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Di (2-Etilhekzil) Fitalatın Pubertal Dönemdeki

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Di (2-Etilhekzil) Fitalatın Pubertal Dönemdeki Dİ (2-ETİLHEKZİL) FİTALATIN PUBERTAL DÖNEMDEKİ ERKEK SIÇANLARDA GENOTOKSİK, HİSTOLOJİK VE BİYOKİMYASAL ETKİLERİNİN ARAŞTIRILMASI INVESTIGATION OF GENOTOXIC, HISTOLOGIC AND BIOCHEMICAL EFFECTS OF DI (2-ETHYLHEXYL) PHTHALATE ON PUBERTAL MALE RATS GÖZDE KARABULUT Prof. Dr. NURHAYAT BARLAS Tez Danışmanı Hacettepe Üniversitesi Lisansüstü Eğitim – Öğretim ve Sınav Yönetmeliğinin Biyoloji Anabilim Dalı için Öngördüğü YÜKSEK LĠSANS TEZĠ olarak hazırlanmıştır. 2014 GÖZDE KARABULUT ’un hazırladığı “Dİ (2-ETİLHEKZİL) FİTALATIN PUBERTAL DÖNEMDEKİ ERKEK SIÇANLARDA GENOTOKSİK, HİSTOLOJİK VE BİYOKİMYASAL ETKİLERİNİN ARAŞTIRILMASI ʺ adlı bu çalışma aşağıdaki jüri tarafından BİYOLOJİ ANABİLİM DALI ’nda YÜKSEK LİSANS TEZİ olarak kabul edilmiştir. Prof. Dr. Dürdane KOLANKAYA Başkan Prof. Dr. Nurhayat BARLAS Danışman Prof. Dr. Güldeniz SELMANOĞLU Üye Prof. Dr. Arzu KOÇKAYA Üye Prof. Dr. Ġsmet ÇOK Üye Bu tez Hacettepe Üniversitesi Fen Bilimleri Enstitüsü tarafından YÜKSEK LİSANS TEZİ olarak onaylanmıştır. Prof. Dr. Fatma SEVĠN DÜZ Fen Bilimleri Enstitüsü Müdürü Canım Aileme… ETİK Hacettepe Üniversitesi Fen Bilimleri Enstitüsü, tez yazım kurallarına uygun olarak hazırladığım bu tez çalışmasında; tez içindeki bütün bilgi ve belgeleri akademik kurallar çerçevesinde elde ettiğimi, görsel, işitsel ve yazılı tüm bilgi ve sonuçları bilimsel ahlak kurallarına uygun olarak sunduğumu, başkalarının eserlerinden yararlanılması durumunda ilgili eserlere bilimsel normlara uygun olarak atıfta bulunduğumu, atıfta bulunduğum eserlerin tümünü kaynak olarak gösterdiğimi, kullanılan verilerde herhangi bir tahrifat yapmadığımı, ve bu tezin herhangi bir bölümünü bu üniversite veya başka bir üniversitede başka bir tez çalışması olarak sunmadığımı beyan ederim. 09/09/2014 GÖZDE KARABULUT ÖZET Dİ (2-ETİLHEKZİL) FİTALATIN PUBERTAL DÖNEMDEKİ ERKEK SIÇANLARDA GENOTOKSİK, HİSTOLOJİK VE BİYOKİMYASAL ETKİLERİNİN ARAŞTIRILMASI Gözde KARABULUT Yüksek Lisans, Biyoloji Bölümü Tez Danışmanı: Prof. Dr. Nurhayat Barlas Eylül 2014, 104 sayfa Bu çalıĢmada, plastikleĢtirici olarak kullanılan ve gündelik hayatta sıkça maruz kalınan 2-etilhekzilfitalat’ın (DEHP) pubertal dönemdeki erkek sıçanlarda genotoksik etkileri Comet testi ile araĢtırılmıĢ, histopatolojik, hematolojik ve biyokimyasal analizler yapılarak incelenmiĢtir. Bu amaçla sıçanlar her grupta 6 hayvan olacak Ģekilde, pozitif kontrol (MMS), yağ kontrol, 100 mg/kg/gün, 200 mg/kg/gün ve 400 mg/kg/gün uygulama gruplarına ayrılmıĢ ve 28 gün süreyle sıçanlara DEHP gavaj yoluyla oral uygulanmıĢtır. Deney sonuna kadar her hafta alınan kan örneklerinde genotoksik analiz yapılmıĢtır. Deney süresince sıçanlarda vücut ağırlığı, yem ve su tüketimleri ölçülmüĢtür. Deney süresi sonunda sıçanlardan alınan serum örneklerinde alanin aminotransferaz (ALT), aspartat transaminaz (AST), üre, ürik asit, trigliserit, glikoz, kreatinin miktarları ölçülmüĢtür. Ayrıca kanda hematolojik analiz yapılmıĢtır. Karaciğer ve böbrek dokularında oluĢabilecek histopatolojik etkiler ıĢık mikroskobunda incelenmiĢ ve değiĢik büyütmeler ile görüntülenmiĢtir. DEHP uygulanan gruplarda DNA hasarı oluĢumu kontrol grubuna göre yüksek bulunmuĢtur. Trigliserit seviyelerinin de kontrol grubuna göre anlamlı olarak yüksek olduğu saptanmıĢtır. Yapılan histopatolojik incelemeler, DEHP’in böbrekte ve karaciğerde histopatolojik değiĢikliklere neden olduğunu ortaya koymuĢtur. Anahtar Kelimeler: Di (2-etilhekzil) Fitalat, Genotoksisite, Comet Testi, Sıçan, Karaciğer, Böbrek. i ABSTRACT INVESTIGATION OF GENOTOXIC, HISTOLOGIC AND BIOCHEMICAL EFFECTS OF DI (2- ETHYLHEXYL) PHTHALATE ON PUBERTAL MALE RATS Gözde KARABULUT Master of Science, Department of Biology Supervisor: Prof. Dr. Nurhayat Barlas September 2014, 104 pages In the present study, we investigated the genotoxic effects of DEHP which is widely used as a plasticizer and people were exposed in daily life on pubertal male rats. These effects were investigated with Comet assay and histopathological, hematological and biochemical analysis were investigated. For this purpose, rats were divided into 5 groups and each group had 6 animals. The dose groups were oil control, positive control (MMS), 100 mg/kg/day, 200 mg/kg/day and 400 mg/kg/day of DEHP. It was administered daily to the rats by oral gavage for 28 days and every week until the end of the experiment, genotoxic effect was measured in blood. During the experiment, body weights and feed and water consumption were recorded. At the end of the experiment blood samples were taken and hematological analysis, alanine aminotransferase (ALT), aspartate aminotransferase (AST), ure, uric acid, trigliserid, glucose, creatinine were determined in serum. Kidney and liver were dissected and tissues were investigated potential histopathologic findings and monitorized in different magnifications by the Leica light microscopy. DNA damage level was statistically different from treatment groups compared to the oil control. Triglycerid levels were also significantly higher in HFCS and sucrose groups. Histopathological examinations revealed histopathological changes in kidney and liver of rats received DEHP. Keywords: Di (2-ethylhexyl) Phthalate, Genotoxicity, Comet Assay, Rat, Liver, Kidney. ii TEŞEKKÜR ÇalıĢmam süresince beni yönlendiren, bilgi ve tecrübelerini benimle paylaĢarak bana her konuda yardımcı olan ve desteğini esirgemeyen çok değerli danıĢman hocam Sayın Prof. Dr. Nurhayat BARLAS’a, Tezimin son halini almasında önerileri ile beni yönlendiren çok değerli jüri üyelerine, Comet testi analizlerinin yapılabilmesi için bize laboratuvarını açan sayın hocam Prof. Dr. Ġsmet ÇOK’a ve analizlerin hepsinde yanımda olup yardımcı olan sevgili uzman biyolog arkadaĢım GülĢen GÖNEY’e Deneylerimde ve her konuda bana yardımcı olup yol gösteren sevgili Egemen FOTO ve AraĢ. Gör. Fatma ZĠLĠFDAR’a, Tezimin deneysel aĢamalarında yanımda olan ve yardımlarını esirgemeyen, değerli arkadaĢlarım Eylül TURASAN, Pınar KAÇAMAK ve Elnaz YOUSEFĠ ARDEBĠLĠ’ye Yardımlarını hiçbir zaman esirgemeyen baĢta oda arkadaĢım AraĢ. Gör. Mehmet KürĢat ġAHĠN olmak üzere tüm Hacettepe Üniversitesi Biyoloji Bölümü Zooloji Anabilim Dalı Öğretim Üyeleri ve Yardımcılarına, Ne zaman arasam hemen koĢarak gelen ve tez içinde ve dıĢında her konuda destek ve yardımcı olan canım kuzenim Damla Ceren AKIN’a, her konuda yanımda olup bana destek olan sonsuz güvendiğim canım teyzelerim Reyhan AKIN ve ġükran ÖZDEMĠR’e, Lisans döneminden beri yanımda olan, iyi ve kötü her anımı paylaĢtığım, her ihtiyacımda bir telefon kadar yakın olan canım dostlarım AraĢ. Gör. Hasan ÇALIġKAN, Zeliha YILMAZ ve AraĢ. Gör. Merve ÖZCAN’a, Yüksek lisansa baĢladığım günden beri birlikte güldüğüm, üzüldüğüm, sevindiğim, gezdiğim, eğlendiğim kısacası her anımı paylaĢtığım değerli dostlarım Derya AKKURT ve AraĢ. Gör. Sevcan ALDEMĠR’e, Beni gittiği her yolculukta yanında götüren ve bu sayede uçak fobimi yenmemi sağlayan, her arayıĢımda bana güç veren ve güldüren, değerli hediyeleriyle beni Ģımartan değerli dostum Av. Ġpek GENÇOĞLU’na, iii Her Ģeyden önemlisi, hayatımın bütün anlarında yanımda olan, maddi ve manevi olarak hep beni destekleyen, güvenini hep hissettiğim canım babam Sermet KARABULUT’a, beni büyütüp bugünlere getiren, her zaman arkamda olduğunu bilip güvenini her zaman hissettiğim, dünyanın en fedakar annesi olan canım annem Nuran KARABULUT’a, her konuda bana akıl ve fikir verip yol gösteren, olgunluğuyla bana çok Ģey katan ve yukarılara çıkaran, onsuz bir hayat düĢünemediğim canım yol arkadaĢım, canımın içi kardeĢim Aslı KARABULUT’a sonsuz teĢekkür ederim. Gözde KARABULUT Ankara, Eylül 2014 iv İÇİNDEKİLER Sayfa ÖZET……………………………………………………………………………………...i ABSTRACT……………………………………………………………………………...ii TEġEKKÜR……………………………………………………………………………...iii ĠÇĠNDEKĠLER…………………………………………………………………………....v 1.GĠRĠġ .............................................................................................................. 1 2. GENEL BĠLGĠLER…………………………………………………………………...3 2.1.ENDOKRĠN BOZUCULAR ........................................................................... 3 2.1.1.Fitalatlar ..................................................................................................... 8 2.1.1.1.Fitalatların Biyotransformasyonları ....................................................... 13 2.1.1.2.Fitalatların Yaptığı Toksik Etkiler .......................................................... 14 2.1.1.3 Di (2-etilhekzil) Fitalat .......................................................................... 16 2.1.1.3.1 DEHP’ in Metabolizması......................................................................19 2.1.1.3.2 DEHP ve Toksisitesi .......................................................................... 20 2.1.1.3.3 DEHP ve DNA Hasarı ........................................................................ 21 2.2. Genetik Toksikoloji……………………………………….................................22 2.2.1. DNA Hasarı..............................................................................................24 2.2.2. Genotoksisitenin Karsinojenezisle ĠliĢkisi.................................................27 2.2.3. Sık Kullanılan Bazı Genotoksisite Testleri...............................................28 2.2.3.1 Kromozomal aberasyon (CA) testi ........................................................ 29 2.2.3.2. Mikroçekirdek Testi.............................................................................. 29 2.2.3.3.KardeĢ Kromatid DeğiĢimi (Sister chromatid Exchange=SCE)............. 29 2.2.3.4. COMET Tekniği (Single Cell Jel Elektroforez Tekniği) ........................ 29
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