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Biomedical subjects

J A Ramos-Vara

Publications and source records attributed to J A Ramos-Vara.

At least 19 recordsLinked to original sources

Immunohistochemical detection of protein gene product 9.5 (PGP 9.5) in canine epitheliotropic T-cell lymphoma (mycosis fungoides).

Protein gene product 9.5 (PGP 9.5), a ubiquitin COOH-terminal hydrolase initially considered specific for neural and neuroendocrine tissues, is expressed in a variety of epithelial and mesenchymal tumors. During immunohistochemical evaluation of a cutaneous epitheliotropic T-cell lymphoma (mycosis fungoides [MF]) in a dog, strong reactivity for PGP 9.5 was observed. This unexpected result prompted us to examine PGP 9.5 immunoreactivity in 13 additional cases of canine mycosis fungoides. All tumors were confirmed as T-cell epitheliotropic lymphoma by histopathology and immunohistochemistry for CD3. Eight of 14 cases were positive for PGP 9.5, with reactivity mainly in the cytoplasm and less commonly in the nucleus. One case had strong reactivity in the cell membrane, sometimes with concurrent paranuclear staining. Immunoreactivity did not correlate with location (epidermal, dermal, and adnexal) of tumor cells. Disease outcome did not vary between PGP 9.5-positive and negative tumors. Although PGP 9.5 immunoreactivity in MF did not predict tumor behavior in these dogs, it has had prognostic value in certain human carcinomas. This unexpected staining of lymphocytes in mycosis fungoides with an antibody to PGP 9.5 demonstrates its presence in nonneuroendocrine tumors and precludes its use as the sole diagnostic marker in discrete cell tumors in the skin.

Animals↗

Listeric meningoencephalomyelitis in a cougar (Felis concolor): characterization by histopathologic, immunohistochemical, and molecular methods.

Listeria monocytogenes has been recognized as an important food-borne pathogen in animals. Records of the disease caused by this bacterium in large felids are, however, rare. The nervous form of listeriosis was diagnosed in a 12-year-old male cougar (Felis concolor) with a several-day history of neurologic disease characterized by excess salivation, head pressing, and circling that progressed to recumbency and death. Microscopically, the main alteration in the brain and spinal cord was a variably severe meningoencephalomyelitis composed mainly of mononuclear cell aggregates with fewer neutrophils. L. monocytogenes was isolated from the brain by microbiological culture, and L. monocytogenes antigen was detected in formalin-fixed, paraffin-embedded sections of brain and spinal cord by immunohistochemical analysis. On the basis of the nucleotide sequence of the 16S rRNA gene, the isolated strain was determined to be serotype 1/2a. Food-borne transmission of the bacterium was suspected, but food was not available for testing.

Animal Diseases↗

Disseminated encephalitozoonosis in captive, juvenile, cotton-top (Saguinus oedipus) and neonatal emperor (Saguinus imperator) tamarins in North America.

Disseminated encephalitozoonosis was diagnosed in 2 sibling, juvenile, cotton-top tamarins (Saguinus oedipus) and 3 sibling, neonatal, emperor tamarins (S. imperator) by use of histologic examination, histochemical analysis, electron microscopy, and polymerase chain reaction (PCR) analysis with nucleotide sequencing. All tamarins were captive born at zoos in North America and died with no premonitory signs of disease. The main pathologic findings were myocarditis (4/5), hepatitis (3/5), interstitial pneumonia (3/5), skeletal myositis (3/5), meningoencephalitis (2/5), adrenalitis (2/5), tubulointerstitial nephritis (1/5), myelitis (1/5), sympathetic ganglioneuritis (1/5), and retinitis (1/5). Central nervous system lesions were the most prominent findings in cotton-top tamarins. The inflammation was predominantly lymphocytic and suppurative in cotton-top tamarins, whereas emperor tamarins had granulomatous or lymphoplasmacytic lesions. Intralesional periodic acid-Schiff-, gram-, or acid-fast (or all 3)-positive, oval-to-elliptical shaped organisms were found in 1 cotton-top and the 3 emperor tamarins. By electron microscopy, these organisms were consistent with microsporidia of the genus Encephalitozoon. E. cuniculi genotype III was detected by PCR analysis and sequencing in paraffin-embedded brain, lung, and bone marrow specimens from the cotton-top tamarins. Although PCR results were negative for one of the emperor tamarins, their dam was seropositive for E. cuniculi by ELISA and Western blot immunodetection. These findings and recent reports of encephalitozoonosis in tamarins in Europe suggest that E. cuniculi infection may be an emerging disease in callitrichids, causing high neonatal and juvenile mortality in some colonies. The death of 2 less than 1-day-old emperor tamarins from a seropositive dam supports the likelihood of vertical transmission in some of the cases reported here.

Adrenal Glands↗

Usefulness of thyroid transcription factor-1 immunohistochemical staining in the differential diagnosis of primary pulmonary tumors of dogs.

In a previous study, we characterized the reactivity of monoclonal antibody (MoAb) 8G7G3/1 to thyroid transcription factor-1 (TTF-1) in canine thyroid tumors. In this study, we have examined the reactivity of this antibody in 120 canine pulmonary tumors, including 78 primary epithelial tumors. Tissues had been fixed in formalin and routinely processed for histopathology. Nuclear staining for TTF-1 was detected in 64.2% of primary pulmonary epithelial tumors. The most common TTF-1-reactive tumor types were bronchioloalveolar carcinomas and bronchogenic carcinomas. Staining was diffuse, heterogeneous, or patchy. Nonpulmonary, metastatic epithelial tumors, except two of two thyroid carcinomas, did not react with antibody 8G7G3/1. Mesotheliomas and other mesenchymal tumors were also negative for this marker. A reduction or loss of reactivity was apparent in pulmonary epithelial tumors archived in paraffin blocks for 7-8 years. There was slight reduction in the number of positive cells or the intensity of the reaction in control tissues fixed longer than 1 week. On the basis of our limited studies and the human literature, it appears that MoAb 8G7G3/1 to TTF-1 is a highly specific (with the exception of thyroid tumors) and moderately sensitive marker for canine pulmonary epithelial tumors.

Animals↗

Technical aspects of immunohistochemistry.

Immunohistochemistry is an integral technique in many veterinary laboratories for diagnostic and research purposes. In the last decade, the ability to detect antigens (Ags) in tissue sections has improved dramatically, mainly by countering the deleterious effects of formaldehyde with antigen retrieval (AR) and increasing sensitivity of the detection systems. In this review, I address these topics and provide an overview of technical aspects of immunohistochemistry, including those related to antibodies (Abs) and Ags, fixation, AR, detection methods, background, and troubleshooting. Microarray technology and the use of rabbit monoclonal Abs in immunohistochemistry are also discussed.

Alcohols↗

Pulmonary neoplasia in two llamas (Lama glama).

Two llamas with pulmonary tumors were examined. Llama No. 1 had multiple nodules throughout the lung that consisted histologically of solid clusters of polygonal to spindle cells with rare glandular differentiation. Intravascular emboli were common. Similar neoplastic masses were present in the kidney, heart, and liver. Immunohistochemically, neoplastic cells were positive for broad-spectrum cytokeratins (CKs), high-molecular weight CKs, CKs 5/6, and vimentin. The diagnosis was pulmonary carcinoma. Llama No. 2 had pulmonary nodules without extrapulmonary involvement. Microscopically, neoplastic cells formed acini lined by simple epithelium and solid cords of squamous cells that sometimes surrounded acini. Neoplastic cells were strongly positive for broad-spectrum CKs and weakly positive for thyroid transcription factor-1. The diagnosis was adenosquamous carcinoma. Pulmonary tumors account for 23% of neoplasms in South American camelids in our laboratory, making this the second most common type of neoplasm after lymphosarcoma.

Animals↗

Immunohistochemical detection of uroplakin III, cytokeratin 7, and cytokeratin 20 in canine urothelial tumors.

Immunohistochemistry for uroplakin III (UP III), cytokeratin 7 (CK 7), and cytokeratin 20 (CK 20) using commercially available antibodies was done in normal canine urinary bladder and 72 canine urinary bladder tumors that had been fixed in formalin and embedded in paraffin. Prolonged fixation (3-28 days) did not significantly alter the immunostaining for UP III. There was moderate reduction in the intensity for CK 7 and CK 20 after 1 week of fixation. UP III was detected in superficial (umbrella) cells and some intermediate cells of the normal urinary bladder, 7 of 7 transitional cell papillomas (TCPs), 50 of 55 transitional cell carcinomas (TCCs), and 4 of 5 metastatic TCCs. Staining was typically outlined in the plasma membrane, but diffuse or focal cytoplasmic staining was also observed. Intracytoplasmic lumina were usually positive for UP III. One squamous cell carcinoma of the bladder, 4 nonepithelial bladder tumors, and 285 nonurothelial tumors from different nonurinary locations were negative for UP III. CK 7 was detected in 7 of 7 TCPs, 53 of 54 TCCs, and 5 of 5 metastatic TCCs. The staining for CK 7 was diffuse cytoplasmic. CK 20 was detected in 1 of 7 TCPs, 37 of 54 TCCs, and 1 of 5 metastatic TCCs. The staining with CK 20 was cytoplasmic and weaker than with antibodies to UP III or CK 7. There was concurrent expression of UP III, CK 7, and CK 20 in 36 of 54 TCCs. UP III is a specific and sensitive marker for canine transitional epithelial (urothelial) neoplasms, detecting 91% of TCCs. Negative results may be observed with anaplastic tumors.

Animals↗

3-methylindole induces transient olfactory mucosal injury in ponies.

Response to 3-methylindole (3MI) varies among species. Mice recover from 3MI-induced bronchiolar epithelial injury but sustain persistent olfactory mucosal injury with scarring and epithelial metaplasia. In contrast, 3MI induces obliterative bronchiolitis in horses and ponies, but olfactory mucosal injury has not been reported. To evaluate the effect of 3MI on equine olfactory mucosa, ponies were dosed orally with 100 mg 3MI/kg (n = 9) or corn oil vehicle (n = 6). All ponies treated with 3MI developed obliterative bronchiolitis with mild olfactory injury. By 3 days after 3MI dosing, olfactory epithelium appeared disorganized with decreased and uneven surface height and scalloping of the basement membrane zone. Epithelial cells of Bowman's glands were hypertrophic. Proliferation of olfactory epithelium and Bowman's glands was supported by an increased mitotic index and positive immunohistochemical staining for proliferating cell nuclear antigen as compared with controls. The activity of 11beta-hydroxysteroid dehydrogenase, an olfactory mucosal cytosolic enzyme localized to sustentacular and Bowman's glandular epithelial cells, was concurrently decreased. By 9 days postdosing, olfactory mucosal lesions had lessened. Results indicate that 3MI transiently injures equine olfactory mucosa without the extensive necrosis, scarring, or metaplasia seen in murine olfactory mucosa or in equine bronchiolar epithelium.

Animals↗

Melan A and S100 protein immunohistochemistry in feline melanomas: 48 cases.

Immunohistochemistry, using a monoclonal antibody to Melan A and a polyclonal antibody to S100 protein, was applied to 48 formalin-fixed, paraffin-embedded specimens of feline melanoma. Forty-two cutaneous, three oral, one mucocutaneous, and two metastatic melanomas comprised the tumors. Thirty-two tumors (67%) were positive for Melan A and 42 (87.5%) were positive for S100. All but one of the tumors that were positive for Melan A were also positive for S100. S100 was detected in 11 of 16 tumors that were negative for Melan A. Seventy-five percent (9 of 12) of amelanotic melanomas were negative for Melan A. Normal adrenal cortex, the cerebellum, and the skin had cells that were positive for Melan A. Sebaceous adenoma was the only nonmelanocytic tumor examined that reacted with antibody to Melan A. Although less sensitive than S100 protein, Melan A is more specific for melanoma and is useful in differentiating feline cutaneous melanoma from the more common pigmented basal cell tumor.

Animals↗

Immunohistochemical detection of thyroid transcription factor-1, thyroglobulin, and calcitonin in canine normal, hyperplastic, and neoplastic thyroid gland.

Immunohistochemistry for thyroid transcription factor-1 (TTF-1), thyroglobulin, and calcitonin was done in normal, hyperplastic, and neoplastic canine thyroid glands that had been fixed in formalin and embedded in paraffin. Prolonged fixation did not significantly alter the immunostaining for TTF-1. Staining for TTF-1 was always nuclear and usually strong. One of two C-cell adenomas, five of five follicular cell adenomas, 5 of 11 C-cell carcinomas, 38 of 42 follicular cell carcinomas, two of five cases of C-cell hyperplasia, two of two cases of follicular epithelial hyperplasia, one of two metastatic C-cell carcinomas, and three of four metastatic follicular carcinomas were positive for TTF-1. One follicular carcinoma that was positive for TTF-1 was negative for thyroglobulin. Thirty-nine of 42 follicular cell carcinomas were positive for thyroglobulin, including two cases that were negative for TTF-1. All C-cell lesions were positive for calcitonin. Of a variety of normal canine tissues and 278 nonthyroid tumors, only normal lung (airway and alveolar epithelial cells) and four of five pulmonary carcinomas were positive for TTF-1. TTF-1 is a good marker of thyroid differentiation and can be used in conjunction with thyroglobulin and calcitonin to increase the detection and differentiation of thyroid tumors and their metastases.

Adenoma↗

Bronchitis and bronchiolitis in a cat with cilia-associated respiratory bacillus-like organisms.

A 10-year-old male domestic shorthair cat died during anesthesia. Grossly, the lungs had multiple nodules corresponding to inflamed airways (bronchitis and bronchiolitis). Microscopically, cuffs and nodular aggregates of lymphocytes and plasma cells surrounded airways. Peribronchiolar fibrosis was also common. Globule leukocytes infiltrated the respiratory epithelium of noninflamed or mildly inflamed bronchi. Argyrophilic and filamentous organisms, consistent with cilia-associated respiratory bacillus-like organisms (CLO), were intermixed with cilia of respiratory epithelium. Ultrastructurally, CLO were longer and thinner than cilia and had a distinct trilaminar membrane, central electron-lucent areas, and no specialized external structures. Silver stained lung sections from 18 additional feline airways revealed similar bacilli in 2/9 normal lungs, 1/7 lungs with bronchitis and bronchiolitis, and 1/2 lungs with pneumonia. The significance of CLO in the pulmonary lesions was not determined.

Animals↗

Mammary duct ectasia in dogs: 51 cases (1992-1999).

OBJECTIVE: To evaluate the clinical and pathologic characteristics of mammary duct ectasia in dogs. DESIGN: Retrospective study. ANIMALS: 51 dogs with mammary duct ectasia. PROCEDURE: Information regarding body condition, history, number and location of affected mammary glands, appearance of lesions, surgical treatment, nonsurgical treatment, and evidence of recurrence or development of mammary neoplasia was obtained from surveys sent to referring veterinarians. Results of information from examination of histologic sections and referring veterinarians were evaluated for all mammary duct ectasia biopsies performed between 1992 and 1999. RESULTS: Duct ectasia was the primary diagnosis in 51 of 1,825 (2.8%) mammary biopsy specimens and comprised 48% of nonneoplastic mammary diseases. Affected dogs were evenly distributed over a range of 1 to 13 years of age, with a mean age at the time of diagnosis of 6.1 +/- 3.1 years. All dogs were female (31 sexually intact, 20 spayed); 10 of 26 had whelped. Duct ectasia was described as nodular (26 dogs), cystic (13), and multiglandular (11) and located in caudal (31) more often than cranial (14) or middle glands (10). Ectasia recurred in 3 dogs. One dog had a history of previously excised mammary adenocarcinoma; another subsequently developed mammary carcinoma. CONCLUSIONS AND CLINICAL RELEVANCE: Duct ectasia affected mature, sexually intact and spayed female dogs over a wide age range. Certain breeds were affected more commonly than expected. Increased risk for mammary neoplasia was not evident. Duct ectasia should be considered as a cause for mammary enlargement, especially in young dogs or when its cystic nature is evident. Mastectomy is usually curative, and neoplasia should be ruled out in dogs with ectasia.

Age Distribution↗

Immunoreactivity of A103, an antibody to Melan A, in canine steroid-producing tissues and their tumors.

The monoclonal antibody A103 to the melanocytic differentiation antigen Melan A stains human steroid-producing cells and their tumors. A total of 200 formalin-fixed, paraffin-embedded canine normal tissues and hyperplastic and neoplastic lesions of the adrenal gland, testis, and ovary were immunohistochemically tested for Melan A with antibody A103. Leydig cell tumors (23/23, 100%), Sertoli cell tumors (14/15, 93%), and adrenocortical adenomas (12/13, 92%) were consistently positive. Adrenocortical carcinomas (23/35, 65%) and granulosa cell tumors (10/17, 59%) were less frequently positive. All pheochromocytomas, seminomas, and dysgerminomas were negative. The pattern of staining was cytoplasmic, but nuclear staining was also frequently seen in normal Leydig cells and their tumors. As in human tumors, immunohistochemistry for Melan A stains many canine steroid-producing tumors and can be used to distinguish these tumors from those of nonstereidogenic cells.

Adenoma↗

Soft tissue sarcomas in the African hedgehog (Atelerix albiventris): microscopic and immunohistologic study of three cases.

Three soft tissue tumors from 2 female hedgehogs were examined microscopically and immunohistochemically. Two tumors involved haired skin and the third one was vaginal. Microscopically, the cutaneous tumors had features of malignant peripheral nerve sheath tumor (MPNST), whereas the vaginal tumor was classified only as a spindle cell sarcoma. Immunohistochemically, all 3 tumors were strongly positive for vimentin and strongly to moderately positive for CD10 and neuron-specific enolase but did not stain with antibody to S100 protein, an antigen typically present in human MPNST The cutaneous tumor from hedgehog no. 1 was examined ultrastructurally and the neoplastic cells resembled fibroblasts. Hedgehog no. 1 was euthanized at the time of the biopsy. The outcome of the other hedgehog was unknown.

Animals↗

Immunohistochemical characterization of canine hyperplastic hepatic lesions and hepatocellular and biliary neoplasms with monoclonal antibody hepatocyte paraffin 1 and a monoclonal antibody to cytokeratin 7.

Immunostaining with monoclonal antibody (MoAb) hepatocyte paraffin 1 (Hep Par 1) and an MoAb to cytokeratin 7 (CK7) was performed on 105 formalin-fixed, paraffin-embedded canine hyperplastic and neoplastic hepatic lesions. Hep Par 1 was detected in 12/12 hyperplastic nodules, 17/17 hepatocellular adenomas, and 37/40 hepatocellular carcinomas. The staining was disseminated, granular, and cytoplasmic. This antibody did not react with normal or neoplastic biliary epithelium. Other hepatic tumors or tumors metastatic to the liver did not bind Hep Par 1 except one metastatic intestinal carcinoma. MoAb to CK 7 stained all hyperplastic biliary epithelium and benign cholangiocellular tumors (5/5) and 14/18 cholangiocellular carcinomas. One hepatocellular carcinoma had cells positive for both Hep Par 1 and CK 7. Liver was the only normal tissue tested that reacted with MoAb Hep Par 1. Only five nonhepatic tumors (one adrenocortical carcinoma, one interstitial cell tumor of the testis, one melanoma, and two salivary adenocarcinomas) of 277 tumors tested had focal/multifocal staining for Hep Par 1. Prolonged fixation did not alter the staining with Hep Par 1. We conclude that Hep Par 1 is a specific and sensitive marker for canine hepatocellular tumors and allows distinction between hepatocellular and biliary neoplasms.

Adenoma, Liver Cell↗

Identification of porcine circovirus in tissues of pigs with porcine dermatitis and nephropathy syndrome.

Thirty-three pigs affected by porcine dermatitis and nephropathy syndrome, 30 from Spain and three from the USA, were investigated in order to detect porcine circovirus (PCV) in their tissues. A standard in situ hybridisation technique using a specific DNA 317-bp probe based on a well-conserved sequence of PCV (which recognises both PCV-1 and PCV-2) was applied to formalin-fixed, paraffin-embedded tissues. Twenty-eight of the 30 Spanish pigs and all three American pigs had PCV in at least one tissue. Viral nucleic acid was detected mainly in lymphoid organs, and especially the lymph nodes. The viral genome was also found, in order of decreasing quantity, in Peyer's patches, tonsil, lung, spleen, kidney, liver, and skin. Viral nucleic acid was located mainly within the cytoplasm of monocyte/macrophage lineage cells, including follicular dendritic cells, macrophages, histiocytes and Kupffer cells. No viral nucleic acid was found in damaged glomeruli or arteriolar walls. In frozen samples available from three Spanish pigs, the virus was identified as type 2 by using the polymerase chain reaction and restriction fragment length polymorphism. Most of the pigs from which serum was available were seropositive against porcine respiratory and reproductive syndrome virus (PRRSV), and PRRSV antigen was detected in the lung of two of the Spanish pigs. These results suggested that PCV is present in tissues of almost all pigs affected by PDNS, and PCV has to be considered as a possible agent involved in the pathogenesis of the syndrome.

Animals↗

Optimization of immunohistochemical methods using two different antigen retrieval methods on formalin-fixed paraffin-embedded tissues: experience with 63 markers.

Formalin fixation produces cross-links between the proteins and the fixative that alter the ability of some antibodies to recognize antigens. We used formalin-fixed, paraffin-embedded tissues to compare two different antigen retrieval methods for 63 antibodies used in the diagnosis of infectious and neoplastic diseases of animal species. Eighty-four percent of the antibodies needed some type of antigen retrieval for optimal results. Of those antibodies, 67.7% were monoclonal and 32.3% were polyclonal. Steam heat was the method of choice for 31 antibodies. Ten antibodies reacted only with steam heat, but 9 antibodies did not react when steam heat was used. Optimal results were obtained with enzyme digestion for 22 antibodies. Only 10 antibodies yielded optimal results without antigen retrieval; 64% of these antibodies were polyclonal. All antibodies against cytokeratins were optimally retrieved with proteinese K. Antigen retrieval appears to be necessary for the majority of antibodies when used with formalin-fixed, paraffin-embedded tissues.

Animal Diseases↗