R. Singh et al., High glucose decreases matrix metalloproteinase-2 activity in rat mesangial cells via transforming growth factor-beta(1), EXP NEPHROL, 9(4), 2001, pp. 249-257
Diabetic nephropathy is characterized by accumulation of mesangial matrix.
Glucose-induced inhibition of matrix-degrading enzymes such as collagenases
is believed to contribute to matrix accumulation. We have previously demon
strated that 72 kDa type IV collagenase activity is decreased in the rat me
sangial cells cultured in high glucose media [Diabetes 1995;44:929-935]. Th
e present studies were designed to investigate if the cytokine transforming
growth factor-beta (1) (TGF-beta (1)) mediates this effect of glucose. Typ
e IV collagenases degrade type IV collagen as well as gelatin (denatured co
llagen) and are thus also called gelatinases. They belong to the family of
matrix metalloproteinases (MMPs); MMP activity is controlled by tissue inhi
bitors of metalloproteinases (TIMPs). The activity of 72 kDa type IV collag
enase, also known as matrix metalloproteinase-2 (MMP-2), was assessed using
three methods: (I)fluoresceinated gelatin degradation assay to detect free
enzyme activity (activity which is present in excess of TIMP-inhibited act
ivity); (2) zymography to measure total (free + TIMP-bound) enzyme activity
; (3) ELISA using specific antibodies to measure MMP-2 levels. TGF-beta (1)
and TIMP-2 levels were also determined by ELISA. Incubation of primary cul
tures of rat mesangial cells for 5 days in 30 vs. 5 mM glucose resulted in
a 3-fold increase in production of total TGF-beta (1), a significant decrea
se in MMP-2 activity and immunoreactive MMP-2 levels, and an increase in TI
MP-2 levels. Addition of exogenous TGF-beta (1) to mesangial cells incubate
d in 5 mM glucose replicated the high glucose effect by producing a signifi
cant decrease in MMP-2 levels with a concurrent increase in TIMP-2 levels.
Furthermore, glucose-induced inhibition of MMP-2 activity was completely bl
ocked by neutralization of TGF-beta (1) with anti-TGF-beta (1) antibody. We
conclude that the decrease in MMP-2 activity induced by glucose loading is
mediated via TGF-beta (1). Copyright (R) 2001 S. Karger AG, Basel.