4. Discussion
Diabetes and the associated comorbidities such as overweight and cardiac diseases are lead causes of human morbidity and mortality. Overweight is a metabolic disorder due to the accumulation of excess dietary calories into visceral fat and the release of high concentrations of free fatty acids into various organs. It represents a state of chronic oxidative stress and low-grade inflammation that may progress to hyperglycemia and type 2 diabetes (Hotamisligil, 2006). The status of the disease diabetes produces vital changes at the intracellular levels (Rifkin and Porte, 1990; Parinandi et al., 1990). An increasing body of evidence has reported that there is a link between vitamin A and diabetes. Moreover, reported the increase in the oxidative stress status in diabetes mellitus (Halliwell and Gutteridge, 2015, Low and Nickander, 1991, Singh et al., 2001).  Few studies used the retinoic acid and suggested that it can have a reno-protective effect in the early stages of diabetic nephropathy through a unique anti‐inflammatory pattern (Cha et al., 2007).
In the present study, streptozotocin induced diabetic mice demonstrated the expected increases in body weight, plasma glucose and total lipids, consisted with the metabolic abnormalities in diabetes patients (Manzato et al., 1993; Merzouk et al., 2000). In the current experiment, treatment with Vitamin A had no significant effects on neither fasting blood glucose nor insulin levels. Nonetheless, previous clinical study proved that Vitamin A caused a transient impairment of glucose metabolism and insulin sensitivity, as indicated by the increases in insulin levels and HOMA-IR indices (Corbetta et al., 2006). Interestingly, Vitamin A in our study showed a significant improvement in body weight, fat mass, and lipid profile. Three weeks after the beginning of the experiment, the untreated diabetic mice significantly gained more weight compared to either the normal control or vitamin A treated group. Subsequently, the rate of weight and fat gain also increased in the diabetic untreated mice while decreased significantly in the treated group especially after week 12 (at the end of experiment). Multiple studies have shown that adipose tissue act on the metabolism and homeostasis of vitamin A. Our results was different than the fact that chronic vitamin A enriched diet feeding can significantly impacted the obesity development both in young and adult obese rats of WNIN/Ob strain, possibly through thermogenic and glucocorticoid pathways without eliciting any toxic symptoms (Jeyakumar and  Vajreswari, 2015). The exact mechanism of this is unclear and need further investigations. In accordance with our results, previous research indicated that retinoids can reduce body weight, subcutaneous and visceral fat of mice fed high fat diet (Geng et al., 2017). A possible mechanism by which vitamin A reduces fat and body mass, as well as increases energy expenditure, is through the activation of uncoupling protein 1 located in the inner mitochondrial membrane of adipocytes (Jeyakumar et al., 2008). Nevertheless, Ilić and his colleagues,2020, stated that vitamin A caused increase in body weight in rats on high fat diet while high dose of vitamin A lowered body weight in rats fed on standard diet. The discrepancy between their results and our results may be due to the difference in dose and duration of treatment as well as the species and gender of animals.
To evaluate the effects of vitamin A on the antioxidant enzymes activities in diabetic-induced mice, the levels of SOD,  CAT,  and GPO in the liver tissues of all groups were estimated. These enzymes act as one unit with each other to decrease any abnormalities of the reactive oxygen species (ROS), that can lead to oxidative stress (Chelikani et al., 2004, Finkel and Holbrook, 2000, Fridovich, 1986, Jang et al., 2000).
In the present research, we demonstrated a significant decrease in the activity of SOD in diabetic induced mice compared with control normal group. Meanwhile, vitamin A showed a significant amelioration in the activity of SOD in the treated diabetic mice compared with untreated diabetic ones. It is well known that second to SOD in defense is the CAT and GPO. (Chelikani et al., 2004, Tiwari et al., 2013). It has been shown that CAT is essential in a pancreatic β-cells protection from oxidative stress. Any deficiency in this activity will lead to ROS elevation, oxidative stress and finally cellular dysfunction that is seen in diabetes mellitus (Góth and Eaton, 2000). GPO works also when the level of oxidative stress is increased, to protects damaging the cells from free radicals (Birben et al., 2012, Tiwari et al., 2013). In our experiment, we demonstrated that the activities of CAT and GPO were significantly decreased in diabetic induced mice compared with those in control normal mice. However, treatment with vitamin A showed a significant increase in their activities compared with untreated diabetic ones. Coinciding with the previous results, other studies proved the hypothesis that oxidative stress in diabetes mellitus can be caused by impaired antioxidant system (Ramakrishna and Jailkhani, 2008, Sindhu et al., 2004). The mechanism of vitamin A on the antioxidant enzymes still understudied. However a possible explanation is that vitamin A can produces protein carriers, such as zinc and copper, that can lead to an increase in their bioavailability inside the cell. These ions can play a role in the activity of multiple enzymes, such as the antioxidant enzymes (Claro da Silva et al., 2016).
To examine lipid peroxidation in our study we measured MDA levels in the serum that serves as a biomarker of lipid damage and oxidative stress (Halliwell and Chirico, 1993). Our results demonstrated a significant elevation in MDA in diabetic induced mice compared with controls. These results are in agreement with previous studies that showed an elevated level of MDA in diabetes mellitus in both human and animal studies (Ramakrishna and Jailkhani, 2008, Sindhu et al., 2004). On the other hand, treatment with vitamin A in diabetic induced mice demonstrated a significant decrease in MDA levels compared with that in untreated diabetic mice. These Results can support the hypothesis that vitamin A can have an antioxidant effect and protection effect on the damage induced by lipid oxidation. Vitamin A’s antioxidant properties have been frequently reported in vivo and in vitro (Palace et al., 1999). Retinol can react with peroxide radicals (ROO •), thereby it interrupts the chain reaction of lipid peroxidation to form hydroperoxides (ROOH). Furthermore, vitamin A is capable of directly reacting with ROS to form a 5,6-epoxide retinoid (Edge et al., 1997).
The present research demonstrated that lipid profile was significantly improved after 12 weeks of treatment with Vitamin A in diabetic induced mice. This amelioration can be explained by the relationship of lipids and the oxidative symptom, however, the definitive role of lipid contribution to the oxidative system has yet to be fully elucidated. In consistent with our results, a clinical study proved that patients with cardiovascular diseases have elucidated Low levels of vitamin A. Moreover, these patients had lower HDL-cholesterol levels indicating the possible beneficial effects of vitamin A on serum lipids as well as cardiovascular diseases (Godala et al., 2017) .
In conclusion, the results of the present study demonstrated some beneficial effects of Vitamin A in streptozotocin-induced diabetic mice. It caused significant improvement in body weight, lipid profile, lipid peroxidation and oxidative stress. These data suggest that Vitamin A supplementation might help in decrease the metabolic complications of diabetes mellitus. However, further studies should be done to examine the appropriate dose of Vitamin A to avoid side effects or toxicity. Also, more clinical research is needed to determine how Vitamin A status affects humans.