Cell-Tak coating interferes with DNA-based normalization of metabolic flux data.

Metabolic flux investigations of cells and tissue samples are a rapidly advancing tool in diverse research areas. Reliable methods of data normalization are crucial for an adequate interpretation of results and to avoid a misinterpretation of experiments and incorrect conclusions. The most common methods for metabolic flux data normalization are to cell number, DNA and protein. Data normalization may be affected by a variety of factors, such as density, healthy state, adherence efficiency, or proportional seeding of cells. The mussel-derived adhesive Cell-Tak is often used to immobilize poorly adherent cells. Here we demonstrate that this coating strongly affects the fluorescent detection of DNA leading to an incorrect and highly variable normalization of metabolic flux data. Protein assays are much less affected and cell counting can virtually completely remove the effect of the coating. Cell-Tak coating also affects cell shape in a cell line-specific manner and may change cellular metabolism. Based on these observations we recommend cell counting as a gold standard normalization method for Seahorse metabolic flux measurements with protein content as a reasonable alternative.

Modification of human pericardium by chemical crosslinking.

Autologous and allogenic human pericardia used as biomaterials for cardiovascular surgery are traditionally crosslinked with glutaraldehyde. In this work, we have evaluated the resistivity to collagenase digestion and the cytotoxicity of human pericardium crosslinked with various concentrations of glutaraldehyde in comparison with pericardium crosslinked by genipin, nordihydroguaiaretic acid, tannic acid, and in comparison with unmodified pericardium. Crosslinking retained the wavy-like morphology of native pericardium visualized by second harmonic generation microscopy. The collagenase digestion products were analyzed using SDS-PAGE, capillary electrophoresis, and a hydroxyproline assay. Glutaraldehyde and genipin crosslinking protected the native pericardium efficiently against digestion with collagenase III. Only low protection was provided by the other crosslinking agents. The cytotoxicity of crosslinked pericardium was evaluated using xCELLigence by monitoring the viability of porcine valve interstitial cells cultured in eluates from crosslinked pericardium. The highest cell index, reflecting both the number and the shape of the monitored cells was observed in eluates from genipin. Crosslinking pericardium grafts with genipin therefore seems to be a promising alternative procedure to the traditional crosslinking with glutaraldehyde, because it provides similarly high protection against degradation with collagenase, without cytotoxic effects.

Analysis by gas chromatography-mass spectrometry of the essential oil from the aerial parts of Pimpinella junoniae Ceb. & Ort., gathered in La Gomera, Canary Islands, Spain.

The essential oil from the aerial parts of Pimpinella junoniae Ceb. & Ort., growing in La Gomera, Canary Islands, Spain, was studied by gas chromatography and gas chromatography-mass spectrometry, and 43 constituents were identified. The major components were found to be alpha-zingiberene (20.6%), alpha-pinene (17.9%), (E)-beta-farnesene (9.3%), ar-curcumene (7.4%), beta-phellandrene (7.0%), beta-bisabolene (6.1%) and epoxypseudoisoeugenyl 2-methylbutyrate (6.0%). The decomposition product of epoxypseudoisoeugenol derivatives, 5-methoxy-2-methylbenzofuran (5.7%), moderate amounts of other arylpropanoids with the pseudoisoeugenol skeleton (total percentage, 5.2%) and other compounds such as beta-sesquiphellandrene (3.0%), cis-beta-guaiene (1.5%), alpha-phellandrene (1.5%) and alpha-bisabolol (1.3%), were also found.

New charge transfer salts based on bis(ethylenedithio)tetrathiafulvalene (ET) and ferro- or antiferromagnetic oxalato-bridged dinuclear anions: syntheses, structures and magnetism of ET5[MM'(C2O4)(NCS)8] with MM' = Cr(III)Fe(III), Cr(III)Cr(III).

Electrochemical combination of the magnetic dinuclear anion [MM'(C2O4)(NCS)8](4-) (MM' = Cr(III)Cr(III), Cr(III)Fe(III)) with the ET organic pi-donor (ET = BEDT-TTF = bis(ethylenedithio)tetrathiafulvalene) gives rise to two new isostructural molecular hybrid salts ET5[MM'(C2O4)(NCS)8], with MM' = CrCr (1), CrFe (2). The molecular structure of compound 1 has been determined by single crystal X-ray diffraction. The particular arrangement of the organic units consists of an unprecedented two-dimensional organic sublattice nearly similar to that observed in kappa-phase structures. For both compounds, the magnetic susceptibility measurements indicate (i) the ET radicals do not contribute to the magnetic moment probably due to the presence of strong antiferromagnetic interaction between them, and (ii) in the anion, the magnetic coupling is antiferromagnetic for 1 (J = -3.65 cm(-1)) and ferromagnetic for 2 (J = 1.14 cm(-1), J being the parameter of the exchange Hamiltonian H = -2JS1S2). The field dependence of the magnetization of compound 2 at 2.0 K gives further evidence of the S = 4 ground-state arising from the interaction between S = 3/2 Cr(III) and S = 5/2 Fe(III). EPR measurements confirm the nature of the magnetic interactions and the absence of any contribution from the organic part, as observed from the static magnetic measurement. Conductivity measurements and electronic band structure calculations show that both salts are semiconductors with low activation energies.

Heptanuclear hydroxo-bridged copper cluster of the dicubane-like type: structural and magnetic characterizations of [Cu7(OH)6Cl2(pn)6(H2O)2](C(CN)3)4Cl2 (pn = 1,3-diaminopropane).

A new polynuclear copper(II) complex [Cu7(OH)6Cl2(pn)6(H2O)2](C(CN)3)4Cl2 with hydroxo-bridging ligands has been prepared; the centrosymmetric cluster cation can be described as two Cu4O3Cl distorted cubane units sharing one copper cation.

Oxalato-bridged dinuclear complexes of Cr(III) and Fe(III): synthesis, structure, and magnetism of [(C2H5)4N]4[MM'(ox)(NCS)8] with MM' = CrCr, FeFe, and CrFe.

A new series of homo- and heterometallic oxalato-bridged dinuclear compounds of formulas [Et4N]4[MM'(ox)(NCS)8] ([Et4N]+ = [(C2H5)4N]+; ox = C2O4(2-)) with MM' = Cr(III)-Cr(III) (1), Fe(III)-Fe(III) (2), and Cr(III)-Fe(III) (3) is reported. They have been structurally characterized by infrared spectra and single-crystal X-ray diffraction. The three compounds are isostructural and crystallize in the orthorhombic space group Cmca with Z = 8, a = 16.561(8) A, b = 13.481(7) A, and c = 28.168(8) A for 1, a = 16.515(2) A, b = 13.531(1) A, and c = 28.289(4) A for 2, a = 16.664(7) A, b = 13.575(6) A, and c = 28.386(8) A for 3. The structure of 3 is made up of a discrete dinuclear anion [CrFe(ox)(NCS)8]4- and four disordered [Et4N]+ cations, each of them located on special positions. The anion, in a crystallographically imposed C2h symmetry, contains metal cations in distorted octahedral sites. The Cr(ox)Fe group, which is planar within 0.02 A, presents an intramolecular metal-metal distance of 5.43 A. Magnetic susceptibility measurements indicate antiferromagnetic pairwise interactions for 1 and 2 with J = -3.23 and -3.84 cm-1, respectively, and ferromagnetic Cr-Fe coupling with J = 1.10 cm-1 for 3 (J being the parameter of the exchange Hamiltonian H = -2JS1S2). The ESR spectra at different temperatures confirm the magnetic susceptibility data.

Minimally invasive coronary artery bypass grafting for myocardial muscle bridging.

Myocardial bridging is a congenital anomaly of the left anterior descending coronary artery (LAD), which is associated with myocardial ischemia and infarction, cardiac arrhythmias, and sudden cardiac death. Two cases are reported of symptomatic myocardial bridging refractory to medical management treated by minimally invasive coronary artery bypass grafting without cardiopulmonary bypass. We conclude that minimally invasive coronary artery bypass techniques are appropriate alternatives to endovascular stent placement, muscle bridge division, or aortocoronary grafting with cardiopulmonary bypass for the management of symptomatic myocardial bridging.

Hemodynamic changes in patients developing effective hypovolemia after total paracentesis.

BACKGROUND/AIMS: In many centers paracentesis is considered the treatment of choice for tense ascites. However, the mechanism of effective hypovolemia after paracentesis, the main complication associated with this procedure, remains unknown. In the current study, systemic hemodynamics was sequentially studied before and after total paracentesis in 46 patients with cirrhosis and tense ascites. The aim of the study was to assess the mechanism of effective hypovolemia after paracentesis. METHODS: Plasma renin activity and aldosterone, mean arterial pressure, cardiac output (ECO-Doppler) and systemic vascular resistance were measured before, and 3 h, 6 h and 6 days after total paracentesis associated with plasma volume expansion. RESULTS: Effective hypovolemia after paracentesis (defined as 50% increase in plasma renin activity up to a level over 4 ng x m(-1) x h(-1) at the 6th day after paracentesis) occurred in 20 cases [plasma renin activity increased from 8+/-17 to 19+/-2.7 ng x m(-1) x h(-1)]. In the remaining 26 cases no changes in plasma renin activity [8.5+/-2.4 vs. 8.7+/-2.2 ng x m(-1) x h(-1)] were observed. The amounts of ascitic fluid volume removed were similar. Effective hypovolemia after paracentesis was associated with a significant decrease in mean arterial pressure (89+/-2 vs. 81+/-3 mmHg) and systemic vascular resistance [1263+/-67 vs. 1014+/-80 dyn x s(-1) x cm(-5)] 6 days after treatment. In contrast, no significant changes in these parameters were observed in patients not developing this complication. In the whole group of patients a significant inverse relation was observed between changes in plasma renin activity and in systemic vascular resistance (r=0.74;p< 0.001). CONCLUSIONS: These results indicate that effective hypovolemia after paracentesis in cirrhosis is predominantly due to an accentuation of the arteriolar vasodilation already present in these patients.