兽医病理学诊断的挑战:一只犬瘙痒与混合细胞浸润（2023）-【一例 ...

王帆

Diagnostic challenge in veterinary pathology: Pruritus in a dog with a mixed cellular infiltrate

兽医病理学诊断的挑战:一只犬瘙痒与混合细胞浸润（2023）

 

作者：Andhika Putra , Justin Stilwell， Troy Mulder, and Frane Banovic

 

翻译：王帆

 

Clinical History

临床病史

An 11-year-old, castrated male German Shepherd dog initially presented with a 9-month history of pruritus and multifocal erythematous skin lesions, alopecia, and mild weight loss. Skin lesions waxed and waned without complete resolution and newer lesions developed in other body regions. Erythematous macules and alopecia began on the ventral abdomen and progressed to the chest and dorsum. The patient received 0.27 mg/kg cetirizine (twice daily for 20 days), 22 mg/kg amoxicillin/clavulanic acid twice daily for 20 days, 0.5 mg/kg oclacitinib (once daily for 30 days), and 6 mg/kg ketoconazole (once daily for 21 days), in the prior 5 months without significant improvement. There had been no changes in the dog’s appetite or water intake. A complete blood cell count and a serum biochemistry profile performed 3 months prior to referral revealed no abnormalities and thyroid hormone T4 was within the normal reference range.

一只11岁已去势雄性德国牧羊犬，最初因瘙痒、多灶性发红、脱毛和轻度体重下降9个月就诊。皮肤病变时好时坏，未完全消退，身体其他部位出现新的皮肤病变。腹部开始出现发红和脱毛，并逐渐进展至胸部和背部。患犬在之前5个月内接受了0.27 mg/kg西替利嗪(每日2次，治疗20日)、22 mg/kg阿莫西林/克拉维酸(每日2次，治疗20日)、0.5 mg/kg奥拉替尼(每日1次，治疗30日)和6 mg/kg酮康唑(每日1次，治疗21日)治疗，但无显著改善。患犬的食欲和饮水量没有变化。转诊前3个月进行的全血细胞计数和血清生化检查未见异常，甲状腺激素T4在正常参考值范围内。

 

Gross Findings

整体检查

The dog was bright, alert, and responsive on physical examination, with abnormalities limited to the integument. Dermatologic examination revealed multifocal to coalescing erythematous macules and patches, and plaques with scale, crust, and erosions to ulcers distributed along the neck, cranio-ventral and lateral thorax, abdomen, lower extremities, footpads, and preputial region (Fig. 1).

在体格检查中，患犬聪明、警觉、有反应，异常仅限于皮肤。皮肤科检查显示，沿颈部、颅腹侧和胸侧、腹部、下肢、爪垫和包皮分布的多灶性至融合性发红和斑片，以及伴有皮屑、结痂和糜烂至溃疡的斑块(图1)。

 

Differential Diagnosis

鉴别诊断

A neoplastic process such as cutaneous lymphoma was the primary differential diagnosis due to the history, age of onset, and clinical presentation. Generalized superficial pyoderma in conjunction with atopic dermatitis and pemphigus foliaceus were considered differential diagnoses. Both superficial pyoderma and pemphigus foliaceus skin lesions can be pruritic and show signs of erythema and crusting. Reactive histiocytosis was considered less likely as a differential diagnosis due to the lack of nodules.

根据病史、发病年龄和临床表现，肿瘤疾病(如皮肤淋巴瘤)是主要的鉴别诊断。全身性浅表脓皮病合并特应性皮炎和落叶型天疱疮被认为是鉴别诊断。浅表脓皮病和落叶型天疱疮的皮肤病变都可能瘙痒，并表现出发红和结痂的症状。由于缺乏结节，反应性组织细胞增生症被认为不太可能是鉴别诊断。

 

Laboratory Findings

实验室结果

Skin cytology using Diff Quick stain revealed an accumulation of neutrophils and few corneocytes. There were no observed acantholytic keratinocytes or microorganisms. Four, 8-mm punch biopsies were obtained from the inguinal, neck, and cranio-ventral and lateral thoracic skin lesions; fixed in formalin; and processed for histological examination. Limited clinical history was received with biopsy submission.

皮肤细胞学使用Diff Quick染色显示中性粒细胞和少数角质形成细胞的聚积。未观察到棘层松解性角质形成细胞或微生物。对腹股沟、颈部、颅腹侧和胸侧皮肤病变进行8 mm打孔采样，共四块;用福尔马林固定;并进行组织学检查。有限的临床病史和活检提交。

 

Microscopic Findings

显微镜检查结果

Microscopic findings included multifocal, poorly delineated, infiltrates of a mixed population of neutrophils, small mature lymphocytes, plasma cells, histiocytes, eosinophils, and mast cells. Inflammation and edema expanded the superficial dermis and loosely surrounded adnexa superficially but did not extend into the subcutis (Fig. 2a). Depending on the section, the epidermis exhibited multifocal erosion, orthokeratotic hyperkeratosis, serocellular crusts, multifocal subcorneal pustules, and moderate hyperplasia (Fig. 2b, c). Apocrine glands were mildly ectatic (Fig. 2a, d). Special stains (Gram, Ziehl-Neelsen, Periodic-Acid-Schiff-Hematoxylin) did not reveal the presence of infectious agents.

显微镜下观察结果包括多灶性、边界不清的中性粒细胞、小的成熟淋巴细胞、浆细胞、组织细胞、嗜酸性粒细胞和肥大细胞混合浸润。炎症和水肿使真皮浅层扩张，附件周围松弛，但未延伸至皮下组织(图2a)。根据切片不同，表皮表现为多灶性糜烂、正角化性角化过度、血清性结痂、多灶性角质层下脓疱和中度增生(图2b、c)。顶浆汗腺轻度扩张(图2a、d)。特殊染色(革兰式、ZN染色、PAS-苏木精)未显示感染性病原的存在。

 

An initial diagnosis of mixed, multifocal, superficial dermatitis was made based on the presence of a loosely organized, mixed inflammatory infiltrate multifocally present within the dermis. The primary differential diagnoses were allergic dermatitis with potential secondary bacterial infection, pemphigus foliaceus with secondary infection, though no acantholytic cells were identified, or another immune mediated process such as reactive histiocytosis, with an atypical histopathologic presentation.

根据真皮层内多灶性组织松散的混合性炎症浸润，初步诊断为混合性、多灶性浅表性皮炎。主要鉴别诊断包括过敏性皮炎伴潜在继发细菌感染、落叶型天疱疮伴继发感染(但未发现棘层松解细胞)或其他免疫介导过程(如反应性组织细胞增生症)，且组织病理学表现不典型。

 

Further Investigation and Diagnosis

进一步调查和诊断

Additional clinical history was discussed between the clinician and the pathologist after apparent discordance between clinical suspicion of a neoplastic process and histopathologic findings of mixed inflammation. Re-evaluation identified atypical, intermediate to large round cells initially interpreted as histiocytes scattered throughout the dermis and surrounding, but not infiltrating, adnexa (Fig. 2d). These round cells, amongst the dermal rarely infiltrated the overlying epidermis. The cells had distinct borders with low to moderate amounts of eosinophilic cytoplasm; round to ovoid, central nuclei with open chromatin; and 1 or 2 prominent nucleoli (Fig. 2e). Anisocytosis and anisokaryosis were mild to moderate with an occasionally high nuclear to cytoplasmic ratio and rare binucleated to multinucleated cells. The mitotic count for the atypical round cells was 4 per 2.37 mm2 with occasional bizarre mitoses. On immunohistochemistry, the atypical round cells exhibited weak to strong, membranous to cytoplasmic immunoreactivity to CD3 (Fig. 2f). The round cells were separated by mixed inflammatory populations, as previously described. Fewer, small mature lymphocytes within the neoplastic population exhibited strong, membranous immunoreactivity to CD20 and CD79.

在临床怀疑肿瘤过程和组织病理学发现混合性炎症之间明显不一致之后，临床医师和病理学家讨论了其他临床病史。重新评估发现了非典型的中间到大圆形细胞，最初解释为组织细胞散在分布于真皮层和周围，但没有浸润附件(图2d)。这些位于真皮之间的圆细胞很少浸润到上面的表皮。细胞边界清楚，胞质低至中等量嗜酸性;圆形到卵圆形，具有开放染色质的中央核;和1个或2个明显的核仁(图2e)。细胞不等和核不等轻至中度，偶见高核质比，罕见双核和多核细胞。不典型圆形细胞的有丝分裂象为4个/ 2.37 mm2，偶有奇怪的有丝分裂象。在免疫组织化学上，不典型圆形细胞对CD3表现出弱到强、细胞膜到细胞质的免疫反应性(图2f)。如前所述，利用混合炎症细胞群分离圆形细胞。少数成熟的小淋巴细胞对CD20和CD79有较强的细胞膜免疫反应性。

 

These findings indicated that both inflammatory and neoplastic processes might be involved. Therefore, a polymerase chain reaction for antigen receptor rearrangement (PARR) test1, (Veterinary Diagnostics, Davis, CA, USA) was pursued on paraffin-embedded sections to confirm the disease process and diagnosis. This test was performed in triplicate using primer sequences to canine T-cell receptor gamma TRG(A), TRG(B), and TRG(C) and analyzed using electrophoresis as previously reported.Sharp clonal spikes were identified in duplicate analysis for TRG(A), with no clonal spikes seen for TRG(B) or TRG(C).

这些结果表明炎症和肿瘤过程可能都参与其中。因此，我们在石蜡包埋切片上进行了抗原受体重排(PARR)聚合酶链反应检测，以确定疾病过程和诊断。该试验使用犬T细胞受体γ TRG(A)， TRG(B)和TRG(C)的引物序列进行3个重复，并使用电泳分析。TRG(A)重复分析发现明显的克隆棘，TRG(B)和TRG(C)未发现克隆棘。

 

Discussion

讨论

The heavily inflamed, mixed cellular infiltrate and significantly limited clinical history provided with the biopsy submission lead to an initial diagnosis of an inflammatory process. Communication of additional clinical history and suspicion of a neoplastic process between the clinicians and pathologists was necessary to reach the final diagnosis of canine inflamed, nonepitheliotrophic cutaneous T-cell lymphoma (NE-CTCL). The diagnosis of inflamed NE-CTCL was supported by a combination of clinically distinctive, progressive, erythematous skin lesions in an aged dog, histopathologic findings of atypical CD3+ neoplastic round cells within a mixed inflammatory cell population, and a clonal PARR test revealing a sharp spike indicating monoclonal cell population.

活检样本提供的严重炎症、混合细胞浸润和明显有限的临床病史导致了炎症过程的初步诊断。为了最终诊断为犬炎性、非趋上皮性皮肤T细胞淋巴瘤(NE-CTCL)，临床医师和病理学家之间需要交流更多的临床病史和对肿瘤过程的怀疑。炎性NE-CTCL的诊断依据如下:临床特征性的、进展性的老年犬发红皮肤病变，组织病理学发现非典型CD3+肿瘤圆细胞在混合炎症细胞群中，克隆性PARR检测显示单克隆细胞群。

 

Pruritus was a unique feature of the clinical presentation not previously associated with cases of inflamed NE-CTCL. Pruritus potentially augmented inflammation due to selftrauma. Pruritus is nonspecific and is commonly associated with allergic or immune-mediate dermatitis, which were suspected from initial interpretation of microscopic findings. Therefore, this case illustrates how skin lesions of inflamed NE-CTCL can be accompanied by pruritus and mimic other inflammatory dermatoses.

瘙痒是一种独特的临床表现，以前与炎性NE-CTCL病例不相关。瘙痒症可能会加剧自我损伤引起的炎症。瘙痒不是特异性的，通常与过敏性或免疫性皮肤病相关，而最初对显微镜检查结果的解读怀疑这些皮肤病。因此，该病例说明了炎性NE-CTCL的皮肤病变如何伴随瘙痒，并与其他炎性皮肤病相似。

 

The neoplastic cells were initially considered histiocytes within a mixed inflammatory cell infiltrate. The overall pattern of chronic, superficial pustular dermatitis was suggestive of allergic or immune-mediated dermatitis (pemphigus foliaceus) with secondary pyoderma or chronic surface trauma obscuring the primary process. Although amastigotes were not identified microscopically, the superficial distribution and heterogeneous inflammation were consistent with cutaneous leishmaniasis. These differentials were ruled out based on a lack of various key gross and microscopic findings and identification of the atypical round cells as CD3+, intermediate to large lymphocytes.

肿瘤细胞最初被认为是混合炎症细胞浸润中的组织细胞。慢性浅表脓疱性皮炎的总体模式提示过敏性或免疫性皮肤病(落叶型天疱疮)，继发脓皮病或慢性表面创伤掩盖了原发过程。虽然显微镜下未发现无鞭毛体，但其浅表分布和异质性炎症符合皮肤利什曼病。基于缺乏各种关键的肉眼和显微镜观察结果，以及不典型圆形细胞被鉴定为CD3+，中间到大淋巴细胞，这些鉴别诊断被排除。

 

Histologically, typical cases of inflamed NE-CTCL are characterized by a homogenous dermal infiltrate of large, atypical and variably CD3+ lymphocyte population. However, the diagnosis of NE-CTCL can be challenging when a heterogeneous cell population is present, which most commonly involves histiocytes, lymphocytes, and eosinophils. In this case, additional inflammatory cells including large numbers of neutrophils, mast cells, and plasma cells, were present and obscured the neoplastic population. Histological differentiation between reactive histiocytosis and inflamed NE-CTCL can be challenging as both canine-reactive histiocytosis and inflamed NE-CTCL are characterized by heterogeneous inflammatory populations.5 Inflammatory and neoplastic conditions may contain low numbers of mitotic figures, though the presence of bizarre mitoses could indicate a neoplastic process. Using an immunohistochemical panel to differentiate histiocytes from poorly differentiated, intermediate to large lymphocytes and clonality testing may improve chances for detecting the neoplastic population in these cases.

组织学上典型的炎性NE-CTCL表现为真皮内大量异型的CD3+淋巴细胞浸润。然而，当存在异质性细胞群(最常见的是组织细胞、淋巴细胞和嗜酸性粒细胞)时，NE-CTCL的诊断可能具有挑战性。在此例中，存在其他炎症细胞，包括大量中性粒细胞、肥大细胞和浆细胞，从而掩盖了肿瘤细胞。反应性组织细胞增生症和炎性NE-CTCL的组织学鉴别可能具有挑战性，因为犬反应性组织细胞增生症和炎性NE-CTCL的特征都是异质性炎性细胞。炎症和肿瘤性疾病可能包含少量的有丝分裂象，但奇异的有丝分裂的存在可能表明肿瘤过程。使用免疫组化组合将组织细胞从低分化、中等到大淋巴细胞的分化，以及克隆性检测可能提高检测这些病例中肿瘤疾病的机会。

 

Lymphocyte clonality assessment using PARR testing has been utilized to discern inflammation and inflamed NE-CTCL. Polymerase chain reaction for antigen receptor rearrangement testing detects clonal expansion of lymphocytes and is a highly sensitive and specific diagnostic tool for canine lymphoma, with around 90% sensitivity and specificity for most T-cell lymphomas. However, lymphocyte clonal proliferation in histopathological skin samples is not an exclusive hallmark of neoplasia and can be observed with other lymphocytic inflammatory skin diseases. Therefore, PARR testing as a single tool is not a standalone diagnostic test to diagnose malignancy, despite its high sensitivity and specificity, and PARR results should always be interpreted in the context of the clinical, histopathological, and immunophenotypic findings.

使用PARR检测的淋巴细胞克隆性评估已被用于识别炎症和炎性NE-CTCL。用于抗原受体重排检测的聚合酶链反应可检测淋巴细胞克隆扩增，是一种对犬淋巴瘤具有高度敏感性和特异性的诊断工具，对大多数T细胞淋巴瘤的敏感性和特异性约为90%。然而，组织病理学皮肤样本中的淋巴细胞克隆性增殖并不是肿瘤的唯一标志，也可在其他淋巴细胞性炎性皮肤病中观察到。因此，尽管PARR检测具有较高的敏感性和特异性，但作为单一工具的PARR检测并不是诊断恶性肿瘤的独立诊断方法，而且应始终结合临床、组织病理学和免疫表型结果来解读PARR结果。

 

The prognosis for inflamed NE-CTCL is generally guarded to poor, with survival time ranging from 1 to 36 months after diagnosis. Dogs with NE-CTCL have longer overall survival times than dogs with epitheliotropic CTCL. At the time of this writing and 4 months after diagnosis, the owners of this dog elected not to pursue any therapy as the dog remained systemically healthy. However, the skin lesions are still present and new skin lesions are still developing.

炎性NE-CTCL的预后一般较差，确诊后生存时间为1-36个月。NE-CTCL患犬比趋上皮性CTCL患犬的总生存时间更长。在撰写本文时和诊断后4个月，这只犬主人选择不进行任何治疗，因为犬保持全身健康。但皮肤病变仍存在，新的皮肤病变仍在发展。

 

Inflamed NE-CTCL represents a diagnostic challenge due to its histopathologic resemblance to inflammatory processes. Providing an adequate clinical history with descriptions and distributions of lesions, and suspected differential diagnoses with the biopsy submission is essential to making an accurate diagnosis. Even so, multiple methodologies may be required for diagnostic confirmation of inflamed NE-CTCL, including immunohistochemistry and PARR testing.

炎性NE-CTCL具有与炎症过程相似的组织病理学特征，给诊断带来了挑战。提供充分的临床病史、病变描述和分布，以及可疑的鉴别诊断和活检是做出准确诊断的关键。尽管如此，炎性NE-CTCL的诊断确认可能需要多种方法，包括免疫组织化学和PARR检测。

 

 

Figure 1. Erythematous macules and patches, and plaques with erosion, crust, and alopecia, cervical and thoracic skin, German Shepherd dog: (a) multifocal to coalescing erythematous macules and patches, and plaques with scaling, crusting, and ulcerations distributed along the neck and (b) higher magnification of similar lesions along the lateral thorax.

图1。颈部和胸部皮肤的发红斑点和斑片，以及伴有糜烂、结痂和脱毛的斑块，德国牧羊犬:(a)沿颈部分布的多灶性至融合性发红斑点和斑片，以及伴有皮屑、结痂和溃疡的斑块;(b)沿胸侧的类似病变放大照片。

 

 

Figure 2. Inflamed, nonepitheliotropic T-cell lymphoma, skin, dog: (a) superficial infiltrates (arrows) expanding the dermis and apocrine gland ectasia (Hematoxylin and eosin [HE]). (b) Subcorneal pustules within the epidermis (arrow) and dermal expansion by edema, neoplastic round cells, and inflammation (HE). (c) Neoplastic round cells and mixed inflammation expand the dermis (HE). (d) Low numbers of inflammatory and neoplastic round cells surround, but do not invade, adnexal structures (HE). (e) Atypical round cells with mixed inflammation and occasional mitotic figures (arrows) (HE). (f) Membranous and cytoplasmic round cell immunolabeling of CD3.

图2。炎性非趋上皮性T细胞淋巴瘤，皮肤，犬:(a)真皮扩张的浅表浸润(箭头)和顶浆汗腺扩张(苏木精-伊红[HE]染色)。(b)表皮层角质层下脓疱(箭头)和由水肿、肿瘤性圆细胞和炎症引起的真皮扩张(HE)。(c)肿瘤性圆细胞和混合炎症使真皮层扩张(HE)。(d)附件结构(HE)周围有少量炎性和肿瘤性圆细胞，但不侵袭。(e)非典型圆形细胞混合炎症，偶见有丝分裂象(箭头)(HE)。(f) CD3免疫标记圆细胞膜和胞质。

 

 

 

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