Research paper
The inhibition of inducible nitric oxide synthase and oxidative stress by agmatine attenuates vascular dysfunction in rat acute endotoxemic model

https://doi.org/10.1016/j.etap.2017.08.009Get rights and content

Highlights

Abstract

Vascular dysfunction leading to hypotension is a major complication in patients with septic shock. Inducible nitric oxide synthase (iNOS) together with oxidative stress play an important role in development of vascular dysfunction in sepsis. Searching for an endogenous, safe and yet effective remedy was the chief goal for this study. The current study investigated the effect of agmatine (AGM), an endogenous metabolite of l-arginine, on sepsis-induced vascular dysfunction induced by lipopolysaccharides (LPS) in rats. AGM pretreatment (10 mg/kg, i.v.) 1 h before LPS (5 mg/kg, i.v.) prevented the LPS-induced mortality and elevations in serum creatine kinase-MB isoenzyme (CK-MB) activity, lactate dehydrogenase (LDH) activity, C-reactive protein (CRP) level and total nitrite/nitrate (NOx) level after 24 h from LPS injection. The elevation in aortic lipid peroxidation illustrated by increased malondialdehyde (MDA) content and the decrease in aortic glutathione (GSH) and superoxide dismutase (SOD) were also ameliorated by AGM. Additionally, AGM prevented LPS-induced elevation in mRNA expression of iNOS, while endothelial NOS (eNOS) mRNA was not affected. Furthermore AGM prevented the impaired aortic contraction to KCl and phenylephrine (PE) and endothelium-dependent relaxation to acetylcholine (ACh) without affecting endothelium-independent relaxation to sodium nitroprusside (SNP). In conclusion: AGM may represent a potential endogenous therapeutic candidate for sepsis-induced vascular dysfunction through its inhibiting effect on iNOS expression and oxidative stress.

Introduction

Sepsis is a serious life threatening manifestation of severe systemic infection. It imposes a medico-economic burden on healthcare systems. The mortality rates associated with sepsis exceeds those of HIV/AIDS, breast cancer and prostate cancer combined (Chaudhry and Duggal, 2014).

Sepsis is manifested by multi-organ dysfunction, most commonly affecting respiratory and cardiovascular systems. This multi-organ dysfunction provoked by sepsis can cause hypovolemia, a decrease in vascular tone and myocardial depression (De Backer et al., 2014).

The increased release of nitric oxide (NO) is observable in sepsis-induced cardiovascular dysfunction and it is believed to be due to the increase in inducible nitric oxide synthase (iNOS) mRNA expression as a part of the body’s self defensive mechanism against endotoxemia. The iNOS expression increases throughout the body in tissues such as endothelial cells, platelets, macrophages and cardiac myocytes (Chuaiphichai et al., 2016, Titheradge, 1999).

Lipid peroxidation and the decrease in superoxide dismutase (SOD) caused by oxidative stress can also account for the deleterious cytotoxic effects of sepsis (Li et al., 2016). The role of mitochondrial dysfunction in sepsis relies not only on reactive oxygen species (ROS) but also on reactive nitrogen species (RNS) that are formed due to excessive NO production (Galley, 2011).

The systemic inflammatory response associated with sepsis can be triggered by infection with bacteria such as Escherichia coli (E.Coli) (Opal et al., 2003) or the bacterial prime endotoxin lipopolysaccharide (LPS) (Ramachandran, 2014). LPS is the main constituent of gram-negative bacterial cell wall. Injecting the experimental animals with LPS is widely used as a simple and reproducible model for sepsis (Poli-de-Figueiredo et al., 2008).

Targeting the excessive release of NO and oxidative stress with a natural endogenous compound was the burgeoning goal behind this study.

Agmatine (AGM), a polycationic amine, is the endogenous metabolite of L‐arginine, formed by the action of arginine decarboxylase enzyme (Tabor and Tabor, 1984). The versatile pharmacological targets of AGM (imidazoline receptors, NO release and α2-receptors among many other targets) make it a reliable compound for treating wide spectrum of disorders such as diabetes mellitus, insulin resistance, endothelial dysfunction, neurodegenerative disorders, addiction, neuropathic pain and many inflammatory disorders (Chang et al., 2010, El-Agamy et al., 2014, El-Awady and Suddek, 2014, Gilad et al., 1996, Keynan et al., 2010, Piletz et al., 2013, Sharawy et al., 2016). In addition, AGM has a potent antioxidant effect shown in its ability to decrease lipid peroxidation and increase SOD activity and reduced glutathione (GSH) content (El-Agamy et al., 2014, Freitas et al., 2014). AGM consumption for long term is believed to be safe (Gilad and Gilad, 2013, Gilad and Gilad, 2014, Keynan et al., 2010).

Since AGM has a potential inhibitory effect on iNOS and oxidative stress, therefore this study investigates its protective effect in LPS-induced vascular dysfunction in rats.

Section snippets

Materials

AGM sulphate, pentobarbital sodium and LPS (E. coli O55:B5) were purchased from Sigma Chemical Co. (Saint Louis, Mo, USA). AGM (1-Amino-4-guanidinobutane sulfate salt) was available as white to off-white powder with purity ≥97%, it is soluble in water (50 mg/ml). AGM was dissolved in saline directly before use. LPS was available as white powder, purified by phenol extraction, and is soluble in water (5 mg/ml). All chemicals and reagents used in this study were procured from approved chemical

Effect of AGM on mortality rate

The LPS injection caused high mortality rate of 50% compared to the control group. The mortality rate dropped to only 5% by injecting the rats with AGM 1 h prior to LPS injection, this change was significantly different compared to the LPS group (Fig. 1). The mortality rate in control and AGM groups was 0%.

Effect of AGM on serum CK-MB, LDH and CRP

The injection of LPS significantly increased the activity of CK‐MB and LDH to 247% and 371% respectively compared to the control group. Preinjection with AGM significantly decreased this

Discussion

In the present study, LPS model in rats was used to demonstrate the protective action of AGM against vascular abnormalities induced by endotoxemia. AGM was selected as an endogenous, natural and safe compound beside its effect as a modulator for NO haemostasis and as an antioxidant in the body.

In this study, injecting LPS to rats caused a profound damage to the myocardial muscle as shown by the elevation of CK-MB and CRP levels, and LDH activity. The CK-MB isozyme is distributed primarily in

Conclusion

The endogenous compound AGM has a promising therapeutic potential against LPS induced vascular endothelial dysfunction and can be helpful in case of systemic shock by its inhibitory effects on iNOS expression and oxidative stress.

References (35)

  • M.A. Titheradge

    Nitric oxide in septic shock

    Biochim. Biophy. Acta (BBA)—Bioenergetics

    (1999)
  • O. Bayraktar et al.

    Effects of S-allyl cysteine on lung and liver tissue in a rat model of lipopolysaccharide-induced sepsis

    Naunyn-Schmiedeberg’s Arch. Pharmacol.

    (2015)
  • D. Bishop-Bailey et al.

    Characterization of the induction of nitric oxide synthase and cyclo-oxygenase in rat aorta in organ culture

    Br. J. Pharmacol.

    (1997)
  • J. Boisrame-Helms et al.

    Endothelial dysfunction in sepsis

    Curr. Vasc. Pharmacol.

    (2013)
  • V. Borutaite et al.

    Nitric oxide from inducible nitric oxide synthase sensitizes the inflamed aorta to hypoxic damage via respiratory inhibition

    Shock (Augusta, Ga.)

    (2005)
  • N. Chaudhry et al.

    Sepsis associated encephalopathy

    Adv. Med.

    (2014)
  • S. Cuzzocrea et al.

    A role for nitric oxide-mediated peroxynitrite formation in a model of endotoxin-induced shock

    J. Pharmacol. Exp. Ther.

    (2006)
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