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- PATIENTS AND METHODS
- AUTHOR CONTRIBUTIONS
The characteristics of the 7 patients with fatal NPSLE are shown in Tables 1 and 2. The majority of the NPSLE patients (6 of 7) were female. Their mean ± SD age was 39.6 ± 13.9 years. Their mean ± SD scores on the disease activity and severity assessments were 35.6 ± 21.0 for the SLEDAI, 17.1 ± 8.6 for the Neuro-SLEDAI, 11.3 ± 5.2 for the SDI, and 3.00 ± 1.15 for the Neuro-SDI. The mean ± SD MRS voxels per patient was 2.86 ± 1.86. The high SLEDAI and SDI scores indicated that these patients had very active and aggressive SLE directly prior to death, and all of them had some involvement of component of NPSLE prior to and/or during the episode associated with their death (Table 1). Table 2 summarizes the serologic profiles of the study patients.
Table 2. Serologic profiles of the study patients with fatal neuropsychiatric systemic lupus erythematosus*
| ||ANA titer||dsDNA titer||Sm||RNP||SSA||SSB||Ribosomal P||aCL||LAC|
|IgG, GPL units||IgM, MPL units||IgA, APL units|
|Summary, %||100||71.4||71.4||14.3||57.1||0.0|| 28.6||57.1||71.7||28.3||71.4|
There were significant differences in the levels of brain neurometabolites in the nonlesional occipitoparietal white matter of the NPSLE patients as compared with the controls. In the NPSLE patients, the absolute concentrations of NAA were 25% lower than those in the controls (mean ± SD 9.15 ± 1.78 mM versus 12.2 ± 0.8 mM [95% confidence interval (95% CI) –30.9, –19.1]; P < 0.01). The creatine level was 6.9% lower (mean ± SD 6.43 ± 0.16 in patients versus 6.90 ± 0.60 mM in controls [95% CI –13.5, –0.1]; P < 0.003), and the choline level was 30.7% higher (mean ± SD 2.51 ± 0.42 in patients versus 1.92 ± 0.32 mM in controls [95% CI 17.2, 44.3]; P < 0.04) in the patients than in the controls. Within the NPSLE group, voxels from lesional tissue (n = 13) as compared with nonlesional tissue (n = 7) assessed by MRS demonstrated 29.8% less NAA (mean ± SD 6.42 ± 2.3 mM in lesional versus 9.15 ± 1.78 mM in nonlesional tissue [95% CI –52.9, –6.8]; P < 0.009), 44.0% less creatine (mean ± SD 3.60 ± 2.40 mM in lesional versus 6.43 ± 0.16 mM in nonlesional tissue [95% CI –74.0, –14.0]; P < 0.001), and 60.2% more choline (mean ± SD 4.02 ± 0.52 mM in lesional versus 2.51 ± 0.42 mM in nonlesional tissue [95% CI 41.0, 79.3]; P < 0.001).
Examination of the MRS spectra revealed several different patterns of neurometabolite disturbance, including: 1) increased levels of lactate associated with acute ischemic injury, and with necrosis related to vascular thrombosis (patients 1 and 3); 2) reduced levels of NAA and creatine associated with old ischemic injury, with reduced cellularity (reduced neuronal–axonal density), cellular debris, and glial hyperplasia (patients 1, 2, 4, 6, and 7); 3) reduced levels of NAA associated with acute diffuse ischemic or cytotoxic cellular (neuronal–axonal) injury, without vascular thrombosis (patients 3 and 5); and 4) reduced levels of NAA and elevated levels of choline associated with chronic cellular rarification (neuronal–axonal dropout) related to chronic inflammation and accumulation of calcium concretions (patient 2). There were also widespread heterogeneous histologic changes, including microinfarcts, microhemorrhages, bland angiopathy, thrombotic angiopathy with platelet and fibrin thrombi, neuronal necrosis in various states of resolution, reduced numbers of axons and neurons, vacuole and space formation among the fibers, reduced numbers of oligodendrocytes, reactive microglia and astrocytes, lipid-laden macrophages, and cyst formation, and in 1 patient, calcium-containing concretions consistent with SLE-associated Fahr's disease (23).
Figure 1 shows paired MRS and histologic data from a patient with NPSLE (patient 4) who had antiphospholipid antibodies, increased SLE disease activity, and dementia. This patient experienced an acute cerebral infarction and died. The MRS spectrum in the region away from the acute infarct was located in the occipitoparietal white matter on neuroimaging, which demonstrated chronic diffuse increased intensity on T2-weighted and fluid-attenuated inversion recovery (FLAIR) images (results not shown). MRS (Figure 1, left) demonstrated reduced NAA levels, reduced creatine levels, and elevated choline levels, which on histologic examination (Figure 1, right), corresponded to linear areas of chronic neuronal and parenchymal hypocellularity associated with chronic vasculopathy of nearby blood vessels.
Figure 1. Magnetic resonance spectroscopy (MRS) and histopathologic assessment of brain tissues in a patient with neuropsychiatric systemic lupus erythematosus (NPSLE) and dementia. Shown are paired data from an MRS performed premortem and histopathologic assessment performed postmortem in NPSLE patient 4, who had antiphospholipid antibodies, increased SLE disease activity, and dementia. This patient died of an acute cerebral infarction. The MRS spectrum (left), which was obtained in minimally abnormal occipitoparietal white matter, demonstrates reduced levels of N-acetylaspartate (NAA), reduced levels of creatine (Cre), and elevated levels of choline (Cho). A myoinositol (mI) peak is also shown. The histopathologic appearance of the same area of the brain (right) shows linear areas of chronic neuronal and parenchymal hypocellularity (short arrows) associated with chronic vasculopathy of nearby blood vessels (arrow) (Luxol fast blue/periodic acid–Schiff stained, original magnification × 100).
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Figure 2 shows paired MRS and histologic data from a patient with NPSLE (patient 1) who had antiphospholipid antibodies, increased SLE disease activity, and multiple cerebral infarctions. This patient experienced an acute cerebral infarction and died. The neuroimaging results were characterized by hyperintensity on the FLAIR and T2-weighted images and by restricted diffusion on the diffusion-weighted image (results not shown). In the first few days after cerebral infarction, the MRS (Figure 2, left) showed the characteristic doublet of increased lactate levels (1.32 ppm) and low NAA levels during the early phase of infarction. During the resolution phase (2 weeks postinfarction), this same region was characterized by further reductions in NAA levels, increased choline and lipid levels, and persistent lactate (results not shown). After the patient died of further cerebrovascular events, the underlying histopathology in the brain (Figure 2, right) was assessed. Brain tissues demonstrated a typical resolving cerebral infarction, with a marked reduction in cellularity (neuronal–axonal numbers), necrotic amorphous material, infiltration with phagocytic cells, and glial hyperplasia. Surrounding blood vessels showed persistent thrombosis.
Figure 2. Magnetic resonance spectroscopy (MRS) and histopathologic assessment of brain tissues in a patient with neuropsychiatric systemic lupus erythematosus (NPSLE) and acute cerebral infarction. Shown are paired data from an MRS performed premortem and histopathologic assessment performed postmortem in NPSLE patient 1, who had antiphospholipid antibodies, increased SLE disease activity, and multiple cerebral infarctions. This patient died of an acute cerebral infarction. Findings of MR imaging (results not shown) were characterized by hyperintensity on the fluid-attenuated inversion recovery and T2-weighted images and restricted diffusion on the diffusion-weighted image. During the first few days after a cerebral infarction, the MRS spectrum (left) showed the characteristic doublet of increased lactate at 1.32 ppm and low levels of N-acetylaspartate (NAA) during the early phase of infarction. Creatine (Cre), choline (Cho), and myoinositol (mI) peaks are also shown. During the resolving phase (2 weeks postinfarction), this same region was characterized by further reductions in NAA levels, increased levels of choline and lipid, and persistent lactate (results not shown). After the patient died of further cerebrovascular events, the histopathologic appearance of the area of the acute infarct of the brain was assessed (right) and demonstrated a typical resolving cerebral infarction, with marked reduction in cellularity (neuronal–axonal numbers) (long arrow), necrotic amorphous material (arrowheads and short arrow), infiltration with phagocytic cells, and glial hyperplasia. Surrounding blood vessels showed persistent thrombosis (thick arrows) (Luxol fast blue/periodic acid–Schiff stained, original magnification × 50).
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Correlation and regression analyses across all samples were then used to explain the MRS versus histopathology relationships. Univariate correlations between the histopathologic findings and the brain neurometabolite levels demonstrated multiple associations. However, multivariate logistic analysis demonstrated that an elevated choline level was independently associated with gliosis (partial correlation coefficient β = 1.005, P < 0.0004), vasculopathy (β = 0.46, P < 0.03), and edema (β = 0.78, P < 0.006) (r = 0.75, P < 0.004 in the total model). Similarly, a reduced creatine level was independently associated with reduced neuronal–axonal density (β = –0.55, P < 0.03) and gliosis (β < –0.64, P < 0.03) (r = 0.72, P < 0.002 in the total model). A reduced level of NAA was independently associated with reduced neuronal–axonal density (r = 0.66, P < 0.001). The presence of lactate was independently associated with the presence of necrosis (β = 0.65, P < 0.0001), microhemorrhages (β = 0.31, P < 0.0001), and edema (β = 0.34, P < 0.0001) (r = 0.996, P < 0.002 in the total model).
- Top of page
- PATIENTS AND METHODS
- AUTHOR CONTRIBUTIONS
In the present study, we compared the levels of brain neurometabolites, as determined by premortem MRS, with the findings of histopathologic examination of the brain, as determined postmortem, in patients with NPSLE. This experimental design, which paired MRS with specific histopathologic assessment, provides a direct, rather than inferred, pathologic basis for the profound changes in neurometabolite levels in patients with NPSLE (5–11, 24–27). The principal changes include reduced NAA and creatine levels and elevated choline (choline-containing compounds) and lactate levels associated with reduced neuronal density, increased glial elements, acute and resolving injury, and in the majority of patients, acute and chronic ischemic or nonspecific vasculopathy (Figures 1 and 2). These histopathologic changes are broadly consistent with the pathologic changes identified by microscopy as reported in classic autopsy series of NPSLE patients (28–35).
The neurometabolite NAA is located almost entirely in neurons and is the strongest peak in the MRS brain spectrum in adults (36–38). The precise function of NAA in neuronal metabolism as an osmolyte and metabolic reserve continues to be explored, but NAA appears to be essential for normal neurocognitive function (21). Reduced levels of NAA have been noted in many diseases, and this is generally interpreted as representing neuronal injury or death (5–14). NAA levels have previously been reported to be reduced in normal-appearing white matter and gray matter, as well as in focal lesions, of patients with NPSLE (5–14, 36–38). Reduced NAA levels in radiologically normal–appearing tissues in NPSLE patients have been correlated with small focal lesions seen on MRI elsewhere in the brain, suggesting that the decline in NAA might also be due to extensive microlesions, most likely microinfarcts, that are too small to be seen on MRI (9, 24). This is further supported by the close association between reduced NAA levels and the presence of IgG antiphospholipid antibodies in NPSLE patients, suggesting thrombotic microinfarcts as a common cause of this decline (26). However, decreased NAA levels have also been noted in patients with generalized seizures, psychosis, or confusional state, which are not always associated with thrombosis, indicating that other causes of injury, including global ischemia and nonischemic cytotoxic effects, might be involved (27).
The present study confirms the idea that the reduced NAA level in patients with NPSLE is a significant finding and is most closely associated with reduced cellularity (reduced neuronal–axonal density). A reduced creatine level was also associated with reduced neuronal–axonal density and the presence of gliosis and inflammation, which are markers of acute and chronic brain injury. These changes were present in both normal-appearing tissues and lesional tissues, but were much worse in the lesions.
Other neuronal metabolites seen on MRS might also play important roles in NPSLE (14, 38). The choline peak represents choline moieties (–N+[CH3]3) that are visible on MRS, primarily phosphocholine, glycerophosphocholine, and choline. Choline levels are often elevated in NPSLE patients and have been postulated to be related to disease activity, stroke, inflammation, or chronic white matter disease (5–14, 36–38). Increased choline is also observed in NPSLE patients without obvious stroke or in the normal-appearing white matter, indicating the presence of preclinical disease or considerable microscopic brain disease not easily identified using conventional imaging modalities (5, 26, 27). There is evidence that increased choline levels might be a measure of disease activity or evidence of reactive brain inflammation, and thus, elevated choline levels in NPSLE patients should be interpreted as an ominous sign (9, 27). This is further supported by the finding that choline levels are elevated in NPSLE patients with cognitive impairment, indicating that the functional impact of this disease may be reflected by both elevated choline and reduced NAA (20, 26). Increased choline was associated histologically with vascular and reactive changes, including gliosis, vasculopathy, inflammatory cell infiltration, and edema, similar to the findings in other diseases in which there are active or reactive brain processes (13, 14, 38).
The presence of elevated upfield peaks at 1.32 ppm arising from lipids, macromolecules, and lactate has previously been associated with elevated disease activity in patients with NPSLE (5–14, 36–38). This is similar to the neurometabolic changes found in diseases associated with persistent or slowly resolving membrane degradation, activation, and demyelination (14, 38). However, although ischemia is often important in NPSLE, definite lactate in nonlesional tissue has not previously been observed on MRS, suggesting that in anything but overt stroke or global ischemia, extensive fixed anaerobic metabolism is not a fundamental characteristic of NPSLE (5–15, 30–33, 36–39). It should also be noted that lactate and other upfield peaks can be observed in patients with nonvascular diseases, including brain abscess, carbon monoxide poisoning, and status epilepticus (36–38). In the present study, the presence of lactate was associated with severe histopathologic changes in the brain, including tissue necrosis, microhemorrhages, and cerebral edema (Figure 2).
Previous autopsy studies in NPSLE have demonstrated highly variable changes, from minimally to grossly abnormal features, with cortical atrophy, gross infarcts, gross hemorrhage, microhemorrhages, ischemic demyelination, multiple sclerosis–like demyelination, and rarely, subdural hematoma, vasculitis, or aneurysms (28–35, 39–46). Fatal cases of NPSLE have also been associated with stroke or complement consumption, leukostasis, leukoaggregation, ischemia, and extensive cortical microinfarcts, indicating acute immune-mediated vascular occlusion (32–40). Leukostasis was not observed in the present study, but infarcts, microinfarcts, neurocytotoxic changes, vasculopathy, and many nonspecific or minimal changes were present. The concept of certain forms of NPSLE manifesting as cerebral edema with only subtle correlates at autopsy is emerging, particularly for the diffuse manifestations of NPSLE, with edema having an ominous significance (3, 47).
Bland vasculopathy, characterized by vascular hyalinization, vessel tortuosity, endothelial proliferation, and perivascular gliosis, is a common, but nonspecific, finding in NPSLE that was also observed in patients in the present study. This is indistinguishable from a distinctive type of cerebral vasculopathy characterized by fibrin thrombi, widespread obstruction by a proliferation of intimal fibrous tissue and myointimal cells, varying stages of recanalization, and in the late stages, fibrous webs across arterial lumens, which may be identified in the small leptomeningeal arteries of patients with high levels of antiphospholipid antibodies or with Libman-Sacks endocarditis (41, 48, 49). Cortical microinfarcts have also been associated with thrombi or emboli due to antiphospholipid antibodies, cardiac lesions, dissection, fibromuscular dysplasia, vasculitis, or atherosclerosis (28–35). Perivascular cuffing of arterioles or venules with inflammatory cells may occur, but there is a general recognition now that true central nervous system vasculitis is rare in SLE (28–35, 40–46). In the present study, despite widespread vasculopathy, thrombosis, and microinfarcts, true vasculitis was observed in only 1 small focus in patient 2.
There are limitations to this study that are related both to the design and to the inherent difficulties in any autopsy study. SLE has highly variable mortality, thus, the predictability of death and the ability to obtain an autopsy within a fixed time interval from the previous imaging study was variable (Table 1). Therefore, in some patients, the histologic changes may have been more evolved and the pathologic changes more severe at autopsy than the MRS findings would suggest. Autopsy studies of this type are biased, since they rely on the death of patients who typically have more severe disease. Accordingly, our results cannot necessarily be translated to less severe or nonfatal cases of NPSLE. Such a study would require studying patients with less active NPSLE who died of other causes.
Furthermore, NPSLE as defined by the ACR case definitions does not include subclinical injury; therefore, altered neurometabolite levels and/or histopathologic changes are not pathognomonic of clinical NPSLE (19). The study design also did not specifically address the role of antineuronal, excitotoxic, or anti–N-methyl-D-aspartic acid receptor antibodies; however, if present, it is likely that these antibodies would amplify the neuronal injury initiated by what appears to be the primary vascular insult evident in these study patients (50). Another limitation of this study is the relatively small number of patients. Clearly, a much larger multisite collaborative study is required to define the individual neurometabolite and histopathologic patterns associated with each NPSLE subtype (19). Finally, this study did not use specific stains and quantitative stereology to precisely quantify neuronal–axonal density, glial numbers, numbers of obstructed blood vessels, and the type, quantity, and immunotypes of inflammatory infiltrates; however, this study provides the scientific justification for including quantitative brain stereology in future autopsy studies in NPSLE (13).
Despite the inherent limitations and technical difficulties of autopsy studies, the findings of the present study confirm that abnormal brain neurometabolites measured premortem by MRS are indicative of significant underlying histopathologic changes consistent with acute and chronic brain injury. The findings of this study further support the concept of NPSLE as an aggressive and progressive brain disease and suggest that abnormal brain neurometabolites measured by MRS may indicate underlying histopathologic evidence of brain injury.