Glutathione transferase mu 2 protects glioblastoma cells against aminochrome toxicity by preventing autophagy and lysosome dysfunction.

Abstract

U373MG cells constitutively express glutathione S-transferase mu 2 (GSTM2) and exhibit (3)H-dopamine uptake, which is inhibited by 2 µM of nomifensine and 15 µM of estradiol. We generated a stable cell line (U373MGsiGST6) expressing an siRNA against GSTM2 that resulted in low GSTM2 expression (26% of wild-type U373MG cells). A significant increase in cell death was observed when U373MGsiGST6 cells were incubated with 50 µM purified aminochrome (18-fold increase) compared with wild-type cells. The incubation of U373MGsiGST6 cells with 75 µM aminochrome resulted in the formation of autophagic vacuoles containing undigested cellular components, as determined using transmission electron microscopy. A significant increase in autophagosomes was determined by measuring endogenous LC3-II, a significant decrease in cell death was observed in the presence of bafilomycin A 1, and a significant increase in cell death was observed in the presence of trehalose. A significant increase in LAMP2 immunostaining was observed, a significant decrease in bright red fluorescence of lysosomes with acridine orange was observed, and bafilomycin A 1 pretreatment reduced the loss of lysosome acidity. A significant increase in cell death was observed in the presence of lysosomal protease inhibitors. Aggregation of TUBA/α-tubulin (tubulin, α) and SQSTM1 protein accumulation were also observed. Moreover, a significant increase in the number of lipids droplets was observed compared with U373MG cells with normal expression of GSTM2. These results support the notion that GSTM2 is a protective enzyme against aminochrome toxicity in astrocytes and that aminochrome cell death in U373MGsiGST6 cells involves autophagic-lysosomal dysfunction.

• Authors: Sandro Huenchuguala, Patricia Muñoz, Patricio Zavala, Mónica Villa, Carlos Cuevas, Ulises Ahumada, Rebecca Graumann, Beston F Nore, Eduardo Couve, Bengt Mannervik, Irmgard Paris, Juan Segura-Aguilar
• Journal: Autophagy (Autophagy) Vol. 10 Issue 4 Pg. 618-30 (Apr 2014) ISSN: 1554-8635 [Electronic] United States
• PMID: 24434817 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Indolequinones
• Protective Agents
• aminochrome 1
• Glutathione Transferase
• glutathione S-transferase Mu 2

Topics

• Astrocytes (metabolism)
• Autophagy (physiology)
• Cell Line
• Glioblastoma (metabolism)
• Glutathione Transferase (metabolism)
• Humans
• Indolequinones (toxicity)
• Lysosomes (metabolism)
• Mitochondria (metabolism)
• Protective Agents (metabolism)