Canine Histiocytic Diseases

CE Article #1

Alastair R. Coomer, BVSc, MS University of Florida

Julius M. Liptak, BVSc, MVetClinStud, FACVSc, DACVS, DECVSa Ontario Veterinary College, University of Guelph

ABSTRACT: Canine histiocytic diseases are an emerging spectrum of diseases characterized by proliferations of histiocytic cells. Nonneoplastic histiocytic disease (reactive histiocytosis, comprising cutaneous and systemic histiocytosis) is uncommon. Neoplastic histiocytic diseases include cutaneous histiocytoma, which is a benign histiocytic tumor, and localized and disseminated histiocytic sarcoma (previously known as malignant histiocytosis), which are malignant diseases.The differentiation of histiocytic diseases can be challenging.This article outlines the characteristics of each disease entity and details the clinicopathologic, histologic, immunohistochemical, prognostic, and therapeutic differences among them.

istiocytic diseases are a commonly diag- reactive (cutaneous or systemic) histiocytosis nosed but poorly understood spectrum (nonmalignant, nonneoplastic disease); (2) cuta- Hof diseases in dogs and other species.1,2 neous histiocytoma (nonmalignant, neoplastic dis- Several different documented canine histiocytic ease); (3) localized histiocytic sarcoma (LHS; proliferative diseases may be variations of the malignant, neoplastic disease); and (4) dissemi- same disease or derived from cells of the same nated histiocytic sarcoma (DHS; malignant, neo- lineage.3 Published case reports and small case plastic disease) (see the box on page 203).2,3,16 series provide some pertinent information on Reactive histiocytosis results from immune system treatment and prognosis, but few concise and dysregulation. Cutaneous histiocytoma is a com- thorough reviews outline the specifics of these mon, benign tumor of the adnexa.17 DHS is rare.17 diseases.1,4–15 Furthermore, much confusion sur- Histiocytes are tissue cells whose functions rounds the terminology, presentation, and true (primarily phagocytosis and antigen presen- prognosis of specific histiocytic diseases. tation) are related to their interactions with lym- PROOF18 phocytes in inflamed tissues. Histiocytes may CLASSIFICATION be classified as macrophages or as dendritic anti- Canine histiocytic diseases have been classi- gen-presenting cells (DAPCs). Macrophages are fied into three major categories: nonmalignant involved in the inflammatory process, contain nonneoplastic, nonmalignant high levels of lysosomal enzymes, and are capa- neoplastic, and malignant neo- ble of varying degrees of phagocytosis.18 The plastic. These three categories term DAPC encompasses follicular dendritic •Take CE tests encompass four syndromes: (1) cells, Langerhans cells, interstitial dendritic cells, • See full-text articles aDr. Liptak is now affiliated with veiled cells, and interdigitating dendritic cells. All Alta Vista Animal Hospital in of these cells have typical dendritic morphology CompendiumVet.com Ottawa, Ontario. and low levels of lysosomal enzymes.18 They are

COMPENDIUM 202 April 2008 Canine Histiocytic Diseases CE 203 mostly incapable of phagocytosis Classification of and play a key role in antigen pres- Histiocytic Diseases 4 entation to lymphocytes. Nonmalignant Nonneoplastic • Reactive histiocytosis SPECIMEN PREPARATION — Cutaneous histiocytosis AND ANALYSIS — Systemic histiocytosis Definitive diagnosis of histio- Nonmalignant Neoplastic cytic disease depends on identify- • Cutaneous histiocytoma ing the standard clinicopathologic and morphologic characteristics of Malignant Neoplastic histiocytes. Tissue specimens sub- • Localized histiocytic sarcoma mitted for immunohistochemistry • Disseminated histiocytic sarcoma and cytology can be snap frozen or fixed with formalin. The advantages of snap freezing are a wide array of available stains and the increased likelihood of a definitive diagnosis. How- ever, the equipment required for snap freezing (e.g., liquid nitrogen, isopen- tane, optimal cutting temperature compound) and for storage and transport of snap-frozen tissue is not widely available to general practitioners. Forma- lin fixation offers the next best alternative.2 Special stains for lysozyme and α1-antitrypsin, when positive, identify cells as being histiocytic rather than lymphoid or epithelial in origin.2 Cytologic characteristics of malignancy are then used to differentiate malignant from benign disease. Cellular phenotype, which can be determined by immunophenotyping, has become the mainstay for definitive diagnosis of histocytic diseases (Table 1). Immunophenotyping involves the use of monoclonal antibodies to differentiate specific surface antigens on lymphocytes and other cells of the immune system.2,5,19 Lymphocytes generally express either CD3 or CD79a and CD18 antigens.2 Histiocytes do not express CD3 or CD79a, but they have 10-fold more CD18 than do lymphocytes.2 Histiocytes, therefore, can be accurately identified as those cells that have characteristic morphology, lack lymphoid immunophenotypic markers, and have abundant CD18.2,6 If the specimen is snap frozen, immunohistochemical markers for CD3, CD11d, CD18, CD45RA, and CD79a are used to definitively determine whether a lesion is of histiocytic origin. Histiocytes in snap-frozen samples should strongly express CD18 but lack expression of CD3, CD11d, CD45RA, and CD79a.18 This panel (CD18+, CD3-, CD11d-, CD45RA-, CD79a-) differentiates histiocytic disease from epitheliotrophic lymphoma (CD3+PROOF and CD79a+), but it cannot differentiate between the different histiocytic diseases because all histiocytes are positive for CD18. If the specimen has been fixed in formalin, then CD18 is the only reliable marker that can be used to diagnose histiocytic disease. Further differentiation of histiocytes into those of macrophage or DAPC origin requires identification of positive CD11b, CD14, and CD68 markers or CD1, CD11c, major histocompatibility complex class II (MHC II), and intercellular adhesion molecule 1 (ICAM-1) markers, respectively.2 DAPCs can also be distinguished as activated or nonactivated based on whether they stain positively or negatively for CD4 and subtypes of CD1, depending on the disease present. These markers can be identified only in snap-frozen specimens. It is unknown whether definitive diagnosis of the cell of origin influ-

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Table 1. Immunohistochemical Markers andTissue PreparationTechniques Used to Identify Cellular Origin Cellular Origin Immunohistochemical Markers Tissue Preparation Leukocyte CD11a, CD44 Frozen CD18, CD45 Fixed or frozen

Lymphocyte CD3, CD79a (Mb-1) Fixed or frozen • B lymphocyte CD21-like Frozen • Reactive lymphocyte CD8 (α, β), TCRαβ Frozen • T lymphocyte TCRγδ Frozen Macrophagea CD11b, CD18, CD14, CD68 Frozen Myeloid DAPCa CD1 (a, b, c), CD11c, CD18, CD54 (ICAM-1), MHC II Frozen Myeloid DAPC (activated)a CD1 (a, b, c), CD4, CD11c, CD54 (ICAM-1), Frozen CD90 (Thy-1), MHC II aHistiocyte ICAM = intercellular adhesion molecule, Mb = B-cell marker, MHC = major histocompatibillity complex, TCR = T-cell receptor ences the clinical management of and prognosis for the of immune system dysregulation and primarily affects various histiocytic diseases in dogs. Clinicians should the skin and subcutis. It is characterized by multiple contact individual commercial veterinary pathology labo- nonpruritic, painless cutaneous plaques and nodules that ratories regarding the availability and preparation of tis- are predominantly located on the head, neck, perineum, sues for immunohistochemical stains (see the box below). scrotum, and extremities.16,18,20–23 In dogs with SH, lesions are also evident in the nasal cavity (mucosae), NONMALIGNANT,NONNEOPLASTIC palpebrae, sclera, lung, spleen, liver, and bone marrow. DISEASE Multiple lymph node involvement is also common.16 Canine reactive histiocytosis, whether systemic histio- SH is more commonly observed in Bernese mountain cytosis (SH) or cutaneous histiocytosis (CH), is a result dogs, rottweilers, and golden and Labrador retrievers than in other breeds. Patient age at diagnosis of SH is Collecting and Submitting Samples usually 4 to 7 years; male dogs are more frequently for Histopathology reported affected than are females. No breed or sex predilections are seen in dogs with CH; in these dogs, When formalin is used, sample collection and processing age at diagnosis ranges from 3 to 9 years. 16,18,20–23 for suspected histiocytic disease is similar to that for traditional surgical pathology submissions (i.e., ink the The major clinical differentiation between CH and margins if necessary and let them dry,place the sample SH is the number of concurrent lesions and the simulta- in formalin, and send it to the laboratory). Clinics or neous multiorgan involvement in dogs with SH.2,18 institutions that can run snap-frozen sections usually accept With SH, because multiple organ systems are involved, freshPROOF (i.e., not fixed) samples and perform the snap freezing clinical signs correspond with lesion location and often themselves. If a clinic has the necessary equipment for snap include anorexia, marked weight loss, stertorous respira- freezing but not the ability to stain samples, then frozen tion, conjunctivitis, and chemosis. Physical examination samples can be submitted to outside laboratories for specific immunohistochemistry.For more guidance on sample reveals multiple cutaneous and subcutaneous nodules or preparation and whether a specific surgical pathology plaques (up to 4 cm in diameter) on the scrotum, nasal laboratory can conduct immunohistochemistry,clinicians planum, nasal apex, and eyelids. The lesions and clinical should call the laboratory,say that they have a patient with course of reactive histiocytosis are generally slowly pro- a suspected histiocytic mass or tumor and would like to gressive. In rare cases, the lesions may regress sponta- conduct immunohistochemistry,and ask whether the neously, especially during the early stages of disease.1,16 laboratory offers special staining and how the sample The skin lesions of CH and SH are histologically should be prepared to maximize the diagnostic yield. identical and are characterized by an angiocentric, pleo- (text continues on page 208)

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Table 2. Immunohistochemistry Panels Used to Diagnose Histiocytic Disease Immunohistochemistry Markers Common Immunohistochemistry Markers Histiocytic Disease to All Histiocytes Specific to Histiocytic Disease Reactive histiocytosis CD11c+, CD18+, MHC II+, ICAM-1+, CD1+, CD4+, CD8+, CD11b+, CD3-, CD45RA-, CD79a- CD90 (Thy-1)+, CD11d- Cutaneous histiocytoma CD11c+, CD18+, MHC II+, ICAM-1+, CD1+, CD4-, CD11d-, CD90 CD3-, CD45RA-, CD79a- (Thy-1)- Localized histiocytic sarcoma CD11c+, CD18+, MHC II+, ICAM-1+, CD1b+, CD1c+, CD4+, CD90 CD3-, CD45RA-, CD79a- (Thy-1)±, CD11d± Disseminated histiocytic sarcoma CD11c+, CD18+, MHC II+, ICAM-1+, CD1b+, CD1c+, CD4+, CD90 CD3-, CD45RA-, CD79a- (Thy-1)±, CD11d±

(continued from page 204) cellular infiltrate composed of lymphocytes, neutrophils, usually prolonged; therefore, patients require prolonged and activated interstitial, myeloid, perivascular DAPCs immunosuppressive therapy. While dogs rarely die of of the dermis.2,3,18 In SH, cutaneous lesions may also be their disease, the need for chronic medication and the characterized by a multinodular, coalescing-to-diffuse frequent recurrence of disease result in most affected pleocellular infiltrate, predominantly in the deep dermis dogs being euthanized.18,21–23 In one report, therapeutic and subcutis. The infiltrate consists of large round-to- success and induction of remission were reported in only oval histiocytes with large monomorphic nuclei. This 6 of 26 dogs with SH treated with cyclosporine A, histologic appearance is consistent with a nonneoplastic leflunomide, or doxorubicin.18 These dogs required reactive process.16 Lesions often form dense perivascular long-term or intermittent therapy to prevent recurrence. cellular cuffs; this is the major histologic difference In this report, systemic corticosteroid therapy failed to between reactive histiocytosis lesions and cutaneous his- induce regression of lesions in most dogs. tiocytoma. Immunophenotyping of snap-frozen specimens reveals dense accumulations of activated DAPCs NONMALIGNANT,NEOPLASTIC that stain positive for the immunohistochemical mark- DISEASE ers CD1, CD4, CD8, CD11b, and CD90 (Thy-1); this Cutaneous histiocytoma is a benign epitheliotropic also supports a nonneoplastic reactive process (Table neoplasm originating from intraepidermal DAPCs 2).2,16 However, these markers are not specific for neo- (Langerhans cells).2 Canine cutaneous histiocytoma plastic or nonneoplastic disease. occurs most frequently in dogs younger than 3 years, In dogs with CH or SH, surgical excision is successful although it is occasionally diagnosed in older dogs.20 A for local disease but does not prevent the development breed predisposition for multiple cutaneous histiocy- of new lesions.18 Immunosuppressive and immunomod- tomas has been reported in shar-peis.16,20 ulatory drugs have been used with mixed results. Cutaneous histiocytomas appear as small, firm, dome- or Because SH and CH are caused by immune system button-shaped dermoepithelial masses of the head, pinnae, dysregulation,PROOF they may respond favorably to immuno- neck, or limbs (Figure 1).2 They are fast growing, nonpru- suppressive therapy; however, this therapy must be ritic, painless, usually solitary, and often less than 2.5 cm in prolonged (i.e., lasting many months).18 Response to diameter.16,20 Solitary lesions may spontaneously regress corticosteroids varies but may be as high as 50% in dogs within 6 weeks; multiple histiocytomas may persist longer with CH.18,24 Cyclosporine A and leflunomide are gen- because of overlap between the regression of old masses erally more successful in treating reactive histiocytosis and development of new masses.16 Persistent nodules may and are necessary in cases refractory to corticosteroids.18 ulcerate and pose a risk for bacterial infection. Regional Ocular lesions may be difficult to resolve and may lymphadenomegaly (due to migrating histiocytoma cells) require ophthalmic preparations of cyclosporine A (1 has been reported with cutaneous histiocytoma and usually drop q12h of 0.2% to 1.0% drops).16 resolves with the skin lesions16; however, if regional lym- Dogs with CH or SH have a guarded prognosis phadenopathy is present, malignant histiocytic disease because the clinical course of reactive histiocytosis is should be suspected.16 Nodal biopsy with immunohisto-

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chemistry is required to rule out malignant histiocytic disease. Cytologic characteristics of samples aspirated from histiocytomas include clumps of round or oval cells with a variable amount of pale basophilic cytoplasm (lacking vacuolation) and round-to-ovoid, eccentrically placed nuclei with finely granular chromatin and inconspicuous nucleoli. Histologically, cutaneous histiocytomas appear as poorly demarcated, nonencapsulated, intracutaneous nodules and are therefore difficult to differentiate from nonepitheliotropic or epitheliotropic cutaneous lymphoma (mycosis fungoides).20 Unlike the perivascular aggregation of reactive histiocytosis, cutaneous histiocytomas are organized into uniform sheets of histiocytes that penetrate the dermis or subcutis. The diffuse popu- Figure 1. A cutaneous histiocytoma lesion on the pinna of a dog. lation of monomorphic round cells with large, round-to- oval, indented or twisted vesicular nuclei may be densely packed in the deeper layers of the dermis.16 Collagen fibers and skin adnexae may be displaced. Because of the MALIGNANTDISEASE epidermal invasion and intraepidermal aggregation of Localized Histiocytic Sarcoma lymphocytes, immunohistochemical phenotyping may be LHS is a rapidly growing, solitary, locally invasive soft required to rule out epitheliotropic lymphoma.16,20 Cuta- tissue mass with moderate-to-high metastatic poten- neous histiocytomas stain positive for CD1 but, unlike tial.2,6,18 LHS has been reported in many breeds, with reactive histiocytosis samples, usually stain negative for flat-coated retrievers, Bernese mountain dogs, Labrador CD4 and CD90 (Thy-1).18 This phenotype is consistent retrievers, and rottweilers being overrepresented.2,11,18 with that of nonactivated epidermal DAPCs18 and is a Most affected dogs are 6 to 11 years of age. major difference between cutaneous histiocytoma and Lesions may arise from a single site. Predilection sites reactive histiocytosis. However, the absence of CD4 include the subcutis and underlying tissues of the staining is not definitive for cutaneous histiocytoma. extremities. Primary lesions may also be found in the While surgical excision or cryosurgery is generally periarticular soft tissues, spleen, lung, brain, nasal cavity, curative, histiocytomas may spontaneously regress and bone marrow.2,3,7,9,18,25–27 Periarticular lesions often within 6 weeks (for solitary lesions) to 10 months (for encircle the joints and involve the joint capsule, tendons, multiple lesions).16,20 Surgical excision is recommended and skeletal muscles.16 LHS is locally invasive, fre- for any solitary mass that has not spontaneously quently invading the deep dermis and the underlying regressed within 3 months.16,20 Because immunosuppres- skeletal muscle and fascia. Metastatic lesions, if present, sive drugs may interfere with spontaneous regression by are seen in draining lymph nodes, with a 60% metastatic inhibitingPROOF T-cell infiltration, their use is contraindi- rate reported in one study.2,3 If organ systems other than cated; hence, every effort should be made to differenti- the skin (e.g., the spleen) are involved, a more aggres- ate cutaneous histiocytoma from nonneoplastic reactive sive, hemophagocytic form of the disease is likely and histiocytosis.16 the metastatic rate is increased, as is the clinical suspi- After surgical excision or spontaneous regression of cion of DHS.20 single or multiple lesions, the prognosis for dogs with Clinical signs are generally related to local disease and histologically confirmed cutaneous histiocytoma is excel- the organ of involvement. Most dogs with LHS present lent. The marked difference between the prognosis for with palpable cutaneous or subcutaneous masses and are this neoplastic disease and that for nonneoplastic reactive not systemically ill, unlike dogs with DHS.3 Thorough histiocytosis is due to the fact that therapy for cutaneous clinical staging includes palpation and aspiration of histiocytoma is curative and limited to one procedure regional lymph nodes, bone marrow aspiration, abdomi- rather than prolonged and immunosuppressive. nal ultrasonography, and three-view thoracic radiogra-

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phy to assess for multiorgan involvement and metastasis and to differentiate LHS from DHS.25,27 The typical histologic findings for LHS include an invasive, highly cellular mass of pleomorphic histiocytic cells. These cells are characterized by anisocytosis with abundant eosinophilic and finely granular to foamy cytoplasm. The nuclei vary markedly in form and size and are often located cen- trally with prominent nucleoli of varying sizes. Multinucleated cells and mitotic figures, including those with abnormal forms, are frequently observed.3,4,7,16 Immunohistochemical evaluation often yields low-to-moderate numbers of lysozyme-positive cells and large numbers of vimentin-positive cells.3,7 All cells should be negative for both T- (e.g., CD3) and B-lymphocyte (e.g., CD79) markers.3,7,28 Nonactivated DAPCs stain positively for the immunohistochemical markers CD1b, CD1c, and, rarely, CD90 (Thy-1).2,10,18 Some histiocytic sarcomas also stain positively for CD11d; however, it is not known whether this finding is unique to histiocytic sarcomas.16 The critical difference between the immunophenotype of malignant LHS and that of benign cutaneous histiocytoma is expression of CD4, a marker for activation of myeloid DAPCs. Many more cells will be CD4 positive in cases of LHS compared with cases of cutaneous histiocytoma.2,18 Aggressive surgical resection of cutaneous LHS is recommended (Figure 2).2 Surgical margins should be the same as those used for soft tissue sarcomas: 3-cm lateral margins and a deep margin of at least one fascial layer.29 Limb amputation is recommended for nonmetastatic periarticular LHS.30 Radiation therapy has not been investigated in the treatment of gross or microscopic incompletely resected LHS, but other round cell tumors (e.g., lymphoma, mast cell tumors) are sensitive to irradiation.2 Hence, full-course fractionated radiation therapy protocols should be considered for incompletely resected or unresectable LHS. Because the potential for distant metastasis is moderate to high, postoperative chemotherapy should also be considered.18 However, the efficacy of different postoperative chemotherapy drugs and protocols in the treatment of dogs with LHS has not been evalu- ated. Based on the current treatment recommendations for DHS, single- agent protocols using either doxorubicin or lomustine are suggested.24 The prognosis for dogs with LHS of the skin and subcutis is unknown, but in one small, retrospective series, neither local tumor recurrence nor metastasis was observed following resection with wide surgical margins and no adjunctive treatment in all five dogs with long-term follow-up.2 How- PROOF2,3 ever, distant metastasis is reported in 38% to 60% of dogs. Time to metastasis has not been reported. In contrast, the prognosis for dogs with LHS of the internal organs (Figure 3), such as the spleen, is poor, with a median survival time (MST) of 1 month and a 0% to 20% 1-year survival time.2,19,31 Poor prognostic factors in dogs with splenic histiocytic sarcomas include lymphoid:fibrohistiocytic ratio, mitotic index, and histologic grade.31,32 Dogs with a lymphoid:fibrohistiocytic ratio of greater than 40% have a 1-year survival probability of 87%; if the ratio is less than 40%, the 1-year survival probability is 55%.31 Dogs with grade III fibrohistiocytic nodules have a 1- year survival rate of 32%, which is significantly worse than the rates of 57% and 61% for dogs with grade I and II lesions, respectively.31 The MST for all

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Figure 2. Digital LHS in a dog.

Before surgical excision.The mass is ulcerated and necrotic. After excision.

origins that are characterized by multinucleated giant cells and commonly diagnosed in the extremities.2,33 This umbrella encompasses fibrosarcomas, leiomyosarcomas, rhabdomyosarcomas, liposarcomas, synovial cell sarcomas, and histiocytic sarcomas. However, this description is somewhat inaccurate in dogs; therefore, the term LHS is preferred by most veterinary pathologists.2,33

Disseminated Histiocytic Sarcoma DHS is the preferred term for the disease historically known as malignant histiocytosis.2 DHS is a systemic proliferation of tissue macrophages (histiocytes) or myeloid DAPCs and their precursors.2,6,15,17 Many reports, however, describe the cellular origin as Figure 3. A solitary omental LHS in a dog. unknown.2,3 In such cases, it is difficult to definitively diagnose DHS. DHS is an aggressive, multisystem disease character- dogs with LHS has not been reported in the literature. ized by multiple tumors in several organs, with the One dog with an LHS of the tendon sheath of the spleen, lung, and bone marrow involved primarily and biceps brachii muscle had a survival time of only 4 the liver and lymph nodes secondarily.2,3,7,17,18,28 Multiple months.3 solid, pale tumors in these organs and others are charac- PROOF3,7,17,18,28 The prognosis for dogs with synovial LHS is very teristic at postmortem examination. Retrievers, poor.30 In a series of 18 dogs with synovial LHS, the rottweilers, and Bernese mountain dogs are overrepre- overall MST was 5.3 months, the MST for dogs under- sented.2,10,18 No sex predisposition has been reported, going amputation (with or without chemotherapy) was and dogs of any age can be affected.2,18 In Bernese 6 months, and the metastatic rate was 91%.30 The use of mountain dogs, DHS is a genetic disease with a poly- postoperative chemotherapy may improve the prognosis genic mode of inheritance.3 for dogs with LHS. Dogs with DHS often have nonspecific clinical signs LHS in dogs may be analogous to malignant fibrous and physical examination findings that indicate multi- histiocytoma in humans. The term malignant fibrous his- system involvement. Cutaneous and subcutaneous tiocytoma has historically been used to describe a collec- lesions are common and must be differentiated from tion of unspecified soft tissue sarcomas of different cell those of LHS based on clinical staging, as described

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above. DHS and SH are differentiated based on immunohistochemistry and clinical staging. Clinical signs specific to organ system involvement include dysp- nea (pulmonary masses with neoplastic involvement of the hilar lymph nodes), lameness (extensive proliferative bone marrow lesions with associated destruction of the surrounding bone), and various neurologic signs (associated with brain and spinal cord lesions).7,34–37 Lym- phadenomegaly, hepatomegaly, and splenomegaly are also often present.2,3,7,17,28 Neurologic signs, such as para- plegia or paralysis, are reported in 38% of dogs.2,3,7,17,28 In addition to clinical staging, diagnosis of DHS is based on abdominal ultrasonography, three-view thoracic radi- ography, bone marrow aspiration and cytology, and clin- Figure 4. Lymph node aspirate (Diff-Quik) from a dog with DHS. The cells are characteristically round to oval with a icopathologic, histopathologic, immunohistochemical, moderate amount of moderately basophilic cytoplasm that 6 and immunophenotypic findings. frequently contains punctuate vacuoles.The large, round-to-oval Radiographs may show pulmonary consolidation, nuclei are eccentrically placed with a coarse chromatin pattern nodular opacities, pleural effusion, diffuse interstitial and prominent nucleoli. infiltrations, and hepatosplenomegaly.17 On ultrasonography, hypoechoic nodules are commonly seen in the spleen; some may distort the splenic margin.25,27 The liver reniform eccentric nuclei with prominent, sometimes is the second most commonly affected organ. Hepatic multiple, nucleoli.38,39 Cytoplasmic hemosiderin granules lesions can be hypoechoic, hyperechoic, or of mixed and vacuolation are common. Cytophagia has been echogenicity.25,27 Other common ultrasonographic find- reported and assists in the diagnosis of DHS.6 Cytology ings include hypoechoic nodules in the kidneys and from bone marrow aspirates shows infiltrates of neo- mesenteric or medial iliac lymphadenomegaly.25,27 Results plastic cells in association with erythroid and myeloid of a recent study suggest that the ultrasonographic hyperplasia.6 appearance of canine abdominal DHS is nonspecific and Histologic examination of grossly abnormal organs that nonhistiocytic disease (e.g., hemangioma, soft tissue reveals infiltration of the organ with sheets of a pleomor- sarcoma) can be ruled out only by cytologic or histo- phic population of atypical histiocytic cells that have pathologic evaluation in these patients.25,27 finely granular or vacuolated eosinophilic cytoplasm and Hematologic findings include cytopenia (e.g., autoim- pleomorphic, anisokaryotic nuclei with pale to granular mune hemolytic anemia, leukopenia, thrombocytopenia) chromatin and prominent nucleoli.2,38,39 Hemosiderin and associated with cellular phagocytosis by neoplastic erythrocytes are occasionally seen in tumor cells, espe- macrophages.6,32 Serum biochemical abnormalities are cially in splenic masses.32 Multinucleate giant cells may be variable, but hyperbilirubinemia has been reported fre- present (Figure 5), and mitotic figures are occasionally quently.32 Dogs with thrombocytopenia may also have a noted.4,6,17,38,39 Poor encapsulation of cellular infiltrates prolonged coagulation profile indicative of (imminent) with necrotic cores that obliterate surrounding normal PROOF17 disseminated intravascular coagulation. Ferritin has tissue architecture has been infrequently reported. shown promise as a biochemical marker in dogs with DAPCs of DHS should stain positive for CD1b, concurrent hyperferritinemia, especially when the owner CD1c, and, rarely, CD90 (Thy-1). LHS also positively elects chemotherapy.14 Serum ferritin levels may be expresses these immunohistochemical markers; therefore, reduced with successful chemotherapy.14 LHS and DHS can only be differentiated based on clini- Cytology from direct or image-guided aspiration of cal presentation, physical examination findings, and accessible masses yields moderate numbers of interme- results of clinical staging.2,25,27 It has been proposed that diate-to-large histiocytic cells, either as single cells or in the origin of malignant cells be confirmed by immuno- small aggregates.38,39 The cells may be round, fusiform, histochemistry. In one human study, the use of additional or irregularly shaped. They contain abundant granular, staining techniques showed that samples from many pale basophilic cytoplasm4,6,17 (Figure 4) and oval to patients originally diagnosed with DHS lacked histio-

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Figure 5. Lymph node aspirate (Wright’s-Giemsa) from the same dog as in Figure 4. The cell at the center of the field exhibits cytomegaly and extreme multinucleation.

cytic markers.40 These patients were subsequently found not to have DHS. We have used immunohistochemistry to diagnose DHS postmortem in a subset of dogs in which multicentric lymphosarcoma was originally diagnosed but did not respond to conventional therapy. An effective treatment for dogs with DHS has not been described. Surgical resection of cutaneous lesions or affected organs (e.g., spleen, liver lobe, lung lobe) is generally not recommended unless it will provide some palliative benefit. Chemotherapy is the recommended treatment, but there are few reports of successful remission, with most dogs having rapid clinical progression and deterioration because of the very aggressive behav- ior of DHS.24 Lomustine (60 to 90 mg/m2 q3–4wk) was used with some success as a single agent in an unpub- lished multiinstitutional study.24 In this study, the response rate in 56 dogs with gross measurable disease treated with lomustine alone was 46%. The median remission duration in the 26 dogs that responded to PROOFtherapy was 85 days, and the MST was 172 days in these dogs. If lomustine-based chemotherapeutic protocols for treatment of DHS are unsuccessful, protocols involving cyclophosphamide, vincristine, prednisone, mitoxantrone, dacarbazine, and/or etoposide have been described.19 A small group of dogs with DHS reportedly responded to an immunotherapy protocol involving treatment with a human cytotoxic T-cell line.3,40 The human equivalent of DHS is disseminated Langer- hans cell histiocytosis, which is an accumulation of tissue histiocytes in one or multiple organs or tissues that generally manifests with extensive multiorgan disease and fail-

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ure.2,41 Treatment is palliative and includes a wide spectrum of drugs (e.g., corticosteroids, chemotherapeutic agents [including vinca alkaloids and antimetabolites]) and immunomodulation (including immune-system up- regulation).41 A trial of the antimetabolite 2-chlorodeoxyadenosine in human patients with refractory, relapsed or widespread disease achieved relative success.41 The prognosis for dogs with DHS is poor. Based on the few available survival data, the MST is 106 days (range: 2 to 884 days) and the 1-year survival rate is 0% to 30%.24,32 Thrombocytopenia, hypoalbuminemia, and histologic evidence of giant cells are all poor prognostic factors.24,32 In some reports, dogs presented with a clinical sign that caused death, and DHS was diagnosed at necropsy; in others, the dog was euthanized at the time of diagnosis or soon after because of rapid clinical deterioration.14,32,34,35 Few reports of treatment, response, and survival rates in dogs with DHS are available.

CONCLUSION Diagnosis, staging, treatment, and prognostication of histiocytic diseases pose a challenge for the clinician. While the initial clinical presentation of the various histiocytic diseases is similar, the management and prognosis vary considerably. Accurate diagnosis and clinical staging are important to determine treatment options and prognosis. There is a distinct difference between dogs with LHS and DHS, and a histologic diagnosis of histiocytic sarcoma should not be interpreted as a poor prognosis until the appropriate staging tests have been conducted to differentiate LHS from DHS.

ACKNOWLEDGMENTS The authors thank Dr. Rebekah Gunn for her assistance with the cytologic images.

REFERENCES 1. Austin BR, Henderson RA. What is your diagnosis? JAVMA 2003;223(11): 1569-1570. 2. AffolterPROOF VK, Moore PF. Localized and disseminated histiocytic sarcoma of dendritic cell origin in dogs. Vet Pathol 2002;39:74-83. 3. Bass M, Gardelle O, Grest P, Bernasconi C. Localized histiocytic sarcoma in a dog: an uncommon diagnosis in forelimb lameness. Vet Comp Orthop Trau- matol 2004;17:48-52. 4. Fant P, Caldin M, Furlanello T, et al. Primary gastric histiocytic sarcoma in a dog—a case report. JAVMA 2004;51:358-362. 5. Sato T,Terui T, Kogawa K, et al. A case of true malignant histiocytosis: identification of histiocytic origin with use of immunohistochemical and immunocytogenic methods. Ann Hematol 2002;81:285-288. 6. Kraje AC, Patton CS, Edwards DF. Malignant histiocytosis in 3 cats. J Vet Intern Med 2001;15:252-256. 7. Chandra AMS, Ginn PE. Primary malignant histiocytosis of the brain in a dog. J Comp Path 1999;121:77-82.

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8. Anjiki T, Wada Y, Honma H, et al. Malignant histiocytosis in cattle. J Vet 36. Rosin A, Moore P,Dubielzig R. Malignant histiocytosis in Bernese mountain Med Sci 2000;62(12):1235-1240. dogs. JAVMA 1986;188(9):1041-1045. 9. Belal A, Kandil A, Allam A, et al. Malignant fibrous histiocytoma: a retro- 37. Wellman ML, Davenport DJ, Morton D, Jacobs RM. Malignant histiocytosis spective study of 109 cases. Am J Clin Oncol 2002;25(1):16-22. in four dogs. JAVMA 1985;187(9):919-921. 10. Morris JS, McInnes EF, Bostock DE, et al. Immunohistochemical and 38. Weiss D. Cytologic evaluation of benign and malignant hemophagocytic dis- histopathological features of 14 malignant fibrous histiocytomas from flat- orders in canine bone marrow. Vet Clin Pathol 2002;31(1):28-34. coated retrievers. Vet Pathol 2002;39:473-479. 39. Brown DE, Thrall MA, Getzy DM, et al. Cytology of canine malignant his- 11. Mellanby RJ, Holloway A, Chantrey J, et al. Immune-mediated haemolytic tiocytosis. Vet Clin Pathol 1994;23(4):118-123. anemia associated with a sarcoma in a flat-coated retriever. J Small Anim Pract 40. Vissoneau S, Cesano A, Tran T, et al. Successful treatment of canine malig- 2004;45:21-24. nant histiocytosis with the human major histocompatibility complex nonre- 12. Walton RM, Brown DE, Burkhard MJ, et al. Malignant histiocytosis in a stricted cytotoxic T-cell line TALL-104. Cancer Res 1997;3:1789-1797. domestic cat: cytomorphologic and immunohistochemical features. Vet Clin 41. Pardanani A, Phyliky R, Chin-Yang Li, Tefferi A. 2-Chlorodeoxyadenosine Pathol 1997;26(2):56-60. therapy for disseminated Langerhans cell histiocytosis. Mayo Clin Proc 13. Hayden DW, Waters DJ, Burke BA, Manivel JC. Disseminated malignant 2003;78:301-306. histiocytosis in a golden retriever: clinicopathologic, ultrastructural, and immunohistochemical findings. Vet Pathol 1993;30(3):256-264. 14. Newlands CE, Houston DM, Vasconcelos DY. Hyperferritinaemia associated ARTICLE #1 CETEST with malignant histiocytosis in a dog. JAVMA 1994;205(6):849-851. This article qualifies for 2 contact hours of continuing CE 15. Smoliga J, Schatzberg S, Peters J, et al. Myelopathy caused by a histiocytic sarcoma in a cat. J Small Anim Pract 2005;46:34-38. education credit from the Auburn University College 16. Affolter VK, Moore PF. Canine histiocytic proliferative disease. 15th Proc of Veterinary Medicine. Subscribers may take AAVD/ACVD 1999:79-86. individual CE tests or sign up for our annual 17. Ramsey IK, McKay JS, Rudorf H, Dobson JM. Malignant histiocytosis in CE program. Those who wish to apply this credit to three Bernese mountain dogs. Vet Rec 1996;138:440-444. 18. Affolter VK, Moore PF. Canine cutaneous and systemic histiocytosis: reactive fulfill state relicensure requirements should consult their histiocytosis of dermal dendritic cells. Am J Dermatopathol 2000;22:40-48. respective state authorities regarding the applicability 19. Spangler WL, Culbertson MR, Kass PH. Primary mesenchymal (nonan- of this program. CE subscribers can take CE tests online giomatous/nonlymphomatous) neoplasms in the canine spleen: anatomic and get real-time scores at CompendiumVet.com. classification, immunohistochemistry, and mitotic activity correlated with patient survival. Vet Pathol 1994;31:37-47. 20. Moore PF, Affolter VK. Canine and feline histiocytic diseases. In: Ettinger 1. Which of the following statements about histio- SJ, Feldman EC, eds. Textbook of Veterinary Internal Medicine. St Louis: Else- cytes is false? vier Saunders; 2005:779-782. a. Histiocytes are tissue macrophages whose functions 21. Paterson S, Boydell P,Pike R. Systemic histiocytosis in the Bernese mountain dog. J Small Anim Pract 1995;36(5):233-236. are related to their interactions with lymphocytes in 22. Moore PF. Systemic histiocytosis of Bernese mountain dogs. Vet Pathol inflamed tissues and consist primarily of phagocytosis 1984;21(6):554-563. and antigen presentation. 23. Mays MB, Bergeron JA. Cutaneous histiocytosis in dogs. JAVMA b. Macrophages are involved in the inflammatory 1986;188(4):377-381. process, have high lysosomal enzyme content, and are 24. Skorupski KA, Clifford CA, Paoloni MC, et al. CCNU for the treatment of dogs with histiocytic sarcoma. J Vet Intern Med 2007;21(1):121-126. capable of varying degrees of phagocytosis. 25. Ramirez S, Douglass J, Robertson I. Ultrasonographic features of canine c. DAPCs have typical dendritic morphology and low abdominal malignant histiocytosis. Vet Radiol Ultrasound 2002;43(2):167-170. levels of lysosomal enzymes, are mostly incapable of 26. Bettini G, Mandrioli L, Brunetti B, et al. Canine splenic pathology: a retro- phagocytosis, and play a key role in antigen presenta- spective study of 109 surgical samples with special emphasis on fibrohistio- tion to lymphocytes. cytic nodules. Eur J Vet Pathol 2001;7:101. 27. Cruz-Arámbulo R, Wrigley R, Powers B. Sonographic features of histiocytic d. Histiocytic phenotype cannot be differentiated using neoplasms in the canine abdomen. Vet Radiol Ultrasound 2004;45:554. immunohistochemistry and is not clinically important. 28. Moore PF, Rosin A. Malignant histiocytosis of Bernese mountain dogs. Vet Pathol 1986;23(1):1-10. 2. Which of the following results of immunohisto- 29. Banks T, Straw R, Thomson M, Powers B. Soft tissue sarcomas in dogs: a chemistry conducted on a formalin-fixed speci- study assessing surgical margin, tumour grade and clinical outcome. Aust Vet men suggests histiocytic disease? PractPROOF2004;34(4):142-147. 30. Craig LE, Julian ME, Ferracone JD. The diagnosis and prognosis of synovial a. CD3+, CD18-, CD45+, CD79- tumors in dogs: 35 cases. Vet Pathol 2002;39:66-73. b. CD3-, CD18+, CD45-, CD79+ 31. Spangler WL, Kass PH. Pathologic and prognostic characteristics of c. CD3+, CD18+, CD45+, CD79+ splenomegaly in dogs due to fibrohistiocytic nodules: 98 cases. Vet Pathol d. CD3-, CD18-, CD45-, CD79- 1998;35:488. 32. Spangler WL, Kass PH. Splenic myeloid metaplasia, histiocytosis, and hyper- splenism in the dog (65 cases). Vet Pathol 1999;36:583. 3. Lesions typical of reactive histiocytosis are 33. Kerlin RL, Hendrick MJ. Malignant fibrous histiocytoma and malignant his- a. nonpruritic and painless, grow slowly, and rarely tiocytosis in the dog—convergent or divergent phenotypic differentiation? Vet regress. Pathol 1996;33:713. b. nonpruritic and painless, grow rapidly, and rapidly 34. Thio T, Hilbe M, Grest P, Pospischil A. Malignant histiocytosis of the brain regress. in three dogs. J Comp Pathol 2006;134(2-3):241-244. 35. Carioto L. Malignant histiocytosis in a Bernese mountain dog presenting as a c. pruritic and painful, grow slowly, and rarely regress. mandibular mass. Can Vet J 1997;38(2):105-107. d. pruritic and painful, grow rapidly, and rapidly regress.

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4. Long-term medical immunosuppression is preferred over local surgical excision for the management of reactive histiocytosis because a. middle-aged dogs are never good surgical candidates. b. multiple lymph node involvement is common in cases of SH. c. distant de novo masses are common with CH. d. b and c

5. A solitary, small, firm, dome-shaped, dermoepithelial mass on the neck of an otherwise healthy 1-year-old shar-pei should raise suspicion for a. SH. c. LHS. b. cutaneous histiocytoma. d. DHS.

6. If the mass described in question 5 were snap frozen and submitted for immunohistochemical staining, results of CD1+, CD11c+, CD18+, MHC II+, ICAM-1+, CD4-, and CD90 (Thy 1)- would be consistent with a a. reactive process such as SH. b. benign neoplastic process such as cutaneous histiocytoma. c. malignant neoplastic process such as LHS. d. malignant neoplastic process such as DHS.

7. Metastasis (local lymph node involvement) has been reported in ___ of cases with LHS. a. less than 1% c. 60% b. 5% to 15% d. more than 90%

8. ______is a poor prognostic factor associated with splenic LHS. a. Lymphoid:fibrohistiocytic cell ratio <40% b. High mitotic index c. High histologic grade d. all of the above

9. DHS and LHS can be distinguished based on a. signalment. b. location of lesions and/or metastasis. c. immunohistochemistry. d.PROOF clinical presentation, physical examination, and clinical staging.

10. Which of the following statements about DHS is true? a. The prognosis for dogs with DHS is poor,with an MST of 106 days and a 0% to 30% 1-year survival rate. b. Thrombocytopenia, hypoproteinemia, and histologic evidence of giant cells are all poor prognostic factors. c. Lomustine has been used with excellent results, achieving median remission rates of more than 85% and longer than 900 days. d. a and b

April 2008 COMPENDIUM