The role of gender and sex hormones in ischemic–reperfusion injury in isolated rat hearts

https://doi.org/10.1016/j.ejphar.2007.01.043Get rights and content

Abstract

To establish potential anti-ischemic effects of testosterone and estradiol on myocardium we used isolated rat hearts in accordance with Langendorff, exposed to 40 min of ischemia and reperfusion. Rats were pretreated for 10 days, males with testosterone and females with estradiol and injuries from those hearts were compared to the hearts where both drugs were applied to the isolated hearts directly. The myocardial injuries were determined by changes in coronary flow, incidence and duration of arrhythmias and lactate dehydrogenase release rates used as markers for level of cardiac injury during reperfusion. Coronary flow in the hearts of animals pretreated with estradiol during reperfusion increased by 68.7 ±3.6% (P < 0.001) and in those pretreated with testosterone by 50.1 ± 2.1% (P < 0.05) vs. control hearts. Lactate dehydrogenase release rates decreased in the hearts of animals pretreated with estradiol by 55.7 ± 1.9% (P < 0.01) vs. controls and by 58.8 ± 3.0 (P < 0.01) vs. directly applied estradiol. Duration of ventricular fibrillation decreased after 10 days application of drugs, from 9.42 ± 0.81 min to 4.58 ± 0.93 min (P < 0.05) with estradiol and from 9.19 ± 1.05 min to 4.65 ± 0.51 min (P < 0.05) with testosterone. The duration of heart arrest decreased in 10 days application of testosterone from 2.42 ± 0.16 min to 20.0 ± 12.26 s (P < 0.01). Hearts from animals pretreated for 10 days with estradiol showed more cardioprotective effects during reperfusion than those pretreated with testosterone. Testosterone pretreatment, despite being less effective in cardioprotection than estradiol, improved coronary flow and decreased arrhythmias as effectively as estradiol.

Introduction

Heart disease is the primary cause of death in developed countries in men and in women over the age of 60. While pre-menopausal women have a low incidence of cardiovascular disease compared to men, mortality in post-menopausal women rises to the same frequency as in men (Adams et al., 1995, Fraser et al., 2000, Gray et al., 2001, Wild and Bartholemew, 1988). This significant gender difference has mostly been attributed to the beneficial role of estrogens (Collins et al., 1993, Gray et al., 2001).

In cardiovascular tissues the protective effects of estrogen and testosterone are manifested through their receptors in coronary arteries, where estrogen directly relaxes arteries via endothelial nitric oxide (NO) mechanism (Dai et al., 2004, Santos et al., 2004, Woodman et al., 2004). It is probable that testosterone relaxes them via a different mechanism as it is in estrogens, also a non-genomic pathway, which does not involve the nuclear androgen receptors and is independent of the vascular endothelium. This response to testosterone is initiated at the specific binding sites in smooth muscle cell membranes. Testosterone directly inhibits voltage-gated calcium channels, with an additional inhibitory action upon store-operated calcium channels (Jones et al., 2004, Yildiz et al., 2005).

Estrogens in myocardium reduce the incidence of post-ischemic ventricular arrhythmias by reducing accumulation of intracellular Ca2+, protecting mitochondrial structures, inhibiting apoptosis, having antioxidant action and interacting with heat stress proteins, thus protecting the heart from injuries (Fraser et al., 2000, Kim et al., 1998, Knowlton and Sun, 2001, Zhai et al., 2000). Both the mitogen activated protein kinase and phosphoinositide-3-kinase are also believed to be involved in the regulation of NO synthesis by estrogen (Gray et al., 2001).

The action of androgens on cardiovascular systems has received relatively little attention and the authors disagree about possible detrimental or protective effects of testosterone in the heart (Pugh et al., 2000). Androgens have been associated with possible proatherogenic effects and an increased cardiovascular risk by adversely affecting the plasma lipid and lipoprotein profile, increasing the risk of thrombosis and cardiac hypertrophy (Adams et al., 1995). On the contrary, short-term administration of testosterone causes vasodilatation in a range of species including humans (Costarella et al., 1996, Crews and Khalil, 1999, Honda et al., 1999, Perusquia et al., 1996, Yue et al., 1995).

To compare the effects of androgens to estrogens on myocardial injuries of isolated rat hearts, we studied their effects on two models. Firstly, the rats were treated with supraphysiological/subtoxic levels of sex hormones each day for 10 days prior to the experiment (males with testosterone, females with estradiol) and then their isolated hearts were subjected to acute ischemia/reperfusion injury. Secondly, isolated rat hearts were perfused with both hormones directly; male hearts with testosterone and female hearts with estradiol and then exposed to 40 min of ischemia and 40 min of reperfusion. Markers for the estimation of ischemia/reperfusion injuries were the level of decrease in the incidence and duration of arrhythmias, the increase in coronary flows and the decrease in lactate dehydrogenase release rates.

Section snippets

Animals and perfusion technique

All experiments were conducted in accordance with guidelines set down by the Veterinary Administration of the Republic of Slovenia (permission No. 323-02-203/00). Male and female Wistar rats (270–330 g and 230–260 g in body weight respectively) were obtained from the Faculty of Medicine, Ljubljana. All animals were 9–12 weeks old, bred under constant housing conditions and fed with standard rat chow in the form of pellets (Krka d.o.o., Novo mesto, Slovenia). A regular circadian cycle was

The coronary flow

The effect of direct application or 10 days pretreatment with estradiol and testosterone on coronary flow was determined in experiments with 40 min ischemia and 40 min of reperfusion.

In hearts where animals were pretreated with both drugs subcutaneously 10 days prior to isolation a rapid increase in coronary flow after the start of reperfusion was induced, and the highest values were reached in the 77th minute of the experiment. The comparable increase in coronary flow was observed after direct

Discussion

The main finding of this study is that both estradiol and testosterone in supraphysiological (subtoxic) levels significantly decreased ischemia/reperfusion injuries of isolated rat hearts in accordance with Langendorff. Reperfusion injuries were reduced both by the direct application of, and 10 days pretreatment with sex hormones prior to the isolated heart experiment. However, protective effects against ischemia/reperfusion injuries were only observed in the male hearts of animals pretreated

Acknowledgements

This work was supported by grant P0-0508-0381 from the Slovenian Ministry of Education, Science and Sport. The authors are grateful to Dani Hamze Jokhadar for assistance in conducting the experiments and evaluation of results and to Prof. Metka V. Budihna for overall help in the research.

References (34)

  • J.K. Crews et al.

    Gender-specific inhibition of Ca2+ entry mechanisms of arterial vasoconstriction by sex hormones

    Clin. Exp. Pharmacol. Physiol.

    (1999)
  • K. Dechering et al.

    Estrogen receptors alpha and beta: two receptors of a kind?

    Current Med. Chem.

    (2000)
  • W.P. Eenadayalu et al.

    Testosterone relaxes coronary arteries by opening the large conductance, calcium activated potassium channel

    Am. J. Physiol, Heart Circ. Physiol.

    (2001)
  • K.M. English et al.

    Testosterone acts as a coronary vasodilator by a calcium antagonistic action

    J. Endocrinol. Investig.

    (2002)
  • F. Er et al.

    Testosterone induces cytoprotection by activating ATP-sensitive K channels in the cardiac mitochondrial inner membrane

    Circulation

    (2004)
  • H. Fraser et al.

    Activation of Ca2+-independent nitric oxide synthase by 17beta-estradiol in post-ischemic rat heart

    Cardiovasc. Res.

    (2000)
  • H. Honda et al.

    Different mechanisms for testosterone-induced relaxation of aorta between normotensive and spontaneously hypertensive rats

    Hypertension

    (1999)
  • Cited by (38)

    • Age and sex differences in outcomes after in-hospital cardiac arrest

      2021, Resuscitation
      Citation Excerpt :

      In a previous study, a higher survival was reported in women in the reproductive age compared to men of similar age.13 Consequently, female sex hormones have been suggested as a potential mechanism that may drive the survival benefit observed in women, which is supported by the pleiotropic effects of estrogen on the cardiovascular system.29,30 In this study, women had a higher odds of survival to 30 days compared to men in the adjusted analyses, which aligns with several previous studies1,4.

    • Cardiac function and tolerance to ischemia/reperfusion injury in a rat model of polycystic ovary syndrome during the postmenopausal period

      2020, Life Sciences
      Citation Excerpt :

      However, the process of reperfusion can induce myocardial reperfusion injury, known as I/R injury [37]. It has been reported that the tolerance to I/R injury can be affected by different factors including, sex steroid hormones especially androgens and estrogens [38]. Several human and animal studies report a more adverse cardiovascular risk profile in subjects with PCOS compared to healthy counterparts during their reproductive years [9,39,40].

    • Sub-therapeutic doses of fluvastatin and valsartan are more effective than therapeutic doses in providing beneficial cardiovascular pleiotropic effects in rats: A proof of concept study

      2017, Vascular Pharmacology
      Citation Excerpt :

      Heart rate (HR) was obtained from oscillations detected in the electrocardiogram, aligned to ventricular pressure values and expressed in beats per min. Left ventricular pressure (LVP), expressed as the difference between systolic and diastolic pressures, was measured continuously with a Millar pressure catheter-transducer as previously described [9]. Data for all parameters measured in the isolated hearts were recorded and processed on a Dewetron acquisition system (Dewetron, Graz, Austria) after analogue-digital conversion (National Instruments, NI PCI-6013, Austin, USA) on a PC hard disk using Dewesoft 6.0 software (Dewetron, Trbovlje, Slovenia).

    • Characteristics and outcome among 14,933 adult cases of in-hospital cardiac arrest: A nationwide study with the emphasis on gender and age

      2017, American Journal of Emergency Medicine
      Citation Excerpt :

      This finding is in agreement with several previous studies [3,5,28,29]. The mechanisms behind this finding can only be speculated on, but it has been hypothesized that female sex hormones might play a role in protecting from ischemic-reperfusion injury and limiting post-resuscitation tumor necrosis factor-α response [30,31]. Other studies, however, have not demonstrated any gender advantage in terms of survival in either OHCA or IHCA, especially in older women [4,17,21,32].

    View all citing articles on Scopus

    Ischemic–reperfusion injury in isolated male rat hearts is reduced only after pretreatment of animals but not with direct administration of testosterone to the isolated heart.

    View full text