Fourteen quinolizidine derivatives, structurally related to the alkaloids lupinine and cytisine and previously studied for other pharmacological purposes, were presently tested for antiarrhythmic, and other cardiovascular effects on isolated guinea pig heart tissues in comparison to well-established reference drugs. According to their structures, the tested compounds are assembled into three subsets: (a) N-(quinolizidinyl-alkyl)-benzamides; (b) 2-(benzotriazol-2-yl)methyl-1-(quinolizidinyl)alkyl-benzimidazoles; (c) N-substituted cytisines. All compounds but two displayed antiarrhythmic activity that was potent for compounds 4, 1, 6, and 5 (in ascending order). The last compound (N-(3,4,5-trimethoxybenzoyl)aminohomolupinane) was outstanding, exhibiting a nanomolar potency (EC50 = 0.017 µM) for the increase in the threshold of ac-arrhythmia. The tested compounds shared strong negative inotropic activity; however, this does not compromise the value of their antiarrhythmic action. On the other hand, only moderate or modest negative chronotropic and vasorelaxant activities were commonly observed. Compound 5, which has high antiarrhythmic potency, a favorable cardiovascular profile, and is devoid of antihypertensive activity in spontaneously hypertensive rats, represents a lead worthy of further investigation.

Ellagitannins may have a beneficial impact in cardiovascular diseases. The aim of the study was to evaluate the effect of high‐fat diet (HFD) and the efficacy of Castanea sativa Mill. bark extract (ENC) on cardiac and vascular parameters. Rats were fed with regular diet, (RD, n = 15), HFD (n = 15), RD + ENC (20 mg/kg/day by gavage, n = 15), and HFD + ENC (same dose, n = 15) and the effects on body weight, biochemical serum parameters, and inflammatory cytokines determined. Cardiac functional parameters and aorta contractility were also assessed on isolated atria and aorta. Results showed that ENC reduced weight gain and serum lipids induced by HFD. In in vitro assays, HFD decreased the contraction force of left atrium, increased right atrium chronotropy, and decreased aorta K+‐induced contraction; ENC induced transient positive inotropic and negative chronotropic effects on isolated atria from RD and HFD rats and a spasmolytic effect on aorta. In ex vivo experiments, ENC reverted inotropic and chronotropic changes induced by HFD and enhanced Nifedipine effect more on aorta than on heart. In conclusion, ENC restores metabolic dysfunction and cardiac cholinergic muscarinic receptor function, and exerts spasmolytic effect on aorta in HFD rats, highlighting its potential as nutraceutical tool in obesity.

The research for innovative treatments against colon adenocarcinomas is still a great challenge. Acacia catechu Willd. heartwood extract (AC) has health-promoting qualities, especially at the gastrointestinal level. This study characterized AC for its catechins content and investigated the apoptosis-enhancing effect in human colorectal adenocarcinoma HT-29 cells, along with its ability to spare healthy tissue. MTT assay was used to describe the time course, concentration dependence and reversibility of AC-mediated cytotoxicity. Cell cycle analysis and AV-PI and DAPI-staining were performed to evaluate apoptosis, together with ROS formation, mitochondrial membrane potential (MMP) changes and caspase activities. Rat ileum and colon rings were tested for their viability and functionality to explore AC effects on healthy tissue. Quantitative analysis highlighted that AC was rich in (±)-catechin (31.5 ± 0.82 mg/g) and (−)-epicatechin (12.5 ± 0.42 mg/g). AC irreversibly decreased cell viability in a concentration-dependent, but not time-dependent fashion. Cytotoxicity was accompanied by increases in apoptotic cells and ROS, a reduction in MMP and increases in caspase-9 and 3 activities. AC did not affect rat ileum and colon rings’ viability and functionality, suggesting a safe profile toward healthy tissue. The present findings outline the potential of AC for colon cancer treatment.

The petals of the saffron crocus (Crocus sativus L.) are considered a waste material in saffron production, but may be a sustainable source of natural biologically active substances of nutraceutical interest. The aim of this work was to study the cardiovascular effects of kaempferol and crocin extracted from saffron petals. The antiarrhythmic, inotropic, and chronotropic effects of saffron petal extract (SPE), kaempferol, and crocin were evaluated through in vitro biological assays. The antioxidant activity of kaempferol and crocin was investigated through the 2 ',7 '-dichlorodihydrofluorescein diacetate (DCFH-DA) assay using rat cardiomyoblast cell line H9c2. The MTT assay was applied to assess the effects of kaempferol and crocin on cell viability. SPE showed weak negative inotropic and chronotropic intrinsic activities but a significant intrinsic activity on smooth muscle with a potency on the ileum greater than on the aorta: EC50 = 0.66 mg/mL versus EC50 = 1.45 mg/mL. Kaempferol and crocin showed a selective negative inotropic activity. In addition, kaempferol decreased the contraction induced by KCl (80 mM) in guinea pig aortic and ileal strips, while crocin had no effect. Furthermore, following oxidative stress, both crocin and kaempferol decreased intracellular ROS formation and increased cell viability in a concentration-dependent manner. The results indicate that SPE, a by-product of saffron cultivation, may represent a good source of phytochemicals with a potential application in the prevention of cardiovascular diseases.

Our research groups have been involved for many years in studies aimed at identifying new active organic compounds endowed with pharmacological properties. In this work, we focused our attention on the evaluation of cardiovascular and molecular drug resistance (MDR) reverting activities of some nitrosubstituted sulphur-containing heterocycles. Firstly, we have examined the effects of 4-nitro-3-(4-methylphenyl)-3,6-dihydro-2H-thiopyran S,S-dioxide 5, and have observed no activity. Then we have extended our investigation to the 3-aryl-4-nitrobenzothiochromans S,S-dioxide 6 and 7, and have observed an interesting biological profile. Cardiovascular activities were assessed for all compounds using ex vivo studies, while the MDR reverting effect was evaluated only for selected compounds using tumor cell lines. All compounds were shown to affect cardiovascular parameters. Compound 7i exerted the most effect on negative inotropic activity, while 6d and 6f could be interesting molecules for the development of more active ABCB1 inhibitors. Both 6 and 7 represent structures of large possible biological interest, providing a scaffold for the identification of new ABCB1 inhibitors.

Oxidative stress (OS) arising from tissue redox imbalance, critically contributes to the development of neurodegenerative disorders. Thus, natural compounds, owing to their antioxidant properties, have promising therapeutic potential. Pres phytum (PRES) is a nutraceutical product composed of leaves- and flowers-extracts of Olea europaea L. and Hibiscus sabdariffa L., respectively, the composition of which has been characterized by HPLC coupled to a UV-Vis and QqQ-Ms detector. As PRES possess antioxidant, antiapoptotic and anti-inflammatory properties, the aim of this study was to assess its neuroprotective effects in human neuroblastoma SH-SY5Y cells and in rat brain slices subjected to OS. PRES (1–50 µg/mL) reverted the decrease in viability as well as the increase in sub-diploid-, DAPI-and annexin V-positive-cells, reduced ROS formation, recovered the mitochondrial potential and caspase-3 and 9 activity changes caused by OS. PRES (50–100 µg/mL) neuroprotective effects occurred also in rat brain slices subjected to H2O2 challenge. Finally, as the neuroprotective potential of PRES is strictly related to its penetration into the brain and a relatively good pharmacokinetic profile, an in-silico prediction of its components drug-like properties was carried out. The present results suggest the possibility of PRES as a nutraceutical, which could help in preventing neurodegenerative diseases.

A new Thymus vulgaris L. solid essential oil (SEO) formulation composed of liquid EO linked to solid excipients has been chemically analysed and evaluated for its intestinal spasmolytic and antispastic effects in ex vivo ileum and colon of guinea pig and compared with liquid EO and excipients. Liquid EO and solid linked EO were analysed by original capillary electrochromatography coupled to diode array detection (CEC-DAD) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodologies. The main bioactive constituents are thymol and carvacrol, with minor constituents for a total of 12 selected analysed compounds. Liquid EO was the most effective in decreasing basal contractility in ileum and colon; excipients addiction permitted normal contractility pattern in solid linked EO SEO. In ileum and colon, the Thymus vulgaris L. solid formulation exerted the relaxant activity on K+-depolarized intestinal smooth muscle as well as liquid EO. The solid essential oil exhibits antimicrobial activity against different strains (Staphylococcus aureus, Streptococcus pyogenes, Pseudomonas aeruginosa, Escherichia coli, Salmonella Thyphimurium, Candida albicans) similarly to liquid oil, with activity against pathogen, but not commensal strains (Bifidobacterium Breve, Lactobacillus Fermentum) in intestinal homeostasis. Therefore, Thymus vulgaris L. solid essential oil formulation can be proposed as a possible spasmolytic and antispastic tool in medicine.

Hypertension has become the leading risk factor for worldwide cardiovascular diseases. Conventional pharmacological treatment, after both dietary and lifestyle changes, is generally proposed. In this review, we present the antihypertensive properties of phytocomplexes from thirteen plants, long ago widely employed in ethnomedicines and, in recent years, increasingly evaluated for their activity in vitro and in vivo, also in humans, in comparison with synthetic drugs acting on the same systems. Here, we focus on the demonstrated or proposed mechanisms of action of such phytocomplexes and of their constituents proven to exert cardiovascular effects. Almost seventy phytochemicals are described and scientifically sound pertinent literature, published up to now, is summarized. The review emphasizes the therapeutic potential of these natural substances in the treatment of the 'high normal blood pressure' or 'stage 1 hypertension', so-named according to the most recent European and U.S. guidelines, and as a supplementation in more advanced stages of hypertension, however needing further validation by clinical trial intensification.

The effects of Castanea sativa Mill. have been studied in high fat diet (HFD) overweight rats. Natural Extract of Chestnut bark (Castanea sativa Mill.) (ENC®), rich in ellagitannins, has been studied in 120 male Sprague-Dawley rats, divided in four groups. Two groups were controls: regular (RD) and HDF diet. Two groups received ENC® (20 mg/kg/day): RD + ENC® and HFD + ENC®. At baseline and at 7, 14 and 21 days, weight gain, serum lipids, plasma cytokines, liver histology, microsomial enzymes and oxidation, intestinal oxidative stress and contractility were studied. HFD increased body weight, increased pro-inflammatory cytokines, induced hepatocytes microvescicular steatosis, altered microsomial, increased liver and intestinal oxidative stress, deranged intestinal contractility. In HFD-fed rats, ENC® exerted antiadipose and antioxidative activities and normalized intestinal contractility, suggesting a potential approach to overweight management associated diseases.

Castanea sativa Mill. wood extract (ENC) has health-promoting qualities such as antibacterial, antiviral, antispasmodic, cardio- and neuroprotective-activities. Its potential towards cancer cells of the central nervous system, however, is still unexplored. This study investigates the apoptotic-enhancing effect of ENC in human neuroblastoma SH-SY5Y cell line. This extract irreversibly decreased cell viability with IC 50 of 142.8±20.2, 109.3±16.6 and 82.5±8.6 μg/mL for 24h, 48 and 72h of treatment, respectively. This was accompanied by cell shrinkage and tendency to round-up. A concentration-dependent increase in sub G0/G1 cells as well as a rise in cells with nuclear fragmentation and chromatin condensation, both typical of apoptosis, was also observed. In conclusion, the present findings provide initial data on the potential of ENC for neuroblastoma treatment and offer the rationale for further studies aimed at elucidating the full mechanism(s) underlying its effects.

BACKGROUND:Constitutive (agonist-independent) activity is a prerogative of many G protein-coupled receptors (GPCRs) including α1-adrenoceptors (α1-ARs). Inhibition of such an activity at α1-AR subtypes by antagonists with negative efficacy is difficult to be adequately tested.METHODS:In the present experimental approach, we compared the activity of three calcium channel blockers (nifedipine, diltiazem and verapamil) and of three potent benzodioxane-based α1-AR antagonists, differing for subtype selectivity and inverse agonist properties, in producing smooth muscle relaxation and negative inotropy under the same test conditions. We selected, as benzodioxane derivatives, (S)-WB4101, inverse agonist with slight α1A/α1B-α1D AR selectivity, and two previously developed analogues. Both of these are potent antagonists at α1D-AR, that is the α1- AR subtype suspected of the highest susceptibility to inverse agonists for its high degree of basal activity, but only one is inverse agonist.RESULTS:We found that all the three benzodioxane-related α1-AR antagonists have significant intrinsic relaxant activity on non-vascular smooth muscle and moderate negative inotropic effect, while they do not relax aorta. Their potency is always lower than that of three calcium channel blockers.CONCLUSIONS:Intrinsic myorelaxant and negative inotropic activity of the three benzodioxane-based α1-AR antagonist is related neither to a particular profile of α1-AR subtype selectivity nor to whether or not being an inverse agonist, but it parallels the calcium antagonists effects indicating a direct interaction of the three α1-AR antagonists with L-type Ca2+ channels.

Acacia catechu Willd. is a plant diffused in India and other Asian countries, where it is used as a traditional medicine for the treatment of several ailments including diarrhea, one of the most common pathologies worldwide. In this study, we determined the chemical composition of Acacia catechu Willd. extract (AC) and evaluated its effect on spontaneous and induced contractility in isolated guinea pig ileum and proximal colon. Preliminary data about its antimicrobial effect against some pathogen agents versus some microbiota intestinal strain have been also reported. Chemical analysis revealed the presence of catechins, such as (-)-Epicatechin and (+)-Catechin. AC extract reduced frequency and amplitude of colon smooth muscle spontaneous contractility, in a concentration-dependent manner. A weaker effect of the extract was exerted toward ileum smooth muscle spontaneous contractility. The observed calcium antagonistic effect was more potent in proximal colon than in ileum. The extract showed a noncompetitive reversible antagonism to carbachol, both in proximal colon and ileum, with a higher potency in proximal colon. The antimicrobial effects of AC extract were observed toward Campylobacter jejuni, Escherichia coli, and Salmonella spp., while Bifido and Lactobacillus were not affected by treatment. These effects, however, occurred at concentrations fivefold higher than those inhibiting ileum and colon contractility. In conclusion, our results suggest that AC affects intestinal contractility without affecting intestinal bacterial flora and this may result in clinical benefits in patients suffering from nonbacterial diarrhea.

The high incidence of vulvo-vaginal candidiasis, combined with the growing problems about azole resistance and toxicity of antifungal drugs, highlights the need for the development of new effective strategies for the treatment of this condition. In this context, natural compounds represent promising alternatives. The cyanobacterium Spirulina platensis, a bluegreen alga, exhibits antimicrobial activities against several microorganisms. Nevertheless, only few data about the antifungal properties of Spirulina platensis are available and its potential toxic effects have not been largely investigated. The aim of this study was to evaluate the in vitro activity of a fully-characterized water extract of Spirulina platensis against 22 strains of Candida spp. Prior to considering its potential topical use, we both investigated whether the extract exerted target activities on guinea pig uterine smooth muscle, and the impact of Spirulina platensis on the dominant microorganisms of the vaginal microbiota (i. e., lactobacilli), in order to exclude possible adverse events. By means of a broth microdilution assay, we found that the microalga extract possesses good antifungal properties (MIC: 0.125-0.5 mg/ml), against all the Candida species with a fungicidal activity. At the concentrations active against candida, Spirulina platensis did not modify the spontaneous basic waves pattern of uterine myometrium as underlined by the absence of aberrant contractions, and did not affect the main health-promoting bacteria of the vaginal ecosystem. Finally, we evaluated the selectivity index of our extract by testing its cytotoxicity on three different cell lines and it showed values ranging between 2 and 16. Further in vivo studies are needed, in particular to evaluate the use of control-release formulations in order to maintain Spirulina platensis concentrations at anti-Candida active doses but below the toxic levels found in the present work.

Castanea sativa Mill (ENC®), containing tannins against 33 Chlamydia strains, was compared to SMAP-29 with inhibitory effect against C. trachomatis and C. pneumoniae. The ENC® activity against Chlamydia spp. was evaluated determining the lowest concentration to achieve more than half reduction of intact chlamydial inclusions versus controls. ENC® reduced all Chlamydia strains tested at 1 µg/mL, while SMAP-29 induced reductions of C. trachomatis and C. pneumoniae infectivity at 10 µg/mL. A great reduction of C. trachomatis, C. pneumoniae, and C. abortus infectivity was achieved with a 10 µg/mL ENC® concentration, whereas their infectivity was almost inhibited at 100 µg/mL ENC® concentration.

Olea europaea L. leaves extract (Oe) and Hybiscus sabdariffa L. flowers extract (Hs) have calcium antagonistic properties. Aim of this work was to study the cardiovascular effects of Pres Phytum(®), a nutraceutical formulation containing a mixture of the two extracts and the excipients, and investigate its possible off-target effects, using in vitro biological assays on guinea pig isolated organs. Cardiovascular effects were assessed using guinea pig atria and aorta. The effects of Pres Phytum on spontaneous gastrointestinal, urinary, and respiratory tracts smooth muscle contractility were evaluated. Pres Phytum exerted a vasorelaxant effect (IC50 = 2.38 mg/mL) and a negative chronotropic effect (IC50 = 1.04 mg/mL) at concentrations lower than those producing smooth muscle spontaneous contractility alterations in the other organs. Compared to Pres Phytum, the mixture did not exert negative inotropic activity, while it maintained a negative chronotropic efficacy (IC50 = 1.04 mg/mL). These experimental data suggest a possible nutraceutical use of this food supplement for the management of preclinical hypertension.

We present a series of oxadiazolothiazinones, selective inotropic agents on isolated cardiac tissues, devoid of chronotropy and vasorelaxant activity. Functional and binding data for the precursor of the series (compound 1) let us hypothesize LTCC blocking activity and the existence of a recognition site specific for this scaffold. We synthesized and tested 22 new derivatives: introducing a para-methoxyphenyl at C-8 led to compound 12 (EC50 = 0.022 μM), twice as potent as its para-bromo analogue (1). For 10 analogues, we extended the characterization of the biological properties by including the assessment of metabolic stability in human liver microsomes and cytochrome P450 inhibition potential. We observed that the methoxy group led to active compounds with low metabolic stability and high CYP inhibition, whereas the protective effect of bromine resulted in enhanced metabolic stability and reduced CYP inhibition. Thus, we identified two para-bromo benzothiazino-analogues as candidates for further studies.

G infections constitute important emerging and re-emerging infective worldwide diseases. They are mostly endemic and show a heterogeneous aetiology. Most water-borne diseases caused by microorganisms induce diarrhoea and determine about 5 million deaths per year. The research on anti-diarrheal tools should be focused on the evaluation of substances and chemically charachterized phytocomplexes able to affect intestinal motility and to exert a prebiotic action. Several plants, such as Castanea sativa Mill., Sanseviera liberica Gerome & Labroy, have been shown to inhibit gut peristalsis, through several mechanisms. Furthermore, disparate classes of natural compounds including hydrolysable tannins and flavonoids, restore intestinal functionality, affecting different molecular networks influencing each other’s. Acacia catechu Willd extract (ACE) has been used in Indian Traditional Medicine to manage several diseases including diarrhoea and other gastrointestinal ailments. This extract was shown to contain high amounts of flavonoids, in particular flavan-3-ols. Furthermore, in vitro biological assays were exerted, using tissues from guinea pigs, to assess ACE effects towards induced and spontaneous intestinal smooth muscle contractility. The results demonstrated that ACE reducecs spontaneous and induced colon and ileal smooth muscle contractility via inhibiting muscarinic and histaminergic receptors. Also ACE effects against several pathogenic and non pathogenic bacteria were tested, showing a selective antibacterial activity towards pathogenic strains including, Staphylococcus aureus, Gram-negative Escherichia coli, Salmonella spp., Campilobacter, without inhibiting. These findings suggest that Acacia may represent a nutraceutical option to manage diarrheal infectious and non infectious disesases.