphological, physiological and bio-chemical aspects.

Topics: Antineoplastic Agents; Carcinoembryonic Antigen; Cell Transformation, Neoplastic; DNA, Neoplasm; Humans; L-Lactate Dehydrogenase; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

Retinoic acid has been recognised as a pivotal compound in cell differentiation, proliferation and malignant transformation. We investigated the effects of all-trans-retinoic acid on cell growth and the expression of retinoid nuclear receptor mRNAs in gastric cancer cells in vitro. Cell growth was quantified by measuring total cellular DNA. The growth of two of the five gastric cancer cell lines tested (SC-M1 and TSGH9201) was inhibited by all-trans-retinoic acid at concentrations ranging from 1 x 10(-8) M to 1 x 10(-6) M. Growth inhibition was associated with G0/G1 phase arrest as determined by flow cytometric analysis. Northern blot analysis showed that all five cell lines expressed mRNA for retinoic acid receptors alpha and retinoic x receptor alpha and beta. Retinoic acid receptor beta mRNA was only expressed in TSGH9201 and TMK-1 gastric cancer cell lines. Two RAR gamma mRNA transcripts (3.2 and 3.0 kb) were detected in SC-M1 and TSGH9201 cells. RA-resistant cells had markedly decreased levels of the 3.2 kb RAR gamma transcript. All-trans-retinoic acid had a cytostatic effect on the growth of some gastric cancer cells, which may be associated with the expression of retinoic acid receptors.

Topics: Cell Cycle; DNA, Neoplasm; Dose-Response Relationship, Drug; Humans; Receptors, Retinoic Acid; RNA, Messenger; Stomach Neoplasms; Time Factors; Tretinoin; Tumor Cells, Cultured

4-hydroxycarbophenyl retiamide (R II) is a new synthetic analogue of retinol, but with lower toxicity than retinoic acid. We studied its induction effects and its effects on some malignant phenotypes of the MFC cell line. The mechanism of these effects was also explored. MFC cells were grown in complete RPMI 1640 medium supplemented with 10(-5) mol/L R II for five passages. By then the cell growth rate slowed down; the rate of 3H-TdR incorporation and the colony-forming capacity of MFC cells decreased; morphologically, the cells became epithelial rather than fibroblastic with various degrees of polarization. Further investigation about the mechanism of these changes was also undergone. First by flow cytometry, it was shown that the R II-treated cells were retained in G1 phase. Second, dot blot hybridization showed a decrease of more than 61% of c-myc mRNA and an increase of more than 52% of v-fos mRNA. The major chromosome distribution changed from 54-56 to 46-54 with an increase in diploid. Scanning microscopic examination showed that the R II-treated cells were covered by numerous microvilli and pseudopodia with round terminal expansion in contrast to the ruffle protrusions and leaf-like pseudopodia of control cells. All the results suggested that R II could reverse some malignant phenotypes of MFC cells.

Topics: Animals; Antineoplastic Agents; Cell Transformation, Neoplastic; Gene Expression Regulation, Neoplastic; Genes, fos; Genes, myc; Mice; Mice, Inbred Strains; Phenotype; RNA, Messenger; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

The growth rate and weight of transplanted tumor of SGC7901 gastric cancer cell line, treated with 10 mumol/L retinoic acid (RA) in nude mice, were significantly reduced in comparison with those of control cells. The morphology of RA-treated cells was changed from ellipse shape with meandering margin of a large nucleus to shuttle shape with smooth margin of a small nucleus. With flow cytometry, it was found that G/O Phase markedly increased after the RA treatment, whereas S and G2 + M phase decreased. Especially the S phase fraction was lower than that of control cells. The above results indicated that RA could reverse biological and morphological phenotypes of the human gastric cancer cells in vitro and could induce differentiation of gastric cancer cells to a certain extent.

Topics: Animals; Cell Transformation, Neoplastic; Flow Cytometry; Humans; Interphase; Mice; Mice, Nude; Neoplasm Transplantation; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

The long-term effects of butylated hydroxyanisole (BHA), in combination with various other chemicals on the development of forestomach lesions in rats were investigated. BHA is a synthetic antioxidant, and the other agents included the glutathione-depleting agent diethylmaleate (DEM), the anti-inflammatory drugs indomethacin (IM), dexamethazone (DEX), 6-aminocaproic acetate (6-ACA) and FOY (gabexate mesilate), and the vitamin all-trans-retinol acetate (RA). Concurrent treatment with BHA (1% in diet) and DEM, IM, DEX or FOY for 52 weeks inhibited development of forestomach epithelial hyperplasia as compared to BHA alone, while simultaneous treatment with RA enhanced hyperplastic development. However, the inhibition by DEX or FOY was only partial and in the DEX case, in particular, might have been due to weight loss. Since the most effective inhibitory influence on BHA-induced forestomach lesions exerted in this 1-year experiment was by DEM, a further 2-year experiment was conducted to confirm whether DEM actually can exert inhibitory effects on BHA (2% in diet)-induced forestomach carcinogenesis. The results demonstrated that induction of forestomach hyperplasias and papillomas by BHA was significantly reduced by combination treatment with DEM. Both multiplicity and incidence of forestomach papillomas were significantly decreased, while squamous cell carcinoma development showed a tendency for decrease only.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Butylated Hydroxyanisole; Dexamethasone; Drug Antagonism; Hyperplasia; Male; Maleates; Papilloma; Rats; Rats, Inbred F344; Stomach; Stomach Neoplasms; Tretinoin

We have previously reported that a glycolipid GalNAc beta 1-4[NeuAc alpha 2-3]Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc-Cer named NGM-1 is present specifically in the human gastric fundic mucosa, but not in other organs. In gastric-cancer tissue and cancer cell lines, this glycolipid completely disappears. These findings imply that NGM-1 is expressed only in well-differentiated fundic mucosa. The purpose of this study is to examine the expression of NGM-1 as a differentiation-related molecule. A gastric cell line AZ521 was cultured in the medium with various reagents which had been reported to induce differentiation in cancer cells. The growth of AZ521 was suppressed by the addition of 0.8% dimethylformamide (DMF) to the medium, but not by addition of dimethylsulfoxide (DMSO), retinoic acid or butyric acid. In the ganglioside fraction of the cells cultured with DMF, a glycolipid regarded as NGM-1 which had not been present before treatment was detected using a monoclonal antibody. Suppression of the proliferation of AZ521 by eliminating the serum from the medium could not induce the expression of NGM-1. A colonic cell line treated with DMF also failed to express the glycolipid. The synthase activity of NGM-1 was elevated in the AZ521 cells treated with DMF, but not with DMSO. These results demonstrate that the expression of NGM-1 is induced by DMF specifically in gastric-cancer cells, and suggest the possibility that NGM-1 is a differentiation-related molecule.

Topics: Butyrates; Butyric Acid; Cell Differentiation; Chromatography, Thin Layer; Dimethyl Sulfoxide; Dimethylformamide; Gastric Fundus; Gastric Mucosa; Glycoproteins; Humans; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

Histamine dose-dependently stimulated cyclic AMP production in human gastric carcinoma cell line MKN-45, and this effect was inhibited by cimetidine but not by pyrilamine. Moreover, not only histamine but also cimetidine displaced the specific binding of [3H]tiotidine to these cells, whereas pyrilamine had no effect. On the other hand, pretreatment of MKN-45 cells with retinoic acid (RA) significantly enhanced histamine-induced increase of cyclic AMP production, although the cyclic AMP response to either forskolin or NaF was not affected. Finally, RA treatment increased the number of histamine receptor without altering its affinity. Thus, it appears that histamine H2-receptors are present on MKN-45 cells, and that RA treatment enhances the action of histamine on these cells by increasing the number of H2-receptors.

Topics: Binding, Competitive; Cell Line; Cimetidine; Cyclic AMP; Histamine; Histamine H2 Antagonists; Humans; Kinetics; Receptors, Histamine H2; Stomach Neoplasms; Tretinoin

Two new retinoic acid esters and retinamides synthesized in China, N-(4-ethoxycarbophenyl)retinamide (RI) and N-(4-carboxyphenyl)retinamide (RII), significantly inhibited carcinogenesis induced in the epithelium of the forestomach of mice by N-nitrososarcosine ethyl ester. RI also markedly inhibited carcinogenesis induced in the epithelium of the oesophagus and forestomach in rats by this ester. No sign of hypervitaminosis was noticed with doses as high as six times the therapeutic dose. RI also inhibited precancerous and cancerous lesions in the nasal cavity and nasopharynx and oesophagus of rats induced by dinitrosopiperazine. In a malignant oesophageal epithelial cell line from rats, RE25-3, established in our laboratory, RI and RII inhibited mitosis, proliferation rate, chromosomal aberrations and incorporation of 3H-thymidine into DNA. The ability to form colonies on agar plates was also inhibited by these two compounds.

Topics: Animals; Antineoplastic Agents; Carcinogens; Esophageal Neoplasms; Female; Mice; Nitrosamines; Rats; Rats, Inbred Strains; Stomach Neoplasms; Tretinoin; Tumor Cells, Cultured

Carcinogenic and promoting effects of RRME as isolated from the pickled vegetables in Linxian County, a high incidence area of esophageal cancer, were studied in mice and rats. RRME alone did not cause tumor in the forestomach of mice and esophagus of rats. When the mice were intubated with a single dose of nitroso-sarcosine-ethylester (NSEE), the incidence of the forestomach carcinoma was only 9.5%. However, when the mice were given gastric doses of RRME after one single dose of NSEE, the incidence was increased to 41.0%. In rats, the tumor incidence was 5.3% in nitroso-methylbenzylamine (NMBzA) group, while in NMBzA kRME group, it was 20.7%. In rats intubated with NSEE for 7 times, no carcinoma appeared in esophageal epithelium; while followed by gastric doses of RRME, the incidence of esophagus carcinoma increased up to 63.2%. The experimental results show that RRME has distinct promoting effect on the process of cocarcinogenesis initiated by NSEE and NMBzA in the forestomach of mice and esophagus of rats, but without carcinogenic effect itself. Retinamide (RI) and massive dose of vitamin C showed an obviously inhibitory effect on promoting action of RRME in rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Antineoplastic Agents; Ascorbic Acid; Dimethylnitrosamine; Esophageal Neoplasms; Female; Mice; Nitroso Compounds; Rats; Rats, Inbred Strains; Stomach Neoplasms; Tretinoin

Topics: Animals; Antineoplastic Agents; Female; Mice; Retinoids; Stomach Neoplasms; Tretinoin

Carcinogenesis induced by 7,12-dimethylbenz[a]anthracene (DMBA) and 12-O-tetradecanoylphorbol 13-acetate (TPA) in the forestomach epithelium of mice was delayed by ethyl all-trans-9-(4-methoxy-2,3,6-trimethylphenyl)-3,7-dimethyl-2,4,6,8-nonatetraenoate (etretinate). Beside the clear inhibitory effect during the promotion phase, however, considerable side effects occurred when administering effective doses.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Body Weight; Carcinogens; Epithelium; Etretinate; Female; Mice; Mice, Inbred C57BL; Neoplasms, Experimental; Stomach Neoplasms; Tetradecanoylphorbol Acetate; Tretinoin

The pathogenesis of papillomas which developed spontaneously in the forestomach of WB X C57BL/6 F1-W/Wv mutant mice was investigated. The thickness of the forestomach epithelium was used as a quantitative index for development of papillomas. From the 15th day after birth, the forestomach epithelium of the W/Wv mice was significantly thicker than that of the congenic +/+ mice. Administration of aromatic retinoic acid analog (ethyl all-trans-9-(4-methoxy-2,3,6-trimethylphenyl)-3,7-dimethyl-3,7-dimethyl-2,4,6,8 -nonatetraenoate) did not suppress development of papillomas. Since papillomas did not appear in the stomach which was removed from the W/Wv embryos and grafted to the s.c. space produced in the back of the adult mice and since a considerable amount of bile reflux preceded development of papillomas, bile reflux may be a cause of papillomas in W/Wv mice.

Topics: Alleles; Animals; Bile Reflux; Biliary Tract Diseases; Etretinate; Female; Genotype; Male; Mice; Mutation; Papilloma; Stomach; Stomach Neoplasms; Tretinoin

Topics: Animals; Humans; Neoplasms; Neoplasms, Experimental; Organ Culture Techniques; Papilloma; Rats; Skin Neoplasms; Stomach Neoplasms; Structure-Activity Relationship; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Antineoplastic Agents; Female; Mice; Neoplasms, Experimental; Stomach Neoplasms; Tretinoin