leukotriene-b4 and Colonic-Neoplasms

Topics: Arachidonic Acid; Caco-2 Cells; Colonic Neoplasms; Dinoprostone; Gene Expression Regulation, Neoplastic; Humans; Leukotriene B4; Lipoxygenases; Phytic Acid; Prostaglandin-Endoperoxide Synthases; RNA, Messenger; Signal Transduction

Epigallocatechin-3-gallate (EGCG), atorvastatin (ATST), and their combination have been previously shown to inhibit colon carcinogenesis in animal models. We further investigated their inhibitory activities in azoxymethane (AOM) and dextran sulfate sodium (DSS)-treated Balb/cJ mice and CD-1 mice in 2 slightly different models. The mice were maintained on the AIN93M diet, or a similar diet containing 0.03%, 0.1%, or 0.3% EGCG; 60-ppm ATST; or a combination of 0.1% EGCG and 60-ppm ATST. Unexpectedly, no significant inhibitory activity was observed, and some of the treatment groups resulted in higher tumor multiplicity. To study the effects of EGCG on colon inflammation, CD-1 or C57BL/6 mice were treated with 1.5% DSS for 7 days and sacrificed 3 days later. DSS induced rectal bleeding and colon shortening; treatment with 0.5% EGCG exacerbated the bleeding and decreased mouse body weight. Dietary 0.5% EGCG also increased serum levels of leukotriene B4 and prostaglandin E2. These results suggest that, in mice bearing colon inflammation, high concentrations of EGCG and ATST enhance colon bleeding and may promote colon carcinogenesis.

Topics: Animals; Atorvastatin; Azoxymethane; Catechin; Colitis; Colon; Colonic Neoplasms; Dextran Sulfate; Dinoprostone; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Gastrointestinal Hemorrhage; Heptanoic Acids; Leukotriene B4; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Pyrroles; Rectum; Weight Loss

Matrix metalloproteinase 7 (MMP7), a metallohydrolase involved in the development of several cancers, is downregulated in the Apc(Min/+) colon cancer mouse model following sulindac treatment. To determine whether this effect is relevant to the human condition, HT-29 human colon cancer cells were treated with sulindac and its metabolites, and compared to results obtained from in vivo mouse studies. The expression of MMP7 was monitored. The results demonstrated that sulindac sulfide effectively downregulated both MMP7 expression and activity. Furthermore, activity-based proteomics demonstrated that sulindac sulfide dramatically decreased the activity of leukotriene A4 hydrolase in HT-29 cells as reflected by a decrease in the level of its product, leukotriene B4. This study demonstrates that the effect of sulindac treatment in a mouse model of colon cancer may be relevant to the human counterpart and highlights the effect of sulindac treatment on metallohydrolases.

Topics: Animals; Antineoplastic Agents; Cell Death; Colonic Neoplasms; Drug Screening Assays, Antitumor; Epoxide Hydrolases; Gene Expression Regulation, Neoplastic; GPI-Linked Proteins; HT29 Cells; Humans; Immunoassay; Leukotriene B4; Matrix Metalloproteinase 7; Matrix Metalloproteinases, Membrane-Associated; Mice; Proteomics; Ribosomal Proteins; RNA, Messenger; Sulindac; Trypsin

Celecoxib, a selective cyclooxygenase-2 (COX-2) inhibitor, has anticancer effect on many cancers associated with chronic inflammation by both COX-2-dependent and COX-2-independent mechanisms. The non-COX-2 targets of celecoxib, however, are still a matter of research. Leukotriene B4 (LTB4) has been implicated in prostate and colon carcinogenesis, but little is known about the potential role of LTB4 in celecoxib-mediated anticancer effect. In this study, we evaluated whether LTB4 was involved in celecoxib-mediated inhibitory effect on human colon cancer HT-29 cells and human prostate cancer PC-3 cells. Our data showed that survival of both cell lines was obviously suppressed after celecoxib treatment for 72 h in a concentration-dependent manner. However, only in HT-29 cells, this inhibitory effect could be reversed by LTB4, which promoted survival of HT-29 cells rather than PC-3 cells. Consistent with these results, lioxygenase (LOX) potent inhibitor nordihydroguaiaretic acid (NDGA) had a higher inhibitory effect on HT-29 cells than PC-3 cells. Additionally, ELISA results showed that celecoxib could suppress expression of LTB4 in both cell lines, whereas, inhibition of PGE2 was only detected in HT-29 cells. These results indicate that the anticancer effect of celecoxib is COX-2-independent in HT-29 and PC-3 cells and in HT-29 cells primarily via down-regulating LTB4 production.

Topics: Antineoplastic Agents; Celecoxib; Cell Line, Tumor; Cell Proliferation; Cell Survival; Colonic Neoplasms; Cyclooxygenase 2 Inhibitors; Humans; Leukotriene B4; Lipoxygenase Inhibitors; Male; Masoprocol; Prostatic Neoplasms; Pyrazoles; Sulfonamides

We investigated the effects of a gamma-tocopherol-rich mixture of tocopherols (gamma-TmT, containing 57% gamma-T, 24% delta-T, and 13% alpha-T) on colon carcinogenesis in azoxymethane (AOM)/dextran sulfate sodium (DSS)-treated mice. In experiment 1, 6-week-old male CF-1 mice were given a dose of AOM (10 mg/kg body weight, i.p.), and 1 week later, 1.5% DSS in drinking water for 1 week. The mice were maintained on either a gamma-TmT (0.3%)-enriched or a standard AIN93M diet, starting 1 week before the AOM injection, until the termination of experiment. In the AOM/DSS-treated mice, dietary gamma-TmT treatment resulted in a significantly lower colon inflammation index (52% of the control) on day 7 and number of colon adenomas (9% of the control) on week 7. gamma-TmT treatment also resulted in higher apoptotic index in adenomas, lower prostaglandin E2, leukotriene B4, and nitrotyrosine levels in the colon, and lower prostaglandin E2, leukotriene B4, and 8-isoprostane levels in the plasma on week 7. Some of the decreases were observed even on day 7. In experiment 2 with AOM/DSS- treated mice sacrificed on week 21, dietary 0.17% or 0.3% gamma-TmT treatment, starting 1 week before the AOM injection, significantly inhibited adenocarcinoma and adenoma formation in the colon (to 17-33% of the control). Dietary 0.3% gamma-TmT that was initiated after DSS treatment also exhibited a similar inhibitory activity. The present study showed that gamma-TmT effectively inhibited colon carcinogenesis in AOM/DSS-treated mice, and the inhibition may be due to the apoptosis-inducing, anti-inflammatory, antioxidative, and reactive nitrogen species-trapping activities of tocopherols.

Topics: Adenocarcinoma; Adenoma; Animals; Antioxidants; Apoptosis; Azoxymethane; Carcinogens; Cell Transformation, Neoplastic; Cocarcinogenesis; Colon; Colonic Neoplasms; Dextran Sulfate; Dinoprost; Dinoprostone; Dose-Response Relationship, Drug; gamma-Tocopherol; Inflammation; Leukotriene B4; Male; Mice; Tyrosine

We investigated whether leukotriene B(4) (LTB(4)) and its signaling pathway play an important role in the progression of human colon cancer via a direct stimulation of cancer cell proliferation. Remarkable expression of LTB(4) receptor 1 (BLT1) in human colon cancer tissues was detected by immunohistochemistry, and Western blot analysis revealed the BLT1 expression in cultured human colon cancer cell lines, Caco2 and HT29. The 5-lipoxygenase inhibitor AA-861 and LTB(4)-receptor antagonist U75302 showed negative effects on survival and proliferation of both Caco2 and HT-29 cells. The inhibition of cell proliferation is due to the apoptosis because nuclear condensation and increased annexin V expression were observed in the cells treated with AA-861 and U75302. Knockdown of BLT1 by small interfering RNA caused the suppression of BLT1 protein, resulting in the inhibition of cancer cell proliferation. Blockade of BLT1 by the receptor antagonist significantly suppresses the LTB(4)-stimulated extracellular signal-regulated kinase (ERK) activation in colon cancer cells. These results indicate that the blockade of the LTB(4)-signaling pathway induces apoptosis via the inhibition of ERK activation in colon cancer cells. The LTB(4)-signaling pathway might be a new therapeutic target for colon cancer.

Topics: Apoptosis; Benzoquinones; Caco-2 Cells; Cell Proliferation; Cell Survival; Colonic Neoplasms; Extracellular Signal-Regulated MAP Kinases; Fatty Alcohols; Glycols; Humans; Leukotriene B4; Lipoxygenase Inhibitors; Receptors, Leukotriene B4; RNA Interference; Signal Transduction

Previous studies indicate that the arachidonic acid-metabolizing enzymes COX-2 and 5-LOX are overexpressed during the process of colonic adenoma formation promoted by cigarette smoke. The aims of the present study were to investigate whether there exists a relationship between COX-2 and 5-LOX, and whether dual inhibition of COX-2 and 5-LOX has an anticarcinogenic effect in the colonic tumorigenesis promoted by cigarette smoke. Results showed that pretreating colon cancer cells with cigarette smoke extract (CSE) promoted colon cancer growth in the nude mouse xenograft model. Inhibition of COX-2 or 5-LOX reduced the tumor size. In the group treated with COX-2-inhibitor, the PGE2 level decreased while the LTB4 level increased. In contrast, in the 5-LOX-inhibitor treated group, the LTB4 level was reduced and the PGE2 level was unchanged. However, combined treatment with both COX-2 and 5-LOX inhibitors further inhibited the tumor growth promoted by CSE over treatment with either COX-2-inhibitor or 5-LOX-inhibitor alone. This was accompanied by the downregulation of PGE2 and LTB4. In an in vitro study, we found that the action of CSE on colon cancer cells was mediated by 5-LOX DNA demethylation. In summary, these results indicate that inhibition of COX-2 may lead to a shunt of arachidonic acid metabolism towards the leukotriene pathway during colonic tumorigenesis promoted by CSE. Suppression of 5-LOX did not induce such a shunt and produced a better response. Therefore, 5-LOX inhibitor is more effective than COX-2 inhibitor, and blocker of both COX-2 and 5-LOX may present a superior anticancer profile in cigarette smokers.

Topics: Adenocarcinoma; Animals; Apoptosis; Arachidonate 5-Lipoxygenase; Cell Proliferation; Colonic Neoplasms; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Cyclooxygenase Inhibitors; Dinoprostone; DNA Methylation; Female; Leukotriene B4; Lipoxygenase Inhibitors; Mice; Mice, Inbred BALB C; Mice, Nude; Neovascularization, Pathologic; Nicotiana; Prostaglandin-Endoperoxide Synthases; RNA, Messenger; Smoke; Tumor Cells, Cultured

Excessive fat consumption is a risk factor for colon carcinogenesis, and green tea consumption may reduce the risk of colon and other cancers. The current study was designed to investigate the effects of green tea and a high-fat diet on arachidonic acid metabolism and aberrant crypt foci formation in an azoxymethane (AOM)-induced colon carcinogenesis mouse model. We also determined whether green tea consumption altered the size of regional fat pads. CF-1 female mice were maintained on either a high-fat (20% corn oil) or a low-fat (5% corn oil) diet. AOM was given subcutaneous at a dose of 7.5 mg/kg body weight at 6 wk and then a dose of 10 mg/kg at 7 wk of age. Two weeks after the second AOM injection, 0.6% green tea (6 mg tea solids/ml) was given as the drinking fluid and continued for 10 wk until the experiment was terminated. In the AOM-treated mice not receiving green tea, the high-fat diet significantly enhanced colonic levels of 5-lipoxygenase, leukotriene A4 hydrolase, and leukotriene B4, but it did not significantly alter prostaglandin E2 levels and aberrant crypt foci formation. In AOM-treated mice on the high-fat diet, green tea significantly decreased colonic levels of cytosolic phospholipase A2, 5-lipoxygenase, and leukotriene B4; green tea treatment also decreased the number of aberrant crypt foci (P < 0.05). The weights of parametrial and retroperitoneal fat pads were increased by the high-fat diet and decreased by green tea treatment. The current results indicate that green tea consumption and dietary fat modulate 5-lipoxygenase-dependent pathway of arachidonic acid metabolism during AOM-induced colon carcinogenesis. Green tea inhibits ACF formation in mice on a high corn oil diet, suggesting its possible inhibitory effect on colon carcinogenesis in populations such as those in Western countries that consume high amounts of fat.

Topics: Adipose Tissue; Animals; Arachidonate 5-Lipoxygenase; Arachidonic Acid; Azoxymethane; Body Weight; Colon; Colonic Neoplasms; Cytosol; Dietary Fats; Dinoprostone; Drinking; Eating; Epoxide Hydrolases; Female; Leukotriene B4; Mice; Organ Size; Phospholipases A; Phospholipases A2; Tea

The relationship between growth and alterations in arachidonic acid (AA) metabolism in human breast (MCF-7) and colon (SW480) cancer cells was studied. Four different fatty acid preparations were evaluated: a mixture of conjugated linoleic acid (CLA) isomers (c9,t11, t10,c12, c11,t13, and minor amounts of other isomers), the pure c9,t11-CLA isomer, the pure t10,c12-CLA isomer, and linoleic acid (LA) (all at a lipid concentration of 16 microg/mL). 14C-AA uptake into the monoglyceride fraction of MCF-7 cells was significantly increased following 24 h incubation with the CLA mixture (P < 0.05) and c9,t11-CLA (P < 0.02). In contrast to the MCF-7 cells, 14C-AA uptake into the triglyceride fraction of the SW480 cells was increased while uptake into the phospholipids was reduced following treatment with the CLA mixture (P < 0.02) and c9,t11-CLA (P < 0.05). Distribution of 14C-AA among phospholipid classes was altered by CLA treatments in both cell lines. The c9,t11-CLA isomer decreased (P < 0.05) uptake of 14C-AA into phosphatidylcholine while increasing (P < 0.05) uptake into phosphatidylethanolamine in both cell lines. Both the CLA mixture and the t10,c12-CLA isomer increased (P < 0.01) uptake of 14C-AA into phosphatidylserine in the SW480 cells but had no effect on this phospholipid in the MCF-7 cells. Release of 14C-AA derivatives was not altered by CLA treatments but was increased (P < 0.05) by LA in the SW480 cell line. The CLA mixture of isomers and c9,t11-CLA isomer inhibited 14C-AA conversion to 14C-prostaglandin E2 (PGE2) by 20-30% (P < 0.05) while increasing 14C-PGF2alpha by 17-44% relative to controls in both cell lines. LA significantly (P < 0.05) increased 14C-PGD2 by 13-19% in both cell lines and increased 14C-PGE2 by 20% in the SW480 cell line only. LA significantly (P < 0.05) increased 5-hydroperoxyeicosatetraenoate by 27% in the MCF-7 cell line. Lipid peroxidation, as determined by increased levels of 8-epi-prostaglandin F2alpha (8-epi-PGF2alpha), was observed following treatment with c9,t11-CLA isomer in both cell lines (P < 0.02) and with t10,c12-CLA isomer in the MCF-7 cell line only (P < 0.05). These data indicate that the growth-promoting effects of LA in the SW480 cell line may be associated with enhanced conversion of AA to PGE2 but that the growth-suppressing effects of CLA isomers in both cell lines may be due to changes in AA distribution among cellular lipids and an altered prostaglandin profile.

Topics: Arachidonic Acid; Breast Neoplasms; Carbon Radioisotopes; Cell Survival; Colonic Neoplasms; Dinoprost; Dinoprostone; Humans; Leukotriene B4; Leukotrienes; Linoleic Acid; Prostaglandin D2; Tumor Cells, Cultured

Several lines of evidence strongly link prostaglandins (PGs) and leukotrienes (LTs) to cancer of the intestine. Several studies have reported a 40-50% reduction in mortality from colorectal cancer in individuals who routinely consume nonsteroidal anti-inflammatory drugs, possibly by inhibiting cyclooxygenase activity. However, the role of eicosanoids in this process is still unclear. The heterozygote Min/+ mouse model, like patients with familial adenomatous polyposis, carries a nonsense mutation in the adenomatous polyposis coli (APC) gene that results in the spontaneous development of intestinal adenomas (100% incidence). This study investigated the association between eicosanoid biosynthesis, intestinal tumor load, and the chemotherapeutic effect of the nonsteroidal anti-inflammatory drug sulindac during early and preexisting phases of tumor growth and development as well as residual effects after drug withdrawal. Administration of sulindac (320 ppm) to Min/+ mice reduced the tumor number by 95% but did not alter the levels of PGE2 and LTB4 in intestinal tissues. Increasing PGE2 and LTB4 levels by 44% with dietary arachidonic acid supplementation had no effect on tumor number or size. When sulindac was added to the arachidonic acid-supplemented diet, tumor number was reduced by 82%, whereas eicosanoid levels remained elevated. In Min/+ mice with established tumors, treatment with sulindac for 4 days reduced tumor number by 75%, and continual administration of sulindac was necessary to maintain a reduced tumor load. In summary, alterations in eicosanoid formation were not correlated with tumor number or size in the Min/+ mouse model; thus, the antitumor effect of sulindac seems to be PG independent.

Topics: Adenomatous Polyposis Coli; Administration, Oral; Animals; Anti-Inflammatory Agents, Non-Steroidal; Arachidonic Acid; Colonic Neoplasms; Dinoprostone; Disease Susceptibility; Genes, APC; Heterozygote; Jejunal Neoplasms; Leukotriene B4; Male; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Mutant Strains; Prodrugs; Prostaglandins; Remission Induction; Sulindac

We studied the influence of cancer cells on the LTB4 production by human polymorphonuclear leukocytes (PMN). The cancer cells were isolated from malignant pleural effusion specimens taken from two patients or from a peritoneal effusion specimen of one patient. While human PMN produced LTB4 following stimulation with A23187, the addition of cancer cells inhibited LTB4, 5-HETE and 12-HETE production by PMN in a cell number-dependent manner, while the cancer cell lines also showed a similar inhibition. The addition of lysate of the breast cancer cells also inhibited in a dose-dependent manner the production of LTB4 by PMN following stimulation with A23187. The addition of arachidonic acid completely reversed the inhibition of PMN-LTB4 production by the addition of the breast cancer cell lysates, thus suggesting inhibition at the phospholipase A2 level. The addition of this lysate to the partially purified human cytosolic PLA2 also inhibited the PLA2 activity. In contrast, the addition of lymphoma cells isolated from metastatic lymphnodes did not inhibit the LTB4 production from PMN. Since LTB4 is one of the important chemotactic factors for PMN and monocytes, these findings suggest that the inhibition of the PLA2 activity by the cancer cells thus results in a reduced production of LTB4 from PMN and contributes to a predisposition to develop severe infection in patients with advanced cancer.

Topics: Ascitic Fluid; Cell Separation; Colonic Neoplasms; Humans; Leukotriene B4; Male; Middle Aged; Neutrophils; Phospholipases A; Phospholipases A2; Pleural Effusion, Malignant

Results from epidemiological studies indicate that chronic administration of aspirin reduces the incidence of colon cancer. The mechanism that accounts for this reduction is not known, but it may be related to the decreased production of prostanoids that results from aspirin inhibition of cyclooxygenase. However, it is not known whether aspirin has a local effect on prostanoid production in the colonic mucosa and whether this effect is dose dependent. In this study, we determined the effect of oral administration of aspirin on the production of the prostanoid prostaglandin E2 (PGE2) in the intact human colonic mucosa. Inhibition of cyclooxygenase could result in an increased availability of arachidonic acid and a corresponding increase in production of other eicosanoids. To determine whether such an effect occurs, we also quantitated the concentration of leukotriene B4 (LTB4) in colonic mucosal samples. Mucosal samples were obtained during sigmoidoscopy from the colons of 17 subjects with a history of colonic cancer prior to and following 60 days of self-administration of 325 mg aspirin/day and again 60 days after administration of 650 mg aspirin/day. PGE2 and LTB4 concentrations were determined by enzyme immunoassay for tissue samples that were flash frozen after removal from the biopsy forceps and also in medium that was collected from tissue samples that were incubated for 4 h following removal from the subject. PGE2 concentrations were decreased significantly in samples collected after 60 days of consumption of 325 mg aspirin. An additional 60 days of consuming 650 mg aspirin/day did not result in a further significant decrease relative to that attained after consumption of 325 mg/day. Similar results were obtained using colonic explants, and the addition of aspirin to medium further reduced PGE2 production. LTB4 in tissue and medium was not significantly different in pre-versus post-aspirin samples, with the exception of an increased concentration in medium samples collected after consumption of 650 mg/day relative to pre-aspirin samples. The results indicate that aspirin affects eicosanoid production in the colonic mucosa of humans, but the effect is most likely restricted to products of the cyclooxygenase-dependent pathway. It appears that 325 mg of aspirin is sufficient to affect PGE2 production and that increasing the dosage to 650 mg daily provides an additional decrease in PGE2 synthesis. However, the higher dosage was associated with a conside

Topics: Adult; Aged; Aged, 80 and over; Antineoplastic Agents; Arachidonic Acid; Aspirin; Colon; Colonic Neoplasms; Cyclooxygenase Inhibitors; Dinoprostone; Female; Humans; Intestinal Mucosa; Leukotriene B4; Lipoxygenase; Male; Middle Aged

Eicosanoids have been implicated in colon carcinogenesis, but very little is known on the potential role of leukotrienes (LTs) and hydroxyeicosatetraenoic acids (HETEs) in this process; such compounds are produced by colonocytes and tumor infiltrating leukocytes. We studied the effect of LTB4, LTB4 methyl ester, LTB5, 12(R)-HETE, 12(S)-HETE and 15(S)-HETE (10(-10), 10(-8), 10(-6) M) on the proliferation rate, the cell cycle distribution, and the rate of apoptosis in HT-29 and HCT-15 human colon carcinoma cells. Our data show that LTB4, a lipoxygenase product, increased the proliferation rate of both cell lines in a time- and concentration-dependent manner. In HT-29 cells the concentration-response curve was bell-shaped (maximal effect at 10(-8) M). The proliferative effects of LTB4 in HT-29 cells were inhibited by SC-41930, a competitive antagonist of LTB4, suggesting the existence of an LTB4 receptor in epithelial cells. The methyl ester of LTB4 stimulated the proliferation of these cells, but LTB5, an isomer of LTB4 derived from eicosapentaenoic acid, did not. Of the HETEs, only 12(R)-HETE, a P-450 product, stimulated the proliferation of both cell lines; the other HETEs, all lipoxygenase products, failed to affect the proliferation of these cells. None of these eicosanoids had any effect on cell cycle distribution or apoptosis in either cell line. Taken together with our previous data showing that PGs stimulate colon cancer cell proliferation (Qiao et al. (1995) Biochim. Biophys. Acta 1258, 215-223), these findings indicate that arachidonic acid products synthesized via at least three different pathways (cyclooxygenase, lipoxygenase, P-450) may not be able to modulate the growth of colon cancer, and suggest a potential role in human colon carcinogenesis for LTB4 and 12(R)-HETE.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Benzopyrans; Cell Division; Colonic Neoplasms; Eicosanoids; Flow Cytometry; Humans; Hydroxyeicosatetraenoic Acids; Interphase; Leukotriene B4; Mitosis; Receptors, Leukotriene; S Phase; Tumor Cells, Cultured

Eicosanoids have been implicated in the pathogenesis of cancer and are known to regulate the expression of antigens of the major histocompatibility complex (MHC). In human colon cancer, we have recently observed that: (a) the expression of MHC class I and II antigens are markedly reduced; and (b) the levels of PGE2, but not of PGF2 alpha and LTB4, are elevated compared to histologically normal mucosa. Therefore, we investigated the effect of PGE2, PGF2 alpha and LTB4 on the regulation of MHC class I antigens in two human colon adenocarcinoma cell lines and in a murine model of colon cancer. None of these eicosanoids had any significant effect on the expression of MHC class I antigens in the human colonocytes or the transcription rate of class I genes, with the exception of LTB4 which only modestly suppressed the transcription rate. Similarly, 16, 16-dimethyl-PGE2 had no effect on the expression of MHC class I genes in the colonocytes of BALB/c mice treated with the carcinogen dimethylhydrazine. We conclude that PGE2, PGF2 alpha and LTB4 did not affect the expression of MHC class I antigens in cultured human colon adenocarcinoma cells, and 16, 16-dimethyl PGE2 did not affect their expression in mice, even when mice were treated with a colon carcinogen. Thus, these eicosanoids are an unlikely regulator of the observed underexpression of MHC class I antigens in human colon cancer.

Topics: 16,16-Dimethylprostaglandin E2; Adenocarcinoma; Animals; Colon; Colonic Neoplasms; Dinoprost; Dinoprostone; Eicosanoids; Gene Expression Regulation, Neoplastic; Genes, MHC Class I; Histocompatibility Antigens Class I; Humans; Leukotriene B4; Male; Mice; Mice, Inbred BALB C; Tumor Cells, Cultured

A method for the analysis of omega-carboxyleukotriene B4 and omega-hydroxyleukotriene B4 in human colonic carcinoma homogenate is described. The hydroxy groups of the leukotriene metabolite were acetylated by acetic anhydride, and the mixture was partially purified on a Sep-Pak C18 cartridge and analysed by reversed-phase HPLC-thermospray MS. Generally, the base ion, [MH-2(60)]+, is produced through elimination of two acetic acid (60 mass units) molecules from the protonated molecular ion. On selected-ion monitoring, standard curves for omega-carboxy- or omega-hydroxyleukotriene B4 showed a linear relationship over the range 72-1500 pmol. The assay based on selected-ion monitoring was applied to an extract from human colonic carcinoma homogenate. When a homogenate of human colonic well-differentiated adenocarcinoma was incubated with NADPH and leukotriene B4 (60.6 nmol) as a substrate, the conversion of precursor leukotriene B4 to omega-carboxyleukotriene B4 or omega-hydroxyleukotriene B4 was 0.33 or 3.17%, respectively. Based on these results, it is suggested that carcinoma cells themselves or leukocytes at the hostsite in a region of human colonic well-differentiated adenocarcinoma are performing omega-oxidation through NADPH-dependent omega-hydroxylation of leukotriene B4.

Topics: Adenocarcinoma; Cell Differentiation; Chromatography, High Pressure Liquid; Colonic Neoplasms; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Male; Mass Spectrometry

Prostaglandins (PG) have been implicated in the pathogenesis of cancer and play an important role in immune regulation. Colon cancer is associated with elevated levels of PGE2, while aspirin, the prototypical inhibitor of PG synthesis, appears to reduce the incidence of colon cancer by 50%. We have observed that in human colon cancer the expression of HLA class I and II antigens is reduced or lost; loss of HLA antigens is suspected to be a mechanism by which the malignant cell escapes the immune surveillance. We investigated the effect of these eicosanoids on the expression of HLA antigens in human colon adenocarcinoma cell lines. PGE2 down-regulated the expression of the class II antigen HLA-DR in SW1116 cells (65% reduction at 2.8 x 10(-8) M). This effect was dose- and time-dependent, reversible, and specific (PGF2 alpha and LTB4 had no effect; the expression of carcinoembryonic antigen and class I genes were not affected). Aspirin induced the expression of HLA-DR in HT29 cells, a cell line not expressing constitutively HLA-DR. The reduction of HLA-DR by PGE2 was accompanied by reduced messenger RNA (mRNA) levels of HLA-DR alpha and reduced transcription of the corresponding gene. In contrast to HLA-DR, none of these three eicosanoids affected the expression of HLA class I genes, as assessed via determination of protein expression by fluorescence flow cytometric analysis and evaluation of the corresponding class I mRNA levels. We conclude that PGE2 specifically down-regulates the expression of HLA-DR, while it does not affect the expression of class I antigens.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenocarcinoma; Aspirin; Base Sequence; Cell Line; Colonic Neoplasms; Dinoprost; Dinoprostone; DNA Primers; Dose-Response Relationship, Drug; Gene Expression Regulation, Neoplastic; HLA-DR Antigens; Humans; Leukotriene B4; Molecular Sequence Data; Polymerase Chain Reaction; RNA, Messenger; Tumor Cells, Cultured

The ability of a human colonic epithelial cell line (CaCo-2) to synthesize leukotriene B4 (LTB4) in response to bile salt stimulation was examined, as was the dependency of such stimulation on the hydrophobic-hydrophilic balance of the bile salts. We demonstrate for the first time in this human intestinal epithelial cell line the ability of bile salts to stimulate synthesis of LTB4. CaCo-2 cell monolayers were incubated with a series of bile salts ranging in concentration from 0.5 microM to 1 mM. This resulted in a dose- and hydrophobicity-dependent increase in LTB4 synthesis. Hydrophobic bile salts (glycine and taurine conjugates of lithocholate and deoxycholate) caused LTB4 synthesis to be stimulated 27% and 35%, respectively, above control levels. In contrast, hydrophilic bile salts (glycine and taurine conjugates of ursodeoxycholate) increased LTB4 synthesis only 11.2% and 16.1%. Under basal conditions pretreatment with dexamethasone significantly inhibited bile salt-induced LTB4 synthesis by 38% compared to control. With more hydrophobic bile salts, chenodeoxycholate and deoxycholate, dexamethasone inhibited LTB4 synthesis to levels significantly below those observed with dexamethasone under basal conditions. Unlike A23187 calcium ionophore-induced LTB4 synthesis, bile salt-induced stimulation of LTB4 synthesis was not found to be dependent on the presence of extracellular calcium. Variations in bile salt stimulation of LTB4 by intestinal epithelial cells could be important in modulating cellular responses. The synthesis of chemotactic factors, such as LTB4, by the human colonic adenocarcinoma epithelial cell line now needs to be extended to normal human intestinal epithelium, as it may play a role in many of the functional disturbances which characterize intestinal inflammatory conditions.

Topics: Adenocarcinoma; Arachidonic Acid; Bile Acids and Salts; Calcium; Colon; Colonic Neoplasms; Dexamethasone; Humans; L-Lactate Dehydrogenase; Leukotriene B4; Tumor Cells, Cultured

Eicosanoids may participate in colon carcinogenesis, as evidenced from work in animal tumor models showing prevention of colon cancer by inhibitors of their synthesis and epidemiologic studies demonstrating reduced risk of colon cancer in long-term users of aspirin and other nonsteroidal antiinflammatory drugs (NSAIDs). The levels of prostaglandin E2 (PGE2), PGF2 alpha, PGI2, thromboxane A2 (TXA2), and leukotriene B4 (LTB4), which represent the cyclooxygenase and 5-lipoxygenase pathways, were determined in 21 pairs of surgically excised human colon cancer and histologically normal mucosa samples 5 to 10 cm away from the tumor. The levels of PGE2 were elevated in colon cancer samples as compared with histologically normal mucosa samples distant from the cancer (p < 0.01), whereas levels of prostacyclin (PGI2) were decreased (p < 0.05). The differences in the levels of PGF2 alpha, TXA2, and LTB4 between normal and malignant tissue were not statistically significant. No statistically significant association was found between the level of each of the eicosanoids assayed and Dukes' stage of colon cancer. These findings, confirming and extending earlier work from tumors and cell culture, suggest that the protective effect of aspirin and other NSAIDs in the development of human colon cancer may be mediated, at least in part, through their inhibition of arachidonic acid metabolism by cyclooxygenase.

Topics: Adult; Aged; Aged, 80 and over; Anti-Inflammatory Agents, Non-Steroidal; Arachidonic Acids; Aspirin; Colonic Neoplasms; Dinoprostone; Eicosanoids; Epoprostenol; Female; Humans; Intestinal Mucosa; Leukotriene B4; Male; Middle Aged; Thromboxane A2; Time Factors

Monolayers of Caco-2 cells, a human enterocyte cell line, were incubated separately with 3H8-labeled preparations of three different lipid mediators of inflammation: 5-hydroxyeicosatetraenoic acid, 12-hydroxyeicosatetraenoic acid, and leukotriene B4. Both [3H8]5-hydroxyeicosatetraenoic and [3H8]12-hydroxyeicosatetraenoic acids were taken up and metabolized by Caco-2 cells, but [3H]leukotriene B4 remained unmetabolized in the incubation medium. [3H]5-hydroxyeicosatetraenoic acid was esterified into cellular phospholipids (15%) and triglycerides (4%) but did not undergo beta-oxidation. When [3H]12-hydroxyeicosatetraenoic acid was incubated with Caco-2 cells, 14% underwent two cycles of beta-oxidation to form [3H]8-hydroxyhexadecatrienoic acid, and 3% underwent three cycles of beta-oxidation to form [3H]6-hydroxytetradecadienoic acid, both of which were released into the media. [3H]12-Hydroxyeicosatetraenoic acid was also esterified into cellular phospholipids (13%), but none was esterified into cellular triglycerides.

Topics: Adenocarcinoma; Arachidonic Acid; Cell Line; Colonic Neoplasms; Culture Media; Esterification; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Tumor Cells, Cultured

Prostaglandin E2 (PGE2) secretion by peripheral blood and tissue-fixed macrophages from patients with colorectal carcinoma was assessed. There was no significant difference between PGE2 production by peripheral blood mononuclear cells between patients with colorectal carcinoma and normal controls. However, secretion of PGE2 by tissue-fixed macrophages from within the colorectal carcinomata in response to opsonised zymosan was significantly higher than in the uninvolved colonic tissue. PGE2 production by tissue-fixed macrophages from within colonic polyps was found to be normal. These results could not be explained on the basis of increased availability of substrate arachidonic acid since addition of excess arachidonic acid resulted in similar findings. The enhanced production of PGE2 correlated with Dukes staging but not the level of differentiation. The production of PGE2 from epithelial cells in response to ionophore A23187 was not significantly enhanced. Leukotriene B4 secretion by intestinal macrophages in response to opsonised zymosan was not significantly elevated in the colonic tumour tissue. Modulation of levels of prostaglandin production within colonic tumours may play a role in the rate of growth and vascularity of these tumours and in the regulation of the local immune response to malignancy.

Topics: Calcimycin; Colon; Colonic Neoplasms; Dinoprostone; Humans; In Vitro Techniques; Leukocytes, Mononuclear; Leukotriene B4; Macrophages; Zymosan

The prostaglandin and leukotriene synthesizing capacity of human gastrointestinal tissues obtained at surgery was investigated using radioimmunoassay for prostaglandin E2, leukotriene B4 and sulfidopeptide leukotrienes. The leukotriene immunoassay data were validated by high-pressure liquid chromatography (HPLC). During incubation at 37 degrees C, fragments of human gastric, jejuno-ileal and colonic mucosa released considerably larger amounts of prostaglandin E2 than of leukotriene B4 and sulfidopeptide leukotrienes. Gastrointestinal smooth muscle tissues released even larger amounts of prostaglandin E2, but smaller amounts of leukotrienes than the corresponding mucosal tissues. Adenocarcinoma tissue released larger amounts of leukotriene B4, sulfidopeptide leukotrienes and prostaglandin E2 than normal colonic mucosa. Ionophore A23187 (5 micrograms/ml) did not stimulate release of prostaglandin E2 from any of the tissues investigated, but enhanced release of leukotriene B4 and sulfidopeptide leukotrienes. HPLC analysis demonstrated that immunoreactive leukotriene B4 co-chromatographed almost exclusively with standard leukotriene B4, while immunoreactive sulfidopeptide leukotrienes consisted of a mixture of leukotrienes C4, D4 and E4. Leukotriene synthesis by human gastrointestinal tissues was inhibited by the lipoxygenase inhibitor nordihydroguaiaretic acid (NDGA) and the dual enzyme inhibitor BW755C (3-amino-1-(trifluoromethylphenyl)-2-pyrazoline hydrochloride). Synthesis of prostaglandin E2 was inhibited by the cyclooxygenase inhibitor indomethacin as well as by BW755C. Incubation of gastrointestinal tissues in the presence of glutathione decreased the amounts of leukotrienes D4 and E4, while release of leukotriene C4 was simultaneously increased. On the other hand, incubation of tritiated leukotriene C4 with incubation media from human gastric or colonic mucosa resulted in conversion of the substrate to [3H]leukotriene D4 and [3H]leukotriene E4. The results indicate the capacity of human gastrointestinal tissues to synthesize the 5-lipoxygenase-derived products of arachidonate metabolism, leukotriene B4 and sulfidopeptide leukotrienes, in addition to larger amounts of prostaglandin E2. Furthermore, considerable activities of the sulfidopeptide leukotriene-metabolizing enzymes gamma-glutamyl transpeptidase and dipeptidase were detected in human gastrointestinal tissues. These enzymes might play an important role in biological inactivation and/or change

Topics: 4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine; Adenocarcinoma; Arachidonic Acid; Arachidonic Acids; Calcimycin; Chromatography, High Pressure Liquid; Colonic Neoplasms; Digestive System; Dinoprostone; Gastric Mucosa; Humans; In Vitro Techniques; Intestinal Mucosa; Leukotriene B4; Masoprocol; Muscle, Smooth; Prostaglandins E; Pyrazoles; SRS-A