THE USE OF TETRAZOLIUM SALTS TO DETERMINE SITES OF DAMAGE TO THE MITOCHONDRIAL ELECTRON TRANSPORT CHAIN IN INTACT CELLS FOLLOWING in vitro: PHOTODYNAMIC THERAPY WITH PHOTOFRIN II

Abstract: A method is described utilizing the tetrazolium salts neotetrazolium chloride (NTC), triphenyltetrazolium chloride (TTC), C, N -diphenyl- N' -4,5-dimethylthiazol-2-yrtetrazolium bromide (MTT) and various substrates to elucidate damage to the mitochondrial electron transport chain of intact cells following in vitro photodynamic therapy (PDT). Using this methodology, a portion of the dark toxicity manifested by Photofrin II (PII) was found to occur prior to entry of electrons into the transport chain through Complex I, as evidenced by the fact that the inhibition of MTT reduction was reversible by the addition of malic acid to the culture media. A second site of dark toxicity was found to be Complex IV (cytochrome oxidase). After photoirradiation of the cells, Complex I was found to be affected since malic acid could no longer reverse the inhibition of MTT reduction but it could be reversed by the addition of succinic acid, whose electrons enter the transport chain at Complex II. A second and more sensitive site of photoirradiation damage was found to be Complex IV. A region near cytochrome C was also affected by photoirradiation but appreciably less so than noted for Complexes I and IV. A kinetic analysis of MTT and TTC reduction following photoirradiation indicated that MTT reduction was sustained at a normal rate for 1 h after which it slowed down and eventually plateaued. In contrast, TTC reduction was found to be inhibited almost immediately indicating Complex IV is extremely susceptible to photoirradiation damage. Compared to other assays of mitochondrial function requiring subcellular fractionation, the use of tetrazolium salts is simpler to perform and can be done using physiologically relevant conditions.     

The anatomic site of photodynamic therapy is a determinant for immunosuppression in a murine model

The role of the irradiation site in the induction of suppression of the contact hypersensitivity (CHS) response following photodynamic therapy (PDT) was examined in a murine model. Laser irradiation on the flanks of nontumor-bearing Photofrin-injected mice caused suppression of the CHS response. If the irradiation was conducted on a subcutaneously implanted foil disc on the flank no immunosuppression occurred, indicating that no suppressive factor(s) of sufficient quantity to cause suppression was released from the skin, but rather irradiation of internal organs was the cause. Irradiation of tumors implanted on the flanks of mice reduced the suppression, suggesting an immunopotentiating effect of PDT. Irradiation on the thigh in the presence or absence of a tumor gave no immunosuppression. These results suggest that the anatomic site of irradiation is one determinant for the elicitation of suppression of the CHS response.     

Proton relaxation enhancement by manganese(III)TPPS4 in a model tumor system

Managanese(III)tetraphenylporphine sulfonate [Mn(III)TPPS4] has been investigated as a tumor specific paramagnetic contrast agent for magnetic resonance imaging (MRI) of L1210 solid tumors in mice. Mn(III)TPPS4 was found to clear rapidly from the blood and concentrate in the kidneys, tumor and liver. Although relatively high ratios of tumor to normal tissues could be obtained (e.g., greater than 90 for tumor/muscle), the kidneys were found to have the highest concentration of the metalloporphyrin at all doses and time periods tested. A significant decrease in the longitudinal relaxation time was measured for excised tissues (kidney, tumor, liver, muscle) from mice that were treated with Mn(III)TPPS4. A linear correlation was observed between the longitudinal relaxation rate determined for L1210 tumor and the corresponding concentration of Mn(III)TPPS4 found at various injected doses and time intervals between the injection and analysis. A small animal radiofrequency receiver coil designed for use with a 0.15-T clinical imager was employed to evaluate the ability of Mn(III)TPPS4 to selectively increase the signal intensity of the implanted L1210 tumor. The images show a conspicuous enhancement in the contrast between the tumor and adjacent tissue upon treatment with this agent. The results indicate that Mn(III)TPPS4 is a useful prototype paramagnetic metalloporphyrin MRI contrast agent with a significant affinity for the L1210 tumor.     

Gårding's Theory of Hyperbolic Polynomials

This paper presents a simple, self‐contained account of Gårding's theory of hyperbolic polynomials, together with a recent convexity result of Bauschke‐Güler‐Lewis‐Sendov and an inequality of Gurvits. This account begins by establishing some new results. The first concerns the existence of a pointwise arrangement of the eigenvalues so that they become global real analytic functions. The second asserts that the associated “branches” are independent of the choice of hyperbolic direction. © 2013 Wiley Periodicals, Inc.     

Thermal Characterizations of Inorganic and Organometallic Polymers

This review emphasizes the breadth of metallic and metallic-like polymers evaluated as to thermal properties. Techniques usefully applied to particular systems are noted with the aim of suggesting their application to other systems.     

Low-lying energy levels of the FeH molecule

Multireference configuration interaction (MRCI) and complete active space second-order perturbation theory (CASPT2) calculations are performed on Fe₂ and Fe^? ₂. Although it is not possible to definitively identify the ground states of Fe₂ and Fe^? ₂, the calculations suggest that the ground state of Fe^? ₂ in ⁸Σ^? _u derived from 3d¹³4σ² _g4σ² _u and that the states observed in photodetachment are the ⁹Σ^? _g and ⁷Σ^? _g states with a 3d¹³4σ² _g4σ¹ _u occupation, but that the ground state of Fe₂ is ⁷Δ_u(3d¹⁴4σ² _g) and is not observed in the photo-detachment spectra.     

Data Formats for Elementary Gas Phase Kinetics,Part 1: Unique Representations of Species at the Molecular Level

Standardized electronic formats for data are needed to efficiently and transparently communicate the results of scientific studies. A format for the unique identification of chemical species is a requirement in the field of chemistry, and the IUPAC International Chemical Identifier (InChI) has been widely adopted for this purpose. The InChI identifier has proved to be very useful. The InChI identifier, however, is currently insufficient to uniquely specify some types of molecular entities at a detailed molecular level needed to fully characterize their chemical nature, to differentiate between chemically distinct conformers, to uniquely identify structures used in quantum chemical calculations, and to completely describe elementary chemical reactions. To address this limitation, we propose an augmented form of InChI, denoted as InChI–ER, which contains additional optional layers that allow the unique and unambiguous identification of molecules at a detailed molecular level. The new layers proposed herein are optional extensions of the existing InChI formalism and, like all other InChI layers, would not interfere with InChI identifiers currently in use. The focus of the present work is the better specification of required molecular entities such as rotational conformations, ring conformations, and electronic states. In companion articles, we propose additional reaction layers using an extended InChI format that will enable the unique identification of elementary chemical reactions, including specification of associated transition states, specification of the changes in bonds that occur during reaction, and classification of reaction types.     

A Simple One-Step Synthesis of Alkyl Benzazol-2-carboxylates

A simple one-step synthesis of alkyl benzazol-2-carboxylates employing alkyl trialkoxyacetate is described.