An Experimental Study of the Effect of Six Weeks of Aerobic Exercise along with Q10 Supplementation on TRF1 and TRF2 Gene Expression in Heart Tissue of Aged Wistar Male Rats

Aging is a process of deterioration over time. On the other hand, the prevalence of heart diseases has increased with the increase in life expectancy. Important genes that play a role in the aging process are related to telomerase activity, which plays an important role in heart health. are considered a strong stimulant for the telomere system. Also, coenzyme Q10 is a fat-soluble vitamin, which improves cardiovascular function. The general purpose of this research is to investigate the effect of six weeks of aerobic training along with Q10 supplementation on TRF1 and TRF2 gene expression. Heart tissue was from aged male Wistar rats.

The current study was a developmental study and its method was an experimental one. The statistical population of the current study was 17 old Wistar male mice aged 88 to 96 weeks with an average weight of 404 grams from Razi Vaccine and Serum Research Company. This research was conducted in the physiology laboratory of Shahid Rajaei Tarbiat University. All mice had the same environmental conditions in terms of air temperature (25±2) degrees Celsius, relative humidity (55±5) and day-night cycle (12-12 hours) of light and darkness. The samples were randomly divided into 5 groups including 3 groups of three (exercise sham-supplement sham-control) and two groups of four (aerobic exercise-aerobic exercise and supplement). Laboratory animals were used and approved by the research ethics committee of the Research Institute of Physical Education and Sports Sciences (Ethics ID: IR.SSRC.REC.1401.051) and conducted in the laboratory of the Faculty of Sports Sciences of Tarbiat University, Shahid Rajaei. This study was conducted with a post-test designfor measuring TRF1 and TRF2 and a pre-test-post-test design for measuring Vpeak and lee index. The independent variables of the research included aerobic exercise and Q10 supplement, and the dependent variables included TRF1 gene and TRF2 gene.At the beginning of the study, the animals were familiarized with the environment and trained on a treadmill for two weeks. Finally, after the last familiarization session and after 48 hours of rest, Vpeak [the maximum amount of VO2 )oxygen consumption( during an exercisetest is increasing in which all VO2max criteria are not fulfilled (11) and the Lee index of all subjects were measured. The incremental test started with a speed of 5 m/min and a speed equivalent to 5 m/min was added every three minutes. The time to reach fatigue was determined by the mice's inability to run on the treadmill. Based on the maximum speed obtained, the training protocol was designed for six weeks and five sessions per week for the training groups, and then the training protocols were started (1, 2). Vpeak is measured every two weeks to determine training intensity (14). The program of moderate intensity aerobic training (MICT) is as follows: in the first week, it was performed with an intensity of 65% of Vpeak for 15 minutes, and in the sixth week, it reached an intensity of 70% of Vpeak in 30 minutes: the subjects before starting the protocol with an intensity of -40 50% of Vpeak was heated for 5 minutes and cooled down after completing the protocol.The sham group was given sesame oil by gavage to give stress. The aerobic exercise-supplement group was given 20 mg of fat-soluble Q10 supplement per kilogram of body weight combined with 0.5 cc of sesame oil by gavage for 42 days. The bodyweight was given to the aerobic exercise group. The supplement was prepared from its manufacturing company called Nano Kimia Salamat (15, 16).48 hours after the end of the protocols and the last training session, Vpeak and Lee's index were measured, and also the height and weight of the mice were measured every week. Nasoanal length[cm]] ×1000 was used. 48 hours after the last training session, the samples were anesthetized with a combination of xylazine 5 to 10 mg/kg and ketamine 50 to 100 mg/kg. Then the heart tissue was extracted and washed with PBS and transferred to a negative 80 centigrade freezer and then transferred to the laboratory for tissue analysis using standard kits for gene expression by PCR method (2, 3, 17).Specialized laboratory processes were performed to measure the expression level of TRF1 and TRF2 genes and to measure telomere length using Real time-PCR method. Quantitative real time-PCR was used to measure the gene expression levels using special primers. Amplification of TRF2 and TRF1 genes as well as GAPDH reference gene was done to measure gene expression by real time PCR based on the standard method. To measure the decrease or increase in the expression of TRF1 and TRF2 genes, its expression was compared with the expression of internal control genes. For this purpose, F and R primers and cDNA were added to the master mix and the final volume was prepared for Real time PCR reaction. The primers used were prepared from Cinagen company, made in Iran. (18, 19) Then the real time PCR process was performed using the QIAGEN machine and each reaction was performed in duplicate. After the reaction, the raw data was extracted and the amount was calculated. Expression of genes was done using method 2-ΔΔCt. (20)After collecting the raw data, the Shapiro-Wilk test was used to check the normality of the data distribution. Then, in order to statistically analyze the data and compare between the groups two-way ANOVA test was used, and because the groups were unequal, the Scheffe test was used. Correlated T-test was used to check the changes of stabilizing variables. All statistical calculations were done using SPSS26 software at a significant level of P≤0.05. Excel2013 software was used to draw graphs.

Based on the obtained results, there was a significant difference between the groups after six weeks of aerobic training along with Q10 supplementation on TRF1 and TRF2 gene expression. (P≤0.05)

In general, the present study showed that aerobic exercise along with Q10 supplementation increases the expression of TRF1 and TRF2 genes in heart tissue. Therefore, according to the above result, it is possible to cautiously recommend aerobic exercise along with Q10 supplementation for the elderly.