Resveratrol Inhibits Proliferation and Induces Autophagy by Blocking SREBP1 Expression in Oral Cancer Cells.

Resveratrol is a polyphenolic antioxidant found in grapes, red wine, and peanuts and has been reported to have anti-neoplastic effects on various cancer types. However, the exact mechanism of its anti-cancer effects in oral cancer is not fully understood and remains controversial. Resveratrol exhibits strong hypolipidemic effects; therefore, we examined its effect on lipid metabolism in oral cancer. Resveratrol significantly reduced cell viability and induced autophagic cell death in oral cancer cells but not in normal cells. This selective effect was accompanied by significantly reduced lipogenesis, which is caused by downregulation of the transcription factor sterol regulatory element-binding protein 1 (SREBP1) gene, followed by downregulation of the epidermal fatty acid-binding protein (E-FABP). It was strongly suggested that resveratrol-induced autophagy resulted from the inhibition of SREBP1-mediated cell survival signaling. Luciferase reporter assay further indicated that resveratrol has a potent and specific inhibitory effect on SREBP1-dependent transactivation. Importantly, resveratrol markedly suppressed the growth of oral cancer cells in an animal xenograft model, without exhibiting apparent cytotoxicity. In conclusion, resveratrol induces autophagy in oral cancer cells by suppressing lipid metabolism through the regulation of SREBP1 expression, which highlights a novel mechanism of the anti-cancer effect of resveratrol.

Down-regulation of Glutathione Peroxidase 4 in Oral Cancer Inhibits Tumor Growth Through SREBP1 Signaling.

BACKGROUND/AIM: This study aimed to elucidate the role of glutathione peroxidase 4 (GPX4) on the sterol regulatory element binding proteins (SREBPs)-proliferation pathway in oral cancer cells, and determine its protein expression in oral cancer tissues. MATERIALS AND METHODS: Quantitative RT-PCR and immunoblot analysis were carried out. Cell viability assay, apoptosis detection assay, immunohistochemistry and GPX4 knockdown were performed. RESULTS: The levels of both GPX4 mRNA and protein were highest in SAS cells. GPX4 knockdown in SAS cells, a human oral squamous cell carcinoma cell line, using GPX4 siRNA resulted in a reduction in cell number, which appeared to be due to non-apoptotic cell death such as ferroptosis. Furthermore, SREBP was clearly down-regulated by GPX4 knockdown in SAS cells. Immunopositivity for GPX4 was revealed on the membrane of human oral squamous cell carcinoma cells, and this was correlated with p53 immunoreactivity. CONCLUSION: GPX4 appears to play an important role in oral cancer proliferation.

The impact of spot size, spacing, pattern, duration and intensity of burns on the photocoagulation index in a geometric simulation of pan-retinal laser photocoagulation.

PURPOSE: To evaluate the impact of spot size, spacing, pattern, duration and intensity of burns on the photocoagulation index, using a geometric simulation of pan-retinal laser photocoagulation. METHODS: Simulations of full-scattered pan-retinal laser photocoagulation were performed on a retinal map, using a geometry-based method. Simulations consisted of 300-, 400- or 500-µm diameter equidistant spots on the retina with 1.0-spot width spacing, as well as 400-µm diameter spots on the retina in an equidistant pattern or grid pattern, with 1.0-, 0.75-, 0.50-, 0.25- or 0-spot width spacing. For each simulation, we calculated the ratio of the total photocoagulated retinal area to the whole retina, termed the photocoagulation index. We recalculated the photocoagulation indexes using the expansion ratios of photocoagulated lesions by different duration and intensity of burns from a previous study. RESULTS: The photocoagulation indexes of the simulated pan-retinal laser photocoagulation with 300-, 400- and 500-µm diameter spots were 20.8%, 20.6% and 21.0%, respectively. The photocoagulation indexes of the 1.0-, 0.75-, 0.50-, 0.25- and 0-spot width spacing configurations of pan-retinal laser photocoagulation burns for the equidistant pattern were 20.6%, 27.1%, 36.7%, 53.2% and 83.1%, respectively, and those for the grid pattern were 17.9%, 23.5%, 31.8%, 46.1% and 72.0%, respectively. The photocoagulation indexes obtained with the equidistant and grid patterns changed (range, 1.7-84.7% and 1.5-73.4%, respectively) when the duration or burn intensity of the pan-retinal photocoagulation was changed. CONCLUSION: This geometric simulation method could evaluate the impact of a range of conditions on the photocoagulation index.

Synergism between α-mangostin and TRAIL induces apoptosis in squamous cell carcinoma of the oral cavity through the mitochondrial pathway.

Mangosteen (Garcinia mangostana) is a tree found in South-East Asia and the pericarp of its fruit has been used in folk medicine for the treatment of many human illnesses. Mangosteen fruit rinds contain a high concentration of xanthone, which is a type of polyphenol. One type of xanthone, α-mangostin, has been reported to exert chemopreventive effects against chemically-induced colon cancer through the decrease of c-Myc expression, suppressing tumor growth in a mouse model of mammary cancer. A recent study demonstrated the inhibitive effect of α-mangostin on the growth of prostate cancer. However, it remains unclear whether α-mangostin induces cell death in oral cancer. The present study examined the impact of α-mangostin on human oral squamous cell carcinoma (HOSCC). Firstly we analyzed the expression of c-Myc in five HOSCC cell lines. The highest expression level of c-Myc mRNA was observed in SAS cells and the lowest in HSC-4 cells. Therefore, SAS cells were treated with α-mangostin, which was found to exert a weak cytocidal effect. Since α-mangostin has been reported to exert synergistic effects on cancers when combined with anticancer drugs, we attempted to evaluate such synergistic effects of α-mangostin when used with a cytokine, tumor necrosis factor (TNF)-related apoptosis­inducing ligand (TRAIL). We found that the combination of α-mangostin with TRAIL induced apoptosis of SAS cells through the mitochondrial pathway via activation of caspase-9 and -3/7, following release of cytochrome c. This apoptosis was induced by S/G2/M-phase arrest. Immunopositivity for c-Myc was observed in the cytoplasm of tumor cells in 16 (40%) of the 40 cases of HOSCC. These data revealed that the combination of α-mangostin and TRAIL may have a considerable potential for the treatment of oral cancer.

Periostitis Ossificans Arising in the Mandibular Bone of a Young Patient: Report of an Unusual Case and Review of the Literature.

Periostitis ossificans, also known as Garré osteomyelitis, is a specific type of chronic osteomyelitis that forms new bone under the periosteum resulting from a periosteal reaction to chronic inflammation or infections. It commonly affects the mandible secondary to odontogenic infection. The therapeutic approach involves eliminating the infectious cause and antibiotic administration. This report describes an unusual case of periostitis ossificans arising from the mandible of an 11-year-old boy. The cause of infection was correlated with a lower right unerupted third molar, which had no obvious connection with the oral cavity. The histologic diagnosis was chronic osteomyelitis with proliferative periostitis. The patient has been followed for 1 year, without any evidence of recurrence. Periostitis ossificans can be diagnostically problematic, and various conditions must be considered in the differential diagnosis.

Simulation of panretinal laser photocoagulation using geometric methods for calculating the photocoagulation index.

PURPOSE: To establish geometrically based methods for simulating panretinal laser photocoagulation (PRP) for the photocoagulation index. METHODS: A formula for calculating the curved surface area of a spherical dome was used for the simulation. If the radius of the dome is c and the height of the dome is h, then the curved surface area (S) of the dome is S = π (c2 + h2). We calculated the area of the whole retina using this formula and the anatomical dimensions of the standard eyeball. To simulate PRP with a 400-µm spot on the retina with 1-spot spacing, we drew 400-µm-diameter circles, separated by 400 µm, on a retinal map. We calculated the ratio of the total retinal photocoagulated area to the whole retina, termed the photocoagulation index, in order to investigate the impact of the extent of the photocoagulated area and the pulse duration on PRP. RESULTS: The whole retinal area was 1,092 mm2. The numbers of spots in the scattered and full-scattered PRP were 1,222 and 1,814, respectively. The photocoagulation index was 14.1% and 20.9% for scattered and full-scattered PRP, respectively. These values changed to 14.3% (5.6%) and 21.3% (8.3%), respectively, for PRP with a 100-ms pulse or a 20-ms pulse. CONCLUSIONS: This method will be useful for investigating the impact of various PRP parameters (duration, spacing, intensity of burns, extent of photocoagulated area, etc.) on the photocoagulation index.

IL-23 promotes growth and proliferation in human squamous cell carcinoma of the oral cavity.

Interleukin (IL)-23 is a heterodimeric cytokine, comprising IL-12p40 and the cloned IL-23-specific p19 subunit, was identified as a cancer-associated cytokine in a recent study. Like IL-12, IL-23 is expressed predominantly by activated dendritic cells and phagocytic cells. These cytokines antagonistically regulate local inflammatory responses in the tumor microenvironment and infiltration by intraepithelial lymphocytes. We have previously demonstrated the expression of IL-23 and its receptors in human oral squamous cell carcinoma (HOSCC) cell lines and tissue. Hence, this study investigated whether IL-23 has a role in the growth and proliferation of oral cancer cells by examining the expression kinetics of IL-23 and NF-kappaB activity, in vitro and in vivo. IL-23, which constitutively expressed in oral cancer, was enhanced by TNF-alpha and IL-23. IL-23 promotes cell proliferation in oral cancer and enhances the transport of nuclear factor-kappaB (NF-kappaB p65, RelA) to the nucleus in HSC-3 cells. Furthermore, luciferase reporter assay showed that IL-23 strongly induces RelA activity, and confirmed this finding by knockdown of IL-23 using RNA interference. Although RelA activity was down-regulated by anti-human IL-23p19 polyclonal antibody, used to neutralize the activity of IL-23, apoptosis was not induced. Immunohistochemistry revealed a weak IL-23 immunoreactivity in the cytoplasm of inflammatory infiltrating cells and in the cancer cells derived from 14 of 40 cases (35%) of oral SCC. In contrast, strong RelA immunoreactivity was observed in 30 of 40 cases of SCC (75%), especially consistent with IL-23 positive cells in SCC tissues. These data suggest that IL-23 up-regulates the growth and cell proliferation of oral cancer by promoting the nuclear transactivation of RelA.

Expression of interleukin-23 and its receptors in human squamous cell carcinoma of the oral cavity.

Interleukin (IL)-23 is a heterodimeric cytokine comprising IL-12p40 and the recently cloned IL-23-specific p19 subunit. Like IL-12, IL-23 is expressed predominantly by activated dendritic cells and phagocytic cells. Both cytokines induce interferon-γ secretion by T cells, and antagonistically regulate local inflammatory responses in the tumor microenvironment as well as the infiltration of intra-epithelial lymphocytes. Although the expression of IL-23 in various organs has been reported, it is unclear whether IL-23 is expressed in oral cancer. The expression of IL-23 and its receptors was examined in human oral squamous cell carcinoma (HOSCC) cell lines and tissue. IL-23 and its receptor mRNAs and proteins were spontaneously expressed, and IL-23 was increased by TNF-α stimulation in the oral cancer cells. A cell proliferation assay confirmed that IL-23 promotes cell proliferation in oral cancer. The localization of IL-23 protein in HOSCC tissue was examined using immunohistochemistry. A positive reaction for anti-IL-23 antibody was weakly observed in the cytoplasm of inflammatory infiltrating cells and cancer cells in HOSCC tissue. Meanwhile, nuclear factor-κB immunoreactivity was strongly positive in HOSCC tissue, which is particularly consistent with the observation of IL-23-positive cells in SCC tissue. These data suggest that IL-23 plays a significant role in the growth and proliferation of oral cancer.

Cimetidine inhibits salivary gland tumor cell adhesion to neural cells and induces apoptosis by blocking NCAM expression.

BACKGROUND: Cimetidine, a histamine type-2 receptor antagonist, has been reported to inhibit the growth of glandular tumors such as colorectal cancer, however the mechanism of action underlying this effect is unknown. Adenoid cystic carcinoma is well known as a malignant salivary gland tumor which preferentially invades neural tissues. We demonstrated previously that human salivary gland tumor (HSG) cells spontaneously express neural cell adhesion molecule (NCAM), that HSG cell proliferation may be controlled via a homophilic (NCAM-NCAM) binding mechanism and that NCAM may be associated with perineural invasion by malignant salivary gland tumors. We further demonstrated that cimetidine inhibited NCAM expression and induced apoptosis in HSG cells. Here, we investigated the effects of cimetidine on growth and perineural/neural invasion of salivary gland tumor cells. METHODS: In this study, we have examined the effect of cimetidine on cancer cell adhesion to neural cells in vitro, one of the critical steps of cancer invasion and metastasis. We have also used an in vivo carcinogenesis model to confirm the effect of cimetidine. RESULTS: We have demonstrated for the first time that cimetidine can block the adhesion of HSG cells to neural cell monolayers and that it can also induce significant apoptosis in the tumor mass in a nude mouse model. We also demonstrated that these apoptotic effects of cimetidine might occur through down-regulation of the cell surface expression of NCAM on HSG cells. Cimetidine-mediated down-regulation of NCAM involved suppression of the nuclear translocation of NF-kappaB, a transcriptional activator of NCAM gene expression. CONCLUSION: These findings suggest that growth and perineural/neural invasion of salivary gland tumors can be blocked by administration of cimetidine via induction of apoptosis and in which NCAM plays a role.

Loss of CYLD might be associated with development of salivary gland tumors.

Molecular studies of cylindromas, which arise from the eccrine or apocrine cells of the skin, have demonstrated frequent alterations at chromosome 16q12-13, recently found to house the cylindromatosis (CYLD) gene. CYLD, a tumor suppressor gene, has deubiquitinating enzyme activity and inhibits the activation of transcription factor NF-kappaB. Loss of the deubiquitinating activity of CYLD is correlated with tumorigenesis. It has been reported that the expression of CYLD is observed in various organs. We demonstrated previously that human salivary gland tumor (SGT) cell line, HSG spontaneously expresses CYLD and also found that adenoid cystic carcinoma (ACC) arising from the hard palate was distinctly positive for CYLD, immunohistochemically. However, it is unclear whether loss of CYLD is associated with development of SGTs. This study examined CYLD function in SGT cells and attempted to clarify whether CYLD is associated with development of SGTs. The expression of CYLD and NF-kappaB mRNAs in HSG cells was increased by TNF-alpha. Translocation of NF-kappaB protein from the cytoplasm to the nucleus in HSG cells peaked at 30 min after TNF-alpha stimulation, then decreased at 60 min, whereas that of CYLD protein increased gradually in a time-dependent manner. Luciferase reporter assay indicated that TNF-alpha induced a 5-fold increase of NF-kappaB-dependent transcription at 4 h, which was further enhanced by knockdown of CYLD using RNA interference. Taken together, these data demonstrated that the levels of both CYLD and NF-kappaB mRNAs accumulated in HSG cells during 24 h after TNF-alpha stimulation, although the NF-kappaB activity in the cells was at least negatively regulated by CYLD. Immunohistochemical examinations revealed that there are several correlations between the expression of CYLD and NF-kappaB-related factors in 17 cases of ACC tissues. These findings suggest that loss of CYLD is associated with development of SGTs.

MTHFR gene polymorphism is susceptible to diabetic retinopathy but not to diabetic nephropathy in Japanese type 2 diabetic patients.

AIMS: Previously, we have proposed that methylenetetrahydrofolate reductase (MTHFR) gene polymorphism (C677T) could be a risk factor for diabetic retinopathy. To support our suggestion, we examined in detail the association of MTHFR polymorphism with diabetic retinopathy and nephropathy in Japanese type 2 diabetic patients. METHODS: Subjects (n=190) were free of cardiovascular diseases and were not on hemodialysis. Retinopathy was assessed according to fundamental differentiation; nephropathy was determined according to urinary albumin level; and MTHFR genotype was determined by polymerase chain reaction-restriction fragment length polymorphism. We also analyzed how hyperglycemia affected these three conditions in 131 patients with glycosylated hemoglobin > or =6.5% and fasting blood sugar > or =110 mg/dl. RESULTS: The frequency of 677T/677T homozygous subjects with retinopathy was higher than the frequencies of the other two genotypes, and a significant difference was observed in the distribution of the genotypes (677C/677C, 41.9%; 677C/677T, 31.1%; 677T/677T, 61.5%; P<.05). The susceptibility of 677T/677T homozygote to retinopathy approached significance [odds ratio (OR)=2.17; 95% confidence interval (95% CI)=0.87-5.42]. However, in the population with hyperglycemia, the 677T/677T homozygote modified the risk for retinopathy (OR=4.30; 95% CI=1.42-13.1), especially the risk for nonproliferative retinopathy. In contrast, the 677T/677T homozygote did not affect the risk for nephropathy (OR=1.17; 95% CI=0.45-3.05), even in subjects with hyperglycemia (OR=1.50; 95% CI=0.50-4.48). CONCLUSIONS: Our results are highly suggestive of an important role for MTHFR genotype in susceptibility to retinopathy under hyperglycemia, but not to nephropathy. Preventive therapies based on MTHFR polymorphism could delay the onset of retinopathy in type 2 diabetic patients.

Cimetidine induces apoptosis of human salivary gland tumor cells.

It has been reported that cimetidine, a histamine type-2 receptor (H2R) antagonist, inhibits the growth of glandular tumors such as colorectal cancer. However, its effects against salivary gland tumors are still unknown. We demonstrated previously that human salivary gland tumor (HSG) cells spontaneously express the neural cell adhesion molecule (NCAM) and also that HSG cell proliferation could be controlled via a homophilic (NCAM-NCAM) binding mechanism and that NCAM may be associated with perineural invasion by malignant salivary gland tumors. In the present study, we investigated the effects of cimetidine via the expression of NCAM on tumor growth and perineural/neural invasion in salivary gland tumor cells. Expression of both NCAM mRNA and protein was found to decrease in a dose-dependent manner upon treatment with cimetidine for 24 h. The MTT assay and confocal laser microscopy clearly showed that HSG cells underwent apoptosis after treatment with cimetidine. Activation of caspases 3, 7, 8 and 9 was observed in HSG cells after cimetidine treatment, thus confirming that the apoptosis was induced by the activated caspases. Apaf-1 activity was also detected in HSG cells in a dose-dependent manner after treatment with cimetidine. We also found that the cimetidine-mediated down-regulation of NCAM expression in HSG cells did not occur via blocking of the histamine receptor, even though H2R expression was observed on HSG cells, as two other H2R antagonists, famotidine and ranitidine, did not show similar effects. We demonstrated for the first time that cimetidine can induce significant apoptosis of salivary gland tumor cells, which express NCAM, at least in part by down-regulation of NCAM expression on the cells. These findings suggest that the growth, development and perineural/neural invasion of salivary gland tumor cells can be blocked by cimetidine administration through down-regulation of NCAM expression, as well as induction of apoptosis.

Expression of CYLD, NF-kappaB and NF-kappaB-related factors in salivary gland tumors.

The cylindromatosis (CYLD) gene was originally identified as a tumor suppressor that is mutated in familial cylindromatosis, an autosomal dominant condition that confers a predisposition to multiple tumors of the skin appendages. CYLD has deubiquitinating enzyme activity and inhibits the activation of transcription factor NF-kappaB. Therefore, loss of CYLD function correlates with tumorigenesis. Expression of CYLD has been detected in various organs, but its expression in salivary gland tumor (SGT) is still unknown. Adenoid cystic carcinoma (ACC) is a well known and typical malignant SGT ACC was previously known as cylindroma in view of its marked histological resemblance to dermal cylindroma. In this study, the expressions of CYLD and NF-kappaB mRNA in HSG, a human SGT cell line, were found to be increased by TNF-alpha stimulation. Immunohistochemistry clearly demonstrated the expression of CYLD and NF-kappaB-related factors in ACC tissue.

Induction of apoptosis in human salivary gland tumor cells by anti-NCAM antibody.

Neural cell adhesion molecule (NCAM) is a type of cell surface glycoprotein and a member of the immunoglobulin superfamily. It has been reported that NCAM may be associated with perineural invasion by malignant salivary gland tumors such as adenoid cystic carcinoma. We have previously demonstrated that NCAM is constitutively expressed in the human salivary gland tumor cell line HSG, in vitro. In the present study, we have aimed to clarify the hypothesis that NCAM-mediated inhibition of salivary gland tumor proliferation is caused by homophilic binding and involves the prevention of signal transduction for perineural invasion using HSG cells. NCAM mRNA and protein expression was found to decrease in a dose-dependent manner upon treatment with the anti-NCAM antibody (MAb NCAM) for 24 h. The MTT assay showed a significant reduction in the number of viable HSG cells. Confocal laser microscopy showed that HSG cells underwent apoptosis after treatment with MAb NCAM. The activation of caspases 3, 7 and 9 was observed in HSG cells after treatment with MAb NCAM, thus confirming that apoptosis was induced by the activated caspases. Apaf-1 activity was also detected in HSG cells in a dose-dependent manner after treatment with MAb NCAM. The up-regulation of TGF-beta1-mediated NCAM expression appeared to lead to the activation of homophilic NCAM binding, further accelerating HSG cell proliferation. In addition, the localization of NCAM in adenoid cystic carcinomas (ACCs) was examined using an immunohistochemical method. NCAM was slightly to moderately positive in 9 of 13 cases (69.2%) of ACC. These findings suggest that NCAM is associated not only with a cell-to-cell adhesion mechanism, but also with tumorigenesis, including growth, development and perineural invasion in human salivary gland tumors.

Expression of RCAS1 and its function in human squamous cell carcinoma of the oral cavity.

Receptor-binding cancer antigen expressed on SiSo cells (RCAS1) is a recently discovered human tumor-associated antigen expressed in a wide variety of cancer tissues. It has been suggested that tumor cells evade from immune surveillance by expression of RCAS1. We investigated whether RCAS1 is expressed in human oral squamous cell carcinomas (HOSCCs) or HOSCC (HSC-2, -3, -4, Ca9-22 and KB) cells and whether tumor cells expressing RCAS1, induce apoptosis in its receptor-positive cells, peripheral blood lymphocytes (PBLs). RCAS1 transcripts and proteins were detected in five HOSCC cell types. Immunohistochemical examinations revealed that RCAS1 was slightly to moderately positive in 32 of 40 cases (80%) of HOSCCs. Furthermore, the frequency of RCAS1 expression in HOSCC increased with advancing tumor stage according to the pTNM system. Confocal laser microscopy and DNA fragmentation assay showed that PBLs, which were stimulated with IL-2, underwent apoptosis by co-culturing with KB cells. It was also confirmed by Western dot blotting that soluble RCAS1 is secreted into culture supernatants of KB cells. The apoptotic tumor-infiltrating lymphocytes (TILs) were detected around RCAS1-positive tumor cells in HOSCCs by TUNEL method. These results suggest that RCAS1 expression might be associated with progression of oral tumors and offer a possible mechanism for oral cancer immune escape.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL/APO2L) and its receptors expression in human squamous cell carcinoma of the oral cavity.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL/APO2L) is a novel member of the tumor necrosis factor cytokine family and a potent inducer of apoptosis in tumor cells. TRAIL is expressed in most normal human cells and tissues, including peripheral blood leukocytes, spleen, lung and prostate. However, TRAIL expression in human neoplasms is largely unknown. In this study, we investigated whether TRAIL and its receptors are expressed in human oral squamous cell carcinomas (HOSCCs) or HOSCC cell lines (HSC-2, HSC-3, HSC-4, Ca9-22 and KB) and whether these cells are sensitive to TRAIL-induced apoptosis. TRAIL and its receptor transcripts and proteins were detected in all HOSCC cell lines. A cell viability (MTT) assay performed after exposure to recombinant human (rh) TRAIL for 16 h showed that KB cells were the most sensitive. Immunohistochemical examinations for TRAIL expression were also carried out in a total of 50 HOSCC cases with various types of differentiation. In 17 of 27 WHO Grade 1 SCCs, the tumor invasive areas with keratinization showed diffuse and moderate TRAIL expression. In 6 of 20 WHO Grade 2 SCCs, TRAIL expression was slightly to moderately detected. In 1 of 3 WHO Grade 3 SCCs, a slight TRAIL expression was observed. Thus, there is considerable heterogeneity of TRAIL expression and susceptibility to TRAIL-induced apoptosis among human oral tumors.

Association of I27L polymorphism of hepatocyte nuclear factor-1 alpha gene with high-density lipoprotein cholesterol level.

The serum level of high-density lipoprotein cholesterol (HDL-c), which protects against the development of atherosclerosis, is under genetic control. However, the genetic components responsible for the serum HDL-c level are yet to be determined. A recent knockout mouse study demonstrated that hepatocyte nuclear factor-1 alpha (HNF-1 alpha) is an essential transcriptional regulator of HDL-c metabolism. In this study, the association of an HNF-1 alpha gene polymorphism, isoleucine (Ile) 27 leucine (Leu), with lipid parameters, in particular with serum HDL-c level, was studied in 356 unrelated Japanese men. Though no significant difference was observed in total cholesterol and triglyceride levels among the three genotypes, the serum HDL-c level was significantly associated with the genotype (P < 0.01, trend test). Subjects with the Ile/Ile genotype had low serum HDL-c levels, and those with the Leu/Leu genotype had high serum HDL-c levels. These results demonstrate that the HNF-1 alpha gene locus is associated with serum HDL-c level and suggest that the Ile27 allele is a risk marker for atherosclerosis.

Changes in intracellular concentrations of polyamines during apoptosis of HL-60 cells.

Possible changes in the intracellular concentrations of polyamines were investigated during the apoptosis of human promyelocytic leukemic HL-60 cells. Treatment of HL-60 cells with gallic acid and epigallocatechin gallate (EGCG) resulted in the rapid decline of the intracellular concentration of putrescine, whereas that of spermidine and spermine was not significantly changed during the first 3 hours after treatments. Irradiation with UVB also selectively reduced the intracellular concentration of putrescine. On the other hand, cytotoxic concentrations of anticancer agents, such as etoposide and doxorubicin, only marginally reduced the intracellular concentration of putrescine during the first 3 hours. A significant decline of putrescine was observed at later stages when DNA fragmentation became more prominent. Three normal human cells (gingival fibroblast, pulp cell, periodontal ligament fibroblast) and human tumor cell lines (squamous cell carcinoma, submandibular carcinoma, malignant malanoma, hepatoma), which showed higher resistance to apoptosis inducers, had significantly higher putrescine concentrations than HL-60 cells. These data suggest that the intracellular concentration of putrescine may be a useful marker for the apoptosis induction or the sensitivity of the cells to apoptosis inducers.