Figure 1

Preferential induction of MMP12 is causal for arterial stiffening after vascular injury.

Male wild-type and MMP12-null mice were subjected to femoral artery injury. (A) Differential expression of MMP mRNAs after femoral artery injury in 4–5 month old wild-type mice. (B) Uninjured (n = 6) and injured (n = 4) femoral arteries from wild-type and MMP12-null mice were isolated, cleaned, and immediately analyzed by AFM. The bar graph shows mean + SE. (C) Cyclin D1 staining of uninjured and injured femoral artery sections. Dashed lines show the internal elastic lamina (IEL) and external elastic laminae (EEL). M; media. NI; neointima. Scale bar = 50 μm. (D) Blind quantification of results in C from wild-type (n = 10) and MMP12-null (n = 8) mice. Statistical significance was determined by chi-square test. (E) In situ elastase activity of cross sections of uninjured and injured femoral arteries. Scale bar = 50 μm. (F) Quantification of results in panel E from wild-type (n = 7) and MMP12-null (n = 5) mice. The bar graph shows mean + SE. (G) Elastin fragmentation in the IEL was detected by autofluorescence imaging (green). Scale bar = 25 μm. Nuclei (blue) were stained with DAPI. (H) Quantification of results in panel G from wild-type (n = 10) and MMP12-null (n = 8) mice. The bar graph shows mean + SE. (I) Cross sections of uninjured and injured femoral arteries of wild-type mice immunostained for MMP12 (red) and SMCs (anti-SMA, green) (n = 8). Nuclei (blue) were stained with DAPI. Dashed lines show the IEL and EEL as determined by autofluorescence. (J) Differentiated and de-differentiated SMCs were serum-starved and incubated with 1 mg/ml heat-inactivated, fatty acid-free BSA in the absence (control) or presence of PDGF for 24 h. MMP12 mRNA levels were determined by RT-qPCR and plotted relative to 18S rRNA. The bar graph shows mean + SD; n = 4. The levels of MMP12 mRNA in the absence of PDGF was set to 1.0. All p values are from two-tailed Mann-Whitney tests.