Diabetes is characterized by the reduction in the glucose uptake into muscle and adipose tissue, which leads to chronic extracellular hyperglycemia and many other complications. This life- threatening disease is mostly associated with high risk of atherosclerosis, retinopathy, cataract formation, nephropathy, peripheral nerve damage, and liver apoptosis [1] [2] [3] [4]. Oxidative stress is a well-recognized factor which involves in pathogenesis of diabetes and its complications. Moreover, progression of microvascular and cardiovascular complications of diabetes are well associate with oxidative stress and reactive oxygen species (ROS) [5]. In other words; increase in ROS is a proven mechanism during hyperglycemia which is followed by activation of pathways resulting in several damages in different organs [6]. Oxidation may also be associated with cataract induced by hyperglycemia [7]. In hyperglycemia; cell death is the consequence of NFκB activation with impairment in the regulation of ROS. Based on the type of cell involved, diabetic patients will face complications in various organs such as liver, kidneys, lung and eyes [8] [9].

There are a number of medications used to treat people with diabetes, but synthetic drugs have

always shown adverse reactions and other undesirable side effects. On the other hand, common treatments for diabetes are mostly designed to control the blood glucose and do not show desirable effects on the complications [10]. In this regard, replacing effective natural therapies instead of chemical and pharmaceutical treatments are useful to control the development of diabetes. Recently, herbal medicines with high antioxidant activities have attracted considerable attention as a potential option to prevent and protect oxidative damage caused by free radical species [11]. Crocus sativus L. commonly known as saffron form Iridaceae family is a popular specie in Iran [12]. Saffron has been recognized for having interesting clinical properties such as being anticonvulsant, antidepressant, anti-inflammatory, and antitumor [13] [14] [15] [16]. The pharmacological activities of saffron are attributed to many of its active constituents including safranal, picrocrocin, crocetin, and crocins [17] [18]. Accumulating evidence has suggested the possibility of saffron being used as a candidate for the treatment of diabetes. Although, hypoglycemic effects of this plant have been studied in diabetic rats, the effect of saffron on different organs remains obscure [19].

Therefore, the present research was undertaken to evaluate the effects of saffron on alteration of oxidative parameters including superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), and malondialdehyde (MDA) on the lens of diabetic rats.

Body weight and FBS of rats are shown in Figure 1 . The baseline weight of the rats at the beginning of the study was similar in all groups. The results showed that at the end of the study, the mean body weight of diabetic adult rats had a 10% reduction compared to the beginning of the study. On the other hand, SAE not only prevented weight loss in diabetic rats but also caused a significant increase (2%) in the body weight of diabetic rats Figure 1A .

Measurement of FBS levels at the end of the study showed a significant (P<0.05) increase in diabetic rats. However, diabetic rats treated with SAE showed a significant (P<0.05) decrease in the level of FBS compared to the diabetic control group Figure 1B .

As shown in Figure 2A and Figure 2B , the activity of SOD and GPx were significantly (P<0.05) reduced in the lens tissues of diabetic rats. However, when treated with SAE the diabetic rats showed a significant increase (P<0.05) in the level of antioxidant enzymes (SOD and CAT), as compared to the diabetic control rats.

Figure 2C indicates that STZ causes a reduction in CAT activity. Although the reduction in CAT activity was not significant, SAE administration was able to increase the activity of this enzyme in the lens of diabetic rats, in comparison to other groups (P<0.05).

Figure 2D shows the measured MDA level in the lens of experimental animals. In diabetic control rats, the level of MDA showed a significant increase (P<0.05) compared to the normal control group. However, the level of MDA in saffron treated rats significantly decreased, in comparison with the diabetic control group.

Diabetes is a serious and growing disorder which is associated with severe acute and chronic complications. Different sources of evidence indicate that oxidative stress has been implicated in diabetic patients [23] [24] [25]. In other words, ROS have been defined as an autocatalytic mechanism that can lead to apoptosis [26]. Therefore, regulation of oxidative stress could be a promising therapeutic strategy used to prevent or delay diabetes complications.

After STZ induction, body weight of diabetic rats begins to decrease [26] [27] [28]. In diabetes, both muscle atrophy and loss of weight are the consequence of the body impairment to use the excessive glucose produced during gluconeogenesis [29]. Results of this study demonstrated that SAE is able to prevent weight loss. Moreover, the hypoglycemic effects of saffron which reverses gluconeogenesis is another underlying mechanism involved in the prevention of weight loss in diabetes [30]. As Figure Figure 2 shows, in the diabetic control group, STZ causes a constant high blood glucose which proves that beta cells of pancreas are destroyed. However, diabetic rats treated with SAE showed a significant reduction in the level of blood glucose. Previous studies have also reported the hypoglycemic effects of saffron along with reversing weight loss. In other words; Saffron ethanolic and aqueous extract have the potential to regulates of insulin, improve blood glucose and weight loss in diabetic rats [31] [10] [32] [19]. In addition, in a study done by Samarghandian et al. (2014), saffron was able to decrease hyperglycemia and ameliorate the status of oxidative stress in rats with diabetic encephalopathy [33]. In this regard, the suggested hypoglycemic mechanism of saffron includes a reduction in insulin resistance, prevention of intestinal glucose absorption, and stimulation of glucose uptake in peripheral tissues [34] [35] [36]. Over-production of ROS, reduction in the level of GSH and vitamin E, and the defects in the activity of antioxidant enzymes, such as SOD and GPx, has a major contribution in the pathogenies of diabetes. There are numerous recent publications which describe the effect of oxidative stress on the damage to the lens fibers in diabetes and cataract is a major cause of blindness in this life-threatening disease. Results of the present experiment indicates that the level of SOD and GPx are significantly (P<0.05) decreased in the lens of diabetic control group in comparison with the normal control group. However, the reduction of CAT in the diabetic control group was not significant compared to the normal control group. Moreover, the level of MDA showed significant (P<0.05) increase in the lens of diabetic rats in comparison with the normal control group. In addition to the results obtained from the present study, there are several reports indicating the alteration of antioxidant enzymes and lipid peroxidation in diabetes. Fecondo et al. (1983) measured the activities of the protective enzymes, SOD, CAT, and GPx in the cortical and nuclear sections of the human lens as well as in calf, rabbit, and rat lens. They have reported a quick decrease in both SOD and GPx in the nuclear region of the lens at the onset of nuclear cataract; however, no changes were reported in the activity of CAT. Based on the findings of Fecondo et al. (1983) inactivation of antioxidant enzymes results in an increase in the level of hydrogen peroxide and superoxide anion and these are the molecules causing oxidative modification in lens proteins observed in nuclear cataracts. In another study, Cekic et al. (1999) reported that in the lens of alloxan-induced diabetic rats the activities of SOD and GPx decreased and the level of CAT and xanthine oxidase increased. Moreover, Balog et al.’s (2001) research showed that the lens of alloxan-induced diabetic rats had lower GPx activity than normal rats. They also represented that lipid peroxidation increased in alloxan-induced diabetic rats. Another study published in 2013 mentioned that 4 weeks after the induction of diabetes with STZ, the level of MDA in the lens tissue of diabetic rats increased while the activity of SOD and GPX decreased [37]. Thiraphatthanavong et al. (2014) studied the effect of combined extract of purple waxy corn and ginger on cataractogenesis and retinopathy in STZ diabetic rats [38]. Findings of their study showed an elevated MDA level and a reduction of SOD and GPx in the lens of diabetic rats. Since oxidative stress is an important factor in the pathogenesis of diabetic complications, agents with high antioxidant capacity are considered as an alternative therapeutic method in this life-treating disease. Saffron is a plant which exhibited significant radical scavenging and antioxidant activity [39]. In the present investigation, SAE administration significantly (P<0.05) increased the level of antioxidant enzymes CAT, GPx, and SOD compared to the diabetic control group. The saffron treated group also showed a significant decrease in the level of MDA in comparison with the diabetic control group. Consistent with the present study, different agents with antioxidant activity have been examined to see if they could prevent cataractogenesis in diabetes. A study on Venoruton, a flavonoid with high antioxidant capacity, showed its protective effect on rat lens with diabetic cataract [40]. Quercetin a flavonoid, have shown potential prevention in cataractogenesis in the rat lens organ [41]. Moreover, the protective effect of traditional antioxidants such as vitamin C, vitamin E, and superoxide mimics has shown promising effect against cataract, possibly through protection of membrane lipids against peroxidation [42].

The results of the present study indicate that the protective effects of saffron on diabetes complications due to its effects on antioxidant enzymes but also due to its inhibition of lipid peroxidation. This study suggests that saffron aqueous extract might be a promising candidate for therapy of chemically induced diabetes and its associated complications.

CAT: Catalase; FBG: Fasting Blood Glucose; GPx: Glutathione peroxidase; MDA: Malondialdehyde; NFκB: Nuclear Factor κB; SAE: Saffron Aqueous Extract; D: Superoxide dismutase; STZ: Streptozotocin

Envisaged and designed the experiments: M Ashrafi, N Kazemipour and S Nazifi; Extract preparation, animal study and measurement of biochemical parameters: M Ashrafi, S Talebanzadeh; Data analysis: M Ashrafi, H Erjaee; Manuscript writing: S Nazifi, H Erjaee. All authors reviewed, commented and approved the final manuscript.

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The authors declare that no competing interests exist.