The Journal of Dermatology Vol. 19: 544-547,1992
Aneuploidy in Chronic Actinic Dermatitis Qiu Bingsen, Liao Kanghuang and Ni Hongzhen* Abstract
DNA content and cell cycle distributions in paraffin-embedded blocks of 19 skin biopsy specimens from 17 patients with chronic actinic dermatitis (CAD) [8 patients showed typical actinic reticuloid (AR)] were estimated by DNA cytometry. Sixty-three percent (12/19) of the skin specimens showed aneuploidy. In the 4 cases with the highest DNA indices (DI), ranging from 1.65 to 1.88 (mean: 1.84), the proportions of cells in Sand G2/M phases were increased, ranging from 15-48% (mean: 20%) and from 64-76% (mean: 70.5%), respectively. In 8 cases with DI ranging from 1.15-1.75 (mean: 1.5), the proportion ofcells in S-phase was also increased, ranging from 30-90% (mean: 81.1%). Histologically, it seems likely that the relatively high frequency of aneuploidy, DI, and proportions of cells in the S and/or G2/M phases were not proportional to epidermal or vascular endothelial hyperplasia, but they might be related to dermal lymphoid infiltration. Abbreviations: CAD: chronic actinic dermatitis, AR: actinic reticuloid, DI: DNA indices, PD: photosensitivity dermatitis, FCM: flow cytometry, CML: cutaneous malignant lymphoma
Key words: aneuploidy; CAD; DI; FCM
Introduction Photosensitivity dermatitis (PD) and actinic reticuloid (AR) are the two extremes of the spectrum of chronic actinic dermatitis (CAD), formerly called PDfAR, which is a common eczematous photodennatosis. The finding of contact allergy that is frequently multiple complicates the clinical picture that may vary from early mild eczematous dermatitis to late severe conditions, such as erythroderma or a pseudolymphomatous state. Patch testing and phototesting are the key investigations, with broad UV waveband sensitivity occurring as a dermatitis rather than a sunburn response. To consider the potential progression of AR to malignant lymphoma, DNA flow cytometry (FCM) of affected formalin-fixed skin from six Received June 23, 1992; accepted for publication July 22, 1992. This project was supported by the National Natural Science Foundation of China. Institute of Dermatology, Huashan Hospital, and *Unit of DNA flow cytometry, Cancer Hospital, Shanghai Medical University, Shanghai, China. Reprint requests to: Qiu Bingsen, M.D., Institute of Dermatology, Huashan Hospital, Shanghai Medical University, Shanghai, 200040, China.
patients with histological and photobiological evidence of AR was studied by Norris et al. (1) in 1989. They found no aneuploidy in any patient. Recently, we estimated DNA content in paraffin-embedded tissue of skin biopsies taken from 17 patients with clinical, histological, and photobiological evidence of CAD by DNA FCM and found higher frequencies of aneuploidy in them.
Patients and Methods 1. Patients: Seventeen patients (sixteen men and one woman) with an age range of 46 to 84 years (median, 46 years) were studied. The duration of illness was 4 months to 20 years (mean: 2 years). They presented with persistent infiltrated papules and plaques on light-exposed skin, often with extension to covered areas. Eight patients manifestated typical AR. All were positive to phototesting, showing decreases of MED to UVB and UVA in 14 cases and of MED to UVBin 3 cases. Histologically, a dermal infiltrate with atypical lymphoid cells in varying numbers was present in most cases. The analysis of DNA content was simultaneously performed on paraffin blocks of skin biopsies taken from all 17 cases with CAD, 70 cases with CML,and
Aneuploidy in Chronic Actinic Dennatitis Table l. Case
Cell cycle distributions and DNA indices in skin specimens from 17 patients with CAD Cell cycle distributions (%) S G2/M
Go/G}
1 2 3
8 9 16
4
{5S 8 12 12 8 8 20 33 59 {62 95 72 73 81 95
5 6 7 8 9 10 11
12 13 14 15 16 17
545
18 15 20 25 48 90 85 85 82 81 77 62 39 36 1 25 22 14 2
74 76 64 67 2 2 3 3 10 10 3 5 2 2 4 3 5 5 3
Follow-up (Years)
DNA indices (Aneuploid)
Present status
1.88 1.87 1.80 1.65
A++ A A+
6 4 10
1.74 1.5 1.49 1.75 1.51 1.5 1.33 1.15
A+ A D A A++ A A A
4 3
L
4 2 3 2 4
L
A+ A A+
1 9 1
L
The mean coefficient of variation was 8. The DI in diploid regions was 1.0 ± 0.2. A=alive in remission, A+=alive with active disease, A++=alive with severe active disease, D=dead due to angina, Leelost to follow-up
24 cases with benign dermatoses (2). When the paraffin blocks were taken for cytometric analysis, HE slides were prepared from the same lesions for histological examination. The normal controls included two paraffin blocks of normal-appearing skin over the parotid region taken from two operated patients with benign parotid cysts. 2. Preparation of samples: Cell nuclei were isolated from the embedded material and stained as described by modified Hedley's method (3, 4). Briefly, two 40 pm sections were cut from the paraffin blocks, dewaxed in xylene, rehydrated through a series of alcohols (100%, 90%, 70% and 50%) and washed in PBS. Sections were then incubated in a 0.5% solution of pepsin adjusted to pH 1.5 with HCI for 50 min at 37°C. Once a minute, tire samples were vortexed vigorously. The nuclei Were treated with 1.0% RNase for 10 min at 37°C. 't'he number of nuclei was then adjusted to 2 x !()6/ml and stained with propidium iodide (0.05%) for 20 min at room temperature in a dark room. 3. FCM analysis: After sample preparation and Staining, nuclear DNA content was analyzed on the
EPICSV (Coulter Company, U.SA). Excitation was at 488 nm with a 5W argon-ion laser. In each sample, the DNA content of an average of 10,000 nuclei was measured, and the data was analyzed by PARA-I FCM program; then the DNA histogram and parameters were obtained. In general, DI is the mean of the ratio of the DNA content of the Go/G} cells in each patient's sample to that of the normal diploid Go/G} cells.
Results Peak DI and cell cycle distributions in skin specimens from 17 patients with CAD are summarized in Table 1. Sixty-three percent (12/19) of the skin specimens in 17 cases with CAD studied were aneuploid (Figs. 1, 2). One patient (case 4) showed aneuploidy in each of two skin specimens, but the proportion of cells in the S or G2/M phases was increased in each one differently. In 12 cases, the DI and the proportions of cells in Sand G2/M phases were correlated. In the 4 cases (case 1-4) with the highest DI,
546
Bingsen et al
L
Fluorescence intensity (proportional to DNA content!
Fluorescence intensity (proportional to DNA content)
Fig. 1. Representative histogram of DNA flow cytometry of paraffin-embedded tissue from normal control skin showing the DNA-diploid pattern.
Fig. 2. Representative histogram of DNA flow cytometry of paraffin-embedded tissue from case 10 of CAD showing a DNA-aneuploid pattern.
ranging from 1.65 to 1.88 (mean: 1.84), the proportions of cells in the Sand G2/M phases were highly increased, ranging from 15-48% (mean: 20%) and from 64-76% (mean: 70.5%), respectively. In 8 cases (case 5-12) with higher DIs ranging from 1.15 to 1.75 (mean: 1.5), the proportion of cells in the S-phase was also increased, ranging from 39-90% (mean: 81.1%). Histologically, the epidermal or vascular endothelial hyperplasia was not proportional to the frequency of aneuploidy. It seems likely that the activity of the disease was not proportional to the DI, proportion of cells in the S or G2/M phases, or aneuploidy.
or G2/M phases were not proportional to epidermal or vascular endothelial hyperplasia, but they might be attributed to dermal lymphoid infiltrates. The pathogenesis of CAD has been suggested in the literature by Norris et al. (1). Immunological effects of UV radiation include local inhibition of antigen presentation by Langerhans cells and induction of circulating antigen-specific suppressor/cytotoxic T-cells. Furthermore, abnormally low circulating helper/suppressor T-cell ratios have been demonstrated in photosensitive conditions such as erythrodermic AR Contact allergic sensitivity in patients with UV hypersensitivity and associated partial impairment of antigenic presentation might result in chronic low-grade antigenic stimulation of the immunological system, particularly lymphocytes, and eventually to the abnormal cellular response of AR Clinically, the skin lesions of CAD manifest a spectrum of morphological changes. At present, AR is believed to have the potential of progressing to malignant reticulosis. One patient with AR developed a reticulum cell sarcoma after 9 years (5) and two aparent sufferers from AR later died of Hodgkin's disease (6, 7). Recently, Dosaka et al. (8) have detected monoclonality of infiltrating lymphocytes in AR. Ralfkiaer et al. (9) have examined immunohistologically the cryostat sections of skin biopsies from five patients with CAD (PD/AR syndrome) by the alkaline phosphatase: anti-
Discussion The results were obtained from simultaneous application of DNA FCM to skin paraffin blocks from 111 cases with clinical, histological and photobiological or immunological evidence of AR, CML, and other benign dermatoses. The mean coefficient ofvariation was 8. The frequency of aneuploidy DI and proportion of cells in S or G 2/M phases in cases with CML and other benign dermatoses were basically correlated with their biological behavior and prognosis. Thus, the data obtained by using this method could be considered to be reliable. Although the epidermis was not separated from the paraffin blocks of these skin biopsies, it seems likely that histologically, the relatively high frequency of aneuploidy, high DI, and increase of proportion of cells in the S
Aneuploidy in Chronic Actinic Dermatitis
alkaline phosphatase staining technique using a panel of 24 monoclonal antibodies against lymphoid cells and their subsets. Their data indicated that a T-cell immune response is operative in cutaneous PDfAR lesions. In previous studies of dermal specimens from cutaneous T-cell lymphomas, it has been demonstrated that DNA histogram abnormalities in the form of hyperdiploid peaks frequently precede the development of definite malignant histology, and it has been suggested that hyperdiploid peaks may be indicative of the presence of malignant cell clones (10). However, the finding of hyperdiploid peaks in benign cutaneous lymphoid infiltrates, patch tests (11), and a range of other benign dermatoses such as eczema, psoriasis, lichen planus (12), and lymphomatoid papulosis (13) clearly demonstrates that hyperdiploid peaks are not necessarily indicative of the presence of separate cell clones. Thus, Ralfkiaer et al. emphasized that considerable caution is necessary in interpreting the diagnostic significance of hyperdiploid DNA histogram abnormalities. Our data, showing relatively high frequencies of aneuploidy, high DI, and increases of proportions of cells in the S or G2/M phases in CAD are not indicative of the presence of separate cell clones. We believe that CAD is similar to lymphomatoid papulosis, which is also considered to indicate malignancy or malignant potential and has shown DNA aneuploidy. References 1) Norris PG, NewtonJA, Camplejohn RS, HawkJLM: A flow cytometric study of actinic reticuloid, Clin Exp Dermatol, 14: 128-131,1989.
547
2) Qiu BS, Ni HZ: Application of DNA flow cytometry from paraffin-embedded tissue to the discrimination of cutaneous malignant lymphoma from pseudolymphoma. (in press) 3) Hedley DW, Friwdlander ML, Taylor IW: Frieication of DNAflow cytometry to paraffin-embedded archival material for the study of aneuploidy and its clinical significance, Cytometry, 6: 327-333, 1985. 4) Mclemor DD, Wall lJ, Stephens LC, Jardine JH: Processing of paraffin-embedded tissues for flow cytometry,] Histotedmol; 10: 109-111,1987. 5) Jenson NE, Sneddon IB: Actinic reticuloid with lymphoma, Brit] Dermatol; 82: 287-291, 1970. 6) Thomsen K: The development of Hodgkin's disease in a patient with actinic reticuloid, Clin Exp Dermatol, 2: 109-113, 1977. 7) Frain-bell W: Photosensitivity dermatitis and actinic reticuloid syndrome, in: Cutaneous Photobiology, Oxford University Press, 1985, p 119. 8) Dosaka N, Okawo H, Fujita M, et al: Detection of monoclonality of cutaneous infiltrating lymphocyte in actinic reticuloid, in Yamada M (eds): Lymplwmas of the Skin, Vol. 8, Hamamatsu 431-31, Japan, 1989, pp 22-25. 9) Ralfkiaer E, Wantzin GL, Stein H, Mason DY: Photosensitive dermatitis with actinic reticuloid syndrome: An immunohistological study of the cutaneous infiltrate, Br] Dermatol, 114: 47-56, 1986. 10) Wantzin GL, Larsen JK, Christensen 1], Rasmussen BB, Thomsen K: Aneuploidy in cutaneous T-cell lymphoma, ActaDermaio-uenereol (Stocklwlm), 63: 283, 1983. 11) Ralfkiaer E, Wantzin GL, Larsen jK, Christensen 1], Thomsen K: Single cell DNA measurements in benign cutaneous lymphoid infiltrates and in positive patch tests, Br] Dermatol; 112: 253-262, 1985. 12) Ralfkiaer E, LarsenJK, Christensen 1], Thomsen K, Wantzin GL: DNA analysis by flow cytometry in cutaneous T-cell lymphomas, Br ] Dermatol, 120: 597-605, 1989. 13) Wantzin GL, Larsen JK, Christensen IJ, Rasmussen BB: DNA analysis by flow in lymphomatoid papulosis, Clin Exp Dermatol, 8: 505-512, 1983.
Aneuploidy in Chronic Actinic Dermatitis Qiu Bingsen, Liao Kanghuang and Ni Hongzhen* Abstract
DNA content and cell cycle distributions in paraffin-embedded blocks of 19 skin biopsy specimens from 17 patients with chronic actinic dermatitis (CAD) [8 patients showed typical actinic reticuloid (AR)] were estimated by DNA cytometry. Sixty-three percent (12/19) of the skin specimens showed aneuploidy. In the 4 cases with the highest DNA indices (DI), ranging from 1.65 to 1.88 (mean: 1.84), the proportions of cells in Sand G2/M phases were increased, ranging from 15-48% (mean: 20%) and from 64-76% (mean: 70.5%), respectively. In 8 cases with DI ranging from 1.15-1.75 (mean: 1.5), the proportion ofcells in S-phase was also increased, ranging from 30-90% (mean: 81.1%). Histologically, it seems likely that the relatively high frequency of aneuploidy, DI, and proportions of cells in the S and/or G2/M phases were not proportional to epidermal or vascular endothelial hyperplasia, but they might be related to dermal lymphoid infiltration. Abbreviations: CAD: chronic actinic dermatitis, AR: actinic reticuloid, DI: DNA indices, PD: photosensitivity dermatitis, FCM: flow cytometry, CML: cutaneous malignant lymphoma
Key words: aneuploidy; CAD; DI; FCM
Introduction Photosensitivity dermatitis (PD) and actinic reticuloid (AR) are the two extremes of the spectrum of chronic actinic dermatitis (CAD), formerly called PDfAR, which is a common eczematous photodennatosis. The finding of contact allergy that is frequently multiple complicates the clinical picture that may vary from early mild eczematous dermatitis to late severe conditions, such as erythroderma or a pseudolymphomatous state. Patch testing and phototesting are the key investigations, with broad UV waveband sensitivity occurring as a dermatitis rather than a sunburn response. To consider the potential progression of AR to malignant lymphoma, DNA flow cytometry (FCM) of affected formalin-fixed skin from six Received June 23, 1992; accepted for publication July 22, 1992. This project was supported by the National Natural Science Foundation of China. Institute of Dermatology, Huashan Hospital, and *Unit of DNA flow cytometry, Cancer Hospital, Shanghai Medical University, Shanghai, China. Reprint requests to: Qiu Bingsen, M.D., Institute of Dermatology, Huashan Hospital, Shanghai Medical University, Shanghai, 200040, China.
patients with histological and photobiological evidence of AR was studied by Norris et al. (1) in 1989. They found no aneuploidy in any patient. Recently, we estimated DNA content in paraffin-embedded tissue of skin biopsies taken from 17 patients with clinical, histological, and photobiological evidence of CAD by DNA FCM and found higher frequencies of aneuploidy in them.
Patients and Methods 1. Patients: Seventeen patients (sixteen men and one woman) with an age range of 46 to 84 years (median, 46 years) were studied. The duration of illness was 4 months to 20 years (mean: 2 years). They presented with persistent infiltrated papules and plaques on light-exposed skin, often with extension to covered areas. Eight patients manifestated typical AR. All were positive to phototesting, showing decreases of MED to UVB and UVA in 14 cases and of MED to UVBin 3 cases. Histologically, a dermal infiltrate with atypical lymphoid cells in varying numbers was present in most cases. The analysis of DNA content was simultaneously performed on paraffin blocks of skin biopsies taken from all 17 cases with CAD, 70 cases with CML,and
Aneuploidy in Chronic Actinic Dennatitis Table l. Case
Cell cycle distributions and DNA indices in skin specimens from 17 patients with CAD Cell cycle distributions (%) S G2/M
Go/G}
1 2 3
8 9 16
4
{5S 8 12 12 8 8 20 33 59 {62 95 72 73 81 95
5 6 7 8 9 10 11
12 13 14 15 16 17
545
18 15 20 25 48 90 85 85 82 81 77 62 39 36 1 25 22 14 2
74 76 64 67 2 2 3 3 10 10 3 5 2 2 4 3 5 5 3
Follow-up (Years)
DNA indices (Aneuploid)
Present status
1.88 1.87 1.80 1.65
A++ A A+
6 4 10
1.74 1.5 1.49 1.75 1.51 1.5 1.33 1.15
A+ A D A A++ A A A
4 3
L
4 2 3 2 4
L
A+ A A+
1 9 1
L
The mean coefficient of variation was 8. The DI in diploid regions was 1.0 ± 0.2. A=alive in remission, A+=alive with active disease, A++=alive with severe active disease, D=dead due to angina, Leelost to follow-up
24 cases with benign dermatoses (2). When the paraffin blocks were taken for cytometric analysis, HE slides were prepared from the same lesions for histological examination. The normal controls included two paraffin blocks of normal-appearing skin over the parotid region taken from two operated patients with benign parotid cysts. 2. Preparation of samples: Cell nuclei were isolated from the embedded material and stained as described by modified Hedley's method (3, 4). Briefly, two 40 pm sections were cut from the paraffin blocks, dewaxed in xylene, rehydrated through a series of alcohols (100%, 90%, 70% and 50%) and washed in PBS. Sections were then incubated in a 0.5% solution of pepsin adjusted to pH 1.5 with HCI for 50 min at 37°C. Once a minute, tire samples were vortexed vigorously. The nuclei Were treated with 1.0% RNase for 10 min at 37°C. 't'he number of nuclei was then adjusted to 2 x !()6/ml and stained with propidium iodide (0.05%) for 20 min at room temperature in a dark room. 3. FCM analysis: After sample preparation and Staining, nuclear DNA content was analyzed on the
EPICSV (Coulter Company, U.SA). Excitation was at 488 nm with a 5W argon-ion laser. In each sample, the DNA content of an average of 10,000 nuclei was measured, and the data was analyzed by PARA-I FCM program; then the DNA histogram and parameters were obtained. In general, DI is the mean of the ratio of the DNA content of the Go/G} cells in each patient's sample to that of the normal diploid Go/G} cells.
Results Peak DI and cell cycle distributions in skin specimens from 17 patients with CAD are summarized in Table 1. Sixty-three percent (12/19) of the skin specimens in 17 cases with CAD studied were aneuploid (Figs. 1, 2). One patient (case 4) showed aneuploidy in each of two skin specimens, but the proportion of cells in the S or G2/M phases was increased in each one differently. In 12 cases, the DI and the proportions of cells in Sand G2/M phases were correlated. In the 4 cases (case 1-4) with the highest DI,
546
Bingsen et al
L
Fluorescence intensity (proportional to DNA content!
Fluorescence intensity (proportional to DNA content)
Fig. 1. Representative histogram of DNA flow cytometry of paraffin-embedded tissue from normal control skin showing the DNA-diploid pattern.
Fig. 2. Representative histogram of DNA flow cytometry of paraffin-embedded tissue from case 10 of CAD showing a DNA-aneuploid pattern.
ranging from 1.65 to 1.88 (mean: 1.84), the proportions of cells in the Sand G2/M phases were highly increased, ranging from 15-48% (mean: 20%) and from 64-76% (mean: 70.5%), respectively. In 8 cases (case 5-12) with higher DIs ranging from 1.15 to 1.75 (mean: 1.5), the proportion of cells in the S-phase was also increased, ranging from 39-90% (mean: 81.1%). Histologically, the epidermal or vascular endothelial hyperplasia was not proportional to the frequency of aneuploidy. It seems likely that the activity of the disease was not proportional to the DI, proportion of cells in the S or G2/M phases, or aneuploidy.
or G2/M phases were not proportional to epidermal or vascular endothelial hyperplasia, but they might be attributed to dermal lymphoid infiltrates. The pathogenesis of CAD has been suggested in the literature by Norris et al. (1). Immunological effects of UV radiation include local inhibition of antigen presentation by Langerhans cells and induction of circulating antigen-specific suppressor/cytotoxic T-cells. Furthermore, abnormally low circulating helper/suppressor T-cell ratios have been demonstrated in photosensitive conditions such as erythrodermic AR Contact allergic sensitivity in patients with UV hypersensitivity and associated partial impairment of antigenic presentation might result in chronic low-grade antigenic stimulation of the immunological system, particularly lymphocytes, and eventually to the abnormal cellular response of AR Clinically, the skin lesions of CAD manifest a spectrum of morphological changes. At present, AR is believed to have the potential of progressing to malignant reticulosis. One patient with AR developed a reticulum cell sarcoma after 9 years (5) and two aparent sufferers from AR later died of Hodgkin's disease (6, 7). Recently, Dosaka et al. (8) have detected monoclonality of infiltrating lymphocytes in AR. Ralfkiaer et al. (9) have examined immunohistologically the cryostat sections of skin biopsies from five patients with CAD (PD/AR syndrome) by the alkaline phosphatase: anti-
Discussion The results were obtained from simultaneous application of DNA FCM to skin paraffin blocks from 111 cases with clinical, histological and photobiological or immunological evidence of AR, CML, and other benign dermatoses. The mean coefficient ofvariation was 8. The frequency of aneuploidy DI and proportion of cells in S or G 2/M phases in cases with CML and other benign dermatoses were basically correlated with their biological behavior and prognosis. Thus, the data obtained by using this method could be considered to be reliable. Although the epidermis was not separated from the paraffin blocks of these skin biopsies, it seems likely that histologically, the relatively high frequency of aneuploidy, high DI, and increase of proportion of cells in the S
Aneuploidy in Chronic Actinic Dermatitis
alkaline phosphatase staining technique using a panel of 24 monoclonal antibodies against lymphoid cells and their subsets. Their data indicated that a T-cell immune response is operative in cutaneous PDfAR lesions. In previous studies of dermal specimens from cutaneous T-cell lymphomas, it has been demonstrated that DNA histogram abnormalities in the form of hyperdiploid peaks frequently precede the development of definite malignant histology, and it has been suggested that hyperdiploid peaks may be indicative of the presence of malignant cell clones (10). However, the finding of hyperdiploid peaks in benign cutaneous lymphoid infiltrates, patch tests (11), and a range of other benign dermatoses such as eczema, psoriasis, lichen planus (12), and lymphomatoid papulosis (13) clearly demonstrates that hyperdiploid peaks are not necessarily indicative of the presence of separate cell clones. Thus, Ralfkiaer et al. emphasized that considerable caution is necessary in interpreting the diagnostic significance of hyperdiploid DNA histogram abnormalities. Our data, showing relatively high frequencies of aneuploidy, high DI, and increases of proportions of cells in the S or G2/M phases in CAD are not indicative of the presence of separate cell clones. We believe that CAD is similar to lymphomatoid papulosis, which is also considered to indicate malignancy or malignant potential and has shown DNA aneuploidy. References 1) Norris PG, NewtonJA, Camplejohn RS, HawkJLM: A flow cytometric study of actinic reticuloid, Clin Exp Dermatol, 14: 128-131,1989.
547
2) Qiu BS, Ni HZ: Application of DNA flow cytometry from paraffin-embedded tissue to the discrimination of cutaneous malignant lymphoma from pseudolymphoma. (in press) 3) Hedley DW, Friwdlander ML, Taylor IW: Frieication of DNAflow cytometry to paraffin-embedded archival material for the study of aneuploidy and its clinical significance, Cytometry, 6: 327-333, 1985. 4) Mclemor DD, Wall lJ, Stephens LC, Jardine JH: Processing of paraffin-embedded tissues for flow cytometry,] Histotedmol; 10: 109-111,1987. 5) Jenson NE, Sneddon IB: Actinic reticuloid with lymphoma, Brit] Dermatol; 82: 287-291, 1970. 6) Thomsen K: The development of Hodgkin's disease in a patient with actinic reticuloid, Clin Exp Dermatol, 2: 109-113, 1977. 7) Frain-bell W: Photosensitivity dermatitis and actinic reticuloid syndrome, in: Cutaneous Photobiology, Oxford University Press, 1985, p 119. 8) Dosaka N, Okawo H, Fujita M, et al: Detection of monoclonality of cutaneous infiltrating lymphocyte in actinic reticuloid, in Yamada M (eds): Lymplwmas of the Skin, Vol. 8, Hamamatsu 431-31, Japan, 1989, pp 22-25. 9) Ralfkiaer E, Wantzin GL, Stein H, Mason DY: Photosensitive dermatitis with actinic reticuloid syndrome: An immunohistological study of the cutaneous infiltrate, Br] Dermatol, 114: 47-56, 1986. 10) Wantzin GL, Larsen JK, Christensen 1], Rasmussen BB, Thomsen K: Aneuploidy in cutaneous T-cell lymphoma, ActaDermaio-uenereol (Stocklwlm), 63: 283, 1983. 11) Ralfkiaer E, Wantzin GL, Larsen jK, Christensen 1], Thomsen K: Single cell DNA measurements in benign cutaneous lymphoid infiltrates and in positive patch tests, Br] Dermatol; 112: 253-262, 1985. 12) Ralfkiaer E, LarsenJK, Christensen 1], Thomsen K, Wantzin GL: DNA analysis by flow cytometry in cutaneous T-cell lymphomas, Br ] Dermatol, 120: 597-605, 1989. 13) Wantzin GL, Larsen JK, Christensen IJ, Rasmussen BB: DNA analysis by flow in lymphomatoid papulosis, Clin Exp Dermatol, 8: 505-512, 1983.
