Tumor cell lines resistant to ALA-mediated photodynamic therapy and possible tools to target surviving cells

We isolated and characterized cell lines resistant to aminolevulinic acid (ALA)-mediated photodynamic therapy (PDT) derived from a murine adenocarcinoma and studied cross resistance with other injuries. The most resistant clones were numbers 4 and 8, which exhibited 6.7- and 4.2-fold increase in resistance respectively. Several characteristics were altered in these clones. A 2-fold increase in cell volume, higher cell spreading, and a more fibroblastic, dendritic pattern, were the morphology features that led us to think they could have different adhesive, invasive or metastatic phenotypes. The amount of porphyrins synthesized per cell in the resistant clones was similar to the parental line but, when it was expressed per mg protein, there was a 2-fold decrease, with a higher proportion of hydrophilic porphyrins. These cells were not cross-resistant to photosensitization with Benzoporphyrin derivative and Merocyanine 540, but exhibited a slight resistance to exogenous protoporphyrin IX treatment. Both clones displayed higher protein content and increased number of mitochondria, together with a higher oxygen consumption. The distinctive features found in the resistant lines led as to think how to exploit the changes induced by PDT treatment to target surviving cells. Those hypoxic cells can be also a preferential target of bioreductive drugs and hypoxia-directed gene therapy, and would be sensitive to treatment with other photosensitizers.Fil: Casas, Adriana Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Perotti, Christian. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Ortel, Bernhard. Harvard Medical School; Estados UnidosFil: Di Venosa, Gabriela Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Saccoliti, María. Gobierno de la Ciudad Autónoma de Buenos Aires. Hospital General de Agudos Carlos Durand; ArgentinaFil: Batlle, Alcira Maria del C.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Hasan, Tayyaba. Harvard Medical School; Estados Unido

Distribution of mosquitoes in the South East of Argentina and first report on the analysis based on 18S rDNA and COI sequences

Although Mar del Plata is the most important city on the Atlantic coast of Argentina, mosquitoes inhabiting such area are almost uncharacterized. To increase our knowledge in their distribution, we sampled specimens of natural populations. After the morphological identification based on taxonomic keys, sequences of DNA from small ribosomal subunit (18S rDNA) and cytochrome c oxidase I (COI) genes were obtained from native species and the phylogenetic analysis of these sequences were done. Fourteen species from the genera Uranotaenia, Culex, Ochlerotatus and Psorophora were found and identified. Our 18S rDNA and COI-based analysis indicates the relationships among groups at the supra-species level in concordance with mosquito taxonomy. The introduction and spread of vectors and diseases carried by them are not known in Mar del Plata, but some of the species found in this study were reported as pathogen vectors

Estado del conocimiento de humedales del norte patagónico (Argentina): aspectos relevantes e importancia para la conservación de la biodiversidad regional State of the knowledge of north Patagonian wetlands (Argentina): major aspects and importance for regional biodiversity conservation

Las áreas de humedal ocupan casi el 5 % del territorio de la Patagonia argentina. En la zona extrandina y el desierto patagónico estos ecosistemas son esenciales para sostener numerosas especies de fauna y flora silvestre. Además, los humedales del norte patagónico permiten el desarrollo de comunidades acuáticas complejas y constituyen el hábitat crítico de grupos de organismos como los peces y los anfibios incluidos en listados de conservación prioritaria. Estos ecosistemas tienen un funcionamiento complejo que depende estrechamente de las fluctuaciones de las variables meteorológicas y son por lo tanto muy sensibles a los efectos del cambio climático. En la actualidad el uso de la tierra, la introducción de especies y la interacción de estos aspectos con el cambio climático constituyen las variables de mayor impacto sobre estos ambientes patagónicos. En este trabajo se presenta información geográfica, geomorfológica y climática de un número de humedales del norte de la Patagonia argentina. Por otra parte, se sintetiza información sobre la flora y la distribución de peces y anfibios con el propósito de enfatizar la importancia de estos ambientes en el mantenimiento de la biodiversidad regional. Por último, se señalan los procesos que actualmente afectan a estos sistemas poniendo en evidencia la necesidad de obtención de conocimientos básicos para el diseño de planes de manejo y conservación de sus comunidades biológicasAlmost 5 % of the lansdscape of Patagonia is occupied by wetlands. In the extra-andean region and the patagonian desert, wetlands are essential for sustaining biodiversity and wildlife. Besides, North patagonian wetlands present complex aquatic communities and provide habitat for threatened species of fishes and amphibians. These ecosystems have complex dynamics mostly driven by climatic fluctuations which make them vulnerable to the effect of climate change. Nowadays, the main impact on patagonian wetland is due to land use, species introduction, and the interaction of these disturbances with climate change. In this paper we summarize geographic, geomorphological and climatic information of several wetlands located in North Patagonia (Argentina). Besides, we present information on aquatic vegetation, fish and amphibian fauna distributions and endemisms with the aim of emphasizing the importance of wetlands to sustain regional biodiversity. Finally, we highlight the processes impacting wetlands at present time and point out the importance of obtaining basic information for management and conservation strategie

Mechanisms of Resistance to Photodynamic Therapy: An Update

Photodynamic Therapy (PDT) involves the combination of light andphotosensitizer (PS). Therefore, it is possible for cells to develop resistance basedon the doses of PS used or the light dose. The data compiled by several authorsmake it clear that the degree of cell resistance to PDT is highly dependent on the PSused; however, no cellular characteristics have yet been identified as predictors ofPDT resistance. The mechanisms by which the treated tissue becomes resistant tothe PS share some similarities to those found in general drug resistance and radioresistance,and they are mainly related to both the bioavailability of the PS and tothe mechanisms of detoxification of the generated reactive oxygen species. Amongthe features related to PDT resistance are: the expression of p-glycoprotein andABCG2 transporters, the abrogation of apoptosis and autophagy, the induction ofantioxidant defences, the induction of HSPs changes in cytoskeleton and adhesion,the induction of cyclooxygenases, the production of nitric oxide and hypoxia; theseare some but not all of the factors involved in the development of resistance. As ageneral rule, all the authors that reported resistance to PDT have attributed this phenomenonto several factors acting in concert. In this chapter, we will review someof the most important aspects related to PDT resistance.Fil: Casas, Adriana Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Perotti, Maria Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Di Venosa, Gabriela Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Batlle, Alcira María del C.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; Argentin

Sensitivity to ALA-PDT of cell lines with different nitric oxide production and resistance to NO cytotoxicity

In this work, we studied the in vitro interactions between aminolevulinic acid (ALA)-mediated photodynamic therapy (PDT) and nitric oxide (NO), as well as the interactions between ALA, porphyrins and some NO donors and precursors. We employed three murine adenocarcinoma cell lines: LM2, which does not produce NO; LM3, which produces NO, and LM3-SNP, a variant of LM3 resistant to NO producing the same amount of NO as the parental. We did not find cross-resistance between NO-induced cytotoxicity and ALA-PDT. In spite of the lower porphyrin synthesis, LM2 cells show the highest sensitivity to ALA-PDT. However, we hypothesised that this is not related to the lack of endogenous NO production, because modulation of NO levels did not modify the response to PDT in any of the cell lines. Two unexpected results were found: the enhancement of NO production from the donor sodium nitroprusside (SNP) induced by ALA in both cells and medium, and the inhibition by ALA of NO production from arginine. We also found that SNP strongly protected the cells from ALA-PDT by impairing porphyrin biosynthesis as a consequence of an inhibition of the enzyme ALA dehydratase. We were not able to evaluate the action of NO derived from SNP because of the unexpected porphyrin impairment. On the other hand, impairment of NO from Arginine driven by ALA, although not modulating in vitro the ALA-PDT response, by increasing in vivo blood flow, may be contributing to the mechanism of tumour cures.Fil: Di Venosa, Gabriela Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Perotti, Christian. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Fukuda, Haydee. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Batlle, Alcira María del C.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Casas, Adriana Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; Argentin

Carbohydrate metabolism and fruit quality are affected in frost-exposed Valencia orange fruit

Exposure of citrus fruit to frost often results in the development of freeze injury during their maturation in planta. This work was aimed to analyze changes in the biochemistry and enzymology of carbohydrate metabolism in freeze-injured orange fruit (Citrus sinensis var. Valencia late) and the involvement of oxidative stress in frost damage. The activities of pyrophosphate-dependent phosphofructokinase, phosphoenolpyruvate carboxylase and several fermentative enzymes increased in frost-exposed (FE) fruit, while NADP-malic enzyme and the mitochondrial isoform of NAD-malate dehydrogenase showed a reduction in their activities. Western blot analysis indicated a correlation between activity levels and protein content. Respiration rate in whole fruit was reduced by 40%, whereas the flavedo showed a more pronounced decline (53%). Volatile compound (i.e. ethanol and acetaldehyde) content was significantly higher in FE fruit than in control, as was that of l-malate (three-fold). Additionally, FE fruit showed a marked decrease in the maturity index (24%) because of a higher titratable acidity (39%). Evidence is presented that oxidative stress is involved in freeze-induced damage of orange fruit, where oxidative damage to lipids and proteins, and a greater electrolyte leakage in the flavedo were also observed. The results suggest that freezing temperatures provoke a notable metabolic switch in citrus fruit toward a fermentative stage, resulting in low-quality fruits.Fil: Falcone Ferreyra, María Lorena.Fil: Perotti, Valeria Elisa.Fil: Figueroa, Carlos Maria.Fil: Sergio Garran.Fil: Perla C. Anderson.Fil: Daniel Vazquez.Fil: Iglesias, Alberto Alvaro.Fil: Podesta, Florencio Esteban

Decreased metastatic phenotype in cells resistant to aminolevulinic acid-photodynamic therapy

Photodynamic therapy (PDT) is a novel cancer treatment utilising a photosensitiser, visible light and oxygen. PDT often leaves a significant number of surviving tumour cells. In a previous work, we isolated and studied two PDT resistant clones derived from the mammary adenocarcinoma LM3 line (Int. J. Oncol. 29 (2006) 397-405). The isolated Clon 4 and Clon 8 exhibited a more fibroblastic, dendritic pattern and were larger than the parentals. In the present work we studied the metastatic potential of the two clones in comparison with LM3. We found that 100% of LM3 invaded Matrigel, whereas only 19 ± 6% and 24 ± 7% of Clon 4 and Clon 8 cells invaded. In addition, 100% of LM3 cells migrated towards a chemotactic stimulus whereas 38 ± 8% and 73 ± 10% of Clones 4 and 8, respectively, were able to migrate. In vivo, 100% of the LM3 injected mice developed spontaneous lung metastasis, whereas none of the Clon 8 did, and only one of the mice injected with Clon 4 did. No differences were found in the proteolytic enzyme profiles among the cells. Anchorage-dependent adhesion was also impaired in vivo in the resistant clones, evidenced by the lower tumour take, latency time and growth rates, although both clones showed in vitro higher binding to collagen I without overexpression of β1 integrin. This is the first work where the metastatic potential of cells surviving to PDT has been studied. PDT strongly affects the invasive phenotype of these cells, probably related to a higher binding to collagen. These findings may be crucial for the outcome of ALA-PDT of metastatic tumours, although further studies are needed to extrapolate the results to the clinic employing another photosensitisers and cell types.Fil: Casas, Adriana Gabriela. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Di Venosa, Gabriela Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Vanzulli, Silvia. Academia Nacional de Medicina de Buenos Aires; ArgentinaFil: Perotti, Christian. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; ArgentinaFil: Mamome, Leandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Rodriguez, Lorena Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Simian, Marina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Juarranz, Angeles. Universidad Autónoma de Madrid; EspañaFil: Pontiggia, Osvaldo Alejandro. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Hasan, Tayyaba. Harvard Medical School; Estados UnidosFil: Batlle, Alcira María del C.. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Investigaciones sobre Porfirinas y Porfirias. Universidad de Buenos Aires. Centro de Investigaciones sobre Porfirinas y Porfirias; Argentin