Matthew D. Silva

Transient and Permanent Resolution of Ischemic Lesions on Diffusion-Weighted Imaging After Brief Periods of Focal Ischemia in Rats Correlation With Histopathology

BACKGROUND AND PURPOSE The early ischemic lesions demonstrated by diffusion-weighted imaging (DWI) are potentially reversible. The purposes of this study were to determine whether resolution of initial DWI lesions is transient or permanent after different brief periods of focal brain ischemia and to evaluate histological outcomes. METHODS Sixteen rats were subjected to 10 minutes (n=7) or 30 minutes (n=7) of temporary middle cerebral artery occlusion or sham operation (n=2). DWI, perfusion-weighted imaging (PWI), and T(2)-weighted imaging (T(2)WI) were performed during occlusion; immediately after reperfusion; and at 0.5, 1.0, 1.5, 12, 24, 48, and 72 hours after reperfusion. After the last MRI study, the brains were fixed, sectioned, stained with hematoxylin and eosin, and evaluated for neuronal necrosis. RESULTS No MRI or histological abnormalities were observed in the sham-operated rats. In both the 10-minute and 30-minute groups, the perfusion deficits and DWI hyperintensities that occurred during occlusion disappeared shortly after reperfusion. The DWI, PWI, and T(2)WI results remained normal thereafter in the 10-minute group, whereas secondary DWI hyperintensity and T(2)WI abnormalities developed at the 12-hour observation point in the 30-minute group. Histological examinations demonstrated neuronal necrosis in both groups, but the number of necrotic neurons was significantly higher in the 30-minute group (95+/-4%) than in the 10-minute group (17+/-10%, P<0.0001). CONCLUSIONS Transient or permanent resolution of initial DWI lesions depends on the duration of ischemia. Transient resolution of DWI lesions is associated with widespread neuronal necrosis; moreover, permanent resolution of DWI lesions does not necessarily indicate complete salvage of brain tissue from ischemic injury.

Temporal evolution of ischemic injury evaluated with diffusion-, perfusion-, and T2-weighted MRI.

Objective: Ischemic lesions seen on diffusion-weighted imaging (DWI) are reversible if reperfusion is performed within minutes after the onset of ischemia. This study was designed to determine whether acute reversibility of DWI abnormalities is transient following brief temporary focal brain ischemia and to characterize the temporal evolution of in vivo ischemic lesions. Methods: Eight rats were subjected to 30 minutes of temporary middle cerebral artery occlusion and underwent diffusion-, perfusion-, and T2-weighted MRI during occlusion; immediately after reperfusion; 30, 60, and 90 minutes after reperfusion; and 12, 24, 48, and 72 hours after reperfusion. Average apparent diffusion coefficient (ADCav) values and the cerebral blood flow index (CBFi) ratio were calculated in both the lateral caudoputamen and overlying cortex at each time point. The size of the in vivo ischemic abnormalities was calculated from the ADCav and the T2 maps. Postmortem triphenyltetrazolium chloride (TTC) staining was used to verify ischemic injury. Results: Both the CBFi ratio and ADCav values declined significantly in the two regions during occlusion. The CBFi ratio recovered immediately after reperfusion and remained unchanged over 72 hours. However, ADCav values returned to normal at 60 to 90 minutes and secondarily decreased at 12 hours after reperfusion as compared with those in the contralateral hemisphere. The extent of the in vivo ischemic lesions maximized at 48 hours and was highly correlated with TTC-derived lesion size. Conclusions: Acute recovery of initial ADCav-defined lesions after reperfusion is transient, and secondary ADCav-defined lesions develop in a slow and delayed fashion.

The macrosphere model: evaluation of a new stroke model for permanent middle cerebral artery occlusion in rats

BACKGROUND AND PURPOSE The suture middle cerebral artery occlusion (MCAO) model is widely used for the simulation of focal cerebral ischemia in rats. This technique causes hypothalamic injury resulting in hyperthermia, which can worsen outcome and obscure neuroprotective effects. Herein, we introduce a new MCAO model that avoids these disadvantages. METHODS Permanent MCAO was performed by intraarterial embolization using six TiO(2) macrospheres (0.3-0.4 mm in diameter) or by the suture occlusion technique. Body temperature was monitored, functional and histologic outcome was assessed after 24 h. Additional 16 rats were subjected to macrosphere or suture MCAO. Lesion progression was evaluated using magnetic resonance imaging (MRI). RESULTS The animals subjected to suture MCAO developed hyperthermia (>39 degrees C), while the temperature remained normal in the macrosphere MCAO group. Infarct size, functional outcome and model failure rate were not significantly different between the groups. Lesion size on MRI increased within the first 90 min and remained unchanged thereafter in both groups. CONCLUSIONS The macrosphere MCAO model provides reproducible focal cerebral ischemia, similar to the established suture technique, but avoids hypothalamic damage and hyperthermia. This model, therefore, may be more appropriate for the preclinical evaluation of neuroprotective therapies and can also be used for stroke studies under difficult conditions, e.g., in awake animals or inside the MRI scanner.

Separating changes in the intra- and extracellular water apparent diffusion coefficient following focal cerebral ischemia in the rat brain.

Selective intracellular (IC) and extracellular (EC) brain water apparent diffusion coefficient (ADC) values were measured in normal and ischemic rat brain. Selective T1‐relaxation enhancement of the EC water, using intracerebroventricular (ICV) infusion of an NMR contrast reagent (CR), was used to separate the IC and EC signal contributions. In the CR‐infused, normal brain (n = 4), T1 = 235 ± 10 ms and T2 = 46 ± 2 ms for IC water (85%) and T1 = 48 ± 8 ms and T2 = 6 ± 2 ms for EC water (15%). Volume‐localized ADCz (z‐gradient axis) values were 0.90 ± 0.02 (EC+IC), 0.81 ± 0.05 (IC), 0.51 ± 0.02 (EC+IC), and 0.53 ± 0.07 (IC), for normal, CR‐infused, ischemic, and ischemic/CR‐infused groups, respectively (ADC values are ×10‐3 mm2/s; n = 5 for each group). Imaging ADCz values were 0.81 ± 0.03 (EC+IC), 0.75 ± 0.05 (IC), 0.51 ± 0.04 (EC+IC), and 0.52 ± 0.05 (IC), respectively, for the same groups. Imaging ADCav (average diffusivity) values for the same groups were 0.70 ± 0.05 (EC+IC), 0.69 ± 0.06 (IC), 0.45 ± 0.06 (EC+IC), and 0.44 ± 0.06 (IC), respectively. These results suggest that the IC water ADC determines the overall water ADC value in normal and ischemic rat brain. Magn Reson Med 48:826–837, 2002.

Secondary decline in apparent diffusion coefficient and neurological outcomes after a short period of focal brain ischemia in rats

This study was designed to characterize the initial and secondary changes of the apparent diffusion coefficient (ADC) of water with high temporal resolution measurements of ADC values and to correlate ADC changes with functional outcomes. Fourteen rats underwent 30 minutes of temporary middle cerebral artery occlusion (MCAO). Diffusion‐, perfusion‐, and T2‐weighted imaging was performed during MCAO and every 30 minutes for a total of 12 hours after reperfusion (n = 6). Neurological outcomes were evaluated during MCAO, every 30 minutes for a total of 6 hours and at 24 hours after reperfusion (n = 8). The decreased cerebral blood flow during MCAO returned to normal after reperfusion and remained unchanged thereafter. The decreased ADC values during occlusion completely recovered at 1 hour after reperfusion. The renormalized ADC values started to decrease secondarily at 2.5 hours, accompanied by a delayed increase in T2 values. The ADC‐defined secondary lesion grew over time and was 52% of the ADC‐defined initial lesion at 12 hours. Histological evaluation demonstrated neuronal damage in the regions of secondary ADC decline. Complete resolution of neurological deficits was seen in 1 rat at 1 hour and in 6 rats between 2.5 and 6 hours after reperfusion; no secondary neurological deficits were observed at 24 hours. These data suggest that (1) a secondary ADC reduction occurs as early as 2.5 hours after reperfusion, evolves in a slow fashion, and is associated with neuronal injury; and (2) renormalization and secondary decline in ADC are not associated with neurological recovery and worsening, respectively. Ann Neurol 2000;48:236–244

Multispectral analysis of the temporal evolution of cerebral ischemia in the rat brain

A major difficulty in staging and predicting ischemic brain injury by magnetic resonance (MR) imaging is the time‐varying nature of the MR parameters within the ischemic lesion. A new multispectral (MS) approach is described to characterize cerebral ischemia in a time‐independent fashion. MS analysis of five MR parameters (mean diffusivity, diffusion anisotropy, T2, proton density, and perfusion) was employed to characterize the progression of ischemic lesion in the rat brain following 60 minutes of transient focal ischemia. k‐Means (KM) and fuzzy c‐means (FCM) classification methods were employed to define the acute and subacute ischemic lesion. KM produced an estimate of lesion volume that was highly correlated with postmortem infarct volume, independent of the age of the lesion. Overall classification rates for KM exceeded FCM at acute and subacute time points as follows: KM, 90.5%, 94.4%, and 95.9%; FCM, 82.4%, 90.6%, and 82.6% (for 45 minutes, 180 minutes, and 24–120 hours post MCAO groups). MS analysis also offers a formal method of combining diffusion and perfusion parameters to provide an estimate of the ischemic penumbra (KM classification rate = 70.3%). J. Magn. Reson. Imaging 2000;12:842–858.

Transient and Permanent Resolution of Ischemic Lesions on Diffusion-Weighted Imaging After Brief Periods of Focal Ischemia in Rats

BACKGROUND AND PURPOSE The early ischemic lesions demonstrated by diffusion-weighted imaging (DWI) are potentially reversible. The purposes of this study were to determine whether resolution of initial DWI lesions is transient or permanent after different brief periods of focal brain ischemia and to evaluate histological outcomes. METHODS Sixteen rats were subjected to 10 minutes (n=7) or 30 minutes (n=7) of temporary middle cerebral artery occlusion or sham operation (n=2). DWI, perfusion-weighted imaging (PWI), and T(2)-weighted imaging (T(2)WI) were performed during occlusion; immediately after reperfusion; and at 0.5, 1.0, 1.5, 12, 24, 48, and 72 hours after reperfusion. After the last MRI study, the brains were fixed, sectioned, stained with hematoxylin and eosin, and evaluated for neuronal necrosis. RESULTS No MRI or histological abnormalities were observed in the sham-operated rats. In both the 10-minute and 30-minute groups, the perfusion deficits and DWI hyperintensities that occurred during occlusion disappeared shortly after reperfusion. The DWI, PWI, and T(2)WI results remained normal thereafter in the 10-minute group, whereas secondary DWI hyperintensity and T(2)WI abnormalities developed at the 12-hour observation point in the 30-minute group. Histological examinations demonstrated neuronal necrosis in both groups, but the number of necrotic neurons was significantly higher in the 30-minute group (95+/-4%) than in the 10-minute group (17+/-10%, P<0.0001). CONCLUSIONS Transient or permanent resolution of initial DWI lesions depends on the duration of ischemia. Transient resolution of DWI lesions is associated with widespread neuronal necrosis; moreover, permanent resolution of DWI lesions does not necessarily indicate complete salvage of brain tissue from ischemic injury.

Neuroprotective effects of a novel broad-spectrum cation channel blocker, LOE 908 MS, on experimental focal ischemia: a multispectral study.

Thirty‐four rats undergoing 90 minutes of temporary middle cerebral artery occlusion were randomly and blindly assigned to vehicle or (RS)‐(3,4‐dihydro‐6,7‐dimethoxyisoquinoline‐1‐γγ1)‐2‐phenyl‐N,N‐di‐2‐(2,3,4‐trimethoxyphenyl)ethyl acetamide (LOE 908 MS; 0.5 mg/kg) i.v. bolus at 30 minutes after arterial occlusion followed by a 5 mg/kg/hr i.v. infusion for 3.8 hours (n =17/group). Perfusion‐, diffusion‐ and T2‐weighted magnetic resonance imaging was performed before treatment and repeatedly after treatment. Multispectral analysis was used to define ischemic abnormalities. The size of the ischemic abnormalities, including the ischemic core and penumbra, was not different between the two groups before treatment. However, a significant difference in ischemic lesion size was detected beginning 1.5 hours after treatment. The size of the ischemic core was significantly smaller in the treatment group, while the size of the ischemic penumbra was similar in the two groups at 85 minutes after arterial occlusion. Postmortem infarct size at 24 hours was significantly smaller in the drug‐treated group than in the placebo group. These results demonstrate that LOE 908 MS can reduce ischemic lesion size, which is probably attributable to inhibition of expansion of the ischemic core. J. Magn. Reson. Imaging 1999;10:138–145.

Transient and Permanent Resolution of Ischemic Lesions on Diffusion-Weighted Imaging After Brief Periods of Focal Ischemia in Rats : Correlation With Histopathology Editorial Comment: Correlation With Histopathology

BACKGROUND AND PURPOSE The early ischemic lesions demonstrated by diffusion-weighted imaging (DWI) are potentially reversible. The purposes of this study were to determine whether resolution of initial DWI lesions is transient or permanent after different brief periods of focal brain ischemia and to evaluate histological outcomes. METHODS Sixteen rats were subjected to 10 minutes (n=7) or 30 minutes (n=7) of temporary middle cerebral artery occlusion or sham operation (n=2). DWI, perfusion-weighted imaging (PWI), and T(2)-weighted imaging (T(2)WI) were performed during occlusion; immediately after reperfusion; and at 0.5, 1.0, 1.5, 12, 24, 48, and 72 hours after reperfusion. After the last MRI study, the brains were fixed, sectioned, stained with hematoxylin and eosin, and evaluated for neuronal necrosis. RESULTS No MRI or histological abnormalities were observed in the sham-operated rats. In both the 10-minute and 30-minute groups, the perfusion deficits and DWI hyperintensities that occurred during occlusion disappeared shortly after reperfusion. The DWI, PWI, and T(2)WI results remained normal thereafter in the 10-minute group, whereas secondary DWI hyperintensity and T(2)WI abnormalities developed at the 12-hour observation point in the 30-minute group. Histological examinations demonstrated neuronal necrosis in both groups, but the number of necrotic neurons was significantly higher in the 30-minute group (95+/-4%) than in the 10-minute group (17+/-10%, P<0.0001). CONCLUSIONS Transient or permanent resolution of initial DWI lesions depends on the duration of ischemia. Transient resolution of DWI lesions is associated with widespread neuronal necrosis; moreover, permanent resolution of DWI lesions does not necessarily indicate complete salvage of brain tissue from ischemic injury.

Effects of signal to noise and parametric limitations on fitting biexponential magnetic resonance (MR) inversion-recovery curves using a constrained nonlinear least-squares algorithm

Ischemic stoke is a local reduction in brain tissue perfusion due to an obstruction (such as a blood clot) of the inflowing arterial blood. Using diffusion-weighted magnetic resonance imaging (DWI), it is possible to visualize the brain tissue affected in the acute (early) stages of ischemic stroke. Due to metabolic changes and disruptions in osmotic equilibrium, water moves from the extracellular to the intracellular compartment, which results in cytotoxic edema (cell swelling) and marked changes in the apparent diffusion coefficient (ADC) of brain tissue water. The mechanisms responsible for these ADC changes are not well understood but are thought to involve changes in the separation of restricting barriers and in the relative contributions from the intra- and extracellular compartments. Yeast cells have been investigated as a model system to provide some insight into intra- and extracellular ADC characteristics. Inversion-recovery MR data were fitted to a biexponential equation using a constrained nonlinear least squares algorithm. An ideal fit by the algorithm is not guaranteed since the goodness of fit is sensitive to the relative fractional compartmental contributions and the relaxation time constants of each compartment. Test data sets were created to determine the effects of signal to noise and system parameters on the goodness of fit. It was determined that the best fit was obtained for systems with equal compartmental contributions, relaxation time constants that differed by more than a factor of 3, and a high signal-to-noise ratio. The test data sets will be used to determine the optimal conditions for the yeast cell preparations and, eventually, the design of in vivo stroke experiments. Knowledge of the ADC characteristics in ischemic stroke can guide future research in therapy design and treatment approach.