Cyclic AMP phosphodiesterase (PDE) partially purified from roots of Vigna mungo exhibited optimum activity at pH 5.5 to 6.0 and maximum enzyme activity at 50 degrees C. Levels of PDE activity in roots remained relatively constant from the first to the eleventh day after germination; on the twelfth day there was a 400% increase in PDE activity. The enzyme was stable for at least 48 hours at 28 degrees C, retaining 92% of its original activity. Plant growth hormones including gibberellic acid, indoleacetic acid and kinetin at 1.0 and 10.0 microM concentrations did not have any significant effect on enzyme activity. Nucleotides tested including cyclic 2'3' AMP, cyclic 2'3' GMP completely abolished enzyme activity at 1.0mM while cyclic 3'5' GMP, cyclic 3'5' GMP, 2'deoxy 5' ATP, 2'deoxy 5'GTP and 5'ADP were also inhibitory to the enzyme. The enzyme was stimulated by Mg2+, Fe2+ and NH4+ while Cu2+ and Fe3+ were inhibitory. Theophylline, caffeine, phosphate, pyrophosphate and EDTA were inhibitory to the enzyme.
Heart failure is the end-stage of all cardiovascular diseases with a ~25% 5-year survival rate, and insufficient mitochondrial energy production to meet myocardial demand is the hallmark of heart failure. Mitochondrial components involved in the regulation of ATP production remain to be fully elucidated. Recently, roles of 2',3'-cyclic nucleotide-3'-phosphodiesterase (CNPase) in the pathophysiological processes of heart diseases have emerged, implicated by evidence that mitochondrial CNPase proteins are associated with mitochondrial integrity under metabolic stress. In this study, a zebrafish heart failure model was established, by employing antisense morpholino oligonucleotides and the CRISPR-Cas9 gene-editing system, which recapitulates heart failure phenotypes including heart dysfunction, pericardial edema, ventricular enlargement, bradycardia, and premature death. The translational implications of CNPase in the pathophysiological process of heart failure were tested in a pressure overload-induced heart hypertrophy model, which was carried out in rats through transverse abdominal aorta constriction (TAAC). AAV9-mediated myocardial delivery of CNPase mitigated the hypertrophic response through the specific hydrolysis of 2'-3'-cyclic nucleotides, supported by the decrease of cardiac hypertrophy and fibrosis, the integrity of mitochondrial ultrastructure, and indicators of heart contractility in the AAV9-TAAC group. Finally, the biometrics of a mitochondrial respiration assay carried out on a Seahorse cellular energy analyzer demonstrated that CNPase protects mitochondrial respiration and ATP production from AngII-induced metabolic stress. In summary, this study provides mechanistic insights into CNPase-2',3'-cyclic nucleotide metabolism that protects the heart from energy starvation and suggests novel therapeutic approaches to treat heart failure by targeting CNPase activity.
Obesity and hyperlipidemia are major risk factors for developing vascular diseases. Bee bread (BB) has been reported to exhibit some biological actions, including anti-obesity and anti-hyperlipidemic. This study aims to investigate whether bee bread can ameliorate vascular inflammation and impaired vasorelaxation activity through eNOS/NO/cGMP pathway in obese rats. Forty male Sprague-Dawley rats were randomly divided into four groups (n = 10/group), namely: control (normal group), obese rats (OB group), obese rats treated with bee bread (0.5 g/kg/day, OB/BB group) and obese rats treated with orlistat (10 mg/kg/day, OB/OR group). The latter three groups were given a high-fat diet (HFD) for 6 weeks to induced obesity before being administered with their respective treatments for another 6 weeks. After 12 weeks of the total experimental period, rats in the OB group demonstrated significantly higher Lee obesity index, lipid profile (total cholesterol, triglyceride, low-density lipoprotein), aortic proinflammatory markers (tumor necrosis factor-α, nuclear factor-κβ), aortic structural damage and impairment in vasorelaxation response to acetylcholine (ACh). Bee bread significantly ameliorated the obesity-induced vascular damage manifested by improvements in the lipid profile, aortic inflammatory markers, and the impaired vasorelaxation activity by significantly enhancing nitric oxide release, promoting endothelial nitric oxide synthase (eNOS) and cyclic guanosine monophosphate (cGMP) immunoexpression. These findings suggest that the administration of bee bread ameliorates the impaired vasorelaxation response to ACh by improving eNOS/NO/cGMP-signaling pathway in obese rats, suggesting its vascular therapeutic role.