الفهرس 
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Abstract
Environmental pollution is the contamination of the ecosystem that causes instability, disorder, harm or discomfort to the physical systems or living organisms. Increasing pollution is a major and global problem. This is due to the use of toxic chemicals or xenobiotic substances or by certain synthetic compounds such as heavy compounds.
Copper (Cu) is a trace element essential for life processes due to its role as acofactor in many enzymes (i.e. cytochrome oxidase, caeruloplasmin, lysil oxidase or superoxide dismutase).
Although Cu is a major essential trace element, excess Cu is extremely toxic, leading to many pathological conditions that are consistent with oxidative damage to membranes and molecules, the balance between deficiency and toxicity reflects its biological role. Copper toxicosis does sometimes occur, and it can even acutely lead to death. In the long term, toxicity is most typically manifested by the development of liver cirrhosis, along with episodes of hemolysis and damage to renal tubules, brain and other organs.
The liver is a major target organ in copper toxicity, where it tends to build up, causing diseases like Indian childhood cirrhosis, idiopathic copper toxicosis and tyrolean infantile cirrhosis.
So, copper toxicity and treatment represented an important area of attention, especially treatment by using of antioxidants to ameliorate the oxidative damage resulted from excessive copper consumption.
Up on the previous observations, the present study was conducted to experimentally study:
• Toxic effect of sub chronic oral administration of two forms of copper (ionic copper and nano sized copper particles) separately on rat liver.
• Compare two forms of copper toxicity.
• Evaluate the ameliorative role of orally administered Green tea extract (GTE) and Alpha lipoic acid (ALA or αLA) against copper induced hepatotoxicity in rats.
- In these study one hundred male albino rats were randomly divided into nine groups as following:
• Group I (C): Includes 10 rats, received distilled water and served as (-ve) control group.
• Group II (Cu): Includes 15 rats, received copper in the form of copper sulphate pentahydrate via oral gavage in a dose of 30 mg/ kg.bwt (it represent 1/10 of oral LD50) and served as (+ve) control group.
• Group III (Cu+G): Includes 10 rats, received copper in the form of copper sulphate pentahydrate via oral gavage in a dose of 30 mg/ kg.bwt plus green tea extract in drinking water in a dose of 5g/L.
• Group IV (Cu+LA): Includes 10 rats, received copper in the form of copper sulphate pentahydrate via oral gavage in a dose of 30 mg/ kg.bwt plus Alpha lipoic acid via oral gavage in a dose of 100mg/kg.bwt.
• Group V (N): Includes 15 rats, received copper in the form of nano sized copper particles suspension via oral gavage in a dose of 40 mg/ kg.bwt (it constitutes about 1/10 of oral LD50 of 25 nm sized copper particles and served as (+ve) control group.
• Group VI (N+G): Includes 10 rats, received copper in the form of nano sized copper particles suspension via oral gavage in a dose of 40 mg/ kg.bwt plus green tea extract in drinking water in a dose of 5g/L.
• Group VII (N+LA): Includes 10 rats, received copper in the form of nano sized particles copper suspension via oral gavage in a dose of 40 mg/ kg.bwt plus Alpha lipoic acid via oral gavage in a dose of 100mg/kg.bwt.
• Both copper (either ionic or nano particles), GTE and ALA were administered only 5 days/week.
• Group VIII (G): Includes 10 rats, received green tea extract in drinking water in a dose of 5g/L.
• Group IX (LA): Includes 10 rats, received Alpha lipoic acid via oral gavage in a dose of 100mg/kg.bwt.
• After the first month 5 rats were randomly selected from each group and sacrificed.
• After the second month 5 rats were randomly selected from each group and sacrificed.
• Stop administration of Copper sulphate pentahyderate or nano sized copper particles in the remaining 5 rats of Group II or Group V and administer distilled water only for one month. Then, they were sacrificed as Withdrawal subgroups [(W Cu) and (W N) respectively].
Twenty four hours after the last dose, rats were anesthetized with alcohol chloroform ether mixture (ACE mixture) in a ratio of 1:2:3 respectively and blood and tissue samples were collected.
- Blood samples were collected and divided in to two parts. First, non heparinzed blood for sera separation which kept at -20 oC in deep freezer till biochemical analysis [liver function evaluation (ALT, Cholesterol, Triglycerides and Total bilirubin measurement)] and the second, heparinzed blood for estimation of metals (Cu, Zn and Fe) concentrations .
- Rat’s livers were isolated then rinced in cold saline and dissected into 3 parts, first for estimation of metals (Cu, Zn and Fe) concentrations, second for biochemical parameters of oxidative stress measurement and third fixed in 10% fermol saline for histopathological study.
The following results were obtained:
1- Liver function evaluation tests:
Both ionic and nano copper intoxicated group showed significant increase in ALT activity, cholesterol and total bilirubin concentrations as well as significant decrease in triglyceride concentrations compared with control values.
While concurrent oral administration of green tea or alpha lipoic acid with both copper compounds induce amelioration of altered parameters to be near that of the control group.
2- Antioxidant enzymes activity:
Both ionic and nano copper intoxicated group showed significant increase in SOD activity and LPO concentration as well as significant decrease in GSH concentrations compared with control values.
While concurrent oral administration of green tea or alpha lipoic acid with both copper compounds induce amelioration of altered parameters to be near that of the control group.
3- Metals (Cu, Fe &Zn) concentrations:
A- Liver metals concentration:
Estimation of metals concentration in liver indicated that, administration of both ionic and nano copper intoxicated group showed significant increase in Cu and Fe concentrations as well as significant decrease in Zn concentration compared with control values.
While concurrent oral administration of green tea or alpha lipoic acid with both copper compounds induced decrease in Cu and Fe and increase in Zn concentrations compared with copper intoxicated groups.
B- Blood metals concentration:
Estimation of metals concentration in blood indicated that, administration of both ionic and nano copper intoxicated group showed significant increase in Cu and Zn and significant decrease in Fe concentrations compared with control values.
While concurrent oral administration of green tea with both copper compounds induced decrease in Cu and Zn and increase in Fe concentrations compared with copper intoxicated groups, in the other hand concurrent oral administration of alpha lipoic acid with both copper compounds induced decrease in Cu and increase in Zn and Fe concentrations compared with copper intoxicated groups.
4- Histopathological findings:
Histopathological examination of copper intoxicated rat’s liver showed degenerative changes in the cytoplasm of the hepatocytes, apoptosis in some individual hepatocytes , fibrosis in the portal area, dilatation in the portal vein and Kupffer cells were impacted by black pigment and proliferated in diffuse manner between the hepatocytes .
While concurrent oral administration of green tea or alpha lipoic acid with both copper compounds induced more or less normal histological picture as some slides showed mild degenerative changes in hepatocytes, slight inflammatory cells infiltrations and kupffer cells proliferation.
In the other hand withdrawal group induced no improvement in the copper intoxicated rat’s liver histological picture.