A novel mitochondrial signaling pathway activated by visible-to-near infrared radiation

The number of cells attached to glass substratum increases if HeLa cell suspension is irradiated with monochromatic visible-to-near infrared radiation before plating (the action spectrum with maxima at 619, 657, 675, 700, 740, 760, 800, 820, 840 and 860 nm). Treating of cell suspension with sodium azide (2 x 10(-5) M), sodium nitroprusside (5 x 10(-5) M), ouabain (1 x 10(-6) M) or amiloride (1.7 x 10(-5) M) before irradiation significantly modifies the spectrum of cell attachment enhancement. A light-induced mitochondrial signaling pathway can be regulated by small ligands directly binding to the catalytic center of cytochrome c oxidase (N(3), NO) as well as by chemicals specifically binding to plasma membrane enzymes (ouabain, amiloride). The comparative analysis of action spectra allows the conclusions that first, Cu(A) and Cu(B) chromophores of cytochrome c oxidase could be involved as photoacceptors and second, various signaling pathways (reaction channels) between cytochrome c oxidase and cell attachment regulation are at work.     

Green and red light emission by upconversion from the near-IR in Yb(3+) doped CsMnBr3

Direct near-IR excitation of Yb(3+) 2F(7/2)-->(2)F(5/2) levels at 10126, 10138, and 10596 cm(-1) in CsMnBr3:0.5%Yb(3+) leads to three types of luminescence at cryogenic temperatures: near-IR Yb(3+) emission and green and red upconverted luminescence. The green luminescence around 20 000 cm(-1) is identified as cooperative Yb(3+) pair upconversion. The broad red upconversion luminescence band centered at 14 700 cm(-1) is ascribed to the 4T(1g)-->6A(1g) transition of Mn(2+). Pulsed measurements indicate a sequence of ground-state absorption and excited-state absorption steps for the red upconversion process. One- and two-color excitation experiments support this, and we conclude that the red upconversion occurs by an exchange mechanism involving Yb(3+) and Mn(2+). The Yb(3+) 2F(5/2)-->(2)F(7/2) near-IR emission around 10 000 cm(-1) is also observed after Mn(2+) excitation at 21 838 cm(-1). This is indicative of a Mn(2+) 4T(1g)--> Yb(3+) 2F(5/2) relaxation process, which is a potential loss process for upconversion efficiency.     

Thermally induced transformation of mammalian red blood cells during hyperthermia

The structural and transport characteristics of membranes are mainly determined by the state of the cytoskeleton. The characteristic changes in morphology of human (adult donor and cord) and rat Red Blood Cells (RBC) and of their membrane, induced by hyperthermia (46-51 degrees C) have been analyzed. Two different types of morphological changes have been observed to take place during hyperthermia in all studied RBC groups. We have observed either formation and exfoliation of spiculas from membrane, resulting in the formation of large (4-5 mum) sphere-like cell body and small (0.5-1.5 mum) vesicles or cell fragmentation with formation of large (3-3.5 mum) vesicles. The two distinct phenomena are likely to be determined by the heterogeneity of the RBC population in terms of cell age. There was noted the difference of cord RBC from the donor ones in temperature value of transformation beginning, as well as the character of deformation and vesicle formation, that may testify to their less thermoresistance. The ultrastructure of the membrane, studied with the freeze-fracturing technique, testifies to an irreversible character of membrane changes. The aggregation of intramembrane particles (IMPs) as a continuous network testifies to the strengthening of the interactions between denatured spectrin and bilayer integral components.     

Mitochondrial and endoplasmic reticulum stress-induced apoptotic pathways are activated by 5-aminolevulinic acid-based photodynamic therapy in HL60 leukemia cells

We studied the mechanism of the cytotoxic effects of 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT; induction with 1 mM ALA for 4 h followed by a blue light dose of 18 J/cm(2)) on the human promyelocytic leukemia cell line HL60 using biochemical and electron microscopy methods. The disruption of mitochondrial membrane potential, deltapsi(m), was paralleled by a decrease in ATP level, unmasking of the mitochondrial antigen 7A6, release of cytochrome c into the cytoplasm, activation of caspases 9 and 3 and cleavage of poly(ADP-ribose) polymerase (PARP). This was followed by DNA fragmentation. These data suggest that ALA-PDT activates the mitochondrial apoptotic pathway. The level of endoplasmic reticulum Ca(2+)-binding chaperones ERp57 and ERp72 and of anti-apoptotic proteins Bcl-2 and Bcl-x(L) was decreased whereas that of Ca(2+)-binding protein calmodulin and the stress protein HSP60 was elevated following ALA-PDT. Inhibition of the initiator caspase 9, execution caspase 3 and Ca(2+)-dependent protease m-calpain, did not prevent DNA fragmentation. We conclude that, in our in vitro model, ALA-based photodynamic treatment initiates several signaling processes in HL60 cells that lead to rapidly progressing apoptosis, which is followed by slow necrosis. Two apoptotic processes proceed in parallel, one representing the mitochondrial pathway, the other involving disruption of calcium homeostasis and activation of the endoplasmic reticulum stress-mediated pathway.     

A near-IR reversible fluorescent probe modulated by selenium for monitoring peroxynitrite and imaging in living cells

We have developed a near-IR reversible fluorescent probe containing an organoselenium functional group that can be used for the highly sensitive and selective monitoring of peroxynitrite oxidation and reduction events under physiological conditions. The probe effectively avoids the influence of autofluorescence in biological systems and gave positive results when tested in both aqueous solution and living cells. Real-time images of cellular peroxynitrite were successfully acquired.     

Application of radiation technology in red blood cells immobilization on polycationic supports

The selective adsorption of blood group antibodies can be realized on immobilized red bloods cells on polycationic supports. Different supports containing dimethyl amino ethyl methacrylate (DMAEMA) has been obtained according to various proceedings : grafting on PVC, bulk polymerization, suspension polymerization.Physical characteristics and specific adsorption yields have been compared.This study has led to perfect specific and performant immunoadsorbents.     

Growth process of polymer aggregates formed by perfluorooctyltriethoxysilane. Time-resolved near-IR and two-dimensional near-IR correlation studies

The polymerization of 1H, 1H, 2H, 2H-perfluorooctyltriethoxysilane in ethanol, catalyzed by 0.5, 1.0 and 2.0 M HCl·H₂O, has been examined using time-resolved near-IR (NIR) and 2D NIR correlation techniques. The time-resolved NIR results have demonstrated that the growth of polymeric aggregates prior to the phase separation proceeds in a two-step process, which depends upon the HCl concentration. Furthermore, it has been found that the 2D NIR correlation data provide direct information on the sequence of the intricate reaction steps and interaction of participating components through hydrogen bonding in the growth process. Received: 25 July 2001 Accepted: 16 October 2001     

Ultraviolet radiation stimulates the release of arachidonic acid from mammalian cells in culture

Abstract— C3H 10T½ cells in culture were prelabelled with [³H]arachidonic acid and exposed to UVB radiation. Almost immediately after irradiation cells released labelled arachidonate metabolites into media in a dose dependent manner. This release was inhibited by removing calcium ions from the system and by the addition of dexamethasone and parabromophenacyl bromide to the system. This suggests that the UVB stimulated release of arachidonic acid from membrane phospholipids is, in part, mediated by a phospholipase A₂ enzyme system. Thin layer chromatographic examination of media extracts revealed a dose dependent UVB stimulation of prostaglandin production by cultured cells.     

The ethanol to acetaldehyde conversion activated by microwave radiation

The reaction of nonoxidative conversion of ethanol to acetaldehyde under conditions of thermal and microwave heating was studied on mixed ZnO—CuO—SiO₂ oxide catalysts modified with additives of Nb and Ta carbides. It is suggested that microwave heating promotes the appearance of “hot” zones at points where the metal-rich nanoparticles of metal carbides intensively absorbing the microwave energy contact active sites of the mixed oxide catalyst ZnO—CuO—SiO₂. Formation of these zones increases ethanol conversion and selectivity for acetaldehyde at moderate (<300°C) temperatures.     

The regeneration of polluted activated carbon by radiation techniques

In this paper, the regeneration of used activated carbon from monosodium glutamate factory was experimented using radiation and acid-alkali chemical cleaning method. Results showed that the activated carbon saturated with pollutants can be wash away easily by flushing with chemical solution prior irradiation. DSC was used to monitor the change of carbon adsorption     

Triazole-based osmium(ii) complexes displaying red/near-IR luminescence: antimicrobial activity and super-resolution imaging

Cellular uptake, luminescence imaging and antimicrobial activity against clinically relevant methicillin-resistant S. aureus (MRSA) bacteria are reported. The osmium(ii) complexes [Os(N^N)₃]²⁺ (N^N = 1-benzyl-4-(pyrid-2-yl)-1,2,3-triazole (1²⁺); 1-benzyl-4-(pyrimidin-2-yl)-1,2,3-triazole (2²⁺); 1-benzyl-4-(pyrazin-2-yl)-1,2,3-triazole (3²⁺)) were prepared and isolated as the chloride salts of their meridional and facial isomers. The complexes display prominent spin-forbidden ground state to triplet metal-to-ligand charge transfer (³MLCT) state absorption bands enabling excitation as low as 600 nm for fac/mer-3²⁺ and observation of emission in aqueous solution in the deep-red/near-IR regions of the spectrum. Cellular uptake studies within MRSA cells show antimicrobial activity for 1²⁺ and 2²⁺ with greater toxicity for the meridional isomers in each case and mer-1²⁺ showing the greatest potency (32 μg mL⁻¹ in defined minimal media). Super-resolution imaging experiments demonstrate binding of mer- and fac-1²⁺ to bacterial DNA with high Pearson''s colocalisation coefficients (up to 0.95 using DAPI). Phototoxicity studies showed the complexes exhibited a higher antimicrobial activity upon irradiation with light.

Cellular uptake, luminescence imaging and antimicrobial activity of facial and meridional isomers of Os(ii) triazole-based complexes against methicillin-resistant S. aureus, MRSA.     

Chemistry at boron: synthesis and properties of red to near-IR fluorescent dyes based on boron-substituted diisoindolomethene frameworks

A general method for the synthesis of difluorobora-diisoindolomethene dyes with phenyl, p-anisole, or ethyl-thiophene substituents has been developed. The nature of the substituents allows modulation of the fluorescence from 650 to 780 nm. Replacement of the fluoro ligands by ethynyl-aryl or ethyl residues is facile using Grignard reagents. Several X-ray molecular structures have been determined, allowing establishment of structure-fluorescence relationships. When the steric crowding around the boron center is severe, the aromatic substituents α to the diisoindolomethene nitrogens are twisted out of coplanarity, and hypsochromic shifts are observed in the absorption and emission spectra. This shift reached 91 nm with ethyl substituents compared to fluoro groups. When ethynyl linkers are used, the core remains flat, and a bathochromic shift is observed. All the fluorophores exhibit relatively high quantum yields for emitters in the 650-800 nm region. When perylene or pyrene residues are connected to the dyes, almost quantitative energy transfer from them to the dye core occurs, providing large virtual Stokes shifts spanning from 8000 to 13,000 cm(-1) depending on the nature of the dye. All the dyes are redox active, providing the Bodipy radical cation and anion in a reversible manner. Stepwise reduction or oxidation to the dication and dianion is feasible at higher potentials. We contend that the present work paves the way for the development of a new generation of stable, functionalized luminophores for bioanalytical applications.     

Nanoprobe implantation into mammalian cells by cationic transfection

Submicron-sized Au particles and Au/SiO(2) nanocomposites (superparticles) as large as 670 nm have been introduced into tsA201 cells with minimal cell trauma by cationic transfection systems. Successful implantations can be characterized by the expression of co-transfected DNA.     

Sequence-specific gene silencing in mammalian cells by alkylating pyrrole-imidazole polyamides

Gene silencing was examined by sequence-specific alkylation of DNA by N-methylpyrrole (Py)-N-methylimidazole (Im) hairpin polyamides. Polyamides ImImPyPygammaImImPyLDu86 (A) and ImImPyPygammaImPyPyLDu86 (B) selectively alkylated the coding regions of the renilla and firefly luciferases, respectively, according to the base pair recognition rule of Py-Im polyamides. Two different plasmids, encoding renilla luciferase and firefly luciferase, were used as vectors to examine the effect of alkylation on gene silencing. Transfection of the alkylated luciferase vectors-by polyamide A or B-into HeLa, 293, and NIH3T3 cells demonstrated that these sequence-specific DNA alkylations lead to selective silencing of gene expression. Next, the vectors were cotransfected into HeLa cells and the cells were treated with polyamide A or B. Selective reduction of luciferase activities was caused by both polyamides. On the basis of this sequence-specific alkylation and gene silencing activity, these alkylating Py-Im polyamides thus have potential as antitumor drugs to target specific gene expression in human cells.     

Transfer of foreign genes by electroporation and their expression in mammalian cells

Expression of foreign genes transferred into mammalian cells by electroporation has been studied. The pX1TK gene, pSV2Neo gene and pUCEJ oncogene have been introduced into MLTK- cells and NIH/3T3 cells, respectively. Stable transformation transient expression of TK gene by MLTK- cells as well as stable and malignant transformation of NIH/3T3 cells have been obtained. Transient expression frequency is about 80% and stable transformation frequency is about 10(-4). Integration of foreign genes into the cellular genome was verified with molecular hybridization. Tumor development was observed after inoculation of transformed cells into nude mice.     

Fast relaxing red and near-IR switchable azobenzenes with chalcogen and halogen substituents: periodic trends,tuneable thermal half-lives and chalcogen bonding

Molecular photoswitches operating in the red to near-IR region with controllable thermal relaxation rates are attractive components for photo-regulating biological processes. Herein, we report the synthesis of red-shifted azobenzenes functionalised with the heavier chalcogens and halogens that meet these requirements for biological application; namely fatigue-resistant photo-switching with red and near IR light and functional handles for further functionalisation for application. We report robust periodic trends for the chalcogen and halogen azobenzene series, and exploit intramolecular chalcogen bonding to tune and redshift the absorption maxima, supported by photo-physical measurements and solid-state structural analysis. Remarkably, the rate of the Z → E thermal isomerisation can be tuned over timescales spanning 10⁷ s by judicious choice of chalcogen and halogen substituents.

The synthesis of red-shifted azobenzenes functionalised with heavy chalcogen and halogen substituents is reported, enabling photo-switching with red and near IR light and Z → E thermal isomerization rates spanning seven orders of magnitude.     

Radioprotection by DMSO in nitrogen-saturated mammalian cells exposed to helium ion beams

The contribution of OH radical-mediated indirect action by particle beams under hypoxic irradiation condition was investigated by using a radical scavenger. V79 cells were irradiated with 150 MeV/nucleon helium ions at an LET of 2.2 keV/μm in the presence or absence of DMSO, and their colony survivals were determined. The contribution of indirect action to cell killing under hypoxic condition was estimated to be 52±9%. We conclude that OH radical-mediated indirect action still has a half in total contribution on cell killing under hypoxic condition.