title: RIP3 expression induces a death profile change in U2OS osteosarcoma cells after 5-ALA-PDT.
abstract: OBJECTIVE
  The receptor-interacting protein 3 (RIP3) has recently been outlined as a key necrosis mediator but is also thought to participate in the regulation of apoptosis. The aim of this study is to compare the cell death profile induced by 5-aminolevulic acid (5-ALA)-mediated photodynamic therapy (PDT) in the RIP3-deficient cell line U2OS and in U2OS cells in which the expression of RIP3 was restored.
  
  
  METHODS
  RIP3-expressing U2OS cells (RIP3-U2OS) were obtained after transfection and antibiotic selection. Wild type and RIP3-U2OS cells were treated by 5-ALA-PDT. Overall cell viability was evaluated and different parameters characteristic of apoptosis, autophagy, and necrosis were studied.
  
  
  RESULTS
  Surprisingly, the survival of RIP3-U2OS cells was higher compared to that of the wild type cells. In addition, RIP3-U2OS cell death was decreased by a zVAD-fmk pre-treatment. A higher cleavage of caspase-3, 7, 8, 9, and PARP was also detected in these cells, pointing out to the activation of caspase-dependent apoptosis. In parallel, a thrust of autophagy was clearly identified in the RIP3-U2OS cells. Conversely, RIP3-U2OS exhibited a lower level of necrosis than the wild types. Interestingly, necrostatin-1 efficiently decreased necrosis level in RIP3-U2OS but not in wild type cells.
  
  
  CONCLUSIONS
  Expression of RIP3 in U2OS cells led to a better survival but also to a death profile change in response to PDT. The apoptotic and autophagic pathways were clearly up-regulated compared to the RIP3-deficient wild type cells. However, induction of necrosis was weaker in the RIP3-U2OS cells. In this context, autophagy is likely to play a protective role against PDT-induced cell death and to allow a better survival of RIP3-U2OS cells. This work also highlights the important role played by RIP3 in the apoptotic pathway, although the modalities are still widely unknown.
authors: Virology and Immunology Unit, GIGA-Research, University of Liège, Liège, Belgium.
authors: Isabelle
authors: I
authors: Coupienne
authors: Grégory
authors: G
authors: Fettweis
authors: Jacques
authors: J
authors: Piette
mesh_terms: D000622:Aminolevulinic Acid / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects*; D001343:Autophagy / Q000187:drug effects*; D015153:Blotting, Western; D001859:Bone Neoplasms / Q000188:drug therapy / Q000473:pathology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D006801:Humans; D009336:Necrosis; D012516:Osteosarcoma / Q000188:drug therapy / Q000473:pathology*; D010778:Photochemotherapy; D017319:Photosensitizing Agents / Q000494:pharmacology* / Q000627:therapeutic use; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000378:metabolism*
chemical_list: D017319:Photosensitizing Agents; D000622:Aminolevulinic Acid; C119705:RIPK3 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases