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Delta-aminolevulinic acid cytotoxic effects on human hepatocarcinoma cell lines.

De Siervi A, Vazquez ES, Rezaval C, Rossetti MV, del Batlle AM - BMC Cancer (2002)

Bottom Line: Because apoptosis is usually associated with DNA fragmentation, the DNA of ALA treated and untreated cells were analyzed.To elucidate the mechanisms of ALA induced apoptosis, we examined its effect on p53 expression.CDK2 and CDK4 protein levels were reduced after ALA treatment at physiological concentrations.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centro de Investigaciones sobre Porfirinas y Porfirias, Argentine National Research Council (CONICET), Department of Biological Chemistry, FCEN, University of Buenos Aires, Argentina. desiervi@qb.fcen.uba.ar

ABSTRACT

Background: Acute Intermittent Porphyria is a genetic disorder of heme metabolism, characterized by increased levels of porphyrin precursors, delta-aminolevulinic acid (ALA) and porphobilinogen (PBG). ALA has been reported to generate reactive oxygen species and to cause oxidative damage to proteins, subcellular structures and DNA. It is known that oxidative stress can induce apoptosis. The aim of this work was to study the cytotoxic effect of ALA on two hepatocarcinoma cell lines.

Results: We have determined the impact of ALA on HEP G2 and HEP 3B hepatocarcinoma cell lines survival as measured by the MTT assay. ALA proved to be cytotoxic in both cell lines however; HEP G2 was more sensitive to ALA than HEP 3B. Addition of hemin or glucose diminished ALA cytotoxicity in HEP G2 cells; instead it was enhanced in HEP 3B cells. Because apoptosis is usually associated with DNA fragmentation, the DNA of ALA treated and untreated cells were analyzed. The characteristic pattern of DNA fragmentation ladders was observed in ALA treated cells. To elucidate the mechanisms of ALA induced apoptosis, we examined its effect on p53 expression. No changes in p53 mRNA levels were observed after exposure of both cell lines to ALA for 24 h. CDK2 and CDK4 protein levels were reduced after ALA treatment at physiological concentrations.

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p53 mRNA expression in HEP G2 cells under ALA treatment. Total RNA was isolated after treatment of cells with ALA (0–5 mM) during 24 h and hybridized with cDNA p53 specific probe and normalized relative to GAPDH. Intensity of bands were analysed with ImageMaster.
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Figure 5: p53 mRNA expression in HEP G2 cells under ALA treatment. Total RNA was isolated after treatment of cells with ALA (0–5 mM) during 24 h and hybridized with cDNA p53 specific probe and normalized relative to GAPDH. Intensity of bands were analysed with ImageMaster.

Mentions: To elucidate the mechanisms of ALA causing apoptosis selectively, we have examined the effect of ALA on p53 mRNA levels. No any change in mRNA p53 levels for 24 h was observed in HEP G2 cells (Figure 5). Only at concentrations as high as 5 mM ALA, p53 mRNA expression was 25% diminished.


Delta-aminolevulinic acid cytotoxic effects on human hepatocarcinoma cell lines.

De Siervi A, Vazquez ES, Rezaval C, Rossetti MV, del Batlle AM - BMC Cancer (2002)

p53 mRNA expression in HEP G2 cells under ALA treatment. Total RNA was isolated after treatment of cells with ALA (0–5 mM) during 24 h and hybridized with cDNA p53 specific probe and normalized relative to GAPDH. Intensity of bands were analysed with ImageMaster.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC101407&req=5

Figure 5: p53 mRNA expression in HEP G2 cells under ALA treatment. Total RNA was isolated after treatment of cells with ALA (0–5 mM) during 24 h and hybridized with cDNA p53 specific probe and normalized relative to GAPDH. Intensity of bands were analysed with ImageMaster.
Mentions: To elucidate the mechanisms of ALA causing apoptosis selectively, we have examined the effect of ALA on p53 mRNA levels. No any change in mRNA p53 levels for 24 h was observed in HEP G2 cells (Figure 5). Only at concentrations as high as 5 mM ALA, p53 mRNA expression was 25% diminished.

Bottom Line: Because apoptosis is usually associated with DNA fragmentation, the DNA of ALA treated and untreated cells were analyzed.To elucidate the mechanisms of ALA induced apoptosis, we examined its effect on p53 expression.CDK2 and CDK4 protein levels were reduced after ALA treatment at physiological concentrations.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centro de Investigaciones sobre Porfirinas y Porfirias, Argentine National Research Council (CONICET), Department of Biological Chemistry, FCEN, University of Buenos Aires, Argentina. desiervi@qb.fcen.uba.ar

ABSTRACT

Background: Acute Intermittent Porphyria is a genetic disorder of heme metabolism, characterized by increased levels of porphyrin precursors, delta-aminolevulinic acid (ALA) and porphobilinogen (PBG). ALA has been reported to generate reactive oxygen species and to cause oxidative damage to proteins, subcellular structures and DNA. It is known that oxidative stress can induce apoptosis. The aim of this work was to study the cytotoxic effect of ALA on two hepatocarcinoma cell lines.

Results: We have determined the impact of ALA on HEP G2 and HEP 3B hepatocarcinoma cell lines survival as measured by the MTT assay. ALA proved to be cytotoxic in both cell lines however; HEP G2 was more sensitive to ALA than HEP 3B. Addition of hemin or glucose diminished ALA cytotoxicity in HEP G2 cells; instead it was enhanced in HEP 3B cells. Because apoptosis is usually associated with DNA fragmentation, the DNA of ALA treated and untreated cells were analyzed. The characteristic pattern of DNA fragmentation ladders was observed in ALA treated cells. To elucidate the mechanisms of ALA induced apoptosis, we examined its effect on p53 expression. No changes in p53 mRNA levels were observed after exposure of both cell lines to ALA for 24 h. CDK2 and CDK4 protein levels were reduced after ALA treatment at physiological concentrations.

Show MeSH
Related in: MedlinePlus