Investigation of apoptotic index and p53 expression in livers of rats in which obstructive jaundice was generated

Objective: Obstructive jaundice is a group of diseases which must be treated surgically. Its morbidity and mortality is very high due to complications of of bile ways attempts applied to patients with obstructive jaundice. Main complications are sepsis and renal failure.Apoptosis is a high level planned cell death mechanism which can occur in DNA-dependent cells in physiologic and pathologic cases. Many tissues and specially skin, intestine and immune system cells need apoptosis for the regular renewal. Apoptosis has a governing role in ischemic heart disease, paralysis, neurodegenerative diseases, sepsis and multi-organ disfunction syndrome. It is well understood that improvement of new pharmacological agents which will be able to block apoptosis will give rise to extraordinary treatment approaches that will lead in sepsis cure.The aim of this work is to determine the level of apoptosis, which is believed to hold an important role in septicemia process that affects mortality and morbidity in obstructive jaundice, in lingers of rats that were experimentally subjected to obstructive jaundice.The material and the method: This experimental study was performed in Experimental Surgery and Research Laboratories at Faculty of Medicine of Afyon Kocatepe University, under the approval of Animal Ethics Board of Faculty of Medicine-Kocatepe University. Twenty male type wistar-albino rats of which weights ranged between 270gr. and 320gr. were used. The experimentals were separated into two goups of eight. Choledoch was isolated in each group (Figure 4) and while surgery was ended at this level in the control group, choledoch was tied with 4-0 silk from two different places and cut between ligatures full fold. At the end of the seventh day blood was taken from the tails of the experiment animals and the results were analyzed statistically. Experiment animals were operated for the second time in the post-operative seventh day for liver sampling and sacrification-aimed histological analysis through the old incision with anaesthesia provided and care was taken to make sure that they do not feel pain. Liver tissue samples taken from the experiment and control groups were fixed by putting them in 10% neutral formalin. To exhibit the p53 expression immunohistochemically, anti-p53 Clone DO-7 (Labvision, Fremont CA, USA) was used as the primer antibody and HRP (=Horse Radish Paroxydase) (Labvision, Fremont CA, USA) as the secondary antibody. Samples taken for the determination of apoptosis were painted by the TUNEL method. In the samples observed under the light microscope, apoptotic and p53-positive cells were evaluated numerically in 10 different regions of every preparation at x20 optics objective zoom. UTHSCSA Image Tool Image Analysis Program was used to count the cells.Findings: At the end of the post-operative seventh day, the averages of direct and indirect bilirubin values in the rats in which obstructive jaundice was generated were found to be as 6.99/11.70 mg/dl, respectively. By Mann-Whitney U test, these results were observed to be statistically meaningful with respect to the control group (P=0.001). In the evaluation of apoptotic cells in liver cells (hepatocytes), apoptotic cells were observed to widely exist in the liver tissue and it was determined that they exhibited dense accumulation in some regions (Figure 1-4). Nevertheless, no apoptotic cell was encountered in the liver tissue of the rats in the control group (Figure 5-6). In the immunohistochemical evaluation made for evaluation of p53 expression in hepatocytes, p53-positive hepatocytes were determined to exist quite widely in the tissue samples taken from the livers of rats in the experiment group (Figure 7-9). Nevertheless, no cell exhibiting p53 expression were found in the tissue samples taken from the livers of rats of the control group.Result: Apoptosis is a closed box that must be investigated and which might make it possible to stop the many disesase processes or accelerate the healing. If the principal effective mechanism in the liver under a certain stress factor is apoptosis, it is definite that it will make a difference in the treatment approach. There is need for new studies for the clarification of this cascade and the changes in the treatment approach. ZVAD-fmk study is a good example for this.Consequently, in this study we determined that in the obstructive jaundice group, both apoptotic index and, as a result of the immunohistochemical studies, p53 expression increases in the liver.