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T Hasan

Publications and source records attributed to T Hasan.

At least 109 records · Page 6Linked to original sources
PubMed1991

Mechanism of photosensitization by microsphere-bound chlorin e6 in human bladder carcinoma cells.

Photodynamic therapy is an experimental method of cancer treatment in which a photosensitizer is administered and subsequently the tumor is irradiated with light. Due to problems of prolonged skin phototoxicity with hematoporphyrin derivative, new photosensitizers and methods of localization are being sought. The goal of this study was to compare the photosensitizer chlorin e6 (Ce6) free and bound to 1-micron-diameter microspheres (MS) for phototoxicity, uptake and efflux characteristics, phagocytosis rates in malignant and benign cells, and effects of NaN3, D2O, and buthionine sulfoximine on phototoxic efficacy. Incubation of MGH-U1 human bladder carcinoma cells with Ce6-MS (0.43 microM Ce6-equivalent; 18 h) and subsequent irradiation using an argon laser-pumped dye laser at a radiant exposure of 20 J/cm2 caused 100% cell death 24 h after irradiation. In contrast, MGH-U1 cells incubated with free Ce6 (0.43 microM; 18 h) remained 100% viable 24 h after irradiation at a radiant exposure of up to 50 J/cm2. The presence of D2O during irradiation increased the phototoxicity to MGH-U1 cells, whereas the presence of NaN3 decreased it; these data support an important role for 1O2. Irradiation of MGH-U1 cells in the presence of the glutathione depleter buthionine sulfoximine also increased the phototoxicity, demonstrating a role for intracellular glutathione and possibly free radical intermediates. The cellular uptake of Ce6 was approximately 50 times lower than that of Ce6-MS at equivalent incubation concentrations. Efflux experiments showed that the phototoxicity of Ce6-MS was reduced by 40% for a 5-h washout time as compared to no washout time. In contrast, for free Ce6, the decrease was 95.3% under identical conditions. Because the total intracellular concentration of Ce6-MS after an efflux time of 5 h was only slightly changed, the decreased phototoxicity is attributed to an altered intracellular localization. Confocal laser scanning fluorescence microscopy data appear consistent with this hypothesis although they are not conclusive. The observed patterns were different at 0 and 5 h. Comparison of the phagocytosis rates of Ce6-MS by carcinoma and benign cells showed that on average 20 MS/cell were phagocytosed by MGH-U1 compared with 2.5 and 8.3 in the benign human fibroblasts and keratinocytes, respectively. After incubation with Ce6-MS (0.1 microM; 18 h) and irradiation at 10 J/cm2 the surviving fraction of MGH-U1 cells was 76.3 +/- 0.95% (mean +/- SE) and 40 +/- 3.49% for fibroblasts. In contrast, for keratinocytes the surviving fraction was 93.5 +/- 0.83%.(ABSTRACT TRUNCATED AT 400 WORDS)

Buthionine Sulfoximine↗
PubMed1991

Photosensitized destruction of human bladder carcinoma cells treated with chlorin e6-conjugated microspheres.

A photosensitizer conjugate, chlorin e6 (Ce6) covalently bound to 1-micron-diameter polystyrene microspheres, has been investigated in the photodynamic destruction of MGH-U1 human bladder carcinoma cells in vitro. The microspheres were taken up avidly by the carcinoma cells; confocal laser scanning fluorescence microscopy showed them to be localized in the cytoplasm, apparently within lysosomes, visualized by labeling with acridine orange. In contrast, fluorescence of unconjugated Ce6 was present within most cellular membranes. Use of Ce6-microsphere conjugates led to a 20-fold-higher mean intracellular concentration, compared with unconjugated Ce6. Cells incubated in the presence of Ce6-microsphere conjugates (0.43 microM equivalent) and subsequently irradiated at 659 nm with a dye laser pumped by an argon-ion laser showed dose-dependent phototoxicity, leading to total inhibition of colony formation at a radiant exposure of 5J/cm2; in contrast, cells incubated with either unconjugated Ce6 (0.43 microM) or unconjugated microspheres before laser irradiation were unaffected. Cells pretreated with Ce6-microsphere conjugates and irradiated in the presence of 90% 2H2O showed significantly increased phototoxicity, an effect consistent with an important role for excited-state singlet oxygen in the mechanism of injury. In solution, however, photosensitized generation of singlet oxygen with Ce6-microsphere conjugates was 9 times less efficient than with unconjugated Ce6. The markedly greater phototoxicity of Ce6-microsphere conjugates compared to unconjugated Ce6 was therefore a consequence of the high intracellular Ce6 concentration attained by phagocytosis of the conjugates and their particular sites of intracellular localization. Thus, these conjugates are an efficient system for the delivery of photosensitizing drugs to carcinoma cells.

Cell Division↗
PubMed1991

Free and conjugated chlorin E6 in the photodynamic therapy of human bladder carcinoma cells.

Photodynamic therapy is an experimental modality for treatment of superficial bladder cancer, and consists of the administration of a photosensitizer and subsequent tumor-irradiation with light. Presently hematoporphyrin derivative (HPD) is the only photosensitizer in experimental clinical use in the United States. Because of the high nonspecific phototoxicity of HPD, new methods of photosensitization have been sought. In this study we compared chlorin e6, free and conjugated to 1-micron.-diameter latex microspheres. Phototoxicity was evaluated on MGH-U1 cells derived from a human bladder carcinoma. MGH-U1 cells were preincubated for 18 hours either with free Ce6 (0.43 microM) or Ce6-microspheres (0.43 microM equivalent in Ce6) and irradiated with an argon-laser-pumped dye laser emitting at 659 nm., over the radiant-exposure range of 5-50 J/cm. At 24 hours after light exposure the cells were observed microscopically for morphological alteration and evaluated for cell death by trypan blue exclusion. Cultures incubated with Ce6-microspheres and subsequently irradiated showed morphologic evidence of cell damage, apparent after irradiation with five J/cm. and a light dose dependent decrease in cell survival. In contrast, cells incubated with free Ce6 at the same concentration of 0.43 microM and subsequently irradiated demonstrated neither detectable morphologic alteration nor change in cell survival. Only cells preincubated with free Ce6 at higher concentration showed morphologic changes. Thus, Ce6-microsphere conjugate was much more efficient at inducing photodynamic destruction of bladder carcinoma cells than was free Ce6.

Carcinoma, Transitional Cell↗
PubMed1991

Protective effects of cadmium chloride against UVB injury in mouse skin and in cultured human cells: a possible role of cadmium-induced metallothionein.

Metallothionein (MT) is a cysteine-rich protein with antioxidant and metal-chelating activities that is readily inducible by exposure to a variety of stimuli including heavy metals such as cadmium (Cd2+). We have investigated the protective effects of Cd2+ treatment on sunburn cell (SBC) induction in mouse skin in vivo and human cell survival in vitro after UVB exposure. The number of SBC in mouse ear skin was significantly reduced in Cd(2+)-treated mice (10 mumol CdCl2/kg) with each UVB dose (25, 50 and 100 mJ/cm2) compared with controls. Delay of UVB exposure after Cd2+ administration (24 h) and higher doses of CdCl2 (10-30 mumol/kg) were more efficient in reducing SBC formation. Human bladder cancer cells (MGH-U1) made tolerant to Cd2+ by repeated low dose exposure, or cells acutely exposed to high Cd2+ concentration showed increased tolerance (cell survival) to UVB injury. Electrophoretic-autoradiographic analysis of [35S]-cysteine-labeled protein synthesized by cultured cells after Cd2+ treatment revealed increased MT-like protein content. These results suggest that MT is inducible by exposure to Cd2+ in our system and may be a photoprotective agent against UVB-induced oxidative damage in mammalian skin.

Animals↗
PubMed1990

Rhodamine 123 phototoxicity in laser-irradiated MGH-U1 human carcinoma cells studied in vitro by electron microscopy and confocal laser scanning microscopy.

Rhodamine 123 (R123) is a permeant, cationic, fluorescent dye that localizes preferentially within mitochondria of living carcinoma cells. MGH-U1 human bladder carcinoma cells incubated in vitro with 10 microM R123 for 30 min and then irradiated at 514.5 nm with an argon ion laser underwent selective, phototoxic injury to mitochondria. Ultrastructurally, treatment with R123 plus irradiation with 10 J/cm2 caused selective, progressive mitochondrial alterations consisting of disruption of cristae, vacuolization, swelling, increasing numbers of ring-shaped and angulated mitochondria at 4 to 8 h after irradiation, and obliteration of many mitochondria at 24 to 48 h. Confocal laser scanning microscopy after treatment with R123 plus irradiation with 10 to 30 J/cm2 demonstrated altered uptake and localization of subsequently administered R123, accompanied by striking mitochondrial fragmentation. Irradiation caused a dose-dependent depletion of extractable R123, due to a photosensitized efflux that began immediately and progressed by 4 h after irradiation with 10 to 30 J/cm2; further uptake after reincubation in the presence of R123 was also quantitatively impaired in cells previously irradiated with 30 J/cm2.

Carcinoma, Transitional Cell↗
PubMed1990

Mechanistic investigation of doxycycline photosensitization by picosecond-pulsed and continuous wave laser irradiation of cells in culture.

In order to elucidate the photophysical mechanisms of cellular phototoxicity sensitized by doxycycline, MGH-U1 human bladder carcinoma cells in vitro were treated with 20.7 microM doxycycline and irradiated with either a pulsed (lambda = 355 nm, pulse duration = 24 ps) or a continuous wave (lambda = 351 nm) laser. Cumulative radiant exposure and irradiance were systematically varied in experiments with both lasers. Phototoxicity was assessed by epifluorescence microscopy of unfixed cells using rhodamine 123 labeling of mitochondria. With the continuous wave source, the cumulative radiant exposure required for induction of phototoxic injury was independent of irradiance. With the 24-ps-pulsed source, a significantly lower cumulative radiant exposure was required to induce the phototoxicity when the peak irradiance was 5.8 x 10(7) or 1.3 x 10(8) watts cm-2 compared with when peak irradiance was either lower (6.0 x 10(6) watts cm-2) or higher (7.6 x 10(8) watts cm-2). The measured fluorescence lifetimes of doxycycline in buffered saline solution were longer than the laser pulse duration of 24 ps. The increased efficiency of photosensitization at the optimal peak irradiance in the ps domain appears to result from sequential multiphoton absorption involving higher excited states of the singlet manifold. At the highest irradiance studied, on the other hand, reduced efficiency of photosensitization is attributed to increased photodegradation of doxycycline from higher excited states by processes such as photoionization. A model consistent with these observations is presented along with calculations, based on simple rate equations, that fit the essentials of the proposed model.

Cell Line↗
PubMed1990

Investigations of a manganese-containing mimic of superoxide dismutase in riboflavin phototoxicity in human cells in vitro.

The activity and specificity of a manganese-containing low molecular weight mimic of superoxide dismutase (manganese desferioxamine (Mndf)) were investigated in riboflavin (Rf) photosensitization in solution and cell culture. In addition to the very high superoxide dismutase-like activity of Mndf at micromolar concentrations, photochemical studies in solution indicated that it could quench excited singlet and triplet states at millimolar concentrations. Human erythrocytes, human lymphocytes and a human bladder carcinoma cell line were used to evaluate the potential of Mndf for in vivo use. The efficacy and toxicity of Mndf in the protection against Rf photoxicity varied between the different cell types.

Deferoxamine↗
PubMed1989

Rhodamine dyes as potential agents for photochemotherapy of cancer in human bladder carcinoma cells.

The phototoxicity in vitro of rhodamine 123 and tetrabromo rhodamine 123 (TBR) was compared, in order to assess their photochemotherapeutic potential. Exposure to 514.5-nm radiation from an argon ion laser caused phototoxicity in MGH-U1 bladder carcinoma cells previously treated with either dye at 10 microM for 30 min. As assessed by colony formation and cellular morphology, TBR was markedly more phototoxic than rhodamine 123, reflecting increased intersystem crossing of TBR to the triplet manifold via spin-orbital coupling induced by the heavy bromine atoms. Photoreactions of TBR very efficiently generated singlet oxygen (1O2) in solution; furthermore, irradiation of TBR-treated cells was significantly more toxic when performed in the presence of deuterium oxide, an enhancer of damage caused by 1O2. Retention of fluorescence in TBR-treated cells was enhanced by irradiation, indicating that a stable photoproduct may be formed in reaction with cellular components.

Antineoplastic Agents↗
PubMed1989

Dynamic aspects of rhodamine dye photosensitization in vitro with an argon-ion laser.

Phototoxicity in cultured human bladder carcinoma cells treated by 514.5-nm argon-ion laser irradiation plus rhodamine-123 (R123) or tetrabromo-R123 (TBR) was assessed counting total cell number and percent viability at 1, 24, 48, 72, and 96 h after irradiation. TBR was markedly more efficient than R123 at causing both reversible and persistent phototoxic cytostasis. Furthermore, TBR photosensitization caused net cytocidal effects at 0.5 J/cm2, which were not seen with R123 photosensitization at up to 40 J/cm2. The reduced phototoxic efficiency of R123 as compared to TBR appears, in part, to reflect the presence of a fraction of cells refractory to R123 photosensitization.

Antineoplastic Agents↗
PubMed1988

Ultrastructure and dynamics of selective mitochondrial injury in carcinoma cells after doxycycline photosensitization in vitro.

Mechanisms and intracellular sites of photosensitized damage were investigated in cultured MGH-U1 cells treated with doxycycline (DOTC). Cells were examined by phase-contrast, fluorescence, and electron microscopy at various times (15 minutes to 24 hours) after ultraviolet irradiation (320-400 nm). DOTC localized selectively within the mitochondria, as shown by colocalized fluorescence with rhodamine 123 (R123). Photosensitization at 1 J/sq cm caused striking swelling of the mitochondrial matrix and disruption of the cristae, accompanied by loss of the ability of mitochondria to concentrate R123. These changes progressed during the first hour after irradiation, and then were followed by partial recovery of mitochondrial ultrastructure and function. At no time were other organelles seen to be affected. It appears that this selective, photosensitized alteration was a consequence of localized and partially reversible damage to the mitochondrial inner membrane. In contrast, exposure to 2-6 J/sq cm caused irreversible injury and necrosis.

Doxycycline↗
PubMed1986

Phototoxicity of the tetracyclines: photosensitized emission of singlet delta dioxygen.

The spectroscopic observation of 1268-nm emission of singlet oxygen photosensitized by tetracyclines in oxygenated solutions at room temperature is reported. In the series demeclocycline, tetracycline, and minocycline, the efficiency of singlet oxygen generation is found to parallel the clinical observation of relative frequency of phototoxicity of these antibiotics, suggesting singlet oxygen generation as the origin of their phototoxicity.

Demeclocycline↗
PubMed1986

Antibody-targeted photolysis: selective photodestruction of human T-cell leukemia cells using monoclonal antibody-chlorin e6 conjugates.

Selective in vitro photodestruction of HPB-ALL human T-cell leukemia cells was accomplished using the photosensitizer chlorin e6 coupled through dextran molecules to an anti-T-cell monoclonal antibody (mAb), anti-Leu-1. Conjugates with mAb/chlorin molar ratios as high as 1:36 retained mAb binding activity, and the absorption spectrum and quantum efficiency for singlet oxygen production of bound chlorin (0.7 +/- 0.2) were unchanged from that of the free photosensitizer. Phototoxicity, as measured by a clonogenic assay and by uptake of ethidium bromide, was dependent on the doses of both mAb-chlorin and 630- to 670-nm light, was enhanced by 2H2O, and was observed only in target populations that bound the mAb. Similarly, free chlorin e6 in solution had no photodynamic effect in amounts 100 times more than that carried by the mAb. For this antibody-targeted system, approximately 10(10) molecules of singlet oxygen were necessary to kill a cell.

Antibodies, Monoclonal↗
PubMed1986

Mitochondrial phototoxicity sensitized by doxycycline in cultured human carcinoma cells.

Cultured MGH-U1 (human urinary bladder carcinoma) cells were treated with doxycycline (DOTC) and long-wave UV radiation (UVA). At UVA doses of 1 J/cm2 and above, the cells showed mitochondrial damage, reflected by altered localization of the fluorescent probe rhodamine-123, and striking vacuolization of the cytoplasm. Cell membrane integrity, as monitored by exclusion of ethidium bromide, was maintained for several hours after mitochondrial damage was evident. These changes were potentiated by irradiation in the presence of deuterium oxide, and diminished by irradiation in the presence of sodium azide. Addition of catalase, superoxide dismutase, or mannitol did not alter the damage threshold. These data indicate that the mitochondrion is an earlier target of DOTC photosensitization than the cell membrane. The critical photochemistry appears to involve singlet oxygen.

Carcinoma, Transitional Cell↗
PubMed1985

Photoaffinity labeling of the tetracycline binding site of the Escherichia coli ribosome. The uses of a high intensity light source and of radioactive sancycline derivatives.

[3H]Tetracycline (TC) has been shown to photoincorporate into the Escherichia coli ribosome. However, the utility of this process for characterizing the TC binding site on the ribosome is diminished by competing side reactions which also lead to incorporation of radioactivity. In this work we first conducted a detailed study of the labeling processes occurring when ribosomes are irradiated in the presence of [3H]TC with a common, rather low intensity, lamp. On the basis of the results of this study we next explored the usefulness for photoaffinity labeling of the TC site of both irradiation with a high-intensity laser and radioactive, functional TC derivatives having different photochemical properties than TC itself. Labeling patterns determined by polyacrylamide gel electrophoretic analysis of ribosomal proteins extracted from photoaffinity-labeled 30S subunits provided strong evidence that these two approaches offer distinct advantages for characterizing the TC binding site.

Affinity Labels↗
PubMed1984

Mechanism of tetracycline phototoxicity.

Studies were made to determine factors important in the phototoxicity mechanism of 7 clinically used tetracyclines (TC). The clinical phototoxicity, the rates of photochemical degradation, and the in vitro phototoxicity of the TCs were qualitatively but not quantitatively correlated. Phototoxicity in vitro was partially oxygen-dependent and possibly singlet oxygen is involved. The contribution of photoproducts to the phototoxic process may be the basis for the reported differences between the in vivo action spectrum and the absorption spectrum of demethylchlorotetracycline. A mechanistic model for in vivo phototoxicity is proposed where the absorption of UVA radiation by TC leads to at least two main processes: (i) photosensitization by the drug of biologic molecules to cause phototoxicity; (ii) production of one or more photoproducts which photosensitize by absorption of visible radiation.

Humans↗