Uticaj subhroničnog opterećenja homocisteinom na koronarnu hemodinamiku i oksidacioni stres kod pacova: efekti sumporovitih aminokiselina (metionina, N-acetilcisteina i cisteina) i neorganskog Na-hidrogensulfida

Abstract

Uvod: Sve je veća istraživačka zainteresovanost za delovanje sumporovitih jedinjenja na kardiovaskularni sistem, pod fiziološkim i patološkim uslovima. Cilj: Cilj ove eksperimentalne studije bio je ispitivanje efekata akutne primene sumporovitih aminokiselina (L-metionina, L-cisteina, N-acetilcisteina) i neorganskog Na-hidrogensulfida na kardiodinamske parametre, koronarni protok i redoks ravnotežu kod pacova sa normalnim i povišenim vrednostima homocisteina u krvi. Materijal i metode: Istraživanje je obuhvatilo 80 Wistar albino pacova koje smo podelili u dve grupe: kontrolna grupa (dvonedeljni tretman sa 0,9 % NaCl) i eksperimentalna grupa (subhronična primena DL-homocisteina, u dozi od 0,45 μmol/g telesne mase, dva puta dnevno, subkutano, u trajanju od 14 dana). Nakon dvonedeljnog tretmana, pacovi su žrtvovani pri čemu su prikupljeni uzorci krvi za analizu koncentracija ukupnog homocisteina i biomarkera sistemskog oksidacionog stresa. Izvršena je procedura izolovanja srca pacova, u cilju uspostavljanja retrogradne perfuzije kroz srce metodom po Langendorfu i sprovođenja akutnog tretmana sumporovitim aminokiselinama (L-Meth, L-Cys, NAC) i neorganskim Nahidrogensulfidom (NaHS). Sumporovite supstance su primenjene u istoj dozi od 0.5 mmol/L, u trajanju od 5 minuta. U levoj komori su kontinuirano praćeni parametri funkcije leve komore: dp/dtmax, dp/dtmin, SLVP, DLVP i HR. Koronarni protok (CF) se određivao floumetrijski. U uzorcima krvne plazme i uzorcima koronarnog perfuzata, spektrofotometrijskim metodama, određivani su biomarkeri oksidacionog stresa: indeks lipidne peroksidacije (TBARS), azot monoksid u formi nitrita (NO2-), superoksid anjon radikal (O2-) i vodonik peroksid (H2O2). U hemolizatu eritrocita su mereni parametri antioksidacionog sistema zaštite: katalaza (CAT), superoksid dismutaza (SOD) i redukovani glutation (GSH). Rezultati: Rezultati istraživanja su prikazani kroz 6 tabela i 125 grafikona. Koncentracije ukupnog homocisteina su bile značajno povišene u eksperimentalnoj u odnosu na kontrolnu grupu pacova, i kao takve izazivaju znatne promene u funkciji kardiovaskularnog sistema. Takođe, akutni efekti primenjenih sumporovitih supstanci su pokazali značajne razlike u odnosu na tretman DL- homocisteinom, na osnovu čega zaključujemo da sumporovite supstance ispoljavaju različito dejstvo na kardiovaskularni sistem u zavisnosti od prisustva homocisteina. Zaključci: Analizom rezultata ovog istraživanja zaključuje se da efekti sumporovitih aminokiselina (L-Meth, L-Cys, NAC) i neorganskog NaHS imaju značajnu ulogu u patofiziologiji kardiovaskularnog sistema pri čemu im je glavna karakteristika dozno-zavistan efekat na funkciju kardiovaskularnog sistema što je i osnovna karakteristika jedinjenja koja u svom sastavu imaju sumpor.

Introduction: There is growing interest in the activity of sulfur-containing compounds on cardiovascular system in physiological and pathological conditions. Aim: Aim of this study was to assess possible differences in the effects of various sulfurcontaining compounds on cardiodynamic parameters, coronary flow and redox balance of cardiovascular system, in its physiological state and in the early onset of hyperhomocysteinemia. Material and methods: This study divided Wistar albino rats into two groups: saline treated (control group) and DL-homocysteine-treated (experimental group). Rats from experimental group were subjected to subchronic subcutaneous administration of DL-homocysteine at dose of 0.45 μmol/g body weight, twice a day for 2 weeks. At the end of this period, rats were sacrificed, and blood samples were collected to be analysed for homocysteine concentration and biomarkers of systemic oxidative stress. Isolated rat hearts were excised and attached to the Langendorff apparatus. To assess the effects of acute administration of L-methionine, Lcysteine, N-acetylcysteine, and sodium hydrogen sulfide, the hearts were perfused individually with each of the mentioned substances at same single dose of 0.5 mmol/L for 5 min. Before, during and after perfusion of isolated rat heart (control, effect and washout) we continuously monitored (after insertion and placement a sensor in the left ventricle) the cardiodynamic parameters: a) dp/dt max - maximum rate of change of left ventricle pressure, b) dp/dt min - minimum rate of change of left ventricle pressure, v) SLVP - systolic left ventricular pressure, G) DLVP - diastolic left ventricular pressure and d) HR - heart rate. In the blood and effluent samples we measured biomarkers of oxidative stress: an index of lipid peroxidation (TBARS), nitric oxide in the form of nitrite (NO2-), superoxide anion radical (O2- ), and hydrogen peroxide (H2O2), and the parameters of antioxidant protection system: catalase (CAT), superoxide dismutase (SOD) and reduced glutathione (GSH). Results: The research results are presented in 6 tables and 125 graphs. Total homocysteine level was significantly higher in the experimental group than in the control group, and the effects of applied sulfur-containing compounds were significantly different in experimental and control groups. DL-homocysteine induced considerable changes in functioning of cardiovascular system even before an increase in plasma homocysteine values, and action of sulfur-containing compounds varied depending on the presence of homocysteine. Conclusions: After analyzing the results of this study, it is concluded that the effects of sulfuric amino acids (L-Meth, L-Cys, NAC) and inorganic NaHS play a significant role in the pathophysiology of the cardiovascular system, whereby their he main characteristic is the dose-dependent effect on the function of the cardiovascular system, which is the dominant quality of sulfur-containing compounds.