- Email: [email protected]
Increased blood levels of NKG2D+CD4+ T cells in patients with alopecia areata
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did not affect serum CXCL10 levels, consistent with observations in our mouse model3 (Supplemental Fig 2; available at http://www.jaad.org). The most common side effects of simvastatin were self-limited myalgia in 4 participants and diarrhea in 2 participants. Three participants had mild, transient transaminitis and 4 had mild creatine phosphokinase elevations, none requiring dose modifications. Only 1 withdrawal, because of vertigo, was thought to be treatment-related. Our study does not support the use of oral simvastatin for the treatment of vitiligo. Disparate findings in this study compared with our mouse model may be a result of dosing limitations in humans because of potential toxicity, which is not a concern in mice. Our failure to show efficacy may also have been influenced by long-standing disease in our participants (responses are best in those with recent onset), small sample size, or lack of sensitive measures of treatment response. The VASI relies on an estimation of affected BSA, with limited sensitivity to change ([4.7% BSA).5 Newer outcome measures may provide more sensitive options for monitoring responses. Simvastatin may induce myopathy at higher doses; however, topical treatment may allow the delivery of sufficiently high local concentrations without systemic toxicity and could be tested in larger studies. The authors thank Dane Netherton for statistical assistance.
Stefan G. Vanderweil, MD, Shinya Amano, MD, PhD, Wei-Che Ko, MS, Jillian M. Richmond, PhD, Michelle Kelley, MSN, RN, Maryanne Makredes Senna, MD, Andrea Pearson, MD, Sandhya Chowdary, MD, Celia Hartigan, RN, MPH, Bruce Barton, PhD, and John E. Harris, MD, PhD Department of Medicine, Division of Dermatology, University of Massachusetts Medical School, Worcester, MA Funding sources: National Institutes of Health (NIH) Clinical and Translational Sciences Award UL1TR000161 through a Pilot Project Grant from the University of Massachusetts Center for Clinical and Translational Sciences; Physician-Scientist Career Development Award from the Dermatology Foundation; National Institute of Arthritis and Musculoskeletal and Skin Diseases, part of the NIH, under Award Number AR061437; and research grants from the Charles H. Hood Foundation, Vitiligo Research Foundation, and Kawaja Family Vitiligo Research Initiative.
Conflicts of interest: None declared. Correspondence to: John E. Harris, MD, PhD, University of Massachusetts Medical School, Lazare Research Bldg, Room 225, 364 Plantation St, Worcester, MA 01605 E-mail: [email protected] REFERENCES 1. Rashighi M, Agarwal P, Richmond JM, et al. CXCL10 is critical for the progression and maintenance of depigmentation in a mouse model of vitiligo. Sci Transl Med. 2014;6(223): 223ra23. 2. Zhao Y, Gartner U, Smith FJ, et al. Statins downregulate K6a promoter activity: a possible therapeutic avenue for pachyonychia congenita. J Invest Dermatol. 2011;131(5): 1045-1052. 3. Agarwal P, Rashighi M, Essien KI, et al. Simvastatin prevents and reverses depigmentation in a mouse model of vitiligo. J Invest Dermatol. 2015;135(4):1080-1088. 4. No€el M, Gagne C, Bergeron J, et al. Positive pleiotropic effects of HMG-CoA reductase inhibitor on vitiligo. Lipids Health Dis. 2004;3:7. 5. Komen L, da Graca V, Wolkerstorfer A, et al. Vitiligo Area Scoring Index and Vitiligo European Task Force assessment: reliable and responsive instruments to measure the degree of depigmentation in vitiligo. Br J Dermatol. 2015; 172(2):437-443. http://dx.doi.org/10.1016/j.jaad.2016.06.015
Increased blood levels of NKG2D1CD41 T cells in patients with alopecia areata To the Editor: Natural killer group 2D (NKG2D) is a key target of not only alopecia areata (AA) but other autoimmune diseases. AA is a TH1-mediated disease driven by interferon- (IFN- ) and IFN- einduced cytokines.1 NKG2D1 cells, including NK, NKT, and CD81 T cells, and NKG2D-activating ligands have a key role in AA pathogenesis. Excessive IFN- secretion by activated NKG2D1 cells can lead to collapse of hair follicle immune privilege.2 It is well known that CD41 T cells highly infiltrate the tissue of the ‘‘bee swarm’’ in AA.3 However, the functional role and expression patterns of NKG2D1CD41 T cell in AA remain largely unknown. In this study, we investigated the changes in blood levels of NKG2D1CD41 T cells in patients with AA. Forty-three patients with AA (18 men and 25 women) and 26 healthy controls (10 men and 16 women) with no scalp lesions in their personal history or on clinical examination were enrolled. The cell surface expression of NKG2D on peripheral blood mononuclear cells was studied by flow cytometry analysis (FACS Aria; BD Biosciences, Franklin Lakes, NJ). Results in each group were expressed as a percentage of CD41 T cells, CD81
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Fig 1. Results of fluorescence-activated cell sorting (FACS) analysis of peripheral blood and natural killer group 2D (NKG2D)1CD41 T, NKG2D1CD81 T, and NKG2D1CD561 NK cells in control and alopecia areata (AA) patient groups. Analysis of FACS data was carried out dividing into 4 times because of a large number of samples. The left bar graph indicates the average fold change of NKG2D1 cells from the 4 experiments (26 healthy controls and 43 patients with AA). NKG2D1 expressed on CD41 T cells was significantly higher in AA patient groups compared to control groups, as with NKG2D1CD81 T cells and NKG2D1CD561 NK cells. The right dot plot shows the representative results taken from the third experiment (10 healthy controls and 10 patients with AA).
Fig 2. Double-immunofluorescence analysis with antienatural killer group 2D (NKG2D; green) and anti-CD4 (red) antibodies. The merged image clearly shows infiltration of NKG2D1CD41 T cells in the lesional hair follicles of patients with alopecia areata. The two white arrows indicate the location of hair follicle.
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T cells, or CD561 NK cells expressing NKG2D. We collected blood samples 4 times, and the FACS analysis was also performed by dividing into 4 times. Therefore, final data were represented as a fold change of each cells expressing NKG2D over the healthy control (control set to 1). The patients were segregated into 2 or 3 subgroups according to AA severity, activity, type, and treatment modalities for subgroup analysis. Statistical analysis was performed using the ManneWhitney U test and KruskaleWallis analysis. Double-labeling immunofluorescence microscopy in 2 representative cases using freshly frozen specimens was also performed. The results showed that the levels of NKG2D1CD81 T cells and NKG2D1CD561 NK cells were higher in the AA group compared with the control group (P \ .05). Interestingly, a 2.49-fold increase in the CD41 T cells subset expressing NKG2D was detected in the peripheral blood from patients with AA compared with healthy controls (P \ .05; Fig 1). Although not statistically significant, the proportion of NKG2D1CD41 T cells was lower in patients who had mild severity AA and in those who received systemic immunosuppressive therapy. Double label immunofluorescence revealed NKG2D1 cells coexpressed CD4 around the hair follicles of the AA lesion (Fig 2). Recently, the role of NKG2D1CD41 T cells in other chronic autoimmune diseases, such as Crohn’s disease and rheumatoid arthritis, was highlighted.4 A subset of CD41 T cells expressing NKG2D was higher in the lamina propria of patients with Crohn’s disease.5 These NKG2D1CD41 T cell clones were functionally active through major histocompatibility complex class I-related chain A (MICA) eNKG2D interactions, producing IFN- and killing targets expressing MICA.5 We hypothesize that NKG2D1CD41 T cells might exhibit a TH1 cytokine profile in patients with AA. Our results suggest that IFN- eproducing NKG2D1CD41 T cells have the potential to become therapeutic targets and biologic markers of disease activity in AA. However, additional studies are needed to characterize the functional properties of NKG2D1CD41 T cells in patients with AA. Yong Hyun Jang, MD, PhD,a Jin Kyeong Choi, PhD,b,c Yun Hwan Jang, MD,a Sun Young Moon, MD,a Weon Ju Lee, MD, PhD,a Seok-Jong Lee, MD, PhD,a Young-Ae Choi, PhD,b Sang-Hyun Kim, PhD,b and Do Won Kim, MD, PhDa Departments of Dermatologya and Pharmacology,b School of Medicine, Kyungpook National University,
Daegu, Korea, and the Molecular Immunology Section,c Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland Drs Jang and J. K. Choi contributed equally to this work. Supported by the Basic Science Research Program through the National Research Foundation of Korea, funded by the Ministry of Education (NRF-2015R1D1A3A01016229 and 2014R1A5A 2009242). The protocol for the study was approved by the institutional review board of the Kyungpook National University Hospital (KNUH 2014-03-003). Conflicts of interest: None declared. Correspondence to: Sang-Hyun Kim, PhD, Department of Pharmacology, Kyungpook National University School of Medicine, 200, Dongduk-ro, Jung-gu, Daegu 700-422, Republic of Korea E-mail: [email protected] Or Do Won Kim, MD, PhD, Department of Dermatology, Kyungpook National University School of Medicine, 200, Dongduk-ro, Jung-gu, Daegu, 41944, Republic of Korea E-mail: [email protected] REFERENCES 1. Gilhar A, Etzioni A, Paus R. Alopecia areata. N Engl J Med. 2012; 366:1515-1525. 2. Paus R, Bertolini M. The role of hair follicle immune privilege collapse in alopecia areata: status and perspectives. J Investig Dermatol Symp Proc. 2013;16:S25-S27. 3. Ito T, Ito N, Saatoff M, et al. Maintenance of hair follicle immune privilege is linked to prevention of NK cell attack. J Invest Dermatol. 2008;128:1196-1206. 4. Killock D. Experimental arthritis: NKG2D: a potential therapeutic target in RA? Nat Rev Rheumatol. 2011;7:438. 5. Allez M, Tieng V, Nakazawa A, et al. CD41NKG2D1 T cells in Crohn’s disease mediate inflammatory and cytotoxic responses through MICA interactions. Gastroenterology. 2007; 132:2346-2358. http://dx.doi.org/10.1016/j.jaad.2016.07.056
Classic and HIV-related Kaposi sarcoma treated with 0.1% topical timolol gel To the Editor: During the last 12 months, we treated 4 cases of cutaneous Kaposi sarcoma (KS) with 0.1% topical timolol gel (Table I). Of the 4 cases, 3 were classic KS, whereas 1 was HIV-associated KS. In the first 3 cases, 0.1% timolol gel was applied twice a day until resolution. Blood pressure, glucose,
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did not affect serum CXCL10 levels, consistent with observations in our mouse model3 (Supplemental Fig 2; available at http://www.jaad.org). The most common side effects of simvastatin were self-limited myalgia in 4 participants and diarrhea in 2 participants. Three participants had mild, transient transaminitis and 4 had mild creatine phosphokinase elevations, none requiring dose modifications. Only 1 withdrawal, because of vertigo, was thought to be treatment-related. Our study does not support the use of oral simvastatin for the treatment of vitiligo. Disparate findings in this study compared with our mouse model may be a result of dosing limitations in humans because of potential toxicity, which is not a concern in mice. Our failure to show efficacy may also have been influenced by long-standing disease in our participants (responses are best in those with recent onset), small sample size, or lack of sensitive measures of treatment response. The VASI relies on an estimation of affected BSA, with limited sensitivity to change ([4.7% BSA).5 Newer outcome measures may provide more sensitive options for monitoring responses. Simvastatin may induce myopathy at higher doses; however, topical treatment may allow the delivery of sufficiently high local concentrations without systemic toxicity and could be tested in larger studies. The authors thank Dane Netherton for statistical assistance.
Stefan G. Vanderweil, MD, Shinya Amano, MD, PhD, Wei-Che Ko, MS, Jillian M. Richmond, PhD, Michelle Kelley, MSN, RN, Maryanne Makredes Senna, MD, Andrea Pearson, MD, Sandhya Chowdary, MD, Celia Hartigan, RN, MPH, Bruce Barton, PhD, and John E. Harris, MD, PhD Department of Medicine, Division of Dermatology, University of Massachusetts Medical School, Worcester, MA Funding sources: National Institutes of Health (NIH) Clinical and Translational Sciences Award UL1TR000161 through a Pilot Project Grant from the University of Massachusetts Center for Clinical and Translational Sciences; Physician-Scientist Career Development Award from the Dermatology Foundation; National Institute of Arthritis and Musculoskeletal and Skin Diseases, part of the NIH, under Award Number AR061437; and research grants from the Charles H. Hood Foundation, Vitiligo Research Foundation, and Kawaja Family Vitiligo Research Initiative.
Conflicts of interest: None declared. Correspondence to: John E. Harris, MD, PhD, University of Massachusetts Medical School, Lazare Research Bldg, Room 225, 364 Plantation St, Worcester, MA 01605 E-mail: [email protected] REFERENCES 1. Rashighi M, Agarwal P, Richmond JM, et al. CXCL10 is critical for the progression and maintenance of depigmentation in a mouse model of vitiligo. Sci Transl Med. 2014;6(223): 223ra23. 2. Zhao Y, Gartner U, Smith FJ, et al. Statins downregulate K6a promoter activity: a possible therapeutic avenue for pachyonychia congenita. J Invest Dermatol. 2011;131(5): 1045-1052. 3. Agarwal P, Rashighi M, Essien KI, et al. Simvastatin prevents and reverses depigmentation in a mouse model of vitiligo. J Invest Dermatol. 2015;135(4):1080-1088. 4. No€el M, Gagne C, Bergeron J, et al. Positive pleiotropic effects of HMG-CoA reductase inhibitor on vitiligo. Lipids Health Dis. 2004;3:7. 5. Komen L, da Graca V, Wolkerstorfer A, et al. Vitiligo Area Scoring Index and Vitiligo European Task Force assessment: reliable and responsive instruments to measure the degree of depigmentation in vitiligo. Br J Dermatol. 2015; 172(2):437-443. http://dx.doi.org/10.1016/j.jaad.2016.06.015
Increased blood levels of NKG2D1CD41 T cells in patients with alopecia areata To the Editor: Natural killer group 2D (NKG2D) is a key target of not only alopecia areata (AA) but other autoimmune diseases. AA is a TH1-mediated disease driven by interferon- (IFN- ) and IFN- einduced cytokines.1 NKG2D1 cells, including NK, NKT, and CD81 T cells, and NKG2D-activating ligands have a key role in AA pathogenesis. Excessive IFN- secretion by activated NKG2D1 cells can lead to collapse of hair follicle immune privilege.2 It is well known that CD41 T cells highly infiltrate the tissue of the ‘‘bee swarm’’ in AA.3 However, the functional role and expression patterns of NKG2D1CD41 T cell in AA remain largely unknown. In this study, we investigated the changes in blood levels of NKG2D1CD41 T cells in patients with AA. Forty-three patients with AA (18 men and 25 women) and 26 healthy controls (10 men and 16 women) with no scalp lesions in their personal history or on clinical examination were enrolled. The cell surface expression of NKG2D on peripheral blood mononuclear cells was studied by flow cytometry analysis (FACS Aria; BD Biosciences, Franklin Lakes, NJ). Results in each group were expressed as a percentage of CD41 T cells, CD81
152 Letters
J AM ACAD DERMATOL
JANUARY 2017
Fig 1. Results of fluorescence-activated cell sorting (FACS) analysis of peripheral blood and natural killer group 2D (NKG2D)1CD41 T, NKG2D1CD81 T, and NKG2D1CD561 NK cells in control and alopecia areata (AA) patient groups. Analysis of FACS data was carried out dividing into 4 times because of a large number of samples. The left bar graph indicates the average fold change of NKG2D1 cells from the 4 experiments (26 healthy controls and 43 patients with AA). NKG2D1 expressed on CD41 T cells was significantly higher in AA patient groups compared to control groups, as with NKG2D1CD81 T cells and NKG2D1CD561 NK cells. The right dot plot shows the representative results taken from the third experiment (10 healthy controls and 10 patients with AA).
Fig 2. Double-immunofluorescence analysis with antienatural killer group 2D (NKG2D; green) and anti-CD4 (red) antibodies. The merged image clearly shows infiltration of NKG2D1CD41 T cells in the lesional hair follicles of patients with alopecia areata. The two white arrows indicate the location of hair follicle.
J AM ACAD DERMATOL
Letters 153
VOLUME 76, NUMBER 1
T cells, or CD561 NK cells expressing NKG2D. We collected blood samples 4 times, and the FACS analysis was also performed by dividing into 4 times. Therefore, final data were represented as a fold change of each cells expressing NKG2D over the healthy control (control set to 1). The patients were segregated into 2 or 3 subgroups according to AA severity, activity, type, and treatment modalities for subgroup analysis. Statistical analysis was performed using the ManneWhitney U test and KruskaleWallis analysis. Double-labeling immunofluorescence microscopy in 2 representative cases using freshly frozen specimens was also performed. The results showed that the levels of NKG2D1CD81 T cells and NKG2D1CD561 NK cells were higher in the AA group compared with the control group (P \ .05). Interestingly, a 2.49-fold increase in the CD41 T cells subset expressing NKG2D was detected in the peripheral blood from patients with AA compared with healthy controls (P \ .05; Fig 1). Although not statistically significant, the proportion of NKG2D1CD41 T cells was lower in patients who had mild severity AA and in those who received systemic immunosuppressive therapy. Double label immunofluorescence revealed NKG2D1 cells coexpressed CD4 around the hair follicles of the AA lesion (Fig 2). Recently, the role of NKG2D1CD41 T cells in other chronic autoimmune diseases, such as Crohn’s disease and rheumatoid arthritis, was highlighted.4 A subset of CD41 T cells expressing NKG2D was higher in the lamina propria of patients with Crohn’s disease.5 These NKG2D1CD41 T cell clones were functionally active through major histocompatibility complex class I-related chain A (MICA) eNKG2D interactions, producing IFN- and killing targets expressing MICA.5 We hypothesize that NKG2D1CD41 T cells might exhibit a TH1 cytokine profile in patients with AA. Our results suggest that IFN- eproducing NKG2D1CD41 T cells have the potential to become therapeutic targets and biologic markers of disease activity in AA. However, additional studies are needed to characterize the functional properties of NKG2D1CD41 T cells in patients with AA. Yong Hyun Jang, MD, PhD,a Jin Kyeong Choi, PhD,b,c Yun Hwan Jang, MD,a Sun Young Moon, MD,a Weon Ju Lee, MD, PhD,a Seok-Jong Lee, MD, PhD,a Young-Ae Choi, PhD,b Sang-Hyun Kim, PhD,b and Do Won Kim, MD, PhDa Departments of Dermatologya and Pharmacology,b School of Medicine, Kyungpook National University,
Daegu, Korea, and the Molecular Immunology Section,c Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland Drs Jang and J. K. Choi contributed equally to this work. Supported by the Basic Science Research Program through the National Research Foundation of Korea, funded by the Ministry of Education (NRF-2015R1D1A3A01016229 and 2014R1A5A 2009242). The protocol for the study was approved by the institutional review board of the Kyungpook National University Hospital (KNUH 2014-03-003). Conflicts of interest: None declared. Correspondence to: Sang-Hyun Kim, PhD, Department of Pharmacology, Kyungpook National University School of Medicine, 200, Dongduk-ro, Jung-gu, Daegu 700-422, Republic of Korea E-mail: [email protected] Or Do Won Kim, MD, PhD, Department of Dermatology, Kyungpook National University School of Medicine, 200, Dongduk-ro, Jung-gu, Daegu, 41944, Republic of Korea E-mail: [email protected] REFERENCES 1. Gilhar A, Etzioni A, Paus R. Alopecia areata. N Engl J Med. 2012; 366:1515-1525. 2. Paus R, Bertolini M. The role of hair follicle immune privilege collapse in alopecia areata: status and perspectives. J Investig Dermatol Symp Proc. 2013;16:S25-S27. 3. Ito T, Ito N, Saatoff M, et al. Maintenance of hair follicle immune privilege is linked to prevention of NK cell attack. J Invest Dermatol. 2008;128:1196-1206. 4. Killock D. Experimental arthritis: NKG2D: a potential therapeutic target in RA? Nat Rev Rheumatol. 2011;7:438. 5. Allez M, Tieng V, Nakazawa A, et al. CD41NKG2D1 T cells in Crohn’s disease mediate inflammatory and cytotoxic responses through MICA interactions. Gastroenterology. 2007; 132:2346-2358. http://dx.doi.org/10.1016/j.jaad.2016.07.056
Classic and HIV-related Kaposi sarcoma treated with 0.1% topical timolol gel To the Editor: During the last 12 months, we treated 4 cases of cutaneous Kaposi sarcoma (KS) with 0.1% topical timolol gel (Table I). Of the 4 cases, 3 were classic KS, whereas 1 was HIV-associated KS. In the first 3 cases, 0.1% timolol gel was applied twice a day until resolution. Blood pressure, glucose,