Biochemical, Cytogenetic and Histopathological studies on the effects of Agrimonia eupatoria extracts on albino male mice

number: 
2419
English
Degree: 
Imprint: 
Biotechnology
Author: 
Orooba Nadhum Harbi Al-bederi
Supervisor: 
Dr. Khulood W. Al-Sammarrae
Dr.Ali H. Ad'hiah
year: 
2009
Abstract:

The study was conducted to evaluate some immunological (total and absolute counts of leucocytes), cytogenetic (mitotic index of bone marrow cells, micronucleus formation in bone marrow cells and sperm-head and- tail abnormalities), biochemical (aspartate amino-transferase; AST, alanine aminotransferase; ALT and alkaline phosphatase; ALP), histopathological (liver) and antitumor (human cervical cancer; HeLa and Rhabdomyosarcoma; RD cell lines and a normal cell line; mouse embryo fibroblast; MEF) effects of agrimony (Agrimonia eupatoria L.) extracts (aqueous and methanol), and their effects on carbon tetrachloride (CCl4)-induced acute hepatic injury, in albino male mice. Additionally, a chemical detection of flavonoids, tannins, alkaloids, glycosides and saponins was carried out and furthermore, flavonoids were detected by a thin layer chromatography (TLC) method. Such evaluations were carried out through three stages. In stage I, three doses (1.0, 2.0 or 3.0 mg/kg) of agrimony extracts and a single dose (3.2 mg/kg) of CCl4 were investigated. The tested materials were injected intraperitoneally as a single dose (0.1 ml) per a day and for 6 days, and then the mice were sacrificed on day 7 for laboratory assessments. In the case of CCl4, it was injected subcutaneously as a single dose (0.1 ml) for one a day, and then the mice were sacrificed in the second day. In stage II, interactions between the ideal dose (3.0 mg/kg) of each extract and CCl4 were carried out. In such interactions . the animals were given CCl4 on day one, while the plant extract was given in day 2 till day 6 (single dose/day), and then, the animals were sacrificed on day 7 for laboratory assessments. In stage III, the percentage growth inhibition (PGI) of five plant concentrations (6, 12, 24, 48 and 96 μg/ml) were assessed in vitro using two tumor cell lines (HeLa and RD) and one normal cell line (MEF). Chemical detections of agrimony extracts revealed that the aqueous extract was positive for flavonoids, tannins and phenolic compounds, while the methanol extract was positive for flavonoids, and tannins. The TLC analysis showed that the aqueous extract manifested four types of flavonoids (rutin, kaempferol, quercetin, isorhamnetin and myricetin), while in the methanol extract five flavonoids were detected (rutin, myricetin, azoleatin, vitexin and iso-orientin). The results revealed that CCl4-treated animals showed significant decreased counts of total and absolute counts of leucocytes, Furthermore, significant increased percentages of induced micronucleus formation and sperm-head and-tail abnormalities and a significant decreased mitotic activity of bone marrow cells were also observed. In contrast, the aqueous and methanol extracts of agrimony were significantly effective in enhancing the values of the investigated parameters, especially the mitotic index, which showed a significant increase, and the spontaneous micronucleus formation and sperm-head and-tail abnormalities, which showed a significant decrease. However, these effects were dependent on dose and type of extract,The results of interactions between CCl4 and both agrimony extracts confirmed the forthcoming effects of the plant, and the two extracts were significantly effective in modulating the immune suppressive and mutagenic effects of CCl4, although the effects were also subjected to the dose, type of extract and the parameter of evaluation. Histopathological examinations of liver sections of animals treated with CCl4 showed marked degenerative areas and necrosis of hepatocytes. Moreover, significant increased serum levels of liver function enzymes (AST, ALT and ALP) were observed in the treated animals. In contrast, biochemical parameters demonstrated the hepatoprotective effect of agrimony extracts through decreasing or normalizing the serum level of these enzymes and repairing the induced cellular damage of liver. However, a histopathological examination of liver sections was still showing some of the CCl4-induced hepatic histological damages. The results of stage III revealed that the five concentrations of the plant extracts showed anti-tumor properties in a concentration-dependent manner, and the methanol extract recorded better values of PGI than aqueous extract in HeLa and RD cell lines, while, less PGI values were recorded in the MEF cell line.