AMELIORATION OF INFLAMMATION AND CARDIOPROTECTIVE EFFECT OF TERMINALIA ARJUNA BARK EXTRACT IN HIGH-FAT DIET RAT MODEL

AMELIORATION OF INFLAMMATION AND CARDIOPROTECTIVE EFFECT OF TERMINALIA ARJUNA BARK EXTRACT IN HIGH-FAT DIET RAT MODEL 

Priya Palanivelu, Vijayakumar Jagadesan, Rajagopalan Vijayaraghavan, Senthilkumar Sivanesan * and Shree Mukilan Pari

Department of Research and Development, Saveetha Medical College, Saveetha Institute of Medical and Technical Sciences, Chennai, Tamil Nadu, India.

ABSTRACT: The objective of this study is to investigate the protective effect of aqueous bark extract of Terminalia arjuna (TA) on high-fat diet-induced cardiovascular changes in Wistar rats using histological, immunohistochemical, and gene expression analysis. Group 1- without treatment served as control, group 2- High fat diet (HFD) received corn oil orally at a dose of 10 mL/kg, group 3 - HFD+atorvastatin (ATV) - 10 mg/kg, group 4 - HFD+TA125 mg/kg , and group 5- HFD+TA250 mg/kg, 6 days a week for 5 weeks. The activity of MMP-9 was measured in blood, Hematoxylin and Eosin staining was analyzed with heart tissue and TNF-α immunohistochemistry and ADIPOQ gene expression analysis were conducted using heart tissue. When Compared to the control group, the HFD group showed higher levels of MMP-9 activity and morphological changes in heart histology. TNF-α expression was also significantly increased in HFD group and down-regulation of ADIPOQ gene expression levels. MMP-9 activity and pathological damage to cardiac tissue including ADIPOQ gene expression were significantly reverted when T. arjuna bark extract was administered along with a high-fat diet. The findings reveal that T. arjuna bark has protective against HFD-induced cardiovascular alterations.

Keywords: ADIPOQ gene, High fat diet, MMP- 9, Rats, Terminalia arjuna, TNF-α immunohistochemistry

INTRODUCTION: Heart disease remains the predominant cause of mortality in the United States as well as worldwide. Even in European nations, cardiovascular disease (CVD) and stroke continue to be the leading causes of death, with coronary heart disease (CHD) and stroke accounting for maximum deaths ¹. It was reported that the accumulation of dietary Trans fatty acids (TFA) in plasma and various tissues can trigger inflammation and oxidative stress ².

Hyperlipidaemia increases the production of peroxynitrite in the heart, resulting in myocardial dysfunction ³. MMPs (matrix metalloproteinases) are a group of zinc-dependent endoproteases that are involved in tissue remodelling as well as protein degradation in extracellular matrix (ECM) ⁴. MMPs are indicators of atherosclerotic plaque instability CHD. The future risk of both advanced cardiovascular disease (CAD) and acute coronary syndrome (ACS) could be predicted based on the increased levels of MMPs, indicating the value of MMPs as a prognostic tool and key biomarker of cardiovascular disease ⁵. Circulating MMP-9 has been reported as a promising biomarker to predict atherosclerotic plaques, plaque instability as well as CVD ^5-7. Statins have pleiotropic effects in vivo, such as influencing signalling systems that inhibit MMPs ⁸. The peripheral levels of MMP-2 and MMP-9 can be used as indicators of heart failure and for identifying individuals who would benefit from a specific therapeutic intervention involving the MMP pathway ⁹. Findings reveal that individuals with hyperlipidaemia produce more reactive oxygen species (ROS) ¹⁰.

This could be due to a fat-rich diet increasing the expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase genes ¹¹, which is the primary enzyme responsible for ROS synthesis ¹² resulting in increased ROS formation in cells ¹³. Low-density lipoprotein (LDL) oxidation is triggered by the presence of ROS, which activates inflammatory cytokines, mediators and chemokines produced by invading resident macrophages. Both ROS and inflammation play a role in the onset and progression of CVD ¹⁴. Atherosclerosis is considered as an inflammatory illness rather than a result of hyperlipidemia ¹⁵.

Recent research has suggested that immune response has a role in the aetiology of atherosclerosis, as evidenced by statins' anti-inflammatory action in the therapy of atherosclerosis ¹⁶. The role of TNF-α in the affected heart tissue support both cardioprotective mechanisms and pathogenesis ¹⁷. Despite the therapeutic benefits seen during targeting of TNF-α pathway in heart disease, it was discerned that a more specific and selective mechanism of TNF-α is required to reduce the disease pathogenesis. The current cholesterol-lowering drugs used to manage CVD pose several side effects which include toxicity and muscle-related problems ^{18, 19}. The risks associated with statins are myopathy, incidence of diabetes, hemorrhagic stroke, memory and cognitive impairment, hepatotoxicity renal toxicity and cataract ^20-22. There is a need for safe and viable therapeutic approach to combat CVD and associated problems. Many pivotal works have shown the importance and value of herbal medicines in treating CVD problems. Doxycycline induced shorter epicardium, decreased myocardial density with hyalinization and vacuolated swelling endocardium are signs of myofibril or functional cardiomyocyte loss and are responsible for left ventricular failure. Co-treatment with TAAqE (Terminalia arjuna aqueous extract) prevented the structural alterations. TAAqE appears to protect against Doxycycline-induced LV dysfunction²³. Terminalia arjuna extract has cardioprotective properties, as evidenced by its ability to reduce heart damage induced by Isoproterenol (ISO), oxidative stress, apoptosis, and restore antioxidant state.

The scavenging of free radicals produced by Isoproterenol-induced stress is thought to be the mechanism of Terminalia arjuna extract cardio-protection. Cardioprotective and antioxidant properties of Terminalia arjuna extract could be seen through its ability to reduce heart damage induced by ISO, involving oxidative stress and apoptosis ²⁴. In this study, T. arjuna rendered cardiac protection by scavenging the free radicals generated by ISO. The cardioprotective effect of Alpinia zerumbet, a natural product with potential antioxidant action in the myocardium ²⁵, shows that there are many avenues for detailed research exploration of molecular mechanisms exhibited by those potential phytochemical drugs.

The bark of Terminalia arjuna, a deciduous tree, has long been thought to be therapeutic for cardiac diseases in Indian medicine ^{26, 27}. Many ayurvedic remedies labelled as cardiotonics have it as an essential ingredient ²⁸. The increased understanding of inflammatory events in atherosclerosis provides a molecular framework for comprehending the therapeutic benefits of medications like statins and T. arjuna, which also have immunomodulatory properties ²⁹. T. arjuna extract increased endogenous antioxidant molecules in the rat heart to protect the myocardium from isoproterenol-induced myocardial ischemia-reperfusion injury ³⁰. Studies support the anti-diabetic properties of TA extract and Emblica officinalis aqueous stem bark extracts ³¹. By reducing the expression of pro-inflammatory cytokines and chemokines, decreasing oxidative stress, enhancing plasma zinc levels and gut microbiota structure, TA extract treatment alleviated disease activity in Trinitrobenzene sulfonic acid (TNBS) induced colitis ³². Chronic high-fat diet (HFD) leads to disease development and decrease in cardiac adiponectin gene expression. Reduced cardiac ADIPOQ causes oxidative stress and inflammation, which leads to heart hypertrophy and reduced cardiac function ³³. A study reported the therapeutic role of Moringa oleifera ethanolic extract in the management of obesity and elimination of cardiometabolic abnormalities in female rats caused by high cholesterol diet. M. oleifera extract improved mRNA expression of leptin, resistin and ADIPOQ genes and functioned directly on visceral fat mass ³⁴. Intervention with TA 500 mg/kg could be a good substitute for NAC (N-acetyl cysteine) in rendering protection from APAP-induced (acetaminophen) hepatotoxicity ³⁵. Although evidence supports the traditional use of TA bark extract for cardio-protection, there is a paucity of animal studies to support the mechanism and cardioprotective benefits of this drug in a high-fat diet model. The aim of to investigate the protective effect of bark extract of T. arjuna on high fat diet induced cardiovascular alterations using immunohistochemical and gene expression analysis in rats.

MATERIALS & METHODS:

Animals: Wistar male rats weighing 180–280 g were used for this study. The rats were housed in polypropylene cages (3 rats per cage) with sterile paddy husk for a week to acclimatize to the laboratory environment (25±2ºC, 40-60% humidity and natural light/dark cycle). The rats were fed a pellet meal (Biogen, Bangalore) and given filtered water while being observed at the Centre for Laboratory and Animal Research (CLAR), Saveetha Institute of Medical and Technical Sciences, Chennai. The bedding husk material was changed on a regular basis to maintain a hygienic environment.

Ethics: The Protocol was approved by the Saveetha Medical College Institutional Animal Ethics Committee (SU/CLAR/RD/005/2019 Dated August 09, 2019). Animals were housed and maintained according to the standards of the "Committee for the Purpose of Control and Supervision of Experiments on Animals" (India).

Experimental Groups: Corn oil was given orally at a dose of 10 mL/kg six times a week as a high-fat diet for five weeks to induce heart damage to induce cardiac effects. A low dose of T. arjuna aqueous bark extract was orally administered at (125 mg/kg) six times a week for five weeks, and a high dose (250 mg/kg) of T. arjuna aqueous bark extract was orally administered six times a week for five weeks. The therapeutic doses were given 30 minutes after the corn oil was administered. The compounds were suspended in 2% DMSO for the administration of atorvastatin. Five groups of animals were used (6 rats each). The rats in Group 1 were used as a control group. For them, instead of corn oil, water was administered orally. The rats in Group 2 were provided with a high-fat diet. The groups from 3 to 5 were given a high-fat diet and atorvastatin (10 mg/kg), aqueous bark extract of T. arjuna (125 mg/kg) and aqueous extract of T. arjuna (250 mg/kg) respectively.

Sample Preparation: Twenty four hours after the last dose, the animals were anesthetized with isofluorane, the blood was collected in vacutainer tubes without anticoagulants from retroorbital puncture. It was allowed to coagulate for 20 minutes before being centrifuged at 3000 rpm at 4°C to separate the serum. The serum was separated and stored at -80°C. The animals were sacrificed by cervical dislocation and a vertical incision was made to access the thoracic cavity. The heart was taken out. The heart was gently cleaned in cold saline, blotted with a Whatmann no.1. filter paper and kept at -80°C for biochemical analysis in sterile plastic vials. The cardiac tissues were also preserved in 10% formalin solution for tissue morphological examinations.

Terminalia arjuna Stem Bark Aqueous Extract Preparation: Herbal Care and Cure Centre (Chennai, India) provided the Terminalia arjuna bark powder. Two litres of distilled water was added to 1 kg fine powder of TA bark in a conical flask. It was boiled for 30 min and allowed to cool slowly. After cooling, it was thoroughly mixed at 37ºC by shaking continuously at 200 rpm. The contents were filtered using a muslin cloth. In a centrifugal evaporator, the filtered extract was dried under reduced pressure at 40°C and refrigerated at 4°C for later use. Approximately 18 % of the yield was obtained. High Performance Thin Layer Chromatography plates were prepared in an optimal solvent system, dried in the air, then scanned at 254 nm with a CAMAG TLC scanner 3. To obtain the sample, the standard stock solution was prepared in HPLC grade methanol. The TA bark extract was dissolved in methanol and sonicated for 10 minutes before being diluted to a final volume of 5 mL. The required concentration of the samples was obtained by dilution of the stock solution. The Phenolic compounds, Flavonoids, Glycosides, Saponin and tannin present in T. arjuna were determined by HPTLC methods. IJPER Journal reviewed our paper (Data unpublished).

Biochemical Analysis: The MMP-9 kit was purchased from Ray Biotech Life Inc, (Peachtree Corners, Georgia). The MMP-9 activity was read at 450 nm using a microplate reader (MINDRAY model, India).

TNF-α Immunohistochemistry Study: TNF-α levels of cardiac tissues were determined by immunohistochemical analysis after the tissues were fixed in fixative for 4 hr and then cryoprocessed with 30% sucrose. The tissues were sectioned into 3-5 µm thick sections, which were then placed in an anti-freeze buffer. After that, the sections were treated with 1 % fixed tissue of H₂O₂ to quench the endogenous peroxidase activity, followed by blocking the nonspecific area by a 5% blocking buffer. After blocking, the sections were incubated overnight at 4°C with an anti-TNF-α antibody (1:1000). The sections were washed to remove the non-specific binding and incubated for 2 hr with a biotinylated secondary antibody (1:500). To visualise the immunopositive cells, the tissues in the slides were stained with diaminobenzidine. Under an inverted microscope, TNF-α immune positive regions were identified, and the acquired images were quantified using Image-J software.

Haematoxylin & Eosin Staining: Paraffin sectioning was used for histopathological evaluation and light microscopic examination of tissues. The tissues were then hydrated and dehydrated in a series of graded alcohols. It was then cleaned with Xylene and chloroform and then fixed in paraffin wax for a rotary microtome. Tissue sections (10µm) were then removed and stored overnight at room temperature. After that, the sections were deparaffinized and moistened with descending alcohol concentrations followed by distilled water. Using haematoxylin and eosin stain, the sections were stained and then washed with ascending concentration of alcohol. Permanent slide was prepared using a DPX mount. The slides were observed under a light microscope (40x) (Olympus microscope) and photomicrographs were taken using a digital camera (Sony RX100M3).

ADIPOQ Gene Expression Analysis: Total RNA was extracted from tissue according to the directions provided by the RNA isolation kit obtained from Invitrogen by Thermo Fisher Scientific. A guanidium isothiocyanate/phenol-chloroform single step extraction kit purchased from Invitrogen by Thermo Fisher Scientific was used to extract total RNA from the tissues. Furthermore, each RNA sample's quality was confirmed using agarose-formaldehyde electrophoresis. RNA samples were kept at -80°C until they were analysed. Electrophoresis on a 1.5 % agarose gel containing ethidium bromide was used to detect polymerase chain reaction (PCR) products. A standard size marker, 100-bpladder (Applied Biosystems, Foster City, CA, USA), was used to confirm the location of the predicted product. After that, the gel was imaged using UV transillumination. A video image analysis system was used to measure the intensity of PCR products (Kodak Digital Science). The signal for each transcript was compared to the signal for -actin mRNA in each sample, and the results were reported as a ratio of the transcript to -actin mRNA.

Statistical Analysis: All values are displayed with mean± standard error. ANOVA was used to test for significance of data of the different groups with Bonforani "t" test for multiple comparison. The significance level was set to P < 0.05.  For statistical analysis and graph plotting, SigmaPlot 14.5 (Systat Software Inc, USA) was utilized.

RESULTS:

Effect of TA on HFD Induced serum MMP 9 Expression Levels: The MMP 9 levels (mean ± SE) of control, HFD, HFD±ATV, HFD±TA125, and HFD±TA250 groups were 123.05 ± 2.07, 344.66 ± 22.48, 165.6 ± 14.56, 197.33 ± 8.34 and 152.2 ± 17.72 ng/mL respectively. The MMP 9 levels of HFD group were 2.1 fold higher than the control. Also, the MMP 9 levels were higher than the reference range of rat species. However, in HFD+TA-treated groups, the MMP 9 levels were reduced as compared with HFD group Fig. 1.

FIG. 1: EFFECT OF ATORVASTATIN (ATV), AND TEST DRUG TERMINALIA ARJUNA AQUEOUS BARK EXTRACT (TA) 125 AND 250 MG/KG ON SERUM MMP 9 EXPRESSION IN HIGH-FAT DIET (HFD) INDUCED RATS. ^aSignificantly different from control group. ^bSignificantly different from hfd group

In comparison with the HFD group, there was 1.6, 1.4, and 1.7 fold decrease in MMP 9 levels in HFD+ATV, HFD+TA 125 and HFD+TA 250 groups. The drug treatment groups showed almost similar results. Although TA250 showed a marginally better effect than TA125 and ATV, the difference was not statistically significant.

Effect of TA on HFD Induced TNF-α changes: The TNF-α positive cells (Mean±SE) of control, HFD, HFD ± ATV, HFD ± TA ± 125, and HFD ± TA250 were 3.31±0.48, 6.96±1.55, 6.22±1.97, 9.26±0.52 and 4.65±1.36  respectively. The TNF-α positive cells of the HFD group increased by 20-fold compared with the control. However, in HFD± TA treated groups, a decrease in the numbers of TNF-α positive cells were seen in Fig. 2 as compared to HFD group. Compared to the HFD group, there was 3, 2 and 4-fold decrease in the TNF-α positive levels in HFD±ATV, HFD±TA 125 and HFD±TA250 groups, respectively.

FIG. 2: REPRESENTATIVE IMAGES OF IMMUNOHISTOCHEMISTRY ANALYSIS FOR TNF-ALPHA. HEART TISSUE STAINED WITH DIAMINOBENZIDINE. X100. FIG. 2A. CONTROL, FIG. 2B. HFD, FIG. C. HFD + STANDARD DRUG, ATORVASTATIN. AND FIG. 2D. HFD + TA 125, FIG. 2E. HFD + TA 250. THE INTENSITY OF THE IMMUNOPOSITIVE SIGNAL IS AS FOLLOWS -HFD>TA125>ATV>TA250>CONTROL (REPRESENTATIVE BAR GRAPH). ^aSignificantly different from control group. ^bSignificantly different from hfd group.

Fig. 2 shows sections stained with diaminobenzidine, showing areas of immunoreactivity for TNF-alpha in heart tissue. Dark brown colour indicates expression of TNF-α positive cells. Samples from the control group did not show any marked areas of TNF-alpha positive expression in Fig. 2A, whereas immunostaining intensity was higher in high fat fed group Fig. 2B. Reduced TNF--α positive cells were seen in HFD+ATV Fig. 3C, HFD+TA125 Fig. 3D and HFD+TA250 Fig. 3E groups.

Effect of TA on Heart ADIPOQ Gene Expression Levels in HFD Induced Rats: The adiponectin gene expression of (mean±SE) of control, HFD.HFD±ATV and HFD±TA250, were 0.57±0.02, 0.19±0.02, 0.41±0.02 and 0.53±0.01 respectively. Fig. 3A and 3B.

The adiponectin gene expression of HFD group decreased by 5 fold compared with the control.

However, in HFD± TA treated groups, the decrease in the gene expression was 5 fold less than the HFD group, in comparison with HFD group, HFD±ATV and HFD±TA250 groups showed 4 and 5-fold decrease in the gene expression levels, respectively.

FIG. 3: GENE EXPRESSION STUDIES OF ADIPONECTIN (ADIPOQ) BY RT-PCR. THE HOUSE KEEPING GENE BETA-ACTIN (ACTB) WAS USED AS A LOADING CONTROL. LANE M- MARKER LANE (100-1000 BP); LANE 1 – CONTROL; LANE 2 –HFD GROUP; LANE 3 –HFD + STANDARD DRUG ATORVASTATIN TREATED; LANE 4 – HFD + TA 250 TREATED. THE REPRESENTATIVE BAR GRAPH BOTTOM FIG. 3B: DEPICTS THE VALUES OF FOLD CHANGE IN VARIOUS GROUPS.

Effect of TA on HFD Induced Histopathology Changes of Heart Tissue: Fig. 4 depicts the H&E stained heart tissues of control, HFD and drug-treated groups. Samples from the control group showed normal histology of cardiac myocardium Fig. 4A. Whereas in HFD induced group Fig. 4B, C & D the heart tissue section showed mononuclear cells (thin arrow) and neutrophil infiltration (arrow head) at the infarct site (Fig. 4B). In Fig. 4B the heart tissue section showed inflammation (long arrow), neutrophil infiltration (broad arrow). Moreover, in HFD group Fig. 4D, the heart tissue architecture was disrupted (long thin arrow) with necrosis of myocardial cells (thick broad arrow). Atorvastatin-treated rats showed less myofibrillar loss, reduced inflammatory cell infiltration and very little interstitial collagen fibres (arrows), as indicated in Fig. 4E and Fig. 4F. Meanwhile, treatment with T. arjuna extract (TA 125 and 250 mg/kg) showed minimal changes in the myocardial cells with few neutrophils (thick arrows) and absence of inflammatory infiltrates Fig. 4G and Fig. 4H respectively.

FIG. 4: REPRESENTATIVE IMAGES OF HISTOPATHOLOGICAL ANALYSIS OF HEART TISSUE STAINED WITH HEMATOXYLIN AND EOSIN CAPTURED AT X40 MAGNIFICATION. SECTION-A (CONTROL), SECTIONS – B, C & D (HFD FED), SECTION-E&F (HFD + STANDARD DRUG ATORVASTATIN TREATED) AND SECTION-G (HFD + TA 125 TREATED), SECTION-H (HFD + TA 250 TREATED)

DISCUSSION: Matrix metalloproteinase-9 (MMP-9) is critical in inflammation and fibrosis in cardiovascular disease ³⁶. High level and activity of MMP-9 has been linked to various cardiovascular disorders. This is because MMP-9 breaks type IV collagen, a major basal membrane constituent of vascular smooth muscle cells and endothelium.

MMP-9 retains a vital role in cell migration and infiltration in atherosclerosis. Furthermore, MMP-9’s degradation of elastin protein has been associated with aneurysm as well as arterial wall stiffness, which can increase the risk of hypertensive crisis ³⁸. The present study shows that TA (250 mg/kg) high dose lowered MMP-9 levels more than ATV treatment, there by suggesting decreased risk of cardiovascular complications and disorders.

Tumor necrosis factor alpha (TNF-α) is a proinflammatory cytokine made by macrophages and monocytes. This protein is involved in a multitude of cell signalling events that can lead to necrosis or apoptosis ³⁹. TNF-α has been shown to play an important role in vascular dysfunction and constrict large blood vessels of the heart ⁴⁰. TNF-α gene polymorphism can induce hypertension ⁴¹.

Additionally, TNF-α has been shown to have cardio depressant effects on cardiac myocytes. By decreasing sarcoplasmic reticulum Ca²⁺ uptake as well as myofilament Ca²⁺ sensitivity, TNF-α can reduce cardiac contractility, thus decreasing cardiac output and tissue oxygen delivery ^{41, 42}. Inactivation of TNF-α can prevent progression of atherosclerotic lesions and improve heart failure patients' health ⁴¹. In comparison to the sepsis group, the levels of inflammatory cytokines were significantly lower in the myocardial tissue of mice treated with sophoridine 20mg/kg, i.p. alone or in combination with dexmedetomidine 25µg/kg, i.p against lipopolysaccharides induced myocardial dysfunction ⁴³. The current study has substantiated that TA extract treatment can decrease TNF-α expression in heart tissue, similar to ATV.

ADIPOQ is an adipokine produced from adipose tissue. While it is usually present at 30 μg/mL in plasma, significantly decreased levels were observed in coronary artery disease ⁴⁴. Previous studies have shown that ADIPOQ exhibits cardioprotective effects through inhibition of inducible nitric oxide synthase (iNOS) and nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) oxidase. In addition, previous studies have demonstrated the ADIPOQ gene’s ability to inhibit cardiomyocyte remodelling by reactive oxygen species ⁴⁵.

The present study shows a statistically significant down-regulation of ADIPOQ gene expression in HFD rats and a statistically significant up-regulation of ADIPOQ gene expression in ATV and TA (250 mg/kg) treatments. This increase in ADIPOQ gene expression by the TA (250 mg/kg) treatment suggests that TA may play a role in cardio-protection.

Therapeutic intervention with Terminalia arjuna ethanol extract or Terminalia arjuna aqueous extract at 250 mg/kg body weight showed considerable cardio-protection in isoproterenol-induced myocardial infarction in rats to biochemical and histological data ⁴⁶. Terminalia arjuna extract may have protected myocytes from apoptosis and necrosis ⁴⁷. The histology evaluation showed minimal changes in the myocardial cells with few neutrophils and absence of inflammatory cells in TA treated HFD-induced rats. Treatment with TA extract showed cellular organization of hepatocytes with reduced necrosis ⁴⁸. T. arjuna extract has also protected renal damage in hyperlipidemic rats ⁴⁹. Therefore, our results illustrate that the Terminalia arjuna bark extract can reduce the histological damage caused due to HFD.

CONCLUSION: In conclusion, the current investigation revealed that T. arjuna aqueous bark extract has therapeutic potential to ameliorate high fat diet-induced cardiovascular alterations and pathological changes.

ACKNOWLEDGEMENT: The authors thank S. Madhan Kumar and P. Praveen Kumar, Laboratory Assistants for their technical support during animal experiments. The authors thank Dr. Indira Kumari, Pathologist from Bright Care research Centre, Chennai, for proving histopathology interpretation.

CONFLICTS OF INTEREST: There is no conflict of interest to disclose.

REFERENCES:

1. Nichols M, Townsend N, Luengo-Fernandez, R, Leal J, Gray A, Scarborough P & Rayner M: European Cardiovascular Disease Statistics 2012.
2. Monguchi T, Hara T, Hasokawa M, Nakajima H, Mori K, Toh R, Irino Y, Ishida T, Hirata KI and Shinohara M: Excessive intake of trans fatty acid accelerates atherosclerosis through promoting inflammation and oxidative stress in a mouse model of hyperlipidemia. Journal of Cardiology 2017; 70(2): 121-7.
3. Yuvaraj S, Ramprasath T, Saravanan B, Vasudevan V, Sasikumar S and Selvam GS: Chrysin attenuates high-fat-diet-induced myocardial oxidative stress via upregulating eNOS and Nrf2 target genes in rats. Molecular and Cellular Biochemistry 2021; 476(7): 2719-27.
4. Raeeszadeh-Sarmazdeh M, Do LD and Hritz BG: Metalloproteinases and their inhibitors: potential for the development of new therapeutics. Cells 2020; 9(5): 1313.
5. Olejarz W, Łacheta D and Kubiak-Tomaszewska G: Matrix metalloproteinases as biomarkers of atherosclerotic plaque instability. International Journal of Molecular Sciences 2020; 21(11): 3946.
6. Li T, Li X, Feng Y, Dong G, Wang Y and Yang J: The role of matrix metalloproteinase-9 in atherosclerotic plaque instability. Mediators of Inflammation 2020; 2020.
7. Zheng X, Zhong C, Zhu Z, Zhang K, Peng H, Xu T, Bu X, Che B, Xu T, Wang A and Chen J: Association between serum matrix metalloproteinase-9 and poor prognosis in acute ischemic stroke patients: The role of dyslipidemia. Nutrition, Metabolism and Cardiovascular Diseases 2021; 31(1): 209-15.
8. Skrzypiec-Spring M, Sapa-Wojciechowska A, Haczkiewicz-Leśniak K, Piasecki T, Kwiatkowska J, Podhorska-Okołów M and Szeląg A: HMG-CoA reductase inhibitor, simvastatin is effective in decreasing degree of myocarditis by inhibiting metalloproteinases activation. Biomolecules 2021; 11(10): 1415.
9. Radosinska J, Barancik M and Vrbjar N: Heart failure and role of circulating MMP-2 and MMP-9. Panminerva Medica 2017; 59(3): 241-53.
10. Lü JM, Lin PH, Yao Q and Chen C: Chemical and molecular mechanisms of antioxidants: experimental approaches and model systems. Journal of Cellular and Molecular Medicine 2010; 14(4): 840-60.
11. Mai A: Oxidative stress in high fat diet-induced metabolic syndrome, hypertension and endothelial dysfunction(Doctoral dissertation, University of Surrey).
12. Castaneda OA, Lee SC, Ho CT and Huang TC: Macrophages in oxidative stress and models to evaluate the antioxidant function of dietary natural compounds. Journal of Food and Drug Analysis 2017; 25(1): 111-8.
13. GervaIs KK, Bernard J, Eric B, Ferdinand DY, Fabrice R, Luc D, Germaine N, Gauze C, Absalome M, Mansour AF and Massara CC: Study of isoforms of nicotinamide adenine dinucleotide phosphate oxidase of the heart in a model of rats fed on several vegetable oils. African Journal of Biochemistry Research 2021; 15(3): 49-59.
14. Cai L, Wang Z, Ji A, Meyer JM and van der Westhuyzen: DR. Scavenger receptor CD36 expression contributes to adipose tissue inflammation and cell death in diet-induced obesity. PloS One 2012; 7(5): 36785.
15. Cai L, Wang Z, Ji A, Meyer JM and van der Westhuyzen DR: Scavenger receptor CD36 expression contributes to adipose tissue inflammation and cell death in diet-induced obesity. PloS one 2012; 7(5): 36785.
16. Sugano R, Matsuoka H, Haramaki N, Umei H, Murase E, Fukami K, Iida S, Ikeda H and Imaizumi T: Polymorphonuclear leukocytes may impair endothelial function: results of crossover randomized study of lipid-lowering therapies. Arteriosclerosis Thrombosis and Vascular Biology 2005; 25(6): 1262-7.
17. Rolski F and Błyszczuk P: Complexity of TNF-α signaling in heart disease. Journal of Clinical Medicine 2020; 9(10): 3267.
18. Ward NC, Watts GF and Eckel RH: Statin toxicity: mechanistic insights and clinical implications. Circulation Research 2019; 124(2): 328-50.
19. Keen HI, Krishnarajah J, Bates TR and Watts GF: Statin myopathy: the fly in the ointment for the prevention of cardiovascular disease in the 21st century. Expert Opinion on Drug Safety 2014; 13(9): 1227-39.
20. Pinal-Fernandez I, Casal-Dominguez M and Mammen AL: Pros y contras de las estatinas. Medicina Clínica Barcelona 2018; 150(10): 398-402.
21. Saxon DR and Eckel RH: Statin intolerance: a literature review and management strategies. Progress in Cardiovascular Diseases 2016; 59(2): 153-64.
22. Bitzur R, Cohen H, Kamari Y and Harats D: Intolerance to statins: mechanisms and management. Diabetes Care. 2013; 36(2): 325-30.
23. Bishop S and Liu SJ: Cardioprotective action of the aqueous extract of Terminalia arjuna bark against toxicity induced by doxorubicin. Phytomedicine 2017; 36: 210-6.
24. Thangaraju MM, Tamatam A, Bhat PV, Deshetty UM, Babusha ST and Khanum F: Terminalia arjuna extract attenuates isoproterenol-induced cardiac stress in Wistar rats via an anti-apoptotic pathway. Proceedings of the National Academy of Sciences, India Section B: Biological Sciences 2020; 90(5): 1101-12.
25. Paulino ET, Ferreira AK, da Silva JC, Costa CD, Smaniotto S, de Araujo-Junior JX, Júnior EF, Bortoluzzi JH and Ribeiro ÊA: Cardioprotective effects induced by hydroalcoholic extract of leaves of Alpinia zerumbet on myocardial infarction in rats. Journal of Ethnopharmacology 2019; 242: 112037.
26. Chopra RN and Chopra IC: Indigenous drugs of India, (Kolkata, Academic Publishers) 1994.
27. Bharani A, Ganguli A, Mathur LK, Jamra Y and Raman PG: Efficacy of Terminalia arjuna in chronic stable angina: a double-blind, placebo-controlled, crossover study comparing Terminalia arjuna with isosorbide mononitrate. Indian Heart Journal 2002; 54(2): 170-5.
28. Dwivedi S: Putative uses of Indian cardiovascular friendly plants in preventive cardiology. Ann National Academy of Medical Science 1996; 32(3&4): 159-75.
29. Halder S, Bharal N, Mediratta PK, Kaur I & Sharma KK: Anti-inflammatory, immunomodulatory and antinociceptive activity of Terminalia arjuna Roxb bark powder in mice and rats. Indian Journal of Experimental Biology 2009; 47: 577-83.
30. Karthikeyan K, Bai BS, Gauthaman K, Sathish KS, Devaraj SN: Cardioprotective effect of the alcoholic extract of Terminalia arjuna bark in an in-vivo model of myocardial ischemic reperfusion injury. Life Sciences 2003; 73(21): 2727-39.
31. Gupta A and Chaphalkar SR: Haemolytic activities and anti-diabetic effect of Terminalia arjuna and Emblica officinalis. European Journal of Pharmaceutical and Medical Research 2016; 3(6): 334-8.
32. Cota D, Mishra S, Shengule S: Beneficial role of Terminalia arjuna hydro-alcoholic extract in colitis and its possible mechanism. Journal of Ethnopharmacology 2019; 230: 117-25.
33. Gupta C, Bubber P, Fahim M, Saidullah B and Omanwar S: Adiponectin in onset and progression of T2DM with cardiac dysfunction in rats. Human & Experimental Toxicology 2020; 39(11): 1463-74.
34. Metwally FM, Rashad HM, Ahmed HH, Mahmoud AA, Raouf ER and Abdalla AM: Molecular mechanisms of the anti-obesity potential effect of Moringa oleifera in the experimental model. Asian Pacific Journal of Tropical Biomedicine 2017; 7(3): 214-21.
35. Kannappan SP, Raghunath G, Sivanesan S, Vijayaraghavan R and Swaminathan M: Antioxidant effect of Terminalia arjuna extract against acetaminophen-induced hepatotoxicity via the regulation of cytochrome P450 2E1, phosphatidylinositol-3-kinase/protein kinase B. Pharmacognosy Magazine 2020; 16(67): 13.
36. Plavelil N, Goldstein R, Klein M, Ho VB, Haigney MC and Hood MN: Tachycardia-Induced Matrix Metalloproteinases Activation Associated with Collagen Type III Cardiac Fibrosis and Heart Failure in Swine. The FASEB Journal 2020; 34(1): 1.
37. Valente FM, de Andrade DO, Cosenso-Martin LN, Cesarino CB, Guimarães SM, Guimarães VB, Lacchini R, Tanus-Santos JE, Yugar-Toledo JC and Vilela-Martin JF: Plasma levels of matrix metalloproteinase-9 are elevated in individuals with hypertensive crisis. BMC Cardiovascular Disorders 2020; 20(1): 1-7.
38. Oh YS: Arterial stiffness and hypertension. Clinical Hypertension 2018; 24(1): 1-3.
39. Li K, Qiu H, Yan J, Shen X, Wei X, Duan M and Yang J: The involvement of TNF-α and TNF-β as proinflammatory cytokines in lymphocyte-mediated adaptive immunity of Nile tilapia by initiating apoptosis. Developmental & Comparative Immunology 2021; 115: 103884.
40. Van Sloten TT: Vascular dysfunction: At the heart of cardiovascular disease, cognitive impairment and depressive symptoms. Artery Research 2017; 19: 18-23.
41. Al-Awsi GR, Al-garawi ED and Abdulhussein HH: Investigation of Tumor Necrosis Factor-Alpha (TNF A) Gene Polymorphism in Patients with Hypertension in Al-Diwaniyah City, Iraq. Journal of Global Pharma Technology 2019; 10(2): 144-8.
42. Bode D, Wen Y, Hegemann N, Primessnig U, Parwani A, Boldt LH, M. Pieske B, R. Heinzel F and Hohendanner F: Oxidative stress and inflammatory modulation of Ca2+ handling in metabolic HFpEF-related left atrial cardiomyopathy. Antioxidants 2020; 9(9): 860.
43. Mao B, Zhou Y and Zhang H: Sophoridine Regulates the NADPH Oxidase-2 Pathway and Potentiates the Cardioprotective Effect of Dexmedetomidine Against Sepsis-Induced Cardiomyopathy. Current Topics in Nutraceutical Research 2021; 19(4).
44. Tao L, Gao E, Jiao X, Yuan Y, Li S, Christopher TA, Lopez BL, Koch W, Chan L, Goldstein BJ and Ma XL: Adiponectin cardioprotection after myocardial ischemia/reperfusion involves the reduction of oxidative/nitrative stress. Circulation 2007; 115(11): 1408-16.
45. Essick EE, Ouchi N, Wilson RM, Ohashi K, Ghobrial J, Shibata R, Pimentel DR and Sam F: Adiponectin mediates cardioprotection in oxidative stress-induced cardiac myocyte remodeling. American Journal of Physiology-Heart and Circulatory Physiology 2011; 301(3): 984-93.
46. Sivakumar V and Rajeshkumar S: Screening of cardioprotective effect of Terminalia arjuna bark in isoproterenol-induced myocardial infarction in experimental animals. Int J Pharma Sci Res 2014; 5(6): 262-8.
47. Borde MK, Mohanty IR, Maheshwari U, Suman R and Deshmukh Y: Natural dipeptidyl peptidase-4 inhibitor Terminalia arjuna mitigates myocardial infarction co-existing with diabetes in experimental rats. J. Diabetes Metab. Disord. Control 2018; 5: 48-56.
48. Ragavan B and Krishnakumari S: Effect of T. arjuna stem bark extract on histopathology of liver, kidney and pancreas of alloxan-induced diabetic rats. African Journal of Biomedical Research 2006; 9(3).
49. Kanthe PS, Patil BS and Das KK: Terminalia arjuna supplementation ameliorates high fat diet-induced oxidative stress in nephrotoxic rats. Journal of Basic and Clinical Physiology and Pharmacology 2021; 22.
    

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    Palanivelu P, Jagadesan V, Vijayaraghavan R, Sivanesan S and Pari SM: Temperature triggered in-situ gelling system for ocular antiviral drug. Int J Pharm Sci & Res 2023; 14(1): 340-48. doi: 10.13040/IJPSR.0975-8232.14(1).340-48.

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