5-lipoxygenase mediates docosahexaenoyl ethanolamide and N-arachidonoyl-L-alanine-induced reactive oxygen species production and inhibition of proliferation of head and neck squamous cell carcinoma cells.

BACKGROUND: Endocannabinoids have recently drawn attention as promising anti-cancer agents. We previously observed that anandamide (AEA), one of the representative endocannabinoids, effectively inhibited the proliferation of head and neck squamous cell carcinoma (HNSCC) cell lines in a receptor-independent manner. In this study, using HNSCC cell lines, we examined the anti-cancer effects and the mechanisms of action of docosahexaenoyl ethanolamide (DHEA) and N-arachidonoyl-L-alanine (NALA), which are polyunsaturated fatty acid (PUFA)-based ethanolamides like AEA. METHODS AND RESULTS: DHEA and NALA were found to effectively inhibit HNSCC cell proliferation. These anti-proliferative effects seemed to be mediated in a cannabinoid receptor-independent manner, since the antagonist of cannabinoid receptor-1 (CB1) and vanilloid receptor-1 (VR1), two endocannabinoid receptors, did not reverse the ability of DHEA and NALA to induce cell death. Instead, we observed an increase in reactive oxygen species (ROS) production and a decrease of phosphorylated Akt as a result of DHEA and NALA treatment. Antioxidants efficiently reversed the inhibition of cell proliferation and the decrease of phosphorylated Akt induced by DHEA and NALA; inhibition of 5-lipoxygenase (5-LO), which is expected to be involved in DHEA- and NALA-degradation pathway, also partially blocked the ability of DHEA and NALA to inhibit cell proliferation and phosphorylated Akt. Interestingly, ROS production as a result of DHEA and NALA treatment was decreased by inhibition of 5-LO. CONCLUSIONS: From these findings, we suggest that ROS production induced by the 5-LO pathway mediates the anti-cancer effects of DHEA and NALA on HNSCC cells. Finally, our findings suggest the possibility of a new cancer-specific therapeutic strategy, which utilizes 5-LO activity rather than inhibiting it.

Activation of K(+) channel by 1-EBIO rescues the head and neck squamous cell carcinoma cells from Ca(2+) ionophore-induced cell death.

Ion channels in carcinoma and their roles in cell proliferation are drawing attention. Intracellular Ca(2+) ([Ca(2+)]i)-dependent signaling affects the fate of cancer cells. Here we investigate the role of Ca(2+)-activated K(+) channel (SK4) in head and neck squamous cell carcinoma cells (HNSCCs) of different cell lines; SNU-1076, OSC-19 and HN5. Treatment with 1 µM ionomycin induced cell death in all the three cell lines. Whole-cell patch clamp study suggested common expressions of Ca(2+)-activated Cl(-) channels (Ano-1) and Ca(2+)-activated nonselective cation channels (CAN). 1-EBIO, an activator of SK4, induced outward K(+) current (ISK4) in SNU-1076 and OSC-19. In HN5, ISK4 was not observed or negligible. The 1-EBIO-induced current was abolished by TRAM-34, a selective SK4 blocker. Interestingly, the ionomycin-induced cell death was effectively prevented by 1-EBIO in SNU-1076 and OSC-19, and the rescue effect was annihilated by combined TRAM-34. Consistent with the lower level of ISK4, the rescue by 1-EBIO was least effective in HN5. The results newly demonstrate the role of SK4 in the fate of HNSCCs under the Ca(2+) overloaded condition. Pharmacological modulation of SK4 might provide an intriguing novel tool for the anti-cancer strategy in HNSCC.

The effect of underwater gait training on balance ability of stroke patients.

[Purpose] The purpose of this study was to investigate the effects of underwater treadmill gait training on the balance ability of stroke patients. [Subjects] Twenty-two patients with stroke were randomly assigned to an underwater treadmill group (n =11) or a control group (n =11). [Methods] Both groups received general rehabilitation for 30 min per session, 5 times per week, over a 4-week period. The underwater treadmill group received additional underwater gait training for 30 min per session, 5 times per week, over the same 4-week period. Static and dynamic balances were evaluated before and after the intervention. [Results] The means of static and dynamic balance ability increased significantly in both groups, but there was no significant difference between the two groups. [Conclusion] Compared to the general rehabilitation program, underwater treadmill gait training was not more effective at improving the balance ability of stroke patients than land-based training.

Lesion activity assessment of early caries using dye-enhanced quantitative light-induced fluorescence.

We aimed to determine whether dye-enhanced quantitative light-induced fluorescence (DEQLF), wherein porous structure of caries lesions is stained with a fluorescent dye, could quantitatively distinguish between active and inactive caries. A total of 126 bovine specimens were prepared to artificially simulate caries activity. Active caries were demineralized with 1% carbopol solution for 3 (A3), 5 (A5), and 10 days (A10). For inactive caries, half specimens in each group were remineralized with 2% NaF and reallocated into three groups (I3, I5, and I10, respectively). Wet specimens were dried with compressed air for 10 s and then dyed with 100-µM sodium fluorescein for 10 s. Fluorescence images of speicmens were captured with a QLF-digital 2 + Biluminator. Fluorescence intensity (ΔG) was measured in fluorescence images of dyed specimens. ΔG between active and inactive groups was compared using independent t-test, and ΔG among active groups (or inactive groups) were compared using ANOVA (α = 0.05). ΔG in the active groups was 33.7-59.0 higher than that in the inactive groups (P < 0.001). Except between I3 and I5, there was significant differences in ΔG according to the demineralization period (P < 0.001). DEQLF might be used to evaluate early caries activity, and longitudinally monitor changes in lesion activity.

The effects of the stromal cell-derived cyclooxygenase-2 metabolite prostaglandin E2 on the proliferation of colon cancer cells.

It is well known that tumor-surrounding stromal tissues support tumor development through secreting soluble factors such as various cytokines, chemokines, and growth factors. It has also been suggested that tumor-associated fibroblast and immune cells have a high expression of cyclooxygenase-2 (COX-2) and produce and secrete several prostaglandins (PGs) to adjacent cancer tissues. From these findings, we assumed that COX-2 inhibition might have an anticancer effect on cancer cells even without COX-2 expression in COX-2-dependent mechanisms through blocking the effect of stroma-derived PGs. Here, because of the complex involvement of various factors in vivo, we investigated this possibility with an in vivo-mimicking model using a Transwell system. To test our hypothesis, we used COX-2-transfected cell lines as stromal cells in our model. When we cocultured cancer cells (LS174T cells without COX-2 expression) with COX-2-high stromal cells in the Transwell membrane system, we observed that the proliferation of cancer cells was promoted and vascular endothelial growth factor synthesis was up-regulated significantly. These effects were blocked completely by COX-2 inhibitors and phosphoinositide-3-kinase inhibitors and partially by the PG E(2) receptor 4 antagonist. Even if some cancer cells did not express COX-2, they were found to have expression of PG receptors and PG-related downstream signaling molecules associated with cell viability. Our observation suggests that these cells can be influenced by PGs derived from stromal tissues. These findings also suggest that COX-2 inhibitors can be used to control the interaction between cancer and surrounding stromal tissues and suppress the proliferation of cancer cells regardless of the expression of COX-2 in cancer cells.

The influence of cyclooxygenase-1 expression on the efficacy of cyclooxygenase-2 inhibition in head and neck squamous cell carcinoma cell lines.

We have previously observed that cyclooxygenase-2 (COX-2) inhibition blocked the production of vascular endothelial growth factor (VEGF) in some head and neck squamous cell carcinoma (HNSCC) cells. However, as some HNSCC cells showed little response to COX-2 inhibition, although they highly expressed COX-2 and prostaglandin E2, we set out to elucidate what made this difference between them and focused on the possibility of the differential expression of COX-1. In western blotting, we found that COX-1 was expressed in SNU-1041 and SNU-1066, but not in SNU-1076 and PCI-50. Only in those cell lines without expression of COX-1 was VEGF production blocked meaningfully by small interfering RNA of COX-2. However, by cotreating with small interfering RNAs of COX-2 and COX-1, VEGF synthesis and prostaglandin E2 were inhibited in SNU-1041 and SNU-1066, similarly in SNU-1076 and PCI-50 with high expression of only COX-2. We also found that there was no difference in the pattern of prostaglandin synthesis between COX-2 and COX-1 through enzyme-linked immunosorbent assay for various prostaglandins. Our study suggests that, as COX-1 and COX-2 express and affect VEGF synthesis in HNSCC cells, we should check COX-1 expression in investigations on cancer treatment by inhibiting COX-2-induced prostaglandins.

Conditionally replicating adenovirus improves gene replication efficiency and anticancer effect of E1-deleted adenovirus carrying TRAIL in head and neck squamous cell carcinoma.

To overcome the low efficiency of gene therapy, we combined a conditionally replicating adenovirus (CRAd) and an adenoviral vector with a therapeutic gene. CRAd has an oncolytic activity in cancer cells with abnormal Rb activity and helps the replication of therapeutic genes incorporated in the E1-deleted adenovirus. We investigated the anticancer effect of a combination of CRAd and adenovirus carrying tumor necrosis factor-related apoptosis inducing ligand (ad-TRAIL). We expected to see increased gene expression in cancer cells as well as an antitumor effect. With the combined application of CRAd and ad-luciferase in head and neck cancer cell lines, we observed considerably increased luciferase activity that was 10- to 50-fold greater than with ad-luciferase alone. The combination of CRAd and ad-TRAIL showed significant suppression of growth in cell lines and increased the sub-G(1) portion of cells 30-fold compared to any single treatment. The expression of TRAIL was highly amplified by the combined treatment and was accompanied by expression of molecules related to apoptosis. In a xenograft animal model, mice treated with CRAd and ad-TRAIL showed complete regression of established tumors, whereas mice treated with CRAd or ad-TRAIL alone did not. In conclusion, this combined strategy using CRAd and adenovirus carrying a therapeutic gene increased the gene transfer rate and enhanced antitumor effects. We expect that this combination strategy could be extended to a multitarget cancer gene therapy by combining multiple adenoviruses and CRAd.

Celecoxib inhibits cell proliferation through the activation of ERK and p38 MAPK in head and neck squamous cell carcinoma cell lines.

It has been observed that several cyclooxygenase-2 (COX-2) inhibitory chemicals might inhibit proliferation of various cancer cells through COX-2-independent action. We also identified that celecoxib more selectively kills cell lines derived from head and neck squamous cell carcinoma (HNSCC) than its non-cancerous counterparts, irrespective of COX-2 expression. Herein, we investigated whether the regulation of mitogen-activated protein kinases activity might be one of the main mechanisms related to a conspicuous COX-2-independent tumor-killing effect of celecoxib in HNSCC cell lines. We assessed the effect of celecoxib on extracellular signal-regulated kinase (ERK), p38, and c-Jun NH2-terminal kinase activity by a transcription factor activation assay then evaluated, if these factors might be involved in the COX-2-independent tumor-killing effect of celecoxib by blocking their activity. We found that the blocking activation of ERK and/or p38 could reverse the celecoxib-induced cell growth inhibition by 50-80% in HNSCC cell lines, but it was not tested in cancer cells of other types. In conclusion, our study suggests that most COX-2-independent tumor-killing action of celecoxib is mediated by the upregulation of ERK and/or p38 activity in HNSCC cells. These results encourage investigation on the underlying mechanisms and detailed outcomes of mitogen-activated protein kinases activation by celecoxib more concisely, for using its excellent tumor-killing effect more safely in the clinical field of cancer treatment.

Clinical assessment of an automated fluorescent plaque index scoring with quantitative light-induced fluorescence.

OBJECTIVE: The aims of this study were to evaluate the clinical applicability of a new fluorescent plaque index scoring (FPI) with the Turesky modified Quigley-Hein plaque index (mQH) and to evaluate its relationship with plaque maturity. METHODS: In total 69 subjects participated in this study. White-light and fluorescent images of anterior teeth were acquired using a Qraycam (AIOBIO, Seoul, Korea). FPI was obtained from fluorescent images using the proprietary software (Q-Ray v.1.39, Inspektor Research System BV, Amsterdam, The Netherlands). Teeth were stained with a two-tone disclosing agent. mQH was used to manually score the combined red and blue disclosed plaque (Combi-mQH) and blue disclosed plaque (Blue-mQH) with the white-light images. Linear relationships between FPI and Combi-mQH (or Blue-mQH) were evaluated by using simple linear regression analysis. Differences of Combi-mQH (or Blue-mQH) with respect to FPI scores were statistically evaluated by using ANOVA with Duncan post hoc correction. RESULTS: FPI showed a moderate positive correlation with Combi-mQH (r = 0.66, P < 0.001) and a high positive correlation with Blue-mQH (r = 0.78, P < 0.001). The model explanatory power (R2) between FPI and Blue-mQH was 60.8 %, which is 16.8 % higher than the explanatory power observed with Combi-mQH (44.0 %). Both Combi-mQH and Blue-mQH increased significantly with increasing FPI score (P < 0.001). CONCLUSION: In this study we found that the FPI scoring system can be used to detect plaque and quantitatively distinguish plaque levels. In addition, FPI was determined to be useful in clinic because of its ability to detect and distinguish old and mature plaque.

Absence of STAT1 disturbs the anticancer effect induced by STAT3 inhibition in head and neck carcinoma cell lines.

The family of signal transducers and activators of transcription (STAT) are transcription factors. Among them, STAT1 is associated with an apoptosis pathway, while STAT3 is associated with tumorigenicity in various cancer cells. In order to investigate the primary roles of STAT1 and STAT3 in head and neck squamous cell carcinoma (HNSCC), we blocked STAT3 with two JAK inhibitors: AG490 (JAK2-STAT3 pathway inhibitor) and JAK total inhibitor. When we inhibited STAT3 with AG490, significant cell death was observed. However, in the case of JAK kinase total inhibitor, no cell growth retardation was observed. We focused on the role of STAT1 in this phenomenon. Suppression of STAT1 by si-RNA resulted in increased cell survival. Furthermore, the growth inhibitory effect of AG490 was reduced by treatment with si-RNA of STAT1. These results reveal that STAT1 is required to promote the tumor killing effect of STAT3 inhibition in HNSCC.

Comparison of fluorescence parameters between three generations of QLF devices for detecting enamel caries in vitro and on smooth surfaces.

BACKGROUND: This study compared two fluorescence parameters (fluorescence loss [ΔF] and red fluorescence gain [ΔR]) among three generations of quantitative light-induced fluorescence (QLF) systems with the aim of determining the validities of these parameters in the three devices for differentiating the severity of enamel caries. METHODS: Forty-one extracted human premolars and molars with suspected enamel caries were selected. Fluorescence images of all teeth were obtained using first-, second-, and third-generation QLF systems (Inspektor Pro, QLF-D, and Qraycam, respectively). Fluorescence parameters were then calculated using proprietary software. All of the specimens were also categorized histologically using polarized-light microscopy (PLM) based on histological levels related to the lesion depth into sound enamel (S), caries limited to the outer half of the enamel (E1), and caries involving the inner half of the enamel (E2). The Mann-Whitney test with Bonferroni correction was used to compare fluorescence parameters among the three generations of systems. The sensitivity, specificity, and area under the receiver operating characteristics curve (AUC) at two thresholds (S/E1 for detecting enamel caries lesions and E1/E2 for differentiating the caries severity) were calculated for evaluating the validities of the fluorescence parameters obtained using all three generations of QLF devices. RESULTS: ΔF did not differ significantly between the devices at any histological level. In addition, ΔF showed large AUCs at the thresholds of S/E1 and E1/E2 (0.97-0.98 and 0.89-0.90, respectively). On the other hand, ΔR was significantly higher for the third-generation device than for the first- and second-generation devices for E2 lesions (P < 0.001). At the S/E1 threshold, ΔR values of the first- and third-generation devices showed larger AUCs (0.96-0.97) compared with that of the second-generation device (0.91), whereas at the E1/E2 threshold the AUC was the largest for the third-generation device (0.87). CONCLUSIONS: The ΔF fluorescence parameter did not differ between the three generations of QLF devices, and showed high validity values. In terms of ΔR, the devices of all generations also showed good diagnostic performance for quantifying and detecting enamel caries lesions, but the third-generation QLF system produced superior results.

Evaluation of tooth wear by estimating enamel thickness with quantitative light-induced fluorescence technology.

BACKGROUND: Various techniques have been suggested to quantitatively assess tooth wear; most have limited clinical application. The first aim of this in vitro study was to estimate the residual enamel thickness of teeth with various degrees of occlusal wear using quantitative light-induced fluorescence (QLF). The second aim was to identify relationships between the fluorescence parameters of QLF and the conventional tooth wear index (TWI) system. METHODS: Sixty-nine extracted permanent premolars and molars with initial stages of tooth wear (TWI score 1a-2: enamel wear to dentin exposure) were used. Two blinded and trained examiners participated in evaluation procedures. Occlusal QLF-digital (QLF-D) images were acquired for selecting area of interest (AOI) and calculating fluorescence for occlusal tooth wear (ΔFwear) of the AOI by the first examiner. Each specimen was cross-sectioned in the buccal-lingual direction. Enamel thickness from images obtained by stereomicroscopy and TWI of each sample was determined by the second examiner. Spearman correlation was used to determine the relationship of ΔFwear with enamel thickness and TWI. ΔFwear values were compared between histological scores with the Mann-Whitney U test. RESULTS: Seventy-six AOIs were analyzed. As enamel thickness decreased, ΔFwear values significantly increased and strongly correlated with enamel thickness (Spearman rho = -0.825, P < 0.001). There were significant differences in ΔFwear values among TWI scores (P < 0.001); ΔFwear strongly correlated with TWI (Spearman rho = 0.753, P < 0.001). CONCLUSIONS: ΔFwear values, which denote fluorescence difference by using QLF, showed a strong correlation with residual enamel thickness and tooth wear severity.

Diagnosis and management of cracked tooth by quantitative light-induced fluorescence technology.

INTRODUCTION: The aim of this case report was to describe the process of diagnosis and treatment of a cracked tooth using quantitative light-induced fluorescence (QLF). CASE REPORT: A 43-year-old male presented at our dental clinic with a complaint of cold pain in #17 tooth. A routine oral examination with radiography was performed for evaluation of the oral condition and treatment planning. Additionally, QLF image capture was performed using Qraycam and Qraypen (AIOBIO, Seoul, Republic of Korea), to collect white-light and fluorescence images. The #17 tooth was observed to have a crack line, showing red fluorescence, from the distal to mesial aspect on the occlusal surface. Even though there was no visible root fracture in the radiographic image, bone loss was observed. Therefore, we performed periodontal treatment. One month later, a root canal treatment was performed because the patient still complained of pain in the #17 tooth. During this treatment, one fluorescent image and one white light image set was captured with the Qraypen. A crack line showing red fluorescence was observed, while the line was not visible to the naked eye. After treatment, the patient has had no complaint related to this tooth for 3 years until today. CONCLUSIONS: Clinically, use of QLF confirmed the presence of a crack before and during a root canal treatment. Therefore, it is postulated that the QLF technology could objectively facilitate the diagnosis and treatment of a cracked tooth.

Association of CD47 with natural killer cell-mediated cytotoxicity of head-and-neck squamous cell carcinoma lines.

OBJECTIVE: Integrin-associated protein (CD47) binds specifically to the inhibitory receptor signal-regulatory protein. This study was designed to evaluate the role of CD47 in natural killer (NK) cell-mediated cytotoxicity against cancer cells. METHODS: Head-and-neck squamous cell carcinoma (HNSCC) cell lines were analyzed for the expression of CD47 and susceptibility to NK cell-mediated killing. Cytolytic activity was assessed by (51)Cr-specific release assays and by measuring cytokine production. RESULTS: HNSCC cell lines that had high CD47 expression showed lower levels of NK cytotoxicity than those with low CD47 expression. After pre-treating cells with neutralizing major histocompatibility complex (MHC) class I or anti-CD47 antibodies, NK cell-mediated cytotoxicity against HNSCC cell lines increased. In addition, when CD47 cDNA was transfected into Caco-2 cells, NK cell-mediated cytotoxicity decreased. CONCLUSION: These findings suggest that CD47 may play an inhibitory role in NK cell-mediated cytotoxicity against cancer cells, implying a possible mechanism of immune escape in human cancer.

Caries detection and quantification around stained pits and fissures in occlusal tooth surfaces with fluorescence.

Occlusal discoloration due to staining frequently occurs on the pits and fissures of teeth. Noncariogenic discoloration (non-CD) refers to the attachment of staining chromogens to sound surfaces, whereas cariogenic discoloration (CD) represents the discoloration of porous structures due to bacterial metabolites and mineral loss from the enamel surface. This study evaluated whether it is possible to distinguish between non-CD and CD on stained occlusal surfaces with fluorescence assessed by the quantitative light-induced fluorescence (QLF) technology. Sixty-two extracted human permanent teeth with suspected discolorations on the pit and fissure were examined. The maximum values of fluorescence loss (ΔFmax) and red fluorescence gain (ΔRmax) were calculated using QLF images. Using histology as the gold standard, it was found that 12 teeth were sound (non-CD), while 50 teeth had enamel and dentine caries (CD). The validity tests at the enamel histological caries level, ΔRmax (ρ = 0.80) were strongly correlated with the histology (P < 0.001). At the optimum threshold (105.0) of ΔRmax, it showed high levels of sensitivity and specificity (0.96 and 0.83, respectively). Therefore, QLF can be used to distinguish non-CD from CD on occlusal surfaces using red fluorescence values with high validity.

Celecoxib enhances the inhibitory effect of 5-FU on human squamous cell carcinoma proliferation by ROS production.

OBJECTIVES: The role of celecoxib in preventing and treating tumors has attracted broad attention in recent years because of its selective and specific inhibition of COX-2 activity. We investigated the inhibitory effects and mechanisms of celecoxib combined with 5-fluorouracil (5-FU) on proliferation of squamous cell carcinoma cells in vivo and in vitro. STUDY DESIGN: Animal study and basic research. METHODS: SNU-1041 and SNU-1076 squamous cell lines and an orthotopic tongue cancer mouse model were used to study growth inhibition with 5-FU enhanced by celecoxib. Sensitivity of cells to drug treatment was analyzed by the MTT assay, and generation of reactive oxygen species (ROS) was measured using dichlorofluorescein diacetate. Phosphorylation of AKT was detected by Western blotting. Survival analysis in the mouse model was assessed according to combination treatment with 5-FU and celecoxib. RESULTS: Reactive oxygen species production in vitro was highest when celecoxib was administered 48 hours after 5-FU treatment. 5-FU-induced inhibition of cell proliferation was enhanced when combined with celecoxib, which was positively correlated with ROS production. Antioxidant treatment reversed 5-FU-inhibited cell proliferation by up to 60%. Cotreatment with celecoxib and 5-FU partially blocked AKT phosphorylation, although no significant changes in total AKT protein levels were detected. An increased survival time was observed in an orthotopic mouse model treated with a combination of celecoxib and 5-FU compared to treatment with either agent alone. CONCLUSION: Celecoxib may have an enhanced anticancer effect in combination with 5-FU. Reactive oxygen species production may be a key mechanism in this combination therapy by inhibiting the AKT pathway. LEVEL OF EVIDENCE: N/A. Laryngoscope, 127:E117-E123, 2017.

Quantitative light-induced fluorescence technology for quantitative evaluation of tooth wear.

Various technologies used to objectively determine enamel thickness or dentin exposure have been suggested. However, most methods have clinical limitations. This study was conducted to confirm the potential of quantitative light-induced fluorescence (QLF) using autofluorescence intensity of occlusal surfaces of worn teeth according to enamel grinding depth in vitro. Sixteen permanent premolars were used. Each tooth was gradationally ground down at the occlusal surface in the apical direction. QLF-digital and swept-source optical coherence tomography images were acquired at each grinding depth (in steps of 100 µm). All QLF images were converted to 8-bit grayscale images to calculate the fluorescence intensity. The maximum brightness (MB) values of the same sound regions in grayscale images before (MBbaseline) and phased values after (MBworn) the grinding process were calculated. Finally, 13 samples were evaluated. MBworn increased over the grinding depth range with a strong correlation (r=0.994, P<0.001). In conclusion, the fluorescence intensity of the teeth and grinding depth was strongly correlated in the QLF images. Therefore, QLF technology may be a useful noninvasive tool used to monitor the progression of tooth wear and to conveniently estimate enamel thickness.

Nitric oxide upregulates the cyclooxygenase-2 expression through the cAMP-response element in its promoter in several cancer cell lines.

We previously showed that nitric oxide (NO) induces overexpression of cyclooxygenase-2 (COX-2) and production of prostaglandin E(2) in cancer cells. Here, we investigated the mechanisms by which NO induces COX-2 expression in cancer cells. We found that the cAMP-response element (CRE) is a critical factor in NO-induced COX-2 expression in all cells tested. We found that in cancer cells, three transcription factors (TFs) - cAMP response element-binding protein (CREB), activating transcription factor-2 (ATF-2) and c-jun, bound the CRE in the COX-2 promoter, and their activities were increased by addition of the NO donor, S-nitroso-N-acetyl-D,L-penicillamine (SNAP). NO-induced activation of soluble guanylate cyclase (sGC), p38 and c-Jun NH(2)-terminal kinase (JNK) upregulated the three TFs, leading to COX-2 overexpression. Addition of dibutyryl-cGMP (db-cGMP) induced COX-2 expression in a manner similar to SNAP; this induction was blocked by a p38 inhibitor (SB202190), but not by a JNK inhibitor (SP600125). NO-induced cGMP was found to activate CREB and ATF-2 in a p38, but not c-jun-dependent manner, while NO induced JNK in a cGMP-independent manner, leading to subsequent activation of c-jun and ATF-2. These results suggest that the low concentrations of endogenous NO present in cancer cell may induce the expression of many genes, including COX-2, which promotes the growth and survival of tumor cells.

Celecoxib can prevent tumor growth and distant metastasis in postoperative setting.

Much evidence suggests that an inflammatory condition provides a microenvironment favorable for tumor growth. One of the main components in the healing wound is the induction of cyclooxygenase-2 (COX-2) and prostaglandins, and many solid tumors have been known to overexpress COX-2. The present study investigated the relationship between surgical wounds and tumor growth and the roles of COX-2 and inflammatory reaction in this microenvironment. We created surgical wounds in syngeneic mice for the implantation of SCC VII murine cancer cell line. Accelerated tumor growth and increased angiogenesis by surgical wounds were clearly observed in C3H/HeJ mice with SCC VII tumor. The COX-2 expression of peritumoral tissues and leukocyte infiltration partly explained the accelerated tumor growth, especially in the early phase after surgical wounding. Celecoxib had a significantly suppressive effect on tumor growth, angiogenesis, and metastasis in tumor-implanted mice with surgical wounds. This tumor-suppressive action of celecoxib did not show any noticeable side effects on the late wound healing and on the gastrointestinal tracts. Prophylactic use of the drug can be advocated in many clinical situations, such as residual tumors or contamination of surgical fields by tumor cells.

Anticancer effects of anandamide on head and neck squamous cell carcinoma cells via the production of receptor-independent reactive oxygen species.

BACKGROUND: The endocannabinoids, anandamide (AEA) and 2-arachidonoyl glycerol (2-AG), are considered promising potential anticancer agents. In this study, we examined the anticancer effects of AEA and 2-AG in head and neck squamous cell carcinoma (HNSCC) cell lines. METHODS AND RESULTS: Our results showed that AEA effectively inhibited proliferation of HNSCC cells whereas 2-AG did not. The anticancer effect of AEA seemed to be mediated by a receptor-independent mechanism. Inhibitors of AEA intracellular transportation and transfection of HNSCC cells with fatty acid amide hydrolase, a key enzyme in AEA metabolism, reversed AEA-dependent inhibition of cell proliferation. We found that cyclooxygenase-2 (COX-2) did not mediate the anticancer effects of AEA; instead we observed an increase in reactive oxygen species (ROS) production after AEA treatment. Moreover, antioxidants partially reversed AEA-dependent inhibition of cell proliferation. CONCLUSION: These findings suggest that AEA might have anticancer effects on HNSCC cells by mediating an increase in ROS levels through a receptor-independent mechanism.