Transcriptomic and proteomic pathways of diabetic and non-diabetic mitochondrial transplantation.

Reduced mitochondrial function increases myocardial susceptibility to ischemia-reperfusion injury (IRI) in diabetic hearts. Mitochondrial transplantation (MT) ameliorates IRI, however, the cardioprotective effects of MT may be limited using diabetic mitochondria. Zucker Diabetic Fatty (ZDF) rats were subjected to temporary myocardial RI and then received either vehicle alone or vehicle containing mitochondria isolated from either diabetic ZDF or non-diabetic Zucker lean (ZL) rats. The ZDF rats were allowed to recover for 2 h or 28 days. MT using either ZDF- or ZL-mitochondria provided sustained reduction in infarct size and was associated with overlapping upregulation of pathways associated with muscle contraction, development, organization, and anti-apoptosis. MT using either ZDF- or ZL-mitochondria also significantly preserved myocardial function, however, ZL- mitochondria provided a more robust long-term preservation of myocardial function through the mitochondria dependent upregulation of pathways for cardiac and muscle metabolism and development. MT using either diabetic or non-diabetic mitochondria decreased infarct size and preserved functional recovery, however, the cardioprotection afforded by MT was attenuated in hearts receiving diabetic compared to non-diabetic MT.

Mitochondrial remodeling and ischemic protection by G protein-coupled receptor 35 agonists.

Kynurenic acid (KynA) is tissue protective in cardiac, cerebral, renal, and retinal ischemia models, but the mechanism is unknown. KynA can bind to multiple receptors, including the aryl hydrocarbon receptor, the a7 nicotinic acetylcholine receptor (a7nAChR), multiple ionotropic glutamate receptors, and the orphan G protein-coupled receptor GPR35. Here, we show that GPR35 activation was necessary and sufficient for ischemic protection by KynA. When bound by KynA, GPR35 activated Gi- and G12/13-coupled signaling and trafficked to the outer mitochondria membrane, where it bound, apparantly indirectly, to ATP synthase inhibitory factor subunit 1 (ATPIF1). Activated GPR35, in an ATPIF1-dependent and pertussis toxin-sensitive manner, induced ATP synthase dimerization, which prevented ATP loss upon ischemia. These findings provide a rationale for the development of specific GPR35 agonists for the treatment of ischemic diseases.

A Large Animal Model for Acute Kidney Injury by Temporary Bilateral Renal Artery Occlusion.

Acute kidney injury (AKI) is associated with higher risk for morbidity and mortality post-operatively. Ischemia-reperfusion injury (IRI) is the most common cause of AKI. To mimic this clinical scenario, this study presents a highly reproducible large animal model of renal IRI in swine using temporary percutaneous bilateral balloon-catheter occlusion of the renal arteries. The renal arteries are occluded for 60 min by introducing the balloon-catheters through the femoral and carotid artery and advancing them into the proximal portion of the arteries. Iodinated contrast is injected in the aorta to assess any opacification of the kidney vessels and confirm the success of the artery occlusion. This is furtherly confirmed by the flattening of the pulse waveform at the tip of the balloon catheters. The balloons are deflated and removed after 60 min of bilateral renal artery occlusion, and the animals are allowed to recover for 24 h. At the end of the study, plasma creatinine and blood urea nitrogen significantly increase, while eGFR and urine output significantly decrease. The need for iodinated contrast is minimal and does not affect renal function. Bilateral renal artery occlusion better mimics the clinical scenario of perioperative renal hypoperfusion, and the percutaneous approach minimizes the impact of the inflammatory response and the risk of infection seen with an open approach, such as a laparotomy. The ability to create and reproduce this clinically relevant swine model eases the clinical translation to humans.

Electron-Nucleus Hyperfine Coupling Calculated from Restricted Active Space Wavefunctions and an Exact Two-Component Hamiltonian.

Exact two-component (X2C) relativistic nuclear hyperfine magnetic field operators were incorporated in X2C ab initio wavefunction calculations at the multireference restricted active space (RAS) level for calculations of nuclear hyperfine magnetic properties. Spin-orbit coupling was treated via RAS state interaction (SO-RASSI). The method was tested by calculations of electron-nucleus hyperfine coupling constants. The approach, implemented in the OpenMolcas program, overcomes a major limitation of a previous SO-RASSI implementation for hyperfine coupling that relied on nonrelativistic hyperfine operators [J. Chem. Theor. Comput. 2015, 11, 538-549] and therefore had limited applicability. Results from calculations on systems with light and heavy main group elements, transition metals, lanthanides, and one actinide complex demonstrate reasonably good agreement with experimental data, where available, as long as the active space can generate sufficient spin polarization.

A Multi-Mode System for Myocardial Functional and Physiological Assessment during Ex Situ Heart Perfusion.

Ex situ heart perfusion (ESHP) has proven to be an important and valuable step toward better preservation of donor hearts for heart transplantation. Currently, few ESHP systems allow for a convenient functional and physiological evaluation of the heart. We sought to establish a simple system that provides functional and physiological assessment of the heart during ESHP. The ESHP circuit consists of an oxygenator, a heart-lung machine, a heater-cooler unit, an anesthesia gas blender, and a collection funnel. Female Yorkshire pig hearts (n = 10) had del Nido cardioplegia (4°C) administered, excised, and attached to the perfusion system. Hearts were perfused retrogradely into the aortic root for 2 hours before converting the system to an isovolumic mode or a working mode for further 2 hours. Blood samples were analyzed to measure metabolic parameters. During the isovolumic mode (n = 5), a balloon inserted in the left ventricular (LV) cavity was inflated so that an end-diastolic pressure of 6-8 mmHg was reached. During the working mode (n = 5), perfusion in the aortic root was redirected into left atrium (LA) using a compliance chamber which maintained an LA pressure of 6-8 mmHg. Another compliance chamber was used to provide an afterload of 40-50 mmHg. Hemodynamic and metabolic conditions remained stable and consistent for a period of 4 hours of ESHP in both isovolumic mode (LV developed pressure: 101.0 ± 3.5 vs. 99.7 ± 6.8 mmHg, p = .979, at 2 and 4 hours, respectively) and working mode (LV developed pressure: 91.0 ± 2.6 vs. 90.7 ± 2.5 mmHg, p = .942, at 2 and 4 hours, respectively). The present study proposed a novel ESHP system that enables comprehensive functional and metabolic assessment of large mammalian hearts. This system allowed for stable myocardial function for up to 4 hours of perfusion, which would offer great potential for the development of translational therapeutic protocols to improve dysfunctional donated hearts.

Mitochondrial transplantation by intra-arterial injection for acute kidney injury.

Acute kidney injury is a common clinical disorder and one of the major causes of morbidity and mortality in the postoperative period. In this study, the safety and efficacy of autologous mitochondrial transplantation by intra-arterial injection for renal protection in a swine model of bilateral renal ischemia-reperfusion injury were investigated. Female Yorkshire pigs underwent percutaneous bilateral temporary occlusion of the renal arteries with balloon catheters. Following 60 min of ischemia, the balloon catheters were deflated and animals received either autologous mitochondria suspended in vehicle or vehicle alone, delivered as a single bolus to the renal arteries. The injected mitochondria were rapidly taken up by the kidney and were distributed throughout the tubular epithelium of the cortex and medulla. There were no safety-related issues detected with mitochondrial transplantation. Following 24 h of reperfusion, estimated glomerular filtration rate and urine output were significantly increased while serum creatinine and blood urea nitrogen were significantly decreased in swine that received mitochondria compared with those that received vehicle. Gross anatomy, histopathological analysis, acute tubular necrosis scoring, and transmission electron microscopy showed that the renal cortex of the vehicle-treated group had extensive coagulative necrosis of primarily proximal tubules, while the mitochondrial transplanted kidney showed only patchy mild acute tubular injury. Renal cortex IL-6 expression was significantly increased in vehicle-treated kidneys compared with the kidneys that received mitochondrial transplantation. These results demonstrate that mitochondrial transplantation by intra-arterial injection provides renal protection from ischemia-reperfusion injury, significantly enhancing renal function and reducing renal damage.

Mitochondrial transplantation for myocardial protection in ex-situ‒perfused hearts donated after circulatory death.

BACKGROUND: Donation after circulatory death (DCD) offers an additional source of cardiac allografts, potentially allowing expansion of the donor pool, but is limited owing to the effects of ischemia. In this study, we investigated the efficacy of mitochondrial transplantation to enhance myocardial function of DCD hearts. METHODS: Circulatory death was induced in Yorkshire pigs (40-50 kg, n = 29) by a cessation of mechanical ventilation. After 20 minutes of warm ischemia, cardioplegia was administered. The hearts were then reperfused on an ex-situ blood perfusion system. After 15 minutes of reperfusion, hearts received either vehicle alone (vehicle [VEH], 10 ml; n = 8) or vehicle containing autologous mitochondria (vehicle with mitochondria as a single injection [MT], 5 × 109 in 10 ml, n = 8). Another group of hearts (serial injection of mitochondria [MTS]; n = 6) received a second injection of mitochondria (5 × 109 in 10 ml) after 2 hours of ex-situ heart perfusion and reperfused for an additional 2 hours. A Sham group (sham hearts; n = 6) did not undergo any warm ischemia. RESULTS: At the end of 4 hours of reperfusion, MT and MTS groups showed a significantly increased left ventricle/ventricular peak developed pressure (p = 0.002), maximal left ventricle/ventricular pressure rise (p < 0.001), fractional shortening (p < 0.001), and myocardial oxygen consumption (p = 0.004) compared with VEH. Infarct size was significantly decreased in MT and MTS groups compared with VEH (p < 0.001). No differences were found in arterial lactate levels among or within groups throughout reperfusion. CONCLUSIONS: Mitochondrial transplantation significantly preserves myocardial function and oxygen consumption in DCD hearts, thus providing a possible option for expanding the heart donor pool.

Mitochondrial transplantation for myocardial protection in diabetic hearts.

OBJECTIVES: Type 2 diabetes causes mitochondrial dysfunction, which increases myocardial susceptibility to ischaemia-reperfusion injury. We investigated the efficacy of transplantation of mitochondria isolated from diabetic or non-diabetic donors in providing cardioprotection from warm global ischaemia and reperfusion in the diabetic rat heart. METHODS: Ex vivo perfused hearts from Zucker diabetic fatty (ZDF fa/fa) rats (n = 6 per group) were subjected to 30 min of warm global ischaemia and 120 min reperfusion. Immediately prior to reperfusion, vehicle alone (VEH) or vehicle containing mitochondria isolated from either ZDF (MTZDF) or non-diabetic Zucker lean (ZL +/?) (MTZL) skeletal muscle were delivered to the coronary arteries via the aortic cannula. RESULTS: Following 30-min global ischaemia and 120-min reperfusion, left ventricular developed pressure was significantly increased in MTZDF and MTZL groups compared to VEH group (MTZDF: 92.8 ± 5.2 mmHg vs MTZL: 110.7 ± 2.4 mmHg vs VEH: 44.3 ± 5.9 mmHg; P < 0.01 each); and left ventricular end-diastolic pressure was significantly decreased (MTZDF 12.1 ± 1.3 mmHg vs MTZL 8.6 ± 0.8 mmHg vs VEH: 18.6 ± 1.5 mmHg; P = 0.016 for MTZDF vs VEH and P < 0.01 for MTZL vs VEH). Total tissue ATP content was significantly increased in both MT groups compared to VEH group (MTZDF: 18.9 ± 1.5 mmol/mg protein/mg tissue vs MTZL: 28.1 ± 2.3 mmol/mg protein/mg tissue vs VEH: 13.1 ± 0.5 mmol/mg protein/mg tissue; P = 0.018 for MTZDF vs VEH and P < 0.01 for MTZL vs VEH). Infarct size was significantly decreased in the MT groups (MTZDF: 11.8 ± 0.7% vs MTZL: 9.9 ± 0.5% vs VEH: 52.0 ± 1.4%; P < 0.01 each). CONCLUSIONS: Mitochondrial transplantation significantly enhances post-ischaemic myocardial functional recovery and significantly decreases myocellular injury in the diabetic heart.

Ab Initio Study of Covalency in the Ground versus Core-Excited States and X-ray Absorption Spectra of Actinide Complexes.

Relativistic multireference ab initio wave function calculations within the restricted active space (RAS) framework were performed to calculate metal and ligand X-ray absorption (XAS) near-edge spectroscopy (XANES) intensities for the metal M4,5 edges of [PuO2(H2O)5]2+, [AnVIO2]2+ (An = U, Np, Pu), and [AmCl6]3- and the Cl K edge of the Am complex. The extent of An(5f)-ligand bonding was determined via natural localized molecular orbital analyses of the relevant spin-orbit coupled multireference states. The calculated spectra are in good agreement with experiments and allow a detailed assignment of the observed spectral features. The XANES M4,5-edge spectra are representative of the actinide orbital covalency in the probed core-excited states, which may be different from the ground-state covalency. An assignment of ground-state An orbital covalency based on XAS spectra should therefore be made with caution.

A diuranium carbide cluster stabilized inside a C80 fullerene cage.

Unsupported non-bridged uranium-carbon double bonds have long been sought after in actinide chemistry as fundamental synthetic targets in the study of actinide-ligand multiple bonding. Here we report that, utilizing Ih(7)-C80 fullerenes as nanocontainers, a diuranium carbide cluster, U=C=U, has been encapsulated and stabilized in the form of UCU@Ih(7)-C80. This endohedral fullerene was prepared utilizing the Krätschmer-Huffman arc discharge method, and was then co-crystallized with nickel(II) octaethylporphyrin (NiII-OEP) to produce UCU@Ih(7)-C80·[NiII-OEP] as single crystals. X-ray diffraction analysis reveals a cage-stabilized, carbide-bridged, bent UCU cluster with unexpectedly short uranium-carbon distances (2.03 Å) indicative of covalent U=C double-bond character. The quantum-chemical results suggest that both U atoms in the UCU unit have formal oxidation state of +5. The structural features of UCU@Ih(7)-C80 and the covalent nature of the U(f1)=C double bonds were further affirmed through various spectroscopic and theoretical analyses.

A Pseudotetrahedral Uranium(V) Complex.

A series of uranium amides were synthesized from N, N, N-cyclohexyl(trimethylsilyl)lithium amide [Li][N(TMS)Cy] and uranium tetrachloride to give U(NCySiMe3) x(Cl)4- x, where x = 2, 3, or 4. The diamide was isolated as a bimetallic, bridging lithium chloride adduct ((UCl2(NCyTMS)2)2-LiCl(THF)2), and the tris(amide) was isolated as the lithium chloride adduct of the monometallic species (UCl(NCyTMS)3-LiCl(THF)2). The tetraamide complex was isolated as the four-coordinate pseudotetrahedron. Cyclic voltammetry revealed an easily accessible reversible oxidation wave, and upon chemical oxidation, the UV amido cation was isolated in near-quantitative yields. The synthesis of this family of compounds allows a direct comparison of the electronic structure and properties of isostructural UIV and UV tetraamide complexes. Spectroscopic investigations consisting of UV-vis, NIR, MCD, EPR, and U L3-edge XANES, along with density functional and wave function calculations, of the four-coordinate UIV and UV complexes have been used to understand the electronic structure of these pseudotetrahedral complexes.

Influence of Substituents on the Electronic Structure of Mono- and Bis(phosphido) Thorium(IV) Complexes.

A series of metallocene thorium complexes with mono- and bis(phosphido) ligands have been investigated with varying hues: (C5Me5)2Th(Cl)[P(Mes)2] (Mes = mesityl = 2,4,6-(CH3)3C6H2; dark red-purple), (C5Me5)2Th[P(Mes)(CH3)]2 (dark red-purple), (C5Me5)2Th(CH3)[P(Mes)2] (dark red-purple), (C5Me5)2Th(CH3)[P(Mes)(SiMe3)] (orange), (C5Me5)2Th(Cl)[P(Mes)(SiMe3)] (orange), (C5Me5)2Th[P(Mes)(SiMe3)]2 (orange), and (C5Me5)2Th[PH(Mes)]2 (pale yellow). While all of these complexes bear a mesityl group on phosphorus, the electronic structure observed differs depending on the other substituent (mesityl, methyl, trimethylsilyl, or hydrogen). This sparked an investigation of the electronic structure of these complexes using 31P NMR and electronic absorption spectroscopy in concert with time-dependent density functional theory calculations.

Calculation of linear and nonlinear optical properties of azobenzene derivatives with Kohn-Sham and coupled-cluster methods.

Linear polarizabilities (α) and second hyperpolarizabilities (γ) of unsubstituted azobenzenes and 'push-pull' azobenzene derivatives are investigated using Kohn-Sham theory (KST) and coupled-cluster (CC) approaches. Various standard exchange-correlation functionals as well as a non-empirically 'tuned' long-range corrected (LC) functional with range-separated exchange are used in the KST calculations. When compared to correlated ab initio calculations and measurements, the tuned functional gives accurate low-energy excitation energies, especially for charge transfer (CT) transitions, and performs well for α. Basis set and solvent effects are also studied. In contrast to expectations, but in agreement with a prior study of π-conjugated systems that do not have low-energy CT excitations, the improvements of the CT excitation energies for the push-pull π-chromophores due to tuning do not go along with clear improvements of γ toward the CC reference data, likely due to the importance of the dynamic electron correlation for this property.

Assessment of Tuned Range Separated Exchange Functionals for Spectroscopies and Properties of Uranium Complexes.

The Kohn-Sham delocalization error (DE) is quantified in select uranium compounds for various functionals and shown to correlate with the magnitude of dative ligand donation into the 5f shell. Range separated exchange functionals are reparametrized to minimize the DE and analyzed for their spectroscopic predictive capabilities. Valence excitation spectra of occupied 5f systems exhibit noticeable improvement upon reparametrization, e.g. UCl6-, UCl62-, and UO2+. Less sensitivity to the reparameterization was observed for closed shell 5f systems and core excitation spectra. A general parametrization is proposed to perform well for valence excitation spectra with small DE.

Magnetic circular dichroism of UCl6- in the ligand-to-metal charge-transfer spectral region.

We present a combined ab initio theoretical and experimental study of the magnetic circular dichroism (MCD) spectrum of the octahedral UCl6- complex ion in the UV-Vis spectral region. The ground state is an orbitally non-degenerate doublet E5/2u and the MCD is a -term spectrum caused by spin-orbit coupling. Calculations of the electronic spectrum at various levels of theory indicate that differential dynamic electron correlation has a strong influence on the energies of the dipole-allowed transitions and the envelope of the MCD spectrum. The experimentally observed bands are assigned to dipole-allowed ligand-to-metal charge transfer into the 5f shell, and 5f to 6d transitions. Charge transfer excitations into the U 6d shell appear at much higher energies. The MCD-allowed transitions can be assigned via their signs of the -terms: Under Oh double group symmetry, E5/2u → E5/2g transitions have negative -terms whereas E5/2u → F3/2g transitions have positive -terms if the ground state g-factor is negative, as it is the case for UCl6-.

Impact of the Kohn-Sham Delocalization Error on the 4f Shell Localization and Population in Lanthanide Complexes.

The extent of ligand to metal donation bonding and mixing of 4f (and 5d) orbitals with ligand orbitals is studied by Kohn-Sham (KS) calculations for LaX3 (X = F, Cl, Br, I), GdX3, and LuX3 model complexes, CeCl6(2-), YbCp3, and selected lanthanide complexes with larger ligands. The KS delocalization error (DE) is quantified via the curvature of the energy for noninteger electron numbers. The extent of donation bonding and 4f-ligand mixing correlates well with the DE. For Lu complexes, the DE also correlates with the extent of mixing of ligand and 4f orbitals in the canonical molecular orbitals (MOs). However, the localized set of MOs and population analyses indicate that the closed 4f shell is localized. Attempts to create situations where mixing of 4f and ligand orbitals occurs due to a degeneracy of fragment orbitals were unsuccessful. For La(III) and, in particular, for Ce(IV), Hartree-Fock, KS, and coupled cluster singles and doubles calculations are in agreement in that excess 4f populations arise from ligand donation, along with donation into the 5d shell. Likewise, KS calculations for all systems with incompletely filled 4f shells, even those with "optimally tuned" functionals affording a small DE, produce varying degrees of excess 4f populations which may be only partially attributed to 5d polarization.