CED
Experimental dermatology • Concise report
Clinical and Experimental Dermatology
CPD
Isoflavones in aglycone solution enhance ultraviolet B-induced DNA damage repair efficiency B. Iovine,1 M. Garofalo,1 M. Orefice,1 V. Giannini,2 F. Gasparri,2 G. Monfrecola3 and M. A. Bevilacqua1 1 Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, Naples, Italy; 2Rottapharm-Madaus Dermo-Cosmetic R & D Division, Monza, Italy; and 3Section of Dermatology, Department of Clinical Medicine and Surgery, University of Naples Federico II, Naples, Italy
doi:10.1111/ced.12290
Summary
The isoflavones daidzein and genistein are natural compounds which have antiinflammatory and photoprotective activities, and may be effective in the repair of ultraviolet (UV)-induced photodamage. In this study, an alcoholic solution of aglycone isoflavones with a genistein:daidzein ratio of 1:4 [Rottapharm (RPH)-aglycone] was examined for its effects on the repair of DNA damage induced by a single dose of UVB irradiation (20 mJ/cm2). For this purpose, human skin cells were first UVBirradiated and then treated with RPH-aglycone. Comet assay analysis was used to estimate the UVB-induced DNA damage at different time points after treatment by measuring the tail moment parameter. We found that treatment with 10 lmol/L RPH-aglycone solution resulted in a significantly reduced tail moment at 1 h after treatment, and 34–35% enhancement of damage repair at 4 h after treatment. These results suggest that isoflavone aglycones are protective against UVB-induced DNA damage.
Solar ultraviolet (UV) rays are responsible for the majority of skin damage. UVA represents the majority of the solar radiation at the earth’s surface, but only 10% of the carcinogenic dose of sunlight. UVB radiation is a minor but active constituent of solar light, has direct and indirect adverse biological effects on the skin, and is a complete carcinogen that is absorbed by DNA and induces damage directly.1,2 The prevention of photodamage has been continuously emphasized in recent years, and the use of natural compounds as photoprotective or repair agents has attracted renewed interest, because of their nontoxic
Correspondence: Professor Maria A. Bevilacqua, Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, 80131 Naples, Italy E-mail: [email protected] Conflict of interest: VG is a Dermo-Cosmetic R & D Specialist for Rottapharm-Madaus and FG is an advisor to Rottapharm-Madaus. The other authors declare that they have no conflicts of interest. Accepted for publication 4 November 2013
ª 2014 British Association of Dermatologists
effects and their ability to modulate multiple pathways.3 The naturally occurring isoflavones daidzein and genistein have been investigated extensively because of their antioxidant, chemopreventive and anticancer activities.4,5 In a previous study, we showed that genistein and daidzein, administered at a specific concentration and ratio, exerted a synergistic photoprotective effect in human skin cells subjected to 60 mJ/cm2 of UVB irradiation.6 In addition, we recently reported that the aglycone and glucoside forms of these isoflavones act as anti-inflammatory and photoprotective agents. Moreover, the effects of the aglycones genistein and daidzein were enhanced when combined in an alcoholic solution in a specific formulation. This solution, referred to as Rottapharm (RPH)-aglycone, is a standardized glycine soy extract that is titrated to 90% isoflavone aglycones, at a ratio of 1:4 genistein: daidzein.7 Based on our previous findings, we hypothesized that RPH-aglycone would affect the efficiency of UVB-induced DNA damage repair.
Clinical and Experimental Dermatology (2014) 39, pp391–394
391
Effects of isoflavones on DNA damage repair B. Iovine et al.
In this study, we analysed the effect of this aglycone solution on the repair of DNA damage in UVB-irradiated (20 mJ/cm2) BJ-5ta human skin cells.
Report To assess the effects of RPH on repair efficiency, BJ-5ta human skin cells were first irradiated with a single dose of UVB (20 mJ/cm2) and then treated with 10 lmol/L of RPH-aglycone solution. The cells were harvested between 1 and 24 h after treatment. We used a comet assay to measure the effect. The comet assay, or single-cell gel electrophoresis assay, is a sensitive method for the evaluation of DNA damage/repair and toxicity at the cellular level. The test involves the encapsulation of cells in low-meltingpoint agarose. The cells are lysed and subjected to electrophoretic migration. Under an electrophoretic field, the damaged cellular DNA is separated from the
10 9
NO UVB UVB 20 mJ RPH 10 uM UVB+RPH
8
***
***
Tail moment
7
%Repair ¼ 100 ½TM%ðUVBÞ=TM%ðUVB þ RPHÞ
***
***
**
**
***
***
6
***
5
***
**
***
4 3 2 1
***
***
***
***
***
1h
2h
4h
6h
12 h
***
0 T0
24 h
Figure 1 Analysis of ultraviolet (UVB)-induced DNA damage by
comet assay. BJ-5ta human skin cells were irradiated with a single dose of 20 mJ/cm2 of UVB and subsequently treated with 10 lmol/L of RPH-aglycone. The analysis of the aglycone solution in alcohol found that the product purity was > 90%. The cells were harvested at 0, 1, 2, 4, 6, 12 and 24 h after treatment. The comet assay procedure (carried out with a CometAssay Kit (Trevigen Inc., Gaithersberg, MD, USA) was performed according to the manufacturer’s instructions. At least 50–100 cells per sample were analysed. Statistical significance was evaluated by performing one-way analysis of variance (ANOVA) using Prism GraphPad software (version 4.0; GraphPad Software Inc. La Jolla, CA, USA). The Shapiro–Wilk normality test was used to verify the normal distribution of the data. The data are reported as the tail moments (TMs) and represent the mean SD of three independent experiments. Red line, control cells; pink line, cells treated with 10 lmol/L RPH-aglycone; green line, cells irradiated with 20 mJ/cm2 UVB; blue line, cells irradiated with 20 mJ/cm2 UVB and then treated with 10 lmol/L RPH-aglycone. Significance: **P < 0.001; ***P < 0.0001.
392
intact DNA, yielding the classic ‘comet tail’. A long and intensely fluorescent tail indicates that DNA fragmentation, i.e. DNA damage, has occurred.7 The effect of 10 lmol/L RPH on the efficiency of DNA repair was evaluated by measuring the tail moment (TM) parameter and calculating the fluorescence intensity. The TM is defined as the product of the tail length and the fraction of total DNA in the tail (%tail DNA 9 tail length). The data analysis was carried out using Comet Score software (Comet Assay IV, Perceptive Instruments, Blois Meadow, Suffolk, UK). We measured the TM in the following samples (Fig. 1): (i) non-irradiated control cells; (ii) cells treated with 10 lmol/L RPH-aglycone solution without irradiation; (iii) cells irradiated with UVB (20 mJ/cm2); and (iv) cells first irradiated and then treated with RPH (10 lmol/L). All of the samples were harvested at different time points between 1 and 24 h after treatment. RPH treatment significantly reduced the TM at 1 and 4 h (Fig. 1). In addition, we evaluated the repair efficiency of the aglycone mixture according to the following formula:
Clinical and Experimental Dermatology (2014) 39, pp391–394
Table 1 shows the results obtained in the samples first irradiated with 20 mJ/cm2 of UVB, then treated with 10 lmol/L of RPH and harvested at different time points. As reported in figure 2a we found an increase of approximately 34–35% in the repair rate at 1 and 4 h after treatment with the RPH solution (Fig. 2a). Fig. 2b shows a comparison between the 20 mJ/cm2 UVB-irradiated cells and the UVB-irradiated cells treated with RPH (10 lmol/L) for 4 h. In a previously published study, we showed that 2 h of pre-treatment with RPH-aglycone (8 and 10 lmol/L) had antioxidant and photoprotective effects on BJ-5ta human skin cells irradiated with 60 mJ/cm2 of UVB.7
Table 1 Percentage DNA repair with time in cells subjected to
irradiation with ultraviolet B (20 mJ/cm2) and treated with 10 lmol/L RPH-aglycone at different time points. Time, h 0 1 2 4 6 12 24
Repair, % 0 34 33.8 35 28 28.7 23.2
ª 2014 British Association of Dermatologists
Effects of isoflavones on DNA damage repair B. Iovine et al.
(a) 40 35
% Repair
30 25 20 15 10 5 0 1
0
2
4
6
12
24
Time (h)
involved in the sunburn reaction, even when applied after UV exposure.10 Therefore, studies of the immune system involvement in DNA damage repair could provide a future line of investigation to determine how RPH reduces the DNA damage. In conclusion, our results suggest that this isoflavone aglycone solution promotes the efficient and rapid repair of UVB-induced DNA damage. In the future, we plan to extend our assessments to additional skin cell types, such as primary human dermal fibroblasts or keratinocytes cell lines, to validate this RPH-aglycone solution as an effective cosmetic ingredient to protect against the effects of sun exposure.
(b)
Acknowledgements We are grateful to the Rottapharm-Madaus DermoCosmetic R & D Division for funding this research and providing the RPH-aglycone solution. 4h
4h
UVB 20 mJ/cm2
+
+
RPH 10 µM
–
+
Figure 2 RPH-aglycone mixture protects against DNA damage.
(a) Graphical analysis of the percentage repair; (b) images of 20 mJ/cm2 ultraviolet (UVB)-irradiated cells with or without RPHaglycone (10 lmol/L) treatment at 4 h.
In the current study, we analysed the effect of 10 lmol/L RPH-aglycone solution on the repair of DNA damage induced by a single dose of UVB irradiation (20 mJ/cm2) in the same human skin cell line. This dose of UVB was chosen based on the results of previous tests, which showed that it did not affect cell viability but did induce DNA damage, consistent with mRNA expression of Gadd45, a gene that is rapidly induced upon DNA damage.6,7 In the comet assays performed in this study, treatment with 10 lmol/L RPH-aglycone resulted in a reduced tail moment. In addition, the percentage of DNA repair after 20 mJ/cm2 UVB-induced DNA photodamage was increased by approximately 34–35% at 2 and 4 h after treatment. Our results highlight two important points. Firstly, treatment of cells with 10 lmol/L RPH-aglycone reduced DNA damage at 1–4 h UVB irradiation. Secondly, the repair activity was promoted by a combination of the two isoflavone aglycones.8 These observations are consistent with recent studies reporting the preventative effects of natural compounds against photodamage.9 Recently, it was shown that a lotion containing genistein readily protected the immune system from both the photosuppression and inflammation
ª 2014 British Association of Dermatologists
Learning points
•
RPH is a standardized glycine soy extract titrated to 90% isoflavone aglycones (1:4 genistein:daidzein). • It is an active isoflavone preparation that protects against UVB-induced DNA damage in BJ-5ta human skin cells. • RPH-aglycone significantly reduced the UVBinduced TM in the comet assay, repairing 34– 35% of the DNA photodamage induced by 20 mJ/cm2 UVB. • RPH-aglycone efficiently and rapidly repairs UVB-induced damage, and it may be an effective dermatological sun protectant.
References 1 Narayanan DL, Saladi RN, Fox JL. Ultraviolet radiation and skin cancer. Int J Dermatol 2010; 49: 978–86. 2 Courdavault S, Baudouin C, Sauvaigo S et al. Unrepaired cyclobutane pyrimidine dimers do not prevent proliferation of UVB-irradiated cultured human fibroblasts. Photochem Photobiol 2004; 79: 145–51. 3 F’guyer S, Afaq F, Mukhtar H. Photochemoprevention of skin cancer by botanical agents. Photodermatol Photoimmunol Photomed 2003; 19: 56–72. 4 Wei H, Saladi R, Lu Y et al. Isoflavone genistein: photoprotection and clinical implications in dermatology. J Nutr 2003; 133(Suppl): 3811–19S.
Clinical and Experimental Dermatology (2014) 39, pp391–394
393
Effects of isoflavones on DNA damage repair B. Iovine et al.
5 Widyarini S, Domanski D, Painter N et al. Photoimmune protective effect of the phytoestrogenic isoflavonoid equol is partially due to its antioxidant activities. Photochem Photobiol Sci 2012; 11: 1186–92. 6 Iovine B, Iannella ML, Gasparri F et al. Synergic effect of genistein and daidzein on UVB-induced DNA damage: an effective photoprotective combination. J Biomed Biotechnol 2011; 2011: 1–8. 7 Iovine B, Iannella ML, Gasparri F et al. A comparative analysis of the photo-protective effects of soy isoflavones in their aglycone and glucoside forms. Int J Mol Sci 2012; 13: 16444–56.
394
Clinical and Experimental Dermatology (2014) 39, pp391–394
8 Lin JY, Tournas JA, Burch JA et al. Topical isoflavones provide effective photoprotection to skin. Photodermatol Photoimmunol Photomed 2008; 24: 61–6. 9 Chadha G, Sathigari S, Parsons DL et al. In vitro percutaneous absorption of genistein from topical gels through human skin. Drug Dev Ind Pharm 2011; 37: 498–505. 10 Widyarini S, Spinks N, Husband AJ et al. Isoflavonoid compounds from red clover (Trifolium pratense) protect from inflammation and immune suppression induced by UV radiation. Photochem Photobiol 2001; 74: 465–70.
ª 2014 British Association of Dermatologists
Experimental dermatology • Concise report
Clinical and Experimental Dermatology
CPD
Isoflavones in aglycone solution enhance ultraviolet B-induced DNA damage repair efficiency B. Iovine,1 M. Garofalo,1 M. Orefice,1 V. Giannini,2 F. Gasparri,2 G. Monfrecola3 and M. A. Bevilacqua1 1 Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, Naples, Italy; 2Rottapharm-Madaus Dermo-Cosmetic R & D Division, Monza, Italy; and 3Section of Dermatology, Department of Clinical Medicine and Surgery, University of Naples Federico II, Naples, Italy
doi:10.1111/ced.12290
Summary
The isoflavones daidzein and genistein are natural compounds which have antiinflammatory and photoprotective activities, and may be effective in the repair of ultraviolet (UV)-induced photodamage. In this study, an alcoholic solution of aglycone isoflavones with a genistein:daidzein ratio of 1:4 [Rottapharm (RPH)-aglycone] was examined for its effects on the repair of DNA damage induced by a single dose of UVB irradiation (20 mJ/cm2). For this purpose, human skin cells were first UVBirradiated and then treated with RPH-aglycone. Comet assay analysis was used to estimate the UVB-induced DNA damage at different time points after treatment by measuring the tail moment parameter. We found that treatment with 10 lmol/L RPH-aglycone solution resulted in a significantly reduced tail moment at 1 h after treatment, and 34–35% enhancement of damage repair at 4 h after treatment. These results suggest that isoflavone aglycones are protective against UVB-induced DNA damage.
Solar ultraviolet (UV) rays are responsible for the majority of skin damage. UVA represents the majority of the solar radiation at the earth’s surface, but only 10% of the carcinogenic dose of sunlight. UVB radiation is a minor but active constituent of solar light, has direct and indirect adverse biological effects on the skin, and is a complete carcinogen that is absorbed by DNA and induces damage directly.1,2 The prevention of photodamage has been continuously emphasized in recent years, and the use of natural compounds as photoprotective or repair agents has attracted renewed interest, because of their nontoxic
Correspondence: Professor Maria A. Bevilacqua, Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, 80131 Naples, Italy E-mail: [email protected] Conflict of interest: VG is a Dermo-Cosmetic R & D Specialist for Rottapharm-Madaus and FG is an advisor to Rottapharm-Madaus. The other authors declare that they have no conflicts of interest. Accepted for publication 4 November 2013
ª 2014 British Association of Dermatologists
effects and their ability to modulate multiple pathways.3 The naturally occurring isoflavones daidzein and genistein have been investigated extensively because of their antioxidant, chemopreventive and anticancer activities.4,5 In a previous study, we showed that genistein and daidzein, administered at a specific concentration and ratio, exerted a synergistic photoprotective effect in human skin cells subjected to 60 mJ/cm2 of UVB irradiation.6 In addition, we recently reported that the aglycone and glucoside forms of these isoflavones act as anti-inflammatory and photoprotective agents. Moreover, the effects of the aglycones genistein and daidzein were enhanced when combined in an alcoholic solution in a specific formulation. This solution, referred to as Rottapharm (RPH)-aglycone, is a standardized glycine soy extract that is titrated to 90% isoflavone aglycones, at a ratio of 1:4 genistein: daidzein.7 Based on our previous findings, we hypothesized that RPH-aglycone would affect the efficiency of UVB-induced DNA damage repair.
Clinical and Experimental Dermatology (2014) 39, pp391–394
391
Effects of isoflavones on DNA damage repair B. Iovine et al.
In this study, we analysed the effect of this aglycone solution on the repair of DNA damage in UVB-irradiated (20 mJ/cm2) BJ-5ta human skin cells.
Report To assess the effects of RPH on repair efficiency, BJ-5ta human skin cells were first irradiated with a single dose of UVB (20 mJ/cm2) and then treated with 10 lmol/L of RPH-aglycone solution. The cells were harvested between 1 and 24 h after treatment. We used a comet assay to measure the effect. The comet assay, or single-cell gel electrophoresis assay, is a sensitive method for the evaluation of DNA damage/repair and toxicity at the cellular level. The test involves the encapsulation of cells in low-meltingpoint agarose. The cells are lysed and subjected to electrophoretic migration. Under an electrophoretic field, the damaged cellular DNA is separated from the
10 9
NO UVB UVB 20 mJ RPH 10 uM UVB+RPH
8
***
***
Tail moment
7
%Repair ¼ 100 ½TM%ðUVBÞ=TM%ðUVB þ RPHÞ
***
***
**
**
***
***
6
***
5
***
**
***
4 3 2 1
***
***
***
***
***
1h
2h
4h
6h
12 h
***
0 T0
24 h
Figure 1 Analysis of ultraviolet (UVB)-induced DNA damage by
comet assay. BJ-5ta human skin cells were irradiated with a single dose of 20 mJ/cm2 of UVB and subsequently treated with 10 lmol/L of RPH-aglycone. The analysis of the aglycone solution in alcohol found that the product purity was > 90%. The cells were harvested at 0, 1, 2, 4, 6, 12 and 24 h after treatment. The comet assay procedure (carried out with a CometAssay Kit (Trevigen Inc., Gaithersberg, MD, USA) was performed according to the manufacturer’s instructions. At least 50–100 cells per sample were analysed. Statistical significance was evaluated by performing one-way analysis of variance (ANOVA) using Prism GraphPad software (version 4.0; GraphPad Software Inc. La Jolla, CA, USA). The Shapiro–Wilk normality test was used to verify the normal distribution of the data. The data are reported as the tail moments (TMs) and represent the mean SD of three independent experiments. Red line, control cells; pink line, cells treated with 10 lmol/L RPH-aglycone; green line, cells irradiated with 20 mJ/cm2 UVB; blue line, cells irradiated with 20 mJ/cm2 UVB and then treated with 10 lmol/L RPH-aglycone. Significance: **P < 0.001; ***P < 0.0001.
392
intact DNA, yielding the classic ‘comet tail’. A long and intensely fluorescent tail indicates that DNA fragmentation, i.e. DNA damage, has occurred.7 The effect of 10 lmol/L RPH on the efficiency of DNA repair was evaluated by measuring the tail moment (TM) parameter and calculating the fluorescence intensity. The TM is defined as the product of the tail length and the fraction of total DNA in the tail (%tail DNA 9 tail length). The data analysis was carried out using Comet Score software (Comet Assay IV, Perceptive Instruments, Blois Meadow, Suffolk, UK). We measured the TM in the following samples (Fig. 1): (i) non-irradiated control cells; (ii) cells treated with 10 lmol/L RPH-aglycone solution without irradiation; (iii) cells irradiated with UVB (20 mJ/cm2); and (iv) cells first irradiated and then treated with RPH (10 lmol/L). All of the samples were harvested at different time points between 1 and 24 h after treatment. RPH treatment significantly reduced the TM at 1 and 4 h (Fig. 1). In addition, we evaluated the repair efficiency of the aglycone mixture according to the following formula:
Clinical and Experimental Dermatology (2014) 39, pp391–394
Table 1 shows the results obtained in the samples first irradiated with 20 mJ/cm2 of UVB, then treated with 10 lmol/L of RPH and harvested at different time points. As reported in figure 2a we found an increase of approximately 34–35% in the repair rate at 1 and 4 h after treatment with the RPH solution (Fig. 2a). Fig. 2b shows a comparison between the 20 mJ/cm2 UVB-irradiated cells and the UVB-irradiated cells treated with RPH (10 lmol/L) for 4 h. In a previously published study, we showed that 2 h of pre-treatment with RPH-aglycone (8 and 10 lmol/L) had antioxidant and photoprotective effects on BJ-5ta human skin cells irradiated with 60 mJ/cm2 of UVB.7
Table 1 Percentage DNA repair with time in cells subjected to
irradiation with ultraviolet B (20 mJ/cm2) and treated with 10 lmol/L RPH-aglycone at different time points. Time, h 0 1 2 4 6 12 24
Repair, % 0 34 33.8 35 28 28.7 23.2
ª 2014 British Association of Dermatologists
Effects of isoflavones on DNA damage repair B. Iovine et al.
(a) 40 35
% Repair
30 25 20 15 10 5 0 1
0
2
4
6
12
24
Time (h)
involved in the sunburn reaction, even when applied after UV exposure.10 Therefore, studies of the immune system involvement in DNA damage repair could provide a future line of investigation to determine how RPH reduces the DNA damage. In conclusion, our results suggest that this isoflavone aglycone solution promotes the efficient and rapid repair of UVB-induced DNA damage. In the future, we plan to extend our assessments to additional skin cell types, such as primary human dermal fibroblasts or keratinocytes cell lines, to validate this RPH-aglycone solution as an effective cosmetic ingredient to protect against the effects of sun exposure.
(b)
Acknowledgements We are grateful to the Rottapharm-Madaus DermoCosmetic R & D Division for funding this research and providing the RPH-aglycone solution. 4h
4h
UVB 20 mJ/cm2
+
+
RPH 10 µM
–
+
Figure 2 RPH-aglycone mixture protects against DNA damage.
(a) Graphical analysis of the percentage repair; (b) images of 20 mJ/cm2 ultraviolet (UVB)-irradiated cells with or without RPHaglycone (10 lmol/L) treatment at 4 h.
In the current study, we analysed the effect of 10 lmol/L RPH-aglycone solution on the repair of DNA damage induced by a single dose of UVB irradiation (20 mJ/cm2) in the same human skin cell line. This dose of UVB was chosen based on the results of previous tests, which showed that it did not affect cell viability but did induce DNA damage, consistent with mRNA expression of Gadd45, a gene that is rapidly induced upon DNA damage.6,7 In the comet assays performed in this study, treatment with 10 lmol/L RPH-aglycone resulted in a reduced tail moment. In addition, the percentage of DNA repair after 20 mJ/cm2 UVB-induced DNA photodamage was increased by approximately 34–35% at 2 and 4 h after treatment. Our results highlight two important points. Firstly, treatment of cells with 10 lmol/L RPH-aglycone reduced DNA damage at 1–4 h UVB irradiation. Secondly, the repair activity was promoted by a combination of the two isoflavone aglycones.8 These observations are consistent with recent studies reporting the preventative effects of natural compounds against photodamage.9 Recently, it was shown that a lotion containing genistein readily protected the immune system from both the photosuppression and inflammation
ª 2014 British Association of Dermatologists
Learning points
•
RPH is a standardized glycine soy extract titrated to 90% isoflavone aglycones (1:4 genistein:daidzein). • It is an active isoflavone preparation that protects against UVB-induced DNA damage in BJ-5ta human skin cells. • RPH-aglycone significantly reduced the UVBinduced TM in the comet assay, repairing 34– 35% of the DNA photodamage induced by 20 mJ/cm2 UVB. • RPH-aglycone efficiently and rapidly repairs UVB-induced damage, and it may be an effective dermatological sun protectant.
References 1 Narayanan DL, Saladi RN, Fox JL. Ultraviolet radiation and skin cancer. Int J Dermatol 2010; 49: 978–86. 2 Courdavault S, Baudouin C, Sauvaigo S et al. Unrepaired cyclobutane pyrimidine dimers do not prevent proliferation of UVB-irradiated cultured human fibroblasts. Photochem Photobiol 2004; 79: 145–51. 3 F’guyer S, Afaq F, Mukhtar H. Photochemoprevention of skin cancer by botanical agents. Photodermatol Photoimmunol Photomed 2003; 19: 56–72. 4 Wei H, Saladi R, Lu Y et al. Isoflavone genistein: photoprotection and clinical implications in dermatology. J Nutr 2003; 133(Suppl): 3811–19S.
Clinical and Experimental Dermatology (2014) 39, pp391–394
393
Effects of isoflavones on DNA damage repair B. Iovine et al.
5 Widyarini S, Domanski D, Painter N et al. Photoimmune protective effect of the phytoestrogenic isoflavonoid equol is partially due to its antioxidant activities. Photochem Photobiol Sci 2012; 11: 1186–92. 6 Iovine B, Iannella ML, Gasparri F et al. Synergic effect of genistein and daidzein on UVB-induced DNA damage: an effective photoprotective combination. J Biomed Biotechnol 2011; 2011: 1–8. 7 Iovine B, Iannella ML, Gasparri F et al. A comparative analysis of the photo-protective effects of soy isoflavones in their aglycone and glucoside forms. Int J Mol Sci 2012; 13: 16444–56.
394
Clinical and Experimental Dermatology (2014) 39, pp391–394
8 Lin JY, Tournas JA, Burch JA et al. Topical isoflavones provide effective photoprotection to skin. Photodermatol Photoimmunol Photomed 2008; 24: 61–6. 9 Chadha G, Sathigari S, Parsons DL et al. In vitro percutaneous absorption of genistein from topical gels through human skin. Drug Dev Ind Pharm 2011; 37: 498–505. 10 Widyarini S, Spinks N, Husband AJ et al. Isoflavonoid compounds from red clover (Trifolium pratense) protect from inflammation and immune suppression induced by UV radiation. Photochem Photobiol 2001; 74: 465–70.
ª 2014 British Association of Dermatologists
