Advanced glycation end-products (AGEs) play a pivotal role in macro- and micro-vascular diabetic complications. We investigated the mechanism by which methylglyoxal (an endogenous generator of AGEs) affects vascular contractility using the isolated artery technique. Contractile responses to vasoconstrictors phenylephrine (PE), angiotensin II (Ang II), vasopressin (VP) and KCl were measured in the isolated rat aorta following one-our exposure to methylglyoxal (50-200 μM). The perfused rat kidney was employed to confirm the effect of methylglyoxal on microvessels. Methylglyoxal-induced changes in cytosolic calcium were measured in the smooth muscle layer of the aorta with the calcium-sensing fluorophore Fluo-4 AM. Methylglyoxal significantly increased maximal contraction of the rat aorta to PE, Ang II and VP. Similar results were seen in response to the depolarizing vasoconstrictor KCl in macro and micro vessels. The methylglyoxal-induced increases in aortic contraction mediated by the agonist and KCl were endothelium independent. Methylglyoxal-induced increases in KCl-dependent aortic contraction were abolished after the removal of extracellular calcium or in the presence of the calcium channel blocker nifedipine. Incubation with the antioxidant N-acetyl-l-cysteine (NAC), apocynin (a nonselective NADPH oxidase (NOX) inhibitor) or chelerythrine (a protein kinase C (PKC) inhibitor) prior to methylglyoxal pre-treatment reversed the methylglyoxal-induced increases in the rat aortic contractility. In conclusion, the formation of AGEs increases vasoconstriction of both macro- and micro-vessels by increasing the voltage-activated calcium entry in vascular smooth muscles in a NOX and PKC dependent manner.

Exaggerated vasoconstriction plays important roles in vascular complication in aging and many diseases like diabetes. Here, we investigated the protective effect of Psiadia punctulata (PP) on advanced glycation end products (AGEs)-induced aggravated vasoconstriction. The effect of total methanol extract of PP leaves (PPT) on AGE-induced vascular injury was studied through bioassay-guided fractionation procedures in order to find the bioactive fraction and isolate the bioactive compounds. Vascular reactivity was studied using the isolated artery technique by adding cumulative concentrations of phenylephrine (PE) or acetyl choline (ACh). In addition, the antiglycating effect, as well as the effect on AGEs intermediates dityrosine and N`-formylkynurenine and their radical scavenging activity were measured. The results showed that PPT alleviated the AGEs-induced aggravated vasoconstriction in a concentration-dependent manner. The bioassay guided fractionation procedures suggested the chloroform fraction (Fr I) to be responsible for the activity. Chemical investigation of this fraction resulted in isolation of four major bioactive compounds that were identified as: umuhengerin (1), gardenin (2), luteolin-3`,4`-dimethyl ether (3), and 5,3`-dihydroxy-6,7,4`,5`-tetramethoxyflavone (4). The four compounds alleviated the exaggerated vasoconstriction in a dose dependent manner. In search for their mechanism of action, we observed that PPT, Fr. I and the isolated compounds did not improve the impaired vasodilation associated with AGEs exposure. PPT, Fr. I and the isolated compounds 1-4 inhibited AGEs formation and their protein oxidation intermediates. Furthermore, PPT, Fr. I and the isolated compounds 1-4 showed weak radical scavenging activity with compound 4 as the most potent. In conclusion, PPT appears to protect against AGEs-induced exaggerated vasoconstriction through antiglycation and radical scavenging activities.

The aim of the present study is to investigate the effect and potential mechanism of action of 6-gingerol on alterations of vascular reactivity in the isolated aorta from diabetic rats. Male Wistar rats were divided into two experimental groups, control and diabetics. Diabetes was induced by a single intraperitoneal injection of streptozotocin (50 mg kg(-1)), and the rats were left for 10 weeks to develop vascular complications. The effect of in vitro incubation with 6-gingerol (0.3-3 μM) on the vasoconstrictor response of the isolated diabetic aortae to phenylephrine and the vasodilator response to acetylcholine was examined. Effect of 6-gingerol was also examined on aortae incubated with methylglyoxal as an advanced glycation end product (AGE). To investigate the mechanism of action of 6-gingerol, the nitric oxide synthase inhibitor Nω-nitro-l-arginine methyl ester hydrochloride (100 μM), guanylate cyclase inhibitor methylene blue (5 μM), calcium-activated potassium channel blocker tetraethylammonium chloride (10 mM), and cyclooxygenase inhibitor indomethacin (5 μM) were added 30 minutes before assessing the direct vasorelaxant effect of 6-gingerol. Moreover, in vitro effects of 6-gingerol on NO release and the effect of 6-gingerol on AGE production were examined. Results showed that incubation of aortae with 6-gingerol (0.3-10 μM) alleviated the exaggerated vasoconstriction of diabetic aortae to phenylephrine in a concentration-dependent manner with no significant effect on the impaired relaxatory response to acetylcholine. Similar results were seen in the aortae exposed to methylglyoxal. In addition, 6-gingerol induced a direct vasodilation effect that was significantly inhibited by Nω-nitro-l-arginine methyl ester hydrochloride and methylene blue. Furthermore, 6-gingerol stimulated aortic NO generation but had no effect on AGE formation. In conclusion, 6-gingerol ameliorates enhanced vascular contraction in diabetic aortae, which may be partially attributed to its ability to increase the production of NO and stimulation of cyclic guanosine monophosphate.

Exaggerated vasoconstriction plays a very important role in the hypertension, a major component of metabolic syndrome (MetS). In the current work, the potential protective effect of methanol extract of fruit hulls of Garcinia mangostana L. on the exaggerated vasoconstriction in MetS has been investigated. In addition, the bioactive fraction and compounds as well as the possible mechanism of action have been illustrated.

Vascular dysfunction predisposes to cardiovascular complications of metabolic syndrome (MetS). The current study investigated the mechanism(s) of curcumin's (CUR) protective effect against vascular reactivity irregularities in MetS. MetS was induced by feeding rats on high fructose high salt diet. Tension studies were undertaken in aortic rings to assess the influence of CUR on vasoconstrictor or vasorelaxant responses. The effect on advanced glycation endproducts (AGEs) was studied by incubating aortic tissues with methylglyoxal, the AGEs precursor, in the absence and presence of CUR. In addition, CUR effects on in-vitro generation of AGEs and diphenyl-2-picrylhydrazyl (DPPH) free radicals were studied. The incubation with CUR for 1 hr produced significant and concentration-dependent alleviation of the exaggerated vasoconstriction observed in aortas isolated from MetS, however failed to improve the concomitant attenuation of vasodilatory responses to ACh in PE-precontracted aortas. By contrast, CUR caused direct concentration-dependent vasodilations of precontracted aortas, effects that were blunted after nitric oxide synthase inhibition by L-NAME. Similar to its effects in MetS aortas, CUR alleviated exaggerated PE vasoconstriction but did not affect impaired ACh vasodilations in AGEs-exposed aortas. In addition, CUR showed significant dose-dependent DPPH free radicals scavenging activity and inhibited both MG and fructose induced AGEs formation at the level of protein oxidation step as evident from the effect on dityrosine and N-formylkyramine. CUR alleviates exaggerated vasoconstriction in MetS through interfering with AGEs formation and AGEs-induced vascular injury. Free radical scavenging and direct vasodilatory activities could also participate in the advantageous vascular actions of CUR.

Multiple risk factors combine to increase the risk of vascular dysfunction in patients suffering from metabolic syndrome (MetS). The current study investigates the extent to which quercetin (Q) and chrysin (CH) protect against vascular dysfunction in MetS rats. MetS was induced by feeding rats a high-salt diet (3%) and fructose-enriched water (10%) for 12 weeks. Thoracic aorta was isolated from MetS rats and from control rats, with the latter being injured by methylglyoxal (MG). Aortae were incubated with CH and Q, and vascular reactivity was evaluated through the analysis of aortic contraction and relaxation in response to PE and ACh, respectively. The formation of advanced glycation end products (AGEs) and the free radical scavenging activity of 1,1-diphenyl-2-picrylhydrazyl (DPPH) were also evaluated following the introduction of CH and Q. The increased vasoconstriction and impaired vasodilation in MetS aortae were significantly ameliorated by Q and CH. Similarly, they ameliorated glycation-associated exaggerated vasoconstriction and impaired vasodilation produced by MG in control aortae. In addition, both Q and CH were effective in reducing the formation of AGEs and inhibition of glycosylation in response to MG or fructose treatment. Finally, Q successfully scavenged DPPH free radicals while CH showed significant vasodilation of precontracted aorta that was inhibited by L-NAME. In conclusion, Q and CH provide protection against vascular dysfunction in MetS by interfering with AGEs formations and AGEs-associated vascular deterioration, with CH being largely dependent on NO-mediated mechanisms of vasodilation.

The aim of the present study was to investigate the effect and possible mechanism of action of zingerone, the main constituent of ginger, on vascular reactivity in isolated aorta from diabetic rats. The results show that incubation of aortae with zingerone alleviates the exaggerated vasoconstriction of diabetic aortae to phenylephrine, as well as the impaired relaxatory response to acetylcholine in a concentration-dependent manner. Furthermore, Zingerone directly relax phenylephrine-precontracted aortae. The vasorelaxatory response is significantly attenuated by the nitric oxide synthase inhibitor Nω-nitro-l-arginine methyl ester hydrochloride and the guanylate cyclase inhibitor methylene blue but no effect of either the potassium channels blocker tetraethylammonium chloride, or the cyclooxygenase inhibitor indomethacin was observed. Zingerone had no effect on advanced glycation end product formation as well. In conclusion, zingerone ameliorates enhanced vascular contraction in diabetic aortae which may be mediated by its vasodilator effect through NO- and guanylate cyclase stimulation.

Quercetin is a naturally occurring flavonol present in many foods. Doxorubicin is an effective anticancer agent despite its dose-limiting cardiovascular toxicity. Herein, we investigated the potential protective effects of quercetin against doxorubicin-induced vascular toxicity and its effect on the therapeutic cytotoxic profile of doxorubicin in breast cancer cell lines. The incubation of isolated aortic rings with doxorubicin produced concentration-dependent exaggeration of vasoconstriction responses to phenylephrine but impaired vasodilation responses to acetylcholine. Coincubation with quercetin completely blocked the exaggerated vasoconstriction responses and the impaired vasodilation. In addition, doxorubicin incubation increased reactive oxygen species generation from the isolated aorta, while coincubation with quercetin inhibited ROS generation back to normal values. On the other hand, quercetin in combination with doxorubicin, doubled the IC50 of doxorubicin alone in MCF-7 cells from 0.4 ± 0.03 to 0.8 ± 0.06 μM. To a lesser extent, the IC50 of doxorubicin did not change after combination with quercetin in MDA-MB-231 cells. These findings indicate a significant antagonistic interaction between quercetin and doxorubicin in the aforementioned cell lines. Only in T47D cells, quercetin combination with doxorubicin was an additive interaction (CI - value = 1.17). Yet, quercetin significantly impaired the immediate phase of intracellular ROS generation by doxorubicin within breast cancer cells from 125.2 ± 3.6% to 102.5 ± 3.9% of control cells. Using annexin-V/FITC staining technique, the quercetin/doxorubicin combination showed a significantly lower percent of apoptotic cells compared to doxorubicin alone treated cells. Cell cycle distribution in breast cancer cells was performed using DNA content flowcytometry after propidium iodide staining. Quercetin induced significant accumulation of cells in the S phase as well as in the G2/M phase within both MCF-7 and MDA-MB-231 cell lines and interfered with doxorubicin-induced cell cycle effects. Interestingly, quercetin was found to inhibit the P-glycoprotein ATPase subunit with a consequent enhanced intracellular concentration of doxorubicin in MDA-MB-231 and T47D cells. In conclusion, quercetin, despite its potent vascular protective activity against doxorubicin, was found to influence doxorubicin-induced antibreast cancer effects via pharmacodynamic as well as cellular pharmacokinetic aspects.

Hydroxyphenylalkanes and diarylheptanoids possess potential therapeutic value in different pathophysiological conditions, such as malignancy. In the current study, naturally isolated hydroxyphenylalkane and diarylheptanoid compounds were investigated for potential chemo-modulatory effects in addition to potential vascular protective roles with doxorubicin. Diarylheptanoids showed stronger antioxidant effects, in comparison to hydroxyphenylalkanes, as demonstrated by DPPH assay and amelioration of CCl₄-induced disturbed intracellular GSH/GSSG balance. Shogaol and 4'-methoxygingerol showed considerable cytotoxic effects against HCT116, HeLa, HepG2 and MCF7 cells, with IC50 values ranging from 3.1 to 19.4 µM. Gingerol significantly enhanced the cytotoxic profile of doxorubicin against HepG₂ and Huh7, cells decreasing its IC50s by 10- and 4-fold, respectively. Cell cycle distribution was studied using DNA cytometry. Doxorubicin alone induced cell accumulation at S-phase and G₂/M-phase, while in combination with gingerol it significantly induced cell cycle arrest at the G₂/M-phase. Additionally, the vascular protective effect of gingerol against doxorubicin (10 µM) was examined on isolated aortic rings. Co-incubation with 6-gingerol (30 µM) completely blocked the exaggerated vasoconstriction and impaired vascular relaxation induced by doxorubicin. In conclusion, despite its relatively weak antioxidant properties, gingerol protected from DOX-induced vascular damage, apparently not through a ROS scavenging mechanism. Besides, gingerol synergized the cytotoxic effects of DOX against liver cancer cells without influencing the cellular pharmacokinetics.