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. 2023 Dec 13;19(1):271.
doi: 10.1186/s12917-023-03789-9.

Differentially expressed platelet activation-related genes in dogs with stage B2 myxomatous mitral valve disease

Qingqing Zhou #  1 , Xiang Cui #  1 , Han Zhou  1 , Shuai Guo  1 , Zhimin Wu  1 , Liyang Li  1 , Jinxin Zhang  1 , Wen Feng  1 , Yingfang Guo  1 , Xiaofei Ma  2 , Yu Chen  1 , Changwei Qiu  1 , Ming Xu  3 , Ganzhen Deng  4
Affiliations

Differentially expressed platelet activation-related genes in dogs with stage B2 myxomatous mitral valve disease

Qingqing Zhou et al. BMC Vet Res. .

Abstract

Background: Peripheral blood carries a reservoir of mRNAs that regulate cardiac structure and function potential. Although it is well recognized that the typical symptoms of Myxomatous Mitral Valve Disease (MMVD) stage B2 are long-standing hemodynamic disorder and cardiac structure remodeling caused by mitral regurgitation, the transcriptomic alterations in blood from such dogs are not understood.

Results: In the present study, comparative high-throughput transcriptomic profiling of blood was performed from normal control (NC) and naturally-occurring MMVD stage B2 (MMVD) dogs. Using Weighted Gene Co-expression Network Analyses (WGCNA), Gene Ontology (GO), and Kyoto Encyclopedia of Gene and Genomes (KEGG), we identified that the turquoise module was the most highly correlated with echocardiographic features and found 64 differentially expressed genes (DEGs) that were significantly enriched in platelet activation related pathways. Therefore, from the turquoise module, we selected five DEGs (MDM2, ROCK1, RIPK1, SNAP23, and ARHGAP35) that, according to real-time qPCR, exhibited significant enrichment in platelet activation related pathways for validation. The results showed that the blood transcriptional abundance of MDM2, ROCK1, RIPK1, and SNAP23 differed significantly (P < 0.01) between NC and MMVD dogs. On the other hand, Correlation Analysis revealed that MDM2, ROCK1, RIPK1, and SNAP23 genes negatively regulated the heart structure parameters, and followed the same trend as observed in WGCNA.

Conclusion: We screened four platelet activation related genes, MDM2, ROCK1, RIPK1, and SNAP23, which may be considered as the candidate biomarkers for the diagnosis of MMVD stage B2. These findings provided new insights into MMVD pathogenesis.

Keywords: Biomarkers; Dog; MDM2; Myxomatous mitral valve; Transcriptomic.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
Correlation analysis and principal component analysis of the samples. A Heatmap of hierarchical clustering of eight samples (3 NC and 5 MMVD) using Pearson’s method. Bold values represent the R2 (square of the Pearson correlation coefficient) for replicates of each sample. All the P values are below 1e−16. B Principal component analysis plot of the samples. The NC group (yellow triangle) and MMVD group (green dot) samples are labelled with different shapes and colors, and a 95% confidence ellipse for each group is shown. The percentages represent the contribution ratio
Fig. 2
Fig. 2
Key modules correlate with clinical traits through WGCNA. A Analysis of the scale-free fit index (left) and the mean connectively (right) for various soft-thresholding powers. B Clustering dendrogram of genes based on dissimilarity measures, together with assigned module colors. C Heatmap of the correlation between the module and clinical traits of MMVD. Each cell contains the correlation coefficient and P-value
Fig. 3
Fig. 3
DEG identification and expression. A Volcano plot of the DEGs between in the NC and MMVD groups. The X-axis represents the fold change of DEG expression and the Y-axis represents the statistical significance of the fold change. Each dot represents DEG. Red dots represent DEGs with significant upregulation; green dots represent DEGs that are significantly downregulated, and blue dots represent DEGs that are not significant. B Cluster heatmap of the DEGs. The rows and columns represent genes and samples, respectively. The legend represents log2FC of gene abundance. Red and blue represent high and low expression, respectively
Fig. 4
Fig. 4
GO and KEGG enrichment analysis of the DEGs and Venn diagrams of candidate genes. A GO enrichment analysis of the DEGs. X-axis represents the top 10 GO terms and the Y-axis represents the statistical significance of fold change. BP (red) represents biological processes; CC (green) represents cellular components; and MF (blue) represents molecular functions. B The top 20 GO categories in the dogs with MMVD. C KEGG classification of 990 DEGs are summarized. D The top 20 KEGGs in dogs with MMVD
Fig. 5
Fig. 5
A Venn diagram of all DEGs, platelet-activation-related DEGs in KEGG and GO analysis and the top 100 hub genes in the turquoise module
Fig. 6
Fig. 6
Real-time PCR validation of the candidate genes in the validation cohort. (**P < 0.01)
Fig. 7
Fig. 7
Correlation analysis between the radiology parameters and the candidate genes in the validation cohort (**P < 0.01)
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