Incorporation of heparin-binding proteins into preformed dextran sulfate-chitosan nanoparticles.

Incorporation of proteins into dextran sulfate (DS)-chitosan (CS) nanoparticles (DSCS NPs) is commonly performed using entrapment procedures, in which protein molecules are mixed with DS and CS until particle formation occurs. As DS is an analog of heparin, the authors examined whether proteins could be directly incorporated into preformed DSCS NPs through a heparin binding domain-mediated interaction. The authors formulated negatively-charged DSCS NPs, and quantified the amount of charged DS in the outer shell of the particles. The authors then mixed the DSCS NPs with heparin-binding proteins (SDF-1α, VEGF, FGF-2, BMP-2, or lysozyme) to achieve incorporation. Data show that for DSCS NPs containing 100 nmol charged glucose sulfate units in DS, up to ~1.5 nmol of monomeric or ~0.75 nmol of dimeric heparin-binding proteins were incorporated without significantly altering the size or zeta potential of the particles. Incorporation efficiencies of these proteins were 95%-100%. In contrast, serum albumin or serum globulin showed minimal incorporation (8% and 4%, respectively) in 50% physiological saline, despite their large adsorption in water (80% and 92%, respectively). The NP-incorporated SDF-1α and VEGF exhibited full activity and sustained thermal stability. An in vivo aerosolization study showed that NP-incorporated SDF-1α persisted in rat lungs for 72 h (~34% remaining), while free SDF-1α was no longer detectable after 16 h. As many growth factors and cytokines contain heparin-binding sites/domains, incorporation into preformed DSCS NPs could facilitate in vivo applications of these proteins.

Up-regulation of the mammalian target of rapamycin complex 1 subunit Raptor by aldosterone induces abnormal pulmonary artery smooth muscle cell survival patterns to promote pulmonary arterial hypertension.

Activation of the mammalian target of rapamycin complex 1 (mTORC1) subunit Raptor induces cell growth and is a downstream target of Akt. Elevated levels of aldosterone activate Akt, and, in pulmonary arterial hypertension (PAH), correlate with pulmonary arteriole thickening, which suggests that mTORC1 regulation by aldosterone may mediate adverse pulmonary vascular remodeling. We hypothesized that aldosterone-Raptor signaling induces abnormal pulmonary artery smooth muscle cell (PASMC) survival patterns to promote PAH. Remodeled pulmonary arterioles from SU-5416/hypoxia-PAH rats and monocrotaline-PAH rats with hyperaldosteronism expressed increased levels of the Raptor target, p70S6K, which provided a basis for investigating aldosterone-Raptor signaling in human PASMCs. Aldosterone (10(-9) to 10(-7) M) increased Akt/mTOR/Raptor to activate p70S6K and increase proliferation, viability, and apoptosis resistance in PASMCs. In PASMCs transfected with Raptor-small interfering RNA or treated with spironolactone/eplerenone, aldosterone or pulmonary arterial plasma from patients with PAH failed to increase p70S6K activation or to induce cell survival in vitro Optimal inhibition of pulmonary arteriole Raptor was achieved by treatment with Staramine-monomethoxy polyethylene glycol that was formulated with Raptor-small interfering RNA plus spironolactone in vivo, which decreased arteriole muscularization and pulmonary hypertension in 2 experimental animal models of PAH in vivo Up-regulation of mTORC1 by aldosterone is a critical pathobiologic mechanism that controls PASMC survival to promote hypertrophic vascular remodeling and PAH.-Aghamohammadzadeh, R., Zhang, Y.-Y., Stephens, T. E., Arons, E., Zaman, P., Polach, K. J., Matar, M., Yung, L.-M., Yu, P. B., Bowman, F. P., Opotowsky, A. R., Waxman, A. B., Loscalzo, J., Leopold, J. A., Maron, B. A. Up-regulation of the mammalian target of rapamycin complex 1 subunit Raptor by aldosterone induces abnormal pulmonary artery smooth muscle cell survival patterns to promote pulmonary arterial hypertension.