Volume 16, Issue 3 (May-Jun 2022)                   mljgoums 2022, 16(3): 19-23 | Back to browse issues page

XML Print


Download citation:
BibTeX | RIS | EndNote | Medlars | ProCite | Reference Manager | RefWorks
Send citation to:

roohbakhsh E, barari A, abbaszadeh H. Effects of Exercise Training and Consumption of Fenugreek Seed Extract on Expression of Oxidised LDL and ROS-related Genes in Patients with Coronary Artery Occlusion. mljgoums 2022; 16 (3) :19-23
URL: http://mlj.goums.ac.ir/article-1-1358-en.html
1- Department of Sport Physiology, Ayatollah Amoli Branch, Islamic Azad University, Amol, Iran
2- Department of Sport Physiology, Ayatollah Amoli Branch, Islamic Azad University, Amol, Iran , alireza54.barari@gmail.com
3- Department of Sport Physiology, Sari Branch, Islamic Azad University, Sari, Iran
Full-Text [PDF 547 kb]   (333 Downloads)     |   Abstract (HTML)  (1100 Views)
Full-Text:   (428 Views)
INTRODUCTION
Cardiovascular disease (CVD) is the leading cause of death in developing countries (1,2). It is estimated that by 2020, CVD will be one of the most common diseases in the world (3). Identifying the risk factors of CVDs plays an important role in preventing sudden cardiac death. Hypertension, low-density lipoprotein (LDL) level, age, smoking, glucose intolerance, diabetes, and physical inactivity have been reported as the main causes of CVD-related mortality (4,5). Regular exercise has been regarded as an effective method of CVD prevention. Research also suggests that using herbal supplements is a good alternative to preventing cardiovascular events. The use of supplements, especially dietary supplements, has long been proposed to prevent physical weakness and improve vigor (6). The tendency of the general population to use herbal medicine has increased in recent years due to the harmful effects of chemical drugs. Fenugreek (Trigonellafoenum-graecum) is a medicinal plant with remarkable healing properties that has long been used in Iranian traditional medicine. It has anti-atherosclerosis, anti-inflammatory as well as blood cholesterol, blood lipids, high blood pressure, and blood triglycerides lowering effects (7).On the other hand, in recent years, high-intensity intermittent exercise training (HIIT) has been recognized as an effective exercise intervention with similar or greater benefits than moderate-intensity intermittent exercise (8,9). For example, it has been reported that HIIT has similar effects to moderate-intensity intermittent exercise on skeletal muscle metabolic adaptations, cardiovascular fitness, and body composition. Both herbal supplements and physical activity are effective in controlling or preventing CVD risk factors, such as LDL level. Clinical studies have shown that the atherogenic properties of oxidized LDL-cholesterol (oxLDL-c) are greater than those of LDL-c (10-12). The mechanism of LDL-c entry into macrophages is not mediated by the LDL-c receptor (13), but several studies have shown that LDL-c, the largest carrier of cholesterol in the blood, is first converted to oxLDL-c, and then enters macrophages by scavenger ii receptors or oxLDL-c receptors (10,11,14). It has been reported that oxLDL-c levels are very high in patients with coronary heart disease (15). Antioxidants inhibit the production of oxLDL-c (16,17). On the other hand, exercise, especially regular moderate-intensity exercise, increases the amount of antioxidants, thereby improving the body's defense system (18, 19).The Centers for Disease Control and the American College of Exercise Medicine have recommended that 30 minutes of moderate-intensity physical activity on most days of the week can improve cardiovascular health in adults (20). Increased oxidative stress and lipoprotein oxidation are associated with coronary artery disease (10). There was also a significant relationship between oxLDL-c levels and CVD-related deaths (21). Under normal conditions, about 2 to 5 percent of mitochondrial oxygen is transferred to free radical oxygen compounds such as superoxide (O2, hydrogen peroxide, hydroxyl, etc.). Oxidative stress is a process caused by excess of free radicals on the surface of cell membranes, which can damage membranes of cells and intracellular organelles, especially mitochondria. Damage to lipid membranes causes cell peroxidation and lipid wall hardening. Under normal conditions, antioxidants convert reactive oxygen species (ROS) into water and prevent the production of free radicals (22, 23). The imbalance between oxidants and antioxidants disrupts the normal function of immune cells (24). High intensity physical activity, especially intense aerobic exercise, increases oxidative stress and lipid peroxidation (25). Some studies have shown that oxLDL-c had significantly reduced in patients with coronary heart disease after a period of moderate-intensity aerobic exercise along with a diet plan (26,27). The present study aimed at investigating effects of intermittent exercise and consumption of fenugreek seed extract on expression of oxLDL and ROS in patients with coronary artery occlusion.


MATERIALS AND METHODS
This was an experimental study with a pretest-posttest design. The statistical population included all men with CVD (aged 55 to 65 years) who had been referred to the Rouhani and Shahid Beheshti hospitals in Babol (north of Iran) in the second half of 1397-98. The subjects had no history of exercise or consumption of fenugreek seed extract for at least six months. Angina pectoris and irreversible heart failure were the exclusion criteria. Overall, 32 subjects were enrolled in the study after medical examination. Written consent was taken, and then the subjects were divided into four groups: control, fenugreek supplementation, exercise, and exercise + fenugreek supplementation.
Exercise was discontinued if the subjects experienced any specific problems or discomforts such as ST segment depression in electrocardiogram, sudden increase in heart rate, and decrease in blood pressure. Patients' records were reviewed by a physician after each session. The research protocol was approved by the Ethics Committee of the Islamic Azad University, Babol Branch, Iran (ethical approval code: IR.IAU.BABOL.REC.1398.091).
The training program consisted of eight weeks of intermittent indoor running for 50-70 minutes, three sessions per week with emphasis on gradual overload. Training intensity ranged between 55%and 65% of heart rate reserve, and rest periods ranged from 35% to 45% of heart rate reserve.
Each training session began with 10 minutes of warm-up (stretching and light and dynamic movements of the whole body) and ended with 10 minutes of cool down. The intensity of the main workout was monitored using a polar heart rate tracking device.
The subjects were instructed to eat a relatively uniform diet containing equal amounts of fruits and vegetables and not to take special supplements. Diets were controlled using a questionnaire.
Fenugreek seeds were obtained from villages around Kermanshah (west of Iran). Then, 1 kg of pulverized seeds powder was soaked in 2 liters of water and ethanol 96% for 48 hours. Next, the mixture was filtered with filter paper and then dried   (22). Subjects in the supplementation groups consumed 10 mg/kg of fenugreek extract daily (at 6 o'clock) for 8 weeks. Fasting blood samples were taken at 24 hours before the intervention and 48 hours after the last day of the intervention. Whole blood was centrifuged at 3,000 rpm for 7 minutes. The white fluid formed between the red blood cell layer at the bottom and the plasma called the weave layer was slowly removed with a weave sampler.
To extract RNA, about 100 μl of buffy coat were mixed with 1 ml of Trizol solution (Sigma-Aldrich, USA) in a microtube free of RNase enzyme. The microtube was centrifuged at 2-8 °C for 15 minutes at 12000 g. Finally, the precipitate was dissolved in DEPC-treated water and stored at -70 °C.
Except for the first stage, which was performed under a conventional hood due to trizol toxicity, all steps were performed under a laminar hood. The extracted RNA was quantitatively analyzed by spectrophotometry and electrophoresis on agarose gel. Next, cDNA synthesis was done using a commercial kit (Fermentas, Germany)(28).
After reverse transcription reaction, in order to amplify the desired fragment and quantitatively evaluate the expression of ROS-related genes, real-time PCR reaction was performed using SYBR Green dye. To determine the efficiency of primers, the LinReg PCR software was used.
Data were described using descriptive statistics. Intragroup and intergroup changes between pretest and posttest stages were analyzed using two-way analysis of variance with repeated measures and Tukey's test. All statistical analysis was carried out using GraphPad Prism 8 and Microsoft Excel at a significant level of <0.05.



RESULTS
The mean level of oxLDL gene expression decreased significantly after the training, fenugreek supplementation, and combination of fenugreek supplementation with training (p<0.0001). The results of Tukey's post hoc test also showed that the mean level of oxLDL gene expression differed significantly between the intervention and control groups (p<0.0001) (Figure (1).


Figure 1.‌ Comparison of mean oxLDL gene expression in different groups between the pretest and posttest stages. Before: 24 hours before interventions, After: 48 hours after the last day of interventions. Control: control group, Train: Exercise group, Shanb: Fenugreek supplementation, Shanb + Train: Exercise + Fenugreek supplementation. a: Significant difference compared to the control group (p<0.05). b: significance difference compared to the fenugreek + exercise group (p <0.05). *: p<0.05, **: p<0.0001.
 The mean level of ROS expression decreased significantly in all intervention groups (p<0.0001). Moreover, the mean level of ROS expression in the posttest stage was significantly lower in all intervention groups compared with the control group (p<0.0001).


Figure 2.‌ Comparison of mean ROS gene expression in different groups between the pretest and posttest stages. a: Significant difference compared to the control group (p<0.05). b: significance difference compared to the fenugreek + exercise group (p <0.05). *: p<0.05), **: p<0.0001.
DISCUSSION

Due to sociocultural modernization and decrease in physical activity, the incidence of diseases caused by physical inactivity, such as CVD, is on the rise increasing (1,2). The present study evaluated the effects of intermittent exercise and fenugreek seed extract supplementation on expression of oxLDL and ROS in patients with coronary artery occlusion. Based on the results, the expression of ROS and oxLDL genes reduced significantly after the intervention. A previous study also reported that oxLDL-c levels reduced significantly in patients with coronary heart disease after a period of moderate-intensity aerobic exercise with a diet plan (28). Under normal conditions, there is a balance between the amount of ROS and antioxidants (29). Disruption of this balance and the increase in ROS, especially during strenuous exercise, causes oxidative stress (25). During intense endurance (aerobic) training, the production of ROS increases, mainly in the mitochondria of active muscle cells (30). It has been shown that fenugreek has anti-atherosclerotic, anti-inflammatory, and antioxidant effects (31). Interval training may lead to overproduction of free radicals (32,33). Some studies on both animals and humans have reported an increase in free radicals production after aerobic or anaerobic training in subjects with coronary artery disease (32,34). The binding of oxLDL to low-density lipoprotein receptor-1 (LOX-1) activates NADPH oxidase by inducing translocation of specific subunits on the cell membrane, leading to a rapid elevationin intracellular ROS, such as hydrogen peroxide and superoxide; the latter decreases intracellular nitric oxide and upregulates LOX-1 expression, thus resulting in further increase in ROS production (33-35).
Flavonoids can regulate some steps of angiogenesis, such as cell migration and the formation of microcapillary tubules (35). It has been demonstrated that fenugreek could improve maximal and submaximal aerobic function (34). In addition, fenugreek extract can significantly reduce the atherogenic index. Fenugreek contains bitter saponins such as protodioscin. Studies have shown the effect of diazine (a form of protodioscin and diosin) on fat and glucose metabolism. Diosgenin increases the amount of PPAR-y in white adipose tissue and induces the differentiation of fat cells and reduces the size of adipocytes. This increases the secretion of adiponectin, which inhibits inflammation in adipocytes (33).
In this study, control of each subject's diet, level of motivation, mental stress, lifestyle, endocrine secretion, as well as genetic and congenital characteristics could not be controlled.
The limitations of the present study were the small number of subjects and the measurement of limited angiogenic indices. Therefore, it is suggested to conduct future studies on larger study populations in order to understand the possible effects of periodic exercises and fenugreek extract supplementation.

 CONCLUSION
Interval training may lead to adaptations faster than continuous aerobic training. The results of this research showed that a combination of interval training and fenugreek seed extract supplementation can reduce oxLDL and expression of ROS-related genes. Therefore, the use of this supplement could have beneficial effects on antioxidant defense of patients with coronary artery occlusion.
ACKNOWLEDGMENTS

The authors would like to thank all participants in the study for their cooperation.
DECLARATIONS
Funding
The authors received no financial support for the research, authorship, and/or publication of this article.
Ethics approvals and consent to participate

Written consent was taken from all subjects prior to participation in the study. The study has been approved by the Ethics Committee of the Islamic Azad University, Babol Branch, Iran (ethical approval code: IR.IAU.BABOL.REC.1398.091).
 Conflict of interest
The authors declare that there is no conflict of interest regarding publication of this article. 
 
Research Article: Original Paper | Subject: Sport Physiology
Received: 2021/01/17 | Accepted: 2021/03/1 | Published: 2022/05/14 | ePublished: 2022/05/14

References
1. Subasi S, Geleccek N, Ozdemir N, Ormen M. Influence of acute resistance and aerobic exercise on plasma homocysteine level and lipid profiles. Turk J Biochem. 2009; 34(1): 9-14.
2. DehghanSH, SharifiGH, Faramarzi M. The effect of eight week low impact rhythmic aerobic training on total plasma homocysteine concentration in older non-athlete women. J MzandaranUniv Med Sci 2009; 19(72): 54-59. [View at Publisher] [Google Scholar]
3. Jabery A, Jazayery A, Mohagheghi A, Rahimi A. Blood homocysteine enhancement in 35- 65 ischemic stricken patience. J Hygiene Res Center 2003; 7(3): 63-67.
4. Gill JM, Caslake MJ, McAllister C, Tsofliou F, Ferrell WR, Packard CJ, et al. Effects of short-term detraining on postprandial metabolism, endothelial function, and inflammation in endurance-trained men: dissociation between changes in triglyceride metabolism and endothelial function. j.gill@bio.gla.ac.uk. J Clin Endocrinol Metab. 2003; 88(9): 4328-35. [View at Publisher] [DOI:10.1210/jc.2003-030226] [PubMed] [Google Scholar]
5. Ridker PM , Rifai N , Rose L , Buring JE , Cook NR. Comparison of C-reactive protein and low-density lipoprotein cholesterol levels in the prediction of first cardiovascular events. New England Journal of Medicine. 2002; 347(20): 1557-1565. [View at Publisher] [DOI:10.1056/NEJMoa021993] [PubMed] [Google Scholar]
6. Abdullaev F. Plant-derived agents against cancer. Pharmacology and therapeutics in the new millennium. New Delhi: Narosa Publishing House. 2001; 345-354. [Google Scholar]
7. KamaliSarvestani A. The combined effect of aerobic exercise and fenugreek supplementation on the lipid profile of inactive obese women, M.Sc. Thesis, 2013
8. Burgomaster KA, Howarth KR, Phillips SM, Rakobowchuk M, Macdonald MJ, McGee SL, et al. Similar metabolic adaptations during exercise after low volume sprint interval and traditional endurance training in humans. J Physiol. 2008; 586: 151-60. [View at Publisher] [DOI:10.1113/jphysiol.2007.142109] [PubMed] [Google Scholar]
9. GibalaMJ. High-intensity interval training: New insights. Chinese Sports Medicine. 2008; (3):31.
10. Steinberg D, Parthasarathy S, Carew TE, KhooJC, Witztum JL. Beyond cholesterol: Modifications of lowdensity lipoprotein that increase its atherogenicity. N Engl J Med. 1989; 320: 915-24. [View at Publisher] [DOI:10.1056/NEJM198904063201407] [PubMed]
11. HeineckeJW. Oxidants and antioxidants in the pathogenesis of atherosclerosis: implications for the oxidized low density lipoprotein hypothesis. Atherosclerosis. 1998; 141: 1-15. [View at Publisher] [DOI:10.1016/S0021-9150(98)00173-7] [PubMed] [Google Scholar]
12. Yla-Herttuala S. Is oxidized low-density lipoprotein present in vivo? CurrOpinLipidol. 1998; 9: 337-44. [View at Publisher] [DOI:10.1097/00041433-199808000-00009] [PubMed] [Google Scholar]
13. Brown MS, Goldstein JL. Lipoprotein metabolism in the macrophage: implications for cholesterol disposition in atherosclerosis. Annu Rev Biochem 1983; 52: 223-61. [View at Publisher] [DOI:10.1146/annurev.bi.52.070183.001255] [Google Scholar]
14. Chisolm GM, Hazen SL, Fox PL, Cathcart MK. The oxidation of lipoproteins by monocytes-macrophages.Biochemical and biological mechanisms. J BiolChem. 1999; 274: 25959-62. [View at Publisher] [DOI:10.1074/jbc.274.37.25959] [PubMed] [Google Scholar]
15. Holvoet P, Vanhaecke J, Janssens S, Van de WerfF,Collen D. Oxidized LDL and malondialdehyde-modified LDL in patients with acute coronary syndromes and stable coronary artery disease. Circulation 1998; 98:1487-94. [View at Publisher] [DOI:10.1161/01.CIR.98.15.1487] [PubMed] [Google Scholar]
16. Erel O. A novel automated direct measurement method for total antioxidant capacity using a new generation,more stable ABTS radical cation. ClinBiochem. 2004; 37: 277-85. [View at Publisher] [DOI:10.1016/j.clinbiochem.2003.11.015] [PubMed] [Google Scholar]
17. Young IS, Woodside IV. Antioxidants in health and disease. J ClinPathol; 2001. 54: 176-86. [View at Publisher] [DOI:10.1136/jcp.54.3.176] [PubMed] [Google Scholar]
18. Clarkson PM, Thompson HS. Antioxidants: what role do they play in physical activity and health? Am J ClinNutr. 2000; 72: 637S-46S. [View at Publisher] [DOI:10.1093/ajcn/72.2.637S] [PubMed] [Google Scholar]
19. Wang JS, Lin CC, Chen JK, Wong MK. Role of chronic exercise in decreasing oxidized LDL-potentiated platelet activation by enhancing platelet-derived no release and bioactivity in rats. Life Sci. 2000; 66: 1937-48. [View at Publisher] [DOI:10.1016/S0024-3205(00)00519-1] [PubMed] [Google Scholar]
20. Petit PR, Sauvaire YD, Hillaire-Buys DM, Leconte OM, Baissac YG, Ponsin G, et al. Steroidsaponins from fenugreek seeds: extraction, purification, and pharmacological investigation on feeding behavior and plasma cholesterol. Steroids.1995; 60(10): 674-680. [View at Publisher] [DOI:10.1016/0039-128X(95)00090-D] [PubMed] [Google Scholar]
21. Yagi K. Lipid peroxides and human diseases. ChemPhys Lipids. 1987; 45: 337-51. [View at Publisher] [DOI:10.1016/0009-3084(87)90071-5] [PubMed] [Google Scholar]
22. Buetner GR, Schafer FQ. Free radicals, oxidants and antioxidants. Teratology. 2000; 62: 234-38. [View at Publisher] [DOI:10.1002/1096-9926(200010)62:43.0.CO;2-9] [PubMed] [Google Scholar]
23. Rumley AG, Paterson JR. Analytical aspects of antioxidants and free radical activity inclinical biochemistry. Ann ClinBiochem; 1998. 35: 181-200. [DOI:10.1177/000456329803500202] [PubMed] [Google Scholar]
24. William D, Mc Ardle-Frank I. Katch. Exercise Physiology. 2nd ed. Philadelphia: Lippincott. 2007; 72-4.
25. Akams A, Best TM. The role of anti oxidants in exercise and disease prevention. Med Sci Sports Exerc;2002. 30(5): 265-71. [View at Publisher] [DOI:10.3810/psm.2002.05.281] [PubMed] [Google Scholar]
26. Hodgson EK, Fridovich I. The interaction of bovine erythrocyte superoxide dismutase with hydrogen peroxide: inactivation of the enzyme. Biochemistry. 1975; 14(24): 5294-9. [DOI:10.1021/bi00695a010] [PubMed]
27. Naghibi S, Maleki J. The effect of exercise training on anaerobic threshold and exercise tolerance in patients with coronary artery disease. medical social. 2011; 17-33. [Google Scholar]
28. Srimahachota S, Wunsuwan R, Siritantikorn A, Boonla C, Chaiwongkarjohn S, Tosukhowong P. Effects of lifestyle modification on oxidized LDL, reactive oxygen species production and endothelial cell viability in patients with coronary artery disease. ClinBiochem. 2010; 43: 858-62. [View at Publisher] [DOI:10.1016/j.clinbiochem.2010.04.056] [PubMed] [Google Scholar]
29. Lloyd PG, Prior BM, Yang HT, TerjungRL. Angiogenic growth factor expression in rat skeletal muscle in response to exercise training. American Journal of Physiology-Heart and Circulatory Physiology. 2003; 284(5): H1668-H78. [View at Publisher] [DOI:10.1152/ajpheart.00743.2002] [PubMed] [Google Scholar]
30. Attaran M, Pas qualotto E, Margaritas I, Palazzetti S, Rousseau AS. Antioxidants supplementation and tapering exercise, improve exercise-induced antioxidant response. Am J Nutrition. 2003; 22(2): 147-56. [View at Publisher] [DOI:10.1080/07315724.2003.10719288] [PubMed] [Google Scholar]
31. Emtiazy M, Oveidzadeh L, Habibi M, Molaeipour L, Talei D, Parvin M, et al. Investigating the effectiveness of the Trigonellafoenum-graecum L.(fenugreek) seeds in mild asthma: A randomized controlled trial. Allergy, Asthma & Clinical Immunology. 2018; 14(1):19. [View at Publisher] [DOI:10.1186/s13223-018-0238-9] [PubMed] [Google Scholar]
32. Arshadi S, Azarbayjani MA, Hajiaghaalipour F, Yusof A, Peeri M, Bakhtiyari S, et al. Evaluation of Trigonellafoenum-graecum extract in combination with swimming exercise compared to glibenclamide consumption on type 2 Diabetic rodents. Food & nutrition research. 2015; 59(1): 29717. [View at Publisher] [DOI:10.3402/fnr.v59.29717] [PubMed] [Google Scholar]
33. yousefi E, Zavoshy R, Noroozi M, Jahani Hashemi H, Zareiy S, Alizade K et al . Effect of oral administration of fenugreek seeds powdered on lipid profile. EBNESINA. 2015; 17 (1) :33-38 [View at Publisher] [Google Scholar]
34. GohZhong Sheng J. Effects of curcumin and fenugreek soluble fiber supplements on submaximal and maximal aerobic performance indices in untrained college-aged subjects. UKnowledge. 2019. [View at Publisher] [DOI:10.3390/jfmk5020034] [Google Scholar]
35. Nisari M, Alpa S, Yilmaz S, Inanc N. Effects of Fenugreek Extract on Total Antioxidant/Oxidant Status at Ehrlich Ascites Tumor Bearing Mice. EJMI. 2020; 4(1):116-122. [DOI:10.14744/ejmi.2020.17680] [Google Scholar]

Add your comments about this article : Your username or Email:
CAPTCHA

Send email to the article author


Rights and permissions
Creative Commons License This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

© 2007 All Rights Reserved | Medical Laboratory Journal

Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.