Search PubMed⌕ Search

Biomedical subjects

N Rucker

Publications and source records attributed to N Rucker.

At least 19 recordsLinked to original sources
PubMed1996

Photodynamic therapy-mediated oxidative stress can induce expression of heat shock proteins.

Photodynamic therapy (PDT) is an experimental cancer therapy inducing tumor tissue damage via photosensitizer-mediated oxidative cytotoxicity. A previous report indicates that oxidative stress induced by hydrogen peroxide or menadione activates the heat shock transcription factor in mouse cells but does not result in either increased transcription or translation of heat shock proteins (HSPs). Our study documents that photosensitizer-mediated oxidative stress can activate the heat shock factor as well as increase HSP-70 mRNA and protein levels in mouse RIF-1 cells. The cellular heat shock response after PDT varied for the different photosensitizers being examined. Treatments using either a chlorin (mono-L-aspartyl chlorin-e6)- or purpurin (tin etio-purpurin)-based sensitizer induced HSP-70 expression, whereas identical photosensitization conditions with a porphyrin (Photofrin)-based sensitizer failed to induce a cellular HSP response. These sensitizers, which generate singlet oxygen as the primary oxidant during photosensitization, were used in experiments under isoeffective treatment conditions. HSP-70 expression after photosensitization was associated with the concomitant induction of thermotolerance in PDT-treated cells. Interestingly, reverse transcription-PCR demonstrated that in vivo PDT treatments of RIF-1 tumors induce expression of HSP-70 for all photosensitizers including Photofrin. These results indicate that photosensitizer-generated singlet oxygen exposure can induce in vitro and in vivo HSP-70 expression, and that specific subcellular targets of PDT (which can differ for various sensitizers) are determinants for HSP-70 activation after oxidative stress.

Animals↗
PubMed1995

Decreased expression and function of alpha-2 macroglobulin receptor/low density lipoprotein receptor-related protein in photodynamic therapy-resistant mouse tumor cells.

Parental and photodynamic therapy (PDT)-resistant mouse, radiation-induced fibrosarcoma cell lines were evaluated using mRNA differential display in an attempt to identify unique transcripts. We detected one transcript that was consistently present in the parental cells but absent in PDT-resistant cells. The transcript was cloned, sequenced, and identified as alpha-2 macroglobulin receptor/low density lipoprotein receptor-related protein (alpha-2 MR/LRP). Northern and Western immunoblot analysis confirmed that receptor expression was present in the parental cell line but barely detectable in PDT-resistant cells. Functionality of the receptor was evaluated by exposing cells to Pseudomonas exotoxin A. alpha-2 MR/LRP is responsible for Pseudomonas exotoxin A internalization, and only the parental cells exhibited toxin-mediated cytotoxicity. The binding and endocytosis of activated alpha-2 macroglobulin and lipoproteins by alpha-2 MR/LRP are consistent with modulating uptake and localization of photosensitizers. Our results demonstrate that PDT-resistant murine tumor cells exhibit minimal alpha-2 MR/LRP activity and suggest that this receptor plays a role in PDT sensitivity by modulating photosensitizer uptake and/or subcellular localization.

ADP Ribose Transferases↗
PubMed1992

Establishment of a Chinese hamster ovary cell line that expresses grp78 antisense transcripts and suppresses A23187 induction of both GRP78 and GRP94.

GRP78, a 78,000 dalton protein residing in the endoplasmic reticulum, is postulated to play important roles in protein folding and cell survival during calcium and other physiological stress. Here we describe the construction of an eukaryotic expression vector for the constitutive expression of grp78 antisense RNA and the creation of a CHO cell line, 78WO, which expresses high levels of the grp78 antisense RNA through amplification of the stably transfected antisense vector. We observed that whereas 78WO maintains a basal level of GRP78 similar to that of control cells, GRP78 is no longer inducible by A23187. The 78WO cells have undergone a compensatory increase in grp78 transcription such that the effects of antisense are cancelled out at the protein level under nonstressed conditions. In these same cells, GRP94, a 94,00 dalton ER protein, is also rendered noninducible by A23187. This provides the first evidence that the regulation of two ER proteins might be coupled such that the failure to induce GRP78 results in the down-regulation of GRP94. The 78WO cell line grows with a doubling time of about 26 hr and exhibits decreased tolerance to A23187, suggesting the GRPs contribute to cell viability under calcium stress. The establishment of this cell line, which can be stably maintained, will provide a useful tool for testing whether the induction of the GRPs is important for protein folding or transport and whether their enhanced synthesis is the cause or consequence of a variety of physiological adaptations.

Animals↗
PubMed1991

Glucose regulated protein induction and cellular resistance to oxidative stress mediated by porphyrin photosensitization.

Photodynamic therapy (PDT) utilizes a tumor localizing porphyrin photosensitizer in the clinical treatment of cancer. At a mechanistic level, porphyrin photosensitization generates reactive oxygen species which initiate oxidative damage to a wide spectrum of biomolecules. Cellular stress proteins are also increased following oxidative stress treatments. In the current study, we examined porphyrin photosensitizing parameters associated with induction of the glucose regulated family of stress proteins. Elevated levels of mRNA encoding glucose regulated proteins (GRPs) as well as increases in GRP protein synthesis were observed for mouse radiation induced fibrosarcoma cells exposed to an extended (16-h) porphyrin incubation prior to light exposure. However, a short (1-h) porphyrin incubation prior to light treatment (designed to produce comparable phototoxicity as PDT using the 16-h porphyrin incubation protocol) was associated with only minimal increases in GRP mRNA levels or GRP protein synthesis. The relationship between GRP levels and PDT sensitivity was examined in radiation induced fibrosarcoma cells pretreated with the calcium ionophore A-23187 in order to overexpress GRPs prior to photosensitization. Resistance to PDT was observed in cells overexpressing GRPs only under photosensitizing conditions associated with the extended porphyrin incubation protocol, and this response was not due to changes in cellular porphyrin uptake. In separate experiments, a transient elevation of GRP mRNA levels was observed in transplanted mouse mammary carcinomas following in vivo PDT treatments. Our results indicate that specific targets of oxidative damage (modulated by porphyrin incubation conditions) instead of generalized cellular exposure to reactive oxygen species are correlated with PDT mediated GRP induction. In this regard, GRP induction may be a useful in vivo biochemical marker of PDT mediated injury. These results also support the hypothesis that GRPs may play a role in modulating sensitivity to cellular stresses including certain types of oxidative injury.

Animals↗
PubMed1991

Direct comparison of in-vitro and in-vivo Photofrin-II mediated photosensitization using a pulsed KTP pumped dye laser and a continuous wave argon ion pumped dye laser.

A pulsed KTP pumped dye laser (25 kHz repetition rate and 470 nsec pulse width) has been compared to a continuous wave argon ion pumped dye laser as the source of 630 nm light during in-vitro and in-vivo Photofrin-II mediated photosensitization studies. Individual experiments documented the effectiveness of each laser system on a) photosensitizer induced cytotoxicity and induction of stress protein synthesis using Chinese hamster fibroblasts; b) photobleaching of Photofrin-II in aqueous solution; c) Photofrin II mediated photosensitization of normal mouse skin; d) Photofrin II mediated photodynamic therapy of a mouse mammary carcinoma; and e) tumor temperature levels generated during laser exposure. Comparable results were obtained for both laser systems in all experiments.

Animals↗
PubMed1991

Increased transcription and translation of heme oxygenase in Chinese hamster fibroblasts following photodynamic stress or Photofrin II incubation.

Porphyrin mediated photosensitization can enhance the transcription and translation of several oxidative stress genes. In this study, we report on the enhanced expression of the gene encoding for heme oxygenase in Chinese hamster fibroblasts by; (1) incubation in Photofrin II; (2) Photofrin II mediated photosensitization; and (3) photosensitization induced by Rose Bengal. Increased expression of heme oxygenase mRNA was accompanied by a concomitant increase in the synthesis of the 34 kDa heme oxygenase protein. Western blot analysis using antibody to heme oxygenase confirmed the immunoreactivity of the 34 kDa protein induced by Photofrin II and PDT. These results demonstrate that heme oxygenase can be activated by non-metalloporphyrins as well as by photosensitization associated with singlet oxygen mediated subcellular injury.

Animals↗
PubMed1990

Porphyrin photosensitivity in cell lines expressing a heat-resistant phenotype.

In vitro sensitivity to porphyrin-mediated photodynamic therapy (PDT) has been examined in cell lines resistant to hyperthermia. Parental (HA-1) and heat-resistant (3012) Chinese hamster fibroblasts as well as parental (RIF-1) and temperature-resistant (TR-4, TR-5, and TR-10) mouse radiation-induced fibrosarcoma cells were evaluated for thermal and PDT sensitivity. Quantitative survival curves were generated, and porphyrin uptake properties were obtained for all cell lines. Significant resistance to hyperthermia (45 degrees C for varying exposure periods) was documented for the 3012 and temperature-resistant RIF cell strains when compared with the parent lines. Normal and heat-resistant clones, however, exhibited comparable levels of porphyrin uptake and photosensitivity. Our results indicate that cross-resistance between hyperthermia and PDT is not observed and that members of the Mr 70,000 heat shock protein family (which are elevated in the thermal-resistant cells and which may be associated with the heat-resistant phenotype) do not play a significant role in modulating PDT sensitivity. Mechanisms of in vitro cytotoxicity appear to be different for PDT and hyperthermia even though possible subcellular targets (such as the plasma membrane) and types of damage (protein denaturation) may be similar for the two modalities.

Animals↗
PubMed1989

Properties and applications of photodynamic therapy.

Photodynamic therapy (PDT) is the treatment of malignant lesions with visible light following the systemic administration of a tumor-localizing photosensitizer. Pharmacological and photochemical properties of the photosensitizer are combined with precise delivery of laser-generated light to produce a treatment which can offer selective tumoricidal action. Hematoporphyrin derivative (HD) and a purified component called Photofrin II are currently being used in clinical PDT. Initial patient results have been encouraging, and considerable interest has developed in the synthesis and evaluation of new photosensitizers with improved photochemical and pharmacological characteristics. In addition, there has been a gradual increase in knowledge related to in vitro and in vivo mechanisms of action of PDT. This report provides an overview of the properties and applications of PDT. Information and data related to drug development, photochemistry, subcellular targets, in vivo responses, and clinical trials of PDT are presented.

Clinical Trials as Topic↗
PubMed1988

Differential cell photosensitivity following porphyrin photodynamic therapy.

Experiments were performed to determine if differences in porphyrin photosensitivity could be observed for cells with varying efficiency in DNA damage repair, as well as for cells which make up components of the vasculature. Photofrin II is undergoing current clinical evaluation for photodynamic therapy of solid tumors, and therefore the retention, dark toxicity, and photosensitizing effects of this drug on human DNA repair-deficient fibroblasts (ataxia telangiectasia and xeroderma pigmentosum) were compared to normal human fibroblasts. In addition, bovine cells of endothelial, smooth muscle, and fibroblast origin were compared for porphyrin retention, toxicity, and photosensitivity. All human fibroblasts exhibited porphyrin-induced dark toxicity, but there were no significant differences in photosensitization or porphyrin retention for any of these cell lines. However, bovine endothelial cells were considerably more photosensitive than smooth muscle or fibroblast cells treated under identical conditions. All bovine cells accumulated similar levels of porphyrin, and therefore the increased sensitivity of the endothelial cells was not due to differences in porphyrin retention. These results provide additional evidence that nuclear damage and/or repair is not a dominant factor in the cytotoxicity induced by porphyrin photosensitization. In addition, these results indicate that endothelial cell photosensitivity may play a role in the vascular damage observed following photodynamic therapy.

Animals↗
PubMed1988

Molecular, cellular, and tissue responses following photodynamic therapy.

Photodynamic therapy (PDT) is being utilized in the treatment of a wide variety of malignant tumors. Results using PDT have been encouraging, and controlled clinical trials are currently being performed. The procedure exploits both the tumor-localizing and -photosensitizing properties of hematoporphyrin derivative or its purified component, Photofrin II. When this porphyrin mixture is administered systemically, it is retained preferentially in tumor tissue as compared to surrounding normal tissue. Localized tumor destruction induced by PDT results from the photochemical generation of cytotoxic oxygen species within the tumor. This review will provide a summary of historical and current research pertaining to molecular, cellular, and tissue responses induced by PDT. Emphasis is placed on information related to the chemistry of current photosensitizers, subcellular targets, preclinical treatment parameters, and clinical responses following PDT.

Animals↗
PubMed1988

Transformation and mutagenic potential of porphyrin photodynamic therapy in mammalian cells.

The transformation and mutagenic potential of porphyrin photodynamic therapy has been examined in mammalian cells. The mutagenic frequency in Chinese hamster cells at the Na+/K+ ATPase locus was measured by resistance to ouabain following treatment with either photodynamic therapy (PDT) or UV irradiation. The C3H 10T 1/2 mouse embryo cell system was used to document the transformation frequency following PDT, UV irradiation, gamma irradiation or exposure to 3-methylcholanthrene (MCA). Treatments with UV irradiation were effective in producing mutants resistant to ouabain, and treatments with UV irradiation, gamma irradiation and MCA generated transformants at frequencies comparable to those which are reported in the literature. However, PDT treatment conditions (which produced a full range of cytotoxicity) did not induce any mutagenic or transformation activity above background levels.

Animals↗
PubMed1986

Expression of potentially lethal damage in Chinese hamster cells exposed to hematoporphyrin derivative photodynamic therapy.

Experiments were performed to determine whether the expression and/or repair of potentially lethal damage could be observed in mammalian cells exposed to hemataporphyrin derivative (HPD) photodynamic therapy (PDT). Photodynamic therapy was combined with posttreatment protocols known to inhibit the repair of potentially lethal damage in cells treated with X-rays, ultraviolet radiation, or alkylating agents. Potentiation of lethal damage from photodynamic therapy was induced by hypothermia (4 degrees C) following short (1 h) or extended (16 h) HPD incubation conditions. Caffeine potentiated the lethal effects of PDT only when cells were incubated with HPD for extended time periods. However, 3-aminobenzamide had no effect on the cytotoxic actions of PDT following either short or extended HPD incubations. Recovery from potentially lethal damage expressed by posttreatment hypothermia was complete within 1 h, while recovery from potentially lethal damage expressed by posttreatment caffeine required time periods of up to 24 h. The lack of effect of 3-aminobenzamide on expression of potentially lethal damage following photodynamic therapy may be related to direct inhibition of adenosine diphosphoribose transferase by photodynamic therapy. These results indicate that the expression and repair of potentially lethal damage can be observed in cells treated with PDT and will vary as a function of porphyrin incubation conditions.

Animals↗
PubMed1985

In vitro and in vivo light dose rate effects related to hematoporphyrin derivative photodynamic therapy.

In vitro and in vivo experiments were performed to evaluate the parameter of light dose rate as it relates to the efficiency of hematoporphyrin derivative (HPD)-induced photosensitization. Exponentially growing Chinese hamster ovary cells were incubated with HPD (25 micrograms/ml) and were then exposed to red light (630 nm) delivered at different dose rates. A total of five dose rates (0.5, 5.0, 15, 23, and 60 milliwatts/sq cm) were examined following a 1-hr HPD incubation, two dose rates (1 and 20 milliwatts/sq cm) were examined after a 12-hr HPD incubation, and three dose rates (0.4, 4, and 40 milliwatts/sq cm) were examined following a 16-hr incubation and a 30-min serum wash protocol. The effect of light dose rate was determined from cell survival curves obtained by standard clonogenic colony formation assays. Similar levels of cellular toxicity were obtained when cells from each HPD incubation group were treated with equal doses of red light delivered at different dose rates. For in vivo experiments, albino mice were given injections of HPD (7.5 mg/kg) and 24 h later the right hind leg of each mouse was treated with localized red light (630 nm). A total dose of 270 J/sq cm was delivered to the right hind leg at dose rates of 5, 25, or 125 milliwatts/sq cm. The resulting acute skin damage induced by HPD photosensitization was scored over a 30-day period, and skin response curves for the three dose rates were obtained. Comparable levels of damage were induced in each of the three experimental groups. The results obtained from both in vitro and in vivo studies indicate that the photosensitizing efficiency of HPD photodynamic therapy is not affected by nonthermal variations in clinically relevant dose rates of delivered light.

Animals↗