1136 Protective Effect of Broccoli and Red Cabbage Against
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Journal of American Science, 2010;6(12) http://www.americanscience.org Protective Effect of Broccoli and Red Cabbage Against Hepatocellular Carcinoma Induced by N- Nitrosodiethylamine in Rats Aml F. M. Morsy*a, Hodaa S. Ibrahima and M. A. Shalabyb a Nutrition and Food Science Department, Faculty of Home Economics, Helwan University, Egypt. b Pharmacology Department Faculty of Veterinary Medicine, Cairo University, Egypt. * [email protected] Abstract: The hepatoprotective effect of broccoli and red cabbage extracts against hepatocellur carcinoma induced by N- Nitrosodiethyamine (NDEA) in male rats were studied. Four groups of rats were used; group (1) was used as a negative control (normal), while rats of the other groups were given NDEA as a single interperoitenial dose with subcutaneous injection of carbon tetrachloride (CCl4) once weekly for six weeks to induce hepatocellular carcinoma. Group (2) was left as a positive control, while groups (3) and (4) were pretreated with broccoli and red cabbage 10%extract, for 12 weeks, respectively. At the end of the experiment, blood samples were taken for biochemical analysis and liver tissues were histopathologically examined. The obtained results revealed that rats with hepatocellur carcinoma (HCC) had significant increase in serum levels of AST, ALT, ALP, total protein, albumin, total and direct bilirubin and malondialdehyide (MDA), as well as significant decrease in reduced glutathione (GSH), glutathione peroxidase (GPX), superoxide dismutase (SOD) and catalase (CAT) enzymes, compared to the normal control group. Liver sections of rats with HCC showed fatty infiltration of hepatocytes, cytomegaly with karyomegaly as well as vesicular active nuclei and presence of more than one nucleolus in some hepatocytes. Oral administration of broccoli and red cabbage extracts caused significant reduction in serum levels of AST, ALT, ALP, total protein, albumin, total and direct bilirubin as well as MDA and produced significant increase in GSH, GPX, SOD and CAT, compared to the positive group. Liver of these rats revealed only slight hydropic degeneration of hepatocytes, while other sections showed apparent normal hepatocytes. This study concluded that broccoli and red cabbage have a protective effect against hepatocellular carcinoma in rats, therefore this study recommends increased dietary intake of broccoli and red cabbage may be beneficial for patients with liver cancer as a preventative measures. [Aml F. M. Morsy, Hodaa S. Ibrahim and M. A. Shalaby. Protective Effect of Broccoli and Red Cabbage Against Hepatocellular Carcinoma Induced by N- Nitrosodiethylamine in Rats. Journal of American Science 2010;6(12):1136-1144]. (ISSN: 1545-1003). http://www.americanscience.org. Keywords: Broccoli; Red cabbage; Liver cancer; Biochemistry; Histopathology; Rat. 1. Introduction: Numerous compounds in the human diet have Hepatocellular carcinoma (HCC) is the most chemoprotective properties against chemical common form of liver cancer in adults, which carcinogens (Wattenberg, 1990). Intake of a diet rich in accounts for about 75% of primary liver cancers. It is fruits and vegetables is associated with a lowering the 5th most common cancer in worldwide and risk of certain types of cancer (Steinmetz and Potter, represents 83% of all cases (Ferlay et al., 2001). In 1991). Green leafy vegetables of all varieties and Egypt, HCC is frequent accounting for 13% of all cruciferous plants such as cabbage, Brussels sprouts, cancer types (Inas, 2005). Liver cancers have cauliflower and broccoli are rich in anti-carcinogens different growth patterns; the first type begins as a (Block et al., 1992; Wargovich, 1999). Cruciferous single tumor that grows larger in hepatic tissue. The vegetables contain a number of bioactive components second type is spread through the liver almost from such as folate, vitamin C, tocopherols, carotenoids, the beginning and is not confined to a single tumor. flavonoids and polyphenols (Price et al., 1998; This is seen most often in people with liver cirrhosis. In Kurilich et al., 1999). the third type, the cancer develops as nodules in Broccoli is a plant of family Brassicaceae several parts of the liver (Strauss, 1995). (formerly Cruciferae) which has large green flower Risk factors for HCC include hepatitis B virus heads (Murray and Lara, 2005). Recently, Elizabeth (HBV), hepatitis C virus (HCV) and aflatoxins are and Marcela, (2009) suggested that broccoli can assumed to play an important role in the high decrease the risk for incidence of cancer. It contains incidence of HCC in Egypt. HBV vaccination of many bioactive, including vitamins C and E, quercetin children and high-risk population must be the priority and kaempferol glycosides. in reducing the incidence of HCC. Measures to reduce Red Cabbage (Brassica oleracea var. capitata f. food spoilage by fungi and the associated dietary rubra) is a type of cabbage, also known as Red Kraut exposure to aflatoxins are a desirable public health or Blue Kraut after preparation (Michaelis et al., goal (Wild and Hall, 2000). http://www.americanscience.org 1136 [email protected] Journal of American Science, 2010; 6(12) http://www.americanscience.org 2008). The breakdown products of glucosinolates, 60%), temperature (20-25°C) and light (12-h light: 12- which present in it such as isothiocyanates are h dark cycle) for one week before starting the considered responsible for the chemopreventive experimental for acclimatization. Rats were divided properties of red cabbage (Lynn et al., 2006). into four groups of ten animals each as follows: Group (1): Served as a control negative (normal rats) The aim of the present study was to investigate the and fed on basal diet for 12 weeks. hepatoprotective effect of broccoli and red cabbage Group (2): Kept as a control positive (with HCC) and extracts against hepatocellur carcinoma induced by N - fed on basal diet for 12 weeks. Nitrosodiethylamine in male rats. Group (3): Fed on the basal diet and given orally 10% broccoli extract using stomach tube 2. Materials and methods: for 12 weeks. Group (4): Fed on the basal diet and given red 2.1 Material: cabbage 10% extract orally by stomach 2.1.1 Rats and Diet: tube for 12 weeks. Male albino rats of Sprague Dawley strain weighing In first six weeks of experimental period, animals of 175±5g were used. Their age between 14-16 weeks groups (2), (3) and (4) were given a single old and were purchased from the Laboratory Animal intraperitoneal dose of NDEA (200 mg/kg b.wt.) Colony, Ministry of Health and Population, Helwan, followed by carbon tetrachloride (CCl4) tetrachloride Egypt. Basal diet constituents were obtained from El- (CCl4) given subcutaneously once weekly in a dose Gomhorya Company, Cairo, Egypt. of 200 mg/kg b.wt. during the other 6 weeks for induction of HCC as described by Sundaresan and 2.1. 2 Chemicals: Subramanian (2003). At the end of the experimental N-Nitrosodiethyamine (NDEA) was purchased from period, blood samples were collected from the portal Sigma Chemical Company, USA. Carbon vein into dry clean centrifuge tubes for serum tetrachloride (CCl4) was obtained from El-Gomhorya º separation. Serum samples were frozen at -10 C until Company, Cairo, Egypt. Biochemical kits for serum chemical analysis. Liver of sacrificed rats were kept analysis were purchased from the Gamma Trade in 10% formalin solution till processed for Company for Pharmaceutical and Chemicals, Dokki, histopathological examination. Egypt. 2. 1.3 Plants: 2.2.4 Biochemical analysis: Broccoli and red cabbage were purchased from the 2. 2.4.1 Determination of liver functions: local market, Cairo, Egypt. The plants were Serum asprtate aminotransferase (AST) and alanine aminotransferase (ALT) activities were determined authenticated in the Botany Department, Faculty of according to the method described by Kaplan, (1984). Agriculture, Cairo University. Serum alkaline phosphatase (ALP) was 2. 2 Methods: calorimetrically determined according to the method 2.2.1 Preparation of plant extracts:- described by Roy (1970). Serum total protein Broccoli and red cabbage vegetables were cleaned, concentration was calorimetrically determined air dried and grinded into a fine powder. The according to the method described by Koller, (1984). powdered plants were extracted with 90% ethyl Serum concentrations of albumin, total and direct alcohol using Soxhlet apparatus and concentrated at bilirubin were determined as described by Kaplan, low temperature(50Ċ) using a Rotary evaporator (1984). apparatus (manufactured in Basil, Switzerland) according to the method described by Mohamed, 2.2.4.2 Determination of malondialdehyde and (2002). Each dried ethanol extract was dissolved reduced glutathione: separately in a mixture of carboxy methylcellulose Serum malondialdehyde (MDA) was determined by and few drops of Tween 80 as a suspending agent to the method of Draper and Hadly, (1990). Reduced obtain 10% concentration liquid extract. glutathione concentration (GSH) in was measured by the method described by Beutler and Kelly, (1963). 2. 2.2 Preparation of basal diet: 2.2.4.3Determination of antioxidant enzymes: The basal diet (AIN-93M) was prepared according The serum levels of glutathione peroxidase (GPX), to Reeves et al., (1993). Diet was formulated to meet superoxide dismutase (SOD) and catalase (CAT) were the recommended nutrients levels for rats. determined by autoanalyzer (Roche-Hitachi, Japan) according to the methods described by Hissin and Hilf, 2. 2.3 Experimental Design: (1976);