Synthesis of Quinoline and Dihydroquinoline Embelin Derivatives as Cardioprotective Agents.

A set of new dihydroquinoline embelin derivatives was obtained from the reaction of the natural benzoquinone embelin (1) with anilines and aromatic aldehydes in the presence of AgOTf. The synthesis of these compounds involves the formation of a Knoevenagel adduct, followed by nucleophilic addition of aniline and subsequent electrocyclic ring closure. The scope of the reaction regarding the aldehydes and anilines was determined. Quinoline derivatives were also obtained from the corresponding dihydroquinolines under oxidation with DDQ. The cardioprotective activity of the synthesized compounds was screened using a doxorubicin-induced cardiotoxicity model in H9c2 cardiomyocytes. Some structure-activity relationships were outlined, and the best activities were achieved with quinoline-embelin derivatives having a 4-nitrophenyl group attached at the pyridine ring. The obtained results indicated that embelin derivatives 4i, 6a, 6d, 6k, and 6m could have potential as cardioprotective agents, as they attenuated a DOX-induced cardiotoxicity effect acting on oxidative stress and apoptosis.

Phenolic and quinone methide nor-triterpenes as selective NLRP3 inflammasome inhibitors.

Dysregulated inflammasome activity, particularly of the NLRP3 inflammasome, is associated with the development of several inflammatory diseases. The study of molecules directly targeting NLRP3 is an emerging field in the discovery of new therapeutic compounds for the treatment of inflammatory disorders. Friedelane triterpenes are biologically active phytochemicals having a wide range of activities including anti-inflammatory effects. In this work, we evaluated the potential anti-inflammatory activity of phenolic and quinonemethide nor-triterpenes (1-11) isolated from Maytenus retusa and some semisynthetic derivatives (12-16) through inhibition of the NLRP3 inflammasome in macrophages. Among them, we found that triterpenes 6 and 14 were the most potent, showing markedly reduced caspase-1 activity, IL-1ß secretion (IC50 = 1.15 µM and 0.19 µM, respectively), and pyroptosis (IC50 = 2.21 µM and 0.13 µM, respectively). Further characterization confirmed their selective inhibition of NLRP3 inflammasome in both canonical and non-canonical activation pathways with no effects on AIM2 or NLRC4 inflammasome activation.

Labdane conjugates protect cardiomyocytes from doxorubicin-induced cardiotoxicity.

The cardiovascular side effects associated with doxorubicin (DOX), a wide spectrum anticancer drug, have limited its clinical application. Therefore, to explore novel strategies with cardioprotective effects, a series of new labdane conjugates were prepared (6a-6c and 8a-8d) from the natural diterpene labdanodiol (1). These hybrid compounds contain anti-inflammatory privileged structures such as naphthalimide, naphthoquinone, and furanonaphthoquinone. Biological activity of these conjugates against DOX-induced cardiotoxicity was tested in vitro and the potential molecular mechanisms of protective effects were explored in H9c2 cardiomyocytes. Three compounds 6c, 8a, and 8b significantly improved cardiomyocyte survival, via inhibition of reactive oxygen species-mediated mitogen-activated protein kinase signaling pathways (extracellular signal-regulated kinase and c-Jun N-terminal kinase) and autophagy mediated by Akt activation. Some structure-activity relationships were outlined, and the best activity was achieved with the labdane-furonaphthoquinone conjugate 8a having an N-cyclohexyl substituent. The findings of this study pave the way for further investigations to obtain more compounds with potential cardioprotective activity.

Dehydroisohispanolone as a Promising NLRP3 Inhibitor Agent: Bioevaluation and Molecular Docking.

Dehydroisohispanolone (DIH), is a labdane diterpene that has exhibited anti-inflammatory activity via inhibition of NF-κB activation, although its potential effects on inflammasome activation remain unexplored. This study aims to elucidate whether DIH modulates NLR family pyrin domain-containing protein 3 (NLRP3) inflammasome in macrophages. Our findings show that DIH inhibited NLRP3 activation triggered by Nigericin (Nig), adenosine triphosphate (ATP) and monosodium urate (MSU) crystals, indicating broad inhibitory effects. DIH significantly attenuated caspase-1 activation and secretion of the interleukin-1ß (IL-1ß) in J774A.1 cells. Interestingly, the protein expressions of NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), pro-caspase-1 and pro-IL-1ß were not affected by DIH treatment. Furthermore, we found that DIH pretreatment also inhibited the lipopolysaccharide (LPS)-induced NLRP3 inflammasome priming stage. In addition, DIH alleviated pyroptosis mediated by NLRP3 inflammasome activation. Similar results on IL-1ß release were observed in Nig-activated bone marrow-derived macrophages (BMDMs). Covalent molecular docking analysis revealed that DIH fits well into the ATP-binding site of NLRP3 protein, forming a covalent bond with Cys415. In conclusion, our experiments show that DIH is an effective NLRP3 inflammasome inhibitor and provide new evidence for its application in the therapy of inflammation-related diseases.

Dehydrohispanolone Derivatives Attenuate the Inflammatory Response through the Modulation of Inflammasome Activation.

The NLRP3 inflammasome plays a critical role in inflammation-mediated human diseases and represents a promising drug target for novel anti-inflammatory therapies. Hispanolone is a labdane diterpenoid isolated from the aerial parts of Ballota species. This diterpenoid and some derivatives have demonstrated anti-inflammatory effects in classical inflammatory pathways. In the present study, a series of dehydrohispanolone derivatives (1-19) was synthesized, and their anti-inflammatory activities toward NLRP3 inflammasome activation were evaluated. The structures of the dehydrohispanolone analogues produced were elucidated by NMR spectroscopy and mass spectrometry. Four derivatives significantly inhibited IL-1ß secretion, with 15 and 18 being the most active (IC50 = 18.7 and 13.8 µM, respectively). Analysis of IL-1ß and caspase-1 expression revealed that the new diterpenoids 15 and 18 are selective inhibitors of the NLRP3 inflammasome, reinforcing the previously demonstrated anti-inflammatory properties of hispanolone derivatives.

Current status of terpenoids as inflammasome inhibitors.

Increasing evidence supports NLRP3 inflammasome as a new target to control inflammation. Dysregulation of NLRP3 inflammasome has been reported to be involved in the pathogenesis of several human inflammatory diseases. However, no NLRP3 inflammasome inhibitors are available in clinic. Terpenoids are natural products with multi-target activities against inflammation. Recent studies have revealed that these compounds are capable of inhibiting the activation of NLRP3 inflammasome in several mouse models of NLRP3 inflammasome-related pathogenesis. Thus, terpenoids represent an interesting pharmacological approach for the treatment of inflammatory diseases as they are endowed with a dual mechanism of inhibition of NF-KB transcription factor and inflammasome activation, both critically involved in their anti-inflammatory effects. This work provides an overview of the current knowledge on the therapeutic potential of terpenoids as NLRP3 inflammasome inhibitors.

α-Hispanolol Induces Apoptosis and Suppresses Migration and Invasion of Glioblastoma Cells Likely via Downregulation of MMP-2/9 Expression and p38MAPK Attenuation.

α-Hispanolol (α-H) is a labdane diterpenoid that has been shown to induce apoptosis in several human cancer cells. However, the effect of α-H in human glioblastoma cells has not been described. In the present work, we have investigated the effects of α-H on apoptosis, migration, and invasion of human glioblastoma cells with the aim of identifying the molecular targets underlying its mechanism of action. The results revealed that α-H showed significant cytotoxicity against human glioma cancer cell lines U87 and U373 in a concentration- and time-dependent manner. This effect was higher in U87 cells and linked to apoptosis, as revealed the increased percentage of sub-G1 population by cell cycle analysis and acquisition of typical features of apoptotic cell morphology. Apoptosis was also confirmed by significant presence of annexin V-positive cells and caspase activation. Pretreatment with caspase inhibitors diminishes the activities of caspase 8, 9, and 3 and maintains the percentage of viable glioblastoma cells, indicating that α-H induced cell apoptosis through both the extrinsic and the intrinsic pathways. Moreover, we also found that α-H downregulated the anti-apoptotic Bcl-2 and Bcl-xL proteins and activated the pro-apoptotic Bid and Bax proteins. On the other hand, α-H exhibited inhibitory effects on the migration and invasion of U87 cells in a concentration-dependent manner. Furthermore, additional experiments showed that α-H treatment reduced the enzymatic activities and protein levels of matrix metalloproteinase MMP-2 and MMP-9 and increased the expression of TIMP-1 inhibitor, probably via p38MAPK regulation. Finally, xenograft assays confirmed the anti-glioma efficacy of α-H. Taken together, these findings suggest that α-H may exert anti-tumoral effects in vitro and in vivo through the inhibition of cell proliferation and invasion as well as by the induction of apoptosis in human glioblastoma cells. This research describes α-H as a new drug that may improve the therapeutic efficacy against glioblastoma tumors.

Amphiphilic Acrylic Nanoparticles Containing the Poloxamer Star Bayfit® 10WF15 as Ophthalmic Drug Carriers.

Topical application of drops containing ocular drugs is the preferred non-invasive route to treat diseases that affect the anterior segment of the eye. However, the formulation of eye drops is a major challenge for pharmacists since the access of drugs to ocular tissues is restricted by several barriers. Acetazolamide (ACZ) is a carbonic anhydrase inhibitor used orally for the treatment of ocular hypertension in glaucoma. However, large ACZ doses are needed which results in systemic side effects. Recently, we synthesized copolymers based on 2-hydroxyethyl methacrylate (HEMA) and a functionalized three-arm poloxamer star (Bayfit-MA). The new material (HEMA/Bayfit-MA) was engineered to be transformed into nanoparticles without the use of surfactants, which represents a significant step forward in developing new ophthalmic drug delivery platforms. Acetazolamide-loaded nanocarriers (ACZ-NPs) were prepared via dialysis (224 ± 19 nm, -17.2 ± 0.4 mV). The in vitro release rate of ACZ was constant over 24 h (cumulative delivery of ACZ: 83.3 ± 8.4%). Following standard specifications, ACZ-NPs were not cytotoxic in vitro in cornea, conjunctiva, and macrophages. In normotensive rabbits, ACZ-NPs generated a significant intraocular pressure reduction compared to a conventional solution of ACZ (16.4% versus 9.6%) with the same dose of the hypotensive drug (20 µg). In comparison to previously reported studies, this formulation reduced intraocular pressure with a lower dose of ACZ. In summary, HEMA:Bayfit-MA nanoparticles may be a promising system for ocular topical treatments, showing an enhanced ocular bioavailability of ACZ after a single instillation on the ocular surface.

Molecular Targets Involved in the Neuroprotection Mediated by Terpenoids.

Natural products and their derivatives represent the most consistently successful source of drug leads. Terpenoids, a structurally diverse group, are secondary metabolites widely distributed in nature, endowed with a wide range of biological activities such as antibacterial, anti-inflammatory, antitumoral, or neuroprotective effects, which consolidate their therapeutic value. During the last decades, and taking into consideration the prevalence of aging-related diseases, research activity into the neuroprotective effects of these types of compounds has increased enormously. Several signaling pathways involved in neuroprotection are targets of their mechanism of action and mediate their pleiotropic protective activity in neuronal cell damage. In the present review, molecular basis of the neuroprotection exerted by terpenoids is presented, focusing on preclinical evidence of the therapeutic potential of diterpenoids and triterpenoids on neurodegenerative disorders. By acting on diverse mechanisms simultaneously, terpenoids have been emphasized as promising multitarget agents.

Semisynthesis and Inhibitory Effects of Solidagenone Derivatives on TLR-Mediated Inflammatory Responses.

A series of nine derivatives (2⁻10) were prepared from the diterpene solidagenone (1) and their structures were elucidated by means of spectroscopic studies. Their ability to inhibit inflammatory responses elicited in peritoneal macrophages by TLR ligands was investigated. Compounds 5 and 6 showed significant anti-inflammatory effects, as they inhibited the protein expression of nitric oxide synthase (NOS-2), cyclooxygenase-2 (COX-2), and cytokine production (TNF-α, IL-6, and IL-12) induced by the ligand of TLR4, lipopolysaccharide (LPS), acting at the transcriptional level. Some structure⁻activity relationships were outlined. Compound 5 was selected as a representative compound and molecular mechanisms involved in its biological activity were investigated. Inhibition of NF-κB and p38 signaling seems to be involved in the mechanism of action of compound 5. In addition, this compound also inhibited inflammatory responses mediated by ligands of TLR2 and TLR3 receptors. To rationalize the obtained results, molecular docking and molecular dynamic studies were carried out on TLR4. All these data indicate that solidagenone derivative 5 might be used for the design of new anti-inflammatory agents.

A hispanolone-derived diterpenoid inhibits M2-Macrophage polarization in vitro via JAK/STAT and attenuates chitin induced inflammation in vivo.

Macrophages are highly plastic cells that adopt different functional phenotypes in response to environmental signals. Classically activated macrophages (M1) exhibit a pro-inflammatory role, mediating host defense against microorganisms or tumor cells; whereas alternatively activated macrophages (M2) perform a range of physiological processes, including inflammation, wound repair and tissue remodeling. Interestingly, M2 macrophages have been involved in pathological settings such as tumor progression, parasitic infection and respiratory disorders. Consequently, the search of new agents able to control macrophage polarization is on the basis of new therapeutic strategies. In the present study, we have evaluated the effect of the hispanolone derivative 8,9-dehydrohispanolone-15,16-lactol (DHHL) on M2 macrophage polarization. Our results reveal that DHHL significantly inhibited IL-4- or IL-13-stimulated M2 macrophage activation, as showed by reduced expression of M2 markers. In addition, DHHL suppressed IL-4-induced STAT-6 and JAK-1 tyrosine phosphorylation, suggesting that this compound inhibited M2 polarization by suppressing the JAK-STAT signaling pathway. Finally, DHHL prevented eosinophil recruitment and the presence of F4/80+-CD206+ M2-like macrophages in an in vivo model of M2 polarization via administration of chitin. Collectively, these results confirm DHHL as a novel regulator of macrophage polarization suitable to design future therapies towards M2-macrophages mediated pathologies.

GQ-11: A new PPAR agonist improves obesity-induced metabolic alterations in LDLr-/- mice.

BACKGROUND: Obesity and insulin resistance/diabetes are important risk factors for cardiovascular diseases and demand safe and efficacious therapeutics. OBJECTIVE: To assess the effects of a new thiazolidine compound-GQ-11-on obesity and insulin resistance induced by a diabetogenic diet in LDL receptor-deficient (LDLr-/-) mice. METHODS: Molecular docking simulations of GQ-11, PPARα and PPARγ structures were performed. Male C57BL/6J LDLr-/- mice fed a diabetogenic diet for 24 weeks were treated with vehicle, GQ-11 or pioglitazone or (20 mg/kg/day) for 28 days by oral gavage. Glucose tolerance test, insulin, HOMA-IR, adipokines (leptin, adiponectin) and the lipid profile were assessed after treatment. Adipose tissue was analysed by X-ray analysis and morphometry; gene and protein expression were evaluated by real-time PCR and western blot, respectively. RESULTS: GQ-11 showed partial agonism to PPARγ and PPARα. In vivo, treatment with GQ-11 ameliorated insulin sensitivity and did not modify subcutaneous adipose tissue and body weight gain. In addition, GQ-11 restored adipokine imbalance induced by a diabetogenic diet and enhanced Glut-4 expression in the adipose tissue. Improved insulin sensitivity was also associated with lower levels of MCP-1 and higher levels of IL-10. Furthermore, GQ-11 reduced triglycerides and VLDL cholesterol and increased HDL-cholesterol by upregulation of Apoa1 and Abca1 gene expression in the liver. CONCLUSION: GQ-11 is a partial/dual PPARα/γ agonist that demonstrates anti-diabetic effects. Additionally, it improves the lipid profile and ameliorates chronic inflammation associated with obesity in atherosclerosis-prone mice.

Novel Nano-Liposome Formulation for Dry Eyes with Components Similar to the Preocular Tear Film.

Dry eye is commonly treated with artificial tears; however, developing artificial tears similar to natural tears is difficult due to the complex nature of tears. We characterized and evaluated a novel artificial tear formulation with components similar to the lipid and aqueous constituents of natural tears. Nano-liposomes, composed in part of phosphatidylcholine, were dispersed in an aqueous solution of bioadhesive sodium hyaluronate. Liposome size, zeta potential, and physicochemical properties of the fresh and stored (4 °C) liposomal formulation were analyzed. In vitro tolerance was tested using human corneal and conjunctival cell lines by exposures of 15 min to 4 h. The tolerance of the liposomal formulation was evaluated in animals (rabbits). The average liposome size was 186.3 ± 7.0 nm, and the zeta potential was negative. The osmolarity of the formulation was 198.6 ± 1.7 mOsm, with a surface tension of 36.5 ± 0.4 mN/m and viscosity of 3.05 ± 0.02 mPa·s. Viability values in the human corneal and conjunctival cell lines were always >80%, even after liposomal formulation storage for 8 weeks. Discomfort and clinical signs after instillation in rabbit eyes were absent. The new formulation, based on phosphatidylcholine-liposomes dispersed in sodium hyaluronate has suitable components and characteristics, including high in vitro cell viability and good in vivo tolerance, to serve as a tear substitute.

8,9-Dehydrohispanolone-15,16-lactol diterpene prevents LPS-triggered inflammatory responses by inhibiting endothelial activation.

Endothelial activation contributes to lung inflammatory disorders by inducing leucocyte recruitment to pulmonary parenchyma. Consequently, vascular-targeted therapies constitute promising strategies for the treatment of inflammatory pathologies. In the present study, we evaluated the effect of 8,9-dehydrohispanolone-15,16-lactol diterpene (DT) on lung endothelium during inflammation. Lung endothelial cells pre-treated with DT and activated with lipopolysaccharide (LPS) or tumour necrosis factor-α (TNF-α) exhibited reduced expression of the pro-inflammatory cytokines Cxcl10, Ccl5 and Cxcl1, whereas the anti-inflammatory molecules IL1r2 and IL-10 were induced. Consistent with this result, DT pre-treatment inhibited nuclear factor κB (NF-κB) nuclear translocation, by interfering with IκBα phosphorylation, and consequently NF-κB transcriptional activity in endothelium activated by LPS or TNF-α. Furthermore, DT, probably through p38 signalling, induced transcriptional activation of genes containing activator protein 1 (AP-1)-binding elements. Inhibition of p38 prevented IL1r2 mRNA expression in endothelium incubated with DT alone or in combination with LPS or TNF-α. Accordingly, conditioned medium (CM) from these cells failed to stimulate leucocytes as measured by a reduction in adhesive ability of the leucocyte cell line J774 to fibronectin (FN). Additionally, DT reduced the expression of the endothelial adhesion molecules E-selectin, vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1) after activation. Similarly, expression of VCAM-1 and ICAM-1 molecules on the lung endothelial layer of C57/BL6 mice pre-treated with DT and challenged with LPS were unchanged. Finally, inhibition of vascular adhesion molecule expression by DT decreased the interaction of J774 cells with lung endothelial cells in an inflammatory environment. Our findings establish DT as a novel endothelial inhibitor for the treatment of inflammatory-related diseases triggered by Gram-negative bacteria or by the associated cytokine TNF-α.

New PPARγ partial agonist improves obesity-induced metabolic alterations and atherosclerosis in LDLr(-/-) mice.

Peroxisome proliferator-activated receptor gamma (PPARγ) regulates multiple pathways involved in the pathogenesis of obesity and atherosclerosis. Here, we evaluated the therapeutic potential of GQ-177, a new thiazolidinedione, on diet-induced obesity and atherosclerosis. The intermolecular interaction between PPARγ and GQ-177 was examined by virtual docking and PPAR activation was determined by reporter gene assay identifying GQ-177 as a partial and selective PPARγ agonist. For the evaluation of biological activity of GQ-177, low-density lipoprotein receptor-deficient (LDLr(-/-)) C57/BL6 mice were fed either a high fat diabetogenic diet (diet-induced obesity), or a high fat atherogenic diet, and treated with vehicle, GQ-177 (20mg/kg/day), pioglitazone (20mg/kg/day, diet-induced obesity model) or rosiglitazone (15mg/kg/day, atherosclerosis model) for 28 days. In diet-induced obesity mice, GQ-177 improved insulin sensitivity and lipid profile, increased plasma adiponectin and GLUT4 mRNA in adipose tissue, without affecting body weight, food consumption, fat accumulation and bone density. Moreover, GQ-177 enhanced hepatic mRNA levels of proteins involved in lipid metabolism. In the atherosclerosis mice, GQ-177 inhibited atherosclerotic lesion progression, increased plasma HDL and mRNA levels of PPARγ and ATP-binding cassette A1 in atherosclerotic lesions. GQ-177 acts as a partial PPARγ agonist that improves obesity-associated insulin resistance and dyslipidemia with atheroprotective effects in LDLr(-/-) mice.

Biological evaluation of angular disubstituted naphthoimidazoles as anti-inflammatory agents.

A series of naphthoimidazoles derivatives (3a-3f) were tested for potential anti-inflammatory activity on lipopolysaccharide (LPS)-treated macrophages. Naphthoimidazole 3e exhibited significant inhibitory effects on nitric oxide (NO) production (IC50 <10µM) and decreased the expression of nitric oxide synthase-2 (NOS-2) and cycloxygenase-2 (COX-2) enzymes. It also inhibited the activation of transcription factor NF-κB. Naphthoimidazole 3e might represent a starting point for the synthesis of new anti-inflammatory naphthoimidazoles derivatives.

Novel biodegradable polyesteramide microspheres for controlled drug delivery in Ophthalmology.

Most of the posterior segment diseases are chronic and multifactorial and require long-term intraocular medication. Conventional treatments of these pathologies consist of successive intraocular injections, which are associated with adverse effects. Successful therapy requires the development of new drug delivery systems able to release the active substance for a long term with a single administration. The present work involves the description of a new generation of microspheres based on poly(ester amide)s (PEA), which are novel polymers with improved biodegradability, processability and good thermal and mechanical properties. We report on the preparation of the PEA polymer, PEA microspheres (PEA Ms) and their characterization. PEA Ms (~15µm) were loaded with a lipophilic drug (dexamethasone) (181.0±2.4µg DX/mg Ms). The in vitro release profile of the drug showed a constant delivery for at least 90days. Based on the data from a performed in vitro release study, a kinetic ocular model to predict in vivo drug concentrations in a rabbit vitreous was built. According to the pharmacokinetic simulations, intravitreal injection of dexamethasone loaded PEA microspheres would provide release of the drug in rabbit eyes up to 3months. Cytotoxicity studies in macrophages and retinal pigment epithelial cells revealed a good in vitro tolerance of the microsystems. After sterilization, PEA Ms were administered in vivo by subtenon and intravitreal injections in male Sprague-Dawley rats and the location of the microspheres in rat eyes was monitored. We conclude that PEA Ms provide an alternative delivery system for controlling the delivery of drugs to the eye, allowing a novel generation of microsphere design.

α-Hispanolol sensitizes hepatocellular carcinoma cells to TRAIL-induced apoptosis via death receptor up-regulation.

Hispanolone derivatives have been previously described as anti-inflammatory and antitumoral agents. However, their effects on overcoming Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) resistance remain to be elucidated. In this study, we analyzed the cytotoxic effects of the synthetic hispanolone derivative α-hispanolol (α-H) in several tumor cell lines, and we evaluated the induction of apoptosis, as well as the TRAIL-sensitizing potential of α-H in the hepatocellular carcinoma cell line HepG2. Our data show that α-H decreased cell viability in a dose-dependent manner in HeLa, MDA-MB231, U87 and HepG2 cell lines, with a more prominent effect in HepG2 cells. Interestingly, α-H had no effect on non-tumoral cells. α-H induced activation of caspase-8 and caspase-9 and also increased levels of the proapoptotic protein Bax, decreasing antiapoptotic proteins (Bcl-2, X-IAP and IAP-1) in HepG2 cells. Specific inhibition of caspase-8 abrogated the cascade of caspase activation, suggesting that the extrinsic pathway has a critical role in the apoptotic events induced by α-H. Furthermore, combined treatment of α-H with TRAIL enhanced apoptosis in HepG2 cells, activating caspase-8 and caspase-9. This correlated with up-regulation of both the TRAIL death receptor DR4 and DR5. DR4 or DR5 neutralizing antibodies abolished the effect of α-H on TRAIL-induced apoptosis, suggesting that sensitization was mediated through the death receptor pathway. Our results demonstrate that α-H induced apoptosis in the human hepatocellular carcinoma cell line HepG2 through activation of caspases and induction of the death receptor pathway. In addition, we describe a novel function of α-H as a sensitizer on TRAIL-induced apoptotic cell death in HepG2 cells.

New indole-thiazolidine attenuates atherosclerosis in LDLr(-/-) mice.

Thiazolidinediones (TZDs) are peroxisome proliferator-activated receptor γ (PPARγ) agonists that improve insulin-mediated glucose uptake and possess beneficial vasculoprotective actions. However, because undesirable side effects are associated with these drugs, novel TZDs are under development. In this study, we evaluated the biological activity of LYSO-7, a new indole-thiazolidine, on PPAR activation, inflammation and atherogenesis using a gene reporter assay, lipopolysaccharide (LPS)-activated RAW 264.7 cell culture, and a low-density lipoprotein receptor knockout (LDLr(-/-)) mouse model of atherosclerosis. LYSO-7 shows low cytotoxicity in RAW 264.7 cells and at 2.5µmol/L induces PPARα and PPARγ transactivation as well as inhibits LPS-induced nitrite production and the mRNA gene expression levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), vascular cell adhesion molecule-1 (VCAM-1) and monocyte chemoattractant protein-1 (MCP-1). In addition, treatment with LYSO-7 reduces the development of atherosclerosis in LDLr(-/-) mice, improves the lipid profile, blood glucose levels, and downregulates CD40 and CD40L expression without affecting the body weight of the animals. Altogether, our data show that LYSO-7 possesses anti-inflammatory properties and that treatment with this TZD attenuates atherosclerosis progression in LDLr(-/-) mice by modulating lipid metabolism and inflammation. Thus, LYSO-7 shows potential as a new drug candidate for the treatment of atherosclerosis.

Anti-inflammatory activity and phenolic profile of propolis from two locations in Región Metropolitana de Santiago, Chile.

ETHNOPHARMACOLOGICAL RELEVANCE: Propolis has long been used as a popular folk medicine due to its wide spectrum of alleged biological and pharmaceutical properties. In Chile, propolis is widely used by folklore medicine as an anti-inflammatory agent; however, this property has not been demonstrated by scientific methods. AIM OF THE STUDY: The objective of this study was to determine the anti-inflammatory activity in vivo and in vitro and to establish the phenolic profile of propolis collected in two localities in Región Metropolitana de Santiago (RM), Chile. MATERIALS AND METHODS: Propolis was collected in the areas of Caleu and Buin, RM Chile. Following that, the samples were unwaxed to obtain the global ethanolic extracts of propolis (EEPs) and, from these, the serial extracts of dichloromethane (EEP-DCMs) and ethanol (EEP-EtOHs). The topic anti-inflammatory effect was evaluated through mice ear edema induced by arachidonic acid (AA) and 12-O-tetradecanoylphorbol-13-acetate (TPA) at a dose of 3 mg/ear. Nitric oxide (NO) measurements were determined spectrophotometrically (Greiss reagent) by the accumulation of nitrite in the medium of macrophages RAW 264.7 stimulated with the lipopolysaccharide (LPS, 1 µg/mL) for 20 h at different concentrations of the EEPs, EEP-DCMs and EEP-EtOHs (6.25-50.00 µg/mL). The content of total phenols and flavonoids were determined through the methods of Folin-Ciocalteau and AlCl3, respectively. The profile of phenolic compounds was determined by HPLC-UV-ESI-MS/MS. RESULTS: The EEP-EtOH (64%) and EEP (59%) of Buin were the most active in the inflammation induced by TPA and AA respectively, being the anti-inflammatory effect stronger than the same Caleu extracts. Regarding the release of NO, all the extracts from the Buin propolis inhibited significantly its release in a concentration-dependent manner, this inhibition was stronger than the extracts from Caleu propolis. CONCLUSIONS: Our research shows for the first time a comparative study of the topical in vivo activity of two Chilean propolis. Both propolis showed in vivo topical anti-inflammatory activity against AA and TPA, the most active was Buin propolis and this difference is due in part to the variations in total phenols and flavonoids content and the phenolic profile. The phenols and flavonoids content of Buin propolis was higher than Caleu propolis. The extracts from Buin propolis result in a lower release of NO.