- Email: [email protected]
Protection by black tea and green tea against UVB and UVA+B induced skin cancer in hairless mice
Mutation Research 422 Ž1998. 191–199
Protection by black tea and green tea against UVB and UVA q B induced skin cancer in hairless mice Ian R. Record ) , Ivor E. Dreosti CSIRO DiÕision of Human Nutrition, PO Box 10041, Gouger St., Adelaide, 5000, South Australia, Australia Received 10 March 1997; revised 24 July 1997; accepted 28 July 1997
Abstract The effects of green and black tea consumption on the early indices of UVB and UVAq B skin damage in hairless mice have been studied in the absence of any chemical tumour initiators or promoters. Black tea consumption was associated with a reduction in the number of sunburn cells in the epidermis of mice 24 h after UVAq B irradiation, although there was no effect of green tea. Other indices of early damage such as necrotic cells or mitotic figures were not affected. Neutrophil infiltration as a measure of skin redness was slightly lowered by tea consumption in the UVB group. Consumption of either green or black tea resulted in significantly fewer skin papillomas and tumours induced by UVA q B light, however black tea provided better protection against UVB-induced tumours than green tea. This study confirms earlier reports that tea consumption can reduce the incidence of skin cancer in hairless mice, and indicates that black tea may afford more protection against simulated solar irradiation than green tea. q 1998 Elsevier Science B.V. All rights reserved. Keywords: Black tea; Green tea; Skin cancer
1. Introduction Australia has one of the highest incidences of skin cancer in the world. Estimates have ranged up to 2.5% in men and 1.7% in women w1x. Despite continuing public education programs there are in excess of 150,000 cases of skin cancer each year. A similar situation exist in the USA where skin cancers represent 55% of all human cancers w2x. In recent years there has been increasing interest in the health benefits of tea, particularly green tea, and the possibility that consumption of the beverage could protect against certain forms of cancer. The general topic of ) Corresponding author. Tel.: q61-8-83038836; Fax: q61-88303-8899; E-mail: [email protected]
tea and cancer has been the subject of several recent reviews w3–5x. In epidemiological studies, green tea appears to be beneficial, whereas no consistent relationship between black tea and cancer has been noted. Considering that almost 80% of the tea produced in the world is consumed as black tea Žwith or without milk., it is perhaps surprising that only a few experimental studies have assessed the anticarcinogenic potential of black tea. All true teas such as these are prepared from the leaves of Camellia sinensis. In the preparation of green tea the leaves are dried and rolled relatively rapidly, whereas leaves used to prepare black tea undergo a ‘fermentation’ process during which naturally occurring polyphenol oxidase converts polyphenols to bisflavanols, theaflavins and the poorly categorised thearubigens
0027-5107r98r$ - see front matter q 1998 Elsevier Science B.V. All rights reserved. PII: S 0 0 2 7 - 5 1 0 7 Ž 9 8 . 0 0 1 9 2 - 4
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
192
which impart the characteristic red–brown colour. Approximately 30% of the dry matter in green tea beverage is composed of catechins, principally Žy.epigallocatechin gallate, whereas the major components of black tea are thearubigens w6x. The green tea catechins are powerful antioxidants, a property which has been linked to the apparent health benefits of whole tea, especially against oxidative damage such as that induced by ultra-violet radiation. More recent studies by Serafini et al. w7x have demonstrated that infusions of green and black tea have similar antioxidant capacities in vitro, even with the addition of milk. Plasma antioxidant capacity appeared to increase equally in volunteers consuming green or black tea, although in a similar study Maxwell and Thorpe w8x were unable to detect an increase in plasma antioxidant capacity in their study of volunteers who had consumed black tea. The comparison of regular and decaffeinated green and black teas by Wang et al. w9x appears to be the only reported comparison between these two types of teas. Most of the animal studies on skin cancer carried out by other workers have focused on green tea, or else the polyphenolic fractions such as Žy.epigallocatechin-3-gallate. In addition most studies using the hairless mouse as a skin cancer model have used a tumour initiator such as 7,12 dimethylbenzwaxanthracene or a promoter, such as 12-O-tetra-
decanoylphorbol-13-acetate ŽTPA. in conjunction with UVB irradiation w9–12x. With this background, we have examined the effects of both black and green tea consumption on early indices of damage Žsunburn cell formation and neutrophil infiltration into the dermis. following the exposure of skin to either UVB radiation, or a mixture of UVA and UVB designed to simulate solar radiation; we have also attempted to obtain more information about the extent to which they protect against skin cancer in the absence of known Žchemical. tumour initiators or promoters.
2. Materials and methods 2.1. Animal care and adherence to guidelines Female SKH-1 mice were purchased from the Animal Resource Service of Murdoch University, Perth, West Australia at approximately 4 weeks of age. All were allowed free access to rodent diet formulated according to the American Institute of Nutrition 1989 guidelines w13x and to either water or one of the teas described below. The experimental protocol and all procedures were approved by CSIRO Division of Human Nutrition Animal Experimentation Ethics Committee and followed the Australian
Table 1 Histological assessment of the skin of hairless mice 24 h after a single exposure to either UVB or mixed UVA q B a Control Žwater.
BT 1.25%
BT 2%
GT 1.25%
GT 2%
Sunburn cells UVB UVA q B
10.1 " 0.90 9.13 " 1.09
9.25 " 0.80 6.35 " 0.86 b
7.85 " 1.00 6.82 " 0.98
7.63 " 0.66 8.34 " 1.18
8.67 " 1.03 7.99 " 0.84
Necrotic cells UVB UVA q B
1.87 " 0.29 0.89 " 0.22
2.33 " 0.36 0.71 " 0.07
1.77 " 0.43 1.01 " 0.23
0.96 " 0.17 b 0.83 " 0.17
2.26 " 0.32 0.86 " 0.16
Mitotic figures UVB UVA q B
0.34 " 0.11 0.70 " 0.18
0.12 " 0.04 0.61 " 0.29
0.46 " 0.30 0.60 " 0.16
0.50 " 0.24 0.74 " 0.34
0.33 " 0.06 0.55 " 0.21
Neutrophils UVB UVA q B
63.8 " 9.9 34.5 " 4.8
37.7 " 7.1 55.6 " 8.6 b
37.6 " 7.0 28.9 " 3.6
45.1 " 5.3 35.7 " 9.7
23.8 " 6.7 30.8 " 4.9
a b
Values are means " S.E.M. for 10 animals per group. Significantly different from any other group P - 0.05.
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
193
Code of Practice for the care and use of animals for scientific purposes. 2.2. Preparation of teas Black ŽBT. and green tea ŽGT. were obtained from TJ Lipton, NJ, USA and prepared as a 2.0% or 1.25% infusion on alternate days using a Braun coffee maker. The dry weight of tea solids was measured regularly by drying portions overnight at 608C. The average solids content over the whole study of the 1.25% infusions was 5.06 " 0.17 mgrml for black tea and 5.12 " 0.13 mgrml for the green tea. In the carcinogenesis study animals were accustomed to the teas by increasing the dose over a seven-day period from 0.5 to 0.75 to 1.25%. Sucrose Ž2%. was added to all drinking fluids to improve palatability. 2.3. UV irradiation Simulated solar irradiation was provided by an array of six UVA tubes ŽSylvania F40BL. around a single UVB tube ŽPhilips FL40SE.. For UVB irradiation the UVA tubes were disconnected. Preliminary studies were carried out to determine the length of time needed to induce a mild erythema by 24 h after
Fig. 2. Time course of induction of tumours and papillomas by UVB alone on the skin of mice given water ŽI., black tea Žl. or green tea ŽB. both at 1.25% as the sole drinking fluid.
exposure. In the first set of studies into the early effects of irradiation animals were exposed for 15 min Ži.e., to twice the minimal erythematous dose wMEDx.. In the carcinogenesis phase of the study the skins of the animals progressively thickened and the time of exposure to reach 1 MED was increased from 5 minrday 5 = rweek to a maximum of 15 minrday. The integrated UVB irradiance Ž280–320 nM. was 2.4 = 10y4 Wrcm2 , and the UVA Ž320– 400 nM. was 1.8 = 10y3 Wrcm2 as measured with a model IL 1700 spectroradiometer ŽInternational Light, Newburyport, MA.. 2.4. Skin histology
Fig. 1. Time course of induction of tumours and papillomas by combined UVAqB on the skin of mice given water ŽI., black tea Žl. or green tea ŽB. both at 1.25% as the sole drinking fluid.
Animals were killed by nembutal overdose and sections of skin were excised from the mid-dorsal surface and fixed in 10% formol-saline pH 7.4 overnight, embedded in paraffin blocks, sectioned at 3 mm and stained in haematoxylin-eosin. Sunburn cells, mitotic figures and necrotic cells were counted in 20 high power microscope fields in three separate epidermal sections per mouse. Separate sections were stained for chloroacetate esterase activity w14x and neutrophils in 20 high power microscope fields in the dermis counted in three sections from each animal.
194
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
2.5. Carcinogenesis Tumour appearance has been expressed progressively as incidence Žpercentage of mice bearing at least one papilloma or tumour. and yield Žaverage number of papillomas or tumours per mouse. as well as burden Žarea of skin affected.. 2.6. Statistical analysis Numbers of sunburn cells, mitotic figures, necrotic cells and neutrophils were compared by analysis of variance ŽANOVA. following a log eŽ X q 0.5. transformation to accommodate zero values. In the tumourigenesis studies differences were evaluated by the Wilcoxon rank sum test. In the case of tumour areas the data were transformed to the square root and then Student’ t-test was used.
3. Results 3.1. Histological studies The UVB and UVArB sunburn cell numbers are presented in Table 1. Black tea at 1.25% inhibited the formation of sunburn cells in the UVArB group only. In the UVB group there was a trend towards a dose–response with black tea Ž P s 0.065.. In unirradiated control mice there were 1.80 " 0.42 sunburn cells per 20 fields. With respect to green tea, there were no significant differences in the skins of the UVArB group, but in the UVB group animals given 1.25% green tea the number of necrotic cells was significantly lower than in all other groups except those given 2% black tea. Unirradiated tissue had 0.18 " 0.12 necrotic cells per 20 fields. The number of mitotic figures in 20 microscope fields was not affected by any dietary treatment. Although the black tea 1.25% UVB figure was lower than any other figure, this observation was not statistically different. Unirradiated mice had 2.48 " 0.44 mitotic figures per 20 fields. Neutrophils in Fig. 3. Numbers of papillomas ŽA., tumours ŽB. and papillomas plus tumours ŽC. on the skins of mice exposed to combined UVAqB on the skin of mice given water ŽI., black tea Žl. or green tea ŽB. both at 1.25% as the sole drinking fluid.
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
the dermis of the animals were counted as an estimate of erythema. Unirradiated animals had 22.7 " 4.7 neutrophils per 20 fields. With the exception of an inexplicably high neutrophil count in the 1.25% black tea group irradiated with UVA q B there were no other significant differences, although a trend to reduced counts was evident with both black and green tea groups exposed to UVB. In the course of this study it was observed that the mice given 2% tea gained less than 2 g during the time they consumed tea, compared with 3.2 g in the water-drinking group and 2.5 g in the 1.25% group. It was therefore decided to use 1.25% tea as the standard for the carcinogenesis study. 3.2. Carcinogenesis After 29 weeks consuming tea Ž1.25%. or water there were no significant differences in weight between animals irradiated with UVA q B, however the animals given black tea and exposed to UVB were significantly Ž P - 0.001. lighter than either of the other dietary treatments Žwater 32.2 " 1.4 g; green tea 32.3 " 0.7 g; black tea 29.1 " 0.2 g.. Initial body weights were 20.5 g. Tumour incidence data are presented in Fig. 1 ŽUVA q B. and Fig. 2 ŽUVB.. For the purposes of this analysis papillomas and ulcerative sites were combined. In some cases papillomas either regressed or were bitten off, so were not counted until they had regrown. In Fig. 1 can be seen that the time for 50% of mice consuming water to develop tumours ŽTDT50 . was 20 weeks. Mice consuming black or green tea reached a TDT50 at 25 weeks. In the UVB group ŽFig. 2. the times were 17 weeks for water, 19 weeks for green tea and 21 weeks for black tea. All mice consuming water developed tumours, and in the UVA q B groups 90% of those consuming green tea and 85% of those drinking black tea had tumours. In the UVB groups 100% of the green tea consumers and 95% of the black tea consumers developed tumours at some stage.
Fig. 4. Numbers of papillomas ŽA., tumours ŽB. and papillomas plus tumours ŽC. on the skins of mice exposed to UVB alone on the skin of mice given water ŽI., black tea Žl. or green tea ŽB. both at 1.25% as the sole drinking fluid.
195
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
196
Table 2 Papillomas and tumours in mice at the end of the study a
UVA q B
UVB
Water Black Tea 1.25% Green Tea 1.25% Water Black Tea 1.25% Green Tea 1.25%
Papillomas
Tumours
Papillomasq tumours
1.00 " 0.06 1.10 " 0.06 1.42 " 0.08 1.89 " 0.46 1.45 " 0.33 1.72 " 0.41
7.06 " 0.42 2.20 " 0.12 b 3.05 " 0.17 b 6.53 " 1.54 2.55 " 0.59 c 5.67 " 1.37
8.06 " 0.47 3.30 " 0.17 b 4.47 " 0.25 b 8.42 " 1.99 4.00 " 0.92 c 7.39 " 1.79
a
Values are means " S.E.M. of 19–20 animals per group. Significantly less than water group P - 0.05. c Significantly less than any other group P - 0.05. b
Fig. 3A,B,C shows the progressive increase in the number of papillomas Ža., tumours Žb. and papillomas plus tumours Žc. as a function of total time of UVA q B irradiation Žapproximately 8 minrday. for 29 weeks. Fig. 4A,B,C shows the corresponding data from the UVB group. Several points of interest emerge from these data. In both the UVA q B and UVB groups given water there is a distinct decrease in the number of papillomas in the last few weeks of exposure, just as the number of tumours begins to increase most rapidly. This decrease probably represents the conversion of papillomas to tumours. In the groups consuming either green or black tea the numbers of papillomas reached a plateau, with little suggestion of a decrease. In the UVA q B groups ŽTable 2. black tea and green tea significantly reduced the tumour burden when compared to the water drinking animals Ž P - 0.0001; P - 0.002, respectively.. There were no significant effects on papilloma burden at this time, although both teas reduced the number of papillomas plus tumours Žgreen P - 0.05; black P - 0.0001.. In the UVB
treated animals, black tea was associated with a significant reduction in the number of tumours and tumours plus papillomas when compared to controls Ž P s 0.0002; P - 0.0001, respectively.. Green tea consumption tended to decrease the combined number of tumours plus papillomas when compared to water Ž P s 0.08., however animals consuming black tea had significantly fewer tumours and tumours plus papillomas than the animals consuming green tea Ž P s 0.05; P - 0.002, respectively.. Comparing yields between the two UV groups it would seem that green tea is more effective at reducing the total numbers of tumours and papillomas in mice exposed to UVA q B than UVB alone Ž P s 0.03.. Average tumour areas were also measured ŽTable 3.. In terms of tumour burden it would appear that black and green teas were equally as effective at reducing the tumour area in the UVA q B irradiated mice Ž P - 0.05., but black tea was clearly superior in the UVB exposed mice Ž P - 0.0001..
4. Discussion Table 3 Average areas of tumours on mice a Tumour area Žmm UVAqB
UVB
a
Water Black tea 1.25% Green tea 1.25% Water Black tea 1.25% Green tea 1.25%
2.
34.9"8.6 20.9"8.2 b 20.3"8.5 b 58.0"11.3 13.1"3.3 c 49.0"9.3
Values are means"S.E.M. of 19–20 animals per group. Significantly less than water group P - 0.05. c Significantly less than any other group P - 0.05. b
We in this laboratory and other workers have previously reported that a number of dietary antioxidants can provide protection against UV-associated free radical damage to skin in vitro and in vivo w15–22x. More recently interest has focused on some of the non-nutrient dietary antioxidants, and in particular various teas as protectants against skin cancer. Wang et al. w10x were the first to study the effects of a green tea polyphenolic fraction on UVB-related photocarcinogenesis in mice. At a dose of 6 mgrday they observed a significant reduction in the number
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
of tumours per mouse, as well as a significant lengthening of the TDT50 . Several more recent studies have also described the effects of green tea polyphenols on chemically-induced and chemicallypromoted skin cancer w23–26x. Protection by green tea and black tea infusions against skin cancer in hairless mice has also been reported w11,12,27x. In the first of these studies w11x it was demonstrated that tea consumption significantly inhibited the formation of skin cancers promoted by TPA and also markedly reduced the intensity of UVB-induced skin lesions. The same group later noted that administration of green tea in the drinking water, or intraperitoneal injection of Žy.-epigallocatechin gallate inhibited tumour growth, also caused chemically-induced and chemically-promoted established papillomas to regress either partially, or in a few cases completely w27x. Similar results were obtained when UVB and TPA were used to produce the tumours and papillomas. Wang et al. w9x extended their studies to mice initiated with 7,12-dimethylbenz wa xanthracene ŽDMBA. and subsequently exposed to UVB. In this experiment black tea was found to protect as effectively as green tea, although the decaffeinated teas seemed to be slightly less effective than regular teas at the highest dose used. In the current study we refrained from using chemical initiators or promoters since some of these, TPA in particular, are known to cause free radical damage to cells. We also compared UVB with mixed UVA q B, since the mixed spectrum more closely simulates solar radiation. The early events Žsunburn cell formation, necrotic cells, mitotic figures and neutrophil infiltration. were not dramatically affected by consumption of tea, although there was a trend for most variables to be reduced in the tea-consuming groups. On exposure to UV irradiation the skin responds in a number of ways. Epidermal cells exposed to sufficient damage can undergo apoptosis or ‘programmed cell death’; this is most obviously manifested as ‘sunburn cells’. A reduction in the number of sunburn cells has been used as an indication of a protective effect of sunscreens. Exposure of epidermal cells to antioxidants has been demonstrated to reduce the number of sunburn cells, presumably because some of the free radicals generated by UV can be removed before they cause excessive damage w15x. The number of
197
necrotic cells also tends to be lower in the tea groups. Following irradiation, cell division is usually delayed somewhat. This provides cells more time in which to affect the repair of cellular and genomic damage. This repair time is reflected in the number of mitotic figures present, so that a reduction in the number of figures may well provide a fair representation of the degree of damage. In this study administration of tea had little effect. Neutrophil infiltration, which is due to increased blood flow in the dermis, also tended to be lower in the tea groups Žexcept for the black tea 1.25% which was inexplicably high., suggesting that tea may be able to reduce the erythema Žredness. associated with sun exposure. Other workers have reported beneficial effects of tea and its catechins against early markers of skin damage, including induction of ornithine decarboxylase activity w28x, contact hypersensitivity and inflammation w29x and lipid peroxidation in mouse skin microsomes in vitro w30x. Previous workers have used chemical initiators or promoters to induce skin cancers, and there appears to be only one other study in which the effects of green and black teas have been compared w9x. In this earlier study there seemed to be little difference between the protective effects of green and black teas, although decaffeinated teas did seem to be somewhat less effective. In the present study black tea provided significantly more protection than green tea, reducing the yield of papillomas and tumours and appearing to delay the onset of papilloma development Žespecially in the animals exposed to UVA q B.. Although there was a plateau effect on papilloma formation in tea-consuming animals, there was a decline in the number of papillomas in the control groups, together with an increase in the number of tumours, suggesting that this reduction could be an artefact, almost certainly involving the conversion of papillomas to tumours. The differences between UVA q B and UVB were small, although there were indications that cell necrosis and the number of mitotic figures were less affected by the combined irradiation regime. The number of tumours and total tumour area in the green tea-drinking groups and the area of tumours in the water group were lower in the UVA q B group than the UVB group. Similar effects of an amelioration of skin tumours in mice has been observed
198
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
before w31x, although later studies have not been entirely confirmatory w32x. Clearly this area requires further research, but in a manner which is beyond the scope of this present study. In recognition of the fact that solar radiation is mixed UVA and UVB future studies will only involve UVA q B exposure. In summary, our results suggest that consumption of either green or black tea may have the potential to reduce some of the early effects of UV exposure, albeit to a relatively small degree. Consumption of either green or black tea was shown to be associated with protection against skin carcinogenesis by UVB and UVA q B, thereby confirming reports from other laboratories. In contrast to one other study involving black tea, we found that black tea was, if anything, more beneficial than green tea. It may be that the use of initiation or promotion protocols by earlier workers will prove to have some bearing on this discrepancy. There may be important implications for human health, since most of the world tea production is black, and much of this is consumed with milk. Clearly further studies are required, both to confirm the difference between black and green tea and to ascertain whether addition of milk to the beverage has any effect.
Acknowledgements The authors wish to thank Ms. Mary Jannes and Ms. Linda Ormond for excellent technical assistance, TJ Lipton, Englewood Cliffs, NJ, for the teas, and Unifoods ŽAustralia. for financial support.
References w1x A. Green, D. Battistutta, A. Russel, C. Frost, Skin cancer: the quiet epidemic, Proc. Mt. Buller Conf. Environmental Radiation, 1996, p. 16 ŽAbs.. w2x American Cancer Society, Cancer facts and figures, Am. Cancer Soc., Washington, D.C., 1991. w3x I.E. Dreosti, Bioactive ingredients: antioxidants and polyphenols in tea, Nutr. Rev. 54 Ž1996. s51–s58. w4x R.A. Goldbohm, M.G.L. Hertog, H.A. M Brants, G. van Poppel, P.A. van den Brandt, Consumption of black tea and cancer risk: a prospective cohort study, J. Nat. Cancer Inst. 88 Ž1996. 93–100.
w5x S.K. Katiyar, H. Mukhtar, Tea consumption and cancer, in: A. Simopoulos ŽEd.., World Rev. Nutr. Diet. Karger, Basel, Vol. 79, 1996, pp. 154–184. w6x H.N. Graham, Green tea consumption, consumption and polyphenol chemistry, Prev. Med. 21 Ž1992. 334–350. w7x M. Serafini, A. Ghiselli, A. Ferro-Luzzi, In vivo antioxidant effects of green and black tea in man, Eur. J. Clin. Nutr. 50 Ž1996. 28–32. w8x S. Maxwell, G. Thorpe, Tea flavonoids have little short term impact on serum antioxidant activity, Br. Med. J. 313 Ž1996. 229. w9x Z.Y. Wang, M.T. Huang, Y.R. Lou, J.G. Xie, K.R. Reuhl, H.L. Newmark, C.T. Ho, C.S. Yang, A.H. Conney, Inhibitory effects of black tea, green tea, decaffeinated black tea and decaffeinated green tea on ultraviolet B light-induced skin carcinogenesis in 7,12-dimethylbenzwaxanthraceneinitiated SKH-1 mice, Cancer Res. 54 Ž1994. 3235–3248. w10x Z.Y. Wang, R. Agarwal, D.R. Bickers, H. Mukhtar, Protection against ultraviolet B radiation-induced photocarcinogenesis in hairless mice by green tea polyphenols, Carcinogenesis 12 Ž1991. 1527–1530. w11x Z.Y. Wang, M.T. Huang, T. Ferraro, C.Q. Wong, Y.R. Lou, K. Reuhl, M. Iatropoulos, C.S. Yang, A.H. Conney, Inhibitory effect of green tea in the drinking water on tumorigenesis by ultraviolet light and 12-O-tetradecanoylphorbol13-acetate in the skin of SKH-1 mice, Cancer Res. 52 Ž1992. 1162–1170. w12x A.H. Conney, Z.Y. Wang, M.T. Huang, C.T. Ho, C.S. Yang, Inhibitory effect of green tea on tumorigenesis by chemicals and ultraviolet light, Prev. Med. 21 Ž1992. 361–369. w13x P.G. Reeves, AIN-76 diet: should we change the formulation?, J. Nutr. 119 Ž1989. 1081–1082. w14x L.D. Leder, The chloroacetate esterase reaction, Am. J. Dermatol. 1 Ž1979. 39–42. w15x Y. Miyachi, T. Horio, S. Imamura, Sunburn cell formation is prevented by scavenging oxygen intermediates, Clin. Exp. Derm. 8 Ž1983. 305–310. w16x I.R. Record, I.E. Dreosti, M. Konstantinopoulos, R.A. Buckley, The influence of topical and systemic vitamin E on ultra violet light-induced skin damage in hairless mice, Nutr. Cancer 16 Ž1991. 219–225. w17x I.R. Record, L. Cavallo, I.E. Dreosti, Protection against UVA and UVB induced lipid peroxidation in liposomes by different forms of vitamins C and E, in: K. Yagi ŽEd.., International Congress Series No. 998; 5th International Conference on Oxygen Radicals. Elsevier, Amsterdam, 1992, pp. 621– 624. w18x I.R. Record, M. Jannes, I.E. Dreosti, Protection by zinc against UVA- and UVB-induced cellular and genomic damage in vivo and in vitro, Biol. Trace Elem. Res. 53 Ž1996. 19–25. w19x J.-P. Perchellet, N.L. Abney, R.M. Thomas, Y.L. Guislain, E.M. Perchellet, Effects of combined treatments with selenium, glutathione and vitamin E on glutathione peroxidase activity, ornithine decarboxylase induction and complete and multistage carcinogenesis in mouse skin, Cancer Res. 47 Ž1987. 477–485.
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199 w20x K.E. Gerrish, H.L. Gensler, Prevention of photocarcinogenesis by dietary vitamin E, Nutr. Cancer 19 Ž1993. 125–133. w21x L.A. Lambert, W.G. Wamer, R.R. Wei, S. Lavu, S.J. Chirtel, A. Kornhauser, The protective but nonsynergistic effect of dietary b-carotene and vitamin E on skin tumourigenesis in Skh mice, Nutr. Cancer 21 Ž1994. 1–12. w22x M.M. Mathews-Roth, N.I. Krinsky, Carotenoids affect development of UV-B induced skin cancer, Photochemistry and Photobiology 46 Ž1987. 507–509. w23x M.-T. Huang, C.-T. Ho, Z.Y. Wang, T. Ferraro, T. Finnegan-Olive, Y.-R. Lou, J.M. Mitchell, J.D. Laskin, H. Newmark, C.S. Yang, A.H. Conney, Inhibitory effect of topical application of a green tea polyphenol fraction on tumour initiation and promotion in mouse skin, Carcinogenesis 13 Ž1992. 947–954. w24x S.K. Katiyar, R. Agerwal, G.S. Wood, H. Mukhtar, Inhibition of 12-O-tetradecanoylphorbol-13-acetate-caused tumour promotion in 7,12-dimethylbenz wax anthracene-initiated SENCAR mouse skin by a polyphenolic fraction isolated from green tea, Cancer Res. 52 Ž1992. 6890–6897. w25x S.K. Katiyar, R. Agarwal, H. Mukhtar, Inhibition of both stage 1 and stage 11 skin tumour promotion in SENCAR mice by a polyphenolic fraction isolated from green tea: inhibition depends on the duration of polyphenol treatment, Carcinogenesis 14 Ž1993. 2641–2643. w26x S.K. Katiyar, R. Agarwal, H. Mukhtar, Protection against malignant conversion of chemically induced benign skin papillomas to squamous cell carcinomas in SENCAR mice
w27x
w28x
w29x
w30x
w31x
w32x
199
by a polyphenolic fraction isolated from green tea, Cancer Res. 53 Ž1993. 5409–5412. Z.Y. Wang, M.-T. Huang, C.-T. Ho, R. Chang, W. Ma, T. Ferraro, K.R. Reuhl, C.S. Yang, A.H. Conney, Inhibitory effect of green tea on the growth of established skin papillomas in mice, Cancer Res. 52 Ž1992. 6657–6665. R. Agarwal, S.K. Katiyar, S.I.A. Zaidi, H. Mukhtar, Inhibition of skin tumour promoter-caused induction of epidermal ornithine decarboxylase in SENCAR mice by polyphenolic fraction isolated from green tea and its individual epicatechin derivatives, Cancer Res. 52 Ž1992. 3582–3588. S.K. Katiyar, C.A. Elmets, R. Agarwal, H. Mukhtar, Protection against ultraviolet-B radiation-induced local and systemic suppression of contact hypersensitivity and edema responses in C3HrHEN mice by green tea polyphenols, Photochemistry and Photobiology 62 Ž1995. 855–861. S.K. Katiyar, R. Agarwal, H. Mukhtar, Inhibition of spontaneous and photo-enhanced lipid peroxidation in mouse epidermal microsomes by epicatachin derivatives from green tea, Cancer Letters 79 Ž1994. 61–66. H. Van Weelden, J.C. van der Leun, Photorecovery by UVA, in: F. Urbach, R.W. Gange ŽEds.., The Biological Effects of UVA Radiation, Praeger, New York, 1986, pp. 147–152. R.J.W. Berg, F.R. de Gruiji, J.C. van der Leun, Interaction between ultraviolet A and ultraviolet B radiations in skin cancer induction in hairless mice, Cancer Res. 53 Ž1993. 4212–4217.
Protection by black tea and green tea against UVB and UVA q B induced skin cancer in hairless mice Ian R. Record ) , Ivor E. Dreosti CSIRO DiÕision of Human Nutrition, PO Box 10041, Gouger St., Adelaide, 5000, South Australia, Australia Received 10 March 1997; revised 24 July 1997; accepted 28 July 1997
Abstract The effects of green and black tea consumption on the early indices of UVB and UVAq B skin damage in hairless mice have been studied in the absence of any chemical tumour initiators or promoters. Black tea consumption was associated with a reduction in the number of sunburn cells in the epidermis of mice 24 h after UVAq B irradiation, although there was no effect of green tea. Other indices of early damage such as necrotic cells or mitotic figures were not affected. Neutrophil infiltration as a measure of skin redness was slightly lowered by tea consumption in the UVB group. Consumption of either green or black tea resulted in significantly fewer skin papillomas and tumours induced by UVA q B light, however black tea provided better protection against UVB-induced tumours than green tea. This study confirms earlier reports that tea consumption can reduce the incidence of skin cancer in hairless mice, and indicates that black tea may afford more protection against simulated solar irradiation than green tea. q 1998 Elsevier Science B.V. All rights reserved. Keywords: Black tea; Green tea; Skin cancer
1. Introduction Australia has one of the highest incidences of skin cancer in the world. Estimates have ranged up to 2.5% in men and 1.7% in women w1x. Despite continuing public education programs there are in excess of 150,000 cases of skin cancer each year. A similar situation exist in the USA where skin cancers represent 55% of all human cancers w2x. In recent years there has been increasing interest in the health benefits of tea, particularly green tea, and the possibility that consumption of the beverage could protect against certain forms of cancer. The general topic of ) Corresponding author. Tel.: q61-8-83038836; Fax: q61-88303-8899; E-mail: [email protected]
tea and cancer has been the subject of several recent reviews w3–5x. In epidemiological studies, green tea appears to be beneficial, whereas no consistent relationship between black tea and cancer has been noted. Considering that almost 80% of the tea produced in the world is consumed as black tea Žwith or without milk., it is perhaps surprising that only a few experimental studies have assessed the anticarcinogenic potential of black tea. All true teas such as these are prepared from the leaves of Camellia sinensis. In the preparation of green tea the leaves are dried and rolled relatively rapidly, whereas leaves used to prepare black tea undergo a ‘fermentation’ process during which naturally occurring polyphenol oxidase converts polyphenols to bisflavanols, theaflavins and the poorly categorised thearubigens
0027-5107r98r$ - see front matter q 1998 Elsevier Science B.V. All rights reserved. PII: S 0 0 2 7 - 5 1 0 7 Ž 9 8 . 0 0 1 9 2 - 4
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
192
which impart the characteristic red–brown colour. Approximately 30% of the dry matter in green tea beverage is composed of catechins, principally Žy.epigallocatechin gallate, whereas the major components of black tea are thearubigens w6x. The green tea catechins are powerful antioxidants, a property which has been linked to the apparent health benefits of whole tea, especially against oxidative damage such as that induced by ultra-violet radiation. More recent studies by Serafini et al. w7x have demonstrated that infusions of green and black tea have similar antioxidant capacities in vitro, even with the addition of milk. Plasma antioxidant capacity appeared to increase equally in volunteers consuming green or black tea, although in a similar study Maxwell and Thorpe w8x were unable to detect an increase in plasma antioxidant capacity in their study of volunteers who had consumed black tea. The comparison of regular and decaffeinated green and black teas by Wang et al. w9x appears to be the only reported comparison between these two types of teas. Most of the animal studies on skin cancer carried out by other workers have focused on green tea, or else the polyphenolic fractions such as Žy.epigallocatechin-3-gallate. In addition most studies using the hairless mouse as a skin cancer model have used a tumour initiator such as 7,12 dimethylbenzwaxanthracene or a promoter, such as 12-O-tetra-
decanoylphorbol-13-acetate ŽTPA. in conjunction with UVB irradiation w9–12x. With this background, we have examined the effects of both black and green tea consumption on early indices of damage Žsunburn cell formation and neutrophil infiltration into the dermis. following the exposure of skin to either UVB radiation, or a mixture of UVA and UVB designed to simulate solar radiation; we have also attempted to obtain more information about the extent to which they protect against skin cancer in the absence of known Žchemical. tumour initiators or promoters.
2. Materials and methods 2.1. Animal care and adherence to guidelines Female SKH-1 mice were purchased from the Animal Resource Service of Murdoch University, Perth, West Australia at approximately 4 weeks of age. All were allowed free access to rodent diet formulated according to the American Institute of Nutrition 1989 guidelines w13x and to either water or one of the teas described below. The experimental protocol and all procedures were approved by CSIRO Division of Human Nutrition Animal Experimentation Ethics Committee and followed the Australian
Table 1 Histological assessment of the skin of hairless mice 24 h after a single exposure to either UVB or mixed UVA q B a Control Žwater.
BT 1.25%
BT 2%
GT 1.25%
GT 2%
Sunburn cells UVB UVA q B
10.1 " 0.90 9.13 " 1.09
9.25 " 0.80 6.35 " 0.86 b
7.85 " 1.00 6.82 " 0.98
7.63 " 0.66 8.34 " 1.18
8.67 " 1.03 7.99 " 0.84
Necrotic cells UVB UVA q B
1.87 " 0.29 0.89 " 0.22
2.33 " 0.36 0.71 " 0.07
1.77 " 0.43 1.01 " 0.23
0.96 " 0.17 b 0.83 " 0.17
2.26 " 0.32 0.86 " 0.16
Mitotic figures UVB UVA q B
0.34 " 0.11 0.70 " 0.18
0.12 " 0.04 0.61 " 0.29
0.46 " 0.30 0.60 " 0.16
0.50 " 0.24 0.74 " 0.34
0.33 " 0.06 0.55 " 0.21
Neutrophils UVB UVA q B
63.8 " 9.9 34.5 " 4.8
37.7 " 7.1 55.6 " 8.6 b
37.6 " 7.0 28.9 " 3.6
45.1 " 5.3 35.7 " 9.7
23.8 " 6.7 30.8 " 4.9
a b
Values are means " S.E.M. for 10 animals per group. Significantly different from any other group P - 0.05.
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
193
Code of Practice for the care and use of animals for scientific purposes. 2.2. Preparation of teas Black ŽBT. and green tea ŽGT. were obtained from TJ Lipton, NJ, USA and prepared as a 2.0% or 1.25% infusion on alternate days using a Braun coffee maker. The dry weight of tea solids was measured regularly by drying portions overnight at 608C. The average solids content over the whole study of the 1.25% infusions was 5.06 " 0.17 mgrml for black tea and 5.12 " 0.13 mgrml for the green tea. In the carcinogenesis study animals were accustomed to the teas by increasing the dose over a seven-day period from 0.5 to 0.75 to 1.25%. Sucrose Ž2%. was added to all drinking fluids to improve palatability. 2.3. UV irradiation Simulated solar irradiation was provided by an array of six UVA tubes ŽSylvania F40BL. around a single UVB tube ŽPhilips FL40SE.. For UVB irradiation the UVA tubes were disconnected. Preliminary studies were carried out to determine the length of time needed to induce a mild erythema by 24 h after
Fig. 2. Time course of induction of tumours and papillomas by UVB alone on the skin of mice given water ŽI., black tea Žl. or green tea ŽB. both at 1.25% as the sole drinking fluid.
exposure. In the first set of studies into the early effects of irradiation animals were exposed for 15 min Ži.e., to twice the minimal erythematous dose wMEDx.. In the carcinogenesis phase of the study the skins of the animals progressively thickened and the time of exposure to reach 1 MED was increased from 5 minrday 5 = rweek to a maximum of 15 minrday. The integrated UVB irradiance Ž280–320 nM. was 2.4 = 10y4 Wrcm2 , and the UVA Ž320– 400 nM. was 1.8 = 10y3 Wrcm2 as measured with a model IL 1700 spectroradiometer ŽInternational Light, Newburyport, MA.. 2.4. Skin histology
Fig. 1. Time course of induction of tumours and papillomas by combined UVAqB on the skin of mice given water ŽI., black tea Žl. or green tea ŽB. both at 1.25% as the sole drinking fluid.
Animals were killed by nembutal overdose and sections of skin were excised from the mid-dorsal surface and fixed in 10% formol-saline pH 7.4 overnight, embedded in paraffin blocks, sectioned at 3 mm and stained in haematoxylin-eosin. Sunburn cells, mitotic figures and necrotic cells were counted in 20 high power microscope fields in three separate epidermal sections per mouse. Separate sections were stained for chloroacetate esterase activity w14x and neutrophils in 20 high power microscope fields in the dermis counted in three sections from each animal.
194
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
2.5. Carcinogenesis Tumour appearance has been expressed progressively as incidence Žpercentage of mice bearing at least one papilloma or tumour. and yield Žaverage number of papillomas or tumours per mouse. as well as burden Žarea of skin affected.. 2.6. Statistical analysis Numbers of sunburn cells, mitotic figures, necrotic cells and neutrophils were compared by analysis of variance ŽANOVA. following a log eŽ X q 0.5. transformation to accommodate zero values. In the tumourigenesis studies differences were evaluated by the Wilcoxon rank sum test. In the case of tumour areas the data were transformed to the square root and then Student’ t-test was used.
3. Results 3.1. Histological studies The UVB and UVArB sunburn cell numbers are presented in Table 1. Black tea at 1.25% inhibited the formation of sunburn cells in the UVArB group only. In the UVB group there was a trend towards a dose–response with black tea Ž P s 0.065.. In unirradiated control mice there were 1.80 " 0.42 sunburn cells per 20 fields. With respect to green tea, there were no significant differences in the skins of the UVArB group, but in the UVB group animals given 1.25% green tea the number of necrotic cells was significantly lower than in all other groups except those given 2% black tea. Unirradiated tissue had 0.18 " 0.12 necrotic cells per 20 fields. The number of mitotic figures in 20 microscope fields was not affected by any dietary treatment. Although the black tea 1.25% UVB figure was lower than any other figure, this observation was not statistically different. Unirradiated mice had 2.48 " 0.44 mitotic figures per 20 fields. Neutrophils in Fig. 3. Numbers of papillomas ŽA., tumours ŽB. and papillomas plus tumours ŽC. on the skins of mice exposed to combined UVAqB on the skin of mice given water ŽI., black tea Žl. or green tea ŽB. both at 1.25% as the sole drinking fluid.
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
the dermis of the animals were counted as an estimate of erythema. Unirradiated animals had 22.7 " 4.7 neutrophils per 20 fields. With the exception of an inexplicably high neutrophil count in the 1.25% black tea group irradiated with UVA q B there were no other significant differences, although a trend to reduced counts was evident with both black and green tea groups exposed to UVB. In the course of this study it was observed that the mice given 2% tea gained less than 2 g during the time they consumed tea, compared with 3.2 g in the water-drinking group and 2.5 g in the 1.25% group. It was therefore decided to use 1.25% tea as the standard for the carcinogenesis study. 3.2. Carcinogenesis After 29 weeks consuming tea Ž1.25%. or water there were no significant differences in weight between animals irradiated with UVA q B, however the animals given black tea and exposed to UVB were significantly Ž P - 0.001. lighter than either of the other dietary treatments Žwater 32.2 " 1.4 g; green tea 32.3 " 0.7 g; black tea 29.1 " 0.2 g.. Initial body weights were 20.5 g. Tumour incidence data are presented in Fig. 1 ŽUVA q B. and Fig. 2 ŽUVB.. For the purposes of this analysis papillomas and ulcerative sites were combined. In some cases papillomas either regressed or were bitten off, so were not counted until they had regrown. In Fig. 1 can be seen that the time for 50% of mice consuming water to develop tumours ŽTDT50 . was 20 weeks. Mice consuming black or green tea reached a TDT50 at 25 weeks. In the UVB group ŽFig. 2. the times were 17 weeks for water, 19 weeks for green tea and 21 weeks for black tea. All mice consuming water developed tumours, and in the UVA q B groups 90% of those consuming green tea and 85% of those drinking black tea had tumours. In the UVB groups 100% of the green tea consumers and 95% of the black tea consumers developed tumours at some stage.
Fig. 4. Numbers of papillomas ŽA., tumours ŽB. and papillomas plus tumours ŽC. on the skins of mice exposed to UVB alone on the skin of mice given water ŽI., black tea Žl. or green tea ŽB. both at 1.25% as the sole drinking fluid.
195
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
196
Table 2 Papillomas and tumours in mice at the end of the study a
UVA q B
UVB
Water Black Tea 1.25% Green Tea 1.25% Water Black Tea 1.25% Green Tea 1.25%
Papillomas
Tumours
Papillomasq tumours
1.00 " 0.06 1.10 " 0.06 1.42 " 0.08 1.89 " 0.46 1.45 " 0.33 1.72 " 0.41
7.06 " 0.42 2.20 " 0.12 b 3.05 " 0.17 b 6.53 " 1.54 2.55 " 0.59 c 5.67 " 1.37
8.06 " 0.47 3.30 " 0.17 b 4.47 " 0.25 b 8.42 " 1.99 4.00 " 0.92 c 7.39 " 1.79
a
Values are means " S.E.M. of 19–20 animals per group. Significantly less than water group P - 0.05. c Significantly less than any other group P - 0.05. b
Fig. 3A,B,C shows the progressive increase in the number of papillomas Ža., tumours Žb. and papillomas plus tumours Žc. as a function of total time of UVA q B irradiation Žapproximately 8 minrday. for 29 weeks. Fig. 4A,B,C shows the corresponding data from the UVB group. Several points of interest emerge from these data. In both the UVA q B and UVB groups given water there is a distinct decrease in the number of papillomas in the last few weeks of exposure, just as the number of tumours begins to increase most rapidly. This decrease probably represents the conversion of papillomas to tumours. In the groups consuming either green or black tea the numbers of papillomas reached a plateau, with little suggestion of a decrease. In the UVA q B groups ŽTable 2. black tea and green tea significantly reduced the tumour burden when compared to the water drinking animals Ž P - 0.0001; P - 0.002, respectively.. There were no significant effects on papilloma burden at this time, although both teas reduced the number of papillomas plus tumours Žgreen P - 0.05; black P - 0.0001.. In the UVB
treated animals, black tea was associated with a significant reduction in the number of tumours and tumours plus papillomas when compared to controls Ž P s 0.0002; P - 0.0001, respectively.. Green tea consumption tended to decrease the combined number of tumours plus papillomas when compared to water Ž P s 0.08., however animals consuming black tea had significantly fewer tumours and tumours plus papillomas than the animals consuming green tea Ž P s 0.05; P - 0.002, respectively.. Comparing yields between the two UV groups it would seem that green tea is more effective at reducing the total numbers of tumours and papillomas in mice exposed to UVA q B than UVB alone Ž P s 0.03.. Average tumour areas were also measured ŽTable 3.. In terms of tumour burden it would appear that black and green teas were equally as effective at reducing the tumour area in the UVA q B irradiated mice Ž P - 0.05., but black tea was clearly superior in the UVB exposed mice Ž P - 0.0001..
4. Discussion Table 3 Average areas of tumours on mice a Tumour area Žmm UVAqB
UVB
a
Water Black tea 1.25% Green tea 1.25% Water Black tea 1.25% Green tea 1.25%
2.
34.9"8.6 20.9"8.2 b 20.3"8.5 b 58.0"11.3 13.1"3.3 c 49.0"9.3
Values are means"S.E.M. of 19–20 animals per group. Significantly less than water group P - 0.05. c Significantly less than any other group P - 0.05. b
We in this laboratory and other workers have previously reported that a number of dietary antioxidants can provide protection against UV-associated free radical damage to skin in vitro and in vivo w15–22x. More recently interest has focused on some of the non-nutrient dietary antioxidants, and in particular various teas as protectants against skin cancer. Wang et al. w10x were the first to study the effects of a green tea polyphenolic fraction on UVB-related photocarcinogenesis in mice. At a dose of 6 mgrday they observed a significant reduction in the number
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
of tumours per mouse, as well as a significant lengthening of the TDT50 . Several more recent studies have also described the effects of green tea polyphenols on chemically-induced and chemicallypromoted skin cancer w23–26x. Protection by green tea and black tea infusions against skin cancer in hairless mice has also been reported w11,12,27x. In the first of these studies w11x it was demonstrated that tea consumption significantly inhibited the formation of skin cancers promoted by TPA and also markedly reduced the intensity of UVB-induced skin lesions. The same group later noted that administration of green tea in the drinking water, or intraperitoneal injection of Žy.-epigallocatechin gallate inhibited tumour growth, also caused chemically-induced and chemically-promoted established papillomas to regress either partially, or in a few cases completely w27x. Similar results were obtained when UVB and TPA were used to produce the tumours and papillomas. Wang et al. w9x extended their studies to mice initiated with 7,12-dimethylbenz wa xanthracene ŽDMBA. and subsequently exposed to UVB. In this experiment black tea was found to protect as effectively as green tea, although the decaffeinated teas seemed to be slightly less effective than regular teas at the highest dose used. In the current study we refrained from using chemical initiators or promoters since some of these, TPA in particular, are known to cause free radical damage to cells. We also compared UVB with mixed UVA q B, since the mixed spectrum more closely simulates solar radiation. The early events Žsunburn cell formation, necrotic cells, mitotic figures and neutrophil infiltration. were not dramatically affected by consumption of tea, although there was a trend for most variables to be reduced in the tea-consuming groups. On exposure to UV irradiation the skin responds in a number of ways. Epidermal cells exposed to sufficient damage can undergo apoptosis or ‘programmed cell death’; this is most obviously manifested as ‘sunburn cells’. A reduction in the number of sunburn cells has been used as an indication of a protective effect of sunscreens. Exposure of epidermal cells to antioxidants has been demonstrated to reduce the number of sunburn cells, presumably because some of the free radicals generated by UV can be removed before they cause excessive damage w15x. The number of
197
necrotic cells also tends to be lower in the tea groups. Following irradiation, cell division is usually delayed somewhat. This provides cells more time in which to affect the repair of cellular and genomic damage. This repair time is reflected in the number of mitotic figures present, so that a reduction in the number of figures may well provide a fair representation of the degree of damage. In this study administration of tea had little effect. Neutrophil infiltration, which is due to increased blood flow in the dermis, also tended to be lower in the tea groups Žexcept for the black tea 1.25% which was inexplicably high., suggesting that tea may be able to reduce the erythema Žredness. associated with sun exposure. Other workers have reported beneficial effects of tea and its catechins against early markers of skin damage, including induction of ornithine decarboxylase activity w28x, contact hypersensitivity and inflammation w29x and lipid peroxidation in mouse skin microsomes in vitro w30x. Previous workers have used chemical initiators or promoters to induce skin cancers, and there appears to be only one other study in which the effects of green and black teas have been compared w9x. In this earlier study there seemed to be little difference between the protective effects of green and black teas, although decaffeinated teas did seem to be somewhat less effective. In the present study black tea provided significantly more protection than green tea, reducing the yield of papillomas and tumours and appearing to delay the onset of papilloma development Žespecially in the animals exposed to UVA q B.. Although there was a plateau effect on papilloma formation in tea-consuming animals, there was a decline in the number of papillomas in the control groups, together with an increase in the number of tumours, suggesting that this reduction could be an artefact, almost certainly involving the conversion of papillomas to tumours. The differences between UVA q B and UVB were small, although there were indications that cell necrosis and the number of mitotic figures were less affected by the combined irradiation regime. The number of tumours and total tumour area in the green tea-drinking groups and the area of tumours in the water group were lower in the UVA q B group than the UVB group. Similar effects of an amelioration of skin tumours in mice has been observed
198
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199
before w31x, although later studies have not been entirely confirmatory w32x. Clearly this area requires further research, but in a manner which is beyond the scope of this present study. In recognition of the fact that solar radiation is mixed UVA and UVB future studies will only involve UVA q B exposure. In summary, our results suggest that consumption of either green or black tea may have the potential to reduce some of the early effects of UV exposure, albeit to a relatively small degree. Consumption of either green or black tea was shown to be associated with protection against skin carcinogenesis by UVB and UVA q B, thereby confirming reports from other laboratories. In contrast to one other study involving black tea, we found that black tea was, if anything, more beneficial than green tea. It may be that the use of initiation or promotion protocols by earlier workers will prove to have some bearing on this discrepancy. There may be important implications for human health, since most of the world tea production is black, and much of this is consumed with milk. Clearly further studies are required, both to confirm the difference between black and green tea and to ascertain whether addition of milk to the beverage has any effect.
Acknowledgements The authors wish to thank Ms. Mary Jannes and Ms. Linda Ormond for excellent technical assistance, TJ Lipton, Englewood Cliffs, NJ, for the teas, and Unifoods ŽAustralia. for financial support.
References w1x A. Green, D. Battistutta, A. Russel, C. Frost, Skin cancer: the quiet epidemic, Proc. Mt. Buller Conf. Environmental Radiation, 1996, p. 16 ŽAbs.. w2x American Cancer Society, Cancer facts and figures, Am. Cancer Soc., Washington, D.C., 1991. w3x I.E. Dreosti, Bioactive ingredients: antioxidants and polyphenols in tea, Nutr. Rev. 54 Ž1996. s51–s58. w4x R.A. Goldbohm, M.G.L. Hertog, H.A. M Brants, G. van Poppel, P.A. van den Brandt, Consumption of black tea and cancer risk: a prospective cohort study, J. Nat. Cancer Inst. 88 Ž1996. 93–100.
w5x S.K. Katiyar, H. Mukhtar, Tea consumption and cancer, in: A. Simopoulos ŽEd.., World Rev. Nutr. Diet. Karger, Basel, Vol. 79, 1996, pp. 154–184. w6x H.N. Graham, Green tea consumption, consumption and polyphenol chemistry, Prev. Med. 21 Ž1992. 334–350. w7x M. Serafini, A. Ghiselli, A. Ferro-Luzzi, In vivo antioxidant effects of green and black tea in man, Eur. J. Clin. Nutr. 50 Ž1996. 28–32. w8x S. Maxwell, G. Thorpe, Tea flavonoids have little short term impact on serum antioxidant activity, Br. Med. J. 313 Ž1996. 229. w9x Z.Y. Wang, M.T. Huang, Y.R. Lou, J.G. Xie, K.R. Reuhl, H.L. Newmark, C.T. Ho, C.S. Yang, A.H. Conney, Inhibitory effects of black tea, green tea, decaffeinated black tea and decaffeinated green tea on ultraviolet B light-induced skin carcinogenesis in 7,12-dimethylbenzwaxanthraceneinitiated SKH-1 mice, Cancer Res. 54 Ž1994. 3235–3248. w10x Z.Y. Wang, R. Agarwal, D.R. Bickers, H. Mukhtar, Protection against ultraviolet B radiation-induced photocarcinogenesis in hairless mice by green tea polyphenols, Carcinogenesis 12 Ž1991. 1527–1530. w11x Z.Y. Wang, M.T. Huang, T. Ferraro, C.Q. Wong, Y.R. Lou, K. Reuhl, M. Iatropoulos, C.S. Yang, A.H. Conney, Inhibitory effect of green tea in the drinking water on tumorigenesis by ultraviolet light and 12-O-tetradecanoylphorbol13-acetate in the skin of SKH-1 mice, Cancer Res. 52 Ž1992. 1162–1170. w12x A.H. Conney, Z.Y. Wang, M.T. Huang, C.T. Ho, C.S. Yang, Inhibitory effect of green tea on tumorigenesis by chemicals and ultraviolet light, Prev. Med. 21 Ž1992. 361–369. w13x P.G. Reeves, AIN-76 diet: should we change the formulation?, J. Nutr. 119 Ž1989. 1081–1082. w14x L.D. Leder, The chloroacetate esterase reaction, Am. J. Dermatol. 1 Ž1979. 39–42. w15x Y. Miyachi, T. Horio, S. Imamura, Sunburn cell formation is prevented by scavenging oxygen intermediates, Clin. Exp. Derm. 8 Ž1983. 305–310. w16x I.R. Record, I.E. Dreosti, M. Konstantinopoulos, R.A. Buckley, The influence of topical and systemic vitamin E on ultra violet light-induced skin damage in hairless mice, Nutr. Cancer 16 Ž1991. 219–225. w17x I.R. Record, L. Cavallo, I.E. Dreosti, Protection against UVA and UVB induced lipid peroxidation in liposomes by different forms of vitamins C and E, in: K. Yagi ŽEd.., International Congress Series No. 998; 5th International Conference on Oxygen Radicals. Elsevier, Amsterdam, 1992, pp. 621– 624. w18x I.R. Record, M. Jannes, I.E. Dreosti, Protection by zinc against UVA- and UVB-induced cellular and genomic damage in vivo and in vitro, Biol. Trace Elem. Res. 53 Ž1996. 19–25. w19x J.-P. Perchellet, N.L. Abney, R.M. Thomas, Y.L. Guislain, E.M. Perchellet, Effects of combined treatments with selenium, glutathione and vitamin E on glutathione peroxidase activity, ornithine decarboxylase induction and complete and multistage carcinogenesis in mouse skin, Cancer Res. 47 Ž1987. 477–485.
I.R. Record, I.E. Dreostir Mutation Research 422 (1998) 191–199 w20x K.E. Gerrish, H.L. Gensler, Prevention of photocarcinogenesis by dietary vitamin E, Nutr. Cancer 19 Ž1993. 125–133. w21x L.A. Lambert, W.G. Wamer, R.R. Wei, S. Lavu, S.J. Chirtel, A. Kornhauser, The protective but nonsynergistic effect of dietary b-carotene and vitamin E on skin tumourigenesis in Skh mice, Nutr. Cancer 21 Ž1994. 1–12. w22x M.M. Mathews-Roth, N.I. Krinsky, Carotenoids affect development of UV-B induced skin cancer, Photochemistry and Photobiology 46 Ž1987. 507–509. w23x M.-T. Huang, C.-T. Ho, Z.Y. Wang, T. Ferraro, T. Finnegan-Olive, Y.-R. Lou, J.M. Mitchell, J.D. Laskin, H. Newmark, C.S. Yang, A.H. Conney, Inhibitory effect of topical application of a green tea polyphenol fraction on tumour initiation and promotion in mouse skin, Carcinogenesis 13 Ž1992. 947–954. w24x S.K. Katiyar, R. Agerwal, G.S. Wood, H. Mukhtar, Inhibition of 12-O-tetradecanoylphorbol-13-acetate-caused tumour promotion in 7,12-dimethylbenz wax anthracene-initiated SENCAR mouse skin by a polyphenolic fraction isolated from green tea, Cancer Res. 52 Ž1992. 6890–6897. w25x S.K. Katiyar, R. Agarwal, H. Mukhtar, Inhibition of both stage 1 and stage 11 skin tumour promotion in SENCAR mice by a polyphenolic fraction isolated from green tea: inhibition depends on the duration of polyphenol treatment, Carcinogenesis 14 Ž1993. 2641–2643. w26x S.K. Katiyar, R. Agarwal, H. Mukhtar, Protection against malignant conversion of chemically induced benign skin papillomas to squamous cell carcinomas in SENCAR mice
w27x
w28x
w29x
w30x
w31x
w32x
199
by a polyphenolic fraction isolated from green tea, Cancer Res. 53 Ž1993. 5409–5412. Z.Y. Wang, M.-T. Huang, C.-T. Ho, R. Chang, W. Ma, T. Ferraro, K.R. Reuhl, C.S. Yang, A.H. Conney, Inhibitory effect of green tea on the growth of established skin papillomas in mice, Cancer Res. 52 Ž1992. 6657–6665. R. Agarwal, S.K. Katiyar, S.I.A. Zaidi, H. Mukhtar, Inhibition of skin tumour promoter-caused induction of epidermal ornithine decarboxylase in SENCAR mice by polyphenolic fraction isolated from green tea and its individual epicatechin derivatives, Cancer Res. 52 Ž1992. 3582–3588. S.K. Katiyar, C.A. Elmets, R. Agarwal, H. Mukhtar, Protection against ultraviolet-B radiation-induced local and systemic suppression of contact hypersensitivity and edema responses in C3HrHEN mice by green tea polyphenols, Photochemistry and Photobiology 62 Ž1995. 855–861. S.K. Katiyar, R. Agarwal, H. Mukhtar, Inhibition of spontaneous and photo-enhanced lipid peroxidation in mouse epidermal microsomes by epicatachin derivatives from green tea, Cancer Letters 79 Ž1994. 61–66. H. Van Weelden, J.C. van der Leun, Photorecovery by UVA, in: F. Urbach, R.W. Gange ŽEds.., The Biological Effects of UVA Radiation, Praeger, New York, 1986, pp. 147–152. R.J.W. Berg, F.R. de Gruiji, J.C. van der Leun, Interaction between ultraviolet A and ultraviolet B radiations in skin cancer induction in hairless mice, Cancer Res. 53 Ž1993. 4212–4217.