الفهرس 
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Abstract
This research was designed to examine the effects of intermittent fasting (IF) as an alternative to or in combination with metformin, an antihyperglycemic drug for the treatment of type 2 diabetes, in managing blood sugar levels and preventing the development of diabetic cardiomyopathy, one of the most serious complications of diabetes, as well as its potential mechanisms of action.
The present study was carried out on 50 adult male rats weighing (200-300gm) in a trial to assess the effects of IF on high fat diet + STZ induced type II diabetic rats. The rats were housed at room temperature with normal standard lighting for two weeks after arrival from the supplier for acclimatization. Up to the start of the trial, they were given a typical diet of commercial rat food and tap water on demand. The next five groups, each with 10 rats, were created from them.
1- Control Group.
Whereby each rat receives only a car (0.01 M citrate buffer, pH 4.5).
2- Diabetic Group
In this experiment, rats were given a high-fat diet (HFD) for two weeks to cause T2DM, which was then followed by an intraperitoneal injection of streptozotocin at a dosage of 35 mg/kg dissolved in freshly made 0.01 M citrate buffer at pH 4. A rat tail blood sample was used to assess the blood glucose three days after injection. A diabetic rat was one whose blood glucose level was below 200 mg/dl.
3- Diabetes + Metformin Group:
Each rat in this study underwent HFD for two weeks before receiving the same intraperitoneal injection of streptozotocin as the control group. Following seven days of streptozotocin injections, each rat got metformin orally for 28 days at a rate of 200 mg/kg/day.
4- Diabetes + Intermittent fasting Group:
Each rat in this study underwent HFD for two weeks before receiving the same intraperitoneal injection of streptozotocin as the control group. Each rat was then starved of food for 16 hours each day for 28 days after receiving a streptozotocin injection for 7 days.
5- Diabetes + Intermittent fasting + Metformin Group:
Each rat in this group had a 2-week high-fat diet (HFD) before receiving the same intraperitoneal injection of streptozotocin as the prior group. Then, after seven days of streptozotocin injections, each rat got metformin at a dosage of 200 mg/kg/day for 28 days, along with 16 hours per day of food deprivation.
Weekly body weight measurements and daily food consumption measurements were taken throughout the research period.
After receiving therapy for four weeks, all groups had an overnight fast followed by decapitation. After blood samples were drawn from the jugular vein, they were allowed to clot at room temperature before being spun for 15 minutes at 3000 rpm in a cooling centrifuge (Hettich centrifuge). The serum layer was then removed and placed into designated Eppendorf tubes, where it was kept until the test at - 20 °C.
Biochemical tests were performed on the following variables in stored serum samples from fasting animals in the research groups:
serum sugar.
Blood insulin
Total cholesterol is included in the serum lipid profile (TC).
Triglycerides (TG).
HDL, high-density lipoprotein (HDL).
Lipids with a low density (LDL).
Lactate dehydrogenase in serum (LDH).
Total creatine kinase in serum (CK).
isoenzyme of serum creatine kinase MB (CK-MB)
Malondialdehyde in serum (MDA).
Antioxidant levels in serum (TAC).
In addition, The left ventricular cardiac samples were promptly fixed in a buffered 10% formalin solution. After that, the specimens were prepared to produce paraffin blocks. Hematoxylin and eosin (H&E) and Masson’s trichrome stains were used to stain serial (5-7 m) slices of heart tissue to confirm the induction of diabetes.
Also, immunohistochemical staining was performed by using the avidin biotin‐ peroxidase technique for measurement of
Beclin-1 (autophagy regulator)
TNF alpha
BCL2
Caspase 3
Collagen
The results of the present study showed that:
High fat diet feeding for 2 weeks followed by low dose STZ (HFD+STZ) resulted in a successful animal model of type II diabetes characterized by:
Significantly elevated blood sugar levels (>200 mg%) coupled with insulin resistance and insulin insufficiency (evidenced by significantly higher HOMA-IR than control group). In the context of hyperphagia, body weight fell as a consequence of poor glycemic control and the loss of insulin’s anabolic action.
This mouse also exhibited hyperlipidemia, as seen by considerably higher levels of total cholesterol, triglycerides, and LDL. While compared to the control group, HDL levels in blood were lower.
Compared to the control group, there were higher levels of CK, CK-MB, and LDH (markers of cardiac damage), as well as higher levels of MDA (a marker of lipid peroxidation), and lower levels of TAC.
Increased levels of the apoptotic marker caspase 3 and decreased levels of the anti-apoptotic marker BCL-2 were found in an immunolabelled investigation. Additionally, compared to the control group, there was an increase in the inflammatory marker TNF alpha, along with an increase in the fibrotic marker collagen and a decrease in beclin (a sign of autophagy).
Studying the effects of metformin on the induced diabetic model, it was found that it produced:
Better glycemic control and normal insulin levels may account for the much lower blood glucose level, HOMA-IR to almost the control level, return insulin to normal levels, decreased food intake, and lower body weight compared to the diabetic group.
Significantly reduced the diabetes-induced hyperlipidemia, as seen by a rise in HDL levels to practically control levels and a substantial decrease in TG, TC, and LDL levels (P 0.001).
The blood levels of CK, CK-MB, and LDH considerably decreased but remained significantly higher than in control rats, indicating attenuation of cardiac damage.
Reduction of oxidative stress due to a considerable DROP in blood MDA levels and a rise in serum TAC levels, which attenuated lipid peroxidation.
When compared to the diabetic group but significantly different from control rats, the immunolabelled investigation showed that metformin was also able to boost BCL-2 and reduce caspase 3, demonstrating its anti-apoptotic activity. Additionally, when compared to the diabetic group but also with a significant difference from control rats, it had anti-inflammatory and antifibrotic benefits, which were shown by lowering TNF alpha and collagen, respectively. Furthermore, metformin restored beclin levels to normal, demonstrating its autophagic impact.
Studying the effects of IF on the induced diabetic model, it was found that it produced:
While lowering the rise in blood glucose levels and insulin resistance in diabetic rats over the course of four weeks, IF, the levels were still considerably greater than in the control group. Additionally, IF therapy produced a substantial difference from control in terms of maintaining insulin levels.
Reversing the hyperlipidemia caused by diabetes was shown by a substantial decrease in TG, TC, and LDL levels (P 0.001) and a rise in HDL levels, however the values were remained considerably different from the control group.
reducing the diabetic-induced heart damage as seen by a decline in CK, CK-MB, and LDH serum levels, however they were remained considerably higher than in control rats.
succeeded in reducing oxidative stress by considerably lowering MDA levels and raising serum TAC levels, although with a significant difference from control rats. This attenuated lipid peroxidation.
When compared to the diabetic group, the immunolabeled investigation demonstrated that IF exhibited an anti-apoptotic impact as shown by a rise in BCL-2 and a reduction in caspase 3, although with a significant difference from control rats. Additionally, IF had anti-inflammatory and antifibrotic benefits, which were shown by a reduction in TNF alpha and collagen when compared to the diabetic group, although remained considerably greater than control rats. Additionally, IF was able to raise beclin, showing that it had an autophagic impact, but it was still lower than control.
Studying the combined effect of both metformin and IF for 4 weeks, on the induced diabetic model, it was found that it produced:
The most effective outcomes were achieved because they were able to dramatically reduce the blood glucose level and HOMA-IR to virtually the control level, restore normal insulin levels, reduce food consumption, and lose more body weight than the diabetes group.
significantly reduced the diabetes-induced hyperlipidemia, as seen by a rise in HDL levels to practically control levels and a substantial decrease in TG, TC, and LDL levels (P 0.001). The blood levels of CK, CK-MB, and LDH were also much lower than in control rats, indicating that this combination therapy was successful in reducing cardiac damage. As they drastically reduced MDA levels and restored normal serum TAC levels, they also succeeded in reducing lipid peroxidation.
In a similar vein, the immunolabelled investigation showed that, in comparison to the diabetic group, they were able to boost BCL-2 above normal and reduce caspase 3 almost back to normal. They also reduce TNF alpha, however it remains greater than the control. Compared to the diabetic group, they reduced collagen back to normal. Additionally, they were able to raise beclin to a level that was noticeably greater than the control.