Peripheral Neuropathy Associated with Malignant Neoplasms in Dogs

Transcription

1 Vet. Pathol (1987) Peripheral Neuropathy Associated with Malignant Neoplasms in Dogs K. G. BRAUND, J. A. MCGUIRE, K. A. AMLING, AND R. A. HENDERSON The Neuromuscular Laboratory of the Scott-Ritchey Research Program, Department of Small Animal Surgery and Medicine, College of Veterinary Medicine, and Department of Research Data Analysis, Auburn University, Auburn, AL Abstract. Common peroneal and ulnar nerves of 2 1 dogs between 2 and 15 years of age with malignant tumors were examined for possible paraneoplastic effects. Percent of abnormalities in single-teased fiber studies exceeded the confidence limits (P = 0.99) established for age-matched control dogs in 16 of 21 dogs (76%) with malignancies, none of which manifested clinical signs of polyneuropathy. The incidence of abnormalities was higher in commmon peroneal nerve (73%) than in ulnar nerve (57%). Lesions were characterized by a mixture of demyelinatiodremyelination and axonal degeneration; however, the former lesions predominated in most affected dogs. The severity of the neuropathy vaned with different tumors, with the most severe lesions observed in a bronchogenic carcinoma (59%), a mammary adenocarcinoma (59%), a malignant melanoma (48%), and an insulinoma (47%). Neoplasms may involve the peripheral neuromuscular system directly by local or metastatic infiltration and compression. However, non-metastatic effects of malignant cancer, also referred to as remote or paraneoplastic effects, have become increasingly common in people as therapy becomes more effective and the patients live longer.i6j7by definition, no tumor cells are seen in the nerves or spinal ganglia.3o Polyneuropathies occurring in dogs with malignant tumors, but unassociated with metastases, have been report- ed,15,24 however the incidence is unknown. This study was undertaken to examine selected peripheral nerves of dogs with various malignant system tumors for possible paraneoplastic lesions. Materials and Methods Twenty-one dogs, 14 males and seven females of various breeds between 2 and 15 years of age (mean = 9.3 years), were studied. All dogs were referred to Auburn University Small Animal Clinic with suspected malignant tumors. All dogs were given detailed clinical and neurological examination~.~~ Dogs were in good physical health and without clinical or hematological evidence of diabetes mellitus, renal disease, hyperadrenocorticism, or hypothyroidism. Tumors were clinically evaluated and graded. Preliminary tumor diagnosis in the majority of dogs was made from biopsy samples. Final confirmation of tumor identification was made following complete necropsy studies. Immediately following death by barbiturate overdose (Beuthanasia-D Special, Burns-Biotec Labs, Oakland, CA), 2 cm segments of the common peroneal nerve and ulnar nerve were sampled at the level of the stifle and elbow joints, respectively. The nerves were stretched on wooden tongue depressors with pins and fixed in a mixture of 4% forma- 16 lidl% glutaraldehyde. After fixation for 12 hours, the nerve samples were divided into halves and also longitudinally and were washed in Millonig s phosphate buffer (ph 7.3) at 4 C overnight. One half was post-fixed in 1% osmium tetroxide for 6 to 8 hours. These samples were then placed in 66% glycerin for 24 hours and stored in 100% glycerin for single teased-fiber preparation as previously de~cribed.~ The second half was post-fixed in 1% osmium tetroxide for 1 hour, washed in phosphate buffer, transferred through graded ethanol solutions and processed for embedding in Spurr s plastic medium. Semi-thin sections (1-2 pm) were cut transversely and stained with paraphenylenediamine. Brain, spinal cord, tumor, and samples of representative extraneural tissues were removed and immersion fixed in buffered 10% formalin. Fixed tissues were embedded in paraffin and sections cut at 4 pm and stained with hematoxylin and eosin. Morphologic studies Approximately 120 single fibers per nerve sample were teased randomly from all fascicles without preselection for size or abnormality. They were histologically classified according to the system of Dyck O (A) Teased fiber of normal appearance, (B) normal fiber with excessive myelin irregularity, wrinkling, or folding, (C) teased fiber with single or multiple regions of nodal lengthening or internodal myelin absence (as evaluated with high-dry light objective), (D) single or multiple C and F abnormalities combined, (E) teased fiber with linear rows of myelin ovoids and balls, (F) teased fiber without site(s) of segmental demyelination but with excessive variability of myelin thickness between internodes (thickness of myelin of the internodes with the thinnest myelin is less than 50% of that of the internode with the thickest myelin), or (G) teased fiber without site(s) of segmental demyelination but with excessive variability of myelin thickness within internodes to form globules or sausages. Fibers

2 Paraneoplastic Neuropathy in Dogs 17 Table 1. Incidence of abnormalities in teased nerve fibers from common peroneal nerves of dogs with malignant tumors. Dog No. Histologic Classification Percent of Abnor- Age (yr) C D E F G malities ne ne ne = not examined ne ne ne ne ne ne Tumor Type Bronchogenic carcinoma Bronchogenic carcinoma Insulinoma Insulinoma 48 8 Malignant melanoma Malignant melanoma Osteosarcoma Thyroid adenocarcinoma Mammary adenocarcinoma Mammary adenocarcinoma Mast cell tumor 5 5 Mast cell tumor Squamous cell carcinoma Metastatic sarcoma Metastatic sarcoma ne ne 7 Metastatic sarcoma 15 7 Myxosarcoma Nasal adenocarcinoma Lymphosarcoma 4 9 Lymphosarcoma ne ne 8 Lymphosarcoma with segmental demyelination and remyelination are represented by the graded lesions C, D, F, and G. Fibers undergoing axonal degeneration are represented by E. Statistical studies Linear and quadratic responses of percent abnormalities in common peroneal nerve and ulnar nerve to age changes were computed for tumor and control groups (the latter consisting of 21 age-matched control dogs for which age-related changes in these peripheral nerves have been rep~rted).~j In order to establish reference, base-line data and to compare the incidence of abnormalities in common peroneal and ulnar nerves of control dogs and of dogs with tumors, confidence limits (P = 0.99) were determined from the control nerves as a function of age. Values for percent abnormalities of dogs in the tumor group that were outside these confidence limits were considered different (P = 0.01) from the values of the control group. Results While some dogs manifested signs of exercise intolerance and vague weakness, none showed evidence of a diffise polyneuropathy or a localized neuropathy (i.e., flaccid paresis, hyporeflexia, hypotonia, and neurogenic muscle atrophy).2 Tumor types and associated incidence of abnormalities in teased nerve fibers from common peroneal and ulnar nerves are outlined in Tables 1 and 2. Major abnormal findings (Fig. 1) included paranodall segmental demyelination (graded lesion C) (Fig. 1 b), remyelination as suggested by the presence of short intercalated internodes (graded lesion F) (Fig. lc), axonal degeneration as manifested by linear rows of myelin ovoids and balls (graded lesion E) (Fig. Id), and myelin globules (graded lesion G) (Fig. le). This last change was seen infrequently. In a few cases, demyelination and remyelination (graded lesion D) were present on the same fiber. Fibers comprising a series of uniformly short internodes, suggestive of fiber regeneration, were seen occasionally. The common peroneal nerve exhibited a quadratic response (P = 0.06) to age in percent abnormalities in the control dogs but a linear response (P = ) in the tumor group. The percent abnormalities remained low at early ages and then increased rapidly at older ages in the controls while there was a linear increase due to age in the tumor group from early to older ages (Fig. 2). The ulnar nerve exhibited a quadratic response (P = ) to age by percent abnormalities in the control group but no response to age in the tumor group (Fig. 3). The graphs indicate that the percentage of abnormalities were clearly outside the 99% confidence limits established for the controls in 73% (1 4 of 19 dogs) of common peroneal nerves and 57% (12 of 21 dogs) of ulnar nerves from the tumor group. Overall, 16 of 21 dogs (76%) had a significantly greater number of lesions

3 18 Braund et al. d A Fig. 1. Teased nerve fiber panel, (a) normal myelinated fiber with node of Ranvier (arrow); (b) paranodal demyelination; (c) intercalated internode. Arrows = nodes of Ranvier; (d) myelin ovoids and balls; (e) myelin globules. Osmium tetroxide. in one or both nerves than age-matched controls. Ten of 16 dogs (62%) had more lesions in both nerves cornpared to controls. In dogs with increased numbers of abnormalities, a combination of demyelination, re- myelination, and axonal necrosis was present in almost every nerve. The incidence of demyelination and remyelination, however, was considerably higher than that of axonal degeneration. In common peroneal Table 2. Incidence of abnormalities in teased nerve fibers from ulnar nerves of dogs with malignant tumors. Dog No. Histologic Classification Percent of Abnor- Age (yr) C D E F G malities Tumor Type Bronchogenic carcinoma Bronchogenic carcinoma Insulinoma Insulinoma Malignant melanoma Malignant melanoma Osteosarcoma Thyroid adenocarcinoma Mammary adenocarcinoma Mammary adenocarcinoma Mast cell tumor Mast cell tumor Squamous cell carcinoma Metastatic sarcoma Metastatic sarcoma Metastatic sarcoma Myxosarcoma Nasal adenocarcinoma Lymphosarcoma Lymphosarcoma Lymphosarcoma

4 Paraneoplastic Neuropathy in Dogs 19 CPN COMMON PERONEAL NERVE A CONTROLS DOGS WITH TUMORS UN 50-1 ULNAR NERVE A CONTROLS 0 DOGS WITH TUMORS I,' 20 - ' e / / 5- -*. O i AGE Fig, 2. Incidepce (percent) of abnormalities in teased nerve fibers of common peroneal nerves from dogs with malignant tumors and from Control dogs. Intempted lines represent 99% confidence limits of age-related abnormalities in controls I5 AGE Fig. 3, Incidence (percent) ofabnorm&ies in teased nerve fibers of ulnar from dogs with malignant tumors and from control dogs. lines represent 99% confidence limits of age-related abnormalities in controls. nerves of the tumor group, intercalated internodes (remyelination) predominated in nine of 14 cases (64%) that were outside the control confidence limits, demyelination in two of 14 (14%) cases, and axonal necrosis in three of 14 cases (2 lo/o). Similarly, in ulnar nerves of the tumor group, intercalated internodes predominated in nine of 12 cases (75%) outside the confidence limits, demyelination in two of 12 cases (1 7%), and axonal necrosis in one of 12 cases (8%). Cross-sectional nerve preparations were examined in order to depict marked depletion of myelinated fibers. Such a change was not observed. In some cases, lesions characterized by focal or multifocal fibers with inappropriately thin myelin sheaths and/or fiber degeneration were seen. No lesions were found in the central nervous system that could account for the peripheral polyneuropathies. The highest percentage of combined abnormalities in teased fibers of either nerve from dogs with malignancies were found in bronchogenic carcinoma (59%), mammary adenocarcinoma (5 9%), malignant melanoma (48%), insulinoma (47%), osteosarcoma (39%), thyroid adenocarcinoma (35.5%), and mast cell tumor (32%). Discussion The results of this study indicate that morphological changes in nerves of dogs with malignant tumors (mean age = 9.3 years) are similar to those reported in normal dogs with increasing age;4 however, abnormalities exceeded the 99% confidence limits established for controls in 16 of 2 1 dogs (76%) with tumors even with the age-effect taken into account. The incidence of the combined abnormalities was higher in common peroneal nerve (73%) than in ulnar nerve (57%). The reason for this discrepancy between the two nerves is not known since no differences were observed in agematched control^.^,^ In the absence of other known metabolic causes of polyneuropathy, i.e., diabetes mellitus6 or hyperadrenoc~rticism,~ or of exposure to environmental toxins such as organophosphate compounds,2 it is suggested that the combined lesions that exceed the established age-related confidence limits represent a remote or paraneoplastic effect of the un-

5 20 Braund et al. derlying malignant tumor. Similar peripheral nerve lesions have been previously reported in dogs with malignant tumor^.'^,^^ It is recognized that comparable lesions (especially axonal degeneration) may occur with certain antineoplastic drugs such as the vinca alkaloid, vincristine.'j4 None of the dogs in this report were treated with antineoplastic drugs and none received radiation therapy which rarely induces a polyneuropathy in people.25 The nature of the lesions, typically characterized by a mixture of demyelinationhemyelnation and axonal degeneration, are similar to those described with sensorimotor polyneuropathy in people with malignancie~.*~j~ The determination of a sensory component in dogs awaits electrophysiological evaluation. Although we have seen no clinical signs in older control dogs with 10 to 15% of combined abnormalities in peripheral nerve^,^ the absence of signs of a neuropathic syndrome in any of our tumor dogs is enigmatic, particularly in the two dogs with 59% of combined abnormalities. However, up to 65% of combined abnormalities have been reported in dogs with subclinical neuropathy associated with diabetes mellitus.6 A possible explanation is that remyelination and not axonal degeneration was the most salient pathological feature, as it was in the diabetic study. The clinical incidence of polyneuropathy in human patients with malignancies has been estimated to be less than 2%.17 This figure increases when specific tumors are considered, i.e., 2.8% for carcinoma of the stomach and 5.3% for carcinoma of the l~ng.~,~ Many more human patients with malignancies have an underlying subclinical polyneuropathy. In different electrophysiological studies, the incidence of subclinical polyneuropathy has ranged from 17% to almost 50%.2L,26,27 The figure of 50% represents those patients with lung carcinoma. In the present study, the incidence of subclinical polyneuropathy, as determined by percent of abnormalities outside the confidence limits established for the controls, appears to be higher than the incidence in people. As previously noted, 14 of 21 dogs (76%) had more lesions than controls. The incidence was 73% for common peroneal nerve and 57% for ulnar nerve. Ten of I6 dogs (62%) had more lesions in both of these nerves as compared to controls. Future electrophysiological studies are needed to further define this incidence. Two of the tumor cases (one with the highest number of abnormalities) were bronchogenic carcinomas. Such tumors have long been recognized for their intimate association with paraneoplastic neuropathies in peo- ple.8,9j7,28 It was evident that different types of malignancies (i.e., mammary adenocarcinoma, malignant melanoma, insulinoma, osteosarcoma, etc.) resulted in a differing incidence of neuropathy, a finding also wellrecognized in human cancer patients.17 In addition, the severity of the neuropathy sometimes varied markedly with tumors of the same type, probably reflecting differences in the stage and duration of the illness. Of all the malignant tumors in our study, lymphosarcoma appeared to have the least neuropathic effect. The pathogenesis of paraneoplastic neuropathy remains elusive. At present, there is no evidence to support a link between cachexia and neuropathy, or an underlying vitamin deficiency.17 No definitive neurotoxin has been isolated and there is no evidence that cancerous cells elaborate hormonal substances that produce a neuropathy.i6 The possible role of a virus remains undetermined. Some of the best evidence suggests an underlying immunological mechanism. Serologic studies have demonstrated the presence of circulating antine~ral,~~ antimyelin basic protein,12 and antimyelin-associated glycoproteinz0 antibodies in a few human patients with malignancies. Nonetheless, it is uncertain whether these observations relate to causal mechanisms or whether they are an epiphenomenon. More recently, immunoreactive myelin-basic protein has been demonstrated in tumor cells of two human patients with carcinomatous neuropathy using immunocytochemical studies.'* In another study, acid extracts from human cancer tissue induced a cell-mediated immune response to myelin-basic protein and allergic encephalomyelitis in guinea pigs. l3 Results of such studies suggest that nervous tissue components may be present in certain tumors. Cross-reactivity between basic proteins of human myelin and of human cancer tissue7 may be the result of a common embryologic derivation. In this regard, many of the malignant tumors present in our dogs, namely melanoma, mast cell tumor, thyroid adenocarcinoma, and bronchogenic carcinoma, may represent tumors of the amine precursor uptake and decarboxylation system (APUD), many cells of which are derived from neural cre~t~*,~~ which also gives rise to cells of the peripheral nervous system. The results of this study suggest that malignant tumors can induce subclinical paraneoplastic lesions in peripheral nerves of dogs. Further studies of this disorder should help to clarify the intensity and incidence of the polyneuropathy with specific tumor types; however, the true incidence may always be uncertain since it will vary with the stage and duration of the illness, the diligence with which it is sought, the techniques of investigation employed, and the criteria for diagnosis.i9 Finally, the development of polyneuropathy in older dogs should alert the clinician and pathologist to search carefully for an underlying malignancy.

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