Circulating MiR-30b-5p is upregulated in Cavalier King Charles Spaniels affected by early myxomatous mitral valve disease

Abstract

There is a growing interest in developing new molecular markers of heart disease in young dogs affected by myxomatous mitral valve disease. The study aimed to measure 3 circulating microRNAs and their application as potential biomarkers in the plasma of Cavalier King Charles Spaniels with early asymptomatic myxomatous mitral valve disease. The hypothesis is that healthy Cavalier King Charles Spaniels have different microRNA expression profiles than affected dogs in American College of Veterinary Internal Medicine (ACVIM) stage B1. The profiles can differ within the same class among subjects of different ages. This is a prospective cross-sectional study. Thirty-three Cavalier King Charles Spaniels in ACVIM stage B1 were divided into three groups (11 younger than 3 years, 11 older than 3 years and younger than 7 years, and 11 older than 7 years), and 11 healthy (ACVIM stage A) dogs of the same breed were included as the control group. Three circulating microRNAs (miR-1-3p, miR30b-5p, and miR-128-3p) were measured by quantitative real-time PCR using TaqMan® probes. Diagnostic performance was evaluated by calculating the area under the receiver operating curve (AUC). MiR-30b-5p was significantly higher in ACVIM B1 dogs than in ACVIM A subjects, and the area under the receiver operating curve was 0.79. According to the age of dogs, the amount of miR-30b-5p was statistically significantly higher in group B1<3y (2.3 folds, P = 0.034), B1 3-7y (2.2 folds, P = 0.028), and B1>7y (2.7 folds, P = 0.018) than in group A. The area under the receiver operating curves were fair in discriminating between group B1<3y and group A (AUC 0.780), between B1 3-7y and A (AUC 0.78), and good in discriminating between group B1>7y and A (AUC 0.822). Identifying dogs with early asymptomatic myxomatous mitral valve disease through the evaluation of miR-30b-5p represents an intriguing possibility that certainly merits further research. Studies enrolling a larger number of dogs with preclinical stages of myxomatous mitral valve disease are needed to expand further and validate conclusively the preliminary findings from this report.

Fig 1. Expression levels of miR-1-3p, miR-30b-5p, and miR-128-3p between groups.

Expression levels of the three miRNAs between groups A and B1 (A-C, respectively) and between A and B1 were divided according to the age at MMVD diagnosis (D-F, respectively). MiR-30b-5p increased 2.4 folds (P < 0.05) in group B1 compared to A (B). Splitting group B1 according to the age of dogs, the expression of miR-30b-5p remained significantly higher (E). Group B1<3 (2.3 folds, P = 0.034), B1 3–7 (2.2 folds, P = 0.028), and B1>7 (2.7 folds, P = 0.018) expressed a higher level of miR-30b-5p than group A. No differences were found in the amount of miR-1-3p (A and D) and miR-128-3p (C and F).

Fig 2. ROC curves for miR-30b-5p.

Discrimination capacity between group A and group B1 (A), group A and group B1<3 (B), group A and group B1 3–7 (C), and group A and B1>7 (D). MiR-30b-5p can discriminate between healthy and asymptomatic MMVD-affected dogs (stage B1).