RETENTION AND PHOTOTOXICITY OF TETRA(4-SULFONATOPHENYL)PORPHINE IN CULTIVATED HUMAN CELLS. THE EFFECT OF FRACTIONATION OF LIGHT

Human cervix carcinoma cells of the line NHIK 3025 were incubated for 18 h with tetra(4-sulfonatophenyl)porphine (TPPS4) and further incubated for 1-29 h in sensitizer free medium before exposure to light. After 1 h in sensitizer free medium only a 20% further loss of TPPS4 was observed within the next 28 h. During the time in sensitizer free medium, each TPPS4 molecule became more efficient in sensitizing single cells to photoinactivation. This enhanced photosensitizing efficiency of TPPS4 correlated well with the enhanced fluorescence yield of TPPS4. In some experiments the cells were exposed to a light dose inactivating 10% of the cells after incubation for 1 h in sensitizer free medium and a second graded light dose given 4-28 h later. Exposure of the cells to the first light dose led to loss of 60% of TPPS4 from the cells. Despite the significant loss of sensitizer from the cells the fluorescence yield of TPPS4 from each cell was found to increase (e.g. by 100% 4 h after light exposure). The enhanced fluorescence yield of cell bound TPPS4 was followed by a 1.6-2.5-fold increase in sensitivity of each cell to second light dose. Thus, a small light dose increased the photosensitivity of TPPS4-loaded NHIK 3025 cells for several hours after the first light exposure. The advantageous effect of light fractionation was reduced by a significantly enhanced loss of sensitizer induced by the first light exposure. The optimal time between the two fractions of light seems to be 30-90 min.     

MITOTIC INHIBITION BY PHENYLPORPHINES AND TETRASULFONATED ALUMINIUM PHTHALOCYANINE IN COMBINATION WITH LIGHT

This work relates to studies on modes of phototoxicity by tetrasulfonated aluminium phthalocyanine (AlPcS4), tetrahydroxy- and monosulfonated meso-tetraphenylporphines (3-THPP and TPPS1) on culture cells. Toxicity at moderate light exposures appears to be related to inhibition of microtubule function. Treatment of human cervix carcinoma cells of the line NHIK 3025 incubated for 18 h with the sensitizers and exposed to light inhibits multiplication for the first hours after light exposure, a significant fraction of the cells accumulating in mitosis. For the first hours after treatment, the mitotic cells were always mainly found in metaphase; generally seen as c-metaphases and three-group metaphases. During this time, anaphase and telophase cells were absent or greatly reduced in number. Indirect immunofluorescence staining of beta-tubulin showed that the spindle apparatus of mitotic cells was perturbed in all cases. The accumulation in mitosis was more extensive after treatment with AlPcS4 and light than after treatment with 3-THPP or TPPS1 and light. This may be related to the great difference in the lipophilic properties of these sensitizers; i.e. AlPcS4 being highly water soluble while TPPS1 and 3-THPP are lipophilic sensitizers. The lipophilicity of several sensitizers has been measured by two different methods, the partition between an aqueous and a lipophilic phase (Triton X-114) and the binding strength to a reverse phase column. The results show that the measured relative lipophilicity of the sensitizers may be influenced by the method of analysis.     

LIGHT INDUCED RELOCALIZATION OF SULFONATED meso-TETRAPHENYLPORPHINES IN NHIK 3025 CELLS AND EFFECTS OF DOSE FRACTIONATION

Human cervix carcinoma cells of the line NHIK 3025 were incubated for 18 h with sulfonated meso-tetraphenylporphines (TPPSn where n = 1, 2a, 2o or 4) followed by 1 h in sensitizer-free medium and then exposed to light. The fluorescing fraction of TPPS4, TPPS2o and TPPS2a has recently been shown to be located intracellularly in extracellular granules which are intracellularly localized in a similar pattern as acridine orange-stained granules, assumed to be endosomes and lysosomes (Berg, K., A. Western, J. Bommer and J. Moan. Photochem. Photobiol. 52, 481-487). Light exposure induced a relocalization of TPPS4 from its granular pattern to mainly the nuclear area while TPPS2o and TPPS2a relocalized mainly to cytoplasmic areas. After the light-induced relocalization TPPS4 became less efficient in sensitizing photoinactivation of cells as measured per fluorescing cellbound TPPS4 molecules while TPPS2a and TPPS2o became more efficient. These changes were independent of the extracellular concentration of TPPSn applied to the cells, except for cells incubated with 75 micrograms/mL TPPS4. These cells became more sensitive to light after a light exposure inactivating 20% of the cells. This increased photosensitivity seems to be related to a 2-2.5 fold increase in the amount of fluorescing cellbound TPPS4 induced by the first light exposure.     

On the dynamics of the TPPS4 aggregation in aqueous solutions: successive formation of H and J aggregates

The dynamics of aggregation of meso-tetrakis (p-sulfonatofenyl) porphyrin (TPPS4) in function of its concentration, pH and ionic strength was studied by optical absorption, fluorescence and resonance light scattering (RLS) techniques. In the region of pH, where TPPS4 exists in biprotonated form, the addition of NaCl induces the TPPS4 aggregation due to the formation of the "cloud" of counter ions around the TPPS4 molecule thus reducing electrostatic repulsion between the porphyrin molecules. The formation of this "cloud" shifts the pKa value to acid region (from 5.0 in the absence of salt to 4.5 at [NaCl] = 0.4 M), reduces the TPPS4 absorption in all spectral range and quantum yield and lifetime of fluorescence (from 0.27 to 0.17 and from 4.00+/-0.04 to 3.00+/-0.03 ns in the absence of salt and in the presence of NaCl, respectively). The aggregation process involves two successive stages: initially H aggregates are formed, which in time are transformed in J ones. The existence of these two stages was confirmed by the fluorescence and RLS data. The kinetics of the formation of J aggregates is characterized by the induction time t1 and the average growth time t2, which depend on both TPPS4 and salt concentrations. The induction period t1 appears as a result of initial formation of H aggregates and their successive transformation in J ones. At very high TPPS4 concentrations, the J aggregates are united in more complex structures such as hollow cylinders or helixes.     

Preferential cytotoxicity for multidrug-resistant K562 cells using the combination of a photosensitizer and a cyanine dye

The cyanine dye 1,1',3,3,3',3'-hexamethylindodicarbocyanine iodide (HIDC) protects K562 leukemia cells from photodynamic membrane damage caused by cis-di(4-sulfonatophenyl)diphenylporphine (TPPS2) and 420 nm light. This wavelength of light is chosen because it is absorbed by TPPS2, but not by HIDC. The photodynamic system studied may be useful as a model for antineoplastic therapy. A subline of K562 leukemia (K562/DOX), expressing the multidrug-resistance (MDR) phenotype, is found to accumulate smaller amounts of HIDC than the parent cell line and thus has less photoprotection. In the absence of added HIDC, the K562/DOX cell line is more resistant to photodynamic cytotoxicity than the K562 cell line. The resistance of the K562/DOX cell line is not due to a smaller accumulation of TPPS2 than the K562 cell line. However, when both cell lines are incubated with HIDC and TPPS2, and then exposed to light, the K562/DOX cell line becomes more sensitive to photodynamic cell damage than the K562 cell line. The combination of a photosensitizer with a cationic or lysomorphotropic photoprotector represents a novel strategy for the eradication of malignant cells expressing the MDR phenotype.     

Cellular uptake and photocytotoxicity of glycoconjugated porphyrins in HeLa cells

Thirty-two glycoconjugated porphyrins were synthesized by a modification of Lindsey method in the presence of Zn(OAc)(2).2H(2)O as a template. The Zn(2+) ion template strategy improved the yield about three-fold in the case of meta-substituted tetraphenylporphyrins. In addition, free-base porphyrins were obtained almost quantitatively by demetalation with 4 M HCl. Sixteen deacetylated glycoconjugated porphyrins were tested as candidate photodynamic therapy (PDT) drugs using HeLa cells. Most of the deacetylated glycoconjugated porphyrins showed higher cellular uptake than tetraphenylporphyrin tetrasulfonic acid (TPPS), and 5,10,15,20-tetrakis[4-(beta-D-arabinopyranosyloxy)phenyl]porphyrin (p-5d) in particular showed 18.5-fold higher uptake than TPPS. The photocytotoxicity of 5,10,15,20-tetrakis[4-(beta-D-glucopyranosyloxy)phenyl]porphyrin (p-5a), p-5d and TPPS was examined with HeLa cells, using a light dose of 16 J/cm(2). These photosensitizers had no cytotoxicity in the dark, but their photocytotoxicity increased in the order of TPPS < p-5a < p-5d. These results suggest p-5d is a good candidate for a PDT drug.     

Microscopic observation of living cells during their exposure to modulated electromagnetic fields

Studying cell behaviour under irradiation with radiofrequency electromagnetic fields (RF-EMF) is often impeded by the difficulty to monitor cell characteristics during irradiation. Here we report the design and the application of a complete device for continuous microscopic observation of cells exposed to modulated EMF similar to mobile phones signals. The system allows the follow up of cell progression into mitosis under controlled temperature and CO(2) environment. Protocols are proposed in which the same cells are the controls before and after the EMF exposure and we demonstrate the interest of the "before exposure" controls. The exposure system was validated by cell endocytosis measurements. While the endocytosis rate was increased, no alteration of mitosis progression and mitosis duration was observed in cells exposed to 900 MHz modulated EMF for 1 h, at 30 degrees C and at a Specific Absorption Rate of 2.2 W/kg.     

ULTRASTRUCTURAL STUDIES ON THE MECHANISM OF THE PHOTODYNAMIC THERAPY OF TUMORS

Abstract Balb/c mice bearing a transplanted MS-2 fibrosarcoma were injected with 2.5 mg kg ¹ of either tetra(4-sulfonatophenyl/porphine (TPPS) in phosphate-buffered saline or 0.5 mg kg⁻¹ of Zn²⁺-phthalocyanine (Zn-Pc) incorporated into unilamellar liposomes of dipalmitoyl-phosphatidylcholine. Chromatographic studies showed that TPPS is mainly transported in the serum by globulins and albumin, while Zn-Pc is specifically bound by lipoproteins. Exposure of the injected mice to red light (300 J cm⁻²) caused extensive tumor necrosis. The ultrastructural analysis of tumor specimens taken from mice at 15 h after PDT showed that TPPS photoinduces a preferential necrosis of the neoplastic cells, while Zn-Pc causes severe photodamage to both the vascular system and the neoplastic cells. The different modes of tumor photosensitization by TPPS and Zn-Pc are discussed on the basis of the transport mechanism of the two dyes.     

The Mode of Cell Death Induced by Photodynamic Treatment Depends on Cell Density

Madison Darby canine kidney II (MDCK II) cells were seeded out at two different densities and incubated with 125 micrograms/mL of the photosensitizer meso-tetra(4-sulfonatophenyl)porphine (TPPS4) for 18 h, washed and irradiated with blue light. Four hours later the cells were studied by fluorescence microscopy. Apoptotic cells were detected by virtue of the distinct condensation and fragmentation of chromatin, and necrotic cells were detected by uptake of propidium iodide. In addition apoptosis was measured by the TdT assay. The fraction of apoptotic cells and the fraction of necrotic cells were determined for both cell densities at various levels of survival. With < 55% total cell death the apoptotic fraction was significantly higher for cells in confluent monolayers than for cells growing in microcolonies at equitoxic doses. Confluent cells were 2.9 times more sensitive than cells in microcolonies partly due to a 1.5 times higher uptake of TPPS4 in monolayer cells. The difference in mode of cell death for the different cell densities was not related to any observable difference in subcellular localization pattern of TPPS4 at equitoxic doses of photodynamic treatment.     

Porphyrin J-aggregates stabilized by ferric myoglobin in neutral aqueous solution

J-aggregates of a diacid form (H4TPPS2-) of 5,10,15,20-tetrakis(4-sulfonatophenyl)porphyrin (H2TPPS4-) were stabilized by binding with ferric myoglobin (metMb) in aqueous solution at neutral pH. The J-aggregates gradually dissociated to monomeric H2TPPS(4-). The average half-lifetime (t1/2) of the J-aggregates in the presence of sufficient amounts of metMb was ca. 3 h in phosphate buffer at pH 7.0 and 25 degrees C. The stabilization of the J-aggregate by metMb is ascribed to encapsulation and fixation of an edge-to-edge structure of the J-aggregate by the relatively rigid protein molecules. The secondary structure of metMb was altered in some extent in the presence of an excess amount of the J-aggregates while no effect on denaturation of metMb was observed with the H2TPPS(4-) monomer or polyacrylate. The hydrophobic nature of the J-aggregate seems to play an important role for denaturation of metMb. The ability of denatured metMb to bind the azide anion was higher than that of natural metMb. The denaturation of metMb by the J-aggregates seems to induce surfacing of hemin leading to an entropy gain in coordination of the N3(-) anion to the iron(III) center.     

UVA‐Induced DNA Damage and Apoptosis in Red Blood Cells of the African Catfish Clarias gariepinus

Ultraviolet‐A light (UVA)‐induced DNA damage and repair in red blood cells to investigate the sensitivity of African catfish to UVA exposure is reported. Fishes were irradiated with various doses of UVA light (15, 30, and 60 min day⁻¹ for 3 days). Morphological and nuclear abnormalities in red blood cells were observed in the fish exposed to UVA compared with controls. Morphological alterations such as acanthocytes, crenated cells, swollen cells, teardrop‐like cells, hemolyzed cells, and sickle cells were observed. Those alterations were increased after 24 h exposure to UVA light and decreased at 14 days after exposure. The percentage of apoptosis was higher in red blood cells exposed to higher doses of UVA light. No micronuclei were detected, but small nuclear abnormalities such as deformed and eccentric nuclei were observed in some groups. We concluded that exposure to UVA light induced DNA damage, apoptosis, and morphological alterations in red blood cells in catfish; however, catfish were found to be less sensitive to UVA light than wild‐type medaka.     

Fluorescence formation during photodynamic therapy in the nucleus of cells incubated with cationic and anionic water-soluble photosensitizers.

The variations of fluorescence during light exposure of the cationic sensitizers methylene blue (MB) and meso-tetra(4N-methylpyridyl)porphyrin (T4MPyP) as well as the anionic meso-tetra(4-sulphonatophenyl)porphyrin (TPPS4) were measured at different intracellular sites using video-intensified microscopy in combination with microspectrofluorometry. Before light exposure the sensitizers were localized in distinct parts of the cytoplasm, especially in fluorescent organelles. During irradiation a drastic fluorescence formation and increase in the cytoplasm and nucleus, which was most pronounced in the nucleoli, could be observed for the cationic sensitizers as well as TPPS4. In the case of MB the increase in fluorescence was concomitant with a spectral shift in the emission spectra. For TPPS4 and T4MPyP the formation of a second species with a Soret band shifted towards longer wavelengths was observed and correlated with the fluorescence increase in the nucleoli. Cell deformations also took place.     

Photochemical internalization enhances the cytotoxic effect of the protein toxin gelonin and transgene expression in sarcoma cells

Further advantages in the treatment of soft-tissue sarcomas will only be achieved by tailoring the adjuvant therapy after surgery. The photochemically directed release of macro-molecules from endosomes and lysosomes into the cytosol is a novel technology, named photochemical internalization (PCI), that has been evaluated for treatment of sarcoma cells in vitro. Two human synovial sarcoma cell lines (SW 982 and CME-1) were treated with the photosensitizer meso-tetraphenylporphine with two sulfonate groups on adjacent phenyl rings (TPPS2a) and a plasmid encoding enhanced green fluorescent protein (EGFP) complexed to poly-L-lysine to investigate the influence of PCI on gene transfer and with 5 micrograms/mL gelonin to investigate PCI of a Type-I ribosome-inactivating protein toxin. In addition, both cell lines were transduced with an Adenovirus serotype 5 encoding the Escherichia coli lacZ gene (AdHCMV-lacZ, expressing beta-galactosidase) and treated with TPPS2a and light to evaluate the effect of PCI on the transduction rate. Photochemically induced transfection with the reporter gene EGFP in CME-1 cells increased from 0% of cells at no light to 40% of the cells after 60 s of light exposure. In contrast, the SW 982 cells showed no enhanced expression of the gene. The fraction of virally transduced cells was about doubled in both cell lines by means of PCI, although the transduction was more efficient in the CME-1 cells. Both cell lines became up to four-fold more sensitive to light when combining photochemical treatment with gelonin incubation. Our experiments showed that PCI induced the endocytic escape of therapeutic substances in cells derived from human soft-tissue sarcomas.     

Plasma membrane associated location of sulfonated meso-tetraphenylporphyrins of different hydrophilicity probed by total internal reflection fluorescence spectroscopy

Sulfonated meso-tetraphenylporphyrins of different hydrophilicity were microspectrofluorimetrically examined in endothelial cells using total internal reflection (TIR) illumination or epi-illumination. Since the penetration depth of the evanescent field during TIR illumination is limited to a few hundred nanometers, photosensitizers were almost selectively examined in close vicinity to the plasma membrane. Pronounced fluorescence signals during TIR illumination were observed for the hydrophilic compounds meso-tetraphenylporphyrin tetrasulfonate (TPPS4) and meso-tetraphenylporphyrin trisulfonate (TPPS3), whereas the more lipophilic compounds meso-tetraphenylporphyrin disulfonate (TPPS2a) and meso-tetraphenylporphyrin monosulfonate (TPPS1) could only be detected under epi-illumination. Irradiation of TPPS1 and TPPS2a in the Soret band led to an increase in fluorescence intensity and formation of a photoproduct with an emission maximum around 610 nm, which was limited to intracellular compartments. In contrast, fluorescence spectra of TPPS3 and TPPS4 obtained by TIR and epi-illumination remained almost unchanged after irradiation in the Soret band. Extralysosomal location of TPPS3 and TPPS4 in close proximity to the plasma membrane was deduced from experiments with the lysosomal markers acridine orange (AO) or lysotracker yellow (LY), which were not detectable under TIR illumination. In conclusion, these results provide for the first time direct evidence for a plasma membrane-associated fraction of the hydrophilic compounds TPPS3 and TPPS4 in living cells.     

Amphiphilic cyclodextrin carriers embedding porphyrins: charge and size modulation of colloidal stability in heterotopic aggregates

The interaction between the anionic 5,10,15,20-tetrakis(4-sulfonatophenyl)-21H,23H-porphyrin (TPPS) and cationic vesicles formed by heptakis(2-omega-amino-O-oligo(ethylene oxide)-6-hexylthio)-beta-cyclodextrin (SC6CDNH2) has been investigated in detail through a combination of elastic light scattering (ELS), quasi-elastic light scattering (QELS), zeta potential measurements, and time-resolved fluorescence anisotropy. ELS experiments provided the first structural characterization of these cationic vesicles both in the absence and in the presence of TPPS porphyrin, modeling the system as a spherical particle described by a single thin shell form factor. The structure of mixed hetero-aggregates is modulated by charge and size of the two components as function of different porphyrin/cyclodextrin (CD) molar ratios. At the limiting molar ratio studied, the absolute value of zeta potential (/zeta/ = 12.5 mV) seems to be a reference value for the formation of stable colloidal CD vesicular aggregates at thermodynamic equilibrium. New insights on the structure of these heterotopic colloids have been obtained by analysis of rotational correlation times at different molar ratios exploiting time-resolved fluorescence anisotropy experiments. At high porphyrin loads, the anisotropy decays behave as monoexponentials and the rotational correlation times (1-2 ns) together with the r(0) values close to zero suggest the presence of small amounts of TPPS embedded in a hydrophobic environment either in monomeric or in aggregated form. At the lower porphyrin/CD molar ratios, the anisotropy decays exhibit a double-exponential behavior showing a predominant component with a slow rotational correlation time (20-25 ns) and limiting anisotropy values of approximately 0.15. This component has been assigned to molecules that are more stabilized onto the CD vesicles, that is, porphyrins embedded into the oligo-ethylene "wall" of the CD vesicles. Scanning near-field optical microscopy of the samples evaporated on glass surfaces gave further insights on the morphology and optical properties of these systems, confirming the embedding of TPPS on the vesicles and evidencing the role of the solvent.     

Intracellular localization of sulfonated meso-tetraphenylporphines in a human carcinoma cell line

The intracellular localization of meso-tetraphenylporphines sulfonated to different degrees (TPPSn), in a human cervix carcinoma cell line (NHIK 3025), was studied by fluorescence microscopy and fluorescence spectroscopy. After an 18 h incubation, TPPS4, TPPS2a and TPPS2o were localized in extranuclear granules. Studies of cells stained with both TPPS4, and acridine orange, which is known to fluoresce red in lysosomes, indicated that these granules were lysosomes. In addition, a fraction of the cellbound TPPS4, TPPS2a and TPPS2o seems to be associated with the plasma membrane. Fluorescence quenching studies of cells doublestained with acridine orange and TPPS4 indicated that TPPS4 is also localized in the nucleus and in the extralysosomal cytoplasm. The intracellular location of TPPS1 differed from that of the other TPPSns studied: In 6 out of 9 experiments fluorescing extranuclear granules were found. A diffuse fluorescence extending from the perinuclear area was also observed.     

Photodamage in a Mitochondrial Membrane Model Modulated by the Topology of Cationic and Anionic Meso‐Tetrakis Porphyrin Free Bases

The photodynamic effects of the cationic TMPyP (meso‐tetrakis [N‐methyl‐4‐pyridyl]porphyrin) and the anionic TPPS4 (meso‐tetrakis[4‐sulfonatophenyl]porphyrin) against PC/CL phosphatidylcholine/cardiolipin (85/15%) membranes were probed to address the influence of phorphyrin binding on lipid damage. Electronic absorption spectroscopy and zeta potential measurements demonstrated that only TMPyP binds to PC/CL large unilamellar vesicles (LUVs). The photodamage after irradiation with visible light was analyzed by dosages of lipid peroxides (LOOH) and thiobarbituric reactive substance and by a contrast phase image of the giant unilamellar vesicles (GUVs). Damage to LUVs and GUVs promoted by TMPyP and TPPS4 were qualitatively and quantitatively different. The cationic porphyrin promoted damage more extensive and faster. The increase in LOOH was higher in the presence of D₂O, and was impaired by sodium azide and sorbic acid. The effect of D₂O was higher for TPPS4 as the photosensitizer. The use of DCFH demonstrated that liposomes prevent the photobleaching of TMPyP. The results are consistent with a more stable TMPyP that generates long‐lived singlet oxygen preferentially partitioned in the bilayer. Conversely, TPPS4 generates singlet oxygen in the bulk whose lifetime is increased in D₂O. Therefore, the affinity of the porphyrin to the membrane modulates the rate, type and degree of lipid damage.     

meso-四(4-磺酸基苯基)卟啉-铬黑T与铁(Ⅲ)混合显色反应的研究

本文研究了以铬黑T作TPPS_4和Fe(Ⅲ)的混合配位体,并在弱酸性条件下运用了铬黑T,首次突破了Fe(Ⅲ))与TPPS_4的成络反应条件:在pH4.0的HAc-NaAc缓冲溶液中,沸水浴加热15min,以1:1:1的组成形成TPPS_4-铬黑T-Fe(Ⅲ)混配络合物,λ_(max)=392um,ε＇=2.07×10~5L·mol~(-1)·cm~(-1),稳定常数为8.7×10~7,Fe(Ⅲ)含量在0～4.5μg/25ml范围内成线性关系.将此方法用于纯铜、茶叶、烟草样品中的痕量铁的测定,获得了较满意的结果.     

Purpurin-18 in combination with light leads to apoptosis or necrosis in HL60 leukemia cells

Photodynamic therapy (PDT), a cancer treatment using a photosensitizer and visible light, has been shown to induce apoptosis or necrosis. We report here that Purpurin-18 (Pu18) in combination with light induces rapid apoptotic cell death in the human leukemia cell line (HL60) at low doses and necrosis at higher concentrations. Cells treated with Pu18 and light under apoptotic conditions exhibited DNA laddering and an increase in both cellular content of subdiploid DNA and externalization of phosphatidylserine (PS), indicating DNA fragmentation and loss of membrane phospholipid asymmetry. In the absence of light activation, Pu18 at nanomolar concentrations had no detectable cytotoxic effect. Caspase-3 activity was increased even after 1 h from treatment with low doses of Pu18 and light. The PS exposure and nuclear features of apoptosis were prevented by treatment of cells before illumination with caspase inhibitors benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (Z-VAD-FMK) and benzyloxycarbonyl-Asp-Glu-Val-Asp-fluoromethylketone (Z-DEVD-FMK). Conversely, the caspase-1 inhibitor, acetyl-Tyr-Val-Ala-Asp-aldehyde (Ac-YVAD-CHO) failed to suppress the apoptosis. No protective effect of the three caspase inhibitors was observed when the cells were exposed to necrotic concentrations of Pu18 and light. Our results show that caspase-3, but not caspase-1, is involved in the signaling of apoptotic events in PDT with Pu18-induced apoptosis of HL60 cells. Moreover, both the time course of PS exposure and the effect of caspase inhibitors on it indicate that it is regulated in the same manner as DNA fragmentation.     

Anion binding to a ferric porphyrin complexed with per-O-methylated beta-cyclodextrin in aqueous solution

5,10,15,20-Tetrakis(4-sulfonatophenyl)porphinato iron(III) (Fe(III)TPPS) forms a very stable 1:2 complex with heptakis(2,3,6-tri-O-methyl)-beta-cyclodextrin (TMe-beta-CD), whose iron(III) center is located at a hydrophobic cleft formed by two face-to-face TMe-beta-CD molecules. Various inorganic anions (X(-)) such as F(-), Cl(-), Br(-), I(-), N(3)(-), and SCN(-) coordinate to Fe(III)TPPS(TMe-beta-CD)(2) to form five-coordinate high-spin Fe(III)TPPS(X)(TMe-beta-CD)(2), while no coordination occurs with ClO(4)(-), H(2)PO(4)(-), NO(3)(-), and HSO(4)(-). Except for F(-), none of the anions investigated coordinate to Fe(III)TPPS in the absence of TMe-beta-CD due to extensive hydration to the anions as well as to Fe(III)TPPS. The present system shows a high selectivity toward the N(3)(-) anion. The thermodynamics suggests that Lewis basicity, hydrophilicity, and shape of an X(-) anion are the main factors to determine the stability of the Fe(III)TPPS(X)(TMe-beta-CD)(2) complex.