Effects on mitochondria of mitochondria-induced nitric oxide release from a ruthenium nitrosyl complex.

The ruthenium nitrosyl complex trans-[Ru(NO)(NH(3))(4)(py)](PF(6))(3) (pyNO), a nitric oxide (NO) donor, was studied in regard to the release of NO and its impact both on isolated mitochondria and HepG2 cells. In isolated mitochondria, NO release from pyNO was concomitant with NAD(P)H oxidation and, in the 25-100 microM range, it resulted in dissipation of mitochondrial membrane potential, inhibition of state 3 respiration, ATP depletion and reactive oxygen species (ROS) generation. In the presence of Ca(2+), mitochondrial permeability transition (MPT), an unspecific membrane permeabilization involved in cell necrosis and some types of apoptosis, was elicited. As demonstrated by externalization of phosphatidylserine and activation of caspase-9 and caspase-3, pyNO (50-100 microM) induced HepG2 cell death, mainly by apoptosis. The combined action of the NO itself, the peroxynitrite yielded by NO in the presence of reactive oxygen species (ROS) and the oxidative stress generated by the NAD(P)H oxidation is proposed to be involved in cell death by pyNO, both via respiratory chain inhibition and ROS levels increase, or even via MPT, if Ca(2+) is present.

Phenotypic switching prevention and proliferation/migration inhibition of vascular smooth muscle cells by the ruthenium nitrosyl complex trans-[Ru(NO)Cl(cyclam](PF6 )2.

OBJECTIVES: Vascular smooth muscle cell (VSMC) migration and proliferation at sites of vascular injury are both critical steps in the development of intimal hyperplasia (IH). Local delivery of nitric oxide (NO) largely prevents these events. Among the NO donors, tetraazamacrocyclic nitrosyl complexes, such as trans-[Ru(NO)Cl(cyclam)](PF6 )2 (cyclamNO), gained attention for their features, which include the possibility of being embedded in solid matrices, and ability to participate in a nitrite/NO catalytic conversion cycle. METHODS: Methods used to evaluate cyclamNO activity: safety margin by NR and MTT; cell proliferation by 3H-thymidine incorporation and proliferating cell nuclear antigen (PCNA) expression; antimigratory properties by transwell and wound healing; prevention of cell phenotypic switching under platelet-derived growth factor type BB (PDGF-BB) stimuli by analysis of alpha smooth muscle actin (α-SMA) expression. KEY FINDINGS: Cell proliferation and migration induced by PDGF-BB were significantly inhibited by cyclamNO. The ~60% reduction on expression of contractile protein α-SMA induced by PDGF-BB revealed VSMC phenotypic switching which is significantly prevented by cyclamNO. Compared to the NO donor sodium nitroprusside, cyclamNO showed to be significantly less cytotoxic. CONCLUSIONS: With great potential to maintain VSMC functionality and prevent IH-associated events, cyclamNO might be a promissory drug for several applications in cardiovascular medicine, as in stents.

Reactivity, photolability, and computational studies of the ruthenium nitrosyl complex with a substituted cyclam fac-[Ru(NO)Cl2(κ3N4,N8,N11(1-carboxypropyl)cyclam)]Cl·H2O.

Chemical reactivity, photolability, and computational studies of the ruthenium nitrosyl complex with a substituted cyclam, fac-[Ru(NO)Cl(2)(κ(3)N(4),N(8),N(11)(1-carboxypropyl)cyclam)]Cl·H(2)O ((1-carboxypropyl)cyclam = 3-(1,4,8,11-tetraazacyclotetradecan-1-yl)propionic acid)), (I) are described. Chloride ligands do not undergo aquation reactions (at 25 °C, pH 3). The rate of nitric oxide (NO) dissociation (k(obs-NO)) upon reduction of I is 2.8 s(-1) at 25 ± 1 °C (in 0.5 mol L(-1) HCl), which is close to the highest value found for related complexes. The uncoordinated carboxyl of I has a pK(a) of ∼3.3, which is close to that of the carboxyl of the non coordinated (1-carboxypropyl)cyclam (pK(a) = 3.4). Two additional pK(a) values were found for I at ∼8.0 and ∼11.5. Upon electrochemical reduction or under irradiation with light (λ(irr) = 350 or 520 nm; pH 7.4), I releases NO in aqueous solution. The cyclam ring N bound to the carboxypropyl group is not coordinated, resulting in a fac configuration that affects the properties and chemical reactivities of I, especially as NO donor, compared with analogous trans complexes. Among the computational models tested, the B3LYP/ECP28MDF, cc-pVDZ resulted in smaller errors for the geometry of I. The computational data helped clarify the experimental acid-base equilibria and indicated the most favourable site for the second deprotonation, which follows that of the carboxyl group. Furthermore, it showed that by changing the pH it is possible to modulate the electron density of I with deprotonation. The calculated NO bond length and the Ru/NO charge ratio indicated that the predominant canonical structure is [Ru(III)NO], but the Ru-NO bond angles and bond index (b.i.) values were less clear; the angles suggested that [Ru(II)NO(+)] could contribute to the electronic structure of I and b.i. values indicated a contribution from [Ru(IV)NO(-)]. Considering that some experimental data are consistent with a [Ru(II)NO(+)] description, while others are in agreement with [Ru(III)NO], the best description for I would be a linear combination of the three canonical forms, with a higher weight for [Ru(II)NO(+)] and [Ru(III)NO].

Cyclam kappa4 to kappa3 ligand denticity change upon mono-n-substitution with a carboxypropyl pendant arm in a ruthenium nitrosyl complex.

The complex fac-[Ru(NO)Cl2(kappa(3)N(4),N(8),N(11)(1-carboxypropyl)cyclam)]Cl.H2O (1-carboxypropyl)cyclam=3-(1,4,8,11-tetraazacyclotetradecan-1-yl)propionic acid) was prepared in a one pot reaction by mixing equimolar amounts of RuNOCl 3 and (1-carboxypropyl)cyclam and was characterized by X-ray crystallography, electrospray ionization tandem mass spectrometry (ESI-MS/MS), elemental analysis, NMR, and electronic and vibrational (IR) spectroscopies. fac-[Ru(NO)Cl 2(kappa(3)N(4),N(8),N(11)(1-carboxypropyl)cyclam)]Cl.H2O crystallizes in the triclinic, space group P1, No. 2, with unit cell parameters of a=8.501(1) A, b=9.157(1) A, c=14.200(1) A, alpha=72.564(5) degrees , beta=82.512(5) degrees , gamma=80.308(5) degrees , and Z=2. The Ru-N interatomic distance and bond angle in the [Ru-NO] unit are 1.739(2) A and 167.7(2) degrees , respectively. ESI-MS/MS shows characteristic dissociation chemistry that initiates by HCl or NO loss. The IR spectrum displays a nu(NO) at 1881 cm(-1) indicating a nitrosonium character. The electronic spectrum shows absorptions bands at 264 nm (log epsilon=3.27), 404 nm (log epsilon=2.53), and 532 nm (log epsilon=1.88). (1)H and (13)C NMR are in agreement with the proposed molecular structure, which shows a very singular architecture where the cyclam ring N (with the carboxypropyl pendant arm) is not coordinated to the ruthenium resulting in a kappa(3) instead of the expected kappa(4) denticity.