The Effect of Six Weeks of Aerobic Training on Cytochrome C Level in the Sensory Part of the Spinal Cord in Wistar Rats with Diabetic Neuropathy

Diabetic neuropathy is the most common diabetic complication. In more than 50% of patients with diabetic neuropathy, significant and irreversible nerve damage occurs before diagnosis. Diabetic neuropathy is associated with increased mortality (2). Half of patients with diabetic neuropathy suffer from neuropathic pain. These painful symptoms are usually severe and often lead to depression, anxiety and sleep disorders and reduced quality of life (2). Oxidative stress caused by hyperglycemia causes damage to mitochondria in nerve cells, but its mechanisms are not fully understood (4, 5). Sequential oxidation reactions in the mitochondria cause unpaired electron leakage in the electron transfer chain in the inner mitochondrial membrane and the production of free radicals (4). Mitochondria are primary targets for ROS-induced injury; because they are the main site of ROS formation in diabetic hyperglycemia (4). Therefore, examining the factors affecting the structural and functional components of mitochondria, such as electron transfer chain components, can provide useful information for the goals and programs of treatment and prevention of neurological damage caused by diabetic neuropathy. Cytochrome C is a small hemoprotein in the inner membrane of the mitochondria; this protein is highly soluble in water and is a key component of the electron transfer chain (6). Velayutham et al. In a study stated that cytochrome C and Fe3 play a significant role in the harmful effects of ischemia / reperfusion and diabetes due to increased production of superoxide radicals (7). On the other hand, insulin therapy has been shown to modulate neurotrophin-dependent treatment of mitochondrial membranes and the expression of genes associated with metabolite pathways and the mitochondrial electron transport chain (8). Although much research has been done on the effect of exercise on diabetic neuropathy, there is no research specifically examining the effect of exercise on cytochrome C as one of the factors affecting the mitochondrial function of the nervous system in mice with diabetic neuropathy; Therefore, the aim of the present study was to determine the effect of six weeks of aerobic exercise on the level of cytochrome C in the sensory part of the spinal cord in rats with diabetic neuropathy.

In the present experimental study, 12 male Wistar rats were divided into 4 groups: healthy exercise, control exercise, control diabetes and control diabetes. In order to induce diabetes, half of the rats became intraperitoneally diabetic by injecting a single dose of streptozotocin (STZ) (55 mg / kg body weight). Forty-eight hours after STZ injection, hyperglycemic rats with serum glucose above 300 mg / dL were considered diabetic. To confirm the neuropathy of rats, the thermal hyperalgesia test was used with a tail-flick device (20). After confirmation of neuropathy, the animals were placed in the diabetic neuropathy group. The training program included 6 weeks of running training on the treadmill in 5 sessions per week. After dissection of the animals, the dorsal part of the spinal cord was analyzed as sensory neurons. Immunohistochemistry was used to measure the amount of cytochrome C protein. One-way analysis of variance and Tukey's post hoc test were used for statistical analysis. Statistical analysis was performed using SPSS software version 26 at a significant level of P ≤ 0.05.

The results showed that six weeks of aerobic training reduced blood glucose in the diabetic group compared to the control diabetic group (P = 0.002), but there was not observed make a significant difference in the weight of rats. The results also showed that after the training period, the level of cytochrome C protein in the healthy exercise group significantly increased compared to the healthy control group (P = 0.041) and in the diabetic group the exercise significantly decreased compared to the control diabetes group (P < 0.001).

The results of the present study showed that induction of diabetes increased the level of cytochrome C in spinal tissue in diabetic neuropathic rats compared to healthy groups. Previous research has shown that diabetic neuropathy is associated with mitochondrial degradation due to stress due to hyperglycemia and oxidative stress due to diabetes (21, 22); in the present study, an increase in cytochrome C in diabetic neuropathy groups was positively correlated with an increase in glucose levels. In the study of the effect of exercise on cytochrome C levels, the results of the present study showed that after six weeks of aerobic exercise, a significant increase in cytochrome C protein content of spinal cord sensory tissue was observed in healthy exercise rats compared to the healthy control group. While in the diabetic exercise group, the results showed a significant decrease in cytochrome C levels compared to the diabetic control group, which showed the difference in the effect of exercise on changes in cytochrome C of sensory neurons in mice with diabetic neuropathy compared to healthy mice. Scientific research has shown that exercise is one of the effective factors in reducing hyperglycemia due to diabetes (11, 25), on the other hand, by controlling blood sugar, the level of inflammatory factors and oxidative stress in these patients is reduced (26, 27); It can be said that aerobic exercise with glycemic control reduces mitochondrial stress and thus reduces cytochrome C as one of the mechanisms of apoptosis signaling (23, 24), in the cells of the spinal cord, Which can prevent the complications of diabetic neuropathy as well as neuropathic pain. The results of the present study also showed that six weeks of aerobic exercise increased the amount of cytochrome C protein in the spinal cord of healthy rats, this was consistent with the results of Yoo et al. (28) and Bashiri and Pourrazi (29) who studied the effect of exercise on cytochrome C in the myocardial of healthy rats. It can be said that the increase in cytochrome C levels in healthy rats is one of the adaptations associated with aerobic exercise and indicates an increase in mitochondrial capacity in ATP production. It can be said that changes in cytochrome C in spinal sensory tissue are different in adapting to six weeks of training in healthy rats with neuropathy, and the increase in cytochrome C in healthy rats is associated with increased mitochondrial capacity in energy production. While the reduction of cytochrome C in rats with diabetic neuropathy is associated with a reduction in hyperglycemia and indicates the protective role of exercise on nerve tissue and reducing neuropathic damage caused by diabetes. According to the results, in addition to controlling hyperglycemia in diabetic patients, aerobic exercise can be used as one of the non-pharmacological and useful treatment methods in reducing the complications of diabetic neuropathy.