Antioxidant and Cytoprotective Properties of Polyphenol-Rich Extracts from Antirhea borbonica and Doratoxylon apetalum against Atherogenic Lipids in Human Endothelial Cells

The endothelial integrity is the cornerstone of the atherogenic process. Low-density lipoprotein (LDL) oxidation occurring within atheromatous plaques lead to deleterious vascular effects including endothelial cell cytotoxicity. The aim of this study was to evaluate the vascular antioxidant and cytoprotective effects of polyphenol-rich extracts from two medicinal plants from the Reunion Island: Antirhea borbonica (A. borbonica), Doratoxylon apetalum (D. apetalum). The polyphenol-rich extracts were obtained after dissolving each dry plant powder in an aqueous acetonic solution. Quantification of polyphenol content was achieved by the Folin–Ciocalteu assay and total phenol content was expressed as g gallic acid equivalent/100 g plant powder (GAE). Human vascular endothelial cells were incubated with increasing concentrations of polyphenols (1–50 µM GAE) before stimulation with oxidized low-density lipoproteins (oxLDLs). LDL oxidation was assessed by quantification of hydroperoxides and thiobarbituric acid reactive substances (TBARS). Intracellular oxidative stress and antioxidant activity (catalase and superoxide dismutase) were measured after stimulation with oxLDLs. Cell viability and apoptosis were quantified using different assays (MTT, Annexin V staining, cytochrome C release, caspase 3 activation and TUNEL test). A. borbonica and D. apetalum displayed high levels of polyphenols and limited LDL oxidation as well as oxLDL-induced intracellular oxidative stress in endothelial cells. Polyphenol extracts of A. borbonica and D. apetalum exerted a protective effect against oxLDL-induced cell apoptosis in a dose-dependent manner (10, 25, and 50 µM GAE) similar to that observed for curcumin, used as positive control. All together, these results showed significant antioxidant and antiapoptotic properties for two plants of the Reunion Island pharmacopeia, A. borbonica and D. apetalum, suggesting their therapeutic potential to prevent cardiovascular diseases by limiting LDL oxidation and protecting the endothelium.

Structural and Functional Changes of Reconstituted High-Density Lipoprotein (HDL) by Incorporation of α-synuclein: A Potent Antioxidant and Anti-Glycation Activity of α-synuclein and apoA-I in HDL at High Molar Ratio of α-synuclein

α-synuclein (α-syn) is a major culprit of Parkinson’s disease (PD), although lipoprotein metabolism is very important in the pathogenesis of PD. α-syn was expressed and purified using the pET30a expression vector from an E. coli expression system to elucidate the physiological effects of α-syn on lipoprotein metabolism. The human α-syn protein (140 amino acids) with His-tag (8 amino acids) was expressed and purified to at least 95% purity. Isoelectric focusing gel electrophoresis showed that the isoelectric point (pI) of α-syn and apoA-I were pI = 4.5 and pI = 6.4, respectively. The lipid-free α-syn showed almost no phospholipid-binding ability, while apoA-I showed rapid binding ability with a half-time (T1/2) = 8 ± 0.7 min. The α-syn and apoA-I could be incorporated into the reconstituted HDL (rHDL, molar ratio 95:5:1:1, palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC):cholesterol:apoA-I:α-syn with the production of larger particles (92 Å) than apoA-I-rHDL (86 and 78 Å) and α-syn-rHDL (65 Å). An rHDL containing both apoA-I and α-syn showed lower α-helicity around 45% with a red shift of the Trp wavelength maximum fluorescence (WMF) from 339 nm, while apoA-I-HDL showed 76% α-helicity and 337 nm of WMF. The denaturation by urea addition showed that the incorporation of α-syn in rHDL caused a larger increase in the WMF than apoA-I-rHDL, suggesting that the destabilization of the secondary structure of apoA-I by the addition of α-syn. On the other hand, the addition of α-syn induced two-times higher resistance to rHDL glycation at apoA-I:α-syn molar ratios of 1:1 and 1:2. Interestingly, low α-syn in rHDL concentrations, molar ratio of 1:0.5 (apoA-I:α-syn), did not prevent glycation with more multimerization of apoA-I. In the lipid-free and lipid-bound state, α-syn showed more potent antioxidant activity than apoA-I against cupric ion-mediated LDL oxidation. On the other hand, microinjection of α-syn (final 2 μM) resulted in 10% less survival of zebrafish embryos than apoA-I. A subcutaneous injection of α-syn (final 34 μM) resulted in less tail fin regeneration than apoA-I. Interestingly, incorporation of α-syn at a low molar ratio (apoA-I:α-syn, 1:0.5) in rHDL resulted destabilization of the secondary structure and impairment of apoA-I functionality via more oxidation and glycation. However, at a higher molar ratio of α-syn in rHDL (apoA-I:α-syn = 1:1 or 1:2) exhibited potent antioxidant and anti-glycation activity without aggregation. In conclusion, there might be a critical concentration of α-syn and apoA-I in HDL-like complex to prevent the aggregation of apoA-I via structural and functional enhancement.

In vitro antioxidant activity of meniran (Phyllantus urinaria) functional drink in human low density lipoprotein (LDL)

Preparation process for meniran (Phillantus urinaria) functional drink (MFD) influences its antioxidant activity. This research aims to understand the phenolic content, DPPH Radical Scavenging Activity (RSA), and LDL oxidation of MFD through various preparation processes. Those preparation processes included soaking fresh meniran (SFM), boiling fresh meniran for 5 minutes (BFM5’), boiling fresh meniran for 10 minutes (BFM10’), and soaking dried meniran (DM). The phenolic content was determined with Folin–Ciocalteu, antioxidant activity was assessed using DPPH and TBARS assay with LDL as the oxidation substrate. An antioxidant references in this research used ascorbic acid. The phenolic content in methods of SFM, BFM5’, BFM10’ and DM were 122±0.022, 182±0.043, 192 ±0.03, and 117 ±0.019 mg GAE/g of meniran respectively. Meanwhile, the DPPH RSA of SFM, BFM5’, BFM10’ and DM accounted for 82.18±0.35, 86.19±0.53, 86.75±0.64 and 69.96% respectively. As comparison, the DPPH RSA of ascorbic acid 50 ppm is 75.65±0.82%. At the same time the optimum inhibition of TBARS formation from BFM5’ and BFM10’ methods were 45.83 % and 48.66%, with MDA concentration in human LDL accounted for 38.30±2.39 and 36.30±1.82 nmol MDA/mg protein, respectively. As comparison, MDA concentration in human LDL added with ascorbic acid 25 ppm accounted for 41.35±2.41 nmol MDA/mg protein. In contrast, the control human LDL was 70.70±2.35 nmol MDA/mg protein. This study concludes that the BFM5’ and BFM10’ methods showed the highest antioxidant properties compared to other methods. All methods showed that MFD extract in concentration more than 25 ppm increased the concentration of MDA in human LDL. Therefore, to produce meniran functional drink in optimum antioxidant properties is best by using BFM5’ and BFM10’ preparation methods in meniran concentration of not more than 25 ppm.

Efektivitas Antioksidan dari Ekstrak Bunga Kasumba Turate (Carthamus tinctorius L.) dan Potensinya Sebagai Antihiperkolesterolemia

Telah dilakukan penelitian untuk menganalisis kandungan metabolit sekunder dan efektivitas antioksidan dari ektrak etanol bunga kasumba turate (Carthamus tinctorius L.), serta potensinya sebagai antihiperkolesterolemia. Bunga kasumba turate yang telah dikeringanginkan, dihaluskan dan dimaserasi dengan pelarut etanol 70%, lalu dievaporasi pelarutnya. Ekstrak etanol (EE) yang diperoleh dipartisi berturut-turut dengan pelarut n-heksana, etil asetat dan air, sehingga diperoleh ekstrak fraksi n-heksana (FH), etil asetat (FEA) dan air (FA). Selanjutnya, EE, FH, FEA, dan FA diuji kandungan metabolit sekundernya (metode Harborne) dan efektivitas antioksidannya (metode DPPH). Hasil penelitian menunjukkan bahwa EE dan FEA mengandung alkaloid, fenolik, flavonoid, saponin, tanin dan triterpenoid. FA mengandung fenolik, flavonoid, saponin, tanin dan triterpenoid. FH mengandung alkaloid, flavonoid dan triterpenoid. Efektivitas antioksidan bunga kasumba turate (dinyatakan dalam IC50) yang tertinggi pada FEA, diikuti FH, FA dan EE, dengan nilai IC50 53,59, 75,45, 77,43, dan 89.,9 µg/mL. Hasil kajian menunjukkan bahwa bunga kasumba turate dapat menghambat oksidasi LDL dan menurunkan kadar kolesterol darah. Penelitian ini menyimpulkan bahwa ekstrak bunga kasumba turate memiliki efektivitas antioksidan yang kuat dan efek antihiperkolesterolemia.Kata kunci: Antihiperkolesterolemia; antioksidan; Carthamus tinctorius L.; etanolAntioxidant Effectiveness of Kasumba Turate Flower Extract (Carthamus tinctorius L.) and Its potential as an AntihypercholesterolemiaABSTRACTResearch has been carried out to analyze the secondary metabolite content and antioxidant effectiveness of the ethanol extract of kasumba turate flower (Carthamus tinctorius L.), as well as its potential as antihypercholesterolemia. Kasumba turate flowers that have been dried and mashed, macerated with 70% ethanol solvent, then evaporated the solvent. The ethanol extract (EE) obtained was partitioned successively with n-hexane, ethyl acetate and water as solvents, so that the extracts of the n-hexane (FH), ethyl acetate (FEA) and water (FA) fractions were obtained. Furthermore, EE, FH, FEA, and FA were tested for their phytochemical content (Harbourne method) and antioxidant effectiveness (DPPH method). The results showed that EE and FEA contained alkaloids, phenolics, flavonoids, saponins, tannins and triterpenoids. FA contains phenolics, flavonoids, saponins, tannins and triterpenoids. FH contains alkaloids, flavonoids and triterpenoids. The antioxidant effectiveness of casumba turate flower (expressed in IC50) was highest in FEA, followed by FH, FA and EE, with IC50 values of 53,59, 75,45, 77,43, and 89,19 g/mL, respectively. The results of the literature review show that kasumba turate flowers can inhibit LDL oxidation and reduce blood cholesterol levels. This study concluded that kasumba turate flower extract has a strong antioxidant effectiveness and antihypercholesterolemic effect.Keywords: Antihypercholesterolemia; antioxidant; Carthamus tinctorius L.; ethanol

Dietary Flavonoids: Cardioprotective Potential with Antioxidant Effects and Their Pharmacokinetic, Toxicological and Therapeutic Concerns

Flavonoids comprise a large group of structurally diverse polyphenolic compounds of plant origin and are abundantly found in human diet such as fruits, vegetables, grains, tea, dairy products, red wine, etc. Major classes of flavonoids include flavonols, flavones, flavanones, flavanols, anthocyanidins, isoflavones, and chalcones. Owing to their potential health benefits and medicinal significance, flavonoids are now considered as an indispensable component in a variety of medicinal, pharmaceutical, nutraceutical, and cosmetic preparations. Moreover, flavonoids play a significant role in preventing cardiovascular diseases (CVDs), which could be mainly due to their antioxidant, antiatherogenic, and antithrombotic effects. Epidemiological and in vitro/in vivo evidence of antioxidant effects supports the cardioprotective function of dietary flavonoids. Further, the inhibition of LDL oxidation and platelet aggregation following regular consumption of food containing flavonoids and moderate consumption of red wine might protect against atherosclerosis and thrombosis. One study suggests that daily intake of 100 mg of flavonoids through the diet may reduce the risk of developing morbidity and mortality due to coronary heart disease (CHD) by approximately 10%. This review summarizes dietary flavonoids with their sources and potential health implications in CVDs including various redox-active cardioprotective (molecular) mechanisms with antioxidant effects. Pharmacokinetic (oral bioavailability, drug metabolism), toxicological, and therapeutic aspects of dietary flavonoids are also addressed herein with future directions for the discovery and development of useful drug candidates/therapeutic molecules.

The Efficacy of Black Chokeberry Fruits against Cardiovascular Diseases

Epidemiological studies have emphasized the association between a diet rich in fruits and vegetables and a lower frequency of occurrence of inflammatory-related disorders. Black chokeberry (Aronia melanocarpa L.) is a valuable source of biologically active compounds that have been widely investigated for their role in health promotion and cardiovascular disease prevention. Many in vitro and in vivo studies have demonstrated that consumption of these fruits is associated with significant improvements in hypertension, LDL oxidation, lipid peroxidation, total plasma antioxidant capacity and dyslipidemia. The mechanisms for these beneficial effects include upregulation of endothelial nitric oxide synthase, decreased oxidative stress, and inhibition of inflammatory gene expression. Collected findings support the recommendation of such berries as an essential fruit group in a heart-healthy diet. The aim of this review was to summarize the reports on the impact of black chokeberry fruits and extracts against several cardiovascular diseases, e.g., hyperlipidemia, hypercholesterolemia, hypertension, as well as to provide an analysis of the antioxidant and anti-inflammatory effect of these fruits in the abovementioned disorders.

Dietary Flavonoids: Cardioprotective Potential with Antioxidant Effects and Their Pharmacokinetic, Toxicological and Therapeutic Concerns

Flavonoids comprise a large group of structurally diverse polyphenolic compounds of plant origin and are abundantly found in human diet such as fruits, vegetables, grains, tea, dairy products, red wine and so on. Major classes of flavonoids include flavonols, flavones, flavanones, flavanols, anthocyanidins, isoflavones, and chalcones. Owing to their potential health benefits and medicinal significance, flavonoids are now considered as an indispensable component in a variety of medicinal, pharmaceutical, nutraceutical, and cosmetic preparations. However, flavonoids play a significant role in preventing cardiovascular diseases (CVDs), which could be mainly due to their antioxidant, antiatherogenic, and antithrombotic effects. Epidemiological and in vitro/in vivo evidences of antioxidant effects support the cardioprotective function of dietary flavonoids. Further, the inhibition of LDL oxidation and platelet aggregation following regular consumption of food containing flavonoids and moderate consumption of red wine might protect against atherosclerosis and thrombosis. A study suggests that daily intake of 100 mg of flavonoids through diet may reduce the risk of developing morbidity and mortality due to coronary heart disease (CHD) by approximately 10%. This review summarizes dietary flavonoids with their sources and potential health implications in CVDs including various redox-active cardioprotective (molecular) mechanisms with antioxidant effects. Pharmacokinetic (oral bioavailability, drug metabolism), toxicological and therapeutic aspects of dietary flavonoids are also addressed herein with future directions for the discovery and development of useful drug candidates/ therapeutic molecules.

The miR-378c-Samd1 circuit promotes phenotypic modulation of vascular smooth muscle cells and foam cells formation in atherosclerosis lesions

MicroRNAs have emerged as key regulators in vascular diseases and are involved in the formation of atherosclerotic lesions. However, the atherosclerotic-specific MicroRNAs and their functional roles in atherosclerosis are unclear. Here, we report that miR-378c protects against atherosclerosis by directly targeting Sterile Alpha Motif Domain Containing 1 (Samd1), a predicted transcriptional repressor. miR-378c was strikingly reduced in atherosclerotic plaques and blood of acute coronary syndrome (ACS) patients relative to healthy controls. Suppression of miR-378c promoted vascular smooth muscle cells (VSMCs) phenotypic transition during atherosclerosis. We also reported for the first time that Samd1 prolonged immobilization of LDL on the VSMCs, thus facilitated LDL oxidation and subsequently foam cell formation. Further, we found that Samd1 contains predicted DNA binding domain and directly binds to DNA regions as a transcriptional repressor. Together, we uncovered a novel mechanism whereby miR-378c-Samd1 circuit participates in two key elements of atherosclerosis, VSMCs phenotypic transition and LDL oxidation. Our results provided a better understanding of atherosclerosis pathophysiology and potential therapeutic management by targeting miR-378c-Samd1 circuit.