To evaluate the long-term effectiveness of traumatic brain injury (TBI) treatment in relation to regional cerebral blood flow (rCBF) in angiogenesis and tissue loss using magnetic resonance (MR) phase gradient mapping (PGM).

The fractional change in blood flow through a vessel can be written as$$f=\frac{\mp R_{pc}}{1-R_{pc}}$$where the relative phase change$$R_{pc}=\frac{\pm (\triangle \phi_{1}-\triangle \phi_{0})}{\triangle \phi_{0}}$$with Δφ₀ and Δφ₁ representing the change in phase between the vessel and the surrounding tissue at two different time points¹. The relative phase change can theoretically be calculated using just the phase difference within the vessels at the two time points¹; however, variations in the MRI scanning procedure (coil tuning, gradient shimming, etc.) can cause additional changes in the phase. By using the change in phase between the vessel and the surrounding tissue, these variations are removed¹. To avoid MRI phase wrapping effects, the change in the phase gradient can be used to calculate the fractional change in rCBF since it’s proportional to the change in phase. The equation$$\triangledown_{x}\phi=\triangledown_{x}P\approx\frac{\triangle P}{\triangle x}$$illustrates this relationship, where φ denotes the wrapped phase and P denotes the real phase². The phase gradient can be calculated using$$\triangledown_{x}\phi=\cos\phi*FT^{-1}[k_{x}*FT(\sin\phi)]-\sin\phi*FT^{-1}[k_{x}*FT(\cos\phi)]$$where FT and FT^-1 refer to the forward and inverse Fourier Transforms and $$k_{x}=\frac{i*\sin(\frac{2\pi l}{L_{x}})}{\triangle x}$$with L_x being the number of voxels in the x-direction and Δx being the voxel size³.

TBIs were performed on six rats using a custom designed controlled cortical impactor⁴. The left fronto-parietal cortex was impacted using a 3mm diameter flat tip driven at a velocity of 2.25m/s, to a depth of 2mm, and for a dwell-time of 250ms after performing a 5mm unilateral craniotomy 0.5mm from the bregma and the lateral suture. Two additional rats received craniotomies, but not TBIs. These rats served as controls. Two days post-TBI, one rat received an injection of chondroitin sulfate glycosaminoglycan hydrogel with neural stem cells and trophic factors (GAG-NSCs-TF), and two rats received the same injection without NSCs (GAG-TF). To evaluate long-term effects, twenty weeks post TBI all rats (2 controls, 3 non-treatments, 3 treatments) were scanned using a MGEMS pulse sequence on a 7T Varian Magnex MRI scanner with parameters: TR=600ms, 8 echoes (TE=10-45ms in 5ms increments), FA=25°, FOV=40x40mm, and 8 slices with thickness=1mm centered on the injury. Phase gradient maps (PGMs) were calculated for each slice in both the readout and phase encoding directions using equations 4 and 5. From these PGMs, phase gradient differences between a vessel and the surrounding tissue were measured across all distinguishable vessels located within the cortex, sub-cortical and basal forebrain areas, and thalamus. It was determined that TE=20ms produced the best contrast for distinguishing vessels within the PGMs, so the third echo was used for all measurements (see Figure 1). Since direct comparisons of the same vessels from the same rats were not made, phase variations in the MRI scanning procedure aren’t automatically removed using equation 2. To account for these variations, measurements were normalized using the phase gradient difference across the third ventricle for each rat. Averages and standard deviations of all normalized phase gradient differences were calculated for both the contralateral and ipsilateral side of the brain to the injury for each region and group, and fractional changes in rCBF between groups were determined using equations 1 and 2.The treatment group consistently showed increases in rCBF compared to the control group in all regions on both the contralateral and ipsilateral sides of the brain, while the non-treatment group consistently showed decreases in rCBF (see Figure 2). The increases in rCBF in the treatment group suggest that angiogenesis is occurring and that tissue loss has been stunted⁵. The decreases in rCBF in the non-treatment group suggest that angiogenesis is not occurring, so greater tissue loss is expected. Brain dissection confirmed increased tissue loss in the non-treatment group compared to the treatment group (see Figure 3). Variations between animals of the same group and the small sample size of each group (n=2 or 3) likely contribute to the significant variations in the changes in rCBF in Figure 2. Further correlation with global CBF measured by arterial spin labeling will be conducted, and histology and staining will be performed to look for angiogenesis. To summarize, changes in rCBF in a rat TBI model were calculated using phase gradient differences measured across vessels within different areas of the rat brain. Rats that received GAG-NSCs-TF or GAG-TF showed increases in rCBF compared to the control group, while rats that did not receive treatment showed decreases in rCBF. Comparing these results with dissection suggests that an increase in CBF relates to a greater effectiveness of treatment concerning reduction in tissue loss and possibly increased angiogenesis.