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Necrosis of Skeletal Muscle due to Adenocarcinoma in a Cat: A Case Report
143:4, 2010
ESVP/ECVP Proceedings 2010
335
P4
P5
STUDY OF POLY(ADP-RIBOSE) POLYMERASES 1 AND 2 (PARP-1 AND PARP-2) IN INDUCED LYMPHOMAS IN P21 CIP1/WAF1 AND P-27 KIP2 MICE J.M. Flores *, R.A. Garcia-Fernandez *, P. Garcia-Palencia *, M.A. Sanchez *, B. Sanchez * and J. Martin-Caballeroy *Complutense University and yParc de la Recerc¸a Biomedica de Barcelona, Spain Introduction: p21cip1/waf1 and p27kip1 are able to arrest the cell cycle at the G1 phase. There is evidence that p21 modulates apoptosis and p27 deficiency is related to thymic hyperplasia and T lymphocyte proliferation. Poly (ADP-ribose) polymerase (PARP) is one of the first mechanisms to participate in DNA repair after irradiation. The aim of this study was to analyze the effect of p21 and p27 proteins on apoptosis using a double-deficient mouse model. Materials and Methods: Twenty-four mice of four different genotypes were g-irradiated to induce thymic lymphomas. Immunohistochemistry was performed using anti-Pax1, anti-CD3, anti-PARP 1, anti-PARP 2, caspase-3 active and Ki67 antibodies. Results: Lymphomas were diagnosed in mice of the following genotypes: p21+/+p27+/+ (n 5 7), p21+/+p27-/- (n 5 3), p21-/-p27 +/+ (n 5 4) and p21-/-p27-/- (n 5 3). In wild-type and p27-null mice all of the tumours were T-cell lymphomas, while in 21-null and double-deficient mice the majority of lymphomas (75%) were B cell in phenotype. We observed strong apoptosis (caspase-3 active) in the thymus of wild-type and p21-null mice, whereas 27-null mice and double knockout mice were more resistant to apoptosis. This result was correlated with the level of immune expression of cleaved PARP-1 in lymphocytes. PARP-2 detection was practically absent in all of the tumours analyzed. No differences were observed in the mitotic (ki67) indices between groups. Conclusions: PARP-1, but not PARP-2, plays an essential role in the apoptosis observed in induced lymphomas in p21-null mice and in a lower proportion in p27-null mice and double-knockout mice.
ESTABLISHMENT AND CHARACTERIZATION OF NEW CELL LINES DERIVED FROM CANINE HAEMANGIOSARCOMAS A. Murai, A. Kodama, K. Maruo, T. Yanai and H. Sakai Gifu University, Japan Introduction: Recently, we created canine haemangiosarcoma (HSA) xenograft models by using nude mice. In the present study we established and characterized the canine HSA xenograft tumour cell lines. Materials and Methods: Tissues of four canine HSA xenograft tumours were minced and digested enzymatically. The cells were cultured in M199 medium with 10% fetal calf serum (FCS) and passaged over 60 times. Uptake of Dil-acetylated low-density lipoprotein (Dil-LDL) by these cells was evaluated. The expression of endothelial markers, angiogenic growth factors and their receptors and angiogenic homeobox genes was investigated by RT-PCR with canine-specific primer sets and immunocytochemical analysis. Results: We established six canine HSA cell lines. RT-PCR and immunocytochemical analyses revealed the expression of von Willebrand factor and/or CD31. Further, all of the cells exhibited active uptake of dil-acetylated LDL. RT-PCR and immunocytochemical analyses revealed the expressions of VEGF-A, bFGF, flt-1, flk-1, FGFR-1, HoxA9, HoxB3, HoxB7, HoxD3, Pbx1 and Meis1. Conclusions: All of the cells expressed endothelial markers, angiogenic growth factors and their receptors and angiogenic homeobox genes similar to that observed in the original xenograft tumour tissues. These cell lines may be useful to study the malignant progression of canine HSA.
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NECROSIS OF SKELETAL MUSCLE DUE TO ADENOCARCINOMA IN A CAT: A CASE REPORT I. Babinska *, J. Szarek *, B. Parysiewicz-Szarek y, M. Gesek * and M. Szweda* *University of Warmia and Mazur and yPrivate Veterinary Practice ‘Pulsar’, Poland Introduction: Coagulation necrosis of skeletal muscle due to embolism of an adenocarcinoma in animals is uncommon. Materials and Methods: Necropsy examination was performed of a 14-year-old female European cat, body weight 3.6 kg, humanely destroyed during diagnostic laparotomy. For several years the cat had received medroxyprogesterone acetate (Provera VetÔ, one 5 mg tablet weekly; Pfizer Poland). Radiography had revealed a mass in the stomach region. Samples of this mass, the alimentary tract and muscles of the abdominal wall were taken for microscopical examination (HE staining). Results: The tumour was present on the mesentery (6 4 3 cm), touching the stomach, spleen and pancreas. The serosal surfaces of the alimentary tract, urinary bladder and mesentery were covered with numerous straw-coloured small, firm nodules. On the capsule of the liver and the spleen there were numerous deposits similar to fibrin. Microscopical examination of the tumour and nodules revealed well- and moderately-differentiated adenocarcinoma, respectively. The cells were columnar, with foamy cytoplasm. Large secretory vacuoles were present in some cells. The nuclei were parabasal and were sometimes arranged chaotically. Numerous mitotic figures and aniso- and macrocaryosis were observed. There was secretion present in the follicles. The parietal peritoneum revealed fibrosis with neoplastic infiltration. Within the muscles of the abdominal wall there were areas of coagulative necrosis that were a consequence of peripheral arterial tumour embolism. Conclusions: Such muscular lesions in animals are usually associated with underlying cardiac disease, whereas in man the association is primarily with pulmonary neoplasia.
A POORLY DIFFERENTIATED PRIMARY HEPATIC SARCOMA IN A CAT E. Loeb *, Y. Semrik *, E. Nachmias y and J. Cullenz *BactoChem Ltd., yDerma, Israel and zCollege of Veterinary Medicine, North Carolina, USA Introduction: A 9-year-old DSH cat in a good physical condition died suddenly and was brought to a private veterinary clinic. The clinical assessment of the veterinarian was a suspicion of intoxication because of the rapid nature of the deterioration in condition and a history of changing food management by the cat’s owners one day before death. Materials and Methods: Necropsy examination was performed and samples from the internal organs were fixed in 10% neutral buffered formalin. Microscopical sections were stained with haematoxylin and eosin. Results: Within the abdominal cavity there was approximately 1 l of free blood. Adjacent to the left ventral liver lobe, a large, solitary mass (15 cm diameter) was found. This had an irregular structure on cross section, with many small blood-filled cavities. Both kidneys had an irregular cortex and were swollen. Microscopically the liver mass was infiltrative, densely cellular and unencapsulated. The cells were spindle-shaped, poorly differentiated and arranged in bundles. They had an eosinophilic finely granular cytoplasm with a large elongate, stippled nucleus with 1e2 nucleoli. Conclusions: The mass had features consistent with an undifferentiated hepatic sarcoma. Immunohistochemical studies are currently being performed to further evaluate this lesion.
ESVP/ECVP Proceedings 2010
335
P4
P5
STUDY OF POLY(ADP-RIBOSE) POLYMERASES 1 AND 2 (PARP-1 AND PARP-2) IN INDUCED LYMPHOMAS IN P21 CIP1/WAF1 AND P-27 KIP2 MICE J.M. Flores *, R.A. Garcia-Fernandez *, P. Garcia-Palencia *, M.A. Sanchez *, B. Sanchez * and J. Martin-Caballeroy *Complutense University and yParc de la Recerc¸a Biomedica de Barcelona, Spain Introduction: p21cip1/waf1 and p27kip1 are able to arrest the cell cycle at the G1 phase. There is evidence that p21 modulates apoptosis and p27 deficiency is related to thymic hyperplasia and T lymphocyte proliferation. Poly (ADP-ribose) polymerase (PARP) is one of the first mechanisms to participate in DNA repair after irradiation. The aim of this study was to analyze the effect of p21 and p27 proteins on apoptosis using a double-deficient mouse model. Materials and Methods: Twenty-four mice of four different genotypes were g-irradiated to induce thymic lymphomas. Immunohistochemistry was performed using anti-Pax1, anti-CD3, anti-PARP 1, anti-PARP 2, caspase-3 active and Ki67 antibodies. Results: Lymphomas were diagnosed in mice of the following genotypes: p21+/+p27+/+ (n 5 7), p21+/+p27-/- (n 5 3), p21-/-p27 +/+ (n 5 4) and p21-/-p27-/- (n 5 3). In wild-type and p27-null mice all of the tumours were T-cell lymphomas, while in 21-null and double-deficient mice the majority of lymphomas (75%) were B cell in phenotype. We observed strong apoptosis (caspase-3 active) in the thymus of wild-type and p21-null mice, whereas 27-null mice and double knockout mice were more resistant to apoptosis. This result was correlated with the level of immune expression of cleaved PARP-1 in lymphocytes. PARP-2 detection was practically absent in all of the tumours analyzed. No differences were observed in the mitotic (ki67) indices between groups. Conclusions: PARP-1, but not PARP-2, plays an essential role in the apoptosis observed in induced lymphomas in p21-null mice and in a lower proportion in p27-null mice and double-knockout mice.
ESTABLISHMENT AND CHARACTERIZATION OF NEW CELL LINES DERIVED FROM CANINE HAEMANGIOSARCOMAS A. Murai, A. Kodama, K. Maruo, T. Yanai and H. Sakai Gifu University, Japan Introduction: Recently, we created canine haemangiosarcoma (HSA) xenograft models by using nude mice. In the present study we established and characterized the canine HSA xenograft tumour cell lines. Materials and Methods: Tissues of four canine HSA xenograft tumours were minced and digested enzymatically. The cells were cultured in M199 medium with 10% fetal calf serum (FCS) and passaged over 60 times. Uptake of Dil-acetylated low-density lipoprotein (Dil-LDL) by these cells was evaluated. The expression of endothelial markers, angiogenic growth factors and their receptors and angiogenic homeobox genes was investigated by RT-PCR with canine-specific primer sets and immunocytochemical analysis. Results: We established six canine HSA cell lines. RT-PCR and immunocytochemical analyses revealed the expression of von Willebrand factor and/or CD31. Further, all of the cells exhibited active uptake of dil-acetylated LDL. RT-PCR and immunocytochemical analyses revealed the expressions of VEGF-A, bFGF, flt-1, flk-1, FGFR-1, HoxA9, HoxB3, HoxB7, HoxD3, Pbx1 and Meis1. Conclusions: All of the cells expressed endothelial markers, angiogenic growth factors and their receptors and angiogenic homeobox genes similar to that observed in the original xenograft tumour tissues. These cell lines may be useful to study the malignant progression of canine HSA.
P6
P7
NECROSIS OF SKELETAL MUSCLE DUE TO ADENOCARCINOMA IN A CAT: A CASE REPORT I. Babinska *, J. Szarek *, B. Parysiewicz-Szarek y, M. Gesek * and M. Szweda* *University of Warmia and Mazur and yPrivate Veterinary Practice ‘Pulsar’, Poland Introduction: Coagulation necrosis of skeletal muscle due to embolism of an adenocarcinoma in animals is uncommon. Materials and Methods: Necropsy examination was performed of a 14-year-old female European cat, body weight 3.6 kg, humanely destroyed during diagnostic laparotomy. For several years the cat had received medroxyprogesterone acetate (Provera VetÔ, one 5 mg tablet weekly; Pfizer Poland). Radiography had revealed a mass in the stomach region. Samples of this mass, the alimentary tract and muscles of the abdominal wall were taken for microscopical examination (HE staining). Results: The tumour was present on the mesentery (6 4 3 cm), touching the stomach, spleen and pancreas. The serosal surfaces of the alimentary tract, urinary bladder and mesentery were covered with numerous straw-coloured small, firm nodules. On the capsule of the liver and the spleen there were numerous deposits similar to fibrin. Microscopical examination of the tumour and nodules revealed well- and moderately-differentiated adenocarcinoma, respectively. The cells were columnar, with foamy cytoplasm. Large secretory vacuoles were present in some cells. The nuclei were parabasal and were sometimes arranged chaotically. Numerous mitotic figures and aniso- and macrocaryosis were observed. There was secretion present in the follicles. The parietal peritoneum revealed fibrosis with neoplastic infiltration. Within the muscles of the abdominal wall there were areas of coagulative necrosis that were a consequence of peripheral arterial tumour embolism. Conclusions: Such muscular lesions in animals are usually associated with underlying cardiac disease, whereas in man the association is primarily with pulmonary neoplasia.
A POORLY DIFFERENTIATED PRIMARY HEPATIC SARCOMA IN A CAT E. Loeb *, Y. Semrik *, E. Nachmias y and J. Cullenz *BactoChem Ltd., yDerma, Israel and zCollege of Veterinary Medicine, North Carolina, USA Introduction: A 9-year-old DSH cat in a good physical condition died suddenly and was brought to a private veterinary clinic. The clinical assessment of the veterinarian was a suspicion of intoxication because of the rapid nature of the deterioration in condition and a history of changing food management by the cat’s owners one day before death. Materials and Methods: Necropsy examination was performed and samples from the internal organs were fixed in 10% neutral buffered formalin. Microscopical sections were stained with haematoxylin and eosin. Results: Within the abdominal cavity there was approximately 1 l of free blood. Adjacent to the left ventral liver lobe, a large, solitary mass (15 cm diameter) was found. This had an irregular structure on cross section, with many small blood-filled cavities. Both kidneys had an irregular cortex and were swollen. Microscopically the liver mass was infiltrative, densely cellular and unencapsulated. The cells were spindle-shaped, poorly differentiated and arranged in bundles. They had an eosinophilic finely granular cytoplasm with a large elongate, stippled nucleus with 1e2 nucleoli. Conclusions: The mass had features consistent with an undifferentiated hepatic sarcoma. Immunohistochemical studies are currently being performed to further evaluate this lesion.