R Guglielmino’s research while affiliated with University of Turin and other places

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Publications (18)


Extranodal γδ-T-cell lymphoma in a dog with leishmaniasis
  • Article

October 2008

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160 Reads

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29 Citations

Veterinary Clinical Pathology

Valentina Foglia Manzillo

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Annalisa Pagano

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Roberta Guglielmino

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[...]

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An 8-year-old intact male mongrel dog with alopecia and weight loss was referred to the Veterinary Faculty of Naples. The dog had pale mucous membranes, enlarged prescapular lymph nodes, and splenomegaly. Laboratory abnormalities included anemia, thrombocytopenia, and hyperglobulinemia. Bone marrow aspirate smears contained numerous Leishmania amastigotes and an immunofluorescent antibody titer was strongly positive (1:1280) for leishmaniasis. The dog was treated with a combination of meglumine antimoniate and allopurinol for 60 days and showed clinical improvement. Two months after the end of treatment the dog was again referred because of relapse of leishmaniasis and the presence of a firm subcutaneous mass on the medial right thigh. Based on cytologic examination of fine needle aspirates of the mass, a diagnosis of large-cell lymphoma was made. Flow cytometry of tumor cells revealed gammadelta-T-cell lymphoma with a CD5+, CD3+, TCRgammadelta+, CD4-, CD8-, CD45RA+ immunophenotype. Using nested PCR, amastigotes were not detected in the neoplastic tissue. An association between leishmaniasis and hematopoietic tumors has been described rarely. gammadelta-T cells may be involved in the host response to this parasite, and prolonged antigenic stimulation and chronic immunosuppression (typical of leishmaniasis) play a crucial role in the etiopathogenesis of T-cell lymphoma.


Effects of road transportation on lymphocyte subsets in calves

March 2008

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40 Reads

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44 Citations

The Veterinary Journal

The effect of transportation on peripheral blood lymphocyte subsets in 24 calves was investigated by flow cytometry. Blood was collected before departure, on arrival, at 24h and 1 week after arrival. Highest leucocyte and neutrophil counts, associated with increased concentrations of cortisol and catecholamines, indicated that stress was maximal upon arrival. At this time, a decrease in the percentages of all T lymphocyte subsets was evident, while they did not decrease as absolute counts. The proportion of CD21(+) cells did not change, indicating that the relative reduction of T lymphocyte subsets was not related to an increase in B lymphocytes. These variations may be due to the increase of a natural killer (NK) cell subset. NK cell expansion, together with increasing lymphocyte count and increasing major histocompatibility complex class II expression, may indicate stress-induced stimulation of the immune system.


Biochemical, ultrastructural and molecular characterization of the triphenyltin acetate (TPTA)-induced apoptosis in primary cultures of mouse thymocytes

August 2006

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20 Reads

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7 Citations

Cell Biology and Toxicology

Triphenyltin acetate (TPTA), a triorganotin compound used in agriculture as a biocide, is immunotoxic in vivo and in vitro. The present study was undertaken to ascertain whether apoptosis might play a role in the TPTA toxicity in vitro. Mouse thymocyte primary cultures were exposed to 0, 4 and 8 micromol/L TPTA; methyl prednisolone (1 micromol/L) was used as a positive control. Cell aliquots were harvested after 0, 1, 2, 4, and 8 h and the presence of early or late apoptotic phenomena was checked by (a) morphological investigations; (b) spectrophotometric quantification of fragmented DNA and agarose gel electrophoresis; (c) cell flow cytofluorometry, using an annexin V-FITC kit; and (d) detection of in situ apoptosis by a colorimetric detection kit (Titer-Tacs). TPTA cytotoxicity was also evaluated using the trypan blue dye exclusion test. Morphological investigation indicated apoptosis and/or necrosis. After 8 h of incubation, cells exposed to 4 micromol/L TPTA showed an increase in DNA fragmentation (on electrophoresis), which was confirmed by spectrophotometry (p < 0.05). Flow cytofluorometry pointed out an early (p < 0.05) increase of annexin V-positive (apoptotic) cells in TPTA-exposed flasks, whereas at least partly contradictory, results were obtained with the Titer-Tacs kit. Overall, these results provide evidence that TPTA, at low concentrations (4 micromol/L) induces early and late apoptotic phenomena, whereas cells exposed to the highest concentrations (8 micromol/L) are likely to undergo necrosis rather than apoptosis.


Flow cytometric patterns in blood from dogs with non-neoplastic and neoplastic hematologic diseases using double labeling for CD18 and CD45

April 2006

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37 Reads

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28 Citations

Veterinary Clinical Pathology

In dogs, flow cytometry is used in the phenotyping of immunologic cells and in the diagnosis of hemic neoplasia. However, the paucity of specific antibodies for myeloid cells and B lymphocytes and of labeled antibodies for multicolor techniques limits the ability to detect all leukocyte subpopulations. This is especially true for neoplastic and precursor cells. CD18 and CD45 are expressed on all leukocytes and are involved in cell activation, and together could be useful in helping determine cell lineage. The purpose of this study was to double label canine blood for CD18 and CD45 and to use the differential expression of antigens to identify leukocyte populations in dogs with non-neoplastic and neoplastic hematologic diseases. A template was developed using blood samples from 10 clinically healthy dogs and a back-gating technique. Differential leukocyte counts obtained with the template were compared with those obtained by manual and automated methods on blood samples from 17 additional healthy dogs. Blood samples obtained from 9 dogs with non-neoplastic (reactive) hematologic diseases and 27 dogs with hemic neoplasia were double stained for CD18 and CD45 using mouse anticanine CD18 monoclonal antibody (mAb) plus phycoerythrin-conjugated rat anticanine CD45 mAb and fluorescein isothiocyanate-conjugated rabbit antimouse IgG. Hemic neoplasms were diagnosed by cell morphology, and immunophenotypic and cytochemical markers. With the double label, neutrophils, eosinophils, monocytes, and T- and B-lymphocytes were identified. In reactive disorders, a population of activated neutrophils with high CD45 and CD18 expression was detected. In hemic neoplasia, cell lineage was easily determined, even in acute leukemia. Double labeling for CD18/CD45 may be useful as a screening method to evaluate hematologic diseases and help determine cell lineage, and to aid in the selection of a panel of antibodies that would be useful for further analysis.



Blood lymphocyte subsets in canine idiopathic pericardial effusion

April 2004

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14 Reads

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3 Citations

Veterinary Immunology and Immunopathology

The immunophenotype of peripheral blood lymphocytes was investigated in 23 dogs diagnosed with idiopathic pericardial effusion in order to provide information about a possible role of the immune system in this pathology. Flow cytometric analysis showed a significant reduction in nearly all lymphocyte subsets examined and a strong, significant (P < 0.001) reduction of the CD4 subset, which gave rise to a significantly lower CD4/CD8 ratio. Our data suggest that an imbalance in the immune system is present during the course of the disease, preferentially affecting the T helper cell response.





Effects of Subclinical Bovine Paratuberculosis onIn-vitro Polymorphonuclear Neutrophil Migration

December 1999

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20 Reads

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10 Citations

Journal of Comparative Pathology

Migration of polymorphonuclear neutrophils (PMNs) [unstimulated or stimulated with zymosan-activated serum (ZAS)] from 18 cows was measured in a microwell filter assay. Of these animals, 10 were subclinically infected with Mycobacterium paratuberculosis and shown by culture to be excreting the organism in the faeces; the remaining eight were clinically normal and negative for M. paratuberculosis on faecal culture. PMN "net migration" (stimulated minus unstimulated cells) of the infected cows was significantly lower than that of the uninfected cows. Migration of unstimulated cells in the infected cows did not differ from that in the uninfected cows. It would therefore appear that the infection influenced only the migratory response of the ZAS-stimulated cells. 1999 Harcourt Publishers Ltd.


Citations (10)


... In canine lymphoma, the most common aberrant patterns include co-expression of B and T antigens such as CD3 and CD79a or CD3 and CD21, aberrant expression of CD34, co-expression of CD4 and CD8 in T cell lymphomas and absence of common markers such as CD45 and CD18 (Wilkerson et al., 2005;Guija et al., 2006;Riondato et al., 2006;Gelain et al., 2008) (Fig. 3). In terms of antigen quantification, the percentage of lymphomas expressing aberrant patterns increases when there is under-expression of CD45 or of CD79a in B cell lymphoma. ...

Reference:

Use of flow cytometric immunophenotyping to refine the cytological diagnosis of canine lymphoma
Utility of flow cytometry in the diagnosis of canine small cell lymphoma
  • Citing Article
  • January 2006

Veterinary Clinical Pathology

... In veterinary medicine, only a limited amount of information is available regarding the pattern of LDH isoenzymes related to lymphoma. An increase in the value of serum LDH has been reported in dogs affected by leukemia [85] and in cats with large granular lymphoproliferative disorders [86]. Bezzecchi et al. [87] found that an increase in LDH 3 was possible indicator of latent canine lymphoma. ...

Large granular lymphoma in a FIV-positive and FeLV-negative cat
  • Citing Article
  • June 1992

Journal of Small Animal Practice

... Interestingly following the M. haemolytica challenge, the level of leukocytes in the peripheral blood remained at statistically higher levels throughout the bacte-rial challenge in the vaccine 1 group as compared to vaccine 2 and the control groups (Fig. 3). In stressed cattle, one of the most interesting findings is the movement of leukocytes, particularly neutrophils from the peripheral blood into the lung [49,50]. In stressed cattle infused with a gram-negative endotoxin, the influx of neutrophils in the lung increases dramatically [49]. ...

Effects of road transportation on lymphocyte subsets in calves
  • Citing Article
  • March 2008

The Veterinary Journal

... In cats, one case report has dealt with feline leishmaniasis with a concurrent nasal squamous cell carcinoma [40]. In dogs, visceral leishmaniosis is associated with transmissible venereal tumors [41][42][43] and, in individual cases, with lymphoma [44,45], fibrosarcoma [45], or adrenocortical adenoma [45]. In the presented dog, leishmaniasis was an incidental finding by evaluating the cytological smears taken to differentiate between an inflammatory lesion and a neoplastic disorder in the mammary glands. ...

Extranodal γδ-T-cell lymphoma in a dog with leishmaniasis
  • Citing Article
  • October 2008

Veterinary Clinical Pathology

... These two mechanisms are probably interrelated and may coexist in the same joint [6]. Leishmania infantum can cause either non-erosive [4,[7][8][9] or erosive [4,8,[10][11][12] arthritis. While the presence of bone and joint lesions due to CanVL in plain radiographs has been reported, computed tomography findings have suggested that joint lesions may be common in CanVL but not so commonly described because of the subclinical characteristic observed in most cases [13]. ...

Osteomyelitis and arthrosynovitis associated with Leishmania donovani infection in a dog
  • Citing Article
  • February 1997

Journal of Small Animal Practice

... In contrast, little is known about the possible involvement of neutrophils during paratuberculosis pathogenesis, due to their short lifespan and the unavailability of the mycobacteria sheltered inside macrophages. Nonetheless, some studies have reported that Map infection could induce an early migration of neutrophils to the infection site and a malfunction during neutrophil recruitment [10][11][12]. Indeed, it has been shown that bovine neutrophils are able to release NETs, a purely extracellular antimicrobial mechanism, in response to the in vitro infection with Map [13]. ...

Effects of Subclinical Bovine Paratuberculosis onIn-vitro Polymorphonuclear Neutrophil Migration
  • Citing Article
  • December 1999

Journal of Comparative Pathology

... Our results of M0 support this observation and associate the ratio to the clinical situation [4]. In both infections, the CD4+/CD8+ ratio and the percentage of cats with normal ratio increased when they were treated with rHuIFN-α, contrarily to studies by others [21,37]. In FeLV+ cats, the positive effect of rHuIFN-α was more evident than in FIV+ cats, and lasted throughout the study, unaffected by the "rebound pattern" mentioned above. ...

Effects of Interferon Alpha (INF-α) Therapy on Peripheral Blood Lymphocyte Subsets from FIV and FeLV Naturally Infected Cats
  • Citing Article
  • October 2003

Veterinary Research Communications

... Madewell 14) on the other hand, identified leukopenia in only 9.3-12.5% of dogs with NHL. In the study carried out by Miniscalco et al. 17) on 48 dogs with malignant lymphoma, leucocytosis was detected at varying rates from mild to very severe (18.3-342 × 10 3 μL) in 33% of the cases. In our study, leucocytosis at varying rates (5.3-91.7 × 10 3 μL) in 30% of the cases and leucopenia in 5% of the cases were found. ...

Clinical Usefulness of Peripheral Blood Lymphocyte Subsets in Canine Lymphoma
  • Citing Article
  • October 2003

Veterinary Research Communications

... Flow cytometry is an important research and clinical instrument that helps to identify, quantify and purify subsets of cells rapidly. In addition, rare cell populations can be measured and the technique is used routinely in human and veterinary medicine in a clinical setting (Chattopadhyay et al., 2004;Comazzi et al., 2006;Lugli et al., 2010;Niemeyer et al., 2001). Therefore, flow cytometry was chosen to identify and quantify Th17 cells based on their phenotype through laser technology (Stelzer et al., 1993;Tarrant, 2005;Wilkerson, 2012). ...

Flow cytometric patterns in blood from dogs with non-neoplastic and neoplastic hematologic diseases using double labeling for CD18 and CD45
  • Citing Article
  • April 2006

Veterinary Clinical Pathology

... The magnitude of DNA fragmentation as a morphological hallmark of late apoptosis was determined using an in situ quantitative colorimetric apoptosis detection system "HT Titer-TACS™ Apoptosis Detection Kit" from R & D Systems (R & D Systems, Minneapolis, MN, USA, Catalogue # 4822-96-K), following the manufacturer's guidelines and well-established contemporary protocols [100][101][102][103]. Briefly, AC-16 cells that were terminally sub-cultured and plated in 96-well microplates to the desired confluence (5 × 10 4 cells/well) and subjected to the respective transfection and experimental interventions (as enunciated earlier) were fixed in situ and subsequently 3 -hydroxyl nick-end-labeled with biotin-conjugated dNTPs (deoxynucleotide triphosphates) using the DNA polymerase, Terminal deoxynucleotidyl Transferase (TdT) [65]. ...

Biochemical, ultrastructural and molecular characterization of the triphenyltin acetate (TPTA)-induced apoptosis in primary cultures of mouse thymocytes
  • Citing Article
  • August 2006

Cell Biology and Toxicology