REV.CHIM.(Bucharest)♦69♦No.1♦2018 http://www.revistadechimie.ro 175 Correlations Between Oxidative Stress and Apoptosis During Anuran Metamorphosis VASILE SIRBU¹ , ANNAMARIA PALLAG 2 , ANA HONIGES 3 , VLADIMIR POROCH 4 *, SABINA IOANA COJOCARU¹ 1 Alexandru Ioan Cuza University of Iasi, Faculty of Biology, 20A,Carol I Blvd., 700505 Iasi, Romania 2 University of Oradea, Faculty of Medicine and Pharmacy, Pharmacy Department, 29 Nicolae Jiga Str., 410028 Oradea, Romania 3 Vasile Goldis Western University of Arad, 94-96, Revolutiei Blvd., 310025 Arad, Romania 4 Grigore T Popa Univeristy of Medicine and Pharmacy Iasi, 16 Universitatii Str., 700115 Iasi, Romania The living organisms can trigger the defense mechanisms against free radicals, by synthesizing different antioxidant enzymes. The present study is focused on establishing some correlation between oxidative stress and the structural changes in cell death at the intestinal larval epithelium level during anuran metamorphosis. Cell death in such conditions may be regarded as the result of an interaction activity in which takes place apoptosis, autophagy, and necrosis, the cell choosing one or more. The amphibian metamorphosis is a complex process, divided into three major periods: prometamorphosis, premetamorphosis and climax. The process ensures the passage of the organism from aquatic to terrestrial life, with dramatic changes in the morphology and structure of some organs. In the climax stages of metamorphosis, a variety of free radicals are produced, starting a numerous cellular oxidation reactions. Keywords: oxidative stress, apoptosis, autophagy, necrosis, anuran amphibians, metamorphosis. *email:vlader2000@yahoo.com; sabina_18ro@yahoo.com The metamorphosis of anuran amphibians represents a study model for different biological mechanisms such as the programmed apoptosis [1]. During the developement, apoptosis occurs normally as a proces to maintain normal cells populations in tissue [2]. This process is related to the morphogenesis [3] and is controlled by the thyroid hormones triiodothyronine and tetraiodothyronine, which in the climax stages induce cellular oxidative stress that lead to larval structures removal and their replacing with juvenile structures that will also function in the adult individuals. The amphibian metamorphosis is a complex process, divided into three major periods: prometamorphosis, premetamorphosis and climax. The process ensures the passage of the organism from aquatic to terrestrial life, with dramatic changes in the morphology and structure of some organs [4-6]. The larval intestine is one of the organs undergoing profound structural alterations in metamorphosis. In these structural reshuffle, the larval epithelium, equipped with an enzymatic set corresponding to microfage nutrition is replaced with juvenile epithelium, equipped with an enzymatic set intended for macrophage, aquatic and terrestrial feeding [7-18]. A big variety of free radicals are produced and they will start a big number of cellular oxidation reactions. The living organisms can trigger the defense mechanisms against free radicals, by synthesizing different antiradicalic enzymes [19-23]. Reactive oxigen species, oxidize biomolecules as proteins, lipids, carbohidrates, DNA and impair normal cellular functions. A shift in balaced between oxidant and antioxidant balance in favour of oxidants, results in oxidative stress. Antioxidant defence system comprise many enzimes known as antioxidant enzymes (superoxide dismutaza, catalase, glutathione peroxidase). The oxidative stress manifests extra and intracellular. Intracellular it acts on the cytoskeleton and on the membranes of different cellular organelles (nucleus, mitochondria, lysosoms), by degrading them [24]. The present study analyses the correlation between the oxidative stress and cells death as a result of oxidative stress, in the complex phenomenon of programmed cell death at Rana temporaria temporaria (L.1758) during metamorphosis. Experimental part Material and methods The biological material used in this study is represented by Rana temporaria temporaria (L.1758) pontes collected from nature. These were brought in the laboratory, were they hatch and were observed during larval development. They were maintained in 10 liters pots which contained dechlorinated tap water. The water temperature was between 18 - 22°C. The metamorphosis stages were established using Taylor si Kollros tables [25]. For each stage, seven individuals were sacrificed. The microdisection was performed on ice, under the binocular magnifying glass, the digestive tube exposed in Sörensen phosphate buffer and photographed. For histological observations, parts of the medium intestine were fixed in glutaraldehyde and osmium tetraoxide, then included in epoxy resins. The semifine sections were stained with toluidine blue. For electron microscopy the contrast was made with uranyl acetate. A part of the larval intestine was ice sections, colored with acridine orange and photographed on a fluorescence microscope. Superoxide dismutase (SOD - E.C. 1.15.1.1) activity was measured according to the method of Winterbourn et al. (1975) [26]. Enzyme activity was determined thanks to the capacity of this enzyme to inhibit Nitrobluetetrazolium reduction by superoxide anions generated after riboflavin photoreduction. The decrease of the absorbance was read at 560 nm using a Shimadzu UV-Vis 1700 spectro- photometer (Japan). Catalase (CAT - E.C. 1.11.1.6) activity was determined using the method of Sinha (Sinha, 1972) [27]. The method