- Email: [email protected]
395 Lack of type 2 innate lymphoid cells promotes a type I driven enhanced immune response in TNCB contact hypersensitivity
ABSTRACTS | Inflammation, Immunity and Infection 394
395
The pivotal role of wide local excision for the treatment of severe hidradenitis suppurativa (Hurley grade III) e retrospective analyses of 74 patients C Posch, B Monshi, T Quint, I Vujic, N Lilgenau and K Rappersberger Dermatology, The Rudolfstiftung Hospital, Vienna, Austria Hidradenitis suppurativa (HS) is a painful, chronic, recurrent inflammatory skin disease that affects terminal hair follicles and apocrine glands. It develops in early adolescence, and is confined to axillary and inguinogenital/gluteal regions. This study aimed to measure the impact of surgery on the individual quality of life in severe grade HS patients. Additionally, parameters such as disease duration, previous therapeutic interventions, postoperative complications (pain, infection, scarring, wound healing deficiency, mobility restrictions), postoperative recurrence and satisfaction with the cosmetic results were collected. Data from 74 patients (40 male, 34 female) with HS Hurley grad III treated with wide local excision and secondary wound healing were evaluated. Most patients had inguinogenital/gluteal disease (n¼51, 68.9%, p<0,001). Inguinogenital/gluteal disease was pronounced in female patients (p¼0.009). Involvement of both, axillary and inguinogenital/gluteal areas were pronounced in male patients (p¼0.018). Most patients (n¼53; 71.6%) had a disease history of more than 5 years at the time of initial presentation at our institution. Wide local excision improved the Dermatology Life Quality Index (DLQI) scores from initially 27.89 (range 2-30; SD¼5.3) to 5.31 (range 0-26; SD 7.38; p<0.001) independent of localization (p¼0.195). 47.3% of patients had postoperative complications, most frequently pain and scarring. Local recurrence rates were calculated with 18.9% from follow-up data covering a period of up to 14 years. 70.3% of patients were highly satisfied with the cosmetic results. From our study we conclude that wide local excision of affected skin significantly improves the quality of life of HS Hurley grad III patients. Further, surgery has the potential to locally heal HS areas with high satisfaction with the cosmetic results. The socio-economic footprint of wide local excision, to date, favors surgery over systemic therapies with anti-inflammatory biologics.
Lack of type 2 innate lymphoid cells promotes a type I driven enhanced immune response in TNCB contact hypersensitivity D Rafei-Shamsabadi1, S Kunz1, S Martin1, C Klose2, Y Tanriver2, S Arnold3, A Diefenbach4, T Halim5, A McKenzie5, T Jakob6, Saskia van de Poel1 and Kalolina Ebert1 1 Department of Dermatology, Freiburg, Germany, 2 Institute of Medical Microbiology, Freiburg, Germany, 3 Institute of Experimental and Clinical Pharmacology and Toxicology, Freiburg, Germany, 4 Institute of Medical Microbiology, Mainz, Germany, 5 MRC, Laboratory of Molecular Biology, Cambridge, United Kingdom and 6 Department of Dermatology and Allergy, Gießen, Germany The role of innate lymphoid cells (ILC) in allergic contact dermatitis has not adequately been addressed. We investigated quantitative changes and functional relevance of ILCs during the elicitation phase of hapten-induced contact hypersensitivity (CHS). EomesGfp/+ x Rorc(gt)CreTg x Rosa26RYfp/+ reporter mice were sensitized and challenged with the hapten TNCB. Ear swelling, ILC numbers and cytokine production were measured at different time points. For functional analysis we depleted ILCs in Rag1-/- mice before recieving sorted T-cells from sensitized donor mice (CD90.1) using either a CD90.2-specific or isotype mAb. In addition CHS was performed in Rora+/floxIl7rCre mice, which selectively lack ILC2. The quantitative analysis of total cell numbers revealed early increases of natural killer (NK) cells in skin and skin draining lymph nodes 24h after allergen challenge, corresponding to the highest ear swelling response and leukocyte infiltrate in the skin. These cells produced high amounts of the type 1 cytokines IFNg and TNFa. ILC1, 2 and 3 showed a delayed increase in total numbers starting from 48h after allergen challenge. ILC2 cells displayed an activated phenotype reflected by increased ICOS expression. Total ILC depletion through CD90.2 mAb administration resulted in a significantly enhanced ear swelling response. Finally, Rora+/ flox Il7rCre mice also displayed increased ear swelling responses after allergen challenge. In conclusion, our data support the concept of NK cells as main innate proinflammatory player and suggests that simultaneously activated ILC2 act as counter regulators in the type 1 dominated immune response of CHS.
396
397
Vd1+T-cells are stress-sentinels in human skin and are implicated in alopecia areata pathogenesis Y Uchida1, J Gherardini1, M Alam2, A Keren3, A Arakawa4, A Rossi5, A Gilhar3, T Kanekura6, M Bertolini1 and R Paus7 1 University of Mu¨nster, Mu¨nster, Germany, 2 Monasterium Laboratory, Mu¨nster, Germany, 3 Technion-Israel Institute of Technology, Haifa, Israel, 4 University of Munich, Munich, Germany, 5 University “La Sapienza”, Rome, Italy, 6 Kagoshima University, Kagoshima, Japan and 7 University of Manchester, Manchester, United Kingdom gd T-cells regulate hair follicle (HF) cycling and execute stress surveillance tasks in murine skin. Yet, their role in human skin physiology remains unknown. Therefore, after confirming the presence of Vd1+T-cells in/around human scalp HFs, we investigated how Vd1+NKG2D+CD69- dermal gd T-cells isolated from human scalp skin, impact on human HFs ex vivo. Despite their autologous nature, co-cultured dermal Vd1+T-cells became activated and exhibited marked HF cytotoxicity against “stressed”, but not against “non-stressed” scalp HFs ex vivo (comparable to autologous CD8+T-cells). These gd T-cells induced HF dystrophy, premature catagen, HF overexpression of the stress-induced NKG2D ligand, MICA, and HF immune privilege (IP) collapse. Most of these phenomena were inhibited by co-administering blocking antibodies against INFg, CD1d, or MICA. Thus, human dermal Vd1+T-cells likely recognize “stressed” HFs via CD1d-presented HF self-antigens and MICA binding to NKG2D+ followed by INFg secretion. This promotes HF dystrophy, HF IP collapse and premature catagen entry. Since this chain of events strikingly mimics alopecia areata (AA) pathogenesis, we also investigated the role of gd T-cells in AA. Indeed, dense infiltrates of NKG2D- and INFg-overexpressing Vd1+T-cells were detected in/around lesional HFs from AA patients and Vd1+T-cells were present around affected HFs in experimentally induced AA lesions in human scalp xenotransplanted onto SCID mice. These data suggest that dermal Vd1+T-cells in human skin serve physiological functions as stress-sensing sentinels, which attain pathological significance in AA. Thus, targeting excessive activities of NKG2D+Vd1+Tcells may be a promising, novel adjunct therapy in AA and related autoimmune diseases.
The TRPA1 ion channel mediates inhibitory effects on imiquimod-induced psoriasiform skin inflammation A´ Keme´ny, S Horva´th, R Komlo´di, A Perkecz, C Gyo¨mo¨rei, E Pinte´r and R Gyulai University of Pe´cs, Pe´cs, Hungary Psoriasis is a chronic immune-mediated inflammatory skin disease with genetic susceptibility. Recent studies have suggested that neuro-immune interactions play important role in its pathomechanism. Apart from being expressed on nociceptive primary afferent nerve fibers, transient receptor potential ankyrin 1 (TRPA1) is present on keratinocytes, melanocytes, and fibroblasts in the skin. Activation of TRP ion channels has been shown to contribute to cutaneous inflammatory process. Our aim was to investigate the role of TRPA1 receptor in imiquimod (IMQ)-induced psoriasiform reaction using genetically engineered TRPA1 knock out mice. Aldara (5% IMQ) and control cream (vaseline) was applied simultaneously in Finn chambers on the shaved back skin of TRPA1 knock out (KO) and wild-type (WT) mice. Treatments were repeated for 4 days. Skin edema/infiltration and blood perfusion were measured with a micrometer and with Laser Speckle Contrast Analysis (LASCA) method, respectively. Dorsal skin samples were collected at different time points, and expression levels of IL-1b, TNF-a, IL-17, IL-22 and IL-23 mRNA were determined by RT-PCR. Histopathological alterations were monitored by semiquantitative scoring on haematoxylin-eosin stained skin sections. Dorsal skin swelling/infiltration was significantly increased in IMQtreated TRPA1 KO mice compared to the TRPA1 WT group, and higher elevation of blood flow was detected in the TRPA1 KO group. Markedly higher mRNA expression of TNFa, IL-1b and IL-22 was observed in the TRPA1 KO mice, while IL-17 and IL-23 mRNA expression changed similarly in KO and WT mice. We found significant histopathological differences between vaseline- or IMQ-treated skin samples in the two mouse strains. In summary, the lack of TRPA1 receptors results in increased skin edema/infiltration and a more pronounced Th1like inflammatory reaction in the IMQ induced psoriasiform dermatitis model. Our results suggest an important role of TRPA1 in the regulation of cutaneous immune responses.
398
399
Novel lymphocyte stimulation tests using ex-vivo expanded skin-infiltrating T cells from severe drug eruptions T Fujiyama1, H Hashizume2, T Umayahara1, T Ito1 and Y Tokura1 1 Dermatology, Hamamatsu University School of Medicine, Hamamatsu, Japan and 2 Dermatology, Shimada City Municipal Hospital, Shimada, Japan Stevens-Johnson syndrome (SJS), toxic epidermal necrolysis (TEN) and drug-induced hypersensitivity syndrome/ drug reaction with eosinophilia and systemic symptoms (DIHS/DRESS) are life-threatening adverse drug reactions. Although determination of the causative drug is critical to prevent the relapse, patch test and lymphocyte stimulation test (LST) usually employed are not necessarily satisfactory in the sensitivity. We sought to explore a novel LST using skin-infiltrating T cells, because antigen-specific, pathogenic T cells seem to infiltrate more preferentially in the skin lesions, but not in the peripheral blood of the patients with severe drug eruptions. Skin-infiltrating T cells were expanded from a punch biopsy specimen of lesional skin with anti-CD3/CD28 antibodies and IL-2. More than 107 T cells/specimen were obtained after two-week expansion. PBMCs from the same patient were used for antigen presenting cells. To investigate the drug-induced T-cell proliferation and cytokine production, CFSE and ELISpot assays were employed, respectively. The T cell responses to the drugs were compared with those of PBMCs. The ELISpot assay showed the number of T cells producing IFN-g was significantly increased in the presence of causative drugs, and there was a greater number of IFN-g+ cells in the skin-derived T cells than in PBMCs. The CFSE assay revealed that both CD8+ and CD4+ T cells proliferated in response to causative drugs in cases of SJS/ TEN and DIHS/DRESS. Notably, the majority of drug-reactive CD8+ T cells produced IFN-g in SJS/TEN cases but not in DIHS/DRESS. Only a low number of IL-4+CD4+ T cells proliferated upon drug stimulation in both SJS/TEN and DIHS/DRESS. The responses of CD4+IL-17+ cells to drugs were variable among the cases. These results suggest that the ex-vivo expanded skininfiltrating T cells represent the pathogenic T cells, and they can be used for determination of causative drugs in severe drug eruptions.
S228 Journal of Investigative Dermatology (2016), Volume 136
Innate immunity of keratynocytes in localized scleroderma I Khamaganova1, N Potekaev2, O Svitich3, L Gankovskaya3, E Malyarenko2 and D Guilaliev1 1 Skin Diseases & Cosmetology, Pirogov Russian National Research Medical University, Moscow, Russian Federation, 2 Moscow Scientific & Practical Centre for Dermatovenerology & Cosmetology, Moscow, Russian Federation and 3 Immunology, Pirogov Russian National Research Medical University, Moscow, Russian Federation Localized scleroderma is a refractory autoimmune disease. The complex activation of the innate immune system may enhance fibrotic changes. The objective of the study was the complex analysis of indicators of innate immunity of the keratinocytes in healthy persons and patients with localized scleroderma. 69 patients aged 19- 72 years old & 6 healthy subjects aged 30- 39 years were examined. Total RNA was isolated from keratinocytes by using a RNeasy Mini Kit (Qiagen, Germany) and kit “RIBO-sorb” (ILS, Russia) according to the manufacturer’s instructions. For cDNA synthesis, 2 ml of total RNA was combined with random primers (Syntol, Russia) and reverse primers for the target genes. For quantitative analysis, real-time PCR was performed, using a SYBR Green Kit for qRT-PCR (Syntol, Russia), according to the manufacturer’s instructions. The primers and probes for the real-time PCR were synthesized by Syntol (Russia). The median of the expression of Toll-liked receptor 2 (TLR2) was significantly reduced both in the affected skin to 863 and in healthy skin in scleroderma patients to 969 in comparison with healthy subjects (4366). The median of the expression of human b defensin 1(hBD1)was significantly increased both in the affected skin to 77356 and in healthy skin in scleroderma patients to 71050 in comparison with healthy subjects (20059). The median of the expression of TNFa was significantly increased both in the affected skin to 76 and in in healthy skin in scleroderma patients to 27 in comparison with healthy subjects (20,5). The investigation showed statistically significant changes of the indicators of innate immunity(TLR2, hBD-1, TNFa) in the skin of scleroderma patients. The data can be further used for the elaboration of scleroderma treatment.
395
The pivotal role of wide local excision for the treatment of severe hidradenitis suppurativa (Hurley grade III) e retrospective analyses of 74 patients C Posch, B Monshi, T Quint, I Vujic, N Lilgenau and K Rappersberger Dermatology, The Rudolfstiftung Hospital, Vienna, Austria Hidradenitis suppurativa (HS) is a painful, chronic, recurrent inflammatory skin disease that affects terminal hair follicles and apocrine glands. It develops in early adolescence, and is confined to axillary and inguinogenital/gluteal regions. This study aimed to measure the impact of surgery on the individual quality of life in severe grade HS patients. Additionally, parameters such as disease duration, previous therapeutic interventions, postoperative complications (pain, infection, scarring, wound healing deficiency, mobility restrictions), postoperative recurrence and satisfaction with the cosmetic results were collected. Data from 74 patients (40 male, 34 female) with HS Hurley grad III treated with wide local excision and secondary wound healing were evaluated. Most patients had inguinogenital/gluteal disease (n¼51, 68.9%, p<0,001). Inguinogenital/gluteal disease was pronounced in female patients (p¼0.009). Involvement of both, axillary and inguinogenital/gluteal areas were pronounced in male patients (p¼0.018). Most patients (n¼53; 71.6%) had a disease history of more than 5 years at the time of initial presentation at our institution. Wide local excision improved the Dermatology Life Quality Index (DLQI) scores from initially 27.89 (range 2-30; SD¼5.3) to 5.31 (range 0-26; SD 7.38; p<0.001) independent of localization (p¼0.195). 47.3% of patients had postoperative complications, most frequently pain and scarring. Local recurrence rates were calculated with 18.9% from follow-up data covering a period of up to 14 years. 70.3% of patients were highly satisfied with the cosmetic results. From our study we conclude that wide local excision of affected skin significantly improves the quality of life of HS Hurley grad III patients. Further, surgery has the potential to locally heal HS areas with high satisfaction with the cosmetic results. The socio-economic footprint of wide local excision, to date, favors surgery over systemic therapies with anti-inflammatory biologics.
Lack of type 2 innate lymphoid cells promotes a type I driven enhanced immune response in TNCB contact hypersensitivity D Rafei-Shamsabadi1, S Kunz1, S Martin1, C Klose2, Y Tanriver2, S Arnold3, A Diefenbach4, T Halim5, A McKenzie5, T Jakob6, Saskia van de Poel1 and Kalolina Ebert1 1 Department of Dermatology, Freiburg, Germany, 2 Institute of Medical Microbiology, Freiburg, Germany, 3 Institute of Experimental and Clinical Pharmacology and Toxicology, Freiburg, Germany, 4 Institute of Medical Microbiology, Mainz, Germany, 5 MRC, Laboratory of Molecular Biology, Cambridge, United Kingdom and 6 Department of Dermatology and Allergy, Gießen, Germany The role of innate lymphoid cells (ILC) in allergic contact dermatitis has not adequately been addressed. We investigated quantitative changes and functional relevance of ILCs during the elicitation phase of hapten-induced contact hypersensitivity (CHS). EomesGfp/+ x Rorc(gt)CreTg x Rosa26RYfp/+ reporter mice were sensitized and challenged with the hapten TNCB. Ear swelling, ILC numbers and cytokine production were measured at different time points. For functional analysis we depleted ILCs in Rag1-/- mice before recieving sorted T-cells from sensitized donor mice (CD90.1) using either a CD90.2-specific or isotype mAb. In addition CHS was performed in Rora+/floxIl7rCre mice, which selectively lack ILC2. The quantitative analysis of total cell numbers revealed early increases of natural killer (NK) cells in skin and skin draining lymph nodes 24h after allergen challenge, corresponding to the highest ear swelling response and leukocyte infiltrate in the skin. These cells produced high amounts of the type 1 cytokines IFNg and TNFa. ILC1, 2 and 3 showed a delayed increase in total numbers starting from 48h after allergen challenge. ILC2 cells displayed an activated phenotype reflected by increased ICOS expression. Total ILC depletion through CD90.2 mAb administration resulted in a significantly enhanced ear swelling response. Finally, Rora+/ flox Il7rCre mice also displayed increased ear swelling responses after allergen challenge. In conclusion, our data support the concept of NK cells as main innate proinflammatory player and suggests that simultaneously activated ILC2 act as counter regulators in the type 1 dominated immune response of CHS.
396
397
Vd1+T-cells are stress-sentinels in human skin and are implicated in alopecia areata pathogenesis Y Uchida1, J Gherardini1, M Alam2, A Keren3, A Arakawa4, A Rossi5, A Gilhar3, T Kanekura6, M Bertolini1 and R Paus7 1 University of Mu¨nster, Mu¨nster, Germany, 2 Monasterium Laboratory, Mu¨nster, Germany, 3 Technion-Israel Institute of Technology, Haifa, Israel, 4 University of Munich, Munich, Germany, 5 University “La Sapienza”, Rome, Italy, 6 Kagoshima University, Kagoshima, Japan and 7 University of Manchester, Manchester, United Kingdom gd T-cells regulate hair follicle (HF) cycling and execute stress surveillance tasks in murine skin. Yet, their role in human skin physiology remains unknown. Therefore, after confirming the presence of Vd1+T-cells in/around human scalp HFs, we investigated how Vd1+NKG2D+CD69- dermal gd T-cells isolated from human scalp skin, impact on human HFs ex vivo. Despite their autologous nature, co-cultured dermal Vd1+T-cells became activated and exhibited marked HF cytotoxicity against “stressed”, but not against “non-stressed” scalp HFs ex vivo (comparable to autologous CD8+T-cells). These gd T-cells induced HF dystrophy, premature catagen, HF overexpression of the stress-induced NKG2D ligand, MICA, and HF immune privilege (IP) collapse. Most of these phenomena were inhibited by co-administering blocking antibodies against INFg, CD1d, or MICA. Thus, human dermal Vd1+T-cells likely recognize “stressed” HFs via CD1d-presented HF self-antigens and MICA binding to NKG2D+ followed by INFg secretion. This promotes HF dystrophy, HF IP collapse and premature catagen entry. Since this chain of events strikingly mimics alopecia areata (AA) pathogenesis, we also investigated the role of gd T-cells in AA. Indeed, dense infiltrates of NKG2D- and INFg-overexpressing Vd1+T-cells were detected in/around lesional HFs from AA patients and Vd1+T-cells were present around affected HFs in experimentally induced AA lesions in human scalp xenotransplanted onto SCID mice. These data suggest that dermal Vd1+T-cells in human skin serve physiological functions as stress-sensing sentinels, which attain pathological significance in AA. Thus, targeting excessive activities of NKG2D+Vd1+Tcells may be a promising, novel adjunct therapy in AA and related autoimmune diseases.
The TRPA1 ion channel mediates inhibitory effects on imiquimod-induced psoriasiform skin inflammation A´ Keme´ny, S Horva´th, R Komlo´di, A Perkecz, C Gyo¨mo¨rei, E Pinte´r and R Gyulai University of Pe´cs, Pe´cs, Hungary Psoriasis is a chronic immune-mediated inflammatory skin disease with genetic susceptibility. Recent studies have suggested that neuro-immune interactions play important role in its pathomechanism. Apart from being expressed on nociceptive primary afferent nerve fibers, transient receptor potential ankyrin 1 (TRPA1) is present on keratinocytes, melanocytes, and fibroblasts in the skin. Activation of TRP ion channels has been shown to contribute to cutaneous inflammatory process. Our aim was to investigate the role of TRPA1 receptor in imiquimod (IMQ)-induced psoriasiform reaction using genetically engineered TRPA1 knock out mice. Aldara (5% IMQ) and control cream (vaseline) was applied simultaneously in Finn chambers on the shaved back skin of TRPA1 knock out (KO) and wild-type (WT) mice. Treatments were repeated for 4 days. Skin edema/infiltration and blood perfusion were measured with a micrometer and with Laser Speckle Contrast Analysis (LASCA) method, respectively. Dorsal skin samples were collected at different time points, and expression levels of IL-1b, TNF-a, IL-17, IL-22 and IL-23 mRNA were determined by RT-PCR. Histopathological alterations were monitored by semiquantitative scoring on haematoxylin-eosin stained skin sections. Dorsal skin swelling/infiltration was significantly increased in IMQtreated TRPA1 KO mice compared to the TRPA1 WT group, and higher elevation of blood flow was detected in the TRPA1 KO group. Markedly higher mRNA expression of TNFa, IL-1b and IL-22 was observed in the TRPA1 KO mice, while IL-17 and IL-23 mRNA expression changed similarly in KO and WT mice. We found significant histopathological differences between vaseline- or IMQ-treated skin samples in the two mouse strains. In summary, the lack of TRPA1 receptors results in increased skin edema/infiltration and a more pronounced Th1like inflammatory reaction in the IMQ induced psoriasiform dermatitis model. Our results suggest an important role of TRPA1 in the regulation of cutaneous immune responses.
398
399
Novel lymphocyte stimulation tests using ex-vivo expanded skin-infiltrating T cells from severe drug eruptions T Fujiyama1, H Hashizume2, T Umayahara1, T Ito1 and Y Tokura1 1 Dermatology, Hamamatsu University School of Medicine, Hamamatsu, Japan and 2 Dermatology, Shimada City Municipal Hospital, Shimada, Japan Stevens-Johnson syndrome (SJS), toxic epidermal necrolysis (TEN) and drug-induced hypersensitivity syndrome/ drug reaction with eosinophilia and systemic symptoms (DIHS/DRESS) are life-threatening adverse drug reactions. Although determination of the causative drug is critical to prevent the relapse, patch test and lymphocyte stimulation test (LST) usually employed are not necessarily satisfactory in the sensitivity. We sought to explore a novel LST using skin-infiltrating T cells, because antigen-specific, pathogenic T cells seem to infiltrate more preferentially in the skin lesions, but not in the peripheral blood of the patients with severe drug eruptions. Skin-infiltrating T cells were expanded from a punch biopsy specimen of lesional skin with anti-CD3/CD28 antibodies and IL-2. More than 107 T cells/specimen were obtained after two-week expansion. PBMCs from the same patient were used for antigen presenting cells. To investigate the drug-induced T-cell proliferation and cytokine production, CFSE and ELISpot assays were employed, respectively. The T cell responses to the drugs were compared with those of PBMCs. The ELISpot assay showed the number of T cells producing IFN-g was significantly increased in the presence of causative drugs, and there was a greater number of IFN-g+ cells in the skin-derived T cells than in PBMCs. The CFSE assay revealed that both CD8+ and CD4+ T cells proliferated in response to causative drugs in cases of SJS/ TEN and DIHS/DRESS. Notably, the majority of drug-reactive CD8+ T cells produced IFN-g in SJS/TEN cases but not in DIHS/DRESS. Only a low number of IL-4+CD4+ T cells proliferated upon drug stimulation in both SJS/TEN and DIHS/DRESS. The responses of CD4+IL-17+ cells to drugs were variable among the cases. These results suggest that the ex-vivo expanded skininfiltrating T cells represent the pathogenic T cells, and they can be used for determination of causative drugs in severe drug eruptions.
S228 Journal of Investigative Dermatology (2016), Volume 136
Innate immunity of keratynocytes in localized scleroderma I Khamaganova1, N Potekaev2, O Svitich3, L Gankovskaya3, E Malyarenko2 and D Guilaliev1 1 Skin Diseases & Cosmetology, Pirogov Russian National Research Medical University, Moscow, Russian Federation, 2 Moscow Scientific & Practical Centre for Dermatovenerology & Cosmetology, Moscow, Russian Federation and 3 Immunology, Pirogov Russian National Research Medical University, Moscow, Russian Federation Localized scleroderma is a refractory autoimmune disease. The complex activation of the innate immune system may enhance fibrotic changes. The objective of the study was the complex analysis of indicators of innate immunity of the keratinocytes in healthy persons and patients with localized scleroderma. 69 patients aged 19- 72 years old & 6 healthy subjects aged 30- 39 years were examined. Total RNA was isolated from keratinocytes by using a RNeasy Mini Kit (Qiagen, Germany) and kit “RIBO-sorb” (ILS, Russia) according to the manufacturer’s instructions. For cDNA synthesis, 2 ml of total RNA was combined with random primers (Syntol, Russia) and reverse primers for the target genes. For quantitative analysis, real-time PCR was performed, using a SYBR Green Kit for qRT-PCR (Syntol, Russia), according to the manufacturer’s instructions. The primers and probes for the real-time PCR were synthesized by Syntol (Russia). The median of the expression of Toll-liked receptor 2 (TLR2) was significantly reduced both in the affected skin to 863 and in healthy skin in scleroderma patients to 969 in comparison with healthy subjects (4366). The median of the expression of human b defensin 1(hBD1)was significantly increased both in the affected skin to 77356 and in healthy skin in scleroderma patients to 71050 in comparison with healthy subjects (20059). The median of the expression of TNFa was significantly increased both in the affected skin to 76 and in in healthy skin in scleroderma patients to 27 in comparison with healthy subjects (20,5). The investigation showed statistically significant changes of the indicators of innate immunity(TLR2, hBD-1, TNFa) in the skin of scleroderma patients. The data can be further used for the elaboration of scleroderma treatment.