sincalide and Stomach-Neoplasms

To investigate the inhibiting effects of multi-target anti-microRNA antisense oligonucleotide (MTg-AMOs) on proliferation and migration of human gastric cancer cells.. Single anti-microRNA antisense oligonucleotides (AMOs) and MTg-AMOs for miR-221, 21, and 106a were designed and transfected into SGC7901, a gastric cancer cell line, to target the activity of these miRNAs. Their expression was analyzed using stem-loop RT-PCR and effects of MTg-AMOs on human gastric cancer cells were determined using the following two assay methods: CCK8 for cell proliferation and transwells for migration.. In the CCK-8 cell proliferation assay, 0.6 μmol/L was selected as the preferred concentration of MTg-AMOs and incubation time was 72 hours. Under these experimental conditions, MTg-AMOs demonstrated better suppression of the expression of miR-221, miR-106a, miR-21 in gastric cancer cells than that of single AMOs (P = 0.014, 0.024; 0.038, respectively). Migration activity was also clearly decreased as compared to those in randomized and blank control groups (28 ± 4 Vs 54 ± 3, P <0.01; 28 ± 4 Vs 59 ± 4, P < 0.01).. MTg-AMOs can specifically inhibit the expression of multiple miRNAs, and effectively antagonize proliferation and migration of gastric cancer cells promoted by oncomirs.

Topics: Cell Growth Processes; Cell Line, Tumor; Cell Movement; Gene Expression Regulation, Neoplastic; Humans; MicroRNAs; Oligodeoxyribonucleotides, Antisense; Sincalide; Stomach Neoplasms; Transfection

Gastric cancer (GC) remains a common cause of cancer death in East Asia. Current treatment strategies for GC, including medical and surgical interventions, are suboptimal. Butyrate, a short-chain fatty acid produced by the intestinal flora, has been reported to be able to inhibit gastric carcinogenesis. This study aimed to investigate the effects of butyrate on human GC and its underlying mechanisms.. Human GC cell lines BGC-823 and SGC-7901, human GC tissues and adjacent normal tissues were used for this study. Cell proliferation was assessed using CCK-8 and EdU staining. TUNEL fluorescence and Annexin V/PI staining were adopted for qualitative and quantitative evaluation of cell apoptosis, respectively. Reactive oxygen species (ROS) assay was performed to analyse mitochondrial function. Real-time q-PCR and western blot were carried out to examine the expression of apoptosis-related genes and the synthesis of apoptosis-related proteins. The association between G protein-coupled receptor 109a (GPR109a) and GC prognosis was analyzed using data from The Cancer Genome Atlas (TCGA).. CCK-8 and EdU staining confirmed inhibitory activities of butyrate against human GC cells. Annexin V/PI staining and TUNEL fluorescence microscopy showed that butyrate promoted GC cell apoptosis. No difference in the expression of GPR109a was found between GC tissues and adjacent normal tissues, and no direct association between GPR109a and GC prognosis was discovered, suggesting that GPR109a may not be a key factor mediating the apoptosis of GC cells. Butyrate increased the synthesis of caspase 9 and decreased BCL-2, the well-known effector and regulator of mitochondria-mediated apoptosis, and significantly induced mitochondrial ROS.. Collectively, our results suggest that butyrate is able to inhibit the proliferation of GC cells and induce GC apoptosis, possibly via a mitochondrial pathway.

Topics: Annexin A5; Apoptosis; Butyrates; Cell Line, Tumor; Cell Proliferation; Humans; Reactive Oxygen Species; Sincalide; Stomach Neoplasms

In addition to its established role in regulating circadian rhythms and reducing inflammation, melatonin has been demonstrated to possess anti-cancer properties. In this study, we investigated the effects of exosomal miRNAs released by melatonin-treated GC cells on gastric cancer.. To identify the potential exosomal miRNAs involved in the treatment of gastric cancer, we performed exosome small RNA sequencing (sRNA-seq) to screen significant changes in 34 exosomal miRNAs in AGS cells before and after melatonin treatment. CCK-8, wound healing, and transwell invasion assays were used to examine the effects of miRNAs on cancer characteristics. Furthermore, a dual-luciferase reporter gene assay was performed to identify the miRNA targets.. Exosomal miR-27b-3p was down-regulated by approximately 1.37-fold following melatonin treatment. The CCK-8 assay revealed a significant increase in cell proliferation in the miR-27b-3p mimic group compared to that in the miR-27b-3p mimic NC group. In the wound healing assay, cells treated with miR-27b-3p mimics displayed significantly more rapid wound closure than that observed in the miR-27b-3p mimic NC group. The transwell invasion assay revealed a substantial increase in the number of invading cells in the miR-27b-3p mimic group compared to that in the miR-27b-3p mimic NC group. Additional analysis revealed that miR-27b-3p directly targets ADAMTS5 and that its up-regulation results in increased proliferation, invasion, and metastasis of GC cells.. Melatonin suppressed the progression of gastric cancer by regulating the exosomal miR-27b-3p-ADAMTS5 pathway. Thus, melatonin represents a promising potential therapeutic agent for patients with gastric cancer.

Topics: Cell Line, Tumor; Cell Proliferation; Exosomes; Humans; Melatonin; MicroRNAs; Sincalide; Stomach Neoplasms

LncRNA ubiquitin-like with PHD and RING finger domains 1 (UHRF1) protein associated transcript (UPAT) regulates the progression of many cancers. However, its role in gastric cancer (GC) is less frequently reported.. In the context of the promoting effect of lncRNA on modulating GC progression, detailed insights into the role and underlying mechanism of UPAT in GC are the foothold in this study.. Overall survival was calculated. The mRNA expressions of UPAT and UHRF1 were measured by qRT-PCR, and the protein expressions of UHRF1, Cyclin D1 and cleaved caspase-3 were determined by western blot. Cell viability, growth, migration and invasion were assessed by CCK-8, colony formation, wound healing and Transwell assays, respectively. Apoptosis rate and cell cycle were assayed by flow cytometry.. UPAT was overexpressed in GC tissue and cell lines. Decreased UPAT level was associated with higher overall survival. Down-regulation of UPAT diminished cell proliferation, Cyclin D1 expression, and migration and invasion rates, increased apoptosis rate and cleaved caspase-3 expression, and blocked cell cycle in AGS and NCI-N87 cells. UPAT expression in GC was positively correlated with UHRF1 expression. UHRF1 overexpression offset the inhibitory effects of UPAT down-regulation on cell proliferation, migration, invasion and cell cycle, and partially reversed the positive effect of UPAT down-regulation on apoptosis.. UPAT might positively regulate the progression of GC via interacting with UHRF1. The UHRF1/UPAT axis revealed in the present study may provide a promising approach to intervene in the progression of GC.

Topics: Caspase 3; CCAAT-Enhancer-Binding Proteins; Cell Line, Tumor; Cyclin D1; Humans; RNA, Long Noncoding; RNA, Messenger; Sincalide; Stomach Neoplasms; Ubiquitin-Protein Ligases; Ubiquitins

ATP-binding cassette E1 (ABCE1) is mainly related to the regulation of viral infection, cell multiplication, and anti-apoptosis. Previous reports confirmed the central role in the regulation of ABCE1 in liver and breast cancer; however, its potential role in gastric adenocarcinoma remains unclear. In our study, siRNA and plasmid were transfected to construct gastric cancer cell lines with low and overexpression of ABCE1, and Western blot, RT-qPCR, and immunohistochemical staining were used to detect ABCE1 expression levels in gastric cancer tissues and cell lines. The effects of ABCE1 on cell growth, metastasis, invasion, cell cycle, and drug resistance were investigated using CCK-8 test, wound healing assay, and clone formation experiment. Functional experiments indicated that si-ABCE1 decreased the proliferation, metastasis, and invasion of gastric adenocarcinoma. Meanwhile, si-ABCE1 has significantly promoted EMT process and enhanced the sensitivity of paclitaxel and cisplatin. In vivo experiments also confirmed that si-ABCE1 group had significantly smaller tumors, and immunohistochemical staining results showed the tumor growth in si-ABCE1 group was reduced obviously. In summary, we found ABCE1 is considered as a crucial role in the evolution of gastric adenocarcinoma and could be a viable therapeutic target for the disease.

Topics: Adenocarcinoma; Adenosine Triphosphate; ATP-Binding Cassette Transporters; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cisplatin; Gene Expression Regulation, Neoplastic; Humans; Paclitaxel; RNA, Small Interfering; Sincalide; Stomach Neoplasms

MicroRNAs (miRNAs) have been reported to play an important role in tumor progression by regulating the expression of target genes.. This study attempted to verify the role of miR-711 in gastric cancer (GC) progression by in vitro and in vivo assays.. The expression of miR-711 in tumor tissues and cells was detected by real-time quantitative PCR (qRT-PCR). Expression of MiR-711 in NCI-N87 and SNU-1 cells was detected by FISH. We transfected GC cells with miR-711 mimics or inhibitors. The effects of miR-711 on the proliferation and metastasis of GC cells were detected by CCK-8, wound healing and transwell assays. Dual-luciferase reporter gene assay was used to verify the targeting relationship between miR-711 and CD44. Xenograft assays was used to verify the regulatory effect of miR-711 on tumor growth.. In GC tissues and cell lines, the expression of miR-711 was down-regulated when compare with adjacent tissues or normal epithelial cells. The results indicated that overexpressing of miR-711 could suppress the GC cell proliferation, migration, and invasion through targeting CD44. The knockdown of CD44 showed similar effects as miR-711 overexpression in GC cells. Moreover, we confirmed these effects in the in vivo assays. Furthermore, we found that miR-711 could play a role by influencing tumor cell stemness.. MiR-711 plays vital roles as a tumor-suppressor by targeting CD44 and may be a therapeutic target for GC treatment.

Topics: Cell Line, Tumor; Cell Movement; Cell Proliferation; Gene Expression Regulation, Neoplastic; Humans; Hyaluronan Receptors; Luciferases; MicroRNAs; Sincalide; Stomach Neoplasms

Circular RNAs (circRNAs) play pivotal roles in malignancies including gastric cancer (GC). We aimed to investigate the biological function and regulatory mechanism of circ_0006089 in GC.. Circ_0006089, microRNA (miR)-143-3p, and polypyrimidine tract-binding protein 3 (PTBP3) expressions were measured via quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) in GC cell lines. Cell proliferative capacity was determined by colony formation and CCK-8 assays. Flow cytometry was employed for measuring cell apoptosis. Cell invasion and migration were measured via transwell and wound-healing assays. Western blot analysis was utilized for detecting protein expressions of E-cadherin, N-cadherin, vimentin, PTBP3, PI3K, p-PI3K, AKT, and p-AKT. Dual-reporter luciferase analysis was conducted to confirm the association between miR-143-3p and circ_0006089 or PTBP3. The role of circ_0006089 in vivo was detected via establishing a mice xenograft model.. Circ_0006089 expression was increased in GC. Circ_0006089 downregulation suppressed the proliferation and metastasis and induced apoptosis of GC cells, which was counteracted by miR-143-3p inhibition or PTBP3 overexpression. In addition, circ_0006089 overexpression could promote GC progression. MiR-143-3p specially bound to circ_0006089 and PTBP3 was targeted by miR-143-3p. Moreover, circ_0006089 could regulate PTBP3 expression and the PI3K/AKT pathway by sponging miR-143-3p. Circ_0006089 knockdown also suppressed tumor growth.. Circ_0006089 regulated miR-143-3p/PTBP3/PI3K/AKT pathway to facilitate GC progression.

Topics: Animals; Cadherins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Gene Expression Regulation, Neoplastic; Humans; Mice; MicroRNAs; Phosphatidylinositol 3-Kinases; Polypyrimidine Tract-Binding Protein; Proto-Oncogene Proteins c-akt; RNA, Circular; Signal Transduction; Sincalide; Stomach Neoplasms; Vimentin

Gastric cancer is one of the most common and deadly cancers worldwide. Basic leucine zipper transcription factor ATF-like 3 (BATF3) plays a key role in tumor immunity. However, the function of BATF3 in gastric cancer remains unclear. Here, we demonstrated BATF3 positively regulated proliferation and radioresistance of gastric cancer cells by regulating S1PR1/STAT3 pathway.. The RNA-seq analyzed the gene expression by UALCAN web portal and Tumor Immune Estimation Resource. RT-qPCR and western blot was performed to verify BATF3 expression in gastric cancer cells. The assays of CCK-8, EdU incorporation and colony formation were used to analyze cell proliferation, and radioresistance in AGS and MKN45 cells. Flow cytometry was used to detect the cell apoptosis of AGS and MKN45 in treatment with si-BATF3 or radiation. Finally, western blot was performed to measure the expression of cell apoptosis-related modules including Bax, cleaved-caspase3, cleaved-PARP and assess the regulation of S1PR1/STAT3 pathway.. BATF3 expression was upregulated in gastric cancer cells. Knockdown of BATF3 suppressed proliferation, radioresistance but promoted the radiation-induced apoptosis of gastric cancer cells through positively regulating S1PR1 expression and STAT3 phosphorylation.. Knockdown of BATF3 inhibits gastric cancer cell growth and radioresistance via S1PR1/STAT3 pathway. BATF3 would become a potential diagnostic indicator for gastric cancer and target of therapeutic treatment.

Topics: Apoptosis; Basic-Leucine Zipper Transcription Factors; bcl-2-Associated X Protein; Cell Line, Tumor; Cell Proliferation; Cell Transformation, Neoplastic; Gene Expression Regulation, Neoplastic; Humans; Poly(ADP-ribose) Polymerase Inhibitors; Signal Transduction; Sincalide; Sphingosine-1-Phosphate Receptors; STAT3 Transcription Factor; Stomach Neoplasms

The antitumor effects of antiplatelet agents in gastric cancer cells are not well known. In this study, the possibility of gastric cancer treatment with an antiplatelet agent, mainly aspirin, was examined both in vivo and in vitro.. For in vivo experiments, tumor-bearing mice were treated by an antiplatelet antibody or aspirin, and the tumor growth was compared. For in vitro experiments, human gastric cancer cell lines were used to confirm the cancer cell growth and inhibition by reducing the platelet count or using aspirin. We also examined several cytokines by using an ELISA assay and conducted microRNA microarray analysis of MKN-45 tumor cells to determine the influence of platelets or aspirin.. In vivo experiments showed that tumor growth was inhibited by halving the circulating platelet count by using an antiplatelet antibody or peroral daily aspirin. In vitro experiments showed that the proliferation rates of gastric cancer cell lines were increased after coincubation with platelets and that the effect was inhibited by aspirin. Although the expression of interleukin-6, platelet-derived growth factor, transforming growth factor-β, and prostaglandin E2 did not correlate with tumor growth inhibition by aspirin, seven microRNAs showed altered expression in cancer cells in response to coincubation with platelets or addition of aspirin. Cells transfected with mir-4670-5p showed a significant increase in proliferation compared to negative control cells.. Our study showed that platelets increased the proliferation of gastric cancer cells and that this increase was inhibited by antiplatelet antibody or aspirin. Mir-4670-5p may play an important role in these responses.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Aspirin; Blood Platelets; Cell Proliferation; Humans; Immunoenzyme Techniques; In Vitro Techniques; Male; Mice; Mice, Inbred BALB C; Mice, Nude; MicroRNAs; Platelet Aggregation Inhibitors; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sincalide; Stomach Neoplasms; Tumor Cells, Cultured

To evaluate the inhibitory effect and mechanism of capecitabine metronomic chemotherapy on gastric cancer cells. In vitro, the effects of 5-fluorouracil (Fu) metronomic chemotherapy on proliferation, apoptosis, tube formation ability, and angiogenesis were detected. In vivo, Ki-67, CD34 and VEGF were detected by immunohistochemical staining (IHC). Flow cytometry was used to detect the percentage of circulating endothelial progenitors (CEPs), and VEGF and PDGF were detected by ELISA in the peripheral blood of nude mice. The proliferation of the SGC-7901 and AGS gastric cancer cell lines in the metronomic 5-Fu group was decreased compared with the control group in vitro. The total length of the small tubes and tubular junction numbers were significantly lower in the metronomic group than the control group. The VEGF and PDGF levels in the cell culture supernatants were lower in the metronomic group than the control group. Compared with the control group, the CEP percentage was decreased in the peripheral blood of tumor-bearing nude mice following treatment with metronomic 5-Fu or capecitabine chemotherapy. No significant changes were found in the conventional or control group. In the peripheral blood of tumor-bearing nude mice, the VEGF and PDGF levels were decreased in the metronomic groups. Metronomic 5-Fu inhibited the proliferation of gastric cancer cells in vitro and in vivo, and their antitumor effects were non-inferior to those of conventional dose chemotherapy, with mild side effects. Thus, tumor inhibition may be attributed to anti-angiogenesis.

Topics: Adenocarcinoma; Administration, Metronomic; Angiogenesis Inhibitors; Animals; Antimetabolites, Antineoplastic; Biomarkers, Tumor; Blood Cell Count; Capecitabine; Cell Line, Tumor; Culture Media; Drug Screening Assays, Antitumor; Endothelial Progenitor Cells; Fluorouracil; Human Umbilical Vein Endothelial Cells; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Morphogenesis; Platelet-Derived Growth Factor; Sincalide; Stomach Neoplasms; Vascular Endothelial Growth Factor A; Xenograft Model Antitumor Assays

Cholecystokinin (CCK) receptors are overexpressed in numerous human cancers, such as pancreatic, colon and gastric cancers. Previous studies have shown that the specific receptor-binding property of CCK for CCK receptors (CCKRs) can be exploited to produce immunotoxins (ITs) that target cancer cells overexpressing CCK receptors.. Construct a new IT-targeting CCKR-overexpressing colon cancers.. To construct the CCKR-targeted IT, a reverse CCK8 peptide was fused with a modified 38-kDa truncated form of the Pseudomonas exotoxin (PE38KDEL). An efficient immunoaffinity purification procedure was used to produce a PE38-based IT. Several analyses, including CCK8 competition and indirect immunofluorescence assays, were performed to confirm the interaction between rCCK8 and CCKR. After cytotoxic assays on several cell lines, the anti-tumor activity of the new IT was detected in nude mice.. The rCCK8PE38 IT showed specific cytotoxicity for two colon cancer cell lines and one gastric cancer cell line. After purification, 18-26 mg of pure rCCK8PE38 per 1 L of culture was obtained. Purified rCCK8PE38 showed high cytotoxicity in colon cancer cell lines with IC50 values of 0.8-3.5 ng/mL. The results of the CCK8 competition and indirect immunofluorescence assays showed that rCCK8 had a specific interaction with CCKR. Nude mice inoculated with HCT-8 tumor xenografts were treated with rCCK8PE38, which efficiently decreased the tumor size in those mice.. All of these data suggest that rCCK8PE38 has potential as a new immunotherapy agent. Furthermore, the results of this study further support the high value of the immunoaffinity method for IT purification procedures.

Topics: Animals; Cell Line, Tumor; Cholecystokinin; Colonic Neoplasms; Exotoxins; Female; Fluorescent Antibody Technique, Indirect; Humans; Immunotherapy; Immunotoxins; Mice; Mice, Nude; Peptide Fragments; Pseudomonas; Receptors, Cholecystokinin; Stomach Neoplasms; Xenograft Model Antitumor Assays

Cinobufacin is used clinically to treat patients with many solid malignant tumors. However, the mechanisms underlying action remain to be detailed. Our study focused on miRNAs involved in cinobufacin inhibition of GC cell proliferation. miRNA microarray analysis and real time PCR identified miR-494 as a significant cinobufacin- associated miRNA. In vivo, ectopic expression of miR-494 inhibited the proliferation and induced apoptosis of BGC-823 cells on CCK-8 and flow cytometry analysis. Further study verified BAG-1 (anti-apoptosis gene) to bea target of miR-494 by luciferase reporter assay and Western blotting. In summary, our study demonstrated that cinobufacin may inhibit the proliferation and promote the apoptosis of BGC-823 cells. Cinobufacin-associated miR-494 may indirectly be involved in cell proliferation and apoptosis by targeting BAG-1, pointing to use as a potential molecular target of cinobufacin in gastric cancer therapy.

Topics: 3' Untranslated Regions; Amphibian Venoms; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Proliferation; DNA-Binding Proteins; Humans; Medicine, Chinese Traditional; MicroRNAs; Sincalide; Stomach Neoplasms; Transcription Factors; Transfection

The aberrant expression of junctional adhesion molecule A (JAM-A), which has a close correlation with the development, progression, metastasis, and prognosis of cancer, has been frequently reported. However, neither JAM-A expression nor its correlation with clinicopathologic variables and patient survival has been defined in gastric cancers. Moreover, little is known about the role of JAM-A in gastric cancer progression. We carried out the present study to investigate the prognostic value of JAM-A expression in gastric cancer patients. Furthermore, the biological roles of JAM-A in gastric cancer progression were also investigated.. We determined JAM-A expression in 167 primary gastric cancer tissues and 94 matched adjacent non-tumor tissues by immunohistochemistry. Transwell migration assays and matrigel invasion assays were used to explore the role of JAM-A in gastric cancer cells migration and invasion. CCK-8 assays were used to examine the effect of JAM-A on the proliferation of gastric cancer cells.. JAM-A was downregulated in gastric cancer tissues. Low JAM-A expression was significantly associated with tumor size, lymphatic vessel invasion, lymph node metastasis, and TNM stage. Low JAM-A expression was also significantly associated with poor disease-specific survival in gastric cancer patients. Multivariate analysis demonstrated low JAM-A expression as an independent factor predicting poor survival. In addition, JAM-A had the effect on inhibition of gastric cancer cells migration and invasion. However, JAM-A had no significant effects on proliferation of gastric cancer cells.. Low JAM-A expression correlates with poor clinical outcome and promotes cell migration and invasion in gastric cancer.

Topics: Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Cell Adhesion Molecules; Cell Movement; Cell Proliferation; Disease Progression; Down-Regulation; Female; Gastrectomy; Humans; Immunohistochemistry; Kaplan-Meier Estimate; Lymph Node Excision; Lymph Nodes; Male; Middle Aged; Neoplasm Invasiveness; Predictive Value of Tests; Prognosis; Receptors, Cell Surface; Sincalide; Stomach Neoplasms; Young Adult

To examine whether the contractile response of gallbladder smooth muscle cells to cholecystokinin is reduced in patients with gallstones, smooth muscle cells were isolated from the gallbladders of patients either with or without gallstones and their direct contractile responses to cholecystokinin-octapeptide (CCK-8) were examined at physiological concentrations. The basal cell lengths were similar in patients with cholesterol gallstones (43 +/- 1.2 micron (SEM)), in patients with black pigment stones (42 +/- 1.1), and in the gallstone-free group (42 +/- 1.3). The contractile responses to CCK-8 at physiological concentrations ranging from 10(-13) to 10(-11) M were significantly greater in the gallstone patients compared with those in the gallstone-free patients, while the responses were similar at 10(-10) and 10(-9) M. No difference in contractile response to CCK-8 was found between the patients with cholesterol gallstones and those with black pigment stones. The effective dose of 50% was 1.6 x 10(-13) M in cholesterol gallstone patients, 3.2 x 10(-14) M in black pigment stone patients, and 1.0 x 10(-12) M in gallstone-free patients. The current in vitro study showed that the contractile responses of isolated smooth muscle cells of the gallbladder to CCK-8 are not impaired in patients with gallstones.

Topics: Analysis of Variance; Cholecystectomy; Cholelithiasis; Female; Gallbladder; Humans; In Vitro Techniques; Male; Middle Aged; Muscle Contraction; Muscle, Smooth; Sincalide; Stomach Neoplasms

Recently, we identified the specific binding site for gastrin on the gastric carcinoid tumor of Mastomys (Praomys) natalensis. In this study, precise characterization of the gastrin binding site on these tumors was performed. Both 125I-human gastrin I (gastrin) and 125I-CCK-8 bound specifically to the cell membrane, and Scatchard analysis revealed a high affinity binding site for each ligand with similar Kd and Bmax values. The specific binding of both 125I-gastrin and 125I-CCK-8 was displaced in a concentration-dependent manner by various related peptides with a relative potency order of CCK-8 > or = gastrin < des(SO3)CCK-8. In addition, L364,718 as well as L365,260 displaced the binding of both ligands with similar potencies. Furthermore, not only gastrin but also CCK-8 increased [Ca2+]i in these tumor cells, the action of both being inhibited by L364,718 as well as by L365,260 (10(-7) M). These results suggest that the carcinoid tumor of Mastomys possesses a high affinity gastrin/CCK binding site coupled to the increase of [Ca2+]i.

Topics: Animals; Carcinoid Tumor; Female; Gastrins; Humans; Kinetics; Male; Muridae; Neoplasm Transplantation; Receptors, Cholecystokinin; Sincalide; Stomach Neoplasms

We report here the cDNA cloning of a putative gastrin receptor from enterochromaffin-like (ECL) carcinoid tumor of Mastomys natalensis. For this study, we used the polymerase chain reaction technique to amplify transmembrane domain sequences related to rat pancreatic cholecystokinin (CCK)-A receptor from the ECL tumor cDNA library. The amino acid sequence deduced from the cloned cDNA showed 85.7% and 49.0% identity to canine parietal cell gastrin receptor and rat pancreatic CCK-A receptor, respectively. Ligand binding studies using COS-7 cells transfected with the cDNA showed the same binding specificity for gastrin and CCK-8 as the gastrin receptor on the Mastomys carcinoid tumor membrane. Both gastrin and CCK-8 elevated free cytosolic calcium concentration in COS-7 cells expressing the cloned receptor. RNA blot analysis revealed the expression of the gastrin receptor in both Mastomys stomach and brain.

Topics: Amino Acid Sequence; Animals; Base Sequence; Calcium; Carcinoid Tumor; Cloning, Molecular; DNA; Dogs; Female; Gastrins; Molecular Sequence Data; Muridae; Pancreas; Parietal Cells, Gastric; Rats; Receptors, Cholecystokinin; Sequence Homology, Nucleic Acid; Sincalide; Stomach Neoplasms; Transfection

Gastrin is known as a trophic factor for some stomach and colorectal cancer cells; however, the roles of gastrin receptors and the intracellular signal transduction pathways by which gastrin regulates cell growth are still unknown. The authors examined the effect of synthetic human gastrin-17 on growth of human stomach cancer cells (the parent line, AGS-P, and two different clones, AGS-10 and AGS-12), which were established (and have been maintained) in our laboratory. Gastrin stimulated growth of AGS-P and AGS-10 cells, which have gastrin receptors, in a dose-dependent fashion. A highly selective gastrin receptor antagonist, JMV 320, inhibited the growth-stimulatory effect of gastrin on AGS-P cells in a dose-dependent fashion. Concentrations of gastrin (10(-8) to 10(-6) M), which stimulated growth of AGS-P cells, did not affect either cyclic adenosine monophosphate production or phosphatidylinositol hydrolysis. Gastrin (10(-11) to 10(-5) M) mobilized calcium from the intracellular organelles to increases intracellular calcium level in AGS-P cells. The AGS-12 clone has no gastrin receptors, and gastrin did not affect growth or mobilization of intracellular calcium in these cells. Our findings indicate that gastrin stimulates growth of AGS cells through a mechanism that involves binding to specific gastrin receptors that are linked to the system for mobilization of intracellular calcium.

Topics: Amino Acid Sequence; Calcium; Gastrins; Hormones; Humans; In Vitro Techniques; Molecular Sequence Data; Receptors, Cholecystokinin; Sincalide; Stomach Neoplasms; Tumor Cells, Cultured

This study deals with the effect of four types of COOH-terminal cholecystokinin (CCK) fragments on the growth of xenotransplantable human gastric cancer (SC-6-JCK, a poorly differentiated adenocarcinoma) whose growth has been promoted by pentagastrin. The growth of the tumor was inhibited using daily s.c. injections of CCK-octapeptide (CCK-8) and glutaryl-CCK-8 at a dose of 500 micrograms/kg body weight. After 30 days of treatment with CCK-8 or glutaryl-CCK-8, a significant decrease was observed in the tumor weight (P less than 0.05) and the tumor size P less than 0.01) in comparison with those of the control. But treatment with CCK-12 and pyroglutamyl-CCK-8 did not produce inhibition of tumor growth. Furthermore the correlation between the effect of CCK-8 on the normal rise in tumor cyclic adenosine 3':5'-monophosphate (cAMP) levels caused by pentagastrin injection and tumor growth was studied. The increase of cAMP by a single i.p. injection of pentagastrin at a dose of 20 micrograms/mouse was significantly inhibited by pretreatment with CCK-8 at concentrations equimolar to pentagastrin (P less than 0.05), while cAMP in the tumor was slightly elevated by a single i.p. injection of CCK-8 alone. Also in the in vitro study, CCK-8 inhibited the increase of cAMP and the activation of cAMP-dependent protein kinase which was stimulated by pentagastrin. These results suggest that proliferation of gastrin-dependent human gastric cancers may be suppressed by CCK in competition with gastrin.

Topics: Adenocarcinoma; Animals; Cholecystokinin; Cyclic AMP; Humans; Mice; Neoplasm Transplantation; Pentagastrin; Protein Kinases; Sincalide; Stomach Neoplasms