The role of CD4 and CD8 cytotoxic T lymphocytes in the formation of viral vesicles

BACKGROUND: Herpetic vesicles caused by herpes simplex virus and varicella zoster virus, and hydroa vacciniforme (HV) are characterized by umbilicated vesicule formation. OBJECTIVES: To understand the histogenesis of umbilicated vesicles in herpetic vesicles and HV, we demonstrated the presence of the virus-associated molecules in the lesions, and the pathogenic role of cytotoxic T-lymphocyte (CTL) immune responses. METHODS: Phenotyping of infiltrating cells was carried out in biopsy specimens from herpes simplex, varicella, herpes zoster and HV, and compared with nonviral contact dermatitis. Viral antigens and Epstein-Barr virus-encoded small nuclear RNA (EBER) were detected by immunostaining and by in situ hybridization, respectively. Infiltrating CTLs expressing granzyme B and granulysin were determined by double immunostaining using confocal laser scanning microscopy. RESULTS: In all herpetic vesicles, the corresponding viral antigens were observed in the cytopathic keratinocytes, and infiltration of lymphoid cells was present in the upper dermis and around the vessels. In all HV lesions studied, EBER+ T cells made up 5-10% of the dermal infiltrates and the dermal infiltrates contained almost no CD56 cells. CTLs expressing granzyme B and granulysin were present in both herpetic and HV lesions, in which they made up 10-30% of the total dermal infiltrates, whereas they comprised less than 5% of the infiltrates of biopsy specimens from nonviral contact dermatitis. Confocal laser microscopic examination demonstrated that both CD4+ and CD8+ T cells expressed granzyme B and granulysin. CONCLUSIONS: CD4+ and/or CD8+ CTLs reactive to the virus-infected cells might be responsible for the histogenesis of herpetic and HV lesions characterized by umbilicated vesicles.     

Histological increase in inflammatory infiltrate in sun-exposed skin of female subjects: the possible involvement of matrix metalloproteinase-1 produced by inflammatory infiltrate on collagen degradation

To investigate morphological changes occurring during cutaneous photoageing, a correlation between the number of infiltrating cells in the dermis and the degree of collagen damage was examined using sections from clinically normal chronically sun-exposed and sun-protected skin of Japanese female subjects. Haematoxylin and eosin-stained sections from 134 sun-exposed (subjects aged 3-82 years) and 73 sun-protected (subjects aged 1-86 years) areas demonstrated a predominant lymphoid cell and to a lesser extent histiocyte infiltration. The mean +/- SD number of lymphoid cells and histiocytes in the sun-exposed skin sections (427.0+/-192.2 and 147.8+/-83.3 cells/mm2, respectively) was significantly higher than in the sun-protected skin sections (292.6+/-98.3 and 125.9+/-59.0 cells/mm2, respectively) (P < 0.001 and P < 0.05, respectively), and the number of lymphoid cells in the sun-exposed skin sections increased significantly with age up to 50 years (r = 0.400, P < 0.001). Sun-exposed skin sections with severe collagen degeneration had a significantly higher number of lymphoid cells than those with slightly degenerated collagen (mean 626.3 vs. 482.4 cells/mm2, P < 0.01). The mean count of mast cells in sun-exposed skin was 202.0 cells/mm2; this did not vary with the age of the subjects or the level of collagen damage. Immunohistochemical studies using 24 frozen sections identified most of the lymphoid cells infiltrating sun-exposed skin as memory T lymphocytes (CD3+, CD4+ and CD45RO+). The number of cells which displayed immunoreactivity to matrix metalloproteinase (MMP)-1 in the sun-exposed skin sections was significantly higher than in the sun-protected skin sections (mean 170.2 vs. 113.6 cells/mm2, P < 0.05). Among these cells were observed CD3 and MMP-1 double-stained T lymphocytes, and T lymphocytes contacting MMP-1-positive cells. These morphological observations suggest that T lymphocytes infiltrating photodamaged skin may play a part in the degeneration and reduction of collagen through MMP-1 activity.     

Immunological activity of photodynamic therapy for genital warts

Background An increasing body of evidence supports the usefulness of photodynamic therapy (PDT) in the treatment of non‐neoplastic pathological conditions, including genital warts. In particular, PDT has demonstrated good clinical cure rates and low recurrence, and is now suggested as a safe alternative means of treating condylomata. Objective To confirm the suitability of aminolaevulinic acid (ALA)‐PDT for the treatment of this condition and to investigate the recruitment and significance of immune cells in lesional areas by immunohistochemical analysis at different time intervals after treatment. Methods Fifteen subjects with histologically proven, recalcitrant condylomata acuminata of the penis, urethra, vulva or perianal area underwent several cycles of PDT following ALA application. Biopsies were taken at baseline and at different intervals during the trial, and infiltrating immune cells, CD3, CD4, CD8, CD1a and CD68, were evaluated by double immunocytochemical alkaline phosphatase antialkaline phosphatase (APAAP) staining. Results Our trial provided a complete cure rate of nine of 15 subjects after five PDT sessions. Perianal lesions showed a particularly rapid remission. While progressing towards total lesion clearance, the immunohistochemical pattern was dominated by dense CD4+ T lymphocytes infiltrating the superficial dermis, accompanied by an accumulation of Langerhans cells. Simultaneously, CD8 began to increase in the lesions of responding patients, and Langerhans cells seemed to migrate towards the dermis. CD68+ macrophages apparently did not participate in the immune inflammatory response. Conclusions This study, to the best of our knowledge, represents the first attempt to clarify the effect of ALA‐PDT on infiltrating immune cells in condylomata acuminata. Our results appear to confirm previously reported clinical data, suggesting that rapid activation of specific immunity in lesional skin, CD4+ T lymphocytes and dendritic cells could be responsible for healing.     

Transfusion-associated graft-versus-host disease: an in situ hybridization analysis of the infiltrating donor-derived cells in the cutaneous lesion.

BACKGROUND: Acute graft-versus-host disease (GVHD) can occur after a blood transfusion. OBJECTIVE: In order to elucidate the pathomechanisms responsible for transfusion-associated GVHD, infiltrating donor-derived cells in a cutaneous lesion were analyzed. METHODS: A skin sample obtained from a 69-year-old woman who developed fatal GVHD after blood transfusions from male donors was studied by performing in situ hybridization (ISH) with a Y-chromosome-specific probe. RESULTS: The cell infiltrates comprised mainly CD3+ T lymphocytes. Immunohistochemistry and ISH in combination demonstrated that 99% (182/184) of the Y-body-positive cells were CD3+. Y bodies were observed in 80% of the CD8+ cells in the epidermis and dermoepidermal junction and in 77 and 45% of the CD8+ and CD4+ cells, respectively, in the dermis. CONCLUSION: These findings suggest that both CD4+ and CD8+ cells of donor origin were involved in the development of cutaneous GVHD.     

固定性药疹急性期免疫组织化学研究

本文用４种鼠抗人白细胞抗原的单克隆抗体，采用标记的链霉亲合素生物素技术对９例固定性药疹患者急性期皮损内浸润的Ｔ细胞及其亚群、Ｂ细胞进行标记。观察到损害内真皮中上部血管周转浸润的细胞中６０％－８０％为Ｔ细胞，其中ＣＤ４＾＋细胞和ＣＤ８＾＋细胞数量大致相等。     

Photoageing shows histological features of chronic skin inflammation without clinical and molecular abnormalities

BACKGROUND: Photodamage is characterized by degradation of collagen and accumulation of abnormal elastin in the superficial dermis. Mast cells and macrophages, which are found in higher numbers in photoaged skin, have been implicated in this process. OBJECTIVES: To analyse the phenotype of haematopoietic-derived infiltrating cells in photodamaged skin. METHODS: Chronically sun-exposed (preauricular) and control sun-protected (postauricular) skin was recovered from eight healthy subjects undergoing cosmetic surgery (facial lifting). RESULTS: Histological analysis showed that sun-exposed skin harboured more infiltrating mononuclear cells than sun-protected skin. Cellular infiltrates were found at the periphery of areas of elastolysis around hair follicles in sun-exposed sites, whereas they were found in the interfollicular dermis around blood vessels and around hair follicles in sun-protected samples. Immunohistochemical analysis revealed an increased number of mast cells, macrophages and CD4+ CD45RO+ T cells in sun-exposed dermis as well as a higher number of CD1a+ dendritic cells in sun-exposed epidermis, compared with the sun-protected samples. Thus photoageing displays histological features of chronic skin inflammation. However, no molecular sign of inflammation was observed and we even found a decreased expression of interleukin-1beta mRNA in sun-exposed compared with sun-protected sites. Furthermore, the patients' skin looked normal and did not display any clinical inflammation. CONCLUSIONS: Collectively, these data show that chronic ultraviolet irradiation induces alterations of innate immune cells which are recruited in sun-exposed skin without being activated.     

Granulysin-producing cytotoxic T cells in the mucocutaneous lesions of Behçet disease: a distinct inflammatory response from erythema nodosum

Background. Cytotoxic T lymphocytes (CTLs) have been recognized as an important effector cell in Behçet disease (BD). Granulysin is a cytolytic granule protein expressed by CTLs and natural killer cells. Aim. To evaluate the involvement of granulysin‐producing T cells in the pathogenesis of BD. Methods. Using immunohistochemistry, lymphocyte subsets expressing granulysin were investigated in mucocutaneous lesions of BD. Serum granulysin levels were assayed by ELISA. Results. Granulysin‐positive cells were seen in specimens from oral ulcers, genital ulcers and acne‐like eruptions, but not erythema nodosum‐like lesions. Both CD4+ and CD8+ T cells expressed granulysin. Serum granulysin levels did not correlate with disease activity in BD. Conclusion. Immune reactions mediated by granulysin‐positive CTLs may play an important role in the pathogenesis of acne‐like eruptions, oral ulcers and genital ulcers in BD.     

Proliferative activity of CD8(+) T cells as an important clue to analyze T cell-mediated inflammatory dermatoses

Abstract To elucidate the pathogenesis of T cell-mediated inflammatory skin diseases, we examined the exact sites where CD8(+) T cells proliferate, correlating them with the localization of antigen-presenting dendritic cells. We performed CD8/Ki-67 double immunohistochemical staining and single staining for CD1a, CD68, and factor XIIIa on sections of paraffin-embedded tissue samples of inflammatory dermatoses in which T lymphocytes are thought to play a crucial role. The dermatoses were lichen planus (12 samples), acute graft-versus host disease (GVHD) (12 samples), chronic GVHD (10 samples), spongiotic dermatitis (8 samples) and psoriasis (7 samples). Labelling for Ki-67 among CD8(+) T cells was predominantly observed in the subepidermal lymphoid infiltrate, and was scanty in the epidermis. This suggested that proliferation of CD8(+) T cells occurred preferentially in the dermis. The labelling index for Ki-67 among dermal and epidermal CD8(+) cells was quite different among the different diseases studied (P < 0.05). They were rich in the subepidermal portion of the dermis of spongiotic dermatitis, acute GVHD and chronic GVHD, but rare in the dermis of psoriasis and lichen planus. A moderate infiltrate was also observed in lesional epidermis of spongiotic dermatitis, acute GVHD and chronic GVHD, whereas they was almost none in the epidermis of psoriasis and lichen planus. CD1a(+) dermal dendritic cells were densely distributed within the lymphoid infiltrate in the affected dermis of spongiotic dermatitis, psoriasis and lichen planus, whereas they were minimal in GVHD. These dermal dendritic cells are candidates as stimulators on T cells in the dermis. In conclusion, the proliferative status of T cells could be an important clue in the elucidation of the pathophysiology of T cell-mediated inflammatory dermatoses. Received: 13 December 2000 / Revised: 24 April 2001 / Accepted: 11 July 2001     

New insights into disease pathogenesis in crusted (Norwegian) scabies: the skin immune response in crusted scabies

Background Crusted scabies is a rare and severely debilitating disease characterized by infestation of the skin with up to millions of Sarcoptes scabiei mites, high total IgG levels, extremely high total IgE levels, and the development of hyperkeratotic skin crusts that may be loose, scaly and flaky or thick and adherent. Objectives To describe crusted scabies skin pathogenesis and identify markers associated with an inappropriate immune response leading to disease progression. Patients/methods Serial sections from skin biopsies obtained from two patients with severe crusted scabies were examined by immunohistochemistry for cell surface markers and inflammatory and regulatory cytokines. Concurrent levels of total B‐ and T‐cell subsets and IgE, IgA, IgM, IgG and IgG subclasses were analysed in the blood. In addition antibody levels were recorded in a further 33 patients with crusted scabies and 14 patients with ordinary scabies. Results A predomination of infiltrating CD8+ T lymphocytes in the dermis was observed compared with minimal helper T lymphocytes (CD4+) and the absence of any B cells. The proportion of T and B lymphocytes and T‐cell subsets in the blood of these patients were within normal ranges, indicating a selective movement of CD8+ T cells into the dermis. Furthermore, strong staining for the inflammatory cytokine interleukin‐1β and anti‐inflammatory cytokine transforming growth factor‐β1 was observed. Elevated levels of IgE, IgG, IgG1, IgG3 and IgG4 were recorded. Conclusions Skin‐homing cytotoxic T cells contribute to an imbalanced inflammatory response in the dermis of crusted scabies lesional skin. This, in combination with the lack of B cells, is contributing to the failure of the skin immune system to mount an effective response resulting in uncontrolled growth of the parasite.     

CD103+T细胞在银屑病皮损中的表达

目的：明确银屑病患者皮肤CD103+T细胞的表达及其与银屑病严重程度的关系。方法：免疫组化检测29例银屑病患者皮损和非皮损皮肤及6名健康对照皮肤中表皮及真皮CD103+T细胞的表达。计算银屑病患者PASI值。结果：CD103+T细胞主要在真皮表达。银屑病患者皮损和非皮损真皮中每个高倍视野CD103+T细胞百分率分别为（26.06%±11.72）%和（12.82±4.5）%（P<0.05）;健康人对照皮肤真皮内CD103+T细胞百分率为（7.47±1.3）%,明显低于银屑病非皮损区（P<0.05）。银屑病患者皮损中,CD103+T的表达与PASI值正相关（P<0.05）。 结论：真皮中CD103+T细胞可能与银屑病的发病及严重程度有关。     

丘疹性蕈样肉芽肿一例

患者男,27岁,因左胸部、左腋窝丘疹3年就诊,无自觉症状,皮损持续不退。组织病理检查示棘层轻度增生肥厚,皮突延长,真皮浅层苔藓样淋巴样细胞浸润,部分移入表皮,形成Pautrier微脓肿。免疫组化示LCA（+）、CD45RO（+）、CD3（+）、CD4（+）、CD8散在（+）、CD20（-）、CD68（-）,CD30（-）。皮肤激光共聚焦显微镜显示棘层中散在有轻度折光的椭圆或圆形细胞影,直径4 ~ 8 μm。诊断：丘疹性蕈样肉芽肿。治疗：外用丙酸氟替卡松乳膏、异维A酸凝胶,窄波紫外线照射,丘疹变软,色泽变淡。     

Mycosis fungoides presenting as pigmented purpuric eruption

A case of pigmented purpuric eruptions evolving to mycosis fungoides during the 4-year follow-up period is described. Clinical manifestation was characterized by petechial lesions with irregular shaped, diffusely pigmented plaques partly sharing morphological similarities with chronic pigmented purpura. Histologically, lymphocytes infiltrated around the capillaries of the superficial dermis with extravasated erythrocytes as well as into the epidermis to form Pautrier microabscesses. Whereas CD4+ cells were observed in the epidermis and upper dermis, CD8+ cells tended to be distributed around the capillaries. Notably, the Rumpel-Leede test revealed extensive punctuate purpura limited to the lesional skin. The aggregation response of platelets was not impaired. Either CD4+ tumor lymphocytes or CD8+ reactive lymphocytes appeared to induce capillary damage resulting in the formation of petechial lesions. Pigmented purpuric eruptions, such as atypical chronic pigmented purpura, is thus an important initial clinical manifestation of mycosis fungoides.     

In situ localization of CD83-positive dendritic cells in psoriatic lesions

BACKGROUND: Dendritic cells (DC) are considered to be the most potent antigen-presenting cells, and CD83 is expressed at a high level on immune-competent, activated and mature DC. Although changes in the number or localization of mature and activated CD83+ DC could be expected in psoriasis, there is little information on such changes. AIM: Morphological identification of CD83+ DC in psoriatic skin lesions. MATERIALS AND METHODS: Immunohistochemical staining was performed in 5 specimens of psoriasis vulgaris and 6 specimens of pustular psoriasis. Formalin-fixed, paraffin-embedded sections were used for examination in this study. The skin sections were pretreated with 0.1% trypsin for 60 min at 37 degrees C prior to immunostaining for CD83. RESULTS: A small but significant subpopulation of CD83+ DC was found in the upper dermis. In addition, CD83+ DC were occasionally scattered in the epidermis. The most common distribution pattern of CD83+ DC was as clusters with mononuclear lymphoid cells in the upper dermis. CD83+ DC were in close contact with lymphocytes. High-intensity staining of CD83 antigens was detected not only on the surface, but also in the cytoplasm of DC. CONCLUSION: These results indicate that activated and mature CD83+ DC may play a role in the immune response in psoriasis and provide in vivo support for the concept that CD83+ DC provide signals for direct intralesional T cell activation.     

Primary cutaneous CD4 positive small/medium T‐cell lymphoma with high proliferation index and CD30‐positive large lymphoid cells

Primary cutaneous CD4 positive small/medium pleomorphic T‐cell lymphoma (SMPTCL) represents a provisional subtype of primary cutaneous T‐cell lymphoma with indolent clinical course. A few aggressive fatal cases with increased proliferation rate and few infiltrating CD8 positive T‐cells have been reported. We describe a case of SMPTCL with an increased proliferation rate, admixed CD30‐positive large lymphoid cells, and few infiltrating CD8 positive T‐cells. The lymphoma cells were positive for CD3, CD4, CD2 and CD5, and negative for CD8. A subset of the lymphoma cells was positive for follicular helper T‐cell markers bcl‐6 and PD‐1. There were approximately 20% CD30‐positive large lymphoid cells, and Ki‐67 showed a moderately high proliferation rate (～40%), mostly in the large lymphoid cells. CD8 infiltrating T‐cells were few (<5%). The patient had an indolent disease with complete response to radiation therapy. To the best of our knowledge, this is the first reported case of SMPTCL with an increased proliferation rate and large CD30+ cells that followed an indolent clinical course.     

儿童药物超敏反应综合征一例

患儿女,10岁9个月,静脉滴注磷霉素后出现全身皮疹,伴发热、浅表淋巴结肿大18d.皮肤科检查:全身皮肤弥漫肿胀性潮红斑,表面覆盖较多灰白色糠皮状黏着性鳞屑.实验室检查:丙氨酸转氨酶、天冬氨酸转氨酶及嗜酸性粒细胞显著升高.皮损组织病理:真皮浅层及皮肤附属器周围散在淋巴细胞浸润.免疫组化:浸润的淋巴细胞以T淋巴细胞为主,未见异形细胞.诊断:药物超敏反应综合征.治疗:予糖皮质激素联合人免疫球蛋白静脉注射及口服环孢素后获得良好疗效.     

Death ligand TRAIL, secreted by CD1a+ and CD14+ cells in blister fluids, is involved in killing keratinocytes in toxic epidermal necrolysis

Toxic epidermal necrolysis (TEN) is characterized by an acute detachment and destruction of keratinocytes, affecting large areas of the skin. It is often related to adverse drug reactions. Previous studies have shown that effector CD8+ T cells, which accumulate in the blister fluid, are functionally cytotoxic and act through a classical perforin/granzyme B pathway. It has recently been shown that these cytotoxic T cells also secrete granulysin peptide, which is lethal to keratinocytes. These cytotoxic T cells exert their killer activity against autologous keratinocytes in the presence of the drug. However, they are unlikely to be the only effectors of TEN. We therefore searched for soluble death factors in the blister fluids that might kill keratinocytes. We found that the amounts of interferon-γ, TRAIL and TNF-α proteins were significantly greater in TEN blister fluids than in all controls (normal sera, TEN sera, burns and Eosinophilic pustular folliculitis blister fluids) and TNF-like weak inducer of apoptosis (TWEAK) amounts are also greater in all controls except burns. We showed that these proteins acted in synergy to induce the death of keratinocytes in vitro. We also found that TRAIL and TWEAK were secreted by CD1a+ and CD14+ cells present in the blister fluids. Thus, in addition to MHC class I-restricted cytotoxic T lymphocytes (CTLs), which lyse keratinocytes, ligands secreted by non-lymphoid cells capable of inducing keratinocyte death in an MHC class I-independent manner, also seem to be present in the blister fluids of patients with TEN.     

Learning from leprosy: insights into contemporary immunology from an ancient disease.

Leprosy provides an ideal model to study immune responses in humans and in skin. Learning from leprosy, we have gained insight into mechanisms of host resistance and susceptibility to infection. New paradigms include the role of Th1/Th2 cytokines, the ability of CD1 to present nonpeptide antigens to T cells, the ability of microbial lipoproteins to stimulate antimicrobial activity in monocytes and the demonstration that T cells can mediate a direct antimicrobial activity through release of granulysin. Together, these findings provide a rationale for developing new strategies to treat and prevent infectious disease.     

颗粒溶素在皮肤病中的研究进展

颗粒溶素是一种溶细胞和促炎症分子,表达于活化的人类细胞毒T细胞(CTLs)和自然杀伤(NK)细胞,参与Stevens-Johnson综合征/中毒性坏死性表皮松解症(SJS/TEN)、银屑病、毛囊炎、扁平苔藓和病毒性水疱等多种皮肤病的发生发展,尤其在SJS/TEN发病过程中是引起角质形成细胞凋亡的重要诱导物。本文就颗粒溶素在皮肤病早期诊断、病情监测及治疗中的应用作一综述。