The effects of all-trans-retinoic acid on cell cycle and alkaline phosphatase activity in pancreatic cancer cells.

Pancreatic cancer is one of the tumors with the highest mortality, poorly responding to available chemotherapeutic agents. The objective of this study was to study the anticancer effects of all-trans retinoid acid, a functional form of vitamin A, on pancreatic cancer cells. Human pancreatic cancer MiaPaCa-2 cells were treated with 1, 5, 10, 20, 30, 40 and 50 microM ATRA for 1, 2, 3, 4, 5 or 6 d, respectively. Cell growth was determined by MTT viability assay. The cell cycle distribution and the alkaline phosphatase (ALP) activity were analyzed by flow cytometry and chemical analyzer, respectively. The results show that ATRA significantly inhibited the growth of MiaPaCa-2 cells at 40 and 50 microM. ATRA arrested pancreatic cancer cells at G0/G1 phase. The sub-G1 peak and DNA fragmentation were observed. There were time and dose dependent increases in alkaline phosphatase activity (ALP), an indicator of cell differentiation, upon treatment with ATRA when compared to controls. In conclusion, ATRA has an inhibitory effect on the cell growth of MiaPaCa-2, and its tumor suppressive effect is by means of cell cycle arrest and apoptosis induction.

Inhibitory effects of dietary curcumin on forestomach, duodenal, and colon carcinogenesis in mice.

Curcumin (diferuloylmethane), a yellow pigment that is obtained from the rhizomes of Curcuma longa Linn., is a major component of turmeric and is commonly used as a spice and food-coloring agent. The inhibitory effects of feeding commercial grade curcumin (77% curcumin, 17% demethoxycurcumin, and 3% bisdemethoxycurcumin) in AIN 76A diet on carcinogen-induced tumorigenesis in the forestomach, duodenum, and colon of mice were evaluated. Administration p.o. of commercial grade curcumin in the diet inhibited benzo(a)pyrene-induced forestomach tumorigenesis in A/J mice, N-ethyl-N'-nitro-N-nitrosoguanidine-induced duodenal tumorigenesis in C57BL/6 mice, and azoxymethane (AOM)-induced colon tumorigenesis in CF-1 mice. Dietary commercial grade curcumin was given to mice at: (a) 2 weeks before, during, and for 1 week after carcinogen administration (during the initiation period); (b) 1 week after carcinogen treatment until the end of the experiment (during the postinitiation period); or (c) during both the initiation and postinitiation periods. Feeding 0.5-2.0% commercial grade curcumin in the diet decreased the number of benzo(a)pyrene-induced forestomach tumors per mouse by 51-53% when administered during the initiation period and 47-67% when administered during the postinitiation period. Feeding 0.5-2.0% commercial grade curcumin in the diet decreased the number of N-ethyl-N'-nitro-N-nitrosoguanidine-induced duodenal tumors per mouse by 47-77% when administered during the postinitiation period. Administration of 0.5-4.0% commercial grade curcumin in the diet both during the initiation and postinitation periods decreased the number of AOM-induced colon tumors per mouse by 51-62%. Administration of 2% commercial grade curcumin in the diet inhibited the number of AOM-induced colon tumors per mouse by 66% when fed during the initiation period and 25% when fed during the postinitiation period. The ability of commercial grade curcumin to inhibit AOM-induced colon tumorigenesis is comparable to that of pure curcumin (purity greater than 98%). Administration of pure or commercial grade curcumin in the diet to AOM-treated mice resulted in development of colon tumors which were generally smaller in number and size as compared to the control group of AOM-treated mice. These results indicate that not only did curcumin inhibit the number of tumors per mouse and the percentage of mice with tumors but it also reduced tumor size. Histopathological examination of the tumors showed that dietary curcumin inhibited the number of papillomas and squamous cell carcinomas of the forestomach as well as the number of adenomas and adenocarcinomas of the duodenum and colon.

Time course for early adaptive responses to ultraviolet B light in the epidermis of SKH-1 mice.

Hairless SKH-1 mice were exposed once to UVB light (180 mJ/cm2), and mechanistically important early adaptive responses in the epidermis were evaluated by immunohistochemical and morphological methods. Interrelationships in the time course for these UVB-induced responses were examined. The number of epidermal cells with DNA strand breaks (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive cells) or with thymine dimers increased to maximal levels within 30 min after UVB. The number of cells with DNA strand breaks located specifically in the basal layer of the epidermis was increased substantially by 3-30 min after UVB and gradually increased further over the next 5.5 hours. DNA strand breaks specifically in the basal layer of the epidermis were increased maximally at 6 h after UVB. The number of epidermal cells with DNA strand breaks or thymine dimers decreased markedly between 12 and 36 h. Pyrimidine (6-4) pyrimidone photodimers (6-4 photoproducts) in isolated epidermal DNA were increased immediately after irradiation of the mice with UVB and decreased markedly during the next 6 h. Exposure to UVB caused a rapid 8-fold increase in the number of epidermal cells with the DNA mismatch repair protein, MSH2 (within 30-60 min), and the level of MSH2-positive cells remained elevated for at least 48 h. These observations suggest a possible role of MSH2 in the repair of UVB-induced DNA damage. The number of epidermal cells with wild-type p53 protein started to increase at 1 h after UVB exposure and reached maximal levels by 8-12 h. The number of p53-positive cells fell markedly between 24 and 48 h. The time course for UVB-induced increases in the number of p53-positive cells was paralleled very closely by the time course for UVB-induced increases in the number of cells with p21(WAF1/CIP1), increases in morphologically distinct apoptotic sunburn cells, and decreases in the number of epidermal cells with bromodeoxyuridine (BrdUrd) incorporation into DNA. Although the start of UVB-induced increases in the number of p21(WAF1/CIP1)-positive cells was similar to that for the increase in p53-positive cells and very high levels of p21(WAF1/CIP1)-positive cells were observed at 8-12 h, maximal increases in p21(WAF1/CIP1)-positive cells were not achieved until 24 h after UVB irradiation (approximately 12 h after the peak value for p53). Myeloperoxidase-positive epidermal cells started to increase by 30 min after UVB exposure, and maximal numbers of myeloperoxidase-positive epidermal cells were observed at 2 h after UVB (18-fold higher than in nonirradiated control mice). An increased level of epidermal peroxidase enzyme activity in the epidermis was also observed from 1 to 24 h after exposure of the mice to UVB. Although neutrophil infiltration into the epidermis was not seen after exposure to UVB, neutrophil infiltration into the dermis (inflammatory response) was observed from 4 to 144 h after UVB exposure. In contrast to the marked inhibitory effect of UVB on BrdUrd incorporation into the DNA of epidermal cells observed at 8-12 h after UVB irradiation (>90% inhibition), BrdUrd incorporation into the DNA of epidermal cells was markedly increased (approximately 30-fold increase in the number of BrdUrd-positive cells) at 48 h after UVB exposure, and increases in epidermal cell layers and epidermal thickness (hyperplasia) were also observed. These later effects were associated with regeneration of the damaged epidermis.

Stimulatory effect of oral administration of green tea or caffeine on ultraviolet light-induced increases in epidermal wild-type p53, p21(WAF1/CIP1), and apoptotic sunburn cells in SKH-1 mice.

Pretreatment of SKH-1 mice with p.o.-administered 0.6% green tea (6 mg of lyophilized tea solids/ml) or 0.044% caffeine (0.44 mg/ml; concentration present in 0.6% green tea) for 2 weeks enhanced UV-induced increases in the number of p53-positive cells, p21(WAF1/CIP1)-positive cells, and apoptotic sunburn cells in the epidermis. These effects of p.o.-administered green tea or caffeine on early adaptive responses to UV provide the first demonstration of in vivo up-regulation of a tumor suppressor gene by a chemopreventive agent. The stimulatory effect of green tea and caffeine on UV-induced increases in the number of p53-positive cells, p21(WAF1/CIP1)-positive cells, and apoptotic sunburn cells may play a role in the inhibitory effects of tea and caffeine on UV-induced carcinogenesis.

Inhibitory effects of black tea, green tea, decaffeinated black tea, and decaffeinated green tea on ultraviolet B light-induced skin carcinogenesis in 7,12-dimethylbenz[a]anthracene-initiated SKH-1 mice.

In a previous study (Z. Y. Wang et al., Cancer Res., 52: 1162-1170, 1992), we found that administration of a water extract of green tea leaves as the sole source of drinking fluid inhibited ultraviolet B light (UVB)-induced carcinogenesis in SKH-1 mice previously initiated with 7,12-dimethylbenz[a]anthracene (DMBA). In the present study, we compared the effects of black tea, green tea, decaffeinated black tea, and decaffeinated green tea on UVB-induced skin carcinogenesis in DMBA-initiated SKH-1 mice. A 1.25% water extract of each kind of tea leaf (1.25 g tea leaf/100 ml water) was prepared by passing 4 liters of hot water through 50 g of tea leaves in a Bunn tea brewing machine. The mean concentrations of solids in multiple samples of 1.25% black tea, green tea, decaffeinated black tea, and decaffeinated green tea analyzed during the course of this study were 4.23, 3.94, 3.66, and 3.53 mg/ml, respectively. These concentrations of tea solids are similar to those present in tea brews ingested by humans. Female SKH-1 mice were treated topically with 200 nmol of DMBA, followed 3 weeks later by irradiation with 30 mJ/cm2 of UVB twice weekly for 31 weeks. UVB-induced formation of skin tumors was markedly inhibited by oral administration of 0.63 or 1.25% black tea, green tea, decaffeinated black tea, or decaffeinated green tea as the sole source of drinking fluid 2 weeks prior to and during 31 weeks of UVB treatment. Administration of each of the eight tea preparations not only inhibited the number of tumors, but tumor size was also markedly decreased. Histopathological examination of each tumor showed that oral administration of the eight tea preparations had a marked inhibitory effect on the formation of UVB-induced keratoacanthomas and carcinomas. Administration of 1.25% black tea, green tea, decaffeinated black tea, or decaffeinated green tea inhibited the number of keratoacanthomas per mouse by 79, 78, 73, or 70%, respectively, and the number of carcinomas per mouse was inhibited by 93, 88, 77, or 72%, respectively. In summary, administration of black tea was comparable to green tea as an inhibitor of UVB-induced skin carcinogenesis in DMBA-initiated SKH-1 mice. Oral administration of decaffeinated black tea or decaffeinated green tea also had a marked inhibitory effect on UVB-induced skin carcinogenesis in DMBA-initiated SKH-1 mice, but these tea preparations were slightly less effective than the regular teas at the high dose level.

Inhibitory effect of green tea in the drinking water on tumorigenesis by ultraviolet light and 12-O-tetradecanoylphorbol-13-acetate in the skin of SKH-1 mice.

Green tea was prepared by extracting 12.5 g of green tea leaves twice with 500 ml of boiling water, and the extracts were combined. This 1.25% green tea extract (1.25 g of tea leaves/100 ml of water) contained 4.69 mg of green tea extract solids per ml and was similar in composition to some green tea beverages consumed by humans. A 2.5% green tea extract (2.5 g of tea leaves/100 ml of water) was prepared similarly. Treatment of female SKH-1 mice with 180 mJ/cm2 of ultraviolet B light (UVB) once daily for 7 days resulted in red sunburn lesions of the skin. The intensity of red color and area of these lesions were inhibited in a dose-dependent fashion by the administration of 1.25 or 2.5% green tea extract as the sole source of drinking water before and during UVB treatment. Treatment of female SKH-1 mice with 180 mJ/cm2 of UVB once daily for 10 days followed 1 wk later by twice weekly application of 12-O-tetradecanoylphorbol-13-acetate for 25 wk resulted in the development of skin tumors. The formation of skin tumors was inhibited by administration of 1.25% green tea extract as the sole source of drinking water prior to and during the 10 days of UVB treatment and for 1 wk after UVB treatment. In additional experiments, female SKH-1 mice were treated with 200 nmol of 7,12-dimethylbenz(a)anthracene followed 3 wk later by irradiation with 180, 60, or 30 mJ/cm2 of UVB twice weekly for 30 wk. UVB-induced formation of skin tumors and increased spleen size were inhibited by administration of 1.25% green tea extract as the sole source of drinking water prior to and during the 30 wk of UVB treatment. In these experiments, treatment of the animals with the green tea extract not only decreased the number of skin tumors but also decreased substantially the size of the tumors. In additional studies, SKH-1 mice were initiated by topical application of 200 nmol of 7,12-dimethylbenz(a)anthracene followed by twice weekly application of 12-O-tetradecanoylphorbol-13-acetate for 25 wk. Administration of 1.25% green tea extract as the sole source of drinking water during promotion with 12-O-tetradecanoylphorbol-13-acetate reduced the number and incidence of skin tumors.

Enhanced skin carcinogenesis in transgenic mice with high expression of glutathione peroxidase or both glutathione peroxidase and superoxide dismutase.

Female transgenic mice (C57BL/6 x CBA/J)F1 with a 1-fold increase in expression of glutathione peroxidase (GP) or with a 1-fold increase in the expression of GP and a 3-4-fold increase in the expression of superoxide dismutase (SOD) had an enhanced carcinogenic response to initiation by 7,12-dimethylbenz[a]anthracene (DMBA) followed by promotion with 12-O-tetradecanoylphorbol-13-acetate (TPA). GP- or GP+SOD-transgenic mice that were initiated by a single topical application of 200 nmol of DMBA followed by promotion with 8 nmol of TPA twice weekly for 30 weeks developed an average of 10.9 or 11.0 skin tumors per mouse and a 100% tumor incidence in comparison with the corresponding nontransgenic mice, which had 3.9 tumors per mouse and an 83% tumor incidence. After stopping TPA application, partial skin tumor regression occurred more rapidly in nontransgenic mice than in either type of transgenic mouse. At 10 weeks after termination of TPA treatment, 9-11% of the tumor-bearing transgenic mice and 26% of the tumor-bearing nontransgenic mice had complete regression of their tumors. Histopathological examination of 96 skin papillomas revealed that the area, location, degree of tumor dysplasia, bromodeoxyuridine labeling index, and p53 protein levels were closely intercorrelated. Further analysis indicated that papillomas with the same grade of dysplasia had a higher bromodeoxyuridine labeling index and a greater p53 protein level in GP- or GP+SOD-transgenic mice than those in nontransgenic mice. The data indicated that overexpression of skin antioxidant enzymes GP or GP+SOD, which are enzymes that are believed to protect cells from oxidative damage by scavenging reactive oxygen species, lead to the increased, rather than the decreased, tumorigenesis in a DMBA/TPA two-stage skin carcinogenesis model.

Effects of tea, decaffeinated tea, and caffeine on UVB light-induced complete carcinogenesis in SKH-1 mice: demonstration of caffeine as a biologically important constituent of tea.

Oral administration of green or black tea inhibited UVB light-induced complete carcinogenesis in the skin of SKH-1 mice. Green tea was a more effective inhibitor than black tea. Oral administration of decaffeinated green or black tea resulted in substantially less inhibitory activity than did administration of the regular teas, and in one experiment, administration of a high-dose level of the decaffeinated teas enhanced the tumorigenic effect of UVB. Oral administration of caffeine alone had a substantial inhibitory effect on UVB-induced carcinogenesis, and adding caffeine to the decaffeinated teas restored the inhibitory effects of these teas on UVB-induced carcinogenesis. In additional studies, topical application of a green tea polyphenol fraction after each UVB application inhibited UVB-induced tumorigenesis. The results indicate that caffeine contributes in an important way to the inhibitory effects of green and black tea on UVB-induced complete carcinogenesis.

Inhibitory effects of orally administered green tea, black tea, and caffeine on skin carcinogenesis in mice previously treated with ultraviolet B light (high-risk mice): relationship to decreased tissue fat.

Treatment of SKH-1 hairless mice with ultraviolet B light (UVB; 30 mJ/cm(2)) twice a week for 22 weeks resulted in tumor-free animals with a high risk of developing malignant and nonmalignant skin tumors during the next several months in the absence of additional UVB treatment (high-risk mice). Oral administration of green tea or black tea (6 mg tea solids/ml) to UVB-pretreated high-risk SKH-1 mice for 23 weeks after stopping UVB treatment decreased the number of tumors/mouse, decreased the size of the parametrial fat pads, and decreased the thickness of the dermal fat layer away from tumors and directly under tumors. Administration of the decaffeinated teas had little or no effect on these parameters, and adding caffeine (equivalent to the amount in the regular teas) to the decaffeinated teas restored their inhibitory effects. Administration of caffeine alone also decreased the number of tumors/mouse, the size of the parametrial fat pads, and the thickness of the dermal fat layer away from tumors and under tumors. Using data from individual mice and linear regression and correlation analysis, we found a highly significant positive correlation between the thickness of the dermal fat layer away from tumors and the number of tumors/mouse (r = 0.34; P = 0.0001), but the correlation between average tumor size/mouse and the thickness of the dermal fat layer away from tumors was weak (r = 0.16; P = 0.034). The results suggested that p.o. administered tea or caffeine may have decreased tumor multiplicity in part by decreasing fat levels in the dermis. Additional analysis revealed that oral administration of caffeinated beverages (green tea, black tea, decaffeinated green tea plus caffeine, decaffeinated black tea plus caffeine, or caffeine alone) decreased the thickness of the dermal fat layer under large tumors to a much greater extent than under small tumors. This is the first demonstration of a close association between inhibition of carcinogenesis and the lowering of tissue fat levels by a chemopreventive agent.

Inhibition of skin tumorigenesis by rosemary and its constituents carnosol and ursolic acid.

A methanol extract of the leaves of the plant Rosmarinus officinalis L. (rosemary) was evaluated for its effects on tumor initiation and promotion in mouse skin. Application of rosemary to mouse skin inhibited the covalent binding of benzo(a)pyrene [B(a)P] to epidermal DNA and inhibited tumor initiation by B(a)P and 7,12-dimethylbenz[a]anthracene (DMBA). Topical application of 20 nmol B(a)P to the backs of mice once weekly for 10 weeks, followed 1 week later by promotion with 15 nmol 12-O-tetradecanoylphorbol-13-acetate (TPA) twice weekly for 21 weeks, resulted in the formation of 7.1 tumors per mouse. In a parallel group of animals that were treated topically with 1.2 or 3.6 mg of rosemary 5 min prior to each application of B(a)P, the number of tumors per mouse was decreased by 54 or 64%, respectively. Application of rosemary to mouse skin also inhibited TPA-induced ornithine decarboxylase activity, TPA-induced inflammation, arachidonic acid-induced inflammation, TPA-induced hyperplasia, and TPA-induced tumor promotion. Mice initiated with 200 nmol DMBA and promoted with 5 nmol TPA twice weekly for 19 weeks developed an average of 17.2 skin tumors per mouse. Treatment of the DMBA-initiated mice with 0.4, 1.2, or 3.6 mg of rosemary together with 5 nmol TPA twice weekly for 19 weeks inhibited the number of TPA-induced skin tumors per mouse by 40, 68, or 99%, respectively. Topical application of carnosol or ursolic acid isolated from rosemary inhibited TPA-induced ear inflammation, ornithine decarboxylase activity, and tumor promotion. Topical application of 1, 3, or 10 mumol carnosol together with 5 nmol TPA twice weekly for 20 weeks to the backs of mice previously initiated with DMBA inhibited the number of skin tumors per mouse by 38, 63, or 78%, respectively. Topical application of 0.1, 0.3, 1, or 2 mumol ursolic acid together with 5 nmol TPA twice weekly for 20 weeks to DMBA-initiated mice inhibited the number of tumors per mouse by 45-61%.

Calcidiol and prostate cancer.

Epidemiological studies suggest that serum calcidiol (25(OH)-Vitamin D3) seems to be associated with several cancers including prostate cancer. We have made several experimental studies in order to clarify the mechanism(s) involved in the association. Calcidiol has been regarded as an inactive prohormone for calcitriol, which possesses the highest biological activity of the Vitamin D metabolites, when it is evaluated on the basis of bioactivity/nmol. However, we found recently that at the physiological concentration calcidiol (100-200 nM) is an active hormone, whereas calcitriol (1alpha,25(OH)2-Vitamin D3) (100 pM) is inactive in human primary prostate stromal cells. Calcidiol is able to inhibit cell growth and to induce or inhibit several genes including 1alpha-hydroxylase and 24-hydroxylase genes. This suggests that calcidiol might be an independent endocrine system involved in the control of cell differentiation and proliferation, whereas calcitriol might be mainly involved in the regulation of calcium and phosphorous balance. Several mechanisms may mediate the action of Vitamin D in the prostate. This is a review of some recent studies on the role of (1) Vitamin D metabolism, (2) growth factors and (3) fatty acid metabolism.

Inhibition by dietary dibenzoylmethane of mammary gland proliferation, formation of DMBA-DNA adducts in mammary glands, and mammary tumorigenesis in Sencar mice.

Dibenzoylmethane (DBM) is a minor constituent of licorice and a beta-diketone analogue of curcumin. Feeding 1% DBM in the diet to Sencar mice during both the initiation and the post-initiation periods strongly inhibited 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumor multiplicity and mammary tumor incidence by 97%. In further in vivo studies to elucidate the possible mechanisms of the inhibitory action of DBM, feeding the 1% DBM in the AIN-76A diet to immature Sencar mice for 4-5 weeks decreased the uterine wet weight by 43%, inhibited the proliferation rate of mammary gland epithelial cells by 53%, uterine epithelium by 23%, and uterine stroma by 77%, when mice were killed during the first estrus phase of estrous cycle. In addition, feeding 1% DBM in the diet to Sencar mice at 2 weeks before, during and 1 week after DMBA treatment (intubation of 1 mg DMBA per mouse once a week for 5 weeks) inhibited formation of total DMBA-DNA adducts in mammary glands by 72% using a post-32P-labeling assay. Thus, feeding 1% DBM diet to Sencar mice inhibited formation of DMBA-DNA adducts in mammary glands and lowered the proliferation rate of the mammary gland in vivo. These results may explain the strong inhibitory actions of dietary DBM on mammary carcinogenesis in mice.

The role of Vitamin D3 metabolism in prostate cancer.

Vitamin D deficiency increases risk of prostate cancer. According to our recent results, the key Vitamin D hormone involved in the regulation of cell proliferation in prostate is 25(OH) Vitamin D3. It is mainly acting directly through the Vitamin D receptor (VDR), but partially also through its 1alpha-hydroxylation in the prostate. A deficiency of 25(OH) Vitamin D is common especially during the winter season in the Northern and Southern latitudes due to an insufficient sun exposure, but Vitamin D deficient diet may partially contribute to it. A lack of Vitamin D action may also be due to an altered metabolism or Vitamin D resistance. Vitamin D resistance might be brought up by several mechanisms: Firstly, an increased 24-hydroxylation may increase the inactivation of hormonal Vitamin D metabolites resulting in a Vitamin D resistance. This is obvious in the cancers in which an oncogenic amplification of 24-hydroxykase gene takes place, although an amplification of this gene in prostate cancer has not yet been described. During the aging, the activity of 24-hydroxylase increases, whereas 1alpha-hydroxylation decreases. Furthermore, it is possible that a high serum concentration of 25(OH)D3 could induce 24-hydroxylase expression in prostate. Secondly, Vitamin D receptor gene polymorphism or defects may result in a partial or complete Vitamin D resistance. Thirdly, an overexpression or hyperphosphorylation of retinoblastoma protein may result in an inefficient mitotic control by Vitamin D. Fourthly, endogenous steroids (reviewed by [D.M. Peehl, D. Feldman, Interaction of nuclear receptor ligands with the Vitamin D signaling pathway in prostate cancer, J. Steroid Biochem. Mol. Biol. (2004)]) and phytoestrogens may modulate the expression of Vitamin D metabolizing enzymes. In summary, the local metabolism of hormonal Vitamin D seems to play an important role in the development and progression of prostate cancer.

Vitamin D and prostate cancer.

Our recent epidemiological study (Ahonen et al., Cancer Causes Control 11(2000) (847-852)) suggests that vitamin D deficiency may increase the risk of initiation and progression of prostate cancer. The nested case-control study was based on a 13-year follow-up of about 19000 middle-aged men free of clinically verified prostate cancer. More than one-half of the serum samples had 25OH-vitamin D (25-VD) levels below 50 nmol/l, suggesting VD deficiency. Prostate cancer risk was highest among the group of younger men (40-51 years) with low serum 25-VD, whereas low serum 25-VD appeared not to increase the risk of prostate cancer in older men (>51 years). This suggests that VD has a protective role against prostate cancer only before the andropause, when serum androgen concentrations are higher. The lowest 25-VD concentrations in the younger men were associated with more aggressive prostate cancer. Furthermore, the high 25-VD levels delayed the appearance of clinically verified prostate cancer by 1.8 years. Since these results suggest that vitamin D has a protective role against prostate cancer, we tried to determine whether full spectrum lighting (FSL) during working hours could increase serum 25-VD concentrations. After 1-month exposure, there was no significant increase in the serum 25-VD level, although there was a bias towards slightly increasing values in the test group as opposed to decreasing values in controls. There was no significant change in the skin urocanic acid production. The possibility to use FSL in cancer prevention is discussed. In order to clarify the mechanism of VD action on cell proliferation and differentiation, we performed studies with the rat and human prostates as well prostate cancer cell lines. It is possible that 25-VD may have a direct role in the host anticancer defence activity, but the metabolism of vitamin D in the prostate may also play an important role in its action. We raised antibodies against human 1alpha-hydroxylase and 24-hydroxylase. Our preliminary results suggest that vitamin D is actively metabolised in the prostate. Vitamin D appears to upregulate androgen receptor expression, whereas androgens seem to upregulate vitamin D receptor (VDR). This may at least partially explain the androgen dependence of VD action. VD alone or administered with androgen causes a suppression of epithelial cell proliferation. VD can activate mitogen-activated kinases, erk-1 and erk-2, within minutes and p38 within hours. Also, auto/paracrine regulation might be involved, since keratinocyte growth factor (mRNA and protein) was clearly induced by VD. Based on these studies, a putative model for VD action on cell proliferation and differentiation is presented.

Increased grooming behavior in mice lacking vitamin D receptors.

Vitamin D is a neuroactive secosteroid with several important functions in the nervous system. Many human and animal findings link alterations in the vitamin D system to various neurological and behavioral disorders. Since grooming is an important element of animal behavior, here we studied whether genetic ablation of vitamin D receptors (VDR) in mice may be associated with altered grooming behaviors. Overall, VDR knockout (VDRko) mice presented longer duration and higher frequency of grooming when tested in the actimeter, open field, elevated plus maze, and horizontal rod tests. Increased grooming did not, however, correlate with unaltered general activity level (actimeter test), anxiety-like behaviors (hole board and elevated plus maze tests), and emotional reactivity index (defecation boli). In general, our results confirm the role of vitamin D and VDR in the regulation of behavior, including grooming, and suggest that increased grooming behavioral phenotype may be associated with genetic ablation of VDR in mutant mice.

Optimal serum calcidiol concentration for cancer prevention.

UNLABELLED: It has been demonstrated in several studies that serum calcidiol (25 OH vitamin D(3)) concentration is in a reversed and linear relationship with cancer risk. However, there are also studies showing no such association and some even suggest the opposite. The risk of pancreatic and oesophageal cancer seems to increase, when serum calcidiol concentration increases. A bias in these studies might be that their basic assumption is linear dependence of cancer on serum calcidiol concentration. Some studies suggest a U-shaped association between the disease and the serum calcidiol concentration. Evidence, in the literature, of the relationship between serum calcidiol concentration and disease is reviewed and an optimal level of 40-80 nmol/L (16-32 ng/ml) is suggested. Serum calcidiol seems to be a better predictor of cancer development than calcitriol (1α, 25 (OH)(2) vitamin D(3)). A calcidiol insufficiency, as well as an insufficient solar exposure, is associated with an increased risk of several solid carcinomas. In a recent study, our group demonstrated that calcidiol is an active hormone in CYP24 (24-hydroxylase) deficient cells. In these cells, calcidiol and calcitriol act synergistically, therefore fluctuations of the serum calcidiol concentration may define the hormonal activity and cancer development. CONCLUSION: Serum calcidiol concentration and the risk of many common diseases and aging phenomena seem to show a U-shaped association suggesting a lower and upper limit for healthy serum calcidiol concentration. An imbalance of hormonal calcidiol rather than that of calcitriol is a risk factor in carcinomas and chronic diseases, which might be prevented by an optimal serum calcidiol concentration. Multiple daily dosing of cholecalcipherol or skin patches could best provide an optimal dosing and stable serum concentration. Alternatively, narrow-band UV-B lamps are a possible optimal solution, when given by trained personnel.

Inhibitory effect of black tea on the growth of established skin tumors in mice: effects on tumor size, apoptosis, mitosis and bromodeoxyuridine incorporation into DNA.

Female CD-1 mice were initiated with a single topical application of 7,12-dimethylbenz[a]anthracene and promoted with 12-O-tetradecanoylphorbol-13-acetate. Mice with established papillomas were then treated with black tea or decaffeinated black tea (approximately 4 mg tea solids/ml) as the sole source of drinking fluid for 11-15 weeks. In four separate experiments, oral administration of black tea inhibited the growth of papillomas (increase in tumor volume/mouse) by an average of 35%, 37%, 41% and 48%, respectively. Studies with decaffeinated black tea gave inconsistent results. In one experiment, administration of decaffeinated black tea inhibited papilloma growth (increase in tumor volume/mouse) by 27%, but in two additional experiments papilloma growth was stimulated by 14% and 193%, respectively. In a separate experiment, skin tumors were generated by treating SKH-1 female mice with ultraviolet B light (UVB; 30 mJ/cm2) twice weekly for 22 weeks, after which UVB administration was stopped. Tumors were allowed to develop during the following 13 weeks, and tumor-bearing mice were then treated with black tea (6 mg/ml tea solids) as the drinking fluid for 11 weeks. In this experiment, tumor growth (increase in tumor volume/mouse) was inhibited by 70%. Histological examination revealed that tea-treated mice had a 58% decrease in the number of nonmalignant tumors (primarily keratoacanthomas)/mouse and a 54% decrease in the number of squamous cell carcinomas/mouse. In addition, administration of black tea decreased the volume per tumor by 60% for nonmalignant tumors and by 84% for carcinomas. Mechanistic studies with tumors from these mice revealed that administration of black tea decreased the bromodeoxyuridine labeling index in squamous cell papillomas, keratoacanthomas and squamous cell carcinomas by 56%, 45% and 35%, respectively, and the apoptosis index was increased by 44%, 100% and 95%, respectively. Administration of black tea decreased the mitotic index in keratoacanthomas and squamous cell carcinomas by 42% and 16%, respectively. The results indicate that oral administration of black tea to tumor-bearing mice inhibited proliferation and enhanced apoptosis in nonmalignant and malignant skin tumors.

Effects of oral administration of tea, decaffeinated tea, and caffeine on the formation and growth of tumors in high-risk SKH-1 mice previously treated with ultraviolet B light.

Treatment of SKH-1 mice with ultraviolet B light (UV-B, 30 mJ/cm2) twice a week for 22-23 weeks resulted in tumor-free animals with a high risk of developing malignant and nonmalignant tumors during the next several months in the absence of further UV-B treatment (high-risk mice). In three separate experiments, oral administration of green tea or black tea (4-6 mg tea solids/ml) as the sole source of drinking fluid for 18-23 weeks to these high-risk mice inhibited the formation and decreased the size of nonmalignant squamous cell papillomas and keratoacanthomas as well as the formation and size of malignant squamous cell carcinomas. In one experiment all these inhibitory effects of tea were statistically significant, whereas in the two other experiments many but not all of the inhibitory effects of tea were statistically significant. The decaffeinated teas were inactive or less effective inhibitors of tumor formation than the regular teas, and adding caffeine back to the decaffeinated teas restored biological activity. Oral administration of caffeine alone (0.44 mg/ml) as the sole source of drinking fluid for 18-23 weeks inhibited the formation of nonmalignant and malignant tumors, and this treatment also decreased tumor size in these high-risk mice.

Some perspectives on dietary inhibition of carcinogenesis: studies with curcumin and tea.

Topical application of curcumin inhibits chemically induced carcinogenesis on mouse skin, and oral administration of curcumin inhibits chemically induced oral, forestomach, duodenal, and colon carcinogenesis. Curcumin and other inhibitors of cyclooxygenase and lipoxygenase are thought to inhibit carcinogenesis by preventing the formation of arachidonic acid metabolites. In contrast to our expectation of a tumorigenic effect of arachidonic acid, we found that treatment of 7,12-dimethylbenz[a]anthracene-initiated mouse skin with very high doses of arachidonic acid twice daily, 5 days a week for 26 weeks, failed to result in tumors. We considered the possibility that some of the cancer chemopreventive effects of curcumin may be related to an effect of this compound on cellular differentiation, and we investigated the effect of curcumin on differentiation in the human promyelocytic HL-60 leukemia cell model system. Although curcumin alone had little or no effect on cellular differentiation, when it was combined with all-trans retinoic acid or 1alpha,25-dihydroxyvitamin D3 a synergistic effect was observed. It is possible that many dietary chemicals in fruits, vegetables, and other edible plants can prevent cancer by synergizing with endogenously produced stimulators of differentiation such as all-trans retinoic acid, 1alpha,25-dihydroxyvitamin D3, and butyrate. More research is needed to test this hypothesis. Administration of green or black tea inhibits carcinogenesis in several animal models, and tumor growth is also inhibited. Several examples were presented of chemopreventive agents that inhibit carcinogenesis in one animal model but enhance carcinogenesis in a different animal model. Greater efforts should be made to understand mechanisms of cancer chemoprevention and to determine whether a potential chemopreventive agent is useful in many experimental settings or whether it is useful in only a limited number of experimental settings.

Effect of curcumin on 12-O-tetradecanoylphorbol-13-acetate- and ultraviolet B light-induced expression of c-Jun and c-Fos in JB6 cells and in mouse epidermis.

Expression of c-jun protein (c-Jun) was observed in normally proliferating JB6 cells but not in confluent cells. Reduction of the serum concentration from 5% to 2% in the cell culture medium caused JB6 cells to enter a quiescent non-proliferating state and down-regulated the expression of c-Jun. Treatment of quiescent JB6 cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) (10 ng/ml) for 24 h markedly stimulated the formation of c-Jun and caused morphological changes. Treatment of JB6 cells with TPA for 48 h resulted in transformed foci with mixed cell populations. Although some cells in these foci expressed high levels of c-Jun, many other cells did not. The increased expression of c-Jun and morphological changes observed at 24 h after treatment of JB6 cells with TPA (10 ng/ml) was inhibited by curcumin (10 nmol/ml). Treatment of JB6 cells with 2.5, 5 or 10 nmol curcumin/ml inhibited the formation of TPA-induced anchorage-independent colonies that grow in soft agar by 31%, 43% and 77%, respectively. Although inhibition of cell proliferation was not observed with 2.5 nmol curcumin/ml, higher concentrations did inhibit cell proliferation. Topical application of 5 nmol TPA to the backs of CD-1 mice once a day for 5 days caused epidermal hyperplasia and the levels of c-Jun were increased in the suprabasal layer of the epidermis and in the muscle layer of the dermis. This treatment also increased c-fos protein (c-Fos) expression in the muscle layer, but there was little or no increase in the expression of c-Fos in the basal or suprabasal layer of the epidermis. Topical application of 10 mumol curcumin together with 5 nmol TPA once a day for 5 days strongly inhibited TPA-induced epidermal hyperplasia and c-Jun and c-Fos expression. A single application of 180 mJ/cm2 of ultraviolet B light (UVB) to the backs of SKH-1 mice caused epidermal hyperplasia and expression of c-Fos and c-Jun in the muscle layer of the dermis and of c-Fos in the suprabasal layer of the epidermis. Maximum effects were observed at 6 days after UVB exposure. Application of 10 mumol curcumin to mouse skin twice a day for 5 days immediately after UVB exposure had only a small/variable inhibitory effect on UVB-induced increases in the expression of c-Fos and c-Jun and on epidermal hyperplasia.(ABSTRACT TRUNCATED AT 400 WORDS)