Magnetic resonance imaging investigation of axonal remodeling and angiogenesis after embolic stroke in sildenafil-treated rats

Abstract

Interaction between angiogenesis and axonal remodeling after stroke was dynamically investigated by MRI in rats with or without sildenafil treatments. Male Wistar rats were subjected to embolic stroke and treated daily with saline (n=10) or with sildenafil (n=11) initiated at 24 h and subsequently for 7 days after onset of ischemia. T(2)(*)-weighted imaging, cerebral blood flow (CBF), and diffusion tensor imaging (DTI) measurements were performed from 24 h to 6 weeks after embolization. T(2)(*) and fractional anisotropy (FA) maps detected angiogenesis and axonal remodeling after stroke, respectively, starting from 1 week in sildenafil-treated rats. Areas demarcated by MRI with enhanced angiogenesis, elevated local CBF, and augmented axonal remodeling were spatially and temporally matched, and FA values were significantly correlated with the corresponding CBF values (R=0.66, P<4 x 10(-5)) at 6 weeks after stroke. Axonal projections were reorganized along the ischemic boundary after stroke. These MRI measurements, confirmed by histology, showed that sildenafil treatment simultaneously enhanced angiogenesis and axonal remodeling, which were MRI detectable starting from 1 week after stroke in rats. The spatial and temporal consistency of MRI metrics and the correlation between FA and local CBF suggest that angiogenesis, by elevating local CBF, promotes axonal remodeling after stroke.

Figures

Figure 1

T2 map acquired at 24 h after stroke (A) demarcated acute ischemic lesion area with values above mean plus two s.d. of contralateral measurements. (B) Final infarction area was determined by T2 map acquired at 6 weeks after stroke. (C) The difference was referred as recovery area after stroke.

Figure 2

With the ROI identified hyperintensity in FA map acquired at 6 weeks after stroke, the group averaged ratios, measurements from the ROI to its contralateral ROI, showed the temporal features of T2∗ (A), CBF (B), and FA (C) in stroke rats with or without treatments. Compared with control rats, sildenafil-treated rats had relative lower T2∗ values, higher CBF and FA values within the 6 weeks after stroke, although similar values at 1 day after stroke. The differences were significant (P < 0.05) during 1 to 4 weeks for T2∗, at 6 weeks for CBF, or at 2, 5, and 6 weeks for FA measurements. Error bars in panels A to C are presented as standard errors. The relative CBF and FA values, individually measured at 6 weeks after stroke in both ischemic boundary and core, exhibited a linear correlation (P <4 × 10−5) with a correlation coefficient of R = 0.66 (D).

Figure 3

In a representative sildenafil-treated rat, T2∗ (A), CBF (B), and FA (C) maps tracked angiogenesis via lower T2∗ values, recovered blood supply with hyperintensity in CBF, and axonal remodeling by higher FA values after stroke, respectively. Red arrows in A2, B2, and C2 indicated that angiogenesis, CBF recovery, and axonal remodeling simultaneously occurred starting from 1 week after stroke, respectively. At 6 weeks after stroke, the low T2∗ value area (D), elevated CBF area (E), and increased FA value area (F) were outlined in individual maps and projected onto a T2 map (G). Fiber tracking by DTI in the treated (H) and in a control (I) rats indicated that the axons were reorganized along ischemic boundary, which were coincident with FA maps of the treated (C7) and the control (J) rats.

Figure 4

With EBA (A) and B&LFB (D) sections stained at 6 weeks after stroke, microvascular (B) and axonal (E) density at 6 weeks after stroke in the angiogenic area demarcated with lower T2∗ values and axonal remodeling area characterized by higher FA values along ischemic boundary were higher than those in the homologous tissue of contralateral hemisphere (C and F), respectively. The nerve fibers after stroke were reorganized along the rim of ischemic boundary in both treated (G) and control (H) rats, as indicated by red arrows. Axonal density along ischemic boundary in the control rat (I) was increased compared with normal tissue, but less than in the treated rat. Scale bar = 100 μm (B).