A modified bioassay system was designed to demonstrate the diffusible nature of endothelium-derived contracting factor(s) released by acetylcholine in the aorta of spontaneously hypertensive rat. In “sandwich”-like layered preparation, isometric tension was recorded from a bioassay strip (without endothelium) in the presence of NG-nitro-L-arginine and tetrahydrobiopterin to selectively potentiate endothelium-dependent contractions. A donor strip (with or without endothelium) was stitched on the bioassay tissue so that it did not directly contribute to the recorded contractions. Acetylcholine induced contractions that occurred only when the donor strip was with endothelium. Superoxide dismutase did not affect but catalase and the combination of superoxide dismutase plus catalase significantly decreased the endothelium-dependent contraction. The contractions in the layered preparations were abolished when the donor strip with endothelium was treated previously with valeryl salicylate, an irreversible cyclooxygenase-1 inhibitor, but remained unaffected when the bioassay strip was treated with the compound. Previous treatment of the bioassay strip alone with S 18886 abolished the contractile response, whereas treatment of the donor strip with endothelium by the selective TP receptor antagonist only produced a moderate inhibition. These results indicate that in the aorta of spontaneously hypertensive rats, endothelium-dependent contractions to acetylcholine involve a diffusible substance(s) released by the endothelium. The production of this contracting factor(s) requires the activation of endothelial cyclooxygenase-1, and its action the activation of TP receptors on the vascular smooth muscle cells.

1 Experiments were designed to investigate whether or not oxygen-derived free radicals mediate endothelium-dependent contractions to acetylcholine in the aorta of spontaneously hypertensive rat (SHR). 2 Isometric tension was measured in aortic rings taken from adult male SHR and Wistar-Kyoto rat (WKY) in the presence of NG-nitro-L-arginine. 3 Endothelium-dependent contractions to acetylcholine were signi

In the aorta of the spontaneously hypertensive rat (SHR), endothelium-dependent contractions are enhanced by inhibitors of NO synthase and scavengers of NO, but not by methylene blue, an inhibitor of guanylyl cyclase, suggesting that the endotheliumderived contracting factor (EDCF) interacts chemically with NO and is inactivated by the latter. However, in view of the relative lack of specificity of methylene blue this hypothesis was re-examined. Acetylcholine-induced endothelium-dependent contractions of isolated rings of SHR aorta were significantly and similarly potentiated by two NOS inhibitors, by two structurally different NO scavengers, by two inhibitors of guanylate cyclase ODQ and NS2028, but to a lesser extent by methylene blue. The contraction of the isolated rat trachea in response to methacholine and the contraction of the rat aorta in response to both 8-isoprostane and KCl were inhibited significantly by methylene blue. Methylene blue binds to the M3 muscarinic receptor subtype but not to the TP receptor. Therefore, methylene blue is an antagonist of the M3 muscarinic receptor subtype, involved in the release of EDCF, and a non-specific inhibitor of TP receptormediated contractions, the receptor involved in the action of EDCF. These inhibitory effects of methylene blue are likely to counteract the effect of the inhibition of soluble guanylate cyclase. These results rule out the hypothesis according to which NO would chemically inactivate EDCF.

This work presented a rapid, inexpensive, reliable, and flexible quantitative immunoassay for cardiac troponin I (cTnI). The assay was based on the concepts of one-step dual monoclonal antibody “sandwich” principle, the low density protein array, the nanogold probe, and the silver enhancement on the gold particles. The capture antibody (IgG1) coated supporting nitrocellulose membrane and the colloidal gold-labeled detection antibody (cAu-IgG2) were prepared before the detection. The detection procedure involved two steps, i.e., immunoreaction and silver amplification. The assay needs only small amounts of serum samples of patients. The whole detection procedure of the assay could be fulfilled within 40 min (much faster than the routine enzyme-linked immunosorbent assay (ELISA) that takes usually at least 3 hours for a turnaround test). The detection results could be easily imaged with a simple flatbed scanner or even observed with the naked eye. The assay showed good specific response to cTnI with very little cross-reactivity to the skeletal isoforms of troponin I (sTnI), cardiac troponin T (cTnT), and myoglobin (Mb). A cut-off value of 0.3 ng/ml was obtained from a reference control group (200 normal serum samples). 588 patients’ serum samples were assayed simultaneously by routine ELISA and this colloidal gold method to test the validity of the method. The data were analyzed using the statistical package SPSS version 11.0 (SPSS Inc.) There was no significant difference between these two assays (P = 0.66 > 0.05). The agreement between this method (≥ or < 0.3 ng/ml) and ELISA was 86%.

AIM: To investigate the changes of cardiac calcium handling proteins and endothelin system in dilated cardiomyopathy (DCM) rats and the effects of perindopril and bisoprolol on the remodeling ventricles. METHODS: DCM rats were employed using a 2-kidney, 1-clip hypertensive and diabetic model. Some of the DCM rats were treated with perindopril and bisoprolol for 3 months, respectively. The ratio of left ventricular weight to body weight (LVW/BW), mRNA expressions of calcium handling proteins and endothelin receptors were determined. The alterations of maximum binding capacity (Bmax) and equilibrium dissociation constant (KD) values of cardiac endothelin receptors (ETR) and its subtypes were detected. RESULTS: Compared with those of normal control, blood pressure, and LVW/BW in the DCM rats were elevated. Sarcoplasmic reticulum calcium pump (SERCA) mRNA expression and SERCA activity decreased in the left ventricle. The ETR Bmax decreased, especially the endothelin receptor A. Endothelin converting enzyme activity and expression were elevated, and mRNA expressions of β1-adrenoreceptor and inositol-3-phosphate receptor in some hearts increased as well. The administration of perindopril and bisoprolol could reverse myocardial hypertrophy and restore the imbalance of calcium handling proteins and endothelin system. CONCLUSION: The disorder of calcium handling proteins and endothelin system existed in the hearts of DCM rats. Treatment of perindopril and bisoprolol could reverse myocardial hypertrophy and changes in DCM rats.