Carol K. Petito

The Consortium to Establish a Registry for Alzheimer's Disease (CERAD). Part II. Standardization of the neuropathologic assessment of Alzheimer's disease

The Neuropathology Task Force of the Consortium to Establish a Registry for Alzheimers Disease (CERAD) has developed a practical and standardized neuropathology protocol for the postmortem assessment of dementia and control subjects. The protocol provides neuropathologic definitions of such terms as “definite Alzheimers disease” (AD), “probable AD,” “possible AD,” and “normal brain” to indicate levels of diagnostic certainty, reduce subjective interpretation, and assure common language. To pretest the protocol, neuropathologists from 15 participating centers entered information on autopsy brains from 142 demented patients clinically diagnosed as probable AD and on eight nondemented patients. Eighty-four percent of the dementia cases fulfilled CERAD neuropathologic criteria for definite AD. As increasingly large numbers of prospectively studied dementia and control subjects are autopsied, the CERAD neuropathology protocol will help to refine diagnostic criteria, assess overlapping pathology, and lead to a better understanding of early subclinical changes of AD and normal aging.

Delayed hippocampal damage in humans following cardiorespiratory arrest.

Transient ischemia in animals produces delayed cell death in vulnerable hippocampal neurons. To see if this occurs in humans, we reexamined brain slides from all patients with anoxic-ischemic encephalopathy and a well-documented cardiorespiratory arrest. Eight patients dying 18 hours or less after cardiac arrest had minimal damage in hippocampus and moderate damage in cerebral cortex and putamen. Six patients living 24 hours or more had severe damage in all four regions. The increase in damage with time postarrest was significant only in the hippocampus. Delayed hippocampal injury now documented in humans provides a target for possible therapy that can be initiated after cardiopulmonary resuscitation.

Vacuolar Myelopathy Pathologically Resembling Subacute Combined Degeneration in Patients with the Acquired Immunodeficiency Syndrome

Twenty of 89 consecutive patients with the acquired immunodeficiency syndrome (AIDS) in whom autopsies were performed over a 3 1/2-year period had a vacuolar myelopathy that was most severe in the lateral and posterior columns of the thoracic cord. Light and electron microscopy showed that vacuoles were surrounded by a thin myelin sheath and appeared to arise from swelling within myelin sheaths. Signs and symptoms referable to the spinal-cord lesions, including paraparesis, often accompanied by spasticity or ataxia (or both), were present in all five patients with marked pathological changes, in five of seven patients with moderate changes, and in two of eight patients with mild changes. Fourteen patients were demented. The clinical presentation was sufficiently distinctive to provide a guide for antemortem diagnosis. Possible causes of the vacuolar changes include uncharacterized viral infection or a metabolic derangement related to selective nutritional deficiency.

Overexpression of Clusterin in Human Breast Carcinoma

Clusterin has been implicated in numerous processes including active cell death, immune regulation, cell adhesion and morphological transformation. The purpose of this study was to examine clusterin expression in a large series of breast carcinomas by immunohistochemistry and in situ hybridization. The study included 40 samples of non-neoplastic glandular epithelia, 42 benign lesions, 15 atypical intraductal hyperplasias, 35 carcinomas in situ, 114 invasive carcinomas, and lymph node metastases from 40 patients. Epithelial normal cells were always negative for clusterin expression and only 19% of the benign lesions presented positive staining. In contrast to the benign lesions, however, the frequency of clusterin positive samples increased in atypical hyperplasias (47%, P = 0.08), intraductal carcinomas (49%, P = 0.01) and invasive carcinomas (53%, P < 0.001). Positive staining presented a cytoplasmic pattern, except in 3 cases of invasive carcinomas which had nuclear staining. Clusterin mRNA by in situ hybridization confirmed the specific cellular pattern of clusterin expression by immunohistochemistry. Clusterin expression was associated with large tumor size (P = 0.04), estrogen and progesterone receptor negative status (P = 0.02 and P = 0.001, respectively) and with the progression from primary carcinoma to metastatic carcinoma in lymph nodes (80% metastatic nodes had positive expression) (P = 0.004). Ten of 15 (67%) primary carcinomas without clusterin expression became positive in lymph node metastases, while most (22 of 25, 88%) of the clusterin-positive primary carcinomas were also immunoreactive in metastases. In survival analysis, clusterin expression did not represent a prognostic indicator by uni- or multivariate analysis. The increased clusterin expression in breast carcinomas tended to correlate inversely with the apoptotic index (P = 0.09) which indicates that clusterin gene expression is not a prerequisite to cellular death by apoptosis. From these results, we suggest that clusterin may have a role in tumorigenesis and progression of human breast carcinomas.

Hydrogen Ions Kill Brain at Concentrations Reached in Ischemia

Elevation of brain glucose before the onset of nearly complete ischemia leads to increased lactic acid within brain. When excessive, such acidosis may be a necessary factor for converting selective neuronal loss to brain infarction from nearly complete ischemia. To examine the potential neurotoxicity of excessive lactic acid concentrations, we microinjected (0.5 μl/min) 150 mM sodium lactate solutions (adjusted to 6.50–4.00 pH) for 20 min into parietal cortex of anesthetized rats. Interstitial pH (pH0) was monitored with hydrogen ion–selective microelectrodes. Animals were allowed to recover for 24 h before injection zones were examined with the light microscope. Injectants produced brain necrosis in a histological pattern resembling ischemic infarction only when pH0 was ≤ 5.30. Nonlethal injections showed only needle tract injuries. Abrupt deterioration of brain acid-base homeostatic mechanisms correlated with necrosis since pH0 returned to baseline more slowly after lethal tissue injections than after nonlethal ones. The slowed return of pH0 to baseline after the severely acidic injections may reflect altered function of plasma membrane antiport systems for pH regulation and loss of brain hydrogen ion buffers.

Selective Glial Vulnerability following Transient Global Ischemia in Rat Brain

Global cerebral ischemia selectively damages neurons, but its contribution to glial cell death is uncertain. Accordingly, adult male. rats were sacrificed by perfusion fixation at 1, 2, 3, 5, and 14 days following 10 minutes of global ischemia. This insult produces CA1 hippocampal neuronal death at post-ischemic (PI) day 3, but minor or no damage to neurons in other regions. In situ end labeling (ISEL) and immunohistochemistry identified fragmented DNA of dead or dying glia and distinguished glial subtypes. Rare ISEL-positive oligodendroglia, astrocytes, and microglia were present in control brain. Apoptotic bodies and ISEL-positive glia significantly increased at PI day 1 in cortex and thalamus (p <0.05) but were similar to controls in other regions and at other PI intervals. Most were oligodendroglia, although ISEL-positive microglia and astrocytes were also observed. These results show that oligodendroglia die rapidly after brief global ischemia and are more sensitive than neurons in certain brain regions. Their selective vulnerability to ischemia may be responsible for the delayed white matter damage following anoxia or CO poisoning or that associated with white matter arteriopathies. Glial apoptosis could contribute to the DNA ladders of apoptotic oligonucleosomes that have been found in post-ischemic brain.

Selective vulnerability of preterm white matter to oxidative damage defined by F2-isoprostanes

Periventricular white matter injury (PWMI) is the leading cause of cerebral palsy and chronic neurological disability in survivors of prematurity. Despite the large number of affected children, the pathogenetic mechanisms related to PWMI remain controversial. Through studies of 33 human autopsy brains, we determined that early PWMI was related to oxidative damage that particularly targeted the oligodendrocyte lineage, whereas other neuronal and glial cell types were markedly more resistant. F2‐isoprostanes, an arachidinate metabolite/lipid peroxidation marker of oxidative damage, were significantly increased in early PWMI lesions but not in cerebral cortex. That deleterious lipid peroxidation accompanied early PWMI was supported by similar increases in F2‐isoprostanes levels in the cerebral cortex from term infants with hypoxic‐ischemic cortical injury. Detection of F4‐neuroprostanes, a neuronal‐specific oxidative damage marker, confirmed that neuroaxonal elements were resistant to injury in cerebral cortex and white matter. Significant protein nitration was not detected in PWMI lesions by 3‐nitrotyrosine staining. Significant cellular degeneration was confirmed in early PWMI lesions by terminal deoxynucleotidyltransferase–mediated dUTP nick end labeling and a marked depletion of oligodendrocyte progenitors of 71 ± 8%. Hence, the predilection of preterm infants for PWMI is related to selective lipid peroxidation–mediated injury of cerebral white matter and targeted death of oligodendrocyte progenitors. Ann Neurol 2005;58:108–120

Ultrastructural characteristics of the brain and blood-brain barrier in experimental seizures.

During experimental seizures, the blood-brain barrier (BBB) is broken; tracer substances such as I131-albumin, Evans blue and horseradish peroxidase (HRP) geographically locate the barrier breakdown primarily in the diencephalon. Using rats, we have induced seizures with electroshocks and demonstrated the breakdown of the BBB with Evans blue and HRP. We have shown that (1) the BBB breakdown is proportional to the number of electroconvulsant shocks (ES) given; (2) the mechanism of increased barrier permeability is primarily by micropinocytosis in the cerebral capillaries, arterioles, and, to a lesser extent, venules; and (3) the stimulus for micropinocytosis and hence BBB breakdown is associated with the abrupt rise in systemic blood pressure and cerebral vasodilatation that accompanies each ES. If the systolic hypertension is abolished via cervical cordotomy, there is little to no breakdown in the BBB.

DNA fragmentation follows delayed neuronal death in CA1 neurons exposed to transient global ischemia in the rat

Apoptosis is an active, gene-directed process of cell death in which early fragmentation of nuclear DNA precedes morphological changes in the nucleus and, later, in the cytoplasm. In ischemia, biochemical studies have detected oligonucleosomes of apoptosis whereas sequential morphological studies show changes consistent with necrosis rather than apoptosis. To resolve this apparent discrepancy, we subjected rats to 10 minutes of transient forebrain ischemia followed by 1 to 14 days of reperfusion. Parameters evaluated in the CA1 region of the hippocampus included morphology, in situ end labeling (ISEL) of fragmented DNA, and expression of p53. Neurons were indistinguishable from controls at postischemic day 1 but displayed cytoplasmic basophilia or focal condensations at day 2; some neurons were slightly swollen and a few appeared normal. In situ end labeling was absent. At days 3 and 5, approximately 40 to 60% of CA1 neurons had shrunken eosinophilic cytoplasm and pyknotic nuclei, but only half of these were ISEL. By day 14, many of the necrotic neurons had been removed by phagocytes; those remaining retained mild ISEL. Neither p53 protein nor mRNA were identified in control or postischemic brain by in situ hybridization with riboprobes or by northern blot analysis. These results show that DNA fragmentation occurs after the development of delayed neuronal death in CA1 neurons subjected to 10 minutes of global ischemia. They suggest that mechanisms other than apoptosis may mediate the irreversible changes in the CA1 neurons in this model.

Dementia with bilateral medial tempora1 lobe ischemia

Neuropathologic examination of two patients with dementia showed chronic bilateral medial temporal lobe ischemic damage that included the hippocampus (particularly the CA-1 region), subiculum, and amygdala. Both patients had several myocardial infarctions, and the relatively circumscribed cerebral injury may have resulted from one or more episodes of global hypoxic ischemia. Focal hippocampal injury has been associated with amnesia. The additional damage to medial temporal lobe structures may have caused the dementia.