Uticaj adrenalina i efedrina na pojavu primarnih oštećenja DNK u limfocitima čoveka in vitro

Abstract

U ovom radu cilj istraživanja je bilo ispitivanje primarnih oštećenja DNK izolovanih limfocita čoveka pod uticajem adrenalina i efedrina. Oštećenja DNK su evaluirana primenom in vitro Komet testa. Ispitivan je širok spektar koncentracija adrenalina i efedrina (u rasponu od 0,0005 μM do 500 μM) u različitim vremenskim intervalima (15 min, 60 min, 120 min, 240 min i 24 sata). Najizraženije oštećenje DNK ustanovljeno je nakon 15 min tretmana adrenalinom, pri čemu su sve koncetracije izuzev najniže (0.0005 μM) dovele do povećane migracije DNK. Nakon 60 min, 120 min, 240 min tretmana adrenalinom indukovano je oštećenje DNK u opsegu od 5 do 300 μM. Najslabiji efekat ispoljen je nakon 24 sata, tako da su samo najviše koncetracije adrenalina (150 μM i 300 μM) dovele do povećanog stepena oštećenja DNK. Radi utvrđivanja mogućeg učešća reaktivnih kiseoničnih vrsta (ROS) u indukovanju DNK oštećenja pod dejstvom adrenalina upotrebili smo antiokisanse katalazu (100 IU i 500 IU) i kvercetin (100 μM i 500 μM). Kotretman limfocita adrenalinom (300 μM) i antioksidansima nakon 15 ili 60 minuta doveo je do značajnog smanjenja količine DNK u repovima kometa. Prema tome, može se zaključiti da adrenalin ispoljava svoje genotoksične efekte indukcijom reaktivnih kiseoničnih vrsta i da se neka oštećenja poprave tokom prva četiri sata nakon tretmana adrenalinom. Za razliku od adrenalina, efedrin nije doveo do povećanja migracije DNK u odnosu na negativnu kontrolu tokom različitih vremenskih intervala. Jedino je koncetracija efedrina od 500 μM nakon 15 minuta tretmana indukovala oštećenje DNK koje je bilo statistički značajno.

The objectives of these investigations were to investigate primary DNA damage in isolated human lymphocytes exposed to adrenaline and ephedrine. DNA damage was evaluated by the in vitro Comet assay. A broad spectrum of adrenaline and ephedrine concentrations (range from 0.0005 μM to 300 μM) were examined in the Comet assay for various treatment times (15 min, 60 min, 120 min, 240 min and 24 h). The most profound DNA damage was observed after 15 min of adrenaline treatment, as all concentrations tested except the lowest one (0.0005 μM) caused an increase in DNA migration. After 1 h, 2 h and 4 h of treatment, adrenaline induced DNA damage at concentration range from 5 μM to 300 μM. The slightest DNA damage was observed after 24 h of adrenaline treatment, therefore only the highest concentrations of adrenaline (150 μM and 300 μM) caused increased level of DNA damage. In order to evaluate the potential contribution of reactive oxygen species (ROS)-induced DNA damage exposed to adrenaline, we used antoxidants catalase (100 IU and 500 IU) and quercetin (100 μM and 500 μM). Co-treatment of lymphocytes with adrenaline (300 μM) and antioxidants for 15 or 60 min, significantly reduced the quantity of DNA in the comet tails. Therefore, it can be concluded that adrenaline exhibits genotoxic effects mainly through induction of reactive oxygen species and that some of the DNA damage is repaired during the first four hours following the treatment with adrenaline