Proteomic analysis of pulmonary arteries and lung tissues from dogs affected with pulmonary hypertension secondary to degenerative mitral valve disease

Abstract

In dogs with degenerative mitral valve disease (DMVD), pulmonary hypertension (PH) is a common complication characterized by abnormally elevated pulmonary arterial pressure (PAP). Pulmonary arterial remodeling is the histopathological changes of pulmonary artery that has been recognized in PH. The underlying mechanisms that cause this arterial remodeling are poorly understood. This study aimed to perform shotgun proteomics to investigate changes in protein expression in pulmonary arteries and lung tissues of DMVD dogs with PH compared to normal control dogs and DMVD dogs without PH. Tissue samples were collected from the carcasses of 22 small-sized breed dogs and divided into three groups: control (n = 7), DMVD (n = 7) and DMVD+PH groups (n = 8). Differentially expressed proteins were identified, and top three upregulated and downregulated proteins in the pulmonary arteries of DMVD dogs with PH including SIK family kinase 3 (SIK3), Collagen type I alpha 1 chain (COL1A1), Transforming growth factor alpha (TGF-α), Apoptosis associated tyrosine kinase (AATYK), Hepatocyte growth factor activator (HGFA) and Tyrosine-protein phosphatase non-receptor type 13 (PTPN13) were chosen. Results showed that some of the identified proteins may play a role in the pathogenesis of pulmonary arterial remodeling. This study concluded shotgun proteomics has potential as a tool for exploring candidate proteins associated with the pathogenesis of PH secondary to DMVD in dogs.

Fig 1. Venn diagram showed the distribution and overlap of identified proteins.

Proteins in the pulmonary arteries (a) and lung tissues (b) of dogs in the control, DMVD and DMVD+PH groups. The proteins that commonly found between groups were pointed out as circle. In the pulmonary arteries, 342 proteins were commonly found in the control and DMVD groups and 576 proteins were commonly found in the DMVD and DMVD+PH groups. In the lung tissues, 692 proteins were commonly found in the control and DMVD groups and 312 proteins were commonly found in the DMVD and DMVD+PH groups.

Fig 2. Volcano plot for the identified proteins that were commonly found.

Protein in the pulmonary arteries and lung tissues of dogs in the control and DMVD groups (a, c) and in the DMVD and DMVD+PH groups (b, d). Each dot represents an individual protein, red dots were significant upregulated proteins and blue dots were significant downregulated proteins compared between groups. Vertical lines indicate 2-fold difference and horizontal line indicates p-value = 0.05.

Fig 3. Venn diagram showed the commonly found proteins.

Proteins in the control and DMVD groups (a) and the commonly found proteins in the DMVD and DMVD+PH groups (b) that expressed in both pulmonary arteries and lung tissues were displayed as circle. Eighteen commonly found proteins in the control and DMVD groups and 16 commonly found proteins in the DMVD and DMVD+PH groups were expressed in both pulmonary arteries and lung tissues.

Fig 4. Involvement of the differentially expressed proteins in the networks of proteins- cardiovascular drugs interactions.

The edge confidence scores were used to assess the strength of these pathway interactions at functional level. The interactions with high edge confidence scores (>0.700) presenting as thick lines were indicated strong relationships. In the pulmonary arteries, the differentially expressed proteins between the control and DMVD groups were not associated with any common cardiovascular drugs (a), whereas the differentially expressed proteins between the DMVD and DMVD+PH groups including SCO-spondin (SSPO; circled) were weakly associated with sildenafil (b). In the lung tissues, the differentially expressed proteins between the control and DMVD groups showed that Glutamate metabotropic receptor 7 (GRM7) and 8 (GRM8) were associated pimobendan (c) while no differentially expressed proteins between the DMVD and DMVD+PH groups were associated with common cardiovascular drugs (d).

Fig 5. Venn diagram showed the uniquely expressed proteins.

Proteins in the pulmonary arteries and lung tissues of dogs in the DMVD groups (a) and the DMVD+PH groups (b). Six uniquely expressed proteins in the DMVD group and 16 uniquely expressed proteins in the DMVD+PH group displayed as circle were expressed in both pulmonary arteries and lung tissues.