Inhibition of Transglutaminase Activity Reduces Extracellular Matrix Accumulation Induced by High Glucose Levels in Proximal Tubular Epithelial Cells

Abstract

Diabetic nephropathy affects 30 -40% of diabetics leading to end-stage kidney failure through progressive scarring and fibrosis. Previous evidence suggests that tissue transglutaminase (tTg) and its protein cross-link product ⑀(␥-glutamyl)lysine contribute to the expanding renal tubulointerstitial and glomerular basement membranes in this disease. Using an in vitro cell culture model of renal proximal tubular epithelial cells we determined the link between elevated glucose levels with changes in expression and activity of tTg and then, by using a highly specific site directed inhibitor of tTg (1,3-dimethyl-2[(oxopropyl)thio]imidazolium), determined the contribution of tTg to glucose-induced matrix accumulation. Exposure of cells to 36 mM glucose over 96 h caused an mRNA-dependent increase in tTg activity with a 25% increase in extracellular matrix (ECM)-associated tTg and a 150% increase in ECM ⑀(␥-glutamyl)lysine cross-linking. This was paralleled by an elevation in total deposited ECM resulting from higher levels of deposited collagen and fibronectin. These were associated with raised mRNA for collagens III, IV, and fibronectin. The specific site-directed inhibitor of tTg normalized both tTg activity and ECMassociated ⑀(␥-glutamyl)lysine. Levels of ECM per cell returned to near control levels with non-transcriptional reductions in deposited collagen and fibronectin. No changes in transforming growth factor ␤1 (expression or biological activity) occurred that could account for our observations, whereas incubation of tTg with collagen III indicated that cross-linking could directly increase the rate of collagen fibril/gel formation. We conclude that Tg inhibition reduces glucose-induced deposition of ECM proteins independently of changes in ECM and transforming growth factor ␤1 synthesis thus opening up its possible application in the treatment other fibrotic and scarring diseases where tTg has been implicated.

Fibronectin-Tissue Transglutaminase Matrix Rescues RGD-impaired Cell Adhesion through Syndecan-4 and β1 Integrin Co-signaling

“…TG2 is translocated to the plasma membrane and was subsequently deposited into the ECM via a non-classical secretory mechanism reportedly dependent on active site conformation and on an intact N-terminal ␤-sandwich domain (4, 5) as well as on its possible association with integrins (6). Deposition of the enzyme into the ECM after cell damage and stress is important in the remodeling and/or stabilization of the several ECM proteins, such as FN (7,8). FN is particularly interesting since TG2 binds to this ECM protein with high affinity promoting wideranging effects on cell-matrix interactions, including the regulation of cell adhesion and migration, matrix assembly, and adhesion-dependent signaling (6,7,9).…”

Transglutaminase 2 is involved in autophagosome maturation

“…4B). 14 It is important to note that R283 specifically inhibits the transglutaminases cross-linking activity.…”

Localization of Tissue Transglutaminase (tTG) in Kidney of ICR-Derived Glomerulonephritis (ICGN) Mice

“…The involvement of tTG is known not only in experimental fibrotic models [15,17,25] but also in human patients [10,27]. In renal biopsy samples from CKD patients, insoluble tTG showed a 14-fold increase and e(g-glutamyl) lysine cross-linking showed an 11-fold increase in contrast to normal renal tissues.…”

“…Also, it inhibits ECM breakdown by conferring resistance to matrix metalloproteinase [18]. In experimental diabetic [25] and non-diabetic renal scarring [15,17], tTG has been proposed as a potential mediator of ECM accumulation. Furthermore, changes in tTg levels have been seen in other types of fibrotic lesions including those affecting the lung [20], liver [7,10], heart [26], and articular cartilage [27].…”