Aldo Pinto

Lung cancer and Toll-like receptors

Lung carcinoma is one of the leading causes of death worldwide. It is a non-immunogenic cancer, resistant to immune surveillance. Toll-like receptors (TLRs) connect the innate to the adaptive immune system. Given that cancerous cells evade the immune system, the activation of TLRs could represent a potential target for cancer therapy. The induction of Th1-like and cytotoxic immunity by TLR signalling could lead to tumour cell death, resulting in tumour regression or arrest. However, basic research and clinical trials revealed that the activation of specific TLRs, such as TLR2, TLR4 and TLR9, do not have any anti-tumour activity in lung carcinoma. Increasing evidence suggests that TLRs are important regulators of tumour biology; however, little is known about their function in lung cancer. Thus, in order to develop new therapeutic approaches, further studies are needed to understand the connection between TLRs and lung cancer progression. This review focuses on the potential mechanisms by which TLR ligands can facilitate or not lung cancer and lung metastases establishment/progression.

Hydrogen Sulfide-Induced Dual Vascular Effect Involves Arachidonic Acid Cascade in Rat Mesenteric Arterial Bed

Hydrogen sulfide (H2S), a novel gaseous transmitter, is considered a physiological regulator of vascular homeostasis. Recent evidence suggests H2S as an endothelium-hyperpolarizing factor (EDHF) candidate. To address this issue, we evaluated the vascular effect of sodium hydrogen sulfide (NaHS), an H2S donor, on the rat mesenteric arterial bed. NaHS concentration-response curve was performed on preconstricted mesenteric arterial bed. To assess the contribution of EDHF, we performed a pharmacologic dissection using indomethacin, NG-nitro-l-arginine methyl ester (l-NAME), or apamin and charybdotoxin as cyclooxygenase, nitric-oxide synthase, and calcium-dependent potassium channel inhibitors, respectively. In another set of experiments, we used 4-(4-octadecylphenyl)-4-oxobutenoic acid, baicalein, or proadifen as phospholipase A2 (PLA2), lipoxygenase, and cytochrome P450 inhibitors, respectively. Finally, an immunofluorescence study was performed to support the involvement of PLA2 in mesenteric artery challenged by NaHS. NaHS promoted a dual vascular effect (i.e., vasoconstriction and vasodilation). l-NAME or baicalein administration affected neither NaHS-mediated vasodilation nor vasoconstriction, whereas apamin and charybdotoxin significantly inhibited NaHS-induced relaxation. Pretreatment with PLA2 inhibitor abolished both the contracting and the relaxant effect, whereas P450 cytochrome blocker significantly reduced NaHS-mediated relaxation. The immunofluorescence study showed that NaHS caused a migration of cytosolic PLA2 close to the nucleus, which implicates activation of this enzyme. Our data indicate that H2S could activate PLA2, which in turn releases arachidonic acid leading, initially, to vasoconstriction followed by vasodilation mediated by cytochrome P450-derived metabolites. Because EDHF has been presumed to be a cytochrome P450 derivative of the arachidonic acid, our results suggest that H2S acts through EHDF release.

Synthesis, characterization and cytotoxicity studies of methoxy alkyl substituted metallocenes

Five titanocene derivatives and one zirconium analogous, having cyclopentadienylethenylmethoxy ligand, were synthesized and fully characterized by NMR, FT-IR, and elemental analysis. Two of these complexes showed a good cytotoxic activity on human breast cancer (MCF-7) cell lines. Moreover, the half-titanocene disclosed also a good cytotoxic activity on human embryonic kidney (HEK-293). Additionally, a study on the rate of hydrolysis of these compounds showed that the leaving groups significantly affect the rate of hydrolysis of cyclopentadienyl groups too. The different activity of synthesized compounds was tentatively related to the rate of hydrolysis.

CpG-ODN increases the release of VEGF in a mouse model of lung carcinoma

Vascular endothelial‐derived growth factor (VEGF) plays a fundamental role in the formation of new vessels within the tumour mass. Increasing evidence has highlighted the involvement of Toll‐like receptors (TLRs) in cancer. Of interest, TLR9 is over‐expressed in human lung carcinoma tissues. The aim of our study was to determine whether TLR9 activation could alter VEGF release in a mouse model of lung carcinoma. Lewis lung carcinoma cells were intravenously (i.v.) inoculated and 10 days later, tumour‐bearing mice were treated with CpG‐ODN (CpG, a TLR9 ligand) or PBS. CpG administration enhanced VEGF release, which was associated with increased tumour lesions in the lung. CpG induced high levels of IL‐6 expression and activation of STAT3 in tumour‐bearing mice. Moreover, CpG induced VEGF release from primary fibroblasts and endothelial cells, which correlated with IL‐6 and TGFβ production. This may explain the large influx of fibroblasts and the production of basic fibroblast growth factor (bFGF) in the tumour mass. The administration of a monoclonal antibody against VEGF A arrested tumour progression and induced a Th1‐like response in CpG‐treated tumour‐bearing mice. In conclusion, our study demonstrates that the combination of CpG with anti‐VEGF monoclonal antibody could be of potential therapeutic in lung carcinoma.

Cl-IB-MECA enhances TNF-α release in peritoneal macrophages stimulated with LPS.

Adenosine receptor A3 (A3R) belongs to the Gi/Gq-coupled receptor family, that leads to the intracellular cAMP reduction and intracellular calcium increase, respectively. A3R is widely expressed and it can play a crucial role in many patho-physiological conditions, including inflammation. Here we investigate the effect of Cl-IB-MECA, A3R agonist, on the production of TNF-α. We found that Cl-IB-MECA enhances LPS-induced TNF-α release in peritoneal macrophages. This effect is reduced by MRS1191, A3R antagonist and by forskolin, activator of adenylyl cyclase. pIκBα increased in LPS+Cl-IB-MECA-treated macrophages, while total IκB kinase-β (IKKβ) reduced. Indeed, p65NF-κB nuclear translocation increased in cells treated with LPS+Cl-IB-MECA. Moreover, IMD 0354, IKKβ inhibitor, significantly abrogated the effect of Cl-IB-MECA on TNF-α release. Inhibition of protein kinase C (PKC) significantly reduced Cl-IB-MECA-induced TNF-α release in LPS-stimulated macrophages. Furthermore, LY-294002, PI3K inhibitor, reduced the TNF-α production enhanced by Cl-IB-MECA, although the phosphorylation status of Akt did not change in cells treated with LPS+Cl-IB-MECA than LPS alone. In summary, these data show that Cl-IB-MECA is able to enhance TNF-α production in LPS-treated macrophages in an NF-κB- dependent manner.

Inhibition of human platelet aggregation in vitro by standardized extract of Wendtia calycina

Wendtia calycina (Griseb.) Griseb., Vivianiaceae, is a Paraguayan herbaceous plant commonly known as burrito. Our previous study indicated that burrito leaves are a very good source of phenylpropanoid glycosides, principally verbascoside. From W. calycina leaves, a standardized, water-soluble extract rich in phenylpropanoid glycosides (WSE) has been developed on an industrial scale to be used as a food supplement, cosmetic, phytomedicine, and ingredient of different formulations. In this study, we investigated the effect of the WSE on human platelet aggregation in vitro induced by adenosine diphosphate (ADP), epinephrine (EPN), collagen (COL) or arachidonic acid (AA). WSE, concentration-dependently, inhibited ADP and EP-induced human platelet aggregation (IC50 were 0.82±0.15 mg/mL and 0.41±0.02 mg/mL, respectively). It did not inhibit collagen-induced platelet aggregation, thus suggesting a selectivity for the ADP-induced platelet activation pathways.