Monocyte chemoattractant protein 1 and CD40 ligation have a synergistic effect on vascular endothelial growth factor production through cyclooxygenase 2 upregulation in gastric cancer

Background Recent studies have reported that expression of monocyte chemoattractant protein 1 (MCP-1) and its receptor (CCR2) and CD40 ligation on mesenchymal cells play important roles in tumor development. Cyclooxygenase 2 (COX-2) has also been shown to contribute to tumor angiogenesis. We examined the interaction between MCP-1 and CD40 ligation in mesenchymal cells in gastric cancer to determine the effect of these factors on vascular endothelial growth factor (VEGF) production via upregulation of COX-2 expression. Methods COX-2, prostaglandin E₂ (PGE₂), and VEGF production were evaluated in CD40 ligand (CD40L)-stimulated macrophages. CD40L and MCP-1 mRNA levels in gastric cancer tissues were evaluated by real-time polymerase chain reaction (PCR). Localizations of MCP-1, CD40L, CD34, CD40, and CCR2 in 34 gastric cancer tissue specimens were evaluated by single-or double-label immunohistochemistry. Results COX-2 expression levels were significantly higher in CD40L-stimulated macrophages and correlated with increased PGE₂ and VEGF production. Addition of MCP-1 to CD40L-stimulated macrophages had a synergistic effect on COX-2 expression and subsequent PGE₂ and VEGF production. CD40L and MCP-1 mRNA levels were significantly higher in poorly differentiated gastric cancers than in H. pylori-infected gastritis patients. High microvessel density was significantly associated with MCP-1 and CCR2 scores and lymph node metastasis. Conclusions MCP-1 and CD40L had a synergistic effect on COX-2 expression and subsequent VEGF production in gastric cancer.     

Anti-metastatic action of non-steroidal anti-inflammatory drugs

Epidemiological studies suggest that nonsteroidal anti-inflammatory drugs (NSAIDs) reduce the incidence and mortality of several types of human cancer. However, the molecular mechanisms by which NSAIDs exert their chemopreventive and anticancer effects are not fully understood. Cyclooxygenase 1 (COX-1) and COX-2 are the main targets for NSAIDs. Recent studies demonstrate that COX-2 is overexpressed in many human cancers and may promote tumorigenesis via: (1) stimulation of cancer cell proliferation; (2) increase of tumor angiogenesis; (3) prevention of cancer cell apoptosis; (4) modulation of immunoregulatory reactions; and (5) enhancement of tumor metastasis. NSAIDs may target the signaling molecules (from upstream activators to downstream effectors) involved in these mechanisms to attenuate the development and progression of cancer. In this review, we discuss the recent findings with regard to the mechanisms by which NSAIDs inhibit tumorigenesis and will specifically focus on the elucidation of NSAID-induced inhibition of tumor metastasis.     

Enhanced cyclooxygenase-2 expression in sporadic and familial adenomatous polyposis of the human colon

BACKGROUND: The cyclooxygenase (COX) enzymes exist in two related but unique isoforms (COX-1 and COX-2) and catalyze the formation of prostaglandins (PGs). COX-1 is constitutively expressed, and is responsible for the synthesis of PGs necessary for gastroprotection and normal renal function. The COX-2 isoform is important in a variety of pathophysiological conditions such as inflammation and tumorigenesis. Numerous studies report that regular use of non-steroidal anti-inflammatory drugs (NSAIDs) can decrease the incidence of some tumor types, including gastrointestinal polyposis. METHODS: In this study, we evaluated COX-1 and COX-2 expression in 30 polyps collected from 10 patients with familial adenomatous polyposis (FAP) and in 18 polyps collected from 18 patients with sporadic adenomatous polyposis (SAP) using COX-1 or COX-2 isoform-specific antibodies. All tissues were formalin-fixed and paraffin-embedded. Immunoreactivity was detected using tyramide signal amplification and evaluated utilizing an immunohistochemical scoring system. RESULTS: COX-2 was minimally detected in the distant non-neoplastic epithelium, which also served as an internal negative control. In comparison, all polyps collected from SAP or FAP patients overexpressed COX-2 in the neoplastic epithelial cells (P < or = 0.002). Additionally, pronounced COX-2 expression was observed in the stromal cells underlying and adjacent to adenomatous lesions. COX-1 immunoreactivity was weak to mild throughout each tissue evaluated and did not change in the neoplastic or stromal cells of the polyps. CONCLUSIONS: COX-2 expression is upregulated in the adenomatous epithelium of SAP and FAP, while the COX-1 isoform appears to be constitutively expressed at low levels in both neoplastic and non-neoplastic regions. The differential expression of COX-1 and COX-2 in these neoplasms suggests that COX-2 rather than COX-1 may play a role in adenoma formation and/or growth in cases of SAP and FAP in humans.     

Dexamethasone Damages the Rat Stomach but Not Small Intestine During Inhibition of COX-1

We previously reported that inhibition of both COX-1 and COX-2 is required for the gastrointestinal ulcerogenic properties of nonsteroidal anti-inflammatory drugs (NSAIDs). Inhibition of COX-1 up-regulates COX-2 expression, and the prostaglandins (PGs) produced by COX-2 help to maintain the mucosal integrity during inhibition of COX-1. In the present study we investigated whether dexamethasone damages rat gastrointestinal mucosa during inhibition of COX-1 and further developed the idea that COX-2 expression is a key event in the ulcerogenic actions of NSAIDs. Dexamethasone was given p.o. in the absence or presence of SC-560 (a selective COX-1 inhibitor), and the stomach or intestine was examined 8 or 24 hr later, respectively. Neither dexamethasone nor SC-560 alone damaged the gastrointestinal mucosa. In the presence of SC-560, however, dexamethasone damaged the stomach but not small intestine. SC-560 decreased PGE₂ levels in both tissues, with a gradual recovery accompanying the up-regulation of COX-2 expression, and both the recovery of PGE₂ levels and the expression of COX-2 were inhibited by dexamethasone. In the animals treated with SC-560, iNOS expression was up-regulated in the intestinal but not the gastric mucosa, and this response was also inhibited by dexamethasone. These results suggest a risk from steroid therapy in the stomach when COX-2 expression is up-regulated. Dexamethasone does not provoke damage in the intestine, despite inhibiting the up-regulation of COX-2 expression under conditions of PG deficiency; at least one of the reasons is that this agent prevents the expression of iNOS, a major factor in the pathogenesis of intestinal lesions.     

Geranylgeranylacetone Induces Cyclooxygenase-2 Expression in Cultured Rat Gastric Epithelial Cells Through NF-κB

Geranylgeranylacetone (GGA) effectively protects the gastric mucosa against noxious agents. The precise mechanisms underlying the gastroprotective actions of GGA are not known. To elucidate the precise mechanism of GGA, the effect of GGA treatment on COX-2 expression in rat gastric epithelial (RGM1) cells was investigated. We used a prostaglandin E₂ (PGE₂) enzyme-linked immunoassay kit and Western blot analysis to measure PGE₂ production and COX-2 induction by GGA treatment in serum-starved RGM1 cells. Gel-shift assay, Western blot analysis, and a reporter assay were performed to determine which COX-2 promoter was involved in GGA-induced COX-2 expression. GGA treatment dose dependently increased COX-2 expression and PGE₂ production. The nuclear factor (NF)-κB sites of the COX-2 gene promoter were critical for GGA-mediated COX-2 expression. GGA induces COX-2 expression and increases PGE₂ production in serum-starved RGM1 cells via activation of the NF-κB sites of COX-2 gene promoters. The first two authors contributed equally to this work.     

Deletion of cytosolic phospholipase A(2) suppresses Apc(Min)-induced tumorigenesis

Although nonsteroidal antiinflammatory drugs (NSAIDs) show great promise as therapies for colon cancer, a dispute remains regarding their mechanism of action. NSAIDs are known to inhibit cyclooxygenase (COX) enzymes, which convert arachidonic acid (AA) to prostaglandins (PGs). Therefore, NSAIDs may suppress tumorigenesis by inhibiting PG synthesis. However, various experimental studies have suggested the possibility of PG-independent mechanisms. Notably, disruption of the mouse group IIA secretory phospholipase A(2) locus (Pla2g2a), a potential source of AA for COX-2, increases tumor number despite the fact that the mutation has been predicted to decrease PG production. Some authors have attempted to reconcile the results by suggesting that the level of the precursor (AA), not the products (PGs), is the critical factor. To clarify the role of AA in tumorigenesis, we have examined the effect of deleting the group IV cytosolic phospholipase A(2) (cPLA(2)) locus (Pla2g4). We report that Apc(Min/+), cPLA(2)(-/-) mice show an 83% reduction in tumor number in the small intestine compared with littermates with genotypes Apc(Min/+), cPLA(2)(+/-) and Apc(Min/+), cPLA(2)(+/+). This tumor phenotype parallels that of COX-2 knockout mice, suggesting that cPLA(2) is the predominant source of AA for COX-2 in the intestine. The protective effect of cPLA(2) deletion is thus most likely attributed to a decrease in the AA supply to COX-2 and a resultant decrease in PG synthesis. The tumorigenic effect of sPLA(2) mutations is likely to be through a completely different pathway.     

Factors Involved in Upregulation of Inducible Nitric Oxide Synthase in Rat Small Intestine Following Administration of Nonsteroidal Anti-inflammatory Drugs

We investigated the functional mechanisms underlying the expression of inducible nitric oxide (NO) synthase (iNOS) in the rat small intestine following the administration of nonsteroidal anti-inflammatory drugs (NSAIDs) and found a correlation with the intestinal ulcerogenic properties of NSAIDs. Conventional NSAIDs (indomethacin, dicrofenac, naproxen, and flurbiprophen), a selective cyclooxygenase (COX)-1 inhibitor (SC-560) and a selective COX-2 inhibitor (rofecoxib) were administered p.o., and the intestinal mucosa was examined 24 hours later. Indomethacin decreased prostaglandin E₂ (PGE₂) production in the intestinal mucosa and caused intestinal hypermotility and bacterial invasion as well as the upregulation of iNOS expression and NO production, resulting in hemorrhagic lesions. Other NSAIDs similarly inhibited PGE₂ production and caused hemorrhagic lesions with intestinal hypermotility as well as iNOS expression. Hypermotility in response to indomethacin was prevented by both PGE₂ and atropine but not ampicillin, yet all these agents inhibited not only bacterial invasion but also expression of iNOS as well, resulting in prevention of intestinal lesions. SC-560, but not rofecoxib, caused a decrease in PGE₂ production, intestinal hypermotility, bacterial invasion, and iNOS expression, yet this agent neither increased iNOS activity nor provoked intestinal damage because of the recovery of PGE₂ production owing to COX-2 expression. Food deprivation totally attenuated both iNOS expression and lesion formation in response to indomethacin. In conclusion, the expression of iNOS in the small intestine following administration of NSAIDs results from COX-1 inhibition and is functionally associated with intestinal hypermotility and bacterial invasion. This process plays a major pathogenic role in the intestinal ulcerogenic response to NSAIDs.     

Inhibition of COX-2 and induction of apoptosis: two determinants of nonsteroidal anti-inflammatory drugs' chemopreventive efficacies in mouse lung tumorigenesis

Recent studies suggested that nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit lung tumorigenesis under conditions that are immunosuppressive. We hypothesized that this inhibition of mouse lung tumorigenesis requires induction of apoptosis and inhibition of COX (cyclooxygenase)-1, COX-2, and the incidence of K-ras mutation. The NSAIDs used in this study include acetylsalicylic acid (ASA) that is anti-inflammatory with COX-1 and COX-2 inhibition and N-[2-(cyclohexyloxy)-4-nitrophenyl]-methanesulfonamide (NS398) that is a specific COX-2 inhibitor. We have previously demonstrated that ASA (147 and 294 mg/kg diet) and NS398 (7 mg/kg diet) inhibited lung tumorigenesis by 31%, 44%, and 34%, respectively, in 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-treated A/J mice. No difference in the incidence and types of K-ras mutations was found between the lung tumors treated with NNK and those treated with NNK/ASA and NNK/NS398. In NNK-treated mice, ASA (394 mg/kg diet) or NS398 significantly increased the apoptotic index, from 0.07 to 0.30 or to 0.33, respectively. ASA (294 mg/kg diet) and NS398 also inhibited the expression of COX-2. Finally, modulation of gene expression by NS398 and ASA (294 mg/kg diet) was determined using Atlas cDNA expression arrays. Expression of cyclin B2 was decreased and expression of Fas-L and BAD were increased in lung tissues treated with both NS398 and ASA. Treatment with NS398 also increased expression of p57kip2 and myosin. These genes modulated by NSAIDs may play a role in mediating the observed chemopreventive effects of the NSAIDs in the mouse lung. Our results demonstrate that lung tumor prevention with NSAIDs involve both the induction of apoptosis and the inhibition of COX-2 expression.     

PPAR-gamma is expressed and NF-kB pathway is activated and correlates positively with COX-2 expression in stromal myofibroblasts surrounding colon adenocarcinomas

Purpose: Accumulated evidence indicates that carcinogenesis is closely associated with the transformation of normal stroma into a ‘reactive’ stromal phenotype. The present study investigated the role of PPARγ, COX-2 and p-IkB-α—important molecular targets of colon cancer chemoprevention—in this stromal remodeling by evaluating and comparing the expression of these factors in stromal myofibroblasts, macrophages and endothelial cells that surround normal colonic mucosa and colon cancer. Methods: Immunohistochemical methodology was employed on archived paraffin-embedded sections prepared from tumors and adjacent normal colon from 45 patients with colon adenocarcinomas. Double immunostaining with the universal marker for myofibroblasts (alpha-smooth muscle actin/α-SMA) as second primary antibody was also performed. Results: Stromal macrophages and endothelial cells expressed these factors both in normal colonic mucosa and colon cancer. By contrast, stromal myofibroblasts expressed PPARγ, COX-2 and p-IkB-α only in colon adenocarcinomas (77.7%, 100% and 100% of cases, respectively) and not in normal colon. COX-2 and p-IkB-α expressions were strongly correlated in these cells (P<0.001). PPARγ, COX-2 and p-IkB-α expression did not correlate with the stage or differentiation of the adenocarcinomas. Conclusions: NF-kB pathway is activated and COX-2 expression is upregulated in stromal myofibroblasts surrounding colon adenocarcinomas compared to normal colon. Induction of COX-2 expression is primarily induced by NF-kB. NSAIDs, selective COX-2 inhibitors and PPARγ ligands may exert their chemoprophylactic properties through direct actions on these cells. G. P. Vandoros and P. A. Konstantinopoulos contributed equally to this work.     

The role of cyclooxygenase selective inhibitors in the gastrointestinal tract

This article reviews the gastrointestinal manifestations of traditional nonsteroidal anti-inflammatory drugs (NSAIDs) and the improved gastrointestinal safety profile of cyclooxygenase selective (COX)-2 inhibitors. By inhibiting the COX enzyme, NSAIDs provide effective analgesia and suppress inflammation in a variety of conditions. Most NSAIDs (nonselective or traditional) not only inhibit prostaglandins at sites of inflammation but also inhibit prostaglandins that have important normal functions in other parts of the body. This may be harmful when normal gastrointestinal mucosal function is impaired and mucosal damage occurs. Although such damage is often trivial and usually not symptomatic, gastrointestinal ulceration may produce pain and, more ominously, lead to bleeding, perforation, or obstruction. A new approach to the gastrointestinal complications of NSAIDs became feasible with the discovery of two isoforms of COX, COX-1 and COX-2, with COX-1 expressed mainly in the gastrointestinal tract. The development of NSAIDs that preferentially inhibit COX-2 offers the promise of relieving pain and inflammation without the side effects attendant to COX-1 blockade. In prospective studies evaluating gastrointestinal ulceration with COX-2-specific NSAIDs, rates of endoscopic ulceration have been equivalent to those with placebo and much lower than those with nonselective NSAIDs. In the recently released studies of gastrointestinal outcomes (perforated, painful, or bleeding ulcers), incidence of clinically relevant ulceration with COX-2 NSAIDs is much lower than that of traditional NSAIDs.     

The role of prostaglandins and other eicosanoids in the gastrointestinal tract

Nonsteroidal anti-inflammatory drugs (NSAIDs) are generally prescribed to ameliorate symptoms associated with acute pain and chronic inflammatory diseases such as arthritis. Recent epidemiologic studies and clinical trials indicate that use of NSAIDs and cyclooxygenase (COX)-2 selective inhibitors are associated with a reduced risk of certain malignancies, especially gastrointestinal cancer. The cyclooxygenase enzymes are the best known targets of NSAIDs; this diverse class of compounds blocks conversion of arachidonic acid to prostanoids. Prostaglandins and other eicosanoids derived from COX-1 and COX-2 are involved in a variety of physiologic and pathologic processes in the gastrointestinal tract. Recent efforts to identify the molecular mechanisms by which COX-2-derived prostanoids exert their proneoplastic effects have provided a rationale for the possible use of NSAIDs alone or in a combination with conventional or experimental anticancer agents for the treatment or prevention of gastrointestinal cancers.     

Enhanced Cyclooxygenase-2 Expression in Sporadic and Familial Adenomatous Polyposis of the Human Colon

Background: The cyclooxygenase (COX) enzymes exist in two related but unique isoforms (COX-1 and COX-2) and catalyze the formation of prostaglandins (PGs). COX-1 is constitutively expressed, and is responsible for the synthesis of PGs necessary for gastroprotection and normal renal function. The COX-2 isoform is important in a variety of pathophysiological conditions such as inflammation and tumorigenesis. Numerous studies report that regular use of non-steroidal anti-inflammatory drugs (NSAIDs) can decrease the incidence of some tumor types, including gastrointestinal polyposis. Methods: In this study, we evaluated COX-1 and COX-2 expression in 30 polyps collected from 10 patients with familial adenomatous polyposis (FAP) and in 18 polyps collected from 18 patients with sporadic adenomatous polyposis (SAP) using COX-1 or COX-2 isoform-specific antibodies. All tissues were formalin-fixed and paraffin-embedded. Immunoreactivity was detected using tyramide signal amplification and evaluated utilizing an immunohistochemical scoring system. Results: COX-2 was minimally detected in the distant non-neoplastic epithelium, which also served as an internal negative control. In comparison, all polyps collected from SAP or FAP patients overexpressed COX-2 in the neoplastic epithelial cells (P     

Functional coupling between secretory phospholipase A2 and cyclooxygenase-2 and its regulation by cytosolic group IV phospholipase A2

Secretory phospholipase A₂ (sPLA₂) is the major effector involved in arachidonic acid (AA) mobilization and prostaglandin E₂ (PGE₂) production during stimulation of P388D₁ macrophages with the inflammatory stimuli bacterial lipopolysaccharide and platelet-activating factor. We herein demonstrate that PGE₂ in stimulated P388D₁ cells is accounted for by the inducible cyclooxygenase (COX)-2. COX-1, though present, appears not to participate significantly in stimulus-induced PGE₂ production in P388D₁ macrophages. Reconstitution experiments utilizing exogenous recombinant sPLA₂ demonstrate that activation of the sPLA₂ at the plasma membrane is highly dependent on previous activation of the cytosolic phospholipase A₂ (cPLA₂). Collectively these results demonstrate (i) that functional coupling exists between sPLA₂ and COX-2 in activated cells, (ii) the critical role that cPLA₂ plays in lipid mediator production, and (iii) that there is crosstalk between cPLA₂ and sPLA₂ in the cell.     

Leukemic cells modulate induction of COX-2 in human stromal fibroblasts

The interaction of cancer cells with surrounding normal tissue cells is of utmost importance for their survival and tumor progression. For these purposes the cancer cells exploit normal tissue responses associated with inflammation and tissue repair. In the immediate tumor microenvironment one of the early stromal markers is cyclooxygenase-2 (COX-2). In this study we evaluated the effect of leukemia cell lines on nemosis-induced COX-2 expression in stromal fibroblasts. We found that THP-1 cells were the most potent leukemic cells (IC50=746) to suppress COX-2 expression. The U-937 cell line exhibited similar suppressive potency (IC50=921), whereas the KG-1 cell line (IC50=3519) was the least potent to affect COX-2 expression in the stromal cells. Our study shows that human leukemic cells can actively participate in modulation of stromal inflammation via inhibition of COX-2 expression. In a co-culture model of leukemia cell lines and stromal fibroblasts, our data suggest that the tumor-stromal interactions are complexly regulated, and the straightforward association of COX-2 expression with tumor progression may require re-evaluation since some tumor cells, e.g. from hematologic malignancies, may differentially modulate inflammation and COX-2 expression.     

Induction of cyclooxygenase-2 in monocyte/macrophage by mucins secreted from colon cancer cells

Up-regulation of cyclooxygenase-2 (COX-2) and overproduction of prostaglandins have been implicated in the initiation and/or progression of colon cancer. However, it is uncertain in which cells and how COX-2 is induced initially in the tumor microenvironment. We found that a conditioned medium of the colon cancer cell line, LS 180, contained a factor to induce COX-2 in human peripheral blood mononuclear cells. This factor was purified biochemically and revealed to be mucins. A small amount of mucins (approximately 100 ng of protein per ml) could elevate prostaglandin E2 production by monocytes. The mucins induced COX-2 mRNA and protein levels of monocytes in a dose- and time-dependent manner, indicating a COX-2-mediated pathway. We also have examined immunohistochemically the localization of COX-2 protein and mucins in human colorectal cancer tissues. It is noteworthy that COX-2-expressing macrophages were located around the region in which mucins were detectable, suggesting that COX-2 also was induced by mucins in vivo. These results suggest that mucins produced by colon cancer cells play a critical role in the initial induction of COX-2 in the tumor microenvironment.     

NSAR und Coxibe: aktueller Stand

NSAIDs (non steroidal anti-inflammatory drugs) are a crucial component for the therapy of pain induced by inflammatory and degenerative joint diseases. Nevertheless their known therapeutic efficacy is contrasted by significant side effects. The recently developed selective COX-2-inhibitors appear to have a better gastrointestinal safety profile, especially relevant to patients with an increased risk for gastrointestinal ulcers and bleeding. However, this effect may similarly be reached by the combination of NSAIDs with proton pump inhibitors. Recent data relate to an elevated myocardial infarction rate in patients using COX-2-inhibitors. This risk may also occur in conventional NSAIDs. Therefore an individual risk calculation is necessary before COX-2-inhibitors or NSAIDs are used. Treatment should be performed with the lowest dosage for the shortest time possible. Combination therapy with salicylic acid seems to abolish the protective effect of COX-2-inhibitors in the GI-tract. Definite risk factors for the treatment with NSAIDs and COX-2-inhibitors have to be defined in further studies providing the best treatment schedule for an individual patient.     

NSAIDs and COX-2-inhibitors: current status

NSAIDs (non steroidal anti-inflammatory drugs) are a crucial component for the therapy of pain induced by inflammatory and degenerative joint diseases. Nevertheless their known therapeutic efficacy is contrasted by significant side effects. The recently developed selective COX-2-inhibitors appear to have a better gastrointestinal safety profile, especially relevant to patients with an increased risk for gastrointestinal ulcers and bleeding. However, this effect may similarly be reached by the combination of NSAIDs with proton pump inhibitors. Recent data relate to an elevated myocardial infarction rate in patients using COX-2-inhibitors. This risk may also occur in conventional NSAIDs. Therefore an individual risk calculation is necessary before COX-2-inhibitors or NSAIDs are used. Treatment should be performed with the lowest dosage for the shortest time possible. Combination therapy with salicylic acid seems to abolish the protective effect of COX-2-inhibitors in the GI-tract. Definite risk factors for the treatment with NSAIDs and COX-2-inhibitors have to be defined in further studies providing the best treatment schedule for an individual patient.     

Correlation between expression of cyclooxygenase-2 and the presence of inflammatory cells in human primary hepatocellular carcinoma: Possible role in tumor promotion and angiogenesis

AIM: To investigate the association of cyclooxygenase-2(COX-2) expression with angiogenesis and the number and type of inflammatory cells (macrophages/Kupffer cells;mast cells) within primary hepatocellular carcinoma (HCC)tissues and adjacent non-tumorous (MT) tissues.METHODS: Immunohistochemistry for COX-2, CD34,CD68 and mast cell tryptase (MCT) was performed on 14 well-characterized series of liver-cirrhosis-associated HCC patients. COX-2 expression and the number of inflammatory cells in tumor lesions and surrounding liver tissues of each specimen were compared. Moreover,COX-2, CD34 staining and the number of inflammatory cells in areas with different histological degrees within each tumor sample were comparatively analyzed.RESULTS: The percentage of COX-2 positive cells was significantly higher in NT tissues than in tumors. COX-2 expression was higher in well-differentiated HCC than in poorly-differentiated tissues. Few mast cells were observed within the tumor mass, whereas a higher number was observed in the surrounding tissue, especially in peri-portal spaces of NT tissues. Abundant macrophages/Kupffer cells were observed in NT tissues, whereas the number of cells was significantly lower in the tumor mass.However, a higher cell number was observed in the well-differentiated tumor and progressively decreased in relation to the differentiation grade. Within the tumor, a positive correlation was found between COX-2 expression and the number of macrophages/Kupffer cells and mast cells. Moreover, there was a positive correlation between CD34 and COX-2 expression in tumor tissues. Comparison between well- and poorly-differentiated HCC showed that the number of CD34-positive cells decreased with dedifferentiation. However, COX-2 was the only independent variable showing a positive correlation with CD34 in a multivariate analysis.CONCLUSION: The presence of inflammatory cells and COX-2 expression in liver tumor suggests a possible relationship with tumor angiogenesis. COX-2 expressing cells and the number of macrophages/Kupffer cells and mast cells decrease with progression of the disease.