Abstract
MRI is a sensitive technique for assessing disease activity in MS.Diseased white matter (WM) can be identified on T2-weighted images, and active disease is reflected by abnormalities in the blood-brain barrier (BBB) shown on T1-weighted images after administration of paramagnetic contrast agents. Active disease may be demonstrated by contrast-enhanced MRI in patients with early, mild relapsing-remitting (RR) MS even during periods of clinical stability, which indicates that MS is an active process even during the early phase of the illness. To examine the amount of abnormal WM at frequent intervals over time, we studied seven mildly affected RRMS patients, all of whom had frequent contrast-enhancing lesions. These RRMS patients were imaged monthly for 26 to 36 months at 1.5 tesla; the area of abnormal increased WM signal was calculated by image-processing software that utilizes both the T2- and T1-weighted images. All patients showed fluctuations over time in amount of abnormal WM signal, which reflected factors such as the amount of BBB breakdown (measured by number or area of enhancing lesions) and measurement error. All seven RRMS patients, however, showed an overall increase in abnormal WM. Because of the fluctuations between individual measurements, the increase was most accurately reflected when the mean of the first 6 months' measurements was compared with the mean of the final 6 months' measurements, or when a linear regression model was applied. Although the accumulation of abnormal WM provides an additional tool for assessing disease activity in MS, its usefulness may be increased by the measurements obtained with additional techniques that are currently available or as yet undeveloped.
NEUROLOGY 1995;45: 1808-1814
MRI provides a sensitive marker of diseased tissue in MS patients. [1-4] Diseased tissue in the CNS of MS patients, associated with either acute inflammatory lesions or long-standing demyelination, can be demonstrated by several MRI techniques. Acute and chronically diseased tissue appear as areas of increased signal intensity on T2-weighted (T2W) or proton density (PD) MR images. [1-4] T1-weighted (T1W) images obtained after injection of paramagnetic contrast agents, most commonly gadopentetate dimeglumine (GdDTPA), permit identification of new or acute lesions. [5] GdDTPA enhancement has been advocated [6-9] as a means of monitoring disease activity because the enhancing lesions are easily recognized and probably represent a critical initial event in the development of a new MS lesion.
The accumulation of abnormal white matter (WM), visualized as high-intensity signals on T2W or PD images, may provide a good measure of overall disease burden and correlate with long-term disability. These longstanding lesions have been less well studied, although a major clinical trial [10] of interferon beta-1b used accumulation of abnormal WM signal as an outcome measure. That study imaged 327 MS patients at 12-month intervals for up to 36 months and found a median increase of 15% in the amount of abnormal WM in patients receiving placebo compared with those receiving beta interferon.
From previous work we know that GdDTPA-enhancing lesions vary on a month-to-month basis in patients with relapsing-remitting (RR) MS. Pathologic studies and previous studies of T2W images led investigators to assume that there is a gradual accumulation of demyelinated tissue in RRMS patients. In the current study, we asked how the bulk of abnormal WM signal changes on MRI if measured on a month-to-month basis in RRMS patients with frequent enhancing lesions over a several-year period. We also assessed how these changes relate to other MRI findings and clinical features.
Methods.
Patients.
This study was reviewed and approved by the National Institute of Neurological Disorders and Stroke (NINDS) Institutional Review Board, and informed consent was obtained from each patient. Seven patients with clinically definite RRMS [11] were entered consecutively as a cohort, which has been described previously. [7,9,12] MRI criteria were not used in patient selection. Patient disability was classified according to the Expanded Disability Status Scale (EDSS). [13] Disability was mild in the RRMS patients, with EDSS scores of 3.0 or less at entrance Table 1.
Table 1. Demographic data describing the patient population
Patient evaluation.
Monthly neurologic evaluations and MRI were performed for 26 to 36 months (mean, 33.29) on the seven RRMS patients. Two healthy volunteers underwent similar MRI intermittently over 20 months. The patients were examined monthly and rated by one neurologist, who used a standardized neurologic examination and assigned an EDSS score based on the functional systems scale. Patients were treated for exacerbations with 1 g of IV methylprednisolone for 3 days followed by 3 weeks of alternate-day oral prednisone when clinically indicated. MRI was delayed by 7 to 10 days after tapering the medication to minimize effects on the imaging.
MRI.
The monthly studies on the General Electric Signa 1.5-tesla unit (General Electric, Milwaukee, WI) consisted of (1) axial oblique PD and T2W images with variable (double) echo (TE 20 msec, TE 100 msec), TR 2,000, 27 slices of 5 mm each, interleaved (contiguous, no gap), field of view (FOV) equals 24 cm, matrix 128 times 256, and two excitations (for the first 15 months, imaging was performed with TE of 80 msec); and (2) axial oblique T1W studies before and after IV injection of GdDTPA (Magnevist 0.1 mmol/kg, Berlex Laboratories, Cedar Knolls, NJ), TE 20 msec, TR 600, 27 slices of 5 mm each, interleaved (contiguous, no gap), FOV equals 24 cm, matrix 192 times 256, and two excitations.
Postcontrast T1W images were consistently begun within 5 minutes after the injection of GdDTPA in order to control for changes in the accumulation of the contrast agent in areas of blood-brain barrier (BBB) breakdown. Although exact slice-to-slice registration was not required by the method of data analysis, because sums of slices rather than individual slice-by-slice comparisons were used, reproducible head positioning was obtained by scout images where the orbital-meatal line was established by placement of one vitamin E capsule in the external auditory meatus and a second at the lateral canthus of the eye (see Validation of methods).
Image evaluation.
MR images from each monthly procedure were transferred from the MR unit to SUN Sparc-Stations (SUN Computers, Palo Alto, CA).
Two-image method.
Analyze 5.0.1 software (Mayo Foundation, Rochester, MN) was used to calculate the areas of interest for both the primary method, which used both T1W and T2W images (hereafter called the two-image method), and the secondary method, which used only the PD images (one-image method). This software allows the operator to place seed points in areas of interest and then expand them to outline larger areas by semi-automated thresholded segmentation through setting signal intensity windows. (As these two methods provide a sum of the areas found on many slices to provide the total for the entire brain, the word "area" has been used consistently to describe the values obtained from these methods.)
For the two-image method used with all patients, the low signal (black) area on T1W images that constitutes the lateral ventricles (LV; not including the third ventricle or the various cisterns) was seeded and the threshold adjusted to segment the appropriate areas on each slice. The pixel values from each slice were summed to obtain the LV area for the brain as a whole. The areas of the high signal intensity (bright white) abnormal WM and the lateral ventricles visualized on the T2W images were seeded and calculated together on a slice-by-slice basis by setting the threshold for the pixel intensities. The sum of the edited T1W slices (the area of the LV) was subtracted from the combined area of abnormal WM signal intensity and LV calculated on T2W images to leave a value for abnormal WM alone.
One-image method.
The one-image method was performed on PD images that were available for the last 15 to 20 months of study. (A sequence change occurred during the course of study whereby a double-echo sequence produced a PD image in addition to the usual T2W images.) In the PD sequence, CSF has lower signal intensity than the higher-intensity, brighter WM lesions, and the latter foci thus can be segmented by pixel intensity thresholds. The area of abnormal WM on the PD images was calculated as described above by placement of seed points and segmentation by pixel intensity thresholds.
Validation of the two-image and one-image methods.
Inter-rater reproducibility.
Two raters other than the principal rater were trained in the two-image method. The area values obtained by these raters differed from those of the primary rater by 5% and 6%. More similar values could be obtained for the T1W lateral ventricle measurements than for the T2W images due to the increased number of judgments required by the rater for the T2W images.
Intra-rater reproducibility.
A single operator performed the area calculations on all the patients for both methods. Area calculations were made from each image at least twice, with a third measurement performed if the first two differed by more than 9%. Images were evaluated generally in chronologic or reverse chronologic order, although occasionally some were evaluated out of order. To examine the effect of head positioning, a patient was placed in the apparatus at two different times during the day with resetting of landmarks. The area of abnormal WM signal intensity obtained from the two sets of T2W images differed by 4%. Thus, summing the likely sources of error, and adding 50% to this total (4% from repositioning plus 6% from inter-rater equal 10%), we would estimate that up to 15% of the month-to-month fluctuations may be attributable to measurement error. The error may be slightly lower for the one-image method because one less step is involved in this method.
Evaluation of controls.
The two-image method assumes that the LV are of similar size whether measured on the T1W or T2W images. This assumption was first evaluated in two healthy controls (data not shown). Calculations of LV area on T1W and T2W images of the normal controls differed by less than 6% in each case. The differences in the size of the acquisition matrices (128 times 256 for T2W images and 192 times 256 for T1W images) and interpolation of the image to a 256 times 256 display matrix for T1W and T2W images could account for this level of disagreement. Although month-to-month variability was observed in these healthy subjects, the LV size covaried between the T1W and T2W images and remained essentially unchanged over the period of study.
Statistical analysis.
A linear regression model with an autoregressive moving average (ARMA) process for the errors was used to test for a linear trend in these MRI data. [14,15] This generalization of the simple linear regression model adjusts for the serial correlation apparent in these data, ie, that the amount of abnormal WM in one scan is affected by the amount of abnormal WM in the scans preceding it in time. This model is described by Equation 1 where Yt and epsilont are the measured data from the MRI and error terms, respectively, at month t. The slope B1 represents the rate of change of Yt, thus the rate of accumulation of abnormal WM. A chi-square test (likelihood ratio) was used to test for a positive slope, ie, accumulation or growth of the characteristic measured on MRI. Models were fitted and tests were made on a patient-by-patient basis. A p value of less than 0.05 was considered to be the threshold for statistical significance.
We used a logistic regression model with autoregressive terms to adjust for clustering or time dependence, [16,17] to examine the overall effect of enhancement demonstrated on T1W imaging during months t and t minus 1 on the probability of having a value at least 15% above the average value for WM signal abnormality for each patient. (Fifteen percent was chosen because this is the presumed maximum measurement error.) The model can be written as Equation 2 where Yit indicates whether the area of abnormal WM at month t is at least 15% above the average amount of abnormal WM area on the ith patient, and where beta1 measures the effect (over all patients) of normalized contrast-enhanced area at time t, and beta2 measures it at time t minus 1, on the probability of having a value at least 15% above an average value of abnormal WM signal at time t. Both the contrast-enhanced areas on T1W images and the abnormal WM signal areas were normalized because the data varied greatly across the seven patients. Normalization was done in two ways: (1) by dividing each monthly contrast-enhanced area and abnormal WM signal area by their corresponding patient average, and (2) by dividing each patient's variables by the average of their six neighboring observations. Each normalization technique produced similar results. Z-tests were used to test for non-zero values of beta1 and beta2.
The data from our seven patients with RRMS were also used to calculate sample sizes for various parallel group designs. The sample sizes required to detect a 100% reduction in the positive slope due to treatment (alpha equals 0.05, power equals 0.8; two-tailed test) were computed [18] with variance estimates obtained by a repeated sampling technique (bootstrapping). [19]
Results.
Assessment of abnormal white matter.
The two-image method, which calculates abnormal WM by subtracting LV area on T1W images from total WM signal abnormality on T2W images, was applied to all seven patients Table 1. This approach was assessed in serial evaluation of two healthy controls. Examples of monthly measurements of abnormal WM are shown for two RRMS patients (patients 3 and 7) in Figure 1. Fluctuations were seen from month to month in the measurements of abnormal WM in each patient.
Figure 1. Graphs of accumulation of abnormal white matter (WM) on T2-weighted images through time calculated by measuring the area of abnormal WM and lateral ventricles (LV) on T2-weighted images and subtracting from it the area of LV on T1-weighted images in two RRMS patients, nos. 3 and 7. Images that were made within a month or less of corticosteroid treatment are indicated by arrows. (Lin Reg [ARMA] equals linear regression with autoregressive moving average; EDSS equals Expanded Disability Status Scale)
The one-image method for measuring abnormal WM, by semiautomated threshold segmentation of PD images, was also performed on four patients over a 15- to 20-month period. A representative example is shown for patient 6 in Figure 2, comparing the two measures of abnormal WM. The magnitude of the month-to-month fluctuation is reduced but still evident in the data obtained from the PD images.
Figure 2. Graph of accumulation of white matter (WM) abnormality over time in RRMS patient 6 compares measurements of abnormal WM on T2-weighted images (two-image method--subtraction of the area of lateral ventricles measured on T1-weighted images) with measurements on proton density images (one-image method).
The month-to-month fluctuations seen in all patients by both methods can be partially explained by a combination of at least three factors as discussed below: (1) measurement error, (2) lesions exhibiting BBB breakdown and thus contrast enhancement, and (3) corticosteroid treatment in some of the patients.
Comparison of abnormal white matter with GdDTPA enhancement.
Because we had monthly data measuring abnormal WM signal area and corresponding data on the area of BBB disruption from contrast-enhanced T1W images, we could compare these data to look for two types of relationships: (1) an association of the amount of enhancing area with the fluctuations in abnormal WM signal area; and (2) the general relationship of the activity level (number or area of contrast-enhancing lesions) with the rate of accumulation of abnormal WM signal area over time.
The amount of abnormal WM as measured by the two-image method was compared with the area of GdDTPA-enhancing lesions for each month because all seven patients had enhancing lesions. Large increases in the area of GdDTPA-enhancing lesions generally were accompanied by large increases in the amount of abnormal WM signal. By use of logistic regression modeling (as discussed in the Methods section), we found that an increase in contrast-enhanced area on T1W imaging relative to a patient's average was associated with a higher probability of having a simultaneous increase (more than 15% higher than the individual patient's average) in abnormal WM signal area (p equals 0.02). No evidence for a predictive effect of the prior month's contrast-enhanced area on the next month's area of abnormal WM signal was found (p equals 0.38).
With regard to the effect of the number or area of enhancing lesions on the rate of accumulation of abnormal WM signal area, our sample size was too small to allow definitive analysis. However, a suggestion of a linear association was found between an increased rate of accumulation in abnormal WM signal and increases in both measures of BBB breakdown--number and area of GdDTPA-enhancing lesions (correlation coefficients: r equals 0.66 for number and r equals 0.89 for area).
Corticosteroid treatment.
During the course of this study, patients were treated with corticosteroids 18 times, with one patient (patient 4) accounting for nine of these instances. In general, MRI was performed on patients 7 to 10 days after stopping corticosteroid treatment, although four MRIs were obtained while the patients were taking 10 to 20 mg of oral prednisone. The effect of corticosteroid treatment on the amount of abnormal WM signal was not consistent, although bulk abnormal WM did appear to decrease in some patients during the months affected by treatment (for patients 3 and 4, shown in Figure 1 and Figure 3, decreases occurred at four of the seven data points potentially affected by corticosteroids).
Figure 3. Comparison of area of GdDTPA enhancement in RRMS patient 4 with accumulation of abnormal white matter (WM) by measurements of T2- and T1-weighted images (two-image method). Arrows indicate data points less than equals to 1 month after corticosteroid treatment.
Accumulation of abnormal white matter.
Measurements of abnormal WM were analyzed in two ways to assess the accumulation of abnormal WM over time. First, after application of the two-image method, the mean of the data obtained from the initial 6 months was compared with the mean of the final 6 months' data in each patient. All seven RRMS patients showed an increase in the mean bulk of abnormal WM and LV size (Figure 4, patients 1 to 7).
Figure 4. Comparison of the mean of the initial 6 months' measurements with the mean of the final 6 months' measurements in each patient. (A) Abnormal white matter on T2-weighted images (calculated from the abnormal white matter and lateral ventricles on T2-weighted images minus the lateral ventricles as measured on T1-weighted images). (B) Lateral ventricular size alone as measured on T1-weighted images.
Second, the data obtained with the two-image model were also evaluated by a linear regression model with an ARMA process for the errors (Equation 1 in Methods). This model is a modified linear regression analysis that provided assessment of the trends in accumulation of abnormal WM in the presence of the apparent serial correlation in these data. By this model, all seven RRMS patients had increases in LV area on T1W images. Increases in three of these patients were statistically significant, and a fourth patient showed a statistical trend (p equals 0.08). The rate of accumulation of abnormal WM signal in the seven RRMS patients is shown in the table. The slopes for three of the RRMS patients are statistically significant and a trend is seen for two additional patients.
Relationship between changes in abnormal WM signal and clinical features.
Four of the seven RRMS patients had increases in baseline EDSS of at least 0.5 during the course of the current study Table 1. All of the RRMS patients had increases in abnormal WM signal. No consistent, significant relationship between the magnitude of change in abnormal WM and clinical worsening was observed in the patients. With the exception of patient 4, who had a very large rate of increase, the rates of increase in abnormal WM were relatively similar between the RRMS patients.
Discussion.
The rationale for the current study was based on two sets of previous studies of MRI in MS patients: (1) serial studies that have demonstrated that the number and area of contrast-enhancing lesions vary from month to month in RRMS patients [7-9]; and (2) studies of T2W images that have shown a gradual accumulation of abnormal high signal intensity WM in RRMS over time. [10] Given these previous studies, we wished to investigate how the bulk of abnormal high signal intensity WM would change if measured on a monthly basis in RRMS patients with frequent contrast-enhancing lesions over 36 months. Secondarily, we wanted to learn how these changes in WM signal related to BBB disruption and clinical features.
With regard to the first goal, the current study indicated that abnormal cerebral WM signal increases over time in RRMS patients, but that these RRMS patients exhibit greater month-to-month variability in the amount of abnormal WM signal than would be expected by measurement error. Two previous studies, [20,21] although not involving such an extensive series, also noted fluctuations in abnormal WM over time. Because comparison of isolated single months' measurements, as established by either technique, revealed month-to-month fluctuations, we used two summary methods to analyze the two-image data more closely as a series. First, the initial 6 months of measurements were averaged and compared with the means of the final 6 months of measurements in each patient. By this method, our data showed an increase in abnormal WM of 20% (8.5% to 27.6%) for all seven RRMS patients over the period of study Figure 4 A.
In the second approach on the two-image data, we demonstrated an increase in abnormal WM over time by a linear regression model with autoregression to account for serial correlation Table 1. All seven RRMS patients had positive slopes by this analysis (three were statistically significant, and an additional two showed a trend). The probability that all seven patients would show a positive slope by chance alone (regardless of the individual p values) was calculated as 0.008 (based on a binomial distribution with a probability of having a positive slope equal to 1/2). (While the data from the one-image method were not analyzed in as much detail, results overall corroborated the findings of the two-image method.)
Application of both these approaches to LV area as measured on T1W images alone yielded similar results. By averaging the first 6 months' measurements and comparison with the last 6 months, LV size also increased over this period, by an average of 12% (range, 3.4 to 25%) Figure 4 B. When we examined the LV data by linear regression, we also found increases similar to the amount of bulk abnormal WM signal. These data support previous reports of atrophy with enlarged LV in long-standing MS. [22]
The most likely sources of the monthly fluctuations in abnormal WM signal area seen in our data are (1) measurement error, which we estimate to be up to 15% of the month-to-month changes in the current study (our measurements of inter- and intra-rater reliability were close to those found by another group of investigators using a similar method [23]); (2) month-to-month changes in the amount of BBB breakdown as measured by contrast-enhancing lesions; and (3) administration of corticosteroids systemically to the RRMS patients for clinical exacerbations. However, there may be additional underlying pathophysiologic mechanisms that are also at work to produce changes in edema or inflammation.
We modeled the relationship of monthly changes in BBB and abnormal WM signal by logistic regression and found that an increase in contrast-enhanced area on T1W images was associated with a higher probability of having a simultaneous increase (greater than 15% above the patient's average) in abnormal WM signal area. We found no evidence, though, for a predictive effect of the amount of BBB breakdown on the amount of the next month's abnormal WM signal area. Abnormalities on T2W images that correspond to enhanced lesions on T1W images may be initially larger on the T2W images due to increased sensitivity of that sequence to edema, although the amount of edema may vary with the age of the lesion. T2W lesions may persist as smaller areas of demyelination and gliosis after the acute inflammation and enhancement disappear on T1W images with closure of the BBB.
Administration of corticosteroids for clinical exacerbations also may contribute to the fluctuations in the amount of abnormal WM signal. Corticosteroid treatment affects the BBB after acute administration, [24] although its effects on abnormal WM are unclear, because previous studies have included comparisons of only two images per patient, without calculation of overall area of abnormal WM. [25] In the current study, corticosteroids did not consistently appear to affect the overall increase in abnormal WM or the month-to-month fluctuations. Corticosteroids were administered only for clinical exacerbations, and in only five of the seven RRMS patients. (See patient 3, Figure 1, and patient 4, Figure 3, where arrows indicate the data points most likely to have been affected by the therapy.) A future study could be planned that might demonstrate a more significant effect on abnormal WM in patients who took higher or more frequent doses.
The substantial month-to-month fluctuations in amount of abnormal WM signal found by both methods has important implications for the use of this type of data to measure outcome in therapeutic trials in MS. An example of how our data would appear if MRI had been performed at intervals of 12 months is seen in Figure 5. Given yearly MRI, a sample size calculation for parallel group design could be made from the data from the two-image method. Sample size estimates were computed as described in Methods, based on our sample of seven RRMS patients. The results suggest that 353 patients would have to undergo MRI twice at 12-month intervals to detect a 100% decrease in the rate of increase, ie, stabilization in abnormal WM due to treatment with the use of a parallel group design (power equals 0.8, alpha equals 0.05). The sample size calculated in this manner for single MRI at 12-month intervals is close to the number of patients enrolled in the multicenter interferon beta trial, [10] which found an effect of the drug on diseased WM burden when patients underwent MRI every 12 months.
Figure 5. Demonstration of how the data obtained from the calculations of area based on the measurement of T2-weighted abnormal white matter area by the two-image method would appear for data points taken only at 12-month intervals in a RRMS patient.
The relationship of burden of abnormal WM to clinical disease state or long-term prognosis remains unclear. Four of the RRMS patients had an increase in their baseline EDSS associated with their increase in bulk abnormal WM Table 1. We also found a suggestion of a linear association between rate of accumulation in abnormal WM signal and both measures of BBB breakdown--area and number of GdDTPA-enhancing lesions. This supports the notion of BBB breakdown as a marker of disease activity, but appears to be influenced by patient 4, who had a great deal of BBB breakdown.
Although T2W images are sensitive to abnormal WM, measurement of diseased WM is cumbersome by these methods. The simpler and faster methods of counting and measuring the area or volume of GdDTPA-enhanced lesions as evidence of disease activity may be more practical for clinical trials from the standpoints of operator ease, sample size, and length of trial. [9,26] However, the current study demonstrates that the amount of abnormal WM signal increases in some RRMS patients, with or without progression in clinical disability, and as reflected in the recent clinical trial of interferon beta-1b, can be an effective outcome measure.
- Copyright 1995 by Advanstar Communications Inc.
REFERENCES
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