الفهرس 
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Abstract
Non-alcoholic fatty liver disease is the most common liver disease. Fatty liver ranges from simple steatosis to non-alcoholic steatohepatitis and may progress to liver-related complications such as cirrhosis, liver cancer. NAFLD is related to several factors such as obesity and diabetes. Till now, no specifically licensed treatment is available.
The aim of the present study is examine the changes that occur in rats after induction of NAFL by high-fat diet for 18 weeks. The levels of these changes include: the changes in body weight and liver weight, the histological structure of the liver tissue, glucose homeostasis, hepatic lipid accumulation, liver function tests, bilirubin and serum lipid profile, as well as glutathione (GSH and oxidized) in the blood and tissues, in addition to alteration in one-carbon metabolism and hepatic global DNA methylation. The aim of this study was also to evaluate the therapeutic effect of one-carbon donors (betaine, choline and folic acid) in NAFL rats.
One hundred and ten male Wistar rats weighing approximately 100-120 g were used in the present study and were divided as follows:
group I (Normal control):
- Ten normal rats were fed standard diet throughout the whole experimental period (18 weeks).
After establishing the fatty liver (after 14 weeks confirmed by US) the NAFL rats (100 rats) were subsequently divided into 4 groups:
group II (Positive control):
- Ten NAFL rats maintained on HFD without treatment for 4 weeks.
group III (Betaine treated group):
- Thirty NAFL rats maintained on HFD and treated orally for 4 weeks with different daily doses of betaine (100, 200, 400 mg/kg); using 10 rats for each dose.
group VI (Choline treated group):
- Thirty NAFL rats maintained on HFD and treated orally for 4 weeks with different daily doses of choline (10, 20, 50 mg/kg); using 10 rats for each dose.
group V (Folic acid treated group):
- Thirty NAFL rats maintained on HFD and treated orally for 4 weeks with different daily doses of folic acid (25, 50, 75 mg/kg); using 10 rats for each dose.
Starting from the end of week 14, fatty liver was confirmed using ultrasound. At the end of the experiment, rats were dissected, blood was collected for biochemical analysis of glucose and insulin, serum lipid profile, liver function tests, bilirubin and liver was divided into small parts for histological examination and to calculate the level hepatic lipid content, s-adenosylmethionine and s-adenisylhomocystiene,global DNA methylation as well as glutathione (GSH and oxidised) in the blood and tissues.
Results revealed that non-alcoholic fatty liver rats had a statistically significant increase in the FBS level compared to the control group, as well as insulin resistance. These disturbances in glucose homeostasis in the blood accompanied by serious changes in the level of lipid profile; high serum levels of total triglycerides and cholesterol, LDL-C and reduced HDL-C. As well as hepatic lipid accumulation that was sharply increased.
The results also pointed to the exposure of NAFL rats to a state of oxidative stress where the serum and tissue contents of reduced glutathione decreased while oxidized glutathione showed a statistically significant increase where oxidative stress resulted in the consumption of reduced glutathione and its transformation into oxidized form.
Epigenetic changes are the key factors in pathogenesis and progression of non-alcoholic fatty liver disease where the NAFL rat results showed a low level of s-adenosylmethionine and a remarkable increase in s-adenosyl homocystiene accompanied by a decrease in DNA methylation.
from the results of the present study it is clear that all the used one-carbon donors had dose-dependent ameliorating effects on the different pathogenic pathways that participated in the induction of NAFL in rats. They all acted to reduce accumulation of fat in the liver, liver function parameters, bilirubin. These biochemical data were confirmed by the histological findings. Also, they improved hepatic redox systems and increased the hepatic global DNA methylation. Although, all the drugs used produced significant effects; there were some discrepancies concerning the extent of the effect and the dose.
The differences between the three one-carbon donors used in the present study were quiet evident on the lipid profile and glycemic parameters. The effects of folic acid and choline were qualitatively similar as they both decreases FBS, insulin and insulin HOMA-IR in a dose dependent matter. They also improved the lipid profile but values of serum TG, cholesterol, LDL-C and HDL-C were still significantly different from normal control values. In contrast, betaine treatment of NAFL rats had no significant effect on FBS level at the lowest doses but surprisingly increased its level in the high dose-treated group as compared to the untreated control group. It also extended the least beneficial effect on the lipid profile parameters.
In folic acid treated NAFL rats, improvements in glucose homeostasis and lipid metabolism in the liver is mediated through the correction of derangements in one-carbon metabolism and the significant surge in SAM/SAH ratio by folic acid. Choline caused similar effects to folic acid, but to a lesser extent. Betaine, another one-carbon donor, is formed from choline and was expected to produce similar effects through the one-carbon metabolism. However, they differentially affected fasting blood sugar.
In this study, we found that after treatment with three one-carbon donors (betaine, choline, folic acid) the percentage of hepatic s-adenosylmethionine / s-adenosyl homocystiene increased with increasing the dose of drugs, accompanied with an increase in methylated DNA.
In conclusion, HFD feeding of rats is a suitable model for NAFLD as it showed the main features of fatty liver at the levels of body and liver weights, histological manifestations, hepatic lipid content, liver function tests, glucose homeostasis, lipid abnormalities, redox balance, one-carbon metabolism and hepatic DNA methylation. The treatment of this NAFL rat model using three one-carbon donors (choline, betaine and folic acid) resulted in significant corrective effects at all levels. The collective comparison between the used drugs indicated that folic acid might have the most prominent effects than other donors.
The results of this study along with the available evidence strongly suggests that it is possible to use betaine, choline and folic acid for the treatment of non-alcoholic fatty liver disease.