In acute stroke patients within 24 hours after symptom onset, we found a significant leakage of the BBB starting at 6 hours after stroke onset. Cerebrovascular risk factors, white matter hyperintensities expressed as Wahlund score13, age or sex showed no association with BBB permeability.
Continuous increase of BBB permeability was described by Strbian et al.2 in rats as well as by Merali et al. in humans7. Strbian et al2 reported the earliest BBB opening 25 minutes after 90 minutes of ischemia-reperfusion up until 5 weeks with 2, 4, 6, 12, 18, 24, 36, 48, 72 hours and 1, 2, 3, 4 weeks follow-up examinations in between. The authors employed two contrast agents: Gd-DTPA (MW 590 Da, Magnevist, Schering, German) for T1 measurements in-vivo and EBA post-mortem (fluorescent dye, Sigma, MW 75.8 kDa, Sigma-Aldrich, Steinheim, Germany). The most pronounced BBB leakage for Gd-DTPA was found between 48h up until 2 weeks after ischemia most pronounced at 6 and 72 hours as well as 2 weeks after reperfusion. For EBA the most pronounced BBB leakage was found at 6 hours as well as 36 - 48 hours after reperfusion followed by a slow decline until the third week. Interestingly, for both markers of BBB leakage the first peak was observed at 6 hours after ischemia despite their different molecular weights. We found a significant BBB leakage at 6-16 hours after stroke onset which coincide with Strbian et al´s2 results in rats. However, we did not find a significant earlier BBB leakage which might be due to the different species, experimental settings with a 4.7 T MR scanner and a bird cage RF coil used in the study by Strbian et al.2 in contrast to our study with a 3 T MR scanner and a 64 channel head coil. Higher field strength might have an influence on the detection of BBB leakage10.
In one of our recent studies14, we also found a more than 100% increase in BBB leakage between the first BBB assessment within 48 hours and the follow-up examination between day 5-7 after symptom onset, supporting the observation of Strbian et al.2, indicating a continuous, however fluctuating increase in BBB permeability.
Merali et al.7 also found a continuous increase in BBB permeability, again most pronounced between 6 to 48 hours after stroke onset, but also in the hyperacute phase, that is within the first 6 hours after ischemia. The authors though, used the parameter permeability-surface area product (PS) instead of Ktrans , which only corresponds to Ktrans under very specific conditions. Only if cerebral blood flow considerably exceeds PS does Ktrans approximately equal PS15. Compared to Merali et al.7 with 20 patients in the hyperacute phase we only had 10 patients and used a different software package for the Patlak analysis, which might contribute to the difference in the results. Furthermore, BBB leakage is not only restricted to the stroke region but was also found in contralesional brain tissue14 in the acute phase of stroke which we found in 6/10 patients in the first group. This might also contribute to the lack of significant difference in BBB permeability in the first 6 hours after symptom onset. When contralesional BBB leakage recovers, as has been reported14, differences in BBB leakage between stroke and contralesional tissue become more pronounced.
As already pointed out, differences in equipment such as MR scanner field strength, coil configuration, duration of ischemia, tracers, experimental design, software packages and kinetic models impact the quantification of BBB leakage2,10, in particular with respect to a probable second BBB opening.
When applying Patlak analysis, which is used in most of the studies, Manning et al.16 found factors yielding erroneous results when assessing BBB permeability. These include the rate of injection of contrast agent (CA). This can result in fast BBB water exchange and thereby in a substantial bias of the permeability surface product and occurs during the early part of the time course of the contrast agent application. A slow injection is less sensitive to plasma flow, since the slower exchange in Gadolinium concentration results in similar arterial, capillary and venous concentrations. Excluding early data points resolves the problem of the fast BBB water exchange, reduces blood flow effects and the sensitivity to plasma flow. Furthermore, a small delay for the bolus injections has significant impact on the estimated parameters and significant errors remain even when early data points are excluded, while slow injection of Gadolinium virtually eliminates the problem. T1 saturation and transverse dephasing caused by the high concentration of a bolus injection can be avoided as well16. Since we employed a slow injection protocol and started the analysis using the very first time points of signal intensity increase we could address these factors mentioned above to minimize bias in our analysis.
Complicating the reported results is the relative lack of longitudinal studies of BBB leakage. One such study was performed by Durukan et al. in rats6 at five time points after reperfusion (2, 24, 48, 72 hours and 1 week) and showed persistent BBB leakage during the whole week. Another longitudinal study by our group in acute stroke patients within 48 hours after stroke onset showed a continuous increase in BBB leakage at 24-hour follow-up14. Follow-up studies in humans are complicated by the repetitive application of Gadolinium with its well-known potential side effects of nephrogenic systemic fibrosis17 and evidence of deposition in certain brain regions18,19.
As previously reported14, we found no association between Ktrans values and HARM sign, another reported marker of BBB disruption, which was observed in 21 out of 53 of patients in this current study. This can be explained by a different pathophysiological mechanism resulting in HARM sign in contrast to an increased Ktrans. Additional factors influencing HARM sign are infarct size (median volume size 0.73, IQR 0.35-1.89 mL), which showed a large variability in our study cohort, as well as risk factors such as hypertension and diabetes20. Both were reported to have an impact on HARM sign and showed no association with Ktrans values in our study using DCE-MRI, again probably due to the two different techniques applied. We also did not find an association between Ktrans and hemorrhagic transformation, but the number of patients was too small (n=3) for any satisfactory analysis and conclusion.
Our study has some limitations, foremost the small number of patients assigned to each time between symptom onset and imaging category. This was largely due to the fact that many patients who were eligible for mechanical thrombectomy did not receive a DCE-MRI scan, as this would have led to unjustified delays in treatment. This also might have introduced a bias with respect to symptom severity in our cohort, which was clearly skewed towards minor strokes as is reflected in the NIHSS score.
Secondly, the patient sample was very heterogeneous regarding stroke location with infra- as well supratentorial strokes as well as pronounced differences in lesion volumes. The latter might also be contributing to the missing evidence of detectable BBB leakage in the hyperacute stage.
Thirdly, the impact of various risk factors on BBB leakage must be interpreted with caution due to the small number of patients. This is despite the fact that the distribution of the number of risk factors was comparable across the different time groups.
A different and novel promising technique is currently emerging for assessing BBB leakage non-invasively: DP-pCASL (diffusion prepared pseudo-continuous arterial spin labeling) using the water exchange rate across the BBB as a means to describe BBB leakage. Shao et al.21 found a good agreement analyzing BBB leakage using DCE-MRI and DP-pCASL as well as high test-retest reproducibility measuring water exchange rate immediately after and about 6 weeks later in elderly participants with cerebral small vessel disease. Using this technique with an endogeneous contrast agent that is water, more insight into the time course of BBB permeability under various conditions may be obtained.