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Hyperkeratotic erythema multiforme variant in 17 dogs
17例犬的角化过度型EM
Frane Banovic | Thierry Olivry | Barbara Artlet | Emily Rothstein | Luc Beco | Monika Linek | Sonja Zabel | Jeanine Peters-Kennedy | Monika Welle | Rebecca Wilkes | Michaela Austel | Keith Linder
翻译：王帆
Abstract
Background: A new canine subgroup defined as ‘old-dog’ or ‘hyperkeratotic’ erythema multiforme (HKEM) with marked hyperkeratosis and parakeratosis has been proposed without any detailed description of larger case series.
Objectives: We report herein the signalment, clinical signs, treatment outcome, and histopathological and immunological findings in 17 dogs with HKEM.
Animals: Inclusion criteria were the presence of (i) scaly skin lesions with or without crusting; and (ii) microscopic lesions typical of EM (i.e. a panepidermal cytotoxic lymphocytic dermatitis with or without basal keratinocyte apoptosis); and (iii) microscopic ortho- and/or parakeratotic hyperkeratosis affecting the interfollicular epidermis.
Materials and Methods: Clinical questionnaires and skin biopsies were reviewed. Polymerase chain reactions for epidermotropic viruses and direct immunofluorescence were performed.
Results: Various breeds were affected with an over-representation of males in their mid-to-late adulthood (median age 9years). Generalised skin lesions included multifocal-to-coalescing, linear and annular macules and plaques with erythema and adherent firm crusting. Microscopic lesions were specific for EM and featured prominent superficial epidermal apoptosis with lymphocytic satellitosis and parakeratosis. No drug triggers were identified. Polymerase chain reactions for canine herpesvirus polymerase gene, canine parvovirus and canine distemper virus were negative in all HKEM and canine erosive EM (15 dogs) biopsies. Lesions failed to respond to oral and/or topical antimicrobials. Complete remission of signs was achieved in 9 of 17 dogs (53%) using immunosuppressive regimens.
Conclusions and Clinical Relevance: Hyperkeratotic erythema multiforme (HKEM) is a chronic, persistent and clinically distinctive erythema multiforme (EM) variant that differs from ‘classic’ vesiculobullous erosive-to-ulcerative EM in dogs.
KEYWORDS dog, erythema multiforme, hyperkeratotic
摘要
背景: 提出了定义为“老年犬”和“角化过度”多形红斑 (HKEM) 伴明显角化过度和角化不全的新亚组，未对更大的犬病例系列进行任何详细描述。
目的: 我们在此报告了17只HKEM犬的特征、临床症状、治疗结果以及组织病理学和免疫学结果。
动物: 入选标准为存在:(i) 皮屑性病变伴或不伴结痂；和 (ii)EM的典型显微镜下病变(即全表皮层细胞毒性淋巴细胞性皮炎伴或不伴基底角质形成细胞的细胞凋亡)；和 (iii) 影响毛囊间表皮的显微镜下角化过度和/或角化不全。
材料和方法: 审查临床问卷和皮肤活检。对亲表皮病毒进行聚合酶链反应和直接免疫荧光检测。
结果: 各种品种都会患病，成年中后期(中位年龄9岁)公犬比例过高。全身性皮肤病变包括多灶性至融合、线性和环形的斑点和斑块，伴皮肤发红和粘附紧密的结痂。显微镜下具有EM特异性病变，以明显的表皮浅层细胞凋亡，伴淋巴细胞卫星现象和角化不全为特征。未发现药物触发因素。犬疱疹病毒聚合酶基因、犬细小病毒和犬瘟热病毒的聚合酶链反应在所有 HKEM 和犬糜烂型EM(15只犬)活检中均为阴性。病变经口服和(或)外用抗菌药治疗无效。使用免疫抑制方案时，9/17只犬 (53%) 的症状完全缓解。
结论和临床相关性: HKEM是一种慢性、持续性和临床上独特的EM亚型，不同于犬的“经典”水疱大疱性糜烂至溃疡性EM。
关键词：犬、多形红斑、角化过度
INTRODUCTION
介绍
Erythema multiforme (EM) is an acute immune-mediated disorder that affects the skin and/or mucous membranes, including the oral cavity. In humans, EM represents a blistering and ulcerative skin disease characterised by target or iris lesions distributed symmetrically on the extremities and trunk; EM is divided into minor (EMm) and major (EMM) forms based on the degree of mucosal involvement and the presence of systemic signs in the latter. The proposed pathomechanism involves T cells activated by antigen (viral, drug)-loaded epithelial cells, which will result in the destruction of antigen-expressing keratinocytes.
多形红斑(EM)是一种急性免疫介导性疾病，影响皮肤和/或粘膜，包括口腔。在人类肿，EM代表一种起泡和溃疡性皮肤病，其特征是四肢和躯干对称分布的靶形或环形病变。EM根据粘膜患病程度分为轻度(EMm)和重度(EMm)两种形式，后者有全身性症状。所提出的病理机制为T细胞被表皮细胞上负载的抗原(病毒、药物)激活，这导致抗原表达角质形成细胞的破坏。
Historically, the diagnosis of EM in veterinary medicine has been driven primarily by histopathological observation of transepidermal cytotoxic lymphocytic dermatitis with cell death occurring in suprabasal and basal epidermal layers. The tendency to equate the ‘tissue reaction pattern’ directly with the disease of EM has resulted in a clinically heterogeneous group of disorders being diagnosed as EM.
历史上，兽医学中EM的诊断主要为组织病理学可见，经表皮细胞毒性淋巴细胞性皮炎，伴表皮基底上层和表皮基底层细胞死亡。将这种“组织反应模式”认为是EM疾病，使得一些临床多样性表现的疾病被诊断为EM.
Canine EMm and EMM, characterised clinically by vesiculobullous, erosive-to-ulcerative skin lesions, resemble, to some degree, their human EM counterparts,yet a new canine subgroup described as ‘old-dog’ or ‘hyperkeratotic’ EM (HKEM) has been proposed without any detailed clinical descriptions. This form of canine EM also is considered idiopathic and is persistent with marked hyperkeratosis and parakeratosis on microscopic examination.
犬EMm和EMM的临床特征为水疱大疱性、糜烂性至溃疡性皮肤病变，在某种程度上与人类EM相似，但有人提出了一个新的犬类亚群，称为“老年犬”或“角化过度”EM (HKEM)，但没有任何详细的临床描述。这种形式的犬EM也被认为是特发性的，在显微镜检查下表现为明显的角化过度和角化不全。
The purpose of this study is to describe the history, clinical features, histopathological and immunopathological findings, and treatment outcome of 17 dogs with skin lesions of canine EM with marked microscopic hyperkeratosis and parakeratosis.
本研究的目的是描述17只显微镜下明显可见角化过度和角化不全皮肤病变的犬EM的病史、临床特征、组织病理学和免疫病理学检结果以及治疗结果。
MATERIAL AND METHODS
材料和方法
Case selection
病例筛选
Cases were identified through an Email request sent to the Vetderm list (vetderm@lists.ncsu.edu). Dogs of any age, sex or breed were included if they presented with (i) scaly skin lesions with or without crusting; and (ii) microscopic lesions typical of EM (i.e. a panepidermal cytotoxic lymphocytic dermatitis with apoptotic keratinocytes predominantly in suprabasal epidermal layers with or without observation of basal keratinocyte apoptosis); and (iii) microscopic ortho- and/or parakeratotic hyperkeratosis affecting the interfollicular epidermis.
病例是通过发送到Vetderm列表(vetderm@lists.ncsu.edu)的电子邮件请求确定的。任何年龄、性别或品种的犬，如果它们出现(i)皮屑皮肤病变，有或没有结痂;和(ii) 镜检可见EM典型病变(即：表皮全层细胞毒性淋巴细胞性皮炎，伴有主要位于基底上表皮层的角质形成细胞的细胞凋亡，有或没有基底层角质形成细胞的细胞凋亡);和(iii)镜检可见毛囊内表皮层的正角化性和/或角化不全性角化过度。
Dogs with macular, vesiculobullous, typical or atypical target skin lesions and histopathological findings of cytotoxic lymphocytic interface dermatitis with keratinocyte apoptosis occurring throughout multiple epidermal layers suggestive of ‘classic’ EMm and EMM were not considered for inclusion.
患犬存在斑点、水疱大疱性，典型或非典型的靶形皮肤病变，以及组织病理学结果为细胞毒性淋巴细胞性界面性皮炎，伴多个表皮层发生角质形成细胞的细胞凋亡，提示“经典型”EMm和EMm的犬不被考虑纳入。
Histopathological and immunopathological investigation
组织病理学和免疫病理学检查
Haematoxylin and eosin-stained histological sections of biopsies of haired skin (17 of 17 cases) and oral mucosa (1 of 17 cases) were reviewed, and lesions were identified and scored subjectively utilising a standard severity scale (see Appendix S1). Direct immunofluorescence (IF) on formalin-fixed paraffin-embedded skin sections for lupus band test (LBT) was performed as described previously.
回顾了有毛皮肤样本(17例中有17例)和口腔粘膜样本(17例中有1例)的苏木精和伊红染色的组织学活检切片，并使用标准严重程度评分表(见附录S1)主观地识别病变并进行评分。使用直接免疫荧光(IF)对福尔马林固定石蜡包包皮肤切片进行狼疮带试验(LBT)，如前所述。
DNA extraction and PCR for canine herpesvirus polymerase gene, parvovirus and distemper virus
犬疱疹病毒聚合酶基因、细小病毒和犬瘟热病毒的DNA提取和PCR
Polymerase chain reactions for canine herpesvirus polymerase (pol) gene, canine parvovirus and canine distemper virus were performed on DNA isolated from formalin-fixed, paraffin-embedded (FFPE) skin tissues from each hyperkeratotic EM dog in this study, and from FFPE blocks of 15 previously published canine erosive EM patients; positive viral FFPE tissues and viral cultures served as controls (see Appendix S1).
聚合酶链反应检测犬疱疹病毒聚合酶(pol)基因、犬细小病毒和犬瘟热病毒，对本研究中每只角化过度EM犬的福尔马林固定石蜡包封(FFPE)皮肤组织的DNA进行分离，并从15只先前发表的犬糜烂型EM患犬的FFPE块中分离，阳性病毒FFPE组织和病毒培养作为对照(见附录S1)。
Clinical management and prognosis
临床管理与预后
Data on signalment, history (including any possible triggers), clinical signs (e.g. time to development of lesions, lesion type and distribution), therapeutic management (e.g. on previous, initial and maintenance immunosuppressive therapy) and treatment outcome for each case were collected from the referring veterinaries and analysed.
从转诊兽医处收集并分析每个病例的特征、病史(包括任何可能的触发因素)、临床症状(如病变发生的时间、病变类型和分布)、治疗管理(如用药史、初期和维持期免疫抑制治疗)和治疗结果。
RESULTS
结果
Seventeen dogs met the inclusion criteria; selected dogs included 16 pure-bred and one cross-bred dog (Table S1). Two of our cases previously were published as short case reports. There was an over-representation of males (male: female ratio of 2.4), with most dogs being either spayed or neutered (13 of 17; 76%). Odds ratios for breed, sex or age predispositions for the development of hyperkeratotic EM could not be estimated, as the subjects were from various North and South American, or European countries, and a control population therefore was not available.
17例犬符合纳入标准。所选的犬包括16例纯种犬和1例杂交犬(表S1)。其中两例在之前以简短病例报告形式发表过。雄性犬占比高(雄性：雌性比例为2.4)，大多数犬已节育(17例犬中有13例;76%)。由于入选病例来自北美、南美或欧洲国家，因无法获得对照组，所以无法估计角化过度EM的品种、性别或年龄易感性。
The age of onset of skin lesions varied between one and 15years (median 9years; mean 8.5years); most dogs (15 of 17; 88%) began exhibiting noticeable lesions in mid-adulthood (i.e. aged five or older).
皮肤病变的发病年龄在1到15岁之间(中位值9岁;平均值8.5岁)，大多数犬(17例中有15例;88%)为中年(即5岁或5岁以上)开始出现明显的病变。
Clinical summary
临床总结
Before the initial visit to the referral clinician, skin lesions were present between 2 and 12months (median 4months), and there was no history of skin lesions waxing and waning or resolving spontaneously. The most common lesions reported by the owners were erythematous, hyperkeratotic and crusting dermatitis; these usually were first noticed on the trunk, abdomen and ear pinnae.
在转诊医生首诊前，皮肤病变存在时间在2至12个月之间(中位值4个月)，无皮肤病变时好时坏或自行消退病史。主诉最常见病变是皮肤发红、角化过度和结痂性皮肤病。这些病变通常首先出现在躯干、腹部和耳廓。
According to our inclusion criteria, skin lesions included multifocal-to-coalescing, linear and annular macules and plaques with erythema and firm adherent crusting (Figures 1 and 2). These lesions often were overlaid with hyperpigmented, dark-coloured or haemorrhagic adherent material (9 of 17 dogs; Figure 1), whereas other dogs exhibited skin lesions covered by predominantly thick adherent yellow waxy-to-greasy material (8 of 17 dogs; Figure 2).
根据我们的纳入标准，皮肤病变包括多灶性至聚集性、线性和环形斑点和斑块，伴皮肤发红和粘附紧实的结痂(图1和图2)。这些病变下方通常有色素沉着、深色或出血性的附着物质(9/17;图1)，而其他犬则表现出皮肤病变被覆着明显的厚的黄色蜡状油腻物质(8/17;图2)。
Upon presentation to the referral clinician, all dogs had lesions on the trunk and ventral abdomen. Lesions also were present on the ear pinnae and head in 16 of 17 cases (94%), respectively, while axillae and distal limbs were affected in 13 (76%) and 14 (82%) dogs, respectively. Mild footpad hyperkeratosis was observed at the margin of the paw pads in three dogs (18%). Sixteen dogs (94%) exhibited mucocutaneous lesions on or around the genitalia, lips, anal area and oral cavity (Figure 3).
转诊前，所有患犬在躯干和腹部腹侧都有病变。17例中有16例(94%)在耳廓和头部也有病变，而腋窝和四肢远端患病的分别有13例(76%)和14例(82%)。在3例犬(18%)的爪垫边缘观察到轻度的爪垫角化过度。16例犬(94%)在生殖器、唇部、肛门区域和口腔或其周围出现皮肤粘膜病变(图3)。
No systemic signs were observed in any dog, apart from recurrent lethargy in 8 of 17 dogs (47%). Four dogs (Dogs 3, 4, 15 and 17) had a history of mild allergic skin disease. By contrast, two dogs developed hyperkeratotic EM skin lesions 4–7months into treatment for immune-mediated thrombocytopenia (Dog 14) and arthritis (Dog 2), respectively. Pruritus and pain at the lesion site were reported in 14 (82%) and 12 dogs (70%), respectively.
除了17例犬中有8例(47%)反复嗜睡外，任何犬都没有观察到全身性症状。4例犬(犬3、4、15和17)有轻度过敏性皮肤病病史。相比之下，两例犬分别在免疫介导性血小板减少症(犬14)和关节炎(犬2)治疗4-7个月后出现角化过度型EM皮肤病变。分别有14例(82%)和12例(70%)报道病变部位有瘙痒和疼痛表现。
A complete blood count revealed mild leucocytosis in two dogs. Serum biochemical values were within reference intervals in 11 dogs, while urinalysis results were unremarkable in all dogs. Two dogs had elevated alkaline phosphatase activity and vacuolated hepatopathy on ultrasound examination, probably as a result of the previous administration of methylprednisolone (Dog 6) and prednisone/azathioprine (Dog 2) by the primary veterinary surgeon.
两例犬全血细胞计数显示有轻度白细胞增多症。11例犬的血清生化值在参考区间内，而所有犬的尿液分析结果均无异常。两例犬出现碱性磷酸酶活性升高，以及超声检查有肝脏空泡样变，可能是由于之前首诊外科医生使用了甲基泼尼松龙(犬6)和泼尼松/硫唑嘌呤(犬2)。
Impression cytological examination revealed degenerate neutrophils and numerous cocci in three dogs (Dogs 1, 3 and 14), which were identified as meticillin susceptible Staphylococcus pseudintermedius by aerobic bacterial culture. One to three courses of systemic antibiotics (e.g. cefpodoxime, cephalexin, marbofloxacin, doxycycline and enrofloxacin) for 14–28days in conjunction with topical antimicrobials were initially provided to 15 of 17 dogs without clinical improvement, and with the new HKEM skin lesions development during the antimicrobial administration. Fungal culture was negative in seven dogs (Dogs 3, 4, 5, 9, 10, 12 and 13).
皮肤压片细胞学检查发现3例犬(犬1、3和14)结果为退行性中性粒细胞和大量球菌，经需氧细菌培养鉴定为甲氧西林敏感型假中间型葡萄球菌。17例犬中有15例最初接受了一到三个疗程的全身性抗生素治疗(如头孢泊肟、头孢氨苄、马坡沙星、多西环素和恩诺沙星)，为期14-28天，联合外用抗菌剂，但无临床改善，在使用抗生素期间出现了新的HKEM皮肤病变。7例犬(犬3、4、5、9、10、12、13)真菌培养阴性。
There was no clear association with the administration of a drug, vaccine or nutraceutical, nor any history of preceding infections in any dog included herein. Dog 2 developed HKEM during the 4month azathioprine treatment, while Dog 15 received meloxicam 4weeks before the onset of HKEM lesions. The discontinuation of both drugs (azathioprine, meloxicam) did not resolve or improve HKEM skin lesions; any association between HKEM and these drugs remains unconfirmed.
所有患犬包括本文患犬的疾病与药物、疫苗或保健品没有明显相关性，与感染病史也没有明显相关性。犬2在硫唑嘌呤治疗4个月期间出现HKEM，而犬15在HKEM病变开始4周前接受美洛昔康治疗。停用两种药物(硫唑嘌呤、美洛昔康)均未消退或改善HKEM皮肤病变，这些药物是否与HKEM有关仍未得到证实。
Histopathological findings
组织病理学结果
Raised plaques contained epidermal hyperkeratosis and a panepidermal cytotoxic dermatitis, both of which were inclusion criteria. Hyperkeratosis (Figure 4) ranged from mild to marked and was due to parakeratosis (16 of 17; 94%) often intermixed with orthokeratosis (12 of 17; 71%) in laminated-to-compact patterns. Epidermal and infundibular hyperplasia mainly were moderate-to marked and regular-to-irregular. Epidermal spongiosis was mild and uncommon (four of 17; 23%).
凸起的斑块包含表皮角化过度和全表皮层细胞毒性皮炎，这两者都是纳入标准。角化过度(图4)范围从轻度到明显，是由于角化不全(16/17;94%)经常混有正角化性角化过度(12/17;71%)呈层状-致密模式。表皮层及毛囊漏斗部增生以中度-明显、规则-不规则为主。表皮层轻微且不常见海绵样水肿(4/17;23%)。
The cytotoxic dermatitis reaction pattern (Figure 4) occurred in areas of epidermal hyperplasia. Here, apoptosis of suprabasal epidermal keratinocytes was typically marked and occurred with apoptosis of basal layer keratinocytes (17 of 17; 100%), which usually was mild, yet was moderate in four cases and marked in one. Clustering of suprabasal epidermal apoptosis occurred commonly (Figure 4). Many cases had small zones of epidermal atrophy (12 of 17; 71%), including the architectural collapse of epidermal layering. Epidermal hydropic degeneration (14 of 17; 82%) often occurred in areas of apoptosis, and mainly was mild. Lymphocyte exocytosis (16 of 17; 94%) and satellitosis of apoptosis (16 of 17; 94%) were moderate-to-marked in nearly all cases. Interestingly, while lymphocytes dominated in the epidermis in all cases, histiocytes were obvious in over half of the patients and sometimes were numerous as satelliting cells (Figure 4). Apoptotic keratinocytes and satelliting leucocytes often were entrapped in the stratum corneum, usually layered with parakeratosis (Figure 4).
细胞毒性皮炎反应模式(图4)发生在表皮增生区域。图中，基底上层表皮角质形成细胞的细胞凋亡明显，且伴有基底层角质形成细胞的细胞凋亡(17/17;100%)，通常为轻度，但有4例为中度，1例有明显症状。基底上层表皮层细胞凋亡常见聚集(图4)。许多病例有小范围的表皮层萎缩(12/17;71%)，包括表皮层结构性萎缩。表皮层水变性(14/17;82%)多发生于细胞凋亡区域，以轻度为主。几乎所有病例都可见中度至明显的淋巴细胞外排(16/17;94%)和细胞凋亡的卫星现象(16/17;94%)。有趣的是，尽管所有病例表皮层卫星现象均以淋巴细胞为主，但半数以上患犬可见组织细胞，有时卫星现象的细胞量多(图4)。角质层常见角质形成细胞的细胞凋亡和白细胞卫星现象，通常位于角化不全层(图4)。
A lymphocytic interface dermatitis reaction pattern (17 of 17; 100%) was mostly mild-to-moderate, while a lichenoid band (16 of 17; 94%) ranged from mild to marked and often was patchy. Despite basal cell cytotoxicity, basal layer mitotic cells were common (16 of 17; 94%) and ranged from mild to marked. Pigmentary incontinence (12 of 13; 92%) was mild, and only sometimes moderate. Basement membrane zone thickening (13 of 17; 76%) was multifocal, patchy, and ranged from mild to marked (Figure 4). Neither dermal fibrosis nor granulation tissue were features. Erosions mostly were absent and, when present, were typically part of a suppurative crusting dermatitis, with or without surface bacterial colonies, which were interpreted to be secondary.
淋巴细胞性界面性皮炎反应模式(17/17;100%)大多为轻度至中度，而苔藓样条带(16/17;94%)的症状从轻微到明显，通常呈片状。尽管基底细胞具有细胞毒性，但基底层常见有丝分裂细胞很(16/17;94%)，范围从轻微到明显。轻度色素失禁(12/13;92%)，有时为中度。基底膜区增厚(13/17;76%)呈多灶性、斑片，范围从轻度到明显(图4)。无真皮纤维化和肉芽组织特征。大多无糜烂，当出现时，通常是代表部分化脓性结痂性皮炎表现，有或没有表面细菌菌落，这被认为是继发性的。
Additional inflammatory patterns were present, and adnexal changes were common and mirrored epidermal changes—see details in Appendix S2.
还存在其他炎症模式，常见附件改变，类似表皮改变-详见附录S2。
Immunopathological findings
免疫病理学结果
Direct IF was performed on paraffin-embedded skin sections from 14 of 17 dogs. A positive LBT was found in seven of 14 (50%) dogs with the anti-immunoglobulin (Ig)G and in one dog (5%) with anti-IgA and IgM. None of the dogs showed positive direct IF for C3. The detection of serum antinuclear antibodies (ANA) was negative in the three dogs tested.
对17例犬中的14例石蜡包埋的皮肤切片进行了直接IF。14例抗免疫球蛋白(Ig)G犬中有7例(50%)抗IgG阳性，1例(5%)抗IgA和IgM阳性。没有一条犬的C3直接IF呈阳性。3例犬的血清抗核抗体(ANA)检测均为阴性。
Polymerase chain reaction for canine herpes (pol)-, distemper and parvovirus
犬疱疹(pol)、犬瘟热和细小病毒的聚合酶链反应
Polymerase chain reaction amplification was performed successfully on each positive-control specimen with the corresponding viral PCR primers; negative controls produced no viral amplicons. No viral amplification was observed for canine herpesvirus (pol), distemper virus and parvovirus in FFPE samples of 16 of 17 HKEM and 15 dogs with erosive EM.
用相应的病毒PCR引物对每个阳性对照标本进行聚合酶链反应扩增成功;阴性对照没有产生病毒扩增子。在17例HKEM和15例糜烂性EM犬的FFPE样本中，未观察到犬疱疹病毒(pol)、犬瘟热病毒和细小病毒的病毒扩增。
Treatment outcome
治疗结果
Information on treatment outcomes was available for all dogs. Spontaneous disappearance of clinical signs was not seen in any patient, and skin lesions had failed to resolve completely after discontinuing oral antibiotics and topical antimicrobials that were provided initially to 15 of 17 dogs. Dogs in which a meticillin-susceptible S. pseudintermedius was cultured were provided with appropriate systemic antibiotics and topical antimicrobial chlorhexidine products for several weeks. Immunomodulating treatment regimens varied widely. Thirteen dogs were treated initially with systemic glucocorticoids alone (3 of 17) or in conjunction with oral azathioprine (1 of 17), mycophenolate mofetil (1 of 17) and ciclosporin (8 of 17); four dogs received ciclosporin or oclacitinib monotherapy. Systemic glucocorticoid monotherapy with prednisone/prednisolone (1.5–2.5 mg/kg once daily) induced a partial improvement of 75% (Dogs 3 and 12) to complete remission (Dog 11); both dogs with partial improvement were lost to follow-up after 30–40days while Dog 11 was maintained on tapered dosage for 7months before being lost to follow-up.
所有的犬都有治疗结果记录。所有患犬都没有出现临床症状自发性消退，17例犬中有15例初期停用口服抗生素和外用抗菌素后，皮肤病变没有出现恢复。对培养结果为有甲氧西林敏感的假中间型葡萄球菌的犬给予适当的全身性抗生素和外用氯己定抗菌产品数周。免疫调节治疗方案差异很大。13例犬最初单独使用全身糖皮质激素(3/17)，或联合口服硫唑嘌呤(1/17)、霉酚酸酯(1/17)和环孢素(8/17)治疗。4例犬接受环孢素或奥拉替尼单药治疗。使用泼尼松/泼尼松龙糖皮质激素单药治疗(1.5-2.5 mg/kg，每日一次)，达到75%的部分改善(犬3和犬12)到完全缓解(犬11)。所有部分缓解患犬均在30-40天后失访，犬11在逐渐减量7个月后失访。
Oral glucocorticoids (1–2 mg/kg/day for 2–4weeks and then tapered) in conjunction with ciclosporin (5– 10 mg/kg once daily) induced complete remission (Dogs 4, 5, 15 and 16; Figure 5) and mild-to-partial improvement (Dogs 1, 7, 10 and 14) in skin lesions of four dogs each. Two dogs (1 and 14) with mild improvement on ciclosporin/glucocorticoids were switched to oral oclacitinib (Dog 1, 0.6 mg/kg twice daily; Dog 14, 1 mg/ kg twice daily) and complete remission was observed after 12 (Dog 1) and six (Dog 14) weeks, respectively.
口服糖皮质激素(1-2 mg/kg/天，持续2 - 4周，然后逐渐减少)，联合环孢素(5 - 10 mg/kg，每日一次)，使4例犬完全缓解(犬4,5,15和16;图5)，4例犬轻度到部分改善(犬1、7、10和14)。2例犬环孢素/糖皮质激素有轻度改善(1和14)，替换成口服奥拉替尼(犬1,0.6 mg/kg，每天两次;犬14,1毫克/公斤，每天两次)，分别在12周(犬1)和6周(犬14)后达到完全恢复。
Dog 2 was continued on azathioprine and prednisone from previous therapy for immune-mediated arthritis yet showed only mild improvement; it was euthanised owing to the poor quality of life (QoL). Oral mycophenolate mofetil (10 mg/kg twice daily) in Dog 17 achieved a complete remission at 3months of therapy with glucocorticoids discontinued after the initial 6weeks; however, the patient was lost to follow-up 4months later.
犬2继续使用硫唑嘌呤和泼尼松治疗免疫介导性关节炎，但仅显示轻度改善，由于生活质量太差，它被安乐死了。犬17口服霉酚酸酯(10毫克/公斤，每天两次)治疗3个月，在治疗初期6周后停止使用糖皮质激素，达到了完全恢复，但是，患犬复诊4个月后失访。
Two dogs initiated on oral ciclosporin alone (5 mg/kg once daily) for two (Dog 9) and eight (Dog 6) months achieved complete and partial remission, respectively. Initial monotherapy with oral oclacitinib (0.6 mg/kg twice daily) in Dog 8 and Dog 13 for 14 and 42days, respectively, resulted in minimal clinical improvement. Clinical improvement was observed after the switch to systemic glucocorticoids, yet as a consequence of the severe glucocorticoid adverse effects and poor QoL, both dogs were humanely euthanised.
两例犬开始单独口服环孢素(5毫克/公斤，每天一次)，持续2个月，达到完全恢复(犬9)，持续8个月，达到部分恢复(犬6)。犬8和犬13初期口服奥拉替尼单药治疗(0.6 mg/kg，每天两次)分别持续14天和42天，临床改善甚微。在改用全身性糖皮质激素后，观察到临床改善，但由于严重的糖皮质激素副作用和生活质量差，两只犬都进行了人道安乐死。
The median time to complete remission in nine of 17 dogs was 2.5 months (range 6–28weeks). In cases where treatment was discontinued or tapered (nine of 17 dogs; 52%), two dogs (of nine) remained lesion-free without any treatment, while skin lesions returned in seven of nine patients (77%); signs eventually disappeared after re-inducing treatment.
17例犬中有9例完全恢复的中位时间为2.5个月(范围6 - 28周)。在停止治疗或逐渐减少治疗的病例中(9/17;52%)，其中两例犬(9例)在没有任何治疗的情况下保持无病变，而9例犬中有7例(77%)皮肤病变复发。在重新诱导治疗后，症状最终消失。
DISCUSSION
讨论
Although there are extensive case series descriptions of dogs with erosive vesiculobullous EM, there are only two case reports of dogs with an apparent hyperkeratotic variant of this disease. Herein, we describe the signalment, clinical signs, treatment outcomes, microscopic and immunopathological characteristics of 17 dogs with HKEM.
虽然有大量的犬糜烂性水疱大疱性EM的病例系列描述，但只有两个病例报告的犬有明显的角化过度型亚型。本文描述了17例HKEM患犬的特征、临床症状、治疗结果、显微特征和免疫病理学特征。
The age of onset of HKEM was variable, with most dogs exhibiting their first skin lesions in mid- to late adulthood, two-thirds developing lesions after 8years of age. The late age of onset in some dogs may explain why some authors initially proposed the terminology of ‘old-dog’ EM.
HKEM发病年龄各不相同，大多数犬在成年中老年后首次出现皮肤病变，三分之二的犬在8岁后出现皮肤病变。一些犬的发病年龄较晚可能解释了为什么一些作者最初命名为了“老年犬”EM术语。
Skin lesions of HKEM featured multifocal-to-coalescing, linear and annular macules and plaques with erythema and adherent firm crusting. In most patients, HKEM skin lesions were generalised and commonly affected sites included the trunk, abdomen, mucocutaneous junctions and concave pinnae. Generalised discoid lupus erythematosus (GDLE) was considered a close mimic of HKEM, yet in the latter skin lesions were more proliferative, without the scarring and skin atrophy observed in the former. Moreover, the prominent suprabasal apoptosis in conjunction with lymphocytic satellitosis observed in HKEM-type reaction patterns is only rarely observed in GDLE.
HKEM皮肤病变以多灶至聚集、线性及环形斑点及斑块伴皮肤发红及粘附致密结痂为特征。在大多数患病动物中，HKEM皮肤病变是全身性的，常见的患病部位包括躯干、腹部、皮肤粘膜交界处和耳廓凹面。全身性盘状红斑狼疮(GDLE)被认为与HKEM相似，但后者皮肤病变更具增生性，没有前者的瘢痕和皮肤萎缩表现。此外，在HKEM型反应模式中观察到的主要以基底上层细胞凋亡和淋巴细胞卫星现象，在GDLE中罕见。
Necrolytic migratory erythema (NME) is an erosive, hyperkeratotic and crusting disease commonly affecting footpads (96% of patients), mucocutaneous junctions and pressure points (e.g. elbow and hocks), with histopathological features of epidermal hyperplasia, severe parakeratotic hyperkeratosis and diffuse intracellular (vacuolar change) and extracellular oedema in the mid-epidermal layers. Some dogs with NME show keratinocyte apoptosis in different epidermal layers on histopathological examination; the aetiology and significance of the phenomenon are currently unknown. In the dogs with HKEM in this report, mild footpad involvement was limited to the margin and serum biochemistry values were within the reference interval, which, together with the HKEM clinical and microscopic features, excluded NME.
坏死松懈性游走性红斑(NME)是一种糜烂性、角化过度和结痂性疾病，常见爪垫(96%的患病动物)、皮肤粘膜交界处和压力点(如肘关节和跗关节)患病，其组织病理学特征为表皮增生、严重的角化不全性角化过度和表皮中层弥散性细胞内(空泡性改变)和细胞外水肿。有些NME患犬在组织病理学检查中显示不同表皮层的角质形成细胞的细胞凋亡。这种现象的病因和意义目前尚不清楚。在本报告中，HKEM患犬中，轻度爪垫患病局限于边缘，血清生化值在参考区间内，加上HKEM的临床和显微特征，排除了NME。
The exact pathomechanism of human and canine EM remains incompletely understood, yet pathogenetic mechanisms of CD8+ lymphocytic cytotoxic responses against keratinocytes are shared in both diseases. In humans, acute self-limited EM typically is precipitated by viral infections, most commonly via DNA particles containing herpes simplex virus DNA pol gene expressed in lesional keratinocytes. The inciting factor of persistent EM, characterised by the widespread and continuous presence of EM skin lesions, remains unknown in many patients. Trigger factors for the development of canine EM in most cases are unknown. The development of erosive-to-ulcerative canine EM typically has been associated with the administration of various drugs. However, this association was most often unproven or speculative in most cases and some dogs with ‘drug-induced EM’ might have had, instead, drug-induced Stevens–Johnson syndrome or toxic epidermal necrolysis. Finally, EM in some dogs can arise without known drug exposure (e.g. ‘idiopathic’ EM).
人类和犬EM的确切发病机制仍不完全清楚，但CD8+淋巴细胞对角质形成细胞的细胞毒性反应的发病机制在这两种疾病中是相同的。在人类中，急性自限性EM通常是由病毒感染引起的，最常见的是通过含有单纯性疱疹病毒DNA pol基因的DNA颗粒在病变角质形成细胞中表达。EM的持续性刺激因素，其特征是广泛和持续存在的EM皮肤病变，在许多患者中仍然未知。大多数犬EM病例发展的触发因素是未知的。糜烂性至溃疡性犬EM的发展通常与各种药物使用有关。然而，在大多数病例中，这种关联通常是未经证实的或推测的，一些患有“药物诱发性EM”的犬可能患有药物诱发性Stevens-Johnson综合征或中毒性表皮坏死松解症。最后，某些EM患犬的可以在没有已知药物使用的情况下发病(例如“特发性”EM)。
In a few dogs, EM was reported to be associated with canine parvovirus. Interestingly, the ‘cytopathic’ epidermal infection with the development of viral inclusion bodies in keratinocytes suggests that these dogs suffered from mucosal and cutaneous parvovirosis rather than virus-associated EM, in which virions are not produced at the site of EM skin lesions. In the present study, there was no viral amplification observed for canine herpesvirus (pol), distemper virus and parvovirus in samples from HKEM and erosive EM samples. A single case report implicated a nutraceutical product as the causative agent for HKEM lesions. By contrast, our study could not confirm the association between HKEM cases and a drug.
在少数患犬中，报道EM与犬细小病毒有关。有趣的是，在角质形成细胞中出现病毒包涵体的“细胞病变”表皮感染提示，这些犬患有粘膜和皮肤细小病毒病，而不是病毒相关的EM，即EM皮肤病变部位不产生病毒颗粒。在本研究中，在HKEM和糜烂EM样本中未观察到犬疱疹病毒(pol)、犬瘟热病毒和细小病毒的病毒扩增。一个单一的病例报告涉及一种保健品是导致HKEM病变的病因。相比之下，我们的研究无法证实HKEM病例与药物之间的关联。
Treatment modalities for EM should focus on addressing the aetiology, eliminating possible trigger factors and when needed, administering immunosuppressive medications to resolve dysregulated immune responses. There have been no clinical trials evaluating treatments of canine EM variants, and a spontaneous resolution of signs was not seen in any of our dogs with HKEM. In human EM skin lesions, T-lymphocyte/natural killer cell cytotoxicity and Janus kinase (JAK)-signal transducer and activator of transcription (STAT) signalling were highly upregulated pathways in a recent transcriptome study. In the present report, various immunomodulatory drugs that target innate immune, lymphocyte responses and JAK–STAT pathways, such as glucocorticoids, ciclosporin, oclacitinib, mycophenolate mofetil or azathioprine, were used initially to treat dogs with HKEM.
EM的治疗方式应侧重于解决病因，消除可能的触发因素，必要时给予免疫抑制药物以解决免疫反应调解异常。目前还没有临床试验评估犬EM的多种治疗方法，本文所有HKEM患犬都没有出现自发性症状消退。在人类EM皮肤病变中，在最近的转录组研究中发现T淋巴细胞/自然杀伤细胞细胞毒性和Janus激酶(JAK)信号传感器和转录激活因子(STAT)信号通路高度上调。在本报告中，针对先天性免疫、淋巴细胞反应和JAK-STAT通路的各种免疫调节药物，如糖皮质激素、环孢素、奥拉替尼、霉酚酸酯或硫唑嘌呤，初期都被用于治疗HKEM患犬。
Ciclosporin, a calcineurin inhibitor, blocks T-cell infiltration, activation, and the subsequent release of inflammatory cytokines interleukin (IL)-2, IL-4, interferon (IFN)-γ and tumour necrosis factor (TNF)-α. In this study, the complete remission of HKEM skin lesions followed treatment with ciclosporin (along with a short course of glucocorticoids at its onset in some dogs) in half of the dogs (5 of 10) given this drug.
环孢素是一种钙调磷酸酶抑制剂，它可以阻止T细胞的浸润、激活以及随后炎症细胞因子白介素(IL)-2、IL-4、干扰素(IFN)-γ和肿瘤坏死因子(TNF)-α的释放。在这项研究中，半数犬(5/10)在接受环孢素治疗后(有些犬发病时，联合使用短疗程糖皮质激素)，HKEM皮肤病变完全缓解。
Oclacitinib, a JAK inhibitor with preferential affinity to JAK1,successfully induced a complete remission in half (two of four) dogs with HKEM. In humans, the oral JAK inhibitors upadacitinib and tofacitinib led to a marked improvement in four patients affected with persistent EM. The effectiveness of JAK inhibitors in EM is likely to be related to their inhibition of interferon-γ (JAK1/2), IL-15 (JAK1/3) and type I interferons (JAK1/ TYK2); however, the precise mechanism of action of JAK inhibitors in human and canine EM warrants further investigations.
奥拉替尼是一种对JAK1具有优先亲和力的JAK抑制剂，在半数用药的HKEM患犬(2/4)中成功达到完全缓解。在人类中，口服JAK抑制剂乌帕替尼和托法替尼可显著改善4例反复性EM患者的症状。JAK抑制剂在EM中的有效性可能与其抑制干扰素-γ (JAK1/2)、IL-15 (JAK1/3)和I型干扰素(JAK1/ TYK2)有关，然而，JAK抑制剂在人类和犬EM中的确切作用机制值得进一步研究。
In conclusion, we report herein 17, predominantly male, dogs with a unique proliferative clinical phenotype of EM characterised by persistent and continuous skin lesions without identifiable viral infection or drug triggers, which we propose to name HKEM. In these dogs, skin lesions appear to respond to a wide range of immunosuppressive regimens, yet many patients experienced relapses upon the tapering of drug dosages. Our limited outcome data suggest that ciclosporin and oclacitinib should be considered as potentially effective therapeutic options for canine HKEM. Further investigations evaluating the pathogenesis of HKEM and responses to medications at the molecular level are warranted.
总之，我们在此报告了17例具有独特的增生性临床表型的EM患犬，主要为雄性，其特征为反复和持续的皮肤病变，没有病毒感染或触发性药物，我们建议将其命名为HKEM。在这些犬中，皮肤病变似乎对多种免疫抑制方案有效，但许多患犬在药物剂量逐渐减少后出现复发。我们有限的结果数据表明，环孢素和奥拉替尼应被认为是犬HKEM潜在有效的治疗选择。有必要进一步研究评估HKEM的发病机制和在分子水平上的药效反应。
FIGURE 1 Initial clinical presentation of dogs with hyperkeratotic erythema multiforme (HKEM). (a–c) Multifocal-to-coalescing, linear and annular hyperpigmented macules, papules and plaques with erythema and adherent, firm dark-coloured (a,b) to haemorrhagic (c) crusting affecting the ventral abdomen and inguinal areas in Dogs 2 and 5.
图1角化过度型多形红斑(HKEM)患犬的初期临床表现。(a -c) 2号和5号犬腹部腹侧和腹股沟区的多灶性至聚集的线状和环形色素沉着斑点、丘疹和斑块，伴皮肤发红和粘附紧密的深色(a,b)至出血性(c)结痂。
FIGURE 2 Initial clinical presentation of dogs with hyperkeratotic erythema multiforme (HKEM). (a) Proliferative multifocal and coalescing erythematous plaques with thick adherent waxy-to-greasy material affecting the ventral abdomen and inguinal areas of Dog 15. (b,c) Close up of the multifocal erythematous plaques with severe, yellow, thick adherent material in Dog 1 (c) and Dog 15 (b).
图2角化过度型多形红斑(HKEM)患犬的初期临床表现。(a)15号犬腹部腹侧和腹股沟区增生性多灶性至聚集性发红斑块，伴粘附厚的蜡质-油腻物质。(b,c)1号犬(c)和15号犬(b)中多灶性发红斑块的特写，严重粘附厚的黄色物质。
FIGURE 3 Initial clinical presentation of dogs with hyperkeratotic erythema multiforme (HKEM). (a,c) Proliferative coalescing erythematous and eroded plaques with thick adherent waxy-to-greasy material affecting inguinal, perigenital and perianal areas in Dog 15. (b) Hyperpigmented multifocal to coalescing macules and plaques with adherent scaling and crusting affecting the concave pinnae and pinna margin in Dog 5.
图3角化过度型多形红斑(HKEM)患犬的初期临床表现。(a,c)15号犬的腹股沟、生殖周围和肛周区域的增生性聚集的发红和糜烂的斑块，伴粘附厚的蜡质-油腻物质。(b)5号犬耳廓凹面和耳廓边缘的多灶性色素沉着至聚集斑点和斑块，粘附皮屑和结痂。
FIGURE 4 Histopathological presentation of epidermal and mucosal changes in canine hyperkeratotic erythema multiforme (HKEM). (a) The margin of a plaque is present that is composed largely of epidermal hyperplasia and hyperkeratosis. ×10. (b) [Inset from (a)] Marked parakeratotic epidermal (asterisks) and infundibular (arrows) hyperkeratosis is present over irregular epidermal and infundibular hyperplasia. A moderate band-like (lichenoid) infiltrate of inflammation is just below the epidermis. ×20. (c) Orthokeratotic hyperkeratosis (arrow) is present occasionally without parakeratosis in a few biopsies. ×10. (d) Higher magnification image shows parakeratosis (asterisk) over epidermal hyperplasia that contains prominent apoptosis (arrows) of keratinocytes at all epidermal levels. Lymphocytic exocytosis and satellitosis of apoptotic cells are marked. The dermis (‘D’) contains numerous lymphocytes and fewer plasma cells in the bandlike (lichenoid) infiltrate below the epidermis. ×40. (e) Apoptotic keratinocytes are individual or grouped, and lymphocytic satellitosis is accompanied by a prominent number of histiocytes (arrows) in some cases. Apoptotic keratinocytes (arrowheads), along with satelliting leucocytes, are carried into the stratum corneum in high numbers in parakeratotic areas. ×40. (f) A markedly thickened basement membrane zone (arrows) is present along with lymphocytic interface dermatitis, seen as jumbling and loss of basal keratinocytes with lymphocyte infiltrate. ×40. (g) Mucosal epithelium contains apoptosis (arrows) at all mucosal epithelial levels with prominent lymphocytic exocytosis and satellitosis. ×40. Haematoxylin and eosin.
图4犬角化过度型多形红斑(HKEM)表皮和粘膜的组织病理学表现。
(a)斑块边缘主要表现为表皮增生和角化过度。×10。
(b)[(a)中小图]表皮层和毛囊漏斗部不规则的增生，明显可见表皮层(星号)和毛囊漏斗部(箭)角化不全性角化过度。表皮层下中度条带状(苔藓样)炎性浸润。×20。
(c)在少数活组织检查中出现正角化性角化过度(箭)偶见无角化不全性角化过度。×10。
(d)高倍放大图像显示表皮增生上的角化不全性角化过度(星号)，所有表皮层都有明显的角质形成细胞的细胞凋亡(箭)。明显有淋巴细胞外排和对细胞凋亡的卫星现象。真皮层(“D”)在表皮层下条带样浸润(苔藓样)中含有大量淋巴细胞，少量浆细胞。×40。
(e)角质形成细胞的细胞凋亡是单个的或成组的，有些病例中淋巴细胞卫星现象也伴有大量的组织细胞(箭)。在角质层的角化不全性角化过度区域，角质形成细胞的细胞凋亡(箭头)与白细胞卫星现象。×40。
(f)基底膜带明显增厚(箭)伴淋巴细胞性界面性皮炎，基底层角质形成细胞混杂不清和缺失，伴淋巴细胞浸润。×40。
(g)在所有粘膜上皮层均有粘膜上皮细胞的细胞凋亡(箭)，并伴有明显的淋巴细胞外排和卫星现象。×40。苏木精和伊红。
FIGURE 5 Clinical presentation of Dog 15 with HKEM before (a) and after treatment at Day 95 (b) with combined/concurrent glucocorticoids and ciclosporin; glucocorticoids were discontinued after 35days and the patient achieved complete remission with only daily ciclosporin.
图5 15号HKEM患犬在治疗前(a)和治疗第95天(b)的临床表现，联合使用/同时使用糖皮质激素和环孢素。治疗35天后停用糖皮质激素，患犬仅每日使用一次环孢素即可达到完全缓解。
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发表于 2023-1-6 22:20:27
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