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The intestines are recognized as the main source of chronic inflammation in chronic kidney disease (CKD) and, among other cells, macrophages are involved in modulating this process as well as in the impaired immune response which also occurs in CKD patients. In this study, we evaluated the effect of Indoxyl Sulfate (IS), a protein bound uremic toxin poorly eliminated by hemodialysis, on inflammatory, oxidative stress and pro-apoptotic parameters, at the intestinal level in mice, on intestinal epithelial cells (IEC-6) and on primary murine peritoneal macrophages. C57BL/6J mice were treated with IS (800 mg/kg i.p.) for 3 or 6 h and histopathological analysis showed that IS induced intestinal inflammation and increased cyclooxygenase-2 (COX-2), nitrotyrosine and Bax expression in intestinal tissue. In IEC-6 cells, IS (125&ndash, 1000 &micro, M) increased tumor necrosis factor-&alpha, levels, COX-2 and inducible nitric oxide synthase expression and nitrotyrosine formation. Moreover, IS increased pro-oxidant, pro-inflammatory and pro-apoptotic parameters in peritoneal macrophages from IS-treated mice. Also, the serum concentration of IS and pro-inflammatory levels of cytokines resulted increased in IS-treated mice. Our results indicate that IS significantly contributes to affect intestinal homeostasis, immune response, and to induce a systemic pro-inflammatory state thus highlighting its potential role as therapeutic target in CKD patients.

Natural products black cumin&mdash, Nigella sativa (N. sativa) and wild garlic&mdash, Allium ursinum (AU) are known for their potential role in reducing cardiovascular risk factors, including antracycline chemotherapy. Therefore, this study investigates the effect of N. sativa and AU water and methanolic extracts in a cellular model of doxorubicin (doxo)-induced cardiotoxicity. The extracts were characterized using Ultraviolet-visible (UV-VIS) spectroscopy, Fourier-transform infrared (FT-IR) spectroscopy, Liquid Chromatography coupled with Mass Spectrometry (LC-MS) and Gas Chromatography coupled with Mass Spectrometry (GC-MS) techniques. Antioxidant activity was evaluated on H9c2 cells. Cytosolic and mitochondrial reactive oxygen species (ROS) release was evaluated using 2&prime, 7&prime, dichlorofluorescin-diacetate (DHCF-DA) and mitochondria-targeted superoxide indicator (MitoSOX red), respectively. Mitochondrial membrane depolarization was evaluated by flow cytometry. LC-MS analysis identified 12 and 10 phenolic compounds in NSS and AU extracts, respectively, with flavonols as predominant compounds. FT-IR analysis identified the presence of carbohydrates, amino acids and lipids in both plants. GC-MS identified the sulfur compounds in the AU water extract. N. sativa seeds (NSS) methanolic extract had the highest antioxidant activity reducing both intracellular and mitochondrial ROS release. All extracts (excepting AU methanolic extract) preserved H9c2 cells viability. None of the investigated plants affected the mitochondrial membrane depolarization. N. sativa and AU are important sources of bioactive compounds with increased antioxidant activities, requiring different extraction solvents to obtain the pharmacological effects.

Oxidative stress is widely accepted as a key factor of doxorubicin (Doxo)-induced cardiotoxicity. There is evidence to indicate that nitrosative stress is involved in this process, and that Doxo interacts by amplifying cell damage. Mitochondrial connexin 43 (mitoCx43) can confer cardioprotective effects through the reduction of mitochondrial reactive oxygen species production during Doxo-induced cardiotoxicity. The present study aimed to evaluate the involvement of mitoCx43 in Doxo-induced nitrosative stress. Rat H9c2 cardiomyoblasts were treated with Doxo in the absence or presence of radicicol, an inhibitor of Hsp90, the molecular chaperone involved in Cx43 translocation to the mitochondria that underlies its role in cardioprotection. FACS analysis and RT-qPCR revealed that Doxo increased superoxide dismutase, and catalase gene and protein expression. As shown by hypodiploid nuclei and confirmed by western blot analysis, Doxo increased caspase 9 expression and reduced procaspase 3 levels, which induced cell death. Moreover, a significant increase in the activation of the NF-κB signaling pathway was observed. It is well known that the increased expression of inducible nitric oxide synthase results in nitric oxide overproduction, which then rapidly reacts with hydrogen peroxide or superoxide generated by the mitochondria, to form highly reactive and harmful peroxynitrite, which ultimately induces nitrotyrosine formation. Herein, these interactions were confirmed and increased effects were observed in the presence of radicicol. On the whole, the data of the present study indicate that an interplay between oxidative and nitrosative stress is involved in Doxo-induced cardiotoxicity, and that both aspects are responsible for the induction of apoptosis. Furthermore, it is demonstrated that the mechanisms that further increase mitochondrial super-oxide generation (e.g., the inhibition of Cx43 translocation into the mitochondria) significantly accelerate the occurrence of cell death.

Chronic kidney disease (CKD) is characterized by an oxidative stress status, driving some CKD-associated complications, even at the gastrointestinal level. Indoxyl Sulfate (IS) is a protein-bound uremic toxin, poorly eliminated by dialysis. This toxin is able to affect the intestinal system, but its molecular mechanism/s in intestinal epithelial cells (IECs) remain poorly understood. This study&rsquo, s aim was to evaluate the effect of IS (31.2&ndash, 250 &micro, M) on oxidative stress in IEC-6 cells and on the intactness of IECs monolayers. Our results indicated that IS enhanced oxidative cell damage by inducing reactive oxygen species (ROS) release, reducing the antioxidant response and affecting Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) nuclear translocation as well its related antioxidant enzymes. In the wound healing assay model, IS reduced IEC-6 migration, slightly impaired actin cytoskeleton rearrangement, this effect was associated with connexin 43 alteration. Moreover, we reported the effect of CKD patients&rsquo, sera in IEC-6 cells. Our results indicated that patient sera induced ROS release in IEC-6 cells directly related to IS sera content and this effect was reduced by AST-120 serum treatment. Results highlighted the effect of IS in inducing oxidative stress in IECs and in impairing the intactness of the IECs cell monolayer, thus significantly contributing to CKD-associated intestinal alterations.

// Michela Terlizzi 1, 2 , Chiara Colarusso 1 , Ada Popolo 1 , Aldo Pinto 1 , Rosalinda Sorrentino 1 1 Department of Pharmacy (DIFARMA), University of Salerno, Salerno, Fisciano, 84084, Italy 2 PhD Program in Drug Discovery and Development, University of Salerno, Salerno, Fisciano, 84084, Italy Correspondence to: Rosalinda Sorrentino, email: rsorrentino@unisa.it Keywords: NLRP3 inflammasome, lung cancer, IL-1α, IL-1β, tumor-associated macrophages Abbreviations: Tumor-associated macrophages: TAMs; interleukin-1: IL-1; NLRP3: nod-like receptor 3 Received: March 03, 2016 Accepted: August 09, 2016 Published: August 12, 2016 ABSTRACT Macrophages highly populate tumour microenvironment and are referred to as tumor-associated macrophages (TAMs). The inflammasome is a multiprotein complex responsible of IL-1 like cytokines release, which biology has been widely studied by using bone-marrow-derived macrophages to mimic a physiological and/or host defense condition. To understand the role of this complex in lung tumor-associated macrophages (TAMs), we isolated and cultured broncho-alveolar lavage (BAL)-derived cells of lung tumor-bearing mice. The stimulation of lung TAMs with LPS+ATP increased the release of IL-1β. The inhibition of NLRP3 by means of glybenclamide significantly reduced IL-1β release. Similarly, C3H-derived, caspase-1 ko and caspase-11 ko TAMs released significantly reduced levels of IL-1β. Moreover, the stimulation of lung TAMs with the sole LPS induced a significant release of IL-1α, which was significantly reduced after caspase-1 pharmacological inhibition, and in TAMs genetically lacking caspase-1 and caspase-11. The inhibition of calpain I/II by means of MDL28170 did not alter IL-1α release after LPS treatment of lung TAMs. To note, the inoculation of LPS-treated bone marrow-derived macrophages into carcinogen-exposed mice increased lung tumor formation. In contrast, the depletion of TAMs by means of clodronate liposomes reduced lung tumorigenesis, associated to lower in vivo release of IL-1α and IL-1β. In conclusion, our data imply lung tumor lesions are populated by macrophages which pro-tumor activity is regulated by the activation of the NLRP3 inflammasome that leads to the release of IL-1α and IL-1β in a caspase-11/caspase-1-dependent manner.

Doxorubicin (DOXO) administration induces alterations in Connexin 43 (Cx43) expression and localization, thus, inducing alterations in chemical and electrical signal transmission between cardiomyocytes and in intracellular calcium homeostasis even evident after a single administration. This study was designed to evaluate if Diazoxide (DZX), a specific opener of mitochondrial KATP channels widely used for its cardioprotective effects, can fight DOXO-induced cardiotoxicity in a short-time mouse model. DZX (20 mg/kg i.p.) was administered 30 min before DOXO (10 mg/kg i.p.) in C57BL/6j female mice for 1–3 or seven days once every other day. A recovery of cardiac parameters, evaluated by Echocardiography, were observed in DZX+DOXO co-treated mice. Western blot analysis performed on heart lysates showed an increase in sarco/endoplasmic reticulum Ca2+-ATPase (SERCAII) and a reduction in phospholamban (PLB) amounts in DZX+DOXO co-treated mice. A contemporary recovery of intracellular Ca2+-signal, detected spectrofluorometrically by means of FURA-2AM, was observed in these mice. Cx43 expression and localization, analyzed by Western blot and confirmed by immunofluorescence analysis, showed that DZX co-treatement increases Cx43 amount both on sarcoplasmic membrane and on mitochondria. In conclusion, our data demonstrate that, in a short-time mouse model of DOXO-induced cardiotoxicity, DZX exerts its cardioprotective effects also by enhancing the amount Cx43.

N-Palmitoyl-ethanolamine (PEA) is an anti-inflammatory component of egg yolk that is usually employed for the prevention of respiratory apparatus virus infection and then frequently used for its efficient anti-inflammatory and analgesic effects in experimental models of visceral, neuropathic, and inflammatory diseases. Nevertheless, data of its use in animal or human therapy are still scarce and further studies are needed. Herein, we report the biological evaluation of a small library of N-palmitoyl-ethanolamine analogues or derivatives, characterized by a protected acid function (either as palmitoyl amides or hexadecyl esters), useful to decrease their hydrolysis rate in vitro and prolong their biological activity. Two of these compounds-namely phenyl-carbamic acid hexadecyl ester (4) and 2-methyl-pentadecanoic acid (4-nitro-phenyl)-amide (5)-have shown good anti-inflammatory and antioxidant properties, without affecting the viability of J774A.1 macrophages. Finally, crystals suitable for X-ray analysis of compound 4 have been obtained, and its solved crystal structure is here reported. Our outcomes may be helpful for a rational drug design based on new PEA analogues/derivatives with improved biological properties.

Among plants, the Lycopersicon esculentum (Solanaceae) is the most important for its beneficial effects on health. Several epidemiological studies have shown the benefits of tomato consumption in the cancer and cardiovascular disease prevention. Tomato products constitute the major source of lycopene, the most potent antioxidant among carotenoids in vitro. In tomatoes leaves are also present many secondary metabolites including phenolic compounds, phytoalexins, protease inhibitors and glycoalkaloids who protect against adverse effects of hosts including fungi, bacteria, viruses, and insects and are involved in host-plant resistance. In this work we evaluated the antiproliferative activity of tomato leaves extract (var. Paul Robenson) in vitro.

This study reports the synthesis and antioxidant activity of some new acetamide derivatives. The compounds’ structures were elucidated by NMR analysis and their melting points were measured. The in vitro antioxidant activity of these compounds was tested by evaluating the amount of scavenged ABTS radical and estimating ROS and NO production in tBOH- or LPS-stimulated J774.A1 macrophages. All compounds were tested for their effect on cell viability by an MTT assay and by a Brine Shrimp Test.

We report the synthesis, antioxidant and antiproliferative activity and a QSAR analysis of synthetic diphenylpropionamide derivatives. Synthesis of these compounds was achieved by direct condensation of 2,2- and 3,3-diphenylpropionic acid and appropriate amines using 1-propylphoshonic acid cyclic anhydride (PPAA) as catalyst. Compound structures were elucidated by NMR analysis and their melting points were measured. The in vitro antioxidant activity of these compounds was tested by evaluating the amount of scavenged ABTS radical and estimating ROS and NO production in LPS stimulated J774.A1 macrophages. All compounds were tested for their effect on viability of cells and results demonstrated that they are not toxic towards the cell lines used. The cytotoxic activity of all compounds was evaluated by a Brine Shrimp Test.

A3 adenosine receptor (A3AR) agonists have been reported to modulate cellular proliferation. This work was aimed to investigate the expression and the possible implication of A3AR in the human thyroid carcinomas. Normal thyroid tissue samples did not express A3 adenosine receptor, while primary thyroid cancer tissues expressed high level of A3AR, as determined by immunohistochemistry analysis. In human papillary thyroid carcinoma cell line, NPA, at concentrations > or =10 microM, the A3AR-selective agonist 2-chloro-N(6)-(3-iodobenzyl)adenosine-5'-N-methylcarboxamide (Cl-IB-MECA) produced inhibition of cell growth, by blocking the G(1) cell cycle phase in a concentration- and time-dependent manner. This effect was well correlated with a reduction of protein expression of cyclins D1 and E2 after 24 hours of Cl-IB-MECA treatment. Moreover Cl-IB-MECA induced dephosphorylation of ERK1/2 in a time- and concentration-dependent manner, which in turn inhibits cell proliferation. The effect of Cl-IB-MECA was not prevented by A3AR antagonists, MRS1191 or MRS1523 or FA385. Furthermore, neither nucleoside transporter inhibitors, Dypiridamole and NBTI, nor the A1, A2A and A2B receptors antagonists were able to block the response to Cl-IB-MECA. Although Cl-IB-MECA has been shown to influence cell death and survival in other systems through an A3AR-mediated mechanism, in NPA cells the growth inhibition induced by micromolar concentrations of Cl-IB-MECA is not related to A3AR activation and hence that its effects on human papillary carcinoma cell line seem to be independent of the presence of this receptor subtype.

// Valentina Bizzarro 1 , Raffaella Belvedere 1 , Maria Rita Milone 2 , Biagio Pucci 2 , Rita Lombardi 2 , Francesca Bruzzese 3 , Ada Popolo 1 , Luca Parente 1 , Alfredo Budillon 2, 3 , Antonello Petrella 1 1 Department of Pharmacy, University of Salerno, Fisciano (SA), Italy 2 Centro Ricerche Oncologiche Mercogliano, Istituto Nazionale Tumori Fondazione G. Pascale – IRCCS, Naples, Italy 3 Experimental Pharmacology Unit, Istituto Nazionale Tumori Fondazione G. Pascale – IRCCS, Naples, Italy Correspondence to: Antonello Petrella, e-mail: apetrella@unisa.it Alfredo Budillon, e-mail: a.budillon@istitutotumori.na.it Keywords: annexin A1, prostate cancer, cell invasion, EMT, cancer stem cells Received: May 18, 2015 Accepted: July 16, 2015 Published: July 28, 2015 ABSTRACT In this study, we have characterized the role of annexin A1 (ANXA1) in the acquisition and maintenance of stem-like/aggressive features in prostate cancer (PCa) cells comparing zoledronic acid (ZA)-resistant DU145R80 with their parental DU145 cells. ANXA1 is over-expressed in DU145R80 cells and its down-regulation abolishes their resistance to ZA. Moreover, ANXA1 induces DU145 and DU145R80 invasiveness acting through formyl peptide receptors (FPRs). Also, ANXA1 knockdown is able to inhibit epithelial to mesenchymal transition (EMT) and to reduce focal adhesion kinase (FAK) and metalloproteases (MMP)-2/9 expression in PCa cells. DU145R80 show a cancer stem cell (CSC)-like signature with a high expression of CSC markers including CD44, CD133, NANOG, Snail, Oct4 and ALDH7A1 and CSC-related genes as STAT3. Interestingly, ANXA1 knockdown induces these cells to revert from a putative prostate CSC to a more differentiated phenotype resembling DU145 PCa cell signature. Similar results are obtained concerning some drug resistance-related genes such as ATP Binding Cassette G2 (ABCG2) and Lung Resistant Protein (LRP). Our study provides new insights on the role of ANXA1 protein in PCa onset and progression.

Doxorubicin is the highly effective anthracycline, but its clinical use is limited by cardiotoxicity and consequent dysfunction. It has been proposed that the etiology of this is related to mitochondrial dysfunction. Connexin 43 (Cx43), the principal protein building block of cardiac gap junctions and hemichannels, plays an important role in cardioprotection. Recent reports confirmed the presence of Cx43 in the mitochondria as well. In this study, the role of mitochondrial Cx43 was evaluated 3 or 6 h after Doxorubicin administration to the rat heart cell line H9c2. Pharmacological inhibition of Hsp90 demonstrated that the mitochondrial Cx43 conferred cardioprotection by reducing cytosolic and mitochondrial reactive oxygen species production, mitochondrial calcium overload and mitochondrial membrane depolarization and cytochrome c release. In conclusion, our study demonstrates that Cx43 plays an important role in the protection of cardiac cells from Doxorubicin-induced toxicity.

Plasmacytoid dendritic cells (pDCs) highly populate lung tumor masses and are strictly correlated to bad prognosis, yet their role in lung cancer is controversial. To understand their role in lung cancer, we isolated pDCs from human samples of lung obtained from non-small cell lung cancer patients undergoing thoracic surgery. Tumor masses presented a higher percentage of pDCs than healthy tissues; pDCs were in the immunosuppressive phenotype, as determined by higher levels of CD33 and PD-L1. Despite higher HLA-A and HLA-D expression, cancerous pDCs did not exert cytotoxic activity against tumor cells but instead promoted their proliferation. In this scenario, cancerous pDCs were able to produce high levels of IL-1α. This effect was observed on the specific activation of the inflammasome absent in melanoma 2 (AIM2), which led to higher cytoplasmic calcium release responsible for calpain activation underlying IL-1α release. The blockade of type I interferon receptor and of AIM2 via the addition of LL-37 significantly reduced the release of IL-1α, which was still high after Nod-like receptor P3 inhibition via glibenclamide. More important, mitochondrial-derived reactive oxygen species sequester diminished AIM2-dependent IL-1α release. Our data demonstrate that lung tumor-associated pDCs are responsive to the activation of AIM2 that promotes calcium efflux and reactive oxygen species from mitochondria, leading to calpain activation and high levels of IL-1α, which facilitate tumor cell proliferation in the lung.

Background: Annexin A1 (ANXA1), a 37 kDa multifunctional protein, is over-expressed in tissues from patients of pancreatic carcinoma (PC) where the protein seems to be associated with malignant transformation and poor prognosis. Methods: The expression and localization of ANXA1 in MIA PaCa-2, PANC-1, BxPC-3 and CAPAN-2 cells were detected by Western Blotting and Immunofluorescence assay. Expression and activation of Formyl Peptide Receptors (FPRs) were shown through flow cytometry/PCR and FURA assay, respectively. To investigate the role of ANXA1 in PC cell migration and invasion, we performed in vitro wound-healing and matrigel invasion assays. Results: In all the analyzed PC cell lines, a huge expression and a variable localization of ANXA1 in sub-cellular compartments were observed. We confirmed the less a ggressive phenotype of BxPC-3 and CAPAN-2 compared with PANC-1 and MIA PaCa-2 cells, through the evaluation of Epithelial-Mesenchymal Transition (EMT) markers. Then, we tested MIA PaCa-2 and PANC-1 cell migration and invasiveness rate which was inhibited by specific ANXA1 siRNAs. Both the cell lines expressed FPR-1 and -2. Ac2-26, an ANXA1 mimetic peptide, induced intracellular calcium release, consistent with FPR activation, and significantly increased cell migration/invasion rate. Interestingly, in MIA PaCa-2 cells we found a cleaved form of ANXA1 (33 kDa) that localizes at cellular membranes and is secreted outside the cells, as confirmed by MS analysis. The importance of the secreted form of ANXA1 in cellular motility was confirmed by the administration of ANXA1 blocking antibody that inhibited migration and invasion rate in MIA PaCa-2 but not in PANC-1 cells that lack the 33 kDa ANXA1 form and show a lower degree of invasiveness. Finally, the treatment of PANC-1 cells with MIA PaCa-2 supernatants significantly increased the migration rate of these cells. Conclusion: This study provides new insights on the role of ANXA1 protein in PC progression. Our findings suggest that ANXA1 protein could regulate metastasis by favouring cell migration/invasion intracellularly, as cytoskeleton remodelling factor, and extracellularly like FPR ligand.

(1) Septic shock represents an important risk factor for patients critically ill. This pathology has been largely demonstrated to be a result of a myriad of events. Glucocorticoids represent the main pharmacological therapy used in this pathology. (2) Previously we showed that ATP-sensitive potassium (KATP) channels are involved in delayed vascular hyporeactivity in rats (24 h after Escherichia coli lipopolysaccharide (LPS) injection). In LPS-treated rats, we observed a significant hyporeactivity to phenylephrine (PE) that was reverted by glybenclamide (GLB), and a significant increase in cromakalim (CRK)-induced hypotension. (3) We evaluated the effect of dexamethasone (DEX 8 mg kg-1 i.p.) whether on hyporeactivity to PE or on hyperreactivity to CRK administration, in vivo, in a model of LPS (8 x 106 U kg-1 i.p.)-induced endotoxemia in urethane-anaesthetised rats. (4) DEX treatment significantly reduced, in a time-dependent manner, the increased hypotensive effect induced by CRK in LPS-treated rats. This effect was significantly (P

Biological activities of different varieties of tomato seed extracts were evaluated to verify the potential antioxidant and/or antiproliferative activity of the bioactive metabolites present in them. Findings demonstrated that among all the varieties investigated (San Marzano Rosso, San Marzano Giallo, Corbarino, Black Tomato and San Marzano/Black Tomato hybrid) San Marzano Rosso seed extract exhibited the highest free radical-scavenging activity with 68% of 2,2-Diphenyl-1-picrylhydrazyl radical inhibition, and the best cytotoxic activity evaluated by using the brine shrimp test (LD50: 23,198 ppm) and 3-(4,5-dimethylthiazol-2-yl)-2,5-phenyl-2H-tetrazolium bromide assay on A375 cell line (IC50: 137.7 μg/mL).

Hyaluronic acid (HA), a macropolysaccharidic component of the extracellular matrix, is common to most species and it is found in many sites of the human body, including skin and soft tissue. Not only does HA play a variety of roles in physiologic and in pathologic events, but it also has been extensively employed in cosmetic and skin-care products as drug delivery agent or for several biomedical applications. The most important limitations of HA are due to its short half-life and quick degradationin vivoand its consequently poor bioavailability. In the aim to overcome these difficulties, HA is generally subjected to several chemical changes. In this paper we obtained an acetylated form of HA with increased bioavailability with respect to the HA free form. Furthermore, an improved radical scavenging and anti-inflammatory activity has been evidenced, respectively, on ABTS radical cation and murine monocyte/macrophage cell lines (J774.A1).

Objectives In this work was to evaluate the cytotoxic activity of a series of monomeric group 3 and lanthanide (N,N,N)-heteroscorpionate-triflate complexes (M (OTf) 2 (cybpamd) (THF)) (Ln = Sc (2), Y (3), La (4), Nd (5), Sm (6), Dy (7), Yb (8); OTf = SO3CF3; cybpamd = N, N′-dicyclohexyl-2,2-bis-(3,5-dimethyl-pyrazol-1-yl)-acetamidinate) having octahedral geometry around the metal atoms on the human epithelial lung adenocarcinoma (A549), human melanoma (A375), human cervical epithelial adenocarcinoma, human embryonic kidney (HEK-293) and murine macrophages (J774.A1) cell lines. Methods All the tested compounds were incubated with cells for 72 h and their growth inhibition assessed by using MTT assay. Key findings On the cell line HEK-293 complexes 5 and 7 show a reasonable activities, while the murine macrophage cell line (J774.A1), only the scandium 2 complex is not very active. All complexes tested are poorly active on human health adenocarcinoma lung epithelial (A549) and human melanoma (A375). Conclusions The group 3 and lanthanide (N,N,N)-heteroscorpionate triflate-complexes (M(OTf)2(cybpamd)(THF)) on murine macrophage (J774.A1) cell line, except that of scandium, show a reasonable activity. On human epithelial cervix adenocarcinoma (HeLa) complexes 3, 5 and 6 are significantly more active than cis-platinum, as well as complex 5 is more active on human embryonic kidney (HEK-293) cell line. All the tested complexes are poorly active on human epithelial lung adenocarcinoma (A549) and human melanoma (A375). The different behaviour of the complexes examined (2–8) let us hypothesize that the cytotoxic activity is related to the molecule as a whole and not only to the ligand or the metal ion separately.

The β 3 -adrenoreceptor plays a major role in lipolysis but the role and distribution of β 3 -receptors in other specific sites have not been extensively studied. β 3 -adrenergic receptors are present not only in adipose tissue but also in human gall bladder, colon, prostate, and skeletal muscle. Recently, β 3 -adrenergic receptor stimulation was shown to elicit vasorelaxation of rat aorta through the NO–cGMP signal transduction pathway. Here we show that β 3 -receptors are present in human corpus cavernosum and are localized mainly in smooth muscle cells. After activation by a selective β 3 -adrenergic receptor agonist, BRL 37344, there was a cGMP-dependent but NO-independent vasorelaxation that was selectively blocked by a specific β 3 -receptor antagonist. In addition, we report that the human corpus cavernosum exhibits basal β 3 -receptor-mediated vasorelaxant tone and that β 3 -receptor activity is linked to inhibition of the RhoA/Rho-kinase pathway. These observations indicate that β 3 -receptors may play a physiological role in mediating penile erection and, therefore, could represent a therapeutic target for treatment of erectile dysfunction.

Brown Cuban propolis (BCP) is the major type of propolis in Cuba; its chemical composition is exclusive and the principal component is nemorosone. In this study we investigated the antiproliferative activity of the ethanol extract of BCP on human breast cancer cell lines. The MTT assay showed a significant antiproliferative activity (P0.01) inhibition of cell growth in the G1 phase of cell cycle, which was mechanism dose- and time-dependent. 17-β Estradiol (10 nM) administration to MCF-7 caused a significant (P Thus it was hypothesized that BCP possesses an estrogen-like activity. It is to be noted, however, that BCP application to MDA-MB 23 1 at 48 h also induced increased cell mortality. Thus, it cannot be ruled out that BCP could not only interact with the ER, but also have an ER-independent activity.