Hyperosmolarity-mediated mitochondrial dysfunction requires Transglutaminase-2 in human corneal epithelial cells.
Hyperosmolar-induced ocular surface cell death is a key mitochondria-mediated event in inflammatory eye diseases. Transglutaminase (TGM)-2, a cross-linking enzyme, is purported to mediate cell death, but its link to mitochondria is unclear. In the cornea, the integrity of the epithelial cells is important for maintaining transparency of the cornea and therefore functional vision. We evaluated the role of TGM-2 and its involvement in hyperosmolarity-stimulated mitochondrial cell death in human corneal epithelial (HCE-T) cells. HCE-T cell lines stably expressing either shRNA targeting TGM-2 (shTG) or scrambled shRNA (shRNA) were constructed. Hyperosmolar conditions reduced viability and increased mitochondrial depolarization in shRNA cells. However, hyperosmolarity failed to induce mitochondrial depolarization to the same extent in shTG cells. Transient overexpression of TGM-2 resulted in very high levels of TGM-2 expression in shTG and shRNA cells. In the case of shTG cells after overexpression of TGM-2, hyperosmolarity induced the same extent of mitochondrial depolarization as similarly treated shRNA cells. Overexpression of TGM-2 also elevated transamidase activity and reduced viability. It also induced mitochondrial depolarization, increased caspase-3/7 and -9 activity, and these increases were partially suppressed by pan-caspase inhibitor Z-VAD-FMK. Corneal epithelial apoptosis via mitochondrial dysfunction after hyperosmolar stimulation is partially dependent on TGM-2. This TGM-2-dependent mechanism occurs in part via caspase-3/7 and -9. Protection against mitochondrial stress in the ocular surface targeting TGM-2 may have important implications in the survival of cells in hyperosmolar stress.
Png
Evelyn
E
Ocular Wound Healing and Therapeutics Laboratory, Singapore Eye Research Institute, Singapore, Singapore.
Samivelu
G K
GK
Yeo
S H
SH
Chew
J
J
Chaurasia
Shyam S
SS
Tong
Louis
L
D049109:Cell Proliferation; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000201:enzymology* / Q000473:pathology*; D019573:Epithelium, Corneal / Q000166:cytology*; D005453:Fluorescence; D019204:GTP-Binding Proteins; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D053078:Membrane Potential, Mitochondrial; D008928:Mitochondria / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D008954:Models, Biological; D009994:Osmolar Concentration; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases / Q000235:genetics / Q000378:metabolism*
D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D019204:GTP-Binding Proteins