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The use of sonication treatment to decellularize aortic tissues for preparation of bioscaffolds

Azhim A, Syazwani N, Morimoto Y, Furukawa KS, Ushida T

J Biomater Appl, 2014 Jul;29(1):130-41.
PMID: 24384523 DOI: 10.1177/0885328213517579

A novel decellularization method using sonication treatment is described. Sonication treatment is the combination of physical and chemical agents. These methods will disrupt cell membrane and release cell contents to external environments. The cell removal was facilitated by subsequent rinsing of sodium dodecyl sulfate detergents. Sonication treatment is used in the preparation of complete decellularized bioscaffolds. The aim of this study is to confirm the usefulness of sonication treatment for preparation of biological scaffolds. In this study, samples of aortic tissues are decellularized by sonication treatment at frequency of 170 kHz in 0.1% and 2% sodium dodecyl sulfate detergents for 10-h treatment time. The relation between decellularization and sonication parameters such as dissolved oxygen concentration, conductivity, and pH is investigated. Histological analysis and biomechanical testing is performed to evaluate cell removal efficiency as well as changes in biomechanical properties. Minimal inflammation response elicit by bioscaffolds is confirmed by xenogeneic implantation and immunohistochemistry. Sonication treatment is able to produce complete decellularized tissue suggesting that these treatments could be applied widely as one of the decellularization method.

Matched MeSH terms: Aorta, Thoracic/metabolism
Fulltext The role of repeatedly heated soybean oil in the development of hypertension in rats: association with vascular inflammation

Ng CY, Kamisah Y, Faizah O, Jaarin K

Int J Exp Pathol, 2012 Oct;93(5):377-87.
PMID: 22974219 DOI: 10.1111/j.1365-2613.2012.00839.x

Thermally oxidized oil generates reactive oxygen species that have been implicated in several pathological processes including hypertension. This study was to ascertain the role of inflammation in the blood pressure raising effect of heated soybean oil in rats. Male Sprague-Dawley rats were divided into four groups and were fed with the following diets, respectively, for 6 months: basal diet (control); fresh soybean oil (FSO); five-time-heated soybean oil (5HSO); or 10-time-heated soybean oil (10HSO). Blood pressure was measured at baseline and monthly using tail-cuff method. Plasma prostacyclin (PGI(2) ) and thromboxane A(2) (TXA(2) ) were measured prior to treatment and at the end of the study. After six months, the rats were sacrificed, and the aortic arches were dissected for morphometric and immunohistochemical analyses. Blood pressure was increased significantly in the 5HSO and 10HSO groups. The blood pressure was maintained throughout the study in rats fed FSO. The aortae in the 5HSO and 10HSO groups showed significantly increased aortic wall thickness, area and circumferential wall tension. 5HSO and 10HSO diets significantly increased plasma TXA(2) /PGI(2) ratio. Endothelial VCAM-1 and ICAM-1 were significantly increased in 5HSO, as well as LOX-1 in 10HSO groups. In conclusion, prolonged consumption of repeatedly heated soybean oil causes blood pressure elevation, which may be attributed to inflammation.

Matched MeSH terms: Aorta, Thoracic/metabolism
Vasorelaxant activities and the underlying pharmacological mechanisms of Gynura procumbens Merr. leaf extracts on rat thoracic aorta

Iqbal Z, Bello I, Asmawi MZ, Al-Mansoub MA, Ahmad A, Jabeen Q, et al.

Inflammopharmacology, 2019 Apr;27(2):421-431.
PMID: 29185178 DOI: 10.1007/s10787-017-0422-4

Previous studies have investigated the cardiovascular activity of Gynura procumbens Merr. single-solvent extracts. The objective of this study was to evaluate the in vitro vasorelaxant properties and the underlying pharmacological mechanisms of serial extracts and fractions of Gynura procumbens (GP). The leaves of GP were serially extracted with petroleum ether, chloroform, methanol and water using the maceration method. Suspended aortic ring preparations were pre-contracted with phenylephrine (PE 1 µM), followed by cumulative addition of GP extracts (0.25-3 mg/mL). The petroleum ether extract (GPPE) was the most potent among the four extracts. Pre-incubation of endothelium-intact aorta with atropine (1 µM), indomethacin (10 µM), methylene blue (10 µM), propranolol (1 µM) and potassium channel blockers such as TEA (1 µM), glibenclamide (10 µM), 4-aminopyridine (1 µM) and barium chloride (10 mM) had no effect on GPPE-induced vasorelaxation. The vasorelaxant effect of GPPE was partly diminished by pretreatment of aortic rings preparations with L-NAME (10 µM) and even more so in endothelium-denuded aortic rings, indicating a minimal involvement of endothelium-dependent pathway in GPPE-induced vasorelaxation. The calcium-induced vasocontractions were antagonized significantly and concentration-dependently by GPPE in calcium free and high potassium medium. These results illustrate that Ca2+ antagonizing actions of GPPE in rat isolated aorta are comparable to that of verapamil and may be mainly responsible for its vasodilation effect. The antioxidant activity of GPPE supports its vasorelaxant effect by attenuating the production of deleterious free radicals and reactive oxygen species in the vasculature.

Matched MeSH terms: Aorta, Thoracic/metabolism
Des-aspartate angiotensin I (DAA-I) reduces endothelial dysfunction in the aorta of the spontaneously hypertensive rat through inhibition of angiotensin II-induced oxidative stress

Loh WM, Ling WC, Murugan DD, Lau YS, Achike FI, Vanhoutte PM, et al.

Vascul Pharmacol, 2015 Aug;71:151-8.
PMID: 25869508 DOI: 10.1016/j.vph.2015.03.011

Des-aspartate angiotensin I (DAA-I), an endogenous nonapeptide, counteracts several effects of angiotensin II on vascular tone. The aim of this study was to investigate the acute protective effect of DAA-I on endothelial function in the spontaneously hypertensive rat (SHR) as well as its effect on angiotensin II-induced contractions and oxidative stress. Aortic rings were incubated with DAA-I (0.1μM) for 30min prior to the assessment of angiotensin II-induced contractions (0.1nM-10μM) in WKY and SHR aortas. Total nitrate and nitrite levels were assessed using a colorimetric method and reactive oxygen species (ROS) were measured by dihydroethidium (DHE) fluorescence and lucigenin-enhanced chemiluminescence. The effect of DAA-I was also assessed against endothelium-dependent and -independent relaxations to acetylcholine and sodium nitroprusside, respectively. Angiotensin II-induced contractions were significantly reduced by DAA-I, losartan and tempol. Incubation with ODQ (soluble guanylyl cyclase inhibitor) and removal of the endothelium prevented the reduction of angiotensin II-induced contractions by DAA-I. Total nitrate and nitrite levels were increased in DAA-I, losartan and tempol treated-SHR tissues while ROS level was reduced by DAA-I and the latter inhibitors. In addition, DAA-I significantly improved the impaired acetylcholine-induced relaxation in SHR aortas whilst sodium nitroprusside-induced endothelium-independent relaxation remained unaffected. The present findings indicate that improvement of endothelial function by DAA-I in the SHR aorta is mediated through endothelium-dependent release of nitric oxide and inhibition of angiotensin II-induced oxidative stress.

Matched MeSH terms: Aorta, Thoracic/metabolism
Mechanisms underlining gender differences in Phenylephrine contraction of normoglycaemic and short-term Streptozotocin-induced diabetic WKY rat aorta

Aloysius UI, Achike FI, Mustafa MR

Vascul Pharmacol, 2012 Sep-Oct;57(2-4):81-90.
PMID: 22172524 DOI: 10.1016/j.vph.2011.11.009

The female gender reduces the risk, but succumbs more to cardiovascular disease. The hypothesis that short-term (8weeks) Streptozotocin-induced diabetes could produce greater female than male vascular tissue reactivity and the mechanistic basis were explored. Aortic ring responses to Phenylephrine were examined in age- and sex-matched normoglycaemic/diabetic rats. The normoglycaemic male tissue contracted significantly more than the normoglycaemic female and the male/female diabetic tissues. Endothelial-denudation, l-NAME or MB reversed these differences suggesting an EDNO-cGMP dependence. 17β-oestradiol exerted relaxant effect on all endothelium-denuded (and normoglycaemic endothelium-intact male) tissues, but not endothelium-intact normoglycaemic female. The greater male tissue contraction is attributable to absent 17β-oestradiol-modulated relaxation. Indomethacin blockade of COX attenuated male normoglycaemic and female diabetic tissue contraction (both reversed by l-NAME), but augmented diabetic male tissue contraction. These data are consistent with the raised contractile TXA(2) and PGE(2) in normoglycaemic male and diabetic female tissues, and the relaxant PGI(2) in diabetic male (and female). The higher levels of PGI(2) in the normoglycaemic and diabetic female perhaps explain their greater relaxant response to Acetylcholine compared to the respective male. In conclusion, there is an endothelium-dependent gender difference in the effect of short term diabetes on vascular tissue reactivity which is COX mediated.

Matched MeSH terms: Aorta, Thoracic/metabolism
Vasorelaxant Action of the Chloroform Fraction of Orthosiphon stamineus via NO/cGMP Pathway, Potassium and Calcium Channels

Yam MF, Tan CS, Ahmad M, Ruan S

Am J Chin Med, 2016;44(7):1413-1439.
PMID: 27785939

Orthosiphon stamineus Benth. (Lamiaceae) is an important plant in traditional folk medicine that is used to treat hypertension and kidney stones. In humans, this plant has been tested as an addition regiment for antihypertensive treatment. Among the treatments for hypertension, O. stamineus had been to have diuretic and vasorelaxant effects in animal models. There is still very little information regarding the vasorelaxant effect of O. stamineus. Therefore, the present study was designed to investigate the vasorelaxant activity and mechanism of action of the fractions of O. stamineus. The vasorelaxant activity and the underlying mechanisms of the chloroform fraction of the 50% methanolic extract of O. stamineus (CF) was evaluated on thoracic aortic rings isolated from Sprague Dawley rats. CF caused relaxation of the aortic ring pre-contracted with phenylephrine in the presence and absence of endothelium, and pre-contracted with potassium chloride in endothelium-intact aortic ring. In the presence of endothelium, both indomethacin (a nonselective cyclooxygenase inhibitor) and [Formula: see text]-[1,2,4]Oxadiazolo[4,3-[Formula: see text]]quinoxalin-1-one (ODQ, selective soluble guanylate cyclase inhibitor) had a small effect on the vasorelaxation response. On the other hand, in the presence of Nω-nitro-L-arginine methyl ester (L-NAME, nitric oxide synthase inhibitor), methylene blue (cyclic guanosine monophosphate lowering agent), tetraethylammonium ([Formula: see text], nonselective calcium activator [Formula: see text] channel blocker), 4-aminopyridine (4-AP, voltage-dependent [Formula: see text] channel blocker), barium chloride ([Formula: see text], inwardly rectifying [Formula: see text] channel blocker), glibenclamide (nonspecific ATP-sensitive [Formula: see text] channel blocker), atropine (muscarinic receptor blocker) and propranolol (β-adrenergic receptor blocker), the vasorelaxant effect significantly reduced the relaxation stimulated by CF. CF was also found to be active in reducing [Formula: see text] release from the sarcoplasmic reticulum and blocking calcium channels.

Matched MeSH terms: Aorta, Thoracic/metabolism
Chronic administration of sodium nitrite prevents hypertension and protects arterial endothelial function by reducing oxidative stress in angiotensin II-infused mice

Ling WC, Mustafa MR, Vanhoutte PM, Murugan DD

Vascul Pharmacol, 2018 03;102:11-20.
PMID: 28552746 DOI: 10.1016/j.vph.2017.05.003

AIM: Endothelial dysfunction accompanied by an increase in oxidative stress is a key event leading to hypertension. As dietary nitrite has been reported to exert antihypertensive effect, the present study investigated whether chronic oral administration of sodium nitrite improves vascular function in conduit and resistance arteries of hypertensive animals with elevated oxidative stress.
METHODS: Sodium nitrite (50mg/L) was given to angiotensin II-infused hypertensive C57BL/6J (eight to ten weeks old) mice for two weeks in the drinking water. Arterial systolic blood pressure was measured using the tail-cuff method. Vascular responsiveness of isolated aortae and renal arteries was studied in wire myographs. The level of nitrite in the plasma and the cyclic guanosine monophosphate (cGMP) content in the arterial wall were determined using commercially available kits. The production of reactive oxygen species (ROS) and the presence of proteins (nitrotyrosine, NOx-2 and NOx-4) involved in ROS generation were evaluated with dihydroethidium (DHE) fluorescence and by Western blotting, respectively.
RESULTS: Chronic administration of sodium nitrite for two weeks to mice with angiotensin II-induced hypertension decreased systolic arterial blood pressure, reversed endothelial dysfunction, increased plasma nitrite level as well as vascular cGMP content. In addition, sodium nitrite treatment also decreased the elevated nitrotyrosine and NOx-4 protein level in angiotensin II-infused hypertensive mice.
CONCLUSIONS: The present study demonstrates that chronic treatment of hypertensive mice with sodium nitrite improves impaired endothelium function in conduit and resistance vessels in addition to its antihypertensive effect, partly through inhibition of ROS production.

Matched MeSH terms: Aorta, Thoracic/metabolism
Modulation of vascular reactivity in normal, hypertensive and diabetic rat aortae by a non-antioxidant flavonoid

Ajay M, Achike FI, Mustafa MR

Pharmacol Res, 2007 May;55(5):385-91.
PMID: 17317209

In this study, we report the effects of a non-antioxidant flavonoid flavone on vascular reactivity in Wistar-Kyoto (WKY) rat isolated aortae. Whether flavone directly modulates vascular reactivity in spontaneously hypertensive rat (SHR) and streptozotocin-induced diabetic-WKY rat isolated aortae was also determined. Thoracic aortic rings were mounted in organ chambers and exposed to various drug treatments in the presence of flavone (10 microM) or its vehicle (DMSO), which served as control. Pretreatment with flavone enhanced relaxant effects to endothelium-dependent vasodilator acetylcholine (ACh) and attenuated contractile effects to alpha(1)-receptor agonist phenylephrine (PE) in WKY aortae compared to those observed in control aortic rings. Flavone had no effect on relaxations to ACh in WKY aortae incubated with either L-NAME or methylene blue, but enhanced relaxations to ACh in WKY aortae incubated with indomethacin or partially depolarized with KCl. Relaxations to ACh are totally abolished in both control or flavone pretreated endothelium-denuded WKY aortae. Flavone attenuated the inhibition by beta-NADH of ACh-induced relaxation in WKY aortae, but it had no significant effect on the transient contractions induced by beta-NADH nor the pyrogallol-induced abolishment of ACh-induced relaxation in WKY aortae. Flavone enhanced endothelium-independent relaxation to sodium nitroprusside (SNP) in both endothelium-intact and -denuded WKY aortae. Flavone enhanced relaxation to ACh and SNP as well as attenuated contractile effects to PE in SHR and diabetic aortae, a finding similar to that observed in normal WKY aortae. From these results, we conclude that flavone modulates vascular reactivity in normal as well as hypertensive and diabetic aortae. These effects of flavone results probably through enhanced bioactivity of nitric oxide released from the endothelium.

Matched MeSH terms: Aorta, Thoracic/metabolism