使用环孢素和甲氨蝶呤治疗一例犬肉芽肿性毛囊壁炎

王帆

Granulomatous mural folliculitis in a dog treated with ciclosporin and methotrexate

使用环孢素和甲氨蝶呤治疗一例犬肉芽肿性毛囊壁炎

作者Pauline Panzuti , Marion Mosca and Didier Pin  

翻译：李诗如

Background – Canine granulomatous mural folliculitis is a very rare cause of scarring alopecia with unknown pathogenesis.

Hypothesis/objectives – To report a case of granulomatous mural folliculitis in a dog treated with ciclosporin (Cs) and methotrexate (MTX).

Animal – A 13-year-old spayed female Pyrenean shepherd dog with a one month history of scarring alopecia.

Methods and results – Histopathological examination revealed a granulomatous and lymphocytic mural and necrotizing folliculitis and furunculosis. Immunochemistry, using antibodies for CD3, CD204, CD206, IBA-1 and CD208, revealed that CD3+ lymphocytes were infiltrating the outer root sheath along with IBA-1+ or CD204+ cells. Ciclosporin (5 mg/kg once daily) and MTX (0.25 mg/kg once weekly then 0.5 mg/kg once weekly) were initiated simultaneously, and Cs was stopped after stabilization of the lesions. The dog’s skin disease was stable for six months.

Conclusions and clinical importance – To the best of the authors’ knowledge, this is the first report of the long-term management of a granulomatous mural folliculitis in a dog. Ciclosporin and MTX appeared to be an effective treatment option. Additional treated cases are needed to assess the effectiveness of MTX in canine immune-mediated diseases.

摘要

背景–犬肉芽肿性毛囊壁炎是一种罕见的瘢痕性脱毛，其病因不明。

假设/目的–报告一例使用环孢素(Cs)和甲氨蝶呤(MTX)治疗的肉芽肿性毛囊壁炎。

动物–一只13岁已绝育雌性比利牛斯山牧羊犬，有一个月的瘢痕性脱毛病史。

方法和结果 – 组织病理学检查显示肉芽肿和淋巴细胞性坏死性毛囊壁炎和疖病。使用CD3、CD204、CD206、IBA-1、CD208抗体进行免疫组化检查，发现CD3+淋巴细胞与IBA-1+、CD204+细胞浸润外根鞘。环孢素 (5 mg/kg，每日一次)和MTX (0.25 mg/kg，每周一次，然后0.5 mg/kg，每周一次)同时使用，待病变稳定后停用Cs。该犬的皮肤病稳定了六个月。

结论和临床重要性–据作者所知，这是第一份关于肉芽肿性毛囊壁炎长期治疗的报告。环孢素和甲氨蝶呤似乎是一种有效的治疗选择。需要更多的治疗病例来评估MTX在犬免疫介导性疾病中的效果。

Introduction

介绍

Granulomatous mural folliculitis is a very rare cause of scarring alopecia in dogs; there are a few reports; little is known about the pathogenesis and long-term management of this disease. In mural folliculitis, inflammation targets the hair follicle walls and is composed mainly of lymphocytes at the level of the  Isthmus or below. Lymphocytic mural folliculitis can be observed in various diseases such as demodicosis, dermatophytosis, and cutaneous and systemic lupus erythematosus. In granulomatous mural folliculitis, histiocytes, macrophages and giant cells infiltrate the outer root sheath of the hair follicle along with lymphocytes.

肉芽肿性毛囊壁炎是一种非常罕见的导致犬瘢痕性脱毛的疾病。虽有少数报告，但关于这种疾病的发病机理和长期治疗知之甚少。在毛囊壁炎中，炎症针对毛囊壁，炎性细胞以淋巴细胞为主多集中在毛囊的峡部或下部。淋巴细胞性毛囊壁炎在多种疾病中都可以观察到，例如蠕形螨病、皮肤癣菌病以及皮肤和系统性皮肤发红狼疮。在肉芽肿性毛囊壁炎中，组织细胞，巨噬细胞和巨细胞与淋巴细胞浸润到毛囊外根鞘。

Methotrexate (MTX, 4-amino-N10-methyl-pteroylglutamic acid) is a folic acid analogue used in various immune-mediated diseases in humans. It appears to be safe and has been used in canine atopic dermatitis. Its reasonable cost could make it an option for owners who cannot afford ciclosporin (Cs). A case is reported herein of granulomatous mural folliculitis managed long-term with Cs and then with weekly subcutaneous MTX.

甲氨蝶呤（MTX，4-氨基-N10-甲基-戊基谷氨酸）是一种叶酸类似物，用于人体的各种免疫介导的疾病。它似乎是安全的，并已用于犬异位性皮炎的治疗。它价格合理，当主人无法负担环孢素（Cs）时，可作为替代。本文是一例关于肉芽肿性毛囊壁炎长期用Cs治疗，然后每周皮下MTX治疗的病例报告。

Case report

病例报告

A 13-year-old spayed female Pyrenean shepherd dog was presented with a one month history of scarring alopecia. Fluralaner (Bravecto MSD Sante Animale; Beaucouze, France) and febantel/pyrantel/praziquantel (Cestem, Ceva Sante Animale; Libourne, France) were administered seven months previously during a routine health check visit.

一只13岁的已绝育雌性比利牛斯山牧羊犬，有一个月的瘢痕性脱毛病史。在七个月前进行常规体检时，服用过氟雷拉纳和非班太尔/ 噻吩嘧啶 / 吡喹酮。

Erythema and nonpruritic alopecia began on the face one month before presentation (Day 0). The dog had received cefalexin, ketoconazole and prednisolone (unknown dosages) per os for three weeks without any improvement. During treatment, the lesions worsened and mild pruritus developed.

一个月前（第0天）面部出现皮肤发红和非瘙痒性脱毛。该犬口服头孢氨苄，酮康唑和泼尼松龙（未知剂量）三周，无任何改善。在治疗过程中，病变恶化，出现轻度瘙痒。

On physical examination, only the skin was affected. Intense erythema, patchy alopecia and irregular mottled leukoderma were observed on the face and neck. Numerous scales and crusts were overlying erosions and ulcerations. All over the body, mildly alopecic lesions, sometimes coalescing, were associated with follicular casts and leukoderma (Figure 1).

经体格检查，仅皮肤患病。在面部和颈部可见明显的皮肤发红，脱毛斑和不规则的斑驳的白斑病。大量的皮屑和结痂覆盖着糜烂和溃疡。轻度脱毛病变遍布全身，有时合并在一起，与毛囊管型和白斑病有关（图1）。

Figure 1. Clinical images of the skin lesions of a dog with granulomatous mural folliculitis.

Patchy alopecia associated with irregular mottled leukoderma was observed on the face and neck. Numerous scales and crusts were overlying erosions and ulceration. Nummular mildly alopecic lesions, sometimes coalescing associated with follicular casts and leukoderma on the trunk and the proximal extremities. (a) General view; (b) face; (c) proximal extremities; (d) trunk.

图1.肉芽肿性毛囊壁炎犬的皮肤病变的临床图像。

在面部和颈部观察到斑脱毛斑，伴有不规则的斑驳的白斑病。大量皮屑和结痂覆盖着糜烂和溃疡。轻度圆形脱毛性病变，有时与躯干和四肢近端的毛囊管型和白斑病结合在一起。 （a）概观； （b）面部； （c）四肢近端； （d）躯干。

Differential diagnoses considered included granulomatous mural folliculitis, cutaneous lupus erythematosus, dermatophytosis, sebaceous adenitis, epitheliotropic T lymphoma with follicular tropism, leishmaniosis, demodicosis and bacterial folliculitis/furunculosis.

考虑的鉴别诊断包括肉芽肿性毛囊壁炎、皮肤红斑狼疮、皮肤癣菌病、皮脂腺炎、趋上皮样T淋巴瘤伴有趋毛囊性、利什曼病、蠕形螨病和细菌性毛囊炎/疖病。

Examination of skin scrapings and plucked hairs ruled out demodicosis and did not reveal any fungal elements. Trichogram revealed follicular casts and hair shaft roots seemed distorted. Impression smears obtained from the erosive skin lesions under the crusts revealed a pyogranulomatous inflammation and no bacteria.

皮肤刮片检查和拔毛镜检排除了蠕形螨病，没有发现任何真菌元素。毛发镜检显示毛囊管型和发干根部似乎变形。从结痂下的糜烂性皮肤病变获得的皮肤压片显示为脓性肉芽肿性炎症且无细菌。

The haematological and serum biochemical findings indicated a mild regenerative, normocytic and normochromic anaemia with a marked increase in alkaline phosphatases (ALP, 837 IU/L; normal range 1–200 IU/L), most likely related to the glucocorticoid administration. Leishmaniosis serological diagnosis was negative (Immunofluorescence antibody test, IFAT).

血液学和血清生化发现表明轻度的再生性、正细胞正色素性贫血，碱性磷酸酶显着增加（ALP，837 IU / L；正常范围1-200 IU / L），最可能与糖皮质激素的给药有关。利什曼病的血清学诊断为阴性（免疫荧光抗体试验，IFAT）。

Histopathological examination of four skin biopsy specimens from lesions on the face, front legs and ventral abdomen revealed a granulomatous and lymphocytic mural and necrotizing folliculitis and furunculosis associated rarely with suppurative hidradenitis (Figures 2 and 3a). Follicular plugging was observed. Sebaceous glands were absent or obliterated by the intense infiltrate. Associated dermal inflammation was dense, superficial, perivascular and composed of lymphocytes, histiocytes with rare neutrophils and mastocytes. Mild interface dermatitis with rare basal cell apoptosis was present. Giant cells were observed. Periodic acid Schiff stain did not reveal any fungal elements on serial sectioning of the four biopsy specimens. Immunochemistry using antibodies for CD3 (Bio-Rad; Marnes-la-Coquette, France), CD204 (Abnova; Taipei, Taiwan), CD206 Novus Biologicals Europe; Abingdon, United Kingdom, IBA-1 (Abcam; Paris, France) and CD208 (DC-LAMP, Dendritics; Lyon, France) revealed that CD3+ lymphocytes were infiltrating the outer root sheath along with IBA-1+ (Ionized calcium binding adaptor molecule 1 for dendritic cells) or CD204+ cells (M1 macrophages) (Figure 3b–d).

从面部、前肢和腹部病变处对四个皮肤活检标本进行组织病理学检查，发现肉芽肿和淋巴细胞坏死性毛囊壁炎和疖病，罕见化脓性汗腺炎（图2和3a）。观察到毛囊堵塞。强烈的浸润使皮脂腺不存在或消失。伴随的皮肤炎症是致密的，浅表的，血管周围的，由淋巴细胞、组织细胞伴少量中性粒细胞和肥大细胞组成。存在轻度的界面性皮炎，罕见基底细胞凋亡。观察到多核巨细胞。高碘酸希夫（Schiff）染色在四个活检标本的连续切片上未显示任何真菌成分。使用CD3、CD204、CD206 、IBA-1和CD208抗体进行免疫组化检查。显示CD3 +淋巴细胞与IBA-1 +（树突状细胞的离子钙衔接分子1）或CD204 +细胞（M1巨噬细胞）浸润外根鞘（图3b–d）。

Figure 2. Histopathological evaluation of granulomatous mural folliculitis in a dog.

Extensive inflammation targeting the hair follicles mainly at the isthmus level, haematoxylin and eosin, *50.

图2。犬肉芽肿性毛囊壁炎的组织病理学评估。

针对毛囊的广泛炎症主要发生在毛囊峡部，苏木精和伊红染色（*50）。

Figure 3. Histopathological evaluation of granulomatous folliculitis in a dog.

(a)        Granulomatous and lymphocytic mural and necrotising folliculitis and furunculosis, haematoxylin and eosin, *200. (b) CD3+ lymphocytes (black arrow) were infiltrating the outer root sheath along with IBA-1+ (Ionized calcium binding adaptor molecule 1 for dendritic cells) (c, arrow head) or CD204+ cells (M1 macrophages) (d, star), immunochemistry *200.

图3。犬肉芽肿性毛囊壁炎的组织病理学评估。

（a）肉芽肿和淋巴细胞性坏死性毛囊壁炎和疖病，苏木精和伊红，200倍。（b）CD3 +淋巴细胞（黑色箭头）与IBA-1 +（树突状细胞的离子钙衔接分子1）浸润外根鞘 （c，箭头）或CD204 +细胞（M1巨噬细胞）（d，星形），免疫组化200倍。

While awaiting the result of the histopathological examination, the dog received clindamycin 11 mg/kg p.o. once daily and antiseptic shampoos (3% chlorhexidine digluconate and 0.5 % climbazole, DOUXO Pyo shampooing, CEVA Sante animale; Libourne, France) every other day for two weeks.

在等待组织病理学检查结果的同时，患犬使用克林霉素，剂量11 mg / kg，口服每日一次，使用抗菌香波（3％的氯己定二葡萄糖酸盐和0.5％氯咪巴唑，DOUXO Pyo香波），隔日一次，连用两周。

On Day 14, the lesions appeared drier but the alopecia and leukoderma were still expanding. At this point, the diagnosis of granulomatous mural folliculitis was made. Ciclosporin was initiated at 5 mg/kg p.o. once daily (Cyclavance, Virbac; Carros, France) and weekly subcutaneous injections of MTX 0.25 mg/kg (Methotrexate Biodim; Paris, France) for two weeks and then 0.5 mg/kg weekly for four months.

在第14天，病变干燥，但脱毛和白斑仍在扩大。此时，诊断为肉芽肿性毛囊壁炎。使用环孢素起始剂量为5 mg/kg 口服每日一次，以及每周一次皮下注射0.25 mg/kg的MTX（甲氨蝶呤），持续两周，然后每周0.5 mg/kg，持续四个月。

Four months (Day 120) after initial presentation, a marked improvement of skin lesions was observed. Hairs and whiskers were regrowing, except for hairs on the face and some areas of the abdomen and the legs. Skin biopsy samples were again collected and granulomatous and lymphocytic mural and necrotising folliculitis and furunculosis associated with suppurative hidradenitis was observed, albeit to a lesser extent compared with the first examination. As the clinical lesions were controlled, the Cs was stopped and only weekly subcutaneous MTX injections at 0.5 mg/kg were continued.

初诊后四个月（第120天），观察到皮肤病变明显改善。除了面部毛发以及腹部和腿部的某些区域外，毛发和胡须都在增长。再次收集皮肤活检样本，并观察到肉芽肿和淋巴细胞性坏死性毛囊壁炎和化脓性汗腺炎，尽管程度较第一次检查要小。由于临床病变的控制，Cs停药，仅每周一次皮下注射0.5 mg / kg MTX。

Two months later (Day 180), the only skin lesions were cicatricial alopecia on the face and in some areas of the abdomen and the legs, giving the impression of shiny smooth skin. The skin was hyperpigmented in hairless areas with some associated leukoderma (Figure 4). Skin biopsy specimens (third set) from cicatricial areas showed striking atrophy of hair follicles. Generally, hair follicle sheaths were reduced to few cords of follicular epithelial cells surrounded with melanophages, a few lymphocytes and histiocytes. Sometimes follicles and sebaceous glands were lacking, leaving only arrector pili muscles and sweat glands. Complete blood counts and serum biochemistry panels, done on a monthly basis, were within the normal range except for ALP that were still elevated (817 IU/ L). The dog continued to improve for six more months; it was then lost to follow-up.

两个月后（第180天），仅存的皮肤病变是面部、腹部和腿部一些区域的瘢痕性脱毛，外观可见皮肤光亮。无毛区域皮肤有色素沉着，伴有一些白斑病（图4）。从瘢痕区域进行皮肤活检采样（第三组）显示出明显的毛囊萎缩。普遍可见，毛囊鞘减少至少量的毛囊上皮细胞，被噬黑素细胞，少量淋巴细胞和组织细胞包围。有些缺乏毛囊和皮脂腺，仅留下竖毛肌和汗腺。每月进行的全血细胞计数和血清生化指标均在正常范围内，但ALP仍升高（817 IU / L）。该犬的皮肤病稳定了六个月。后续随访未跟进。

Figure 4. Clinical images of the skin lesions of a dog with granulomatous mural folliculitis after six months of weekly subcutaneous methotrexate and two months after stopping oral ciclosporin.

Cicatricial alopecia was present on the face and on the abdomen giving the impression of shiny smooth skin. Skin was hyperpigmented in hairless areas, sometimes associated with leukoderma. (a) General view; (b) face; (c) trunk; (d) ventral abdomen.

图4.每周皮下注射甲氨蝶呤六个月后和停止口服环孢素后两个月后，肉芽肿性毛囊壁炎犬的皮肤病变的临床图像。

面部和腹部瘢痕性脱毛，外观可见皮肤光亮。 皮肤在无毛区域色素沉着，有些伴有白斑病。（a）全身照；（b）面部；（c）躯干；（d）腹部。

Discussion

讨论

The present report describes a case of granulomatous mural folliculitis, managed long-term with Cs and MTX. In this case, the therapeutic plan was driven mostly by economic reasons because MTX was cheaper than Cs. Treatment was initiated with Cs because Cs is effective in three to six weeks, whereas MTX may take up to two to three months. Cs was stopped after several months of treatment when the lesions stabilized. No adverse effects due to any of the treatments were reported by the owner.

本报告描述了一例肉芽肿性毛囊壁炎，使用Cs和MTX长期管理。在本病例中，治疗计划主要是出于经济原因，因为MTX比Cs便宜。Cs开始治疗是因为Cs在三至六周内有效，而MTX可能需要两至三个月。治疗几个月后，病 变定后，Cs就停止使用。宠主未报告由于任何治疗引起的不良影响。

The weekly subcutaneous injection of MTX was well tolerated by the dog. One could question the real effectiveness of MTX in the present case as Cs and MTX were given together. Moreover, the behaviour of granulomatous mural folliculitis over time is not well known. Persistent elevation of ALP was noticed from the beginning and remained stable over time. As the dog was not showing any clinical signs or other serum biochemistry abnormality, further investigation was not done.

犬每周皮下注射MTX耐受性良好。本病例中，当Cs和MTX一起使用时，人们可能会质疑MTX的实际疗效。 此外，肉芽肿性毛囊壁炎随时间的表现方式尚不明确。 从一开始就注意到ALP持续升高，并且随着时间的推移保持稳定。由于该犬没有显示任何临床症状或其他血清生化异常，因此未做进一步调查。

Only a few cases of granulomatous mural folliculitis in dogs have been reported. A few were associated with adverse drug reactions. Other cases showed improvement with immunosuppressive therapy supporting an immune-mediated mechanism. Clinical lesions include patchy and multifocal, or coalescing alopecia associated with variable erythema, follicular plugging, scaling, crusting and sometimes leukoderma and leukotrichia. In more severe cases, erosion or ulceration may occur related to the mild interface dermatitis. Advanced cases reveal a very shiny smooth alopecic skin. Pruritus is not a common feature and often is associated with secondary bacterial infection. Histological findings include lymphocytes and histiocytes surrounding and infiltrating hair follicles at the isthmus level or below. In old lesions, disruption of the follicular wall and furunculosis may be associated with multinucleated giant cells.Sebaceous glands are often obliterated or totally destroyed, most probably as collateral damage, rather than being specific targets of the inflammation.

仅有少数犬肉芽肿性毛囊壁炎病例报道。有些与药物不良反应有关。其他病例显示免疫抑制治疗有所改善，支持了一种免疫介导的机制。临床病变包括斑片状以及多灶性或合并性脱毛，伴有不同程度皮肤发红、毛囊堵塞、皮屑、结痂，有时还伴有白斑病和白毛症。在更严重的病例中，可能出现与轻度界面性皮炎有关的糜烂或溃疡。晚期病例显示出脱毛皮肤非常光亮。瘙痒不常见，通常与继发性细菌感染有关。组织学发现包括在毛囊峡部或下段及周围有淋巴细胞和组织细胞浸润。在陈旧病变中，毛囊壁破裂和疖病可能伴有多核巨细胞。皮脂腺通常被破坏或完全消失，最有可能是间接损害，而不是炎症的特定目标。

Anecdotal reports have shown the effectiveness of MTX in scarring alopecia in humans but its effectiveness in immune-mediated diseases in dogs has not been reported. MTX seems to be well-tolerated in dogs, although mild gastrointestinal signs can be observed 24– 48 h after administration. A minor decrease in leukocytes was described, still within the reference range, transient elevations in liver enzymes, especially alanine aminotransferase, c-glutamyl transferase and bilirubin, were observed.The bioavailability appears to be higher following subcutaneous administration compared to the oral route and, in the present authors’ clinical experience, subcutaneous administration also helped in decreasing gastrointestinal adverse effects.

传闻报道显示，MTX对人类瘢痕性脱发有效，但尚未报道其在犬免疫介导疾病中的功效。虽然患犬对MTX耐受性良好，但在给药后24-48小时内可观察到轻度的胃肠道症状。白细胞轻度下降，但仍在参考范围内，观察到肝酶，尤其是丙氨酸氨基转移酶，c-谷氨酰胺基转移酶和胆红素的短暂升高。与口服相比，皮下注射后的生物利用度似乎更高，并且在本作者的临床经验中，皮下给药也有助于减少胃肠道不良反应。

In this case report, few aspects differ from the literature. First, drugs, vaccines and other triggers were not suspected. Second, the persistent mild pruritus was not associated with either bacterial or Malassezia infection.

在本案例报告中，几乎没有什么方面与文献有所不同。首先，没有怀疑药物、疫苗和其他诱因。其次，持续性轻度瘙痒与细菌感染或马拉色氏菌感染均无关。

Histological findings also included suppurative hidradenitis that was observed on the two first sets of skin biopsies but absent in the last one and may have been related to bacterial complications. Finally, immunophenotyping of the inflammatory population in the granulomatous mural folliculitis case herein demonstrated that CD3+ lymphocytes were infiltrating the outer root sheath along with IBA-1+ (dendritic cells) or CD204+ (M1 macrophages). This is not observed in classic bacterial furunculosis secondary to luminal folliculitis in which the histiocytes do not infiltrate the hair follicle walls.

组织学发现还包括化脓性汗腺炎，这是在前两组皮肤活检中观察到的，但在最后一组中没有，可能与细菌并发症有关。最后，本文肉芽肿性毛囊壁炎患犬的炎症免疫表型研究表明，CD3 +淋巴细胞与IBA-1 +（树突状细胞）或CD204 +（M1巨噬细胞）浸润外根鞘。这在典型的继发于毛囊腔毛囊炎的细菌性毛囊炎中没有观察到，组织细胞通常不浸润毛囊壁。

To the best of the authors’ knowledge, this is the first report of long-term management of a granulomatous mural folliculitis with Cs and MTX. MTX appears to be effective in this condition but further clinical trials are needed to assess its effectiveness in canine autoimmune diseases.

据作者所知，这是首例使用Cs和MTX长期管理肉芽肿性毛囊壁炎的报道。MTX在这种疾病中似乎有效，但是需要进一步的临床试验来评估其在犬自体免疫性疾病中的有效性。           

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