In vivo quantification of transvascular water exchange during the acute phase of permanent stroke.

After an ischemic insult, the affected cerebral tissue undergoes a series of changes associated with initially reversible damages that eventually evolve into irreversible tissue damage (1). Biophysical alterations due to ischemic injury include cell swelling, reduced water diffusion, disruption of the blood-brain-barrier (BBB), and loss of cell membrane integrity (2). Almost immediately after ischemic attack, cytotoxic edema, characterized by cellular swelling occurs, leading to an increase in intracellular fluid and a concomitant reduction in extracellular/extravascular water content. The primary mechanism of such changes is believed to a decrease in the intracellular ATP that drives the sodium–potassium pump, and a resultant increase in intracellular sodium concentration, which together create an osmotic pressure gradient. The early increase in cellular volume has been frequently documented, as have changes in both intra- and extracellular water diffusion properties (3,4). Infarcted tissue in acute ischemia typically exhibits restricted water diffusion (i.e., apparent diffusion coefficient: ADC) when examined with diffusion-weighted magnetic resonance imaging (DWI) (5). At later phases, vasogenic edema, associated with local disruption of the blood-brain-barrier (BBB), follows the cytotoxic edema (1). The dysfunctional BBB leads to the extravasation of protein-rich plasma, with subsequent accumulation of vascular fluid in the interstitial space. BBB disruption arises when tight junctions between endothelial cells disengage, during which the neoformation and enhanced activity of pinocytic vesicles are frequently observed (6). Both hydrostatic and osmotic forces created by blood perfusion and the interstitial accumulation of various biological molecules induce the extravasation of intravascular fluid (7). Because vasogenic edema accompanies the extravasation of protein, spatial and temporal leakage profiles of staining dye or various imaging contrast agents have been used to identify the degree of vasogenic dysfunction. Along these same lines, various strategies have been used to evaluate the vascular permeability. However, such techniques have been highly limited by the fact that measurement is only possible when the damaged BBB allows the extravasation of extrinsic markers (e.g., Gd-DTPA) (8,9). Therefore, given that extrinsic markers usually are not capable of crossing the BBB at acute stages of stroke, it has not been feasible to observe the progressive weakening of the BBB. We described here a novel way to quantify acute BBB leakage in cerebral ischemia by using intrinsic water molecules as a measurement marker. Recent evidence has revealed that the expression of perivascular aquaporin proteins (e.g., AQP4) that form water channels may be involved in water movement across the BBB interface and are disturbed as early as 1 hour after cerebral ischemia (10,11). We posit that water movement regulated by perivascular water channel proteins is probably impaired and/or dysfunctional during stroke progression and can be used as a measurement target of interest. To monitor early BBB impairment, we propose an MRI technique based on the difference in the longitudinal relaxation rate (i.e., 1/T1) between the intra- and extravascular spaces. A difference of T1 relaxation rates can be created by intravenous administration of a strictly intravascular long circulating contrast agent. Then, the MRI signal is affected by the water exchange between tissue compartments that consist of intra- and extravascular spaces. The effects of water exchange on MRI signal intensity are also dependent on the MRI pulse sequence and pulse sequence parameters as well as compartmental sizes. Our measurement technique exploits the dependence of the MRI signal intensity acquired with 3D SPGR on the flip angle (α) to identify the disrupted BBB integrity. We demonstrated that this relatively simple MRI technique provides a non-invasive means for understanding and quantifying the water movement across the BBB interface. Water molecules that traverse the vascular membrane offer a unique opportunity not only for observing the presence of vascular damage at early stages of stroke but also for understanding how such impairment evolves into a serious vascular dysfunction at later stages. Previously, we reported that the transvascular water movement (i.e., between intra- and extravascular spaces) in rat brains can be quantified by using a widely available MRI pulse sequence (i.e., 3D SPGR) and an intravascular contrast agent (i.e., Gd-PGC) (12,13). We have also shown that the degree of intra-/extravascular water movement (i.e., transvascular water exchange) reflects the integrity of the BBB and that the transvascular water exchange is elevated in brain tumor vasculature (12). In the current study, we investigated time-dependent hyper-acute changes of various biophysical parameters in a mouse stroke model of permanent middle cerebral artery occlusion (MCAo). Specifically, we hypothesized that vascular integrity is distinctively altered within an hour of ischemic onset, concurrently accompanying cytotoxic edema. We performed two-compartment model simulations to optimize MRI parameters and the contrast agent dose to achieve measurement accuracy. Furthermore, adjustment of the measured water exchange index (WEI) was made, considering time-dependent biophysical changes such as reduced diffusion and cellular swelling that may affect the measurement accuracy. By directly examining the movements of water molecules through the vascular membrane, our goal was to devise a convenient and practical methodology for assessing BBB damage during stroke progression without acquiring the leakage profile of extrinsic molecules into the interstitial space, and thereby, to provide a possible predictor of ensuing vasogenic events.