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Fig. 2. NO/cGMP increase mesoangioblast migration and engrafting in vivo. (A,B) GFP-expressing mesoangioblasts (5x105) pretreated for 12 hours without (NT) or with DETA-NO (20 µM), IMN (50 µM), 8-Br-cGMP (1 mM) or DETA-NO plus ODQ (1 µM) were injected into the right femoral artery of 4-month-old 
SG-null mice. After 6 hours quadriceps (Qd), tibialis anterior (TA) and gastrocnemius (Gs) muscles, as well as liver, spleen and lung were collected and the number of mesoangioblasts migrated into them calculated by quantitative real-time PCR for GFP. Values are expressed as the percentage of the total injected cells (n=3). (C) GFP-expressing mesoangioblasts (5x105) pretreated as indicated above were injected into the tibialis anterior muscles of 4-month-old SG-null mice. After 12 hours muscles were recovered. Apoptosis of GFP-expressing mesoangioblasts was assessed by the TUNEL technique (images from one out of three reproducible experiments). Also shown is the DAPI staining of the nuclei and its overlay with the GFP and TUNEL staining. Arrows in the merge panels show selected GFP, TUNEL and DAPI-positive cells. The red arrow indicates the specific cell for which the GFP, TUNEL, DAPI and merge stainings are magnified in the insets. (D,E) Same conditions as in C, except that mice were sacrificed three weeks later, tibialis muscles were recovered and SG expression was analysed by real-time PCR (D, n=3) and western blot analysis (E, showing both a representative western blot image and densitometric values, n=3). The graph in E reports the ratio of densitometric values of SG vs. those of GAPDH used as an internal loading control. Triple asterisks and crosses, P<0.001 vs. NT and DETA-NO-treated cells, respectively; error bars in A, B, D and E, s.e.m.). Bar in C, 400 µm. The inset in the panel is at 6x magnification.
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