Apoptosis in the transplanted canine transmissible venereal tumor during growth and regression phases

Abstract

Twelve male, mongrel, adult dogs were subcutaneously transplanted with cells originated from two canine transmissible venereal tumors (TVT). The aim was to demonstrate and to quantify the occurrence of apoptosis in the TVT regression. After six months of transplantation, a tumor sample was obtained from each dog, being six dogs with TVT in the growing phase and six in the regression phase as verified by daily measurements. Samples were processed for histological and ultrastructural purposes as well as for DNA extraction. Sections of 4µm were stained by HE, Shorr, methyl green pyronine, Van Gieson, TUNEL reaction and immunostained for P53. The Shorr stained sections went through morphometry that demonstrated an increase of the apoptotic cells per field in the regressive tumors. It was also confirmed by transmission electron microscopy, which showed cells with typical morphology of apoptosis and by the TUNEL reaction that detected in situ the 3'OH nick end labeling mainly in the regressive tumors. The regressive TVTs also showed an intensified immunostaining for P53 besides a more intense genomic DNA fragmentation detected by the agarose gel electrophoresis. In conclusion, apoptosis has an important role in the regression of the experimental TVT in a way that is P53-dependent.Keywords: dog, venereal tumor, apoptosis, cell death RESUMO Doze cães, adultos, machos e sem raça definida foram transplantados subcutaneamente, na região hipogástrica, com células originadas de dois tumores venéreos transmissíveis caninos (TVT). O objetivo do estudo foi demonstrar e quantificar a ocorrência de apoptose na regressão do TVT. Após seis meses

Insertion of the LINE-1 element in the C-MYC gene and immunoreactivity of C-MYC, p53, p21 and p27 proteins in different morphological patterns of the canine TVT

“…It should be highlighted that there was no amplification in the other samples considered in this study. Although Spin et al (2010) have confirmed that this change is characteristic of TVT when compared to the analysis of this same rearrangement in the blood of healthy animals, they did not mention if such occurrence is specific for this neoplastic type. However, the absence of the LINE-1/C-MYC molecular rearrangement in a canine mastocytoma sample of this study suggests that such molecular change is specific to TVT.…”

“…The MYC.S and LINE.A primers prepared for this research were specific for the identification of the amplification of a DNA fragment including approximately 340pb, being in accordance with the methodology by Spin et al (2010) and Fonseca et al (2012). However, it differed from the results presented by other authors like Park et al (2006), who identified a 550pb amplification responsible for the formation of the LINE-1/C-MYC rearrangement in TVT.…”

“…Four-micrometer histological sections were made from the AT2 fragment included in paraffin, and then, they were distended on histological slides and stained by Hematoxylin and Eosin (HE). The histomorphological evaluation of TVT followed the criteria by Santos et al (2008).…”

Cell proliferation markers in the transplanted canine transmissible venereal tumor

“…Similar results were observed during TVT spontaneous (Pérez et al, 1998) and chemotherapy (Gonzalez et al, 2000) induced regression of natural occurrence. This result summed up to the highest apoptotic index observed and this phase (Santos et al, 2008) plays an important role in the natural involution of the TVT.…”

“…The tumor cells were obtained from two mongrel adult male dogs with TVT of natural occurrence, according to Santos et al (2008). Imprints of the tumors were fixed in methanol and stained with May Grünwald-Giemsa (Luna, 1968) to confirm the cytology of the TVT.…”