The objective of the current study was to examine the efficiency of using either flax/pumpkin or purslane/pumpkin seed mixture (components of ω-3 and ω-6) on hyperlipidemia, kidney function and as immunomodulators in rats fed high cholesterol diets. Previous study by Lecumberri et al.[26
] reported that rats fed high cholesterol diet showed significant increase in body weight and other organ weights, thus leads to secondary complications clinically. In our study, body and relative organs weight gain of hypercholesterolemic rats were decreased significantly upon treatment with flax/pumpkin or purslane/pumpkin seed mixture. The hypolipidemic and antiatherogenic effects of flax/pumpkin or purslane/pumpkin seed may be responsible for the beneficial action of these seeds on body weight gain and liver weights. Purslane/pumpkin seed mixture seems to exert a protective effect against overweight. However the pumpkin alone in each mixture may be responsible for reduction in body weight. A study by Hyounjeong et al.[27
] screened various plant sources for their antiobesity activity and demonstrated that pumpkin has strong antiobesity effects in a high fat diet-induced obesity animal model. This mainly due to its effect on synthesis and degradation of lipid products in the body, also it was considered as metabolic regulator of lipogenic and lipolytic pathways and altimately as antiobesity agent.
Cholesterol- enriched diet resulted in a significant increase in total cholesterol, total lipids, phospholipids and triacylglycerol in plasma and liver this accompanied by increased serum LDL-C level, with decreased circulating HDL-C, thus providing a model for dietary hyperlipidemia[28
]. The increase of lipid parameters has been shown to be a strong risk factor for coronary heart diseases in many populations .The high cholesterol level in liver and plasma may be due to increased uptake of exogenous cholesterol and subsequent deposition and decreased cholesterol catabolism as evidenced by a reduction in bile acid production and turnover of bile acids. The metabolism of free and ester cholesterol are impaired in liver, spleen and thymus tissue and the rate of turnover was specifically decreased in all tissues of hyperlipidemic rats. Lipid deposition is a major clinical complication of hyperlipidemia[29
] which is consistent with the present study.
Our results indicated that both flax/pumpkin and purslane/pumpkin seed mixture rich in polyunsaturated fatty acids had a strong hypolipidemic, hypotriglyceridemic and hypocholesterolemic effects in plasma and liver of rats with a reduction of plasma LDL-C levels and an increase in HDL-C levels. Furthermore, the atherogenic index markedly decreased due to significant reduction in LDL/HDL ratio in both groups fed hypercholesterolemic diet supplemented with either flax/pumpkin or purslane/pumpkin seed mixture. Results are agreed with Feoli et al.[30
] who stated that the increase in HDL-C or HTR ratio is one of the most important criteria of anti-hypercholesterolemic agent. The decrease of plasma cholesterol by administration of flaxseed was ascribed to the decrease of both free and esterified cholesterol.
Lignans, fibre and vegetable proteins present in the flaxseed could play major roles in reducing serum cholesterol in animal models and/or in humans[31
]. Therefore, the effect of flaxseed in decreasing serum cholesterol does not seem to be due only to its C18:3 content, but rather to the synergistic action of its constituents. This was reported by Cintra et al.[32
] who found lower serum lipid levels in normocholesterolemic and hypercholesterolemic subjects who were fed flaxseed oil. Fibers are reported to decrease plasma LDL-C levels by interrupting the cholesterol and bile acid absorption and increasing LDL receptor activity. The decline in hepatic cholesterol levels in flax/pumpkin hypercholesterolemic group indicated the possible influence of relatively higher fiber content of seed mixture. In fact, dietary fiber are known to interfere with cholesterol absorption and enterohepatic bile circulation and resulted in depletion of hepatic cholesterol pools[33
]. In addition, diets rich in fibers are known to reduce triacylglycerol levels by inhibition of hepatic lipogenesis[34
]. Moreover, numerous studies have demonstrated that high levels of HDL-C are associated with a lower incidence of cardiovascular diseases . The increase in HDL-C observed in our study, might be due to stimulation of pre-β HDL-C and reverse cholesterol transport as demonstrated by previous study[35
]. High HDL-C levels could potential contribute to its anti-atherogenic properties, including its capacity to inhibit LDL oxidation and protect endothelial cells from the cytotoxic effects of oxidized LDL[36
]. The anti-atherogenic effect of flax and pumpkin seed mixture found in our study might be due to the presence of polyunsaturated fatty acids, phytosterols, tocopherols and β-carotene[37
]. The major total fatty acids present in seed mixture are unsaturated fatty acids such as oleic acid,linolenic acid and linolenic acid , which play a crucial role in reducing blood cholesterol in human and rats Movahedian et al.[38
] showed that addition of purslane leaf extract to the cholesterol-enriched diet improved the hypercholesterolemia induced by a high-cholesterol diet in rabbits. The results showed that upon administration of hydroalcoholic extract of purslane for 12 weeks significantly decreased the serum total cholesterol, LDL-C and VLDL-C at doses of 200, 400 and 800 mg kg-1
bw in comparison with the hypercholesterolemic group. But serum HDL-C was elevated insignificantly in hypercholesterolemic rabbits treated with the above doses of purslane extract. Purslane treated animals also showed a decrease in the atherogenic index with respect of hypercholesterolemic groups, which is generally believed to be beneficial since the HDL level inversely correlated with coronary heart disease and reduction in this ratio is considered as an anti atherosclerotic factor.
The kidney functioning capacity was assessed in this study by measuring the levels of electrolytes, creatinine and urea in the serum of the animals. The presence of significant effect of the cholesterol-enriched diet on the serum concentrations of sodium, potassium and phosphorus of the animals suggest that, the abnormal functioning of the organ in relation to these electrolytes were affected. Kidneys remove metabolic wastes such as urea, uric acid, creatinine and ions, so optimum chemical composition of body fluids is maintained. The concentrations of the metabolites increase in blood during renal diseases or renal damage may due to high activities of xanthine oxidase, lipid peroxidation, and increased triacylglycerol and cholesterol levels[39
Creatinine, synthesized in the liver, passes into the circulation where it is taken up almost entirely by the skeletal muscles. Its retention in the blood is an evidence of kidney impairment[40
]. Therefore, the reduced levels of creatinine in serum may imply that, the seed mixture had interfered with creatinine metabolism and its eventual excretion from the blood. Urea is the main product of protein catabolism. The increase in serum urea level in hypercholesterolemic control group indicates impairment in the normal kidney function of the animal, as the mechanism of removing it from the blood might have been affected. It may also be an indication of dysfunction at the glomerular and tubular levels of the kidney, it is well known that, many biochemical and histopathological findings confirmed renal damage in hypercholesterolemia conditions[41
]. Flax seed is the richest natural source of plant lignans, which are platelet-activating factor (PAF)-receptor antagonists. Platelet-activating factor plays a key role in the mediation of inflammation, mitogenesis, and alteration of glomerular permselectivity. Thus, flaxseed could potentially inhibit various mechanisms associated with the progression of renal diseases[42
]. A study by Dkhill et al.[43
] showed that purslane administration at 1.5mg/kg purslane aqueous juice for 12 days caused significant decrease in urea and creatinine respectively. Schaefer et al.[44
] have recommended many medicinal plants used already in traditional medicine, experimental and clinical, and nephroprotective effects among them are flax and pumpkin seeds mixture rich in PUFAs and antioxidant compounds in animals.
The current study clearly showed that the levels of IgG and IgM were significantly increased (P<0.05) in hypercholesterolemic rats as compared with healthy control. It is clear that consuming seed mixtures either F/P or P/P had positive impacts to the immunity status of hypercholesterolemic rats by significant reduction in rat serum immunoglobulines to bring them near the normal levels. The capacity of dietary n-3 polyunsaturated fatty acids (PUFAs) suppress inflammation-associated processes, has made them attractive candidates for both the prevention and amelioration of a variety of organ-specific and systemic diseases. n-3 PUFAs suppress proinflammatory cytokine production, lymphocyte proliferation, cytotoxic T cell activity, natural killer cell activity, macrophage-mediated cytotoxicity, neutrophil/monocyte chemotaxis, and antigen presentation[45
]. Evidence that these cellular effects indeed impact immune function in vivo is reflected in n-3 PUFA attenuation of mediator production, leukocyte homing, delayed-type hypersensitivity and acute inflammatory responses in experimental animals in which human inflammation and autoimmune diseases are modeled. n-3 PUFAs appear to mediate these effects via both eicosanoid-dependent and eicosanoid-independent pathways. Taken together, these anti-inflammatory and immunomodu-lating activities have led to the evaluation and application of n-3 PUFAs for prevention and treatment of inflammatory and autoimmune diseases, of particular interest here are those studies that have focused on the kidney[46
]. Results by Sun et al.[47
] showed that increasing the supply of linolenic acid to the small intestine of animal models linearly modulate the immunity. On the other hand, it is reported that, 100 g of fresh Purslane leaves (about 1 cup) contain 300–400 mg of alpha-linolenic acid, is important in preventing heart attacks and strengthening the immune system.