Thyroid carcinomas found incidentally in the cervical lymph nodes: do they arise from heterotopic thyroid tissues?

PURPOSE: Thyroid carcinomas have been found incidentally in the cervical lymph nodes during surgery for head and neck squamous cell carcinoma (SCC). Such carcinomas have been considered a metastatic focus for malignant transformation of heterotopic thyroid tissue. We report on cases of so-called occult thyroid carcinoma in the cervical lymph nodes, and review the relevant literature. PATIENTS AND METHODS: We encountered 3 cases of incidental papillary carcinoma in the cervical lymph nodes of patients with oral SCC, and consequently reviewed 75 previously reported cases. RESULTS: Among 148 patients with oral SCC who had undergone cervical lymph node dissection, 3 were diagnosed with occult thyroid carcinoma. Papillary carcinomas were found in 3, 10, and 3 lymph nodes in cases 1, 2, and 3, respectively. Computed tomography showed 2 tumor-like shadows and 1 calcified mass in the thyroid gland in cases 2 and 3, respectively. These shadows did not enlarge during the 3 to 5 years of observation, and all patients are alive, without any events related to the neck and thyroid gland. Among the reviewed cases, approximately two fifths were histopathologically or clinically free from thyroid carcinoma. Progressive thyroid carcinoma was not detected in any patient. CONCLUSIONS: We propose the possibility that thyroid carcinoma in the cervical lymph nodes is not necessarily metastatic, but may occasionally arise from heterotopic thyroid tissue.

Enhancement of apoptotic damage of squamous cell carcinoma cells by inhibition of the mitochondrial DNA repairing system.

Mitochondrial DNA (mtDNA) repair systems are thought to be associated with the susceptibility of cancer cells to anticancer agents. The present study investigated the relationship between the susceptibility to gamma-rays and the mtDNA repair ability of oral squamous cell carcinoma (OSC) cell lines. The levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and mtDNA common deletion in both nuclear and mitochondrial DNA of OSC-2, OSC-3 and OSC-6 cells (radio-sensitive cell lines) after gamma-ray-irradiation were higher than those of OSC-1, OSC-4 and OSC-5 cells (radio-resistant cell lines). Compared with OSC-2, OSC-3 and OSC-6 cells, OSC-1, OSC-4 and OSC-5 cells had higher levels of activity of phosphoinositide-3 kinase (PI-3K)/Akt and more strongly expressed 8-hydroxyguanine DNA glycosylase (OGG1), DNA polymerase gamma (POLG) and mitochondrial transcription factor A (Tfam). Down-regulation of these mtDNA-repair-associated molecules by the RNA interference technique enhanced the susceptibility of OSC-2 and OSC-5 cells to gamma-rays, and the expression of Tfam and POLG was down-regulated by inhibitors of PI-3K/Akt signaling. These results indicate that the inhibition of mtDNA repair capacity by PI-3K/Akt signal inhibitors and OGG1 down-regulator in cancer cells may be a useful strategy for cancer treatment when combined with ionizing irradiation and chemotherapeutic drugs.

Multi-institutional phase II trial of S-1 in patients with oral squamous cell carcinoma.

The aim of this study was to investigate the efficacy and safety of an oral fluoropyrimidine anticancer agent, S-1, in patients with oral squamous cell carcinoma. Patients with pathologically confirmed squamous cell carcinoma and at least one measurable lesion were enrolled. Oral administration of S-1 (40 mg/m2 twice daily) for 28 days was followed by a 14-day rest period. A total of 41 consecutive eligible patients were enrolled in the study between October 2002 and August 2004. The sites of the primary tumor were the gingiva (n=18), the tongue (n=12), the palate (n=5), the oral floor (n=4), the buccal mucosa (n=1), and the labial mucosa (n=1). A median of two cycles of treatment (range, 1-5) was administered. A complete response was achieved in nine patients and a partial response in eight patients, for an overall response rate of 41.5% (95% confidence interval, 26.4-56.5%). The 3-year survival rate was 76.4% (95% confidence interval, 62.8-90.0%). Although grade 3 anemia and anorexia occurred in two patients each (4.9%), and grade 3 neutropenia, thrombocytopenia, nausea, vomiting, stomatitis, and diarrhea in one patient each (2.4%), no grade 4 toxicities were observed. S-1 exhibits definite antitumor activity in patients with oral squamous cell carcinoma and is well tolerated.

The endogenous reactive oxygen species promote NF-kappaB activation by targeting on activation of NF-kappaB-inducing kinase in oral squamous carcinoma cells.

Reactive oxygen species (ROS) could stimulate or inhibit NF-kappaB pathways. However, most results have been obtained on the basis of the exogenous ROS and the molecular target of ROS in NF-kappaB signalling pathways has remained unclear. Here, the oral squamous carcinoma (OSC) cells, with a mild difference in the endogenous ROS level, were used to investigate how slight fluctuation of the endogenous ROS regulates NF-kappaB activation. This study demonstrates that NF-kappaB-inducing kinase (NIK) is a critical target of the endogenous ROS in NF-kappaB pathways. The results indicate that ROS may function as a physiological signalling modulator on NF-kappaB signalling cascades through its ability to facilitate the activity of NIK and subsequent NF-kappaB transactivation. In addition, the data are useful to explain why the altered intracellular microenvironment related to redox state may influence biological behaviours of cancer cells.

EGCG-targeted p57/KIP2 reduces tumorigenicity of oral carcinoma cells: role of c-Jun N-terminal kinase.

The green tea polyphenol epigallocatechin-3-gallate (EGCG) regulates gene expression differentially in tumor and normal cells. In normal human primary epidermal keratinocytes (NHEK), one of the key mediators of EGCG action is p57/KIP2, a cyclin-dependent kinase (CDK) inhibitor. EGCG potently induces p57 in NHEK, but not in epithelial cancer cells. In humans, reduced expression of p57 often is associated with advanced tumors, and tumor cells with inactivated p57 undergo apoptosis when exposed to EGCG. The mechanism of p57 induction by EGCG is not well understood. Here, we show that in NHEK, EGCG-induces p57 via the p38 mitogen-activated protein kinase (MAPK) signaling pathway. In p57-negative tumor cells, JNK signaling mediates EGCG-induced apoptosis, and exogenous expression of p57 suppresses EGCG-induced apoptosis via inhibition of c-Jun N-terminal kinase (JNK). We also found that restoration of p57 expression in tumor cells significantly reduced tumorigenicity in athymic mice. These results suggest that p57 expression may be an useful indicator for the clinical course of cancers, and could be potentially useful as a target for cancer therapies.

The involvement of hypoxia-inducible factor-1alpha in the susceptibility to gamma-rays and chemotherapeutic drugs of oral squamous cell carcinoma cells.

The transcription factor hypoxia-inducible factor-1alpha (HIF-1alpha) is the key regulator that controls the hypoxic response of mammalian cells. The overexpression of HIF-1alpha has been demonstrated in many human tumors. However, the role of HIF-1alpha in the therapeutic efficacy of chemotherapy and radiotherapy in cancer cells is poorly understood. In this study, we investigated the influence of HIF-1alpha expression on the susceptibility of oral squamous cell carcinoma (OSCC) cells to chemotherapeutic drugs (cis-diamminedichloroplatinum and 5-fluorouracil) and gamma-rays. Treatment with chemotherapeutic drugs and gamma-rays enhanced the expression and nuclear translocation of HIF-1alpha, and the susceptibility of OSCC cells to the drugs and gamma-rays was negatively correlated with the expression level of HIF-1alpha protein. The overexpression of HIF-1alpha induced OSCC cells to become more resistant to the anticancer agents, and down-regulation of HIF-1alpha expression by small interfering RNA enhanced the susceptibility of OSCC cells to them. In the HIF-1alpha-knockdown OSCC cells, the expression of P-glycoprotein, heme oxygenase-1, manganese-superoxide dismutase and ceruloplasmin were downregulated and the intracellular levels of chemotherapeutic drugs and reactive oxygen species were sustained at higher levels after the treatment with the anticancer agents. These results suggest that enhanced HIF-1alpha expression is related to the resistance of tumor cells to chemo- and radio-therapy and that HIF-1alpha is an effective therapeutic target for cancer treatment.

[Glandular odontogenic cyst: report of two cases with cytokeratin 18 expression].

OBJECTIVE: To report two cases of glandular odontogenic cyst and examine its cytokeratin 18,19 expression. METHODS: Two cases of glandular odontogenic cyst were reported and studied. The cytokeratin 18, 19 expression in these two cases were also investigated using immuno-histochemical staining as well as in the situ hybridization of the cyst epithelium. RESULTS: Histo-pathological examination revealed that ciliated columnar cells, squamous cells and low-columnar cells were found in the superficial layer of the lining epithelium. Several minor salivary glands, mainly composed of seromucous cells were observed near the satellite cyst. CK18 were expressed in all layers of the lining epithelium of varying intensity. CK18 was negative in lining epithelium of the daughter cyst, but CK19 was positive. CK18-mRNA was expressed in all the layers of the lining epithelium, the salivary glands and daughter cysts. CONCLUSIONS: Histological features and CK18 expression may be indicative of the possibility of salivary glandular and odontogenic differentiation.

[Cytokeratin18, 13 and their gene expression in post-operative maxillary cyst linings with metaplastic epithelium].

OBJECTIVE: To study the cytokeratin 18 and 13 and their gene (CK) expression in post-operation maxillary cyst linings with metaplastic epithelium. METHODS: CK expressions were examined with immunohistochemistry in 46 post-operative maxillary cyst (POMC) which were lined with pseudostriated columnar cells only (13 cases), both kinds of columnar and squamous cells (30 cases) and squamous cells only (3 cases). RESULTS: The expressions of CK8, CK13 and CK18 were observed in 39, 9 and all of the 43 columnar epithelial linings, respectively. Metaplastic squamous epithelia expressed more CK13 and less CK18 and CK8. Of the 33 metaplastic linings, 24 expressed CK8, 23 CK13 and 26 linings expressed CK18. The expression of CK13- and CK18-mRNA was generally correlated with the protein expression level. By in situ hybridization, CK18-mRNA expression was observed not only in 26 metaplastic linings which were positive for CK18 protein but also in five of the seven metaplastic linings which did not express CK18 protein. In addition, RT-PCR revealed an expression of CK18-mRNA in all metaplastic squamous linings although the expression level was weaker than that in the columnar epithelial linings. The CK13-mRNA was expressed in a fashion inverse to the CK18-mRNA. CONCLUSIONS: These results indicate that CK18-mRNA is preserved through metaplasia although the protein expression decreases and metaplastic squamous cells differentiate with a decrease of CK18 and an increase of CK13 expression.

Tight junction protein claudin-1 enhances the invasive activity of oral squamous cell carcinoma cells by promoting cleavage of laminin-5 gamma2 chain via matrix metalloproteinase (MMP)-2 and membrane-type MMP-1.

Although adherent junctions have been extensively studied, the role of tight junctions in cancer cell invasion is not sufficiently explored. We investigated whether claudin-1, a component of tight junctions, regulated invasion activity in oral squamous cell carcinoma (OSC) cells. The expression of claudin-1, activity of matrix metalloproteinase (MMP)-2, and cleavage of laminin-5 gamma2 chains were assessed by Western blot analysis, immunohistochemistry, and zymography in OSC cell lines (OSC-4 and NOS-2, highly invasive; OSC-7, weakly invasive) and their xenografts in severe combined immunodeficient (SCID) mice. The influence of claudin-1 small interfering RNA (siRNA) on the invasion activity of the cell lines was also investigated. Compared with OSC-7, both OSC-4 and NOS-2 more strongly expressed claudin-1 and possessed high activities of MMP-2 and MMP-9. Tumors formed in the tongues of SCID mice xenografted with OSC-4, NOS-2, and OSC-7 immunohistochemically revealed strong, moderate, and weak expression of laminin-5 gamma2 chains, respectively, and laminin-5 gamma2 chains were secreted in the conditioned medium of the cancer cells in parallel with the in vivo results. Claudin-1 siRNA largely suppressed the invasion of OSC-4 and decreased the activation of MMP-2, the expression of membrane-type MMP-1 (MT1-MMP), and the cleavage of laminin-5 gamma2. In addition, not only antibodies against MT1-MMP and epidermal growth factor receptor (EGFR) but also MMP-2 and EGFR inhibitors strongly suppressed the invasion activity of OSC-4. These results suggest that claudin-1 up-regulates cancer cell invasion activity through activation of MT1-MMP and MMP-2, which results in enhanced cleavage of laminin-5 gamma2 chains.

Concomitant chemo-radio-immunotherapy has a lethal therapeutic effect on tongue carcinomas independent of the clinical stage and histological characteristics of the tumor.

We examined the effect of concomitant chemo-radio-immunotherapy on 80 patients with tongue carcinoma. Disappearance of the tumor without recurrence was observed in 21 patients (38.9%) in intravenous infusion chemotherapy group (A) and in 20 patients (76.9%) in intra-arterial infusion chemotherapy group (B) (P<0.005). A total of 41 patients (51.3%) were free from the tumor after the combined therapy. Along with the good therapeutic effect, oral function was preserved with minimal impairment of speech and mastication. Tumor stage, the mode of tumor cell invasion and tumor cell differentiation were not correlated with the therapeutic effect. In addition, the expression of p53, p21(Cip1/WAF1) and proliferating cell nuclear antigen did not differ between the patients with lethal and non-lethal effects. The 5-year-survival rate was 56.8% in Group A, 76.9% in Group B and 59.6% overall. Thus, combined chemo-radio-immunotherapy, especially intra-arterial infusion, may bring a universal therapeutic effect in tongue carcinoma regardless of the tumor stage and the expression of cell phase-regulating proteins.

Antimicrobial peptides enhance the candidacidal activity of antifungal drugs by promoting the efflux of ATP from Candida cells.

OBJECTIVES: To establish a novel strategy of fungal infection control. METHODS: We examined the influences of antimicrobial peptides including a synthesized short lactoferrin peptide (FKCRRWQWRM, Peptide 2; Pep2) on the synthesis of Candida cell wall polysaccharides, ergosterol synthesis, membrane permeability and the efflux of ATP. RESULTS: Colony formation of Candida albicans was synergistically suppressed by a combination of low concentrations of each drug and peptide. All peptides and amphotericin B, but not itraconazole, revealed weak inhibitory activities against ergosterol synthesis and the peptides weakly suppressed the synthesis of Candida cell wall components, glucan, mannan and chitin. Cell membrane permeability was not only increased by these peptides but also clearly increased by both amphotericin B and itraconazole. ATP efflux was however up-regulated by low concentrations of the peptides, especially by Pep2 and Hst5, although both antifungal drugs did not exert any influence on ATP efflux. The expression of the Candida drug resistance genes 1 and 2 (CDR1 and CDR2) was increased by both drugs, but this increase was suppressed by each peptide. In addition, larger amounts of amphotericin B and itraconazole remained in Candida cells in the presence of Pep2 or Hst5 due to the lower excretion. The effects of both peptides on ATP efflux and increase of intercellular amphotericin B and itraconazole were blocked by anion channel inhibitors 4,4'-diisothiocyanatestilbene-2, 2'-disulphonic acid and 5-nitro-2-(3-phenylpropylamino) benzoic acid. CONCLUSIONS: The examined peptides, especially Pep2 and Hst5, enhance the candidacidal activity of antifungal drugs by promoting anion channel-associated ATP efflux from Candida cells and decreasing efflux of the drugs, which could be useful clinical applications.

Mechanism of HIF-1alpha-dependent suppression of hypoxia-induced apoptosis in squamous cell carcinoma cells.

The transcriptional factor hypoxia-inducible factor-1 (HIF-1) plays an important role in solid tumor cell growth and survival. Overexpression of HIF-1alpha has been demonstrated in many human tumors and predicts a poor response to chemoradiotherapy. We examined the HIF-1alpha-induced survival pathways in human oral squamous cell carcinoma cell (OSCC) lines. The results showed that forced expression of HIF-1alpha suppressed hypoxia-induced apoptosis of OSCC lines by inhibiting cytochrome c release from mitochondria. Overexpression of HIF-1alpha inhibited the generation of reactive oxygen species (ROS), elevation of intracellular Ca(2+) concentration, reduction of mitochondrial membrane potential, and cytosolic accumulation of cytochrome c, which resulted in the inactivation of caspase-9 and caspase-3. In addition, antiapoptotic Bcl-2 and Bcl-X(L) levels were increased and pro-apoptotic Bax and Bak levels were decreased in the HIF-1alpha-overexpressing OSCC line. Overexpression of HIF-1alpha also increased the levels of phosphorylation of Akt and extracellular signal-regulated kinases (ERK). These findings indicate that HIF-1alpha prevents apoptotic cell death through two mechanisms, including inhibition of cytochrome c release and activation of Akt and ERK.

Inhibition of autoantigen expression by (-)-epigallocatechin-3-gallate (the major constituent of green tea) in normal human cells.

Autoimmune disorders, characterized by inflammation and apoptosis of target cells leading to tissue destruction, are mediated in part by autoantibodies against normal cellular components (autoantigens) that may be overexpressed. For example, antibodies against the autoantigens SS-A/Ro and SS-B/La are primary markers for systemic lupus erythematosus and Sjögren's syndrome. Recently, studies in animals demonstrated that green tea consumption may reduce the severity of some autoimmune disorders, but the mechanism is unclear. Herein, we sought to determine whether the most abundant green tea polyphenol, (-)-epigallocatechin-3-gallate (EGCG), affects autoantigen expression in human cells. Cultures of pooled normal human primary epidermal keratinocytes and of an immortalized human salivary acinar cell line were incubated with 100 microM EGCG (a physiologically achievable level for topical application or oral administration) for various time periods and then analyzed by cDNA microarray analysis, reverse transcription-polymerase chain reaction, and Western blotting for expression of several major autoantigen candidates. EGCG inhibited the transcription and translation of major autoantigens, including SS-B/La, SS-A/Ro, coilin, DNA topoisomerase I, and alpha-fodrin. These findings, taken together with green tea's anti-inflammatory and antiapoptotic effects, suggest that green tea polyphenols could serve as an important component in novel approaches to combat autoimmune disorders in humans.

DNA identification of the pathogen of candidal aspiration pneumonia induced in the course of oral cancer therapy.

Aspiration of oropharyngeal bacteria and fungi is occasionally suspected in patients with pneumonia. A patient with oral carcinoma underwent chemoradioimmunotherapy and, about 4 weeks from the start of the therapy, the patient suffered from severe oral mucositis induced by chemoradiotherapy, and candidal pneumonia was subsequently induced. The candidal pneumonia was insufficiently improved by potent antifungal drugs, taking a lethal course. Randomly amplified polymorphic DNA analysis and DNA sequence examination of strains isolated from the oral cavity 1 week before the onset of pneumonia and autopsied lung revealed the identity of both strains as Candida albicans, and the DNA analysis supported aspiration of oral Candida. These results indicate that the pathogen of the pneumonia, C. albicans, was aspirated from the oral cavity and that oral Candida is easily aspirated and becomes the pathogen of pneumonia.

Green tea polyphenol-induced epidermal keratinocyte differentiation is associated with coordinated expression of p57/KIP2 and caspase 14.

Epigallocatechin-3-gallate (EGCG), the most abundant polyphenol in green tea, exerts chemopreventive effects by selectively inducing apoptosis in tumor cells. In contrast, EGCG accelerates terminal differentiation in normal human epidermal keratinocytes (NHEK) mediated partially by up-regulation of p57/KIP2, a cyclin-dependent kinase inhibitor that confers growth arrest and differentiation. However, it is unclear if EGCG modulates caspase 14, a unique regulator of epithelial cell terminal differentiation associated with cornification. Here, we examined the effect of EGCG on caspase 14 expression in NHEK and correlated the protein and mRNA expression of p57/KIP2 with those of caspase 14 in either normal keratinocytes or p57/KIP2-expressing tumor cells (OSC2, an oral squamous cell carcinoma cell line). Additionally, paraffin-embedded normal and untreated psoriatic (aberrant keratinization) skin sections from humans were assessed for caspase 14 by immunohistochemistry. In NHEK, EGCG induced the expression of caspase 14 mRNA and protein levels within a 24-h period. The expression of p57/KIP2 in OSC2 cells was adequate to induce caspase 14 in the absence of EGCG; this induction of caspase 14 was down-regulated by transforming growth factor-beta1. In human psoriatic skin samples, caspase 14 staining in the upper epidermis was reduced, especially in nuclear areas. These results suggest that, in addition to p57/KIP2, EGCG-induced terminal differentiation of epidermal keratinocytes involves up-regulation of caspase 14. Further understanding of how EGCG modulates cellular differentiation may be useful in developing green tea preparations for selected clinical applications.

Regulation of fungal infection by a combination of amphotericin B and peptide 2, a lactoferrin peptide that activates neutrophils.

To establish a novel strategy for the control of fungal infection, we examined the antifungal and neutrophil-activating activities of antimicrobial peptides. The duration of survival of 50% of mice injected with a lethal dose of Candida albicans (5 x 10(8) cells) or Aspergillus fumigatus (1 x 10(8) cells) was prolonged 3 to 5 days by the injection of 10 microg of peptide 2 (a lactoferrin peptide) and 10 microg of alpha-defensin 1 for five consecutive days and was prolonged 5 to 13 days by the injection of 0.1 microg of granulocyte-monocyte colony-stimulating factor (GM-CSF) and 0.5 microg of amphotericin B. When mice received a combined injection of peptide 2 (10 microg/day) with amphotericin B (0.5 microg/day) for 5 days after the lethal fungal inoculation, their survival was greatly prolonged and some mice continued to live for more than 5 weeks, although the effective doses of peptide 2 for 50 and 100% suppression of Candida or Aspergillus colony formation were about one-third and one-half those of amphotericin B, respectively. In vitro, peptide 2 as well as GM-CSF increased the Candida and Aspergillus killing activities of neutrophils, but peptides such as alpha-defensin 1, beta-defensin 2, and histatin 5 did not upregulate the killing activity. GM-CSF together with peptide 2 but not other peptides enhanced the production of superoxide (O2-) by neutrophils. The upregulation by peptide 2 was confirmed by the activation of the O2- -generating pathway, i.e., activation of large-molecule guanine binding protein, phosphatidyl-inositol 3-kinase, protein kinase C, and p47phox as well as p67phox. In conclusion, different from natural antimicrobial peptides, peptide 2 has a potent neutrophil-activating effect which could be advantageous for its clinical use in combination with antifungal drugs.

Protective effects of EGCG on salivary gland cells treated with gamma-radiation or cis-platinum(II)diammine dichloride.

Dysfunction of salivary glands is often associated with aging and cancer therapy. Green tea polyphenols were previously found to protect normal epithelial cells from reactive oxygen species, and to induce apoptosis in tumor cells. The current study investigated whether -(-) epigallocatechin-3-gallate (EGCG), the major green tea polyphenol, protects normal salivary gland cells from the effects of gamma-irradiation and the chemotherapy drug cis-platinum(II)diammine dichloride (CDDP). Human immortalized salivary acinar and ductal cells, and oral squamous cell carcinoma cells were irradiated with gamma-rays or treated with CDDP, with or without pretreatment with EGCG, followed by MTT and BrdU incorporation assays. The results demonstrated that EGCG protected the normal salivary gland cells from chemical or irradiation-induced damage. However, protection of oral cancer cells by EGCG was also observed if EGCG was at physiologically achievable salivary concentrations but not at higher concentrations, suggesting that the combination of green tea consumption with cancer therapy requires further evaluation.

Reactive oxygen species (ROS) control the expression of Bcl-2 family proteins by regulating their phosphorylation and ubiquitination.

We examined the influence of ROS on the phosphorylation and complex formation of Bcl-2 family proteins in Mn-superoxide dismutase (SOD) antisense-transfected squamous cell carcinoma cells, OSC-4 cells. The increase of intracellular ROS level induced by cis-diamminedichloroplatinum (CDDP) and gamma-ray treatment was greater in antisense-transfected cells than in control vector-transfected cells, and apoptosis was more extensively induced in the former. Antisense-transfected cells expressed high levels of Bax and Bak, but low levels of Bcl-2 and Bcl-XL when treated with CDDP, peplomycin, 5-fluorouracil or gamma-rays. After treatment with these agents, the phosphorylation of protein kinase A, Bcl-2 (Thr56) and Bad (Ser155) was increased, especially in antioxidant (N-acetylcysteine and pyrrolidine dithiocarbamate)-pretreated control cells, but the phosphorylation levels were very low in the antisense-transfected cells. Bcl-2 ubiquitination was increased, but ubiquitination of Bad and Bax was decreased in the antisense-transfected cells, although their ubiquitination was increased by the antioxidants. These results reveal that ROS induce apoptosis by regulating the phosphorylation and ubiquitination of Bcl-2 family proteins, resulting in increased proapoptotic protein levels and decreased antiapoptotic protein expression.

Roles of catalase and hydrogen peroxide in green tea polyphenol-induced chemopreventive effects.

The green tea polyphenol (-)-epigallocatechin-3-gallate (EGCG) possesses promising anticancer potential. Although in vivo studies unveiled the metabolic routes and pharmacokinetics of EGCG and showed no adverse effects, in vitro studies at high concentrations demonstrated oxidative stress. EGCG causes differential oxidative environments in tumor versus normal epithelial cells, but the roles that EGCG, hydrogen peroxide (H2O2), and intracellular catalase play in the epithelial system are largely unknown. The current study employed enzyme activity assays, reactive oxygen species quantification, and immunoblotting to investigate whether EGCG-induced differential effects correlate with levels of key antioxidant enzymes and H2O2. It was found that normal human keratinocytes with high catalase activity are least susceptible to H2O2, whereas H2O2 caused significant cytotoxicity in oral carcinoma cell lines. However, the EGCG-induced differential effects could not be duplicated by H2O2 alone. The addition of exogenous catalase failed to completely prevent the EGCG-induced cytotoxicity and rescue the EGCG-induced growth arrest in the tumor cells. The antioxidant N-acetyl-L-cysteine rescued the tumor cells from H2O2-induced damage only, but not from EGCG-induced mitochondrial damage. Finally, alterations in catalase or superoxide dismutase activities were not observed upon EGCG exposure. In conclusion, although endogenous catalase may play a role in response to H2O2-induced cytotoxicity, the EGCG-induced cytotoxic effects on tumor cells mainly result from sources other than H2O2.

Decreased excretion of antimicrobial proteins and peptides in saliva of patients with oral candidiasis.

BACKGROUND: Antimicrobial peptides in saliva appear to play a crucial role in the regulation of oral Candida growth, and study on antimicrobial excretion in saliva and oral candidiasis appears useful for the analysis of pathophysiology of oral candidiasis. METHODS: To clarify the role of saliva in the regulation of oral Candida growth, the levels of antimicrobial proteins and peptides and their excretion rates were examined in saliva obtained from 50 patients with oral candidiasis and 35 healthy individuals. RESULTS: The inhibitory activities of patients' saliva against Candida adhesion with HeLa cells and against Candida growth (radiolabeled glucose incorporation) were lower than those of saliva from the healthy controls. The salivary levels of lactoferrin (Lf; 11 +/- 9 microg/ml), secretory immunoglobulin A (sIgA; 160 +/- 37 microg/ml), beta-defensin 1 (375 +/- 37 ng/ml), and beta-defensin 2 (412 +/- 51 ng/ml) in the patients were largely lower than those in the control group (33 +/- 14 microg/ml, 204 +/- 51 microg/ml, 452 +/- 89 ng/ml, and 530 +/- 142 ng/ml, respectively), although the transferrin (Tf) and secretory component (SC) levels were almost same in both groups, and alpha-defensin 1 was slightly increased in the patient group (660 +/- 115 ng/ml vs. 467 +/- 168 ng/ml). In addition, the excretion rates of the proteins and peptides were largely decreased in the patients (Tf: 14 +/- 2 microg/10 min vs. 34 +/- 7 microg/10 min; Lf: 18 +/- 11 microg/10 min vs. 139 +/- 43 microg/10 min; sIgA: 300 +/- 132 microg/10 min vs. 900 +/- 207 microg/10 min; SC: 112 +/- 46 microg/10 min vs. 292 +/- 64 microg/10 min; alpha-defensin 1: 1223 +/- 431 ng/10 min vs. 2044 +/- 612 ng/10 min; beta-defensin 1: 687 +/- 243 ng/10 min vs. 1985 +/- 295 ng/10 min; and beta-defensin 2: 784 +/- 299 ng/10 min vs. 2288 +/- 278 ng/10 min). CONCLUSION: These results conclusively suggest that oral candidiasis is associated with salivary gland hypofunction and that decreases of salivary antibacterial proteins induce Candida overgrowth.