@2024 Afarand., IRAN

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ISSN: 2252-0805    The Horizon of Medical Sciences 2016;22(3):229-235

Background
Diabetes type I is one of the most common endocrine diseases. In this disease, insulin secretion from pancreatic beta islets decreases and it leads to dysfunction in carbohydrate, fat and protein metabolism [1].
Previous Studies
Nowadays turning to new drugs with fewer side effects is spreading. L-carnitine is a substance with a similar structure to amino acid and it functions like vitamins. It is made of lysine and methionine amino acids in the body and it transfers fatty acids into the mitochondria and increases their oxidation. This substance can boost up the activity of pyruvate dehydrogenase and glucose`s catabolism [2]. ... [3-7].
Aim(s)
This study aimed to investigate the effect of oral prescription of these two materials on the amount of blood glucose and lipid peroxidation of brain and liver tissue in diabetic rats.
Research type
This study is experimental.
Research society, place and time
This study was administered on 50 Wistar male rats weighing 180 to 200 grams.
Sampling method and number
The rats were placed under standard conditions in laboratory animals` breeding center of Zabol University, Faculty of Veterinary Medicine. The cages were plastic with metal reticular lids kept in the special position of the cages on a leg. The cages were changed and cleaned every 4 days. The rats had access to water and foods of their own (Javaneh Khorasan Company, Iran).
Used devices&materials
The rats were randomly divided into five groups: 1) control group (healthy rats), 2) negative control group (diabetic rats with alloxan), 3) under treatment diabetic group with 300 mgs per kg of L-carnitine, 4) under treatment diabetic group with 300 mgs per kg of acetyl L-carnitine and 5) under treatment diabetic group with 300 mgs per kg of acetyl L-carnitine and the same amount of L-carnitine. The solutions were fed to rats for 30 days. To prepare L-carnitine and acetyl L-carnitine solutions, physiologic saline was used. The saline was fed to the rats of control group using a special gavage syringe. The solutions` doses were selected based on previous studies and initial investigations [8, 9]. Experimental infusion of diabetes in rats: the experimental model of diabetes mellitus type I (insulin-dependent diabetes) was caused by intraperitoneal injection of alloxan in male rats. Alloxan powder (sigma; USA) was dissolved in distilled water. Prepared solution was injected by an insulin syringe at a dose of 110 mgs per kg, intraperitoneally. Diabetes symptoms appeared after 3 days including polydipsia, polyuria and weight loss. To ensure that the rats are diabetic their blood sugar levels were controlled after taking blood from their tails by glucometer. If the level of their fasting blood glucose had been more than 140 mgs per DL, they would have been considered as diabetic rats; otherwise, the injection must have repeated [10]. At the end of the experiment, blood samples were taken from the rats` hearts and amounts of blood glucose and biochemical parameters of serum were all measured. To get the serum levels of total cholesterol, triglycerides, HDL-C (high-density lipoprotein) and serum creatinine, biochemical kits (Pars Azmoon; Iran) were employed according to their instructions. Cholesterol and triglycerides were measured using enzymes. In order to measure serum HDL-C, precipitator solution containing human anti-lipoprotein antibody for precipitating chylomicrons, LDL-C (low density lipoprotein) and VLDL-C (very low density lipoprotein) were used. Then, the amount of blue complex created through combining 4-amino anti-pyrine and HDL-C was measured by an auto analyzer (Selectra pro M; Netherlands) [11]. The rats` fasting serum glucose level was determined using a glucometer at the start of the experiment and a biochemical kit at the end of test (Pars Azmoon; Iran). ALT (alanine aminotransferase) and AST (aspartate aminotransferase) enzymes were tested using auto analyzer (Selectra pro M; Netherlands) based on instructions of diagnostic kits (Pars Azmoon; Iran). Measuring tissue level of malondialdehyde (MDA): at the end of the experiment and decapitating (killing animals with ethics), liver and brain samples were removed and after washing them with cold saline, the weight of each sample was measured. The samples were homogenized with Tris buffer and the homogenized solution was centrifuged. To prevent degradation of proteins and enzymes all the above stages were performed in 4 ° C and in the refrigerators of Zabol Faculty of Veterinary Medicine. After centrifugation step, the upper transparent solution was separated from the rest of the solution using a pipette and the lower section of it was thrown away. The upper transparent solution was used to measure the amount of tissue MDA. The level of tissue MDA was assessed by determining the amount of reactive substances with thiobarbituric acid and the instructions of the kit (Anzan Chemistry; Iran). The basis of this kit is measuring MDA through the thiobarbituric method using a spectrophotometer which was described by Okawa in 1997 [12]. According to the instruction of the kit, 100 micro liters of homogenized tissue, deionized distilled water and 0.9% sodium chloride were poured into the test tube (the volume ratio of 1: 1: 1). The test tube was kept for 40 min at 37 ° C. Then, the reaction was stopped using 1 ml of 0.8 molar HCl containing 12% trichloroacetic acid. After adding 1 cc of 1% thiobarbituric acid solution, the solution boiled for 20 minutes and cooled down at room`s temperature. Cooled solution was centrifuged for 30 min at 3000 rounds per minute. The solution`s light absorbance was evaluated at the wavelength of 532 nm using a UV / VIS- 2100 spectrophotometer (UNICO; USA) [12]. All the results were expressed as mean. The normality of distribution of data was examined using the Kolmogorov-Smirnov test. To compare the groups to each other, ANOVA and then complementary Tukey test were deployed.
Findings by Text
The density of blood serum`s fasting glucose decreased significantly in the group under treatment with L-carnitine after 30 days compared to the negative control group (untreated diabetic group) (p< 0.05). In the diabetic group that were under treatment with L-carnitine, the levels of serum`s cholesterol and triglyceride were significantly lower than untreated diabetic group (p< 0.001). L-carnitine could significantly increase the HDL levels compared to the negative control group (p< 0.001). Treating diabetic rats with acetyl L-carnitine did not have any significant effect on the levels of blood`s triglycerides and cholesterol, but it increased the amount of HDL significantly compared to the untreated diabetic group (p< 0.001). Prescribing L-carnitine decreased the amount of serum creatinine compared to the diabetic control group (p< 0.05). In contrast, acetyl L-carnitine had no significant effect on serum creatinine. Treating diabetic rats with L-carnitine reduced the levels of liver enzymes compared to the untreated diabetic group, but it could not bring the amount of these enzymes to the control group. The levels of liver enzymes in the group treated with Acetyl L-carnitine had no significant difference with the untreated diabetic group. The amount of liver enzymes in the group receiving Acetyl-L-carnitine simultaneously with L-carnitine was higher than the group treated merely with L-carnitine (p< 0.05). Prescribing Acetyl-L-carnitine significantly decreased the MDA levels of brain tissue compared to the untreated diabetic group (p< 0.05) but had no significant effect on the MDA of liver tissue. L-carnitine did not have any significant impact on lipid peroxidation of brain tissue, but reduced the MDA levels of liver tissue. Prescribing a combination of L-carnitine and acetyl L-carnitine not only strengthened the effect of two substances, but also reduced the levels of tissue MDA to an extend less than prescribing each of the drugs alone (Table 1).
Main comparison to the similar studies
... [13, 14]. In this study, prescription of L-carnitine (300 mgs per kg) could decrease the amount of MDA of liver tissue as the main indicator of lipid peroxidation and this was similar to the results of a study by Mahfooz et al. [15]. …[16-23] MDA of brain tissue in the group treated with acetyl L-carnitine was significantly less than he untreated diabetic group. This is consistent with the findings of the study by Liu et al. [16] and Cover et al. [24]. ... [25-26].
Suggestions
It is suggested to use high doses of L-carnitine in future studies. Moreover, in further researches time period of prescribing L-carnitine and acetyl L-carnitine is better to be longer and the amount of glycated hemoglobin to be measured.
Limitations
Not evaluating the activities of liver`s antioxidant enzymes such as superoxide dismutase, catalase and GSH-Px is one of the limitations of this study.
Conclusions
Prescribing L-carnitine is more effective than acetyl L-carnitine in reducing secondary side effects of diabetes. Besides, simultaneous prescription of these two substances is not recommended.
Acknowledgments
Many thanks are addressed to cooperation of Mr. Mahdi Mirshekar for performing statistical calculations and Mr. Mahmoud Salehi Moghadam who was responsible for the breeding center of laboratory animals in Zabol University.
Conflict of interest
Non-declared
Ethical Permissions
The protocol of this study was approved and conducted based on international laws about laboratory animals and documents related to ethics committee in medical research (document No. : BP-QP-106-01).
Funding Sources
The study was based on the doctoral thesis of Ms. Shaqayeq Hajian Shahri supported by Basic Sciences of Zabol University, Faculty of Veterinary Medicine.

TABLES and CHARTS

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