A. MMP14 protein expression in aging kidney. Expression of MMP-14 is increased in aging mice kidneys in a time-dependent manner (FVB mouse strain). n=5 for 1-month-old, n=2 for 10 month-old, and n=7 for 15 month-old mice.
B. Western blot analysis of TG2 fragmentation: TG2 fragmentation by MMP14. TG2 from guinea pig liver (100 ng) was incubated for 0, 1, and 16 hours (lines 1, 2, and 3, respectively) in serum-free conditioned medium collected from HEK293T cells that were transfected with pcDNA-ΔMT1 construct containing the extracellular domain of human MMP14. The products were blotted and detected with monoclonal TG2 antibody. Note that the band intensity of the smaller fragments (53kDa and 22 kDa) increases along with the duration of incubation. Lane 4 represents conditioned medium (CM) (10 ul) that was blotted with monoclonal MMP14 antibody.
C. The ~53kDa fragment is absent in TG2−/− mice. Monoclonal anti-TG2 antibodies detecting multiple bands on WB in B. ~53 kDa fragment of TG2 were undetectable in the TG2−/− kidneys. n=3 animals per group.