Search

Membrane proteins are present in both cardiac T-tubule (TTM) and outer surface membrane (OSM), although at a different density. Classical pharmacology does not allow to explore the function of a membrane protein separately in OSM vs. TTM. Here, we developed a technology based on size exclusion to explore the function of β-adrenergic receptors (β-ARs) located in the OSM. We synthetized a PEG-Iso molecule by covalent linking between isoprenaline (Iso) and a 5000 Da PolyEthylene-Glycol (PEG). Using confocal microscopy, we show that PEGylation constrains molecules outside the T-tubule network. PEG-Iso produced similar effects as Iso on I ca,L , sarcomere shortening and Ca 2+ transients. However, PEG-Iso increased [cAMP] i with a lower efficacy than Iso, produced a much lower stimulation of nuclear PKA activity than Iso but a larger stimulation of cytosolic PKA at equivalent levels of [cAMP] i . Our results show that activation of OSM β-ARs is sufficient to activate cytosolic PKA and excitation-contraction coupling, but insufficient to activate nuclear PKA or nuclear protein phosphorylation for which additional activation of TTM β-ARs is needed.

Tuberculosis is an infectious disease, affecting mostly lungs, that is still considered a health global problem as it causes millions of deaths worldwide. Current treatment is effective, however, associated with severe adverse effects due to the high doses of each anti-tuberculosis drug daily administrated by oral therapy. For the first time, a pyrazinoic acid biodegradable nanoconjugate was synthesized and developed for pulmonary administration in an attempt to reduce the administered doses by achieving a high drug payload and controlled release at the target site. The conjugate was synthesized by coupling pyrazinoic acid on carboxylic groups of poly(malic acid), which is a biodegradable and biocompatible polymer, and posteriorly self-assembled into nanoconjugates. Characterization confirmed the formation of nanometric, spherical and negatively charged pyrazinoic acid nanoconjugate (NC-PA). NC-PA was stable for 60 days at 4 and 37 ºC and able to deliver PA in a sustained release manner over time. On macrophages, they exhibited no cell toxicity for a wide range of concentrations (from 1 to 100 µg/mL), demonstrating the safety of NC-PA. In addition, the nanoconjugate was efficiently taken up by RAW 264.7 cells over 6 hours reaching a maximum value after 3h of incubation. In conclusion, innovative nanoconjugates are a promising alternative to deliver drugs directly to the lungs and contributing to improving tuberculosis therapy.

Prostate cancer is the most important cancer in terms of incidence and the sixth in terms of mortality for men throughout the world according to the International Agency for Research on Cancer [1]. At the moment, this cancer is treated with hormone therapy, but is this only effective for 24-36 months before the patient develops resistance. Two of the few known cytotoxic molecules active against metastatic hormone-refractory prostate cancer are docetaxel (DOC) [2] and mitoxantrone (MIT) [3]. The encapsulation of these two molecules with different mechanisms of action against tumor cells into biocompatible and biodegradable polymeric nanoparticles (NP) is an original therapeutic approach to avoid the side effects of chemotherapy. The aim of this work was to develop a multifunctional drug delivery system based on polylactic acid-polyethylene glycol (PLA- PEG) block copolymer loaded with two different anti-cancer agents. The presence of PEG delays their capture by the macrophages and decreases the clearance by the reticuloendothelial system. These properties allow them to circulate longer in the body and reach the tumor by the EPR (Enhanced Permeation and Retention) effect [4]. The PLA-PEG copolymer was synthesized by ring-opening polymerization [5]. Finally, the anti-cancer drugs docetaxel and mitoxantrone were encapsulated by a nanoprecipitation technique [6]. Under these conditions, small and monodispersed NPs of around 70 nm were obtained. The colloidal stability of the NPs was verified by dynamic light scattering measurements. The antitumor efficacy of the NPs against two different human prostate cancer cells PC-3 and LnCaP was evaluated by MTS method. The effectiveness of each type of nanoparticle was evaluated separately as well as mixtures of the two types of nanoparticle in different ratios in order to observe any synergy, additivity or antagonism. The results showed the efficiency of all types of nanoparticle with at least 50 % of cell activity reduction in the first 48 h at 125 µg/mL of NPs. Moreover, a synergistic effect was observed in two different combinations of MIT and DOC leading to less than 10 % of cell activity against PC-3 prostate cells in the first 96 h of treatment. Thus, the proposed combination of chemotherapic agents was shown to improve the effectiveness of this kind of nanosized delivery system.