RU2568365C1 - Method for correction of endothelial dysfunction with combination of rosuvastatin and thioctic acid in hypooestrogen-l-name-induced nictic oxide deficiency - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: method involves simulating a dysfunction by performing a bilateral ovariectomy in Wistar male rats. On the 43rd postoperative days, L-nitro-arginine-methyl ester (L-NAME) is administered intragastrically in a dose of 25 mg/kg daily during 7 days. The induced dysfunction is corrected by daily intragastric administration of Rosuvastatin 0.85 mg/kg and thioctic acid 50 mg/kg during 7 days concurring with the administration of L-NAME into the rats.

EFFECT: method provides activating the correction of the endothelial dysfunction in hypooestrogenemia.

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Description

The invention relates to medicine, in particular to experimental cardiopharmacology, and can be used to correct endothelial dysfunction.

A known method for the correction of endothelial dysfunction using rosuvastatin related to statins, described by E.I. Krasilnikova, E.M. Nifontov, E.G. Sergeeva, T.V. Antonova, A.A. Redneck, O.V. Galkina, M.A. Romanova “Indices of endothelial adhesion dysfunction and factors of immune inflammation in patients with hypertension in combination with coronary heart disease: dynamics during treatment with Crestor.” Arterial hypertension - 2009.- T.15.- No. 3.- p.268-274, which consists in the oral administration of this drug in patients to correct endothelial dysfunction.

The main disadvantage of this method is the narrower pathogenetic orientation of the action of rosuvastatin in endothelial dysfunction, associated with its pleiotropic effects, in particular a decrease in the level of proinflammatory cytokine - interleukin-6, which is associated with the development of endothelial dysfunction, while one of the reasons is increased oxide biodegradation Nitrogen as a result of peroxidation remains uncompensated. In addition, in the model of hypoestrogen-L-NAME induced NO deficiency used, the development of endothelial dysfunction is associated not only with increased biodegradation, but also with a decrease in the bioavailability of nitric oxide as a result of increased oxidative stress due to hypoestrogenism. It has been established that women during menopause have many factors that can play an important role in stimulating oxidative stress. One of these factors is plasma renin activity (ARP) and, accordingly, the activity of the renin-angiotensin system (RAS). In a study by James et al. In which ARP was measured over the course of 9 years in men and women, ARP levels were higher in women after menopause than before menopause (James GD, Sealey JE, Muller F et al. Renin Relationship to Sex , Race and Age in Normotensive Population. J Hypertension 1986; 4 (Suppl 5): S387-S389). Angiotensin-2 administered as high (Rajagopalan S, Kerry S, Munzel T et al. Angiotensin ll-mediated hypertension in the ratincreases vascular superoxide production via membrane NADH / NADPH oxidase activation. Contribution to alterations of vasomotor tone. J Clin Invest 1996 ; 97: 1916-1923) and in physiological doses (Reckelhoff JF, Zhang H, Srivastava K et ai. Subpressor doses of angiotensin II increases-plasma F2-isoprostanes in rats. Hypertension 2000; 35: 476-479), is capable of stimulate oxidative stress, as evidenced by an increase in plasma levels of P2-isoprostanes. In addition, studies have shown that P2-isoprostane, in addition to being a renal vasoconstrictor itself (Yura T, Fukunaga M, Kahn R et al. Free-radical-generated F2-isoprostane stimulates cell-proliferation and endothelin-1 expression in endothelial cells. Kidney Int 1999; 56: 471-478), enhances the vasoconstrictor effect of AT2 (Sametz W, Grobuschek T, Hammer-Kogler S et al. Influence of Isoprostanes on Vasoconstrictor Effects of Noradrenaline and Angiotensin II. Eur J Pharm 1999; 378 : 47-55). Both substances, AT2 and endothelin, increase superoxide synthesis by stimulating subunits of NADP (H) oxidase (Sametz W, Grobuschek T, Hammer-Kogler S et al. Influence of Isoprostanes on Vasoconstrictor Effects of Noradrenaline and Angiotensin II. Eur J Pharm 1999; 378 : 47-55). Superoxide binds to nitric oxide (N0), which suppresses its biological activity (i.e., vasodilation) (Sametz W, Grobuschek T, Hammer-Kogler S et al. Influence of Isoprostanes on Vasoconstrictor Effects of Noradrenaline and Angiotensin II. Eur J Pharm 1999; 378: 47-55), therefore, a decrease in the bioavailability of NO can lead to vasoconstriction and an increase in blood pressure. In addition, an increase in the activity of oxidative stress, as evidenced by high levels of not only P2-isoprostane, but also plasma malondialdehyde (markers of oxidative stress) (Helmersson J, Mattsson P, Basu S. Prostaglandin F (2alpha) metabolite and F2-isoprostane excretion in migraine. Clin Sci (Lond) 2002; 102: 39-43), (Signorelli SS, Neri S, Sciacchitano S et al. Duration of menopause and behavior of mal-ondialdehyde, lipids, lipoproteins and carotid wall artery intima-media thickness. Maturitas 2001; 39: 39-42).

 Therefore, monotherapy of endothelial dysfunction with rosuvastatin is considered insufficient, since it does not affect the pathogenesis of endothelial dysfunction associated with activation of oxidative stress, and leads to the search for more effective methods of pharmacotherapy, one of which is the combination of rosuvastatin with an antioxidant.

Closest to the claimed solution is a method for the correction of endothelial dysfunction with L-NAME-induced nitric oxide deficiency, including modeling of endothelial dysfunction by daily intraperitoneal administration of L-nitro-arginine-methyl ether to male rats of the Wistar strain at a dose of 25 mg / kg and the simultaneous correction of dysfunction with a combination of statin and antioxidant (RU No. 2438188 C1, publ. 12/27/2011).

The main disadvantage of this method is the absence of resveratrol action that improves angioneuronal blood flow, which is one of the mechanisms of endothelioprotective action.

The technical result of the invention is a method for the correction of endothelial dysfunction, including the use of rosuvastatin, related to statins, and thioctic acid, which has antioxidant activity.

The technical result is achieved by the fact that modeling of endothelial dysfunction by daily intraperitoneal administration of L-nitro-arginine-methyl ether to female rats of the Wistar line at a dose of 25 mg / kg is carried out simultaneously with correction of dysfunction with a combination of statin and antioxidant, and in modeling endothelial dysfunctions additionally form a hypoestrogenic state within 6 weeks by performing bilateral ovariectomy, and rosuvastatin is used as a statin, which is administered nutrizheludochno once daily at a dose of 0.85 mg / kg and used as an antioxidant thioctic acid which is introduced together with rosuvastatin at a dose of 50 mg / kg by evaluating the degree of correction of endothelial dysfunction ratio endothelium and endothelium dependent vasodilation.

An additional introduction of thioctic acid leads to the activation of the correction of endothelial dysfunction, since it is an antioxidant and prevents the biodegradation of nitric oxide in the vascular endothelium by free radicals.

The method is carried out as follows.

 The experiments were carried out on female white rats weighing 200-250 g of the Wistar line at the age of 3.5 months. To model endothelial dysfunction, rats were anesthetized with chloral hydrate (300 mg / kg) and bilateral ovariectomy was performed. For this, the anterior abdominal wall was opened under aseptic conditions, the tubular and peritoneal sections of the ovaries were ligated using atraumatic suture material, after which the latter were excised. The wound was sutured in layers. On day 43 (6 weeks after the operation), an inhibitor of NO synthase N-nitro-L-arginine methyl ether (L-NAME, Sigma) was administered intraperitoneally once a day at a dose of 25 mg / kg in a volume of 1 ml / kg within 7 days (n = 10 animals). The animals of the intact group are injected with physiological NaCl in the same volume (n = 10 animals). Thioctic acid and rosuvastatin, as well as their combination, are administered daily intragastrically (via a probe) at doses of 50 mg / kg / day and 0.85 mg / kg / day, respectively, for 7 days simultaneously with L-NAME.

On day 8 under anesthesia (chloral hydrate 300 mg / kg and zolet 150 mg / kg, intraperitoneally), the animal was taken in an experiment and, at the first stage, blood pressure and blood pressure responses to endothelium-dependent (acetylcholine at a dose of 40 μg / kg) and endothelium-independent (nitroprusside at a dose of 30 μg / kg) vasodilation by introducing a catheter into the carotid artery. Pharmacological agents were administered bolus into the right femoral vein.

Systolic blood pressure (SBP), diastolic blood pressure (DBP) and heart rate (HR) were recorded using a TSD104A sensor and MP150 hardware and software complex, manufactured by Biopac System, Inc., USA.

 To assess the degree of development of endothelial dysfunction in experimental animals and its correction with the studied drugs, the coefficient of endothelial dysfunction (QED) was calculated:

$$\mathrm{TO}ED=\frac{S_{\mathrm{BUT}DNP}}{S_{\mathrm{BUT}D\text{OH}}},$$

where QED is the coefficient of endothelial dysfunction; S _{AD BP} - the area of the triangle above the blood pressure recovery curve when performing a functional test with the introduction of sodium nitroprusside, S _{AD AX} - the area of the triangle above the blood pressure recovery curve when performing a functional sample with the introduction of acetylcholine at a dose of 40 μg / kg (patent RU No. 2301015 of 20.06. 2007, bull. No. 17).

When statistical data processing is calculated, the average value, the standard deviation. Differences are considered significant at p <0.05.

EXAMPLE OF SPECIFIC PERFORMANCE

Blockade of NO synthase caused by 7-day administration of L-NAME to rats led to a disruption in the relationship between vasodilating and vasoconstrictor mechanisms for regulating vascular tone, as evidenced by the results of vascular tests for endothelium-dependent (acetylcholine) and endothelium-independent (increased sodium vasodilatase and nitroprusside) 0.8 ± 0.1 in intact animals up to 4.6 ± 0.7 (p <0.05).

In the group of animals where only rosuvastatin was administered against the background of the administration of the NO synthase inhibitor L-NAME, this ratio corresponded to a value of 1.7 ± 0.2, and in the group with additional administration of thioctic acid to rosuvastatin, 1.4 ± 0.3, which is less than in the group of animals with the introduction of L-NAME.

Blood pressure in intact animals was: systolic (SBP) - 128.1 ± 6.0 mm Hg, diastolic (DBP) - 95.7 ± 4.0 mm Hg. With the introduction of L-NAME against the background of ovariectomy, there was an increase in blood pressure to 169.9 ± 7.3 mm Hg. and 123.2 ± 7.5 mm Hg respectively.

The introduction of rosuvastatin on the background of hypoestrogenism and L-NAME reduced blood pressure to 151.2 ± 6.2 mm Hg and 113.8 ± 3.3 mmHg respectively (p <0.05), and the introduction of a combination of rosuvastatin and thioctic acid led to a greater decrease in blood pressure, which amounted to 135.5 ± 2.9 mm Hg, DBP - 97.6 ± 7.1 mm Hg. Art. respectively (p <0.05).

The dynamics of systolic (SBP), diastolic (DBP) blood pressure, and the coefficient of endothelial dysfunction in the experimental groups of animals are shown below in table 1.

Table 1

Indicator
Group GARDEN,
mmHg. DBP
mmHg. QED
rel. Intact 128.1 ± 6.0 95.7 ± 4.0 0.8 ± 0.1 Ovariectomy + L-NAME 169.9 ± 7.3 * 123.2 ± 7.5 * 4.6 ± 0.7 * Ovariectomy + L-NAME +
rosuvastatin 151.2 ± 6.2 * 113.8 ± 3.3 * 1.7 ± 0.2 ** Ovariectomy + L-NAME + Rosuvastatin + Thioctic Acid 135.5 ± 2.9 * 97.6 ± 7.1 * 1.4 ± 0.3 **

Note: QED - coefficient of endothelial dysfunction; * - p <0.05 in comparison with the intact group; ** - p <0.05 compared with the L-NAME group.

In addition, the research results are illustrated by graphic images showing the dynamics of indicators of systolic (SBP), diastolic (DBP) blood pressure, coefficient of endothelial dysfunction in experimental groups of animals.

Thus, the obtained results allow us to ascertain the activation of the pharmacological correction of endothelial dysfunction with rosuvastatin when it is combined with thioctic acid, since it is an antioxidant and prevents the biodegradation of nitric oxide in the vascular endothelium by free radicals.

Claims (1)

A method for the correction of endothelial dysfunction in case of hypoestrogen-L-NAME-induced nitric oxide deficiency, including modeling of dysfunction by bilateral ovariectomy of female Wistar rats and for 43 days after surgery, intraperitoneal injection of L-nitro-arginine methyl ester into the daily for 7 days a dose of 25 mg / kg and correction of dysfunction by daily intragastric administration of rosuvastatin at a dose of 0.85 mg / kg and thioctic acid at a dose of 50 mg / kg for 7 days, coinciding with the introduction of rats L-NAME.