acid decreases total ROS and improves NO level in EA.hy926 cells.

<p>10 µM SA was pre-treated for 24 h prior to incubation of cells with 300 µM of H₂O₂ for 4 h. (A) Intracellular total ROS level was measured by the fluorescent probe DCFH-DA and the images were obtained by fluorescence microscopy. (B) NO level was measured by the fluorescent probe DAR-4M AM and the images were obtained by fluorescence microscopy. The representative images from three independent experiments are shown. (C) Total ROS fluorescence intensity value was calculated using Adobe Photoshop version 7.0. (D) Fluorescence intensity measurement against NO was calculated using Adobe Photoshop version 7.0. RFU: Relative Fluorescence Unit. Values are expressed as mean ± SEM. ^*<i>P</i><0.05 vs control; ^#<i>P</i><0.05 vs H₂O₂.</p

Creating partial ischemia in chick embryo vascular bed using ligation model.

<p>(A) Image showed the embryo with the whole yolk material has been placed in the sterile glass bowls. The bowls were covered with sterile glass leads and placed in a chamber maintaining 37°C and relative humidity. (B) Image represents the exact approach taken to block the blood flow to the specific area of the chick embryo vascular bed. Black arrow showed the suture that has ligated the right vitelline artery thus blocking the blood flow in the surrounding vascular bed. The box showed the ischemic area of the vascular bed.</p

Sinapic acid prevents deregulated expression of cardiovascular genes.

<p>(A) Relative expression fold changes of TGF-β and β-MHC mRNAs in heart. (B) Relative expression negative fold change of eNOS mRNA in aorta. (C) Differential eNOS protein expression in aorta and its normalized value with β-actin. Values are expressed as means ± SD. All experiments were done in triplicates. ^*<i>P</i><0.05 vs control; ^#<i>P</i><0.05 vs L-NAME.</p

HIF-1α level in ischemic EC.

<p>(A) HIF-1α production was measured in EC treated for different time under ischemia. Cells were grown on collagen coated cover glasses and placed on the top of the ischemic vascular bed of chick embryo. RNA was isolated from the ischemia treated EC using RNA extraction kit. Loading of total RNA for cDNA conversion was normalized by running a gel for total RNA and calculating the band intensities for each set. After amplification, the band intensities from the representative images were calculated and plotted as a graph. Ischemia induced HIF-1α expression in EC in a time dependent manner. **P<0.001 significantly different than 0 min. (n = 5) (B) RL was administered to evaluate its effect on ischemia induced HIF-1α expression by EC. EC grown on collagen coated cover glasses were placed on ischemic chick embryo vascular bed followed by RNA extraction from the EC using RNA extraction kit. Loading of total RNA for cDNA conversion was normalized by running a gel for total RNA and calculating the band intensities for each set. RL attenuated ischemia induced HIF-1α expression by EC. **P<0.001 versus ischemic. (n = 5).</p

Sinapic acid improved cardiovascular function in experimental hypertensive rats.

<p>(A) Evaluation of ventricular function in heart of various experimental groups. (B) Cumulative concentration-response curves of Ach induced relaxation in endothelium-intact aortic rings. (C) Cumulative concentration-response curves of SNP induced relaxation in endothelium-intact aortic rings. Values are expressed as mean ± SD. n = 6 per group. ^*<i>P</i><0.05 vs control; ^#<i>P</i><0.05 vs L-NAME.</p

Effect of ischemia on GSH level in vitelline vascular tissue.

<p>(A) Level of GSH was measured using CMFDA fluorescent probe. The ischemic vessels were incubated with CMFDA probe for 30 min. Next, the images were acquired with DP71 CCD camera. Images are the representative of 5individual experiments. (B) The fluorescent intensities of the images were calculated using Adobe Photoshop 7.0 and plotted. Fluorescent intensities were considered as directly proportional to the level of GSH in the vessels. **P<0.001 versus non-ischemic. (n = 5) (C) Ischemic tissues were probed with CMFDA for 30 min. Tissues were then homogenized and centrifuged at 6500 g for 5 min. The supernatants were then read at excitation/emission of 485/515 nm. Values obtained were normalized with the weight of the tissues. **P<0.001 versus non-ischemic. (n = 5).</p

Mapping lipid peroxidation in ischemic vascular bed.

<p>(A) Representative image showed the way section has been incised out from the vascular bed of chick embryo to map the lipid peroxidation status in the ischemic vascular bed. (B) Lipid peroxidation was measured by following the TBA protocol. After ligation of right vitelline artery, embryos were incubated for 2 h. An equal six sections of the chick embryo vascular bed was taken and the lipid peroxidation was measured. Data were normalized by taking the weight of the tissues.*P<0.05 and **P<0.001 versus C,D,E and F. (n = 5).</p

Sinapic acid restores nitric oxide metabolites level and ACE activity in experimental hypertensive rats.

<p>(A) Estimation of nitric oxide metabolites level in various experimental groups. (B) Assessment of ACE activity in various experimental groups. Values are expressed as mean ± SD. n = 6 per group. ^*<i>P</i><0.05 vs control; ^#<i>P</i><0.05 vs L-NAME.</p

Sinapic acid prevented oxidative stress.

<p>U^*  =  enzyme concentration required to inhibit the chromogen produced by 50% in one minute under standard condition. U^#  =  μM of H₂O₂ consumed/minute. U^$  =  μg of GSH utilized/minute. Values are expressed as mean ± SD.</p><p>^*<i>P</i><0.05 vs control;</p>#<p><i>P</i><0.05 vs L-NAME; n  =  6 per group.</p><p>Sinapic acid prevented oxidative stress.</p

Studying the angiogenesis pattern in vessels adjacent to ischemic area.

<p>(A) Angiogenesis pattern of the vessels adjacent to the ischemic area were followed for 3 h. The images of the vessels adjacent to the ischemic area were taken with Nikon Cool Pix camera adapted to a stereo microscope. Images were converted to gray scale and presented. Inset showed the magnified images from ischemia group which revealed formation of new vessels in the area. Yellow arrows showed the formation of new vessels in the area. Images are the representative of 5 individual experiments. (B) Analysis of the images were carried out using Angioquant software. Number of separate vessel complexes was analyzed by the software for different time point. Data presented as fold increase with time. **P<0.001 versus respective time point under non-ischemia. (C) Similarly, the software calculated the length of the vessels in different time of incubation. Data presented as fold increase with time. *P<0.05 and **P<0.001 versus respective time point under non-ischemia. (D) The size of the vessels was calculated by the Angioquant software. Data presented as fold increase with time. **P<0.001 versus respective time point under non-ischemia. (E) The number of junctions in the vessels were calculated by the software and plotted. *P<0.05 and **P<0.001 versus respective time point under non-ischemia.</p