BNP or NT-proBNP evaluation is the strongest indicator to confirm the diagnosis of congestive heart failure in humans [14]. These factors are also considered as prognostic parameters [15]. Several studies have reported that circulating BNP is increased in dogs with CHF due to MMVD. Our findings are consistent with those reported in previous studies. Also, in humans, BNP is an important diagnostic indicator for identifying patients with subclinical (asymptomatic) left ventricular dysfunction [14]. In this regard, different results have been reported in dogs with MMVD. Asano et al. described that plasma BNP was high only in dogs with moderate and severe CHF [16]. Häggström et al. reported that the BNP concentrations were increased (approximately twice) only in dogs with decompensated heart failure compared to normal and asymptomatic dogs [17]. However, MacDonald et al. reported that plasma BNP was high in dogs with moderate to severe MMVD in the absence of CHF. They also found that plasma BNP was higher in dogs with CHF due to MMVD [18]. In the present study, the mean serum BNP levels increase with the severity of MMVD. Our results showed that the mean serum BNP levels were significantly increased for all MMVD groups compared to control dogs.
In dogs, various studies have shown that circulating ANP levels increase with the severity of MMVD and the highest levels of ANP are seen in patients with stages C/D [4, 17, 19]. Our findings were consistent with those results. In our study, although the mean serum ANP levels in stage B2 (subclinical MMVD), C and D groups were significantly higher than the control group, the mean serum ANP concentrations did not differ significantly between the stage B1 (earliest subclinical MMVD) and control groups. Different reports have been presented in this regard. Some reports indicated that plasma ANP level was significantly higher for all MMVD groups than control dogs [4, 20], while a study in Cavalier King Charles Spaniels with MMVD found that plasma ANP levels were not increased significantly in asymptomatic dogs; but in dogs with decompensated heart failure, the mean ANP level was 3 to 7 times higher than control dogs [17]. These differences between the results of the present study with other reports may be due to differences in the BNP and ANP measurement methods, populations studied as well as the criteria used to classify the different stages of MMVD. Regarding the ANP measurement method, in our study, serum ANP was measured by a canine-specific ELISA kit, but in other studies, plasma ANP was determined using chemiluminescent enzyme immunoassay for human αANP [4], and radioimmunoassay [17].
Our results showed that increased serum BNP levels were observed in all cardiac patients except 3 patients in stage B1 group. However, in these 3 patients, BNP concentrations were within maximum normal values. Therefore, an increase in BNP level was observed in 87.5% (21/24) of patients. Although BNP concentrations were elevated in 100% of dogs with stages C, D and B2, high BNP was observed in 72.72% of dogs with stage B1. Regarding ANP, elevated ANP levels were found in only 27.27% (3/11) of patients in stage B1, while increased serum ANP levels were observed in 66.66% (4/6) of patients in the stage B2 group. However, in the C and D groups, 100% of patients had an increase in ANP. Therefore, increased ANP was also observed in 58.33% (14/24) of patients with MMVD. Because both the increase in BNP and the percentage of patients who showed an increase in serum BNP were significantly higher, serum BNP appears to have a greater diagnostic value than serum ANP in dogs with MMVD. In human, the indication for measuring BNP or NT-proBNP is detection of asymptomatic (occult or subclinical) heart disease. Our findings are consistent with the results of human research. Our results suggest that serum BNP may be a useful diagnostic marker for MMVD in dogs and can be used to determine the subclinical (asymptomatic) stages of B1 and B2 MMVD. Therefore, early diagnosis in asymptomatic patients can probably prevent the progression of the disease [14].
Echocardiographic variables and VHS are useful indices of disease severity in dogs with MMVD [2]. A previous study has found that only LA/AO ratio had a significant variable with the plasma ANP, NT-proBNP and NT-proANP [21]. However, in our study, serum natriuretic peptide (BNP and ANP) concentrations had a significant positive correlation with echocardiographic variables (LA/AO ratio, LVIDd, LVIDdN) and VHS. Mitral regurgitation is the earliest hemodynamic event. The progression of the disease and the increasing severity of valvular regurgitation generate a volume overload of the left heart, leading to left atrial and ventricular remodeling. In advanced stages of MMVD, associated volume overload promotes progressive valvular regurgitation, left atrial and left ventricular remodeling, atrial rupture, and CHF [22]. However, the difference between BNP and ANP secretion sources in normal and heart injury is still unclear, it seems that ventricles are the important source of natriuretic secretion, especially BNP, but ANP and BNP are mainly produced by the atria in healthy humans [23], and CHF patients [18]. In left ventricular dysfunction, when chronic hemodynamic pressure or volume overload occurs (such as ventricular hypertrophy), the ventricular myocytes undergo phenotypic modifications and re-express several fetal genes, including ANP and BNP [23, 24].
In humans, hyperlipidemia is a major risk factor for cardiovascular disease. In this regard, elevated LDL-C [25], and decreased HDL-C lead to atherogenic hyperlipidemia [26]. In the current study, mean HDL-C concentrations decreased and mean LDL-C concentrations increased in all dogs with acquired MMVD (B1, B2, C and D stages) compared to the control group. Additionally, the lowest mean of HDL-C and the highest mean of LDL-C were observed in the stage D group. Research has shown that elevated LDL-C in the blood leads to their accumulation as plugs in the vascular intimal. Fat particles, calcium and tissue macrophages play a major role in plug formation. However, plug formation reduces the elasticity and narrows the internal space of the arteries and eventually leads to blockage of the arteries or may even lead to rupture and eventually blood clots in the area [26]. HDL-C has anti-atherogenic effects. The anti-atherogenic effects of HDL-C are mediated through the reverse transport of cholesterol [27], and carriers of LDL- reducing potential oxidative proteins such as paraoxonase [28], and platelet activating factor acetyl hydrolase [29], as well as antioxidative and anti-inflammatory effects [30]. Therefore, it maintains endothelial integrity by using the above mechanisms [30]. HDL-C has also been shown to have antithrombotic and profibrinolytic effects [7]. These protective effects may be attenuated by lower blood HDL concentrations, which increases the risk of cardiovascular disease [30]. Although dogs appear to be resistant to atherosclerosis due to their heart structure (a well-developed coronary lateral circulation) [31], and lipoprotein composition [32], it has previously been described that arteriosclerotic changes and small myocardial infarcts are common in geriatric dogs with or without primary mitral regurgitation [33]. Also, Falk et al. reported that dogs with naturally MMVD had significantly more arterial changes and fibrosis in the myocardium than control dogs [34]. Therefore, Coronary arterial disease should also be considered in dogs with mitral regurgitation, which may contribute to reduced myocardial contractility [10]. However, the clinical and diagnostic value of blood lipoproteins in dogs with MMVD is still unclear. Our findings showed that dyslipidemia in dogs with MMVD (all stages) is mainly related to high LDL and low HDL.
We found a significant negative correlation between serum concentrations of BNP and HDL-C. This finding is similar to Takeuchi and Sata’s observation on human patients with cardiovascular disease [35]. Although, their result was not statistically significant. However, Lupattelli et al. reported a positive correlation between HDL-C and BNP in hyperlipemic humans [7]. Regarding LDL-C, our results showed that there is a significant positive correlation between BNP and LDL-C. In the present study, serum ANP, like BNP, had a negative correlation with HDL-C and a positive correlation with LDL-C. But it seems that high LDL-C is more involved in the pathogenesis of MMVD than low HDL-C. Since in the present study, LDL-C, but not HDL-C, had a significant positive correlation with echocardiographic variables, high LDL-C is more involved in the pathogenesis of MMVD than low HDL-C. However, further research is needed to determine the role of dyslipidemia on serum natriuretic peptides.