Monika E. Czerwińska

Oleuropein and oleacein may restore biological functions of endothelial progenitor cells impaired by angiotensin II via activation of Nrf2/heme oxygenase-1 pathway

Endothelial progenitor cells (EPCs) are responsible for neovascularization of ischaemic tissue and may participate in re-endothelization of an injured arterial wall. There is evidence that angiotensin II, by an increase of gp91phox expression and induction of ROS generation, accelerates cell senescence and impairs functions of EPCs. Oleacein is a main phenolic compound from olive oil, whereas oleuropein is present in olive leaves. Both compounds possess antioxidative, hypotensive and anti-inflammatory properties and show beneficial activity on the cardiovascular system. In this study, we examined whether oleoeuropein and oleacein could protect EPCs against impairment of their functions due to angiotensin-induced cell senescence. CD31(+)/VEGFR-2(+) cells were isolated from young healthy volunteers blood samples and cultured on fibronectin-coated plates with angiotensin (1.0μM) in presence or absence of increasing concentrations (from 1.0 to 10.0 μM) of oleoeuropein or oleacein. As compared to angiotensin II-treated cells, EPCs exposed to oleacein or oleuropein prior to angiotensin II showed a significant increase of proliferation and telomerase activity, and a decrease in the percentage of senescent cells and intracellular ROS formation. Oleacein and oleuropein restored migration, adhesion and tube formation of EPCs diminished by angiotensin II in a concentration-dependent manner. This effect was related to NF-E2-related factor 2 (Nrf2) transcription factor activation and the increase of heme oxygenase-1 (HO-1) expression.

Oenothera paradoxa defatted seeds extract and its bioactive component penta-O-galloyl-β-D-glucose decreased production of reactive oxygen species and inhibited release of leukotriene B4, interleukin-8, elastase, and myeloperoxidase in human neutrophils.

In this study, we analyzed ex vivo the effect of an aqueous extract of Oenothera paradoxa defatted seeds on the formation of neutrophil-derived oxidants. For defining active compounds, we also tested lypophilic extract constituents such as gallic acid, (+)-catechin, ellagic acid, and penta-O-galloyl-β-D-glucose and a hydrophilic fraction containing polymeric procyanidins. The anti-inflammatory potential of the extract and compounds was tested by determining the release from activated neutrophils of elastase, myeloperoxidase, interleukin-8 (IL-8), and leukotriene B4 (LTB4), which are considered relevant for the pathogenesis of cardiovascular diseases. The extract of O. paradoxa defatted seeds displays potent antioxidant effects against both 4β-phorbol-12β-myristate-α13-acetate- and formyl-met-leu-phenylalanine-induced reactive oxygen species production in neutrophils with IC50 values around 0.2 μg/mL. All types of polyphenolics present in the extract contributed to the extract antioxidant activity. According to their IC50 values, penta-O-galloyl-β-D-glucose was the more potent constituent of the extract. In cell-free assays, we demonstrated that this effect is partially due to the scavenging of O2- and H2O2 oxygen species. The extract and especially penta-O-galloyl-β-D-glucose significantly inhibit elastase, myeloperoxidase IL-8, and LTB4 release with an IC50 for penta-O-galloyl-β-D-glucose of 17±1, 15±1, 6.5±2.5, and around 20 μM, respectively. The inhibition of penta-O-galloyl-β-D-glucose on reactive oxygen species and especially on O2- production, myeloperoxidase, and chemoattractant release may reduce the interaction of polymorphonuclear leukocyte with the vascular endothelium and by that potentially diminish the risk of progression of atherosclerosis development.

Chemical Composition, Antioxidative and Anti-Inflammatory Activity of Extracts Prepared from Aerial Parts of Oenothera biennis L. and Oenothera paradoxa Hudziok Obtained after Seeds Cultivation

In the present study we investigated the chemical composition of extracts prepared from aerial parts of Oenothera paradoxa Hudziok and Oenothera biennis L. and their antioxidative and anti-inflammatory activities. Ultra high pressure liquid chromatography (UHPLC)-DAD-MS/MS studies showed that both extracts contain a wide variety of polyphenols (39 identified constituents) among which macrocyclic ellagitannin turned out to be the main constituent. During the in vitro studies, using noncellular models, both extracts scavenged reactive oxygen species (ROS) in a concentration-dependent manner, and the lowest SC(50) values were obtained for O(2)(-) and H(2)O(2). Both extracts inhibited ROS production by stimulated human neutrophils. The stronger activity in the case of formyl-met-leu-phenylalanine stimulation suggests that both extracts may act through the receptor-dependent pathway. O. paradoxa extract and O. biennis extract exhibited anti-inflammatory activity by the inhibition of hyaluronidase and lipoxygenase in a concentration-dependent manner. The stronger activity of O.biennis extract toward lipoxygenase may be explained by its higher oenothein B content.

Antioxidant and anti-inflammatory flavonol glucuronides from Polygonum aviculare L.

A series of 11 flavonol glucuronides were isolated from the herb of Polygonum aviculare L. (Ph.Eur) of which 8 were reported for the first time from the Polygonum species. Three acetylated kaempferol and isorhamnetin glucuronides were isolated from a natural source for the first time. All compounds, including the new ones, were characterized using 1D and 2D NMR spectra as well as high resolution mass spectrometry. The influence of all isolated compounds on the production of reactive oxygen species, as well as on elastase release by human neutrophils, was evaluated in in vitro studies. The results showed that all investigated compounds at physiologically achievable concentrations within the range of 0.5-10 μM significantly inhibited the production of reactive oxygen species as well as elastase release in human neutrophils model and should be considered as responsible for anti-inflammatory activity of the P. aviculare herb. The chemotaxonomic value of isolated compounds was also discussed.

Phytochemistry, pharmacology and traditional uses of different Epilobium species (Onagraceae): A review

ETHNOPHARMACOLOGICAL RELEVANCE The Epilobium genus (willowherb) comprises of ca. 200 species of herbaceous plants distributed around the world. Infusions prepared form willowherbs have been traditionally used externally in skin and mucosa infections and in the treatment of benign prostate hyperplasia. Nowadays extracts from different Epilobium species are widely used by patients, however the lack of clinical studies does not allow to fully establish their efficacy. The present review summarizes published data on phytochemistry, ethnopharmacological use and pharmacological studies concerning willowherb species investigated throughout past few decades. MATERIALS AND METHODS Literature survey was performed using Scopus, PubMed, Web of Science and Reaxys databases looking for papers and patents focused on chemical composition and bioactivity of Epilobium species. Systematic research in ethnopharmacological literature in digitalized sources of academic libraries was also carried out. RESULT The chemical composition of different Epilobium species and their bioactivities are described. The detailed information on constituents isolated and detected by chromatographic methods is given. The studies show that polyphenols are main compounds occurring in Epilobium herb among which flavonoids, phenolic acids and tannins (oenothein B and oenothein A) are dominating constituents. The extracts and some isolated compounds from Epilobium sp. were shown to possess antimicrobial, anti-proliferative, anti-inflammatory, analgesic and antioxidative activities. Because many studies suggest that oenothein B as dominating constituent may be responsible for Epilobium sp. pharmacological effects, its documented bioactivities were also described. CONCLUSIONS The pharmacological studies performed on Epilobium justify the traditional use of this species in external and in gastrointestinal inflammations. As far as the treatment of benign prostate hyperplasia (BPH) is considered, in the literature, there are some reports indicating that Epilobium extracts have a beneficial effect for this disorder, but the number of in vitro studies is not sufficient and the in vivo studies are not conclusive or too preliminary to draw a final conclusion about the efficacy of Epilobium preparations. More in vitro, in vivo and clinical studies to confirm this mode of action are strongly needed. Epilobiums extracts have also documented antioxidative and potential anti-inflammatory properties. Oenothein B can be considered as responsible for some of Epilobium pharmacological properties. Because of the lack of clinical data further studies are needed to provide an evidence base for traditional uses of plant materials belonging to the Epilobium genus.

Components of a standardised olive leaf dry extract (Ph. Eur.) promote hypothiocyanite production by lactoperoxidase.

We investigated in vitro the ability of a standardised olive leaf dry extract (Ph. Eur.) (OLE) as well as of its single components to circumvent the hydrogen peroxide-induced inhibition of the hypothiocyanite-producing activity of lactoperoxidase (LPO). The rate of hypothiocyanite (⁻OSCN) formation by LPO was quantified by spectrophotometric detection of the oxidation of 5-thio-2-nitrobenzoic acid (TNB). By using excess hydrogen peroxide, we forced the accumulation of inactive enzymatic intermediates which are unable to promote the two-electronic oxidation of thiocyanate. Both OLE and certain extract components showed a strong LPO-reactivating effect. Thereby an o-hydroxyphenolic moiety emerged to be essential for a good reactivity with the inactive LPO redox states. This basic moiety is found in the main OLE components oleuropein, oleacein, hydroxytyrosol, caffeic acid as well as in different other constituents including the OLE flavone luteolin. As LPO is a key player in the humoral immune response, these results propose a new mode of action regarding the well-known bacteriostatic and anti-inflammatory properties of the leaf extract of Olea europaea L.

Inhibition of human neutrophils NEP activity, CD11b/CD18 expression and elastase release by 3,4-dihydroxyphenylethanol-elenolic acid dialdehyde, oleacein

Polyphenols, such as oleacein (3,4-DHPEA-EDA; 3,4-dihydroxyphenylethanol-elenolic acid dialdehyde), are believed to play a role in the prevention of cardiovascular diseases. Due to an increase of neutrophil mediators in acute myocardial infarction the aim of this study was to establish the effect of oleacein on neutral endopeptidase (NEP) activity and other functions of human neutrophils, such as elastase, MMP-9 and IL-8 production. The effect on CD62L and CD11b/CD18 expression on neutrophils was also determined. Oleacein with a concentration of 100 μM inhibited NEP activity, elastase, MMP-9 and IL-8 release from neutrophils by 77.7 ± 2.7%, 21.3 ± 7.8%, 22.7 ± 4.2% and 25.2 ± 5.6%, respectively. Oleacein with a concentration of 50 μM suppressed CD11b/CD18 expression by 63.6 ± 3.1% and to a lesser extent, increased CD62L expression by 27.3 ± 8.3% on the surface of neutrophils, in comparison with stimulated cells. Oleacein by inhibiting NEP activity, adhesion molecules expression and elastase release might play a role in the protective effects of olive oil against endothelial injuries.

Effects of an Aqueous Extract from Leaves of Ligustrum vulgare on Mediators of Inflammation in a Human Neutrophils Model

Leaves of Ligustrum vulgare (common privet) have been used for treatment of oropharyngeal inflammations or as antirheumatic, diuretic, and hypotensive agents in folk medicine in southern Europe. Taking into account that neutrophils are involved in the inflammation, the aim of the study was to determine the effect of an aqueous extract prepared from leaves of Ligustrum vulgare on neutrophil functions. The extract was characterized by the HPLC-DAD-MSn method. The inhibition of reactive oxygen species production by formyl-met-leu-phenylalanine- or phorbol 12-myristate 13-acetate-stimulated neutrophils was determined using luminol- or lucigenin-dependent chemiluminescence. The effect on myeloperoxidase, metalloproteinase 9, and interleukin 8 production by neutrophils was measured by an enzyme-linked immunosorbent assay. Neutrophil elastase release was established spectrophotometrically. The expression of adhesion molecules on neutrophils was analyzed with flow cytometry. The main compounds detected were flavonoids, phenylpropanoids, hydroxycinnamates, and secoiridoids. The inhibition of oxidative burst by the extract was comparable in both stimuli models (formyl-met-leu-phenylalanine: IC50 = 18.2 ± 4.0 µg/mL; phorbol 12-myristate 13-acetate: IC50 = 19.8 ± 3.0 µg/mL). The extract in the concentration range of 5-50 µg/mL inhibited neutrophil elastase release by 23.9-34.1 % and myeloperoxidase release by 24.2-37.4 %. The inhibitory effect on metalloproteinase 9 and interleukin 8 production was around 20 %. The extract in the highest concentration modulated the expression of L-selectin and β2 integrin. Our results partly support the traditional use of common privet leaves as an anti-inflammatory agent.

Oleacein enhances anti-inflammatory activity of human macrophages by increasing CD163 receptor expression.

BACKGROUND Oleacein (dialdehydic form of decarboxymethyl elenolic acid linked to hydroxytyrosol; 3,4-DHPEA-EDA) have been proven to possess antioxidant and anti-inflammatory activity. PURPOSE In this study, we examined whether oleacein could increase CD163 and IL-10 receptor expression as well as HO-1 intracellular secretion in human macrophages. METHODS Effect of oleacein (10 and 20 μmol/l) or oleacein together with complexes of haemoglobin (Hb) and haptoglobin 1-1 (Hp11) or haptoglobin 2-2 (Hp22) on expression of IL-10 and CD163 receptor was determined by Flow Cytometry. Expression of CD163mRNA was measured by real-time quantitative RT-PCR. Heme oxygenase 1 (HO-1) intracellular secretion in macrophages was investigated by enzyme-linked immunosorbent assay (ELISA). RESULTS Oleacein (OC) together with complexes HbHp11 or HbHp22 stimulated the expression of CD163 (30-100-fold), IL-10 (170-300-fold) and HO-1 secretion (60-130-fold) after 5 days of coincubation. The 2-fold (24 h), 4-fold (48 h) increase of CD163 mRNA level and its final (72 h) decrease was also observed. CONCLUSION Our results suggested that oleacein enhances anti-inflammatory activity of complexes haemoglobin with haptoglobin 1-1 and 2-2 and could play a potential role in the prevention of inflammatory disease related to atherosclerosis.

Quantitative Determination of Secoiridoids and Phenylpropanoids in Different Extracts of Ligustrum Vulgare L. Leaves by a Validated HPTLC–Photodensitometry Method

INTRODUCTION The genus Ligustrum (Oleaceae) is distributed in Europe and Asia (south China and Korea), where it is used to prevent hypertension, sore throats, inflammation and diabetes. The main groups of compounds in extracts of Ligustrum vulgare are biologically active secoiridoids and phenylpropanoids. OBJECTIVES The aim of the study was primarily the development and validation of a HPTLC-photodensitometry method for separation and determination of secoiridoids (oleacein, oleuropein) and phenylpropanoids (echinacoside) in different extracts prepared from leaves of L. vulgare. A secondary issue was the quantitative screening of oleacein, oleuropein and echinacoside in extracts from leaves collected at different stages of plant growth (from May to September). METHODS A HPTLC-photodensitometry method was developed and validated for quantification of oleuropein, oleacein and echinacoside in plant extracts (aqueous and ethanolic extract, decoction, infusion). Silica gel was used as the stationary phase and dichloromethane:methanol:formic acid:water (80:25:1.5:4, v/v/v/v) as the mobile phase. RESULTS The HPTLC-photodensitometry method developed for quantification of oleacein, oleuropein and echinacoside was specific, accurate and precise. The presence of oleacein was detected in aqueous extracts, whereas oleuropein was present, in particular, in ethanolic extracts, decoctions and infusions. Echinacoside was detected in all the extracts prepared. The content of secoiridoids was variable from May to September, whereas the amount of echinacoside increased in this term. CONCLUSION The developed and validated HPTLC-photodensitometry method allowed performing fast screening of quantitative profiles of oleacein, oleuropein and echinacoside in preparations of privet leaves.