İn vitro lipopolisakkarit uyarımlı karaciğer yangı modelinde alfa-terpineol'ün etkinliğinin araştırılması
Investigation of the effectiveness of alpha-terpineol in the in vitro lipopolysaccaride stimulated liver inflammation model
Yazar: AYŞE PINAR TUNÇER
Danışman: DOÇ. DR. ALTUĞ KÜÇÜKGÜL
Hatay Mustafa Kemal Üniversitesi / Sağlık Bilimleri Enstitüsü / Biyokimya Ana Bilim Dalı
Derece: Yüksek Lisans, 2020
Konu: Biyokimya = Biochemistry
Dizin: Alfa terpineol = Alpha terpineol ; Karaciğer = Liver ; Karaciğer hastalıkları = Liver diseases ; Lipopolisakkaritler = Lipopolysaccharides ; İn vitro = In vitro ; İnflamasyon = Inflammation

Özet:
In Vitro Lipopolisakkarit Uyarımlı Karaciğer Yangı Modelinde Alfa-terpineol'ün Etkinliğinin Araştırılması
 Terpinollerin birçok çalışmada antioksidan özelliklerinden dolayı antikanser, antikonvülsant ve antiülser gibi biyolojik etkileri ortaya konulmuştur. Araştırmanın amacını Ecoli'den izole edilen LPS (lipopolisakkarit)'in karaciğer hücrelerine etkin konsantrasyonda uygulanarak oluşturulan yangı modelinde α-terpineol'ün anti-inflamatuar ve anti-apoptotik gibi biyofonksiyonları araştırılması oluşturdu.
Bu amaçla deneme materyali olarak sürekli hücre hatlarından insan orijinli HepG2 hücreleri materyal olarak seçildi. Etken madde olarak seçilen LPS ve α-terpineol, ticari olarak temin edildi ve hücrelere farklı konsantrasyonlarda 24 saat uygulandı.  Her iki etken maddenin etkin konsantrasyonları, hücre canlılık testleriyle gerçekleştirildi.  Süre sonunda alınan mRNA örneklerinden ise, TNF-α, IL-1β ve IL-10,  Kaspaz 3, Bax ve Bcl-2 gen ekspresyon düzeyleri gerçek zamanlı kuantatif polimeraz zincir reaksiyonuyla (qRT-PCR) araştırıldı.
Araştırmada elde edilen sonuçlara göre LPS'in farklı konsantrasyonları hücrelere uygulanarak kontrol grubuna göre %11.5 oranında hücre kayıplarına neden olan 50 ng/ml konsantrasyonu modelleme için seçildi. Ancak α-terpineolün LPS ile birlikte kullanıldığında 10 μM konsantrasyonunda hücre kayıplarını % 2.12 oranında önlediği bulundu. Çalışmada gen analizleri değerlendirildiğinde LPS'in TNF-α ve IL-1β gen ekspresyonlarını arttırdığı, α-terpineol uygulamasının ise bu durumu tersine çevirdiği tespit edildi. Yine IL-10 gen ekspresyonu LPS tarafından baskılanırken, α-terpineol tarafından anlamlı düzeyde uyarıldığı da ortaya konuldu. Bununla birlikte çalışmada LPS'in kaspaz 3 ve Bax gen ekspresyonlarını arttırdığı, ancak α-terpineol'ün bu stimülasyonu baskıladığı tespit edildi. Ayrıca buna ilaveten anti-apoptotik Bcl-2 gen ekspresyonları LPS tarafından baskılanırken, α-terpineol tarafından uyarıldığı bulundu.
Araştırmadan elde edilen verilerle, monoterpen α-terpineol'ün kısa süreli ve düşük olan  proliferatif konsantrasyonunun özellikle patojen kaynaklı  karaciğer   rahatsızlıklarında alternatif bir tedavi ajanı olabileceği görüldü.  Bununla birlikte konunun tam aydınlatıabilmesi için in vivo ve daha ileri düzey araştırmalara ihtiyaç duyulduğu kanısına varıldı.
Anahtar Kelimeler: Lipopolisakkarit,  inflamasyon, α-terpineol, apoptoz.

Abstract:
Due to the antioxidant properties of terpineols, biological effects such as anticancer,
anticonvulsant and antiulcer have been revealed in many studies. The aim of the research
was to investigate the biofunctions of α-terpineol such as anti-inflammatory and
antiapoptotic in the inflammatory model created by applying LPS isolated from Ecoli to liver
cells at an effective concentration.
For this purpose, HepG2 cells of human origin from continuous cell lines were
selected as the test material. LPS and α-terpineol, selected as the active ingredient, were
commercially available and applied to the cells at different concentrations for 24 hours.
Effective concentrations of both active ingredients were carried out by cell viability tests.
TNF-α, IL-1β and IL-10, caspase 3, Bax and Bcl-2 gene expression levels were investigated
by real-time quantitative polymerase chain reaction (qRT-PCR).
According to the results obtained in the study, different concentrations of LPS were
applied to the cells and 50 ng / ml concentration that caused 11.5 % cell loss compared to
the control group was selected for modeling. However, when used together with LPS, αterpineol was found to prevent cell loss at a concentration of 10 μM at a rate of 2.12%. When
gene analysis was evaluated in the study, it was found that LPS increased gene expressions
of TNFα and IL-1β, and α-terpineol application reversed this situation. The other IL-10 gene
expression was also suppressed by LPS and it was significantly stimulated by α-terpineol.
Besides, in the study, it was found that LPS increased caspase 3 and Bax gene expressions,
while α-terpineol significantly suppressed this stimulation. In addition,Bcl-2 gene
expressions were found to be suppressed by LPS and α-terpineol application stimulated this
gene expression.
With the data obtained from the study, it was seen that the short-term and low
proliferative concentration of monoterpene α-terpineol may be an alternative treatment agent
especially in pathogen-induced liver disorders. However, it was concluded that in vivo and
more advanced studies are needed to fully illuminate the subject.
Keywords: Lipopolysaccharide, inflammation, α-terpineol, apoptosis. 

Investıgatıon of the Effectıveness of Alpha-terpineol in the In Vitro lipopolysaccaride Stimulated Liver Inflammation Model
	Due to the antioxidant properties of terpineols, biological effects such as anticancer, anticonvulsant and antiulcer have been revealed in many studies. The aim of the research was to investigate the biofunctions of α-terpineol such as anti-inflammatory and antiapoptotic in the inflammatory model created by applying LPS isolated from Ecoli to liver cells at an effective concentration.
	For this purpose, HepG2 cells of human origin from continuous cell lines were selected as the test material. LPS and α-terpineol, selected as the active ingredient, were commercially available and applied to the cells at different concentrations for 24 hours. Effective concentrations of both active ingredients were carried out by cell viability tests. TNF-α, IL-1β and IL-10, caspase 3, Bax and Bcl-2 gene expression levels were investigated by real-time quantitative polymerase chain reaction (qRT-PCR).
	According to the results obtained in the study, different concentrations of LPS were applied to the cells and 50 ng / ml concentration that caused 11.5 % cell loss compared to the control group was selected for modeling. However, when used together with LPS, α-terpineol was found to prevent cell loss at a concentration of 10 μM at a rate of 2.12%. When gene analysis was evaluated in the study, it was found that LPS increased gene expressions of TNFα and IL-1β, and α-terpineol application reversed this situation. The other IL-10 gene expression was also suppressed by LPS and it was significantly stimulated by α-terpineol. Besides, in the study, it was found that LPS increased caspase 3 and Bax gene expressions, while α-terpineol significantly suppressed this stimulation. In addition,Bcl-2 gene expressions were found to be suppressed by LPS and α-terpineol application stimulated this gene expression.
	With the data obtained from the study, it was seen that the short-term and low proliferative concentration of monoterpene α-terpineol may be an alternative treatment agent especially in pathogen-induced liver disorders. However, it was concluded that in vivo and more advanced studies are needed to fully illuminate the subject.
Keywords: Lipopolysaccharide, inflammation, α-terpineol, apoptosis.