Protective Effect of Fennel Oil on Cyclophosphamide Inhibited Spermatogenesis and Induced Oxidative Stress in Albino Rats

Introduction: Cyclophosphamide (CPA) is an anticancer drug .Fennel (Foeniculum vulgare Mill) essential oil is a traditional medicine used against many diseases. Aim. The present work studied the effect of fennel oil against testicular damage and oxidative stress induced by the anticancer drug, cyclophosphamide (CPA) in albino rats. Methods. Animals were divided into 4 groups: group1, control, group2, orally given fennel oil, group3 treated with CPA and group4 treated with CPA and fennel oil. The testes were removed for histological and immune histochemical preparation. Blood was collected and sera were prepared for hormonal and biochemical analysis. Results. The results revealed that CPA caused histological alterations in the testis including decrease in diameter and germinal epithelial height of the seminiferous tubules, degeneration of germ cells, cytoplasmic vacuolation and congestion of blood vessels. Cell proliferation marker was decreased and apoptotic marker caspase-3 was decreased. Biochemical results revealed decrease in the hormones LH and testosterone. Moreover, the serum activity of the antioxidant enzymes, SOD, CAT was decreased and the lipid peroxidation marker, DMA was increased. Treating rats with CPA and fennel oil caused an improvement in the histological structure of the testis. There was an increase in LH ,testosterone,SOD and CAT, while MDA level decreased. Conclusion. It is concluded that administration of fennel oil exhibited protective effects against CPAinduced reproductive toxicity in male rats. The protective effect of fennel oil might be due to induction of antioxidant defense systems by one or more of its constituents. DOI : 10.1302/issn.2576-6694.jbbs-17-1742 Corresponding author: Saber A.Sakr, Department of Zoology, Faculty of Science, Menoufia University, Shebin El-kom, Egypt, E.mail.: sabsak@yahoo.com


Introduction
Cyclophosphamide (CPA) is a nitrogen mustard alkylating agent, and anti-neoplastic drug. It is used to treat Hodgkin's disease, lymphomas, leukemia, granulomatosis, severe rheumatoid arthritis, and lupus erythematosus. It is also used in combination with other drugs to treat breast cancer, leukemia, and ovarian cancer. The drug also has immunosuppressive action when it has used in smaller doses (1). CPA acts by modifying and cross linking purine bases in DNA, thus inhibiting DNA, RNA and protein synthesis and death of rapidly dividing cells. CPA has been found to cause many toxic effects including haemorrhagic cystitis, pulmonary fibrosis, irreversible a zospermia in man, gastrointestinal bleeding and hepatotoxicity (2) Cardiotoxicity is a major problem with people treated with higher dose regimens (3). In addition, CPA was found to affect male reproduction. Wetzel (4) Indicated that men exposed to CPA may develop oligo spermia or azoospermia associated with increased gonadotropin release. Turk et al., (5) (6). This herb is traditionally used as treatment for colic, wind, irritable bowel, kidneys, spleen, liver, lungs, suppressing appetite, breast enlargement, promoting menstruation, improving digestive system, milk flow and increasing urine flow (7). The chief component of fennel, anethole, had anticarcinogenic and anti-inflammatory effects through modulation tumor necrosis factorinduced cellular processes (8) and antimicrobial properities (9). Furthermore, fennel has a bronchodilatory effect (10) as well as immunomodulatory activities by enhancing natural killer cell functions, the effectors of the innate immune response (11).
Fennel extracts was found to increase male fertility and is consider as a novel medicine for treatment of infertility (12,13). In view of these considerations, the present work investigated the effect of fennel oil on cyclophosphamide-induced testicular toxicity in albino rats.

Materials and Methods
Chemical and Plant used:

Statistical Analysis
Data were expressed as mean ± standard deviation (SD). The significance of differences means was evaluated by using independent sample t test. All statistical analysis was performed using SPSS statistical version 16 software package.

Histological Observations
The 2-The Expression of Casepase-3 In control group, casepase-3 was expressed in cytoplasm of few leydige cells as brown color. The cytoplasm of germ cells stained weakly (Fig 7-A).
Animals given fennel oil for six weeks showed expression of casepase-3 in cytoplasm of spermatocytes nearly similar to control group (Fig 7-B).     comparison with CPA group (Fig 9). A significant decrease in CAT,SOD and GSH activities were recorded in sera of rats treated with CPA for 3 and 6 weeks. Animals treated with CPA and fennel oil showed a significant increase in these enzymes (Fig.11-14).

Discussion
In the present study, results revealed that treating rats with CPA induced many histological alteration in the testis and these alterations were more   (12) reported that fennel oil resulted in amelioration in testicular tissue lesions and decrease of apoptosis in rats exposed to tobacco smoke. Fennel treatment was found to improve sperm quality, and spermatogenic cells apoptosis in obese rats (13). The level of testosterone and LH increased. Ibrahim (42) showed that there were significant increases over the control in testosterone level in the serum of rats treated with fennel oil. The author added that the improvement effect of fennel oil on testicular function as indicated by the testosterone may be attributed to the powerful active components of the fennel oil.