Dolors Fondevila

Histological and immunohistochemical study of clinically normal skin of Leishmania infantum-infected dogs

Skin lesions are the most usual manifestation of canine leishmaniosis. The aim of this study was to investigate the histological pattern and parasite load in clinically normal skin of Leishmania-infected dogs. Two groups of Leishmania-infected dogs were studied. Group A consisted of 15 symptomless animals which, although seronegative or only mildly seropositive, gave a positive polymerase chain reaction (PCR) for Leishmania in the skin. Group B consisted of 20 clinically affected dogs which were highly seropositive and PCR-positive. Biopsies of normal skin from all dogs were processed for routine histology and Leishmania immunohistochemistry. The study demonstrated microscopical lesions and the presence of parasites in the skin from dogs of group B, but not group A. The results cast doubt on the relevance of infected but symptomless dogs in the epidemiology of canine leishmaniosis. In contrast, however, the clinically normal skin of sick dogs harbours the parasite and probably plays a role in the transmission of leishmaniosis.

The tumor suppressor SirT2 regulates cell cycle progression and genome stability by modulating the mitotic deposition of H4K20 methylation

The establishment of the epigenetic mark H4K20me1 (monomethylation of H4K20) by PR-Set7 during G2/M directly impacts S-phase progression and genome stability. However, the mechanisms involved in the regulation of this event are not well understood. Here we show that SirT2 regulates H4K20me1 deposition through the deacetylation of H4K16Ac (acetylation of H4K16) and determines the levels of H4K20me2/3 throughout the cell cycle. SirT2 binds and deacetylates PR-Set7 at K90, modulating its chromatin localization. Consistently, SirT2 depletion significantly reduces PR-Set7 chromatin levels, alters the size and number of PR-Set7 foci, and decreases the overall mitotic deposition of H4K20me1. Upon stress, the interaction between SirT2 and PR-Set7 increases along with the H4K20me1 levels, suggesting a novel mitotic checkpoint mechanism. SirT2 loss in mice induces significant defects associated with defective H4K20me1-3 levels. Accordingly, SirT2-deficient animals exhibit genomic instability and chromosomal aberrations and are prone to tumorigenesis. Our studies suggest that the dynamic cross-talk between the environment and the genome during mitosis determines the fate of the subsequent cell cycle.

Detection of T lymphocytes in canine tissue embedded in paraffin wax by means of antibody to CD3 antigen

This report describes the immunocytochemical detection of CD3 antigen by means of a polyclonal antibody in sections of formol-fixed canine lymphoid tissue, embedded in paraffin wax. In all hybrid organs the lymphocytes in the T cell regions showed an intense immune reaction, located particularly in the cytoplasmic membrane and at the cytoplasmic border.

Immunohistochemical detection of CD31 antigen in normal and neoplastic canine endothelial cells

This report describes the immunohistochemical detection of von Willebrands factor (vWf) and CD31 antigen in paraffin wax-embedded, formalin-fixed tissue sections of canine normal organs and vascular neoplasms. CD31 antigen and vWf were detected in endothelial cells of all organs examined, except for the endothelia of the renal glomeruli, which were negative for vWf. All haemangiomas examined (15) were positive for both markers. Eleven of 15 haemangiosarcomas were positive for vWf and all 15 expressed the CD31 antigen. All other neoplasms investigated (fibrosarcomas, schwannomas, haemangiopericytomas) were negative for both markers. It is concluded that immunohistochemical detection of CD31 antigen is of value for studying vascular disorders of the dog in routinely processed tissue.

Epidermal immunocompetence in canine leishmaniasis

The antigen-presenting cell function of the epidermis was investigated immunocytochemically in 16 dogs with different types of skin lesions induced by Leishmania infection. The degree of epidermal immunocompetence was evaluated according to the presence of Langerhans cells (LC) and keratinocytes expressing class II major histocompatibility complex (MHC II) molecules on the one hand, and the relative numbers of macrophages, T cells, and parasites in the dermis on the other, as described for human cutaneous leishmaniasis. In alopecic dermatitis, appropriate numbers of LC and MHC II-positive keratinocytes were shown to be associated with a mild T cell infiltration without significant numbers of parasites. By contrast, when the epidermis lacked antigen-presenting cells, as occurred in nodular lesions, macrophages and parasites massively infiltrated the dermis. Ulcerative lesions showed intermediate patterns of inflammation. These results suggest that dogs with alopecic dermatitis develop an effective control of the infection, whereas those with a generalized nodular disease mount an impaired immune response against the parasite. Skin lesions in dogs infected by Leishmania might not only have a prognostic value, but also represent a suitable model to study the natural course of human cutaneous leishmaniasis.

Canine mucosal leishmaniasis

Four dogs infected with Leishmania had proliferative lesions on the mucosae of the penis, tongue, oral cavity, prepuce, or nose. These mucosal, nodular lesions produced by parasites of the genus Leishmania have not been described previously in the dog. Leishmaniasis should be considered in the differential diagnosis of tumor-like lesions of mucous membranes.

Canine cutaneous spindle cell tumours with features of peripheral nerve sheath tumours: a histopathological and immunohistochemical study.

In veterinary medicine, the term peripheral nerve sheath tumour is usually restricted to neoplasms that are closely associated with an identified nerve. Thirty-three cases of canine cutaneous tumours previously classified as spindle cell tumours with features resembling peripheral nerve sheath tumours were examined. Two histological patterns were identified: dense areas of spindle shaped cells resembling the Antoni A pattern and less cellular areas with more pleomorphic cells resembling the Antoni B pattern. Immunohistochemically, all tumours uniformly expressed vimentin and 15/33 (45.4%) had scattered and patchy expression of S-100. Laminin expression was found in 25/33 (75.7%) tumours and collagen IV labelling occurred in 14/33 (42.4%). Expression of protein gene product 9.5 was detected in 31/33 (93.9%) of tumours and neuron specific enolase labelling was present in 27/33 (81.8%). Glial fibrillary acidic protein was only expressed within the cytoplasm of some large multinucleated cells in one tumour. These findings suggest that any cutaneous tumour with one of the two histopathological patterns described above should be described as a cutaneous peripheral nerve sheath tumour and that expression of S-100, laminin and collagen IV may be used to define a schwannoma.

Cutaneous mucinosis in shar-pei dogs is due to hyaluronic acid deposition and is associated with high levels of hyaluronic acid in serum

Cutaneous mucinosis affects primarily shar-pei dogs. Hyaluronic acid (HA) is considered to be the main component of mucin and CD44 is the major cell surface receptor of HA, necessary for its uptake and catabolism. The aims of this study were to identify the composition of the mucin in cutaneous mucinosis of shar-pei dogs, investigate the correlation between the deposition of HA and the expression of CD44, and determine whether shar-pei dogs with cutaneous mucinosis presented with elevated levels of serum HA. In skin biopsies, the mucinous material was stained intensely with Alcian blue and bound strongly by the hyaluronan-binding protein. No correlation was found between the degree of HA deposition in the dermis and the expression of CD44 in the skin of shar-pei dogs affected or unaffected by cutaneous mucinosis. A clear positive correlation was found between the existence of clinical mucinosis and the serum HA concentration. In control dogs, serum HA ranged from 155.53 to 301.62 microg L(-1) in shar-pei dogs; without mucinosis it ranged from 106.72 to 1251.76 microg L(-1) and in shar-pei dogs with severe mucinosis it ranged between 843.51 to 2330.03 microg L(-1). Altogether, the results reported here suggest that mucinosis of shar-pei dogs is probably the consequence of a genetic defect in the metabolism of HA.

Detection of Leishmania infection in paraffin-embedded skin biopsies of dogs using polymerase chain reaction

Canine leishmaniosis (CL) is a severe systemic infectious disease of the dog caused by protozoan parasites of the genus Leishmania. The disease is endemic in Mediterranean countries, parts of east Africa, India, and Central and South America. Classic CL is as a chronic wasting disease with anemia, intermittent pyrexia, arthritis, and generalized lymphadenopathy. The skin is one of the main organs affected by CL,17 and cutaneous lesions are highly variable. The most common cutaneous lesion is generalized scaling leading to large areas of alopecia.5,6 Clinical signs of CL are extremely variable, and diagnosis is often difficult. The diagnosis is usually made by direct observation of the parasite in bone marrow smears or by testing for specific antibodies in serum. In CL-endemic areas, however, pathologists often receive skin biopsies of dogs suspected of suffering from CL. Histopathologic pictures are not specific for CL and range from diffuse granulomatous dermatitis to lichenoid or pustular dermatitis.5,6 Under these conditions, a definitive diagnosis can only be established by detection of amastigotes of Leishmania. Often, however, very few amastigotes are present, and they may be overlooked in routine tissue examination.6 Immunohistochemistry has substantially improved the histopathologic diagnosis of CL,3,4 although doubtful cases still emerge when the presence of Leishmania amastigotes cannot be conclusively demonstrated. Recently, oligonucleotide primers with various specificities for Leishmania detection have been described,8,9,14 and the polymerase chain reaction (PCR) has been used to detect Leishmania DNA in bone marrow samples or fresh biopsies and in paraffin-embedded human skin biopsies.7 The aim of this study was to assess the ability of PCR to detect Leishmania DNA in paraffin-embedded canine skin biopsy specimens and to compare the results with those found by immunohistochemistry. Samples from 35 dogs were studied. Ten had confirmed CL diagnosed by clinical profile and course of disease, presence of anti-Leishmania antibodies, and observation of parasites in bone marrow. In addition, skin biopsies were taken from all 10 dogs, fixed in 4% formaldehyde, and embedded in paraffin, and sections were stained with hematoxylin and eosin. Immunohistochemical detection of Leishmania was accomplished using a standard immunoperoxidase protocol.4 A polyclonal anti-Leishmania antibody obtained from a rabbit was used as the first antiserum. In skin biopsies from these 10 dogs, lesions consistent with CL (an accumulation of macrophages in the superficial and deep dermis with some plasma cells, lymphocytes, and polynuclear neutrophils scattered between the macrophages) were observed, and im-

Cutaneous neosporosis during treatment of pemphigus foliaceus in a dog.

A 4-year-old, intact male rottweiler was presented with a 10-day history of papulonodular dermatitis. At the time of presentation, the dog was receiving prednisone and azathioprine to treat pemphigus foliaceus. Cutaneous neosporosis was diagnosed by immunohistochemistry on skin biopsy specimens and a high serum antibody titer to Neospora caninum by Neospora agglutination test. Electron microscopy examination of skin specimens further supported the diagnosis. Clindamycin therapy, together with withdrawal of immunosuppressive medication, resulted in prolonged clinical remission. This report documents cutaneous neosporosis in an adult dog and suggests that immunosuppressive therapy might be a predisposing factor.