Mutation and up-regulation of TP53 in ovine pulmonary adenocarcinoma lung cells as a model of human lung cancer

Abstract

Ovine pulmonary adenocarcinoma (OPA) is a model of human lung cancer‎ and fatal viral disease that causes neoplasia in sheep respiratory cells. ‎In the current study, 986 lung samples was inspected in the slaughterhouse, and finally twenty OPA ‎ lung organs were clinically diagnosed and five healthy lung organs were assigned as the control sample. Three SSCP patterns were detected for the affected lungs animals in comparison with the healthy lungs. In addition, sequencing results indicated three different single point mutations in exon 4 of TP53 within infected lungs, whereas no mutations were observed in exon 9 of this gene. Real-time PCR results showed up-regulation of the TP53 gene in all the infected lung cells compared to healthy cells. There was significant correlation between the mutations in exon 4 and OPAand can be used as a useful tool in determining the mechanism of lung cancer.

Keywords: Mutation; Ovine pulmonary adenocarcinoma; Pathology; TP53 gene; Up-regulation.

Fig. 1

Macroscopic view of the lung suffering from ovine pulmonary adenocarcinoma (OPA). An affected lung with multifocal nodules (a) and a light gray, local extensive mass (b) (the diaphragmatic lobe of left lung)

Fig. 2

Microscopic view of the lung suffering from OPA using Hematoxylin and Eosin stain. A) Histopathologic section of OPA, well-differentiated pulmonary carcinoma with lepidic and papillary patterns (bar = 300 µm). B) Alveoli (a) and terminal brochiol (b) are lined with cuboidal or columnar neoplastic epithelial cells and in papillary pattern, papillary projections into the alveoli and bronchioles are observed in lepidic pattern (bar = 60 µm). C and D) Neoplastic cells were observed in parafollicular zone of bronchial and mediastinal lymph nodes (bars = 300 µm)

Fig. 3

A) The PCR products of exons 4 of TP53 gene on 1.50% agarose gel. M: DNA ‎Ladder (Jena Biosciences, Jena, Germany); Lanes 1-5: The PCR products in infected samples to OPA; lanes 6-10: PCR products of healthy samples. Both infected and healthy samples showed 310 bp‎fragment. ‎B) The PCR products of exons 9 of TP53 gene on 1.50% agarose gel.‎‎ M: DNA Ladder ‎(Jena Biosciences); Lanes 1-5: PCR products in infected samples to OPA; Lanes 6-10: PCR products of healthy samples. Both infected and healthy samples showed 193 bp‎fragment

Fig. 4

A) The SSCP patterns of exons 9 and 4 of TP53 gene in the healthy samples. M: DNA ‎ Ladder, 50-1500 bp (Jena Biosciences); observed SSCP patterns of exons 9 and 4 in healthy samples. B) Some observed patterns of exon 9 of TP53 gene in the infected and healthy samples. Lanes 1-3: SSCP patterns in the healthy samples, Lanes 4-10: SSCP patterns in the infected samples. It was not observed different SSCP patterns among healthy and infected samples. ‎C) Some SSCP patterns of exons 4 of TP53 gene in the infected and healthy samples. M: DNA Ladder, 50-‎1500 bp (Jena Biosciences); Lane 1: A patterns of SSCP; Lane 2: B patterns of SSCP, and Lane 3: C patterns of SSCP; Lanes 4-5: SSCP pattern of healthy samples

Fig. 5

The TP53 gene relative expression in the observed A, B, and C patterns in the ‎infected samples. Expression of p53 gene in the infected samples content C pattern has highest. All of the infected samples (with A, B, and C patterns) have significant differences compared to control groups (p ≤ 0.05). The error bars show mean ± SEM.‎ ^abc TP53 gene relative expression with different subscript was significantly different at p < 0.05