Dominantly Inherited Alzheimer Network Trials Unit (DIAN-TU): Trial Satisfaction and Attitudes towards Future Clinical Trials.

BACKGROUND: Clinical trial satisfaction is increasingly important for future trial designs and is associated with treatment adherence and willingness to enroll in future research studies or to recommend trial participation. In this post-trial survey, we examined participant satisfaction and attitudes toward future clinical trials in the Dominantly Inherited Alzheimer Network Trials Unit (DIAN-TU). METHODS: We developed an anonymous, participant satisfaction survey tailored to participants enrolled in the DIAN-TU-001 double-blind clinical trial of solanezumab or gantenerumab and requested that all study sites share the survey with their trial participants. A total of 194 participants enrolled in the trial at 24 study sites. We utilized regression analysis to explore the link between participants' clinical trial experiences, their satisfaction, and their willingness to participate in upcoming trials. RESULTS: Survey responses were received over a sixteen-month window during 2020-2021 from 58 participants representing 15 study sites. Notably, 96.5% of the survey respondents expressed high levels of satisfaction with the trial, 91.4% would recommend trial participation, and 96.5% were willing to enroll again. Age, gender, and education did not influence satisfaction levels. Participants reported enhanced medical care (70.7%) and pride in contributing to the DIAN-TU trial (84.5%). Satisfaction with personnel and procedures was high (98.3%). Respondents had a mean age of 48.7 years, with most being from North America and Western Europe, matching the trial's demographic distribution. Participants' decisions to learn their genetic status increased during the trial, and most participants endorsed considering future trial participation regardless of the DIAN-TU-001 trial outcome. CONCLUSION: Results suggest that DIAN-TU-001 participants who responded to the survey exhibited high motivation to participate in research, overall satisfaction with the clinical trial, and willingness to participate in research in the future, despite a long trial duration of 4-7 years with detailed annual clinical, cognitive, PET, MRI, and lumbar puncture assessments. Implementation of features that alleviate barriers and challenges to trial participation is like to have a high impact on trial satisfaction and reduce participant burden.

Effects of Vascular Risk Factors on the Association of Blood-Based Biomarkers with Alzheimer's Disease.

Background: Comorbidities may influence the levels of blood-based biomarkers for Alzheimer's disease (AD). We investigated whether differences in risk factors or comorbid conditions might explain the discordance between clinical diagnosis and biomarker classifications in a multi-ethnic cohort of elderly individuals. Aims: To evaluate the relationship of medical conditions and other characteristics, including body mass index (BMI), vascular risk factors, and head injury, with cognitive impairment and blood-based biomarkers of AD, phosphorylated tau (P-tau 181, P-tau 217), in a multi-ethnic cohort. Methods: Three-hundred individuals, aged 65 and older, were selected from a prospective community-based cohort for equal representation among three racial/ethnic groups: non-Hispanic White, Hispanic/Latino and African American/Black. Participants were classified into four groups based on absence (Asym) or presence (Sym) of cognitive impairment and low (NEG) or high (POS) P-tau 217 or P-tau 181 levels, determined previously in the same cohort: (Asym/NEG, Asym/POS, Sym/NEG, Sym/POS). We examined differences in individual characteristics across the four groups. We performed post-hoc analysis examining the differences across biomarker and cognitive status. Results: P-tau 217 or P-tau 181 positive individuals had lower BMI than P-tau negative participants, regardless of symptom status. Symptomatic and asymptomatic participants did not differ in terms of BMI. BMI was not a mediator of the effect of P-tau 217 or P-tau 181 on dementia. Frequencies of other risk factors did not differ between the four groups of individuals. Conclusions: Participants with higher levels of P-tau 217 or P-tau 181 consistent with AD had lower BMI regardless of whether the individual was symptomatic. These findings suggest that weight loss may change with AD biomarker levels before onset of cognitive decline. They do not support BMI as a confounding variable. Further longitudinal studies could explore the relationship of risk factors with clinical diagnoses and biomarkers.

Increased number of Purkinje cell dendritic swellings in essential tremor.

BACKGROUND AND PURPOSE: Essential Tremor (ET) is among the most prevalent neurologic disorders. Growing clinical and neuro-imaging evidence implicates cerebellar dysfunction in the pathogenesis of ET and emerging postmortem studies have identified structural changes in the cerebellum, particularly in Purkinje cells. In this study we systematically quantified focal Purkinje cell dendritic swellings (DS) in 20 ET vs. 19 control brains. METHODS: In each brain, a standard parasagittal neocerebellar tissue block was harvested. DS were quantified in one 7-µm thick section stained with Luxol Fast Blue/Hematoxylin and Eosin (LH&E) and one section stained with Bielschowsky method. RESULTS: The number of DS were higher in cases than controls by LH&E (1.50 ± 1.79 vs. 0.05 ± 0.23, P = 0.002) and Bielschowsky methods (2.70 ± 3.10 vs. 0.37 ± 0.50, P = 0.002). The number of DS was correlated with the number of torpedoes and marginally inversely correlated with the number of Purkinje cells. CONCLUSION: The current study documents and quantifies an additional structural abnormality in the ET cerebellum, adding to the growing list of such changes in this disease. The mechanisms that underlie this and other structural changes observed in ET are currently unknown, and they deserve additional exploration.

Agrin isoforms and their role in synaptogenesis.

Agrin is thought to mediate the motor neuron-induced aggregation of synaptic proteins on the surface of muscle fibers at neuromuscular junctions. Recent experiments provide direct evidence in support of this hypothesis, reveal the nature of agrin immunoreactivity at sites other than neuromuscular junctions, and have resulted in findings that are consistent with the possibility that agrin plays a role in synaptogenesis throughout the nervous system.

Microtubule protein preparations from C6 glial cells and their spontaneous polymer formation.

C6 cell tubulin is indistinguishable from hog brain tubulin with respect to its molecular weight, amino acid composition, and colchicine-binding activity. Moreover, microtubule assembly systems from both sources form the same structures: rings, ribbons, tubules, and drug-induced polymers. There is, nevertheless, a difference between the cultured cell and brain systems which lies in the nature of their microtubule-associated accessory proteins. C6 microtubule preparations exhibit few rings at 0 degrees C, have low polymerization yield, and have a low content of accessory proteins. The addition of brain accessory proteins enhances the numbers of rings, and the yield of microtubules, to levels comparable with those of brain preparations. The polymerizing ability of C6 microtubule protein decays much faster than that of brain, but it can be restored by the addition of brain accessory protein. The results suggest that C6 accessory proteins are more labile than their brain counterparts.

Neural differentiation, NCAM-mediated adhesion, and gap junctional communication in neuroectoderm. A study in vitro.

We studied the development of NCAM and gap junctional communication, and their mutual relationship in chick neuroectoderm in vitro. Expression of NCAM, as detected by monoclonal and polyclonal antibodies, and development of junctional communication, as detected by extensive cell-to-cell transfer of 400-500-D fluorescent tracers, occurred in cultures from stage-2 embryos onward. Both expressions presumably required primary induction. The differentiating cells formed discrete fields of expression on the second to third day in culture, with the NCAM fields coinciding with the junctional communication fields delineated by the tracers. Other neural differentiations developed in the following order: tetanus toxin receptors, neurofilament protein, and neurite outgrowth. Chronic treatment with antibody Fab fragments against NCAM interfered with the development of communication, suggesting that NCAM-mediated adhesion promotes formation of cell-to-cell channels. Temperature-sensitive mutant Rous sarcoma virus blocked (reversibly) communication and the subsequent development of neurofilament protein and neurites, but expression of NCAM continued.

A Randomized, Double-Blind, Phase 2 Study of the Effects of the Vaccine Vanutide Cridificar with QS-21 Adjuvant on Immunogenicity, Safety and Amyloid Imaging in Patients with Mild to Moderate Alzheimer's Disease.

BACKGROUND: Vanutide Cridificar (ACC-001), a novel investigational immunotherapeutic vaccine designed to elicit antibodies against the N-terminal peptide 1-7 of the amyloid-beta peptide, believed to be important in the pathogenesis of Alzheimer's disease (AD). OBJECTIVES: To evaluate the immunogenicity, safety and impact of ACC-001 with Quillaja saponaria (QS-21) adjuvant on the reduction of brain fibrillar amyloid burden, assayed by positron emission tomography (PET) imaging, in patients with mild to moderate AD. DESIGN: Randomized, phase 2, interventional study. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT01284387. PARTICIPANTS: Individuals with mild to moderate Alzheimer's disease (Mini-Mental State Examination scores 18-26; measurable amyloid burden in the expected range, on the screening 18F-florbetapir PET scan; and a Rosen modified Hachinski ischemic score ≤4). INTERVENTION: Participants were randomized to 3 µg or 10 µg ACC-001 (each in combination with 50 µg QS-21) or placebo (without QS-21). MEASUREMENTS: Primary endpoint was the change from baseline to week 104 in cerebral amyloid burden as measured by the global cortical average (GCA) standard value uptake ratio (SUVR) based on the brain 18F-florbetapir PET composite cortical SUVR between each ACC-001+QS-21 dose compared with placebo. Secondary endpoints included safety, immunogenicity and pharmacodynamics. Exploratory endpoints included cognitive and functional efficacy, and health outcome measures. RESULTS: Of 126 randomized patients (placebo: 40; ACC-001 3 µg+QS-21: 43; and ACC-001 10 µg+QS-21: 43), 125 received study treatment; 92 (73%) completed the study. Change in 18F-florbetapir PET GCA SUVR, was not significantly different between either of the two ACC-001+QS-21 treatment groups and placebo (3 µg +QS-21 vs. placebo diff=-0.03, p=0.54; 10 µg +QS-21 vs. placebo diff=-0.08, p=0.07), but the trend was numerically consistent with a dose response. The geometric mean peak anti-Aß IgG titers were slightly higher in the 10 µg than the 3 µg group. The proportion of responders was similar in both dose groups of ACC-001+QS-21. The cerebrospinal fluid (CSF) p-tau changes from baseline in both active treatment groups were not statistically different from placebo, but were numerically consistent with a dose response (3 µg +QS-21 vs. placebo diff=-3.2, p=0.57; 10 µg +QS-21 vs. placebo diff=-7.0, p=0.19). The vMRI showed statistically significant faster treatment-related decrease in brain volume in the 10 µg group but was not significant in the 3 µg group, compared with placebo (3 µg diff =-1.3 mL/year, p=0.50; 10 µg diff=-4.2 mL/year, p=0.02). Measured plasma Aß levels increased in parallel with peak anti-Aß titers after each injection. Amyloid-related imaging abnormalities with edema/effusion (ARIA-E) were more frequent in patients who received ACC-001+QS-21 than placebo (6% vs. 0%) but none were symptomatic. The most common treatment-emergent adverse events in the active groups were injection reactions, and occurred more frequently in the ACC-001+QS-21 groups than the placebo (48% vs 8%), the majority of which were mild and transient. CONCLUSIONS: Primary biomarker efficacy endpoints were not statistically significant in either dose group. The numerical decreases in 18F-florbetapir PET GCA SUVR suggests a dose-related trend for greater reductions in fibrillar amyloid burden in the ACC-001+QS-21 10 µg group compared with placebo. Likewise, while not significant, there was a numerical trend of decreased CSF p-tau levels with ACC-001, possibly consistent with a downstream effect in the ACC-001+QS-21 group. Insufficient antibody titers or quality, insufficient power to detect a difference, or too short duration of follow up may be reasons why a statistically significant response was not observed. Brain volume measures showed faster volume loss in the 10 µg treatment group, similar to the effect seen in few earlier AD immunotherapy trials which may suggest removal of amyloid and resultant decrease in inflammation. No new, unexpected safety signals were detected.

Extracellular Tau Oligomers Produce An Immediate Impairment of LTP and Memory.

Non-fibrillar soluble oligomeric forms of amyloid-ß peptide (oAß) and tau proteins are likely to play a major role in Alzheimer's disease (AD). The prevailing hypothesis on the disease etiopathogenesis is that oAß initiates tau pathology that slowly spreads throughout the medial temporal cortex and neocortices independently of Aß, eventually leading to memory loss. Here we show that a brief exposure to extracellular recombinant human tau oligomers (oTau), but not monomers, produces an impairment of long-term potentiation (LTP) and memory, independent of the presence of high oAß levels. The impairment is immediate as it raises as soon as 20 min after exposure to the oligomers. These effects are reproduced either by oTau extracted from AD human specimens, or naturally produced in mice overexpressing human tau. Finally, we found that oTau could also act in combination with oAß to produce these effects, as sub-toxic doses of the two peptides combined lead to LTP and memory impairment. These findings provide a novel view of the effects of tau and Aß on memory loss, offering new therapeutic opportunities in the therapy of AD and other neurodegenerative diseases associated with Aß and tau pathology.

Increased androgen binding capacity in sebaceous glands in scalp of male-pattern baldness.

Sebaceous glands were isolated by manual dissection under a microscope from surgical specimens of scalp skin with male pattern baldness and skin specimens of hairy and bald scalp obtained at autopsy. The 800 X g pellet (nuclear fraction) and the 164,000 X g supernatant fraction (cytosol) of homogenates of the sebaceous glands were used for measurements of androgen binding characteristics, using dextran-coated charcoal and sucrose gradient methods. Scatchard plots showed high affinity binding for [3H]dihydrotestosterone (DHT) and [3H]methyltrienolone (R1881). Nuclei prepared from bald scalp contained greater total androgen binding capacity than nuclei of hairy scalp, although Kd values of type I binding were similar (0.68 vs 0.56 nM, respectively). On sucrose gradient, the binding protein from cytosol was found in the 7 to 8S density range. Androgen binding by cytosol of sebaceous glands of hairy scalp had Kd of 1.89 +/- .79 and 2.05 +/- .56 nM for DHT and R1881, respectively, and Bmax of 18.7 +/- 4.4 and 20.0 +/- 4.6 fmol/mg protein for DHT and R1881, respectively. Cytosol from sebaceous glands of bald scalp had Kd values approximately half those of hairy scalp, and Bmax values 50%-100% higher. The bound 3H labeled DHT and R1881 could be partially displaced by testosterone (40-50%), moxestrol (28-32%), promegestone (19-26%), and delta 4-androstenedione (6-12%), but not by dehydroepiandrosterone. These data demonstrate the presence of specific androgen binding protein in sebaceous glands, and that sebaceous glands of bald scalp have greater binding affinity and capacity for androgens than those in hairy scalp. This difference may explain the greater androgenic response in androgenic alopecia.

delta 5-3 beta-hydroxysteroid dehydrogenase activity in sebaceous glands of scalp in male-pattern baldness.

Sebaceous glands were isolated by manual dissection using a stereomicroscope from skin specimens of bald scalp of men with male-pattern baldness undergoing hair transplant or scalp reduction surgery and also from specimens taken from hairy and bald areas of scalp at autopsy of adult male victims of accidental death within 3 h post mortem. Homogenates of the isolated glands exhibited activities of delta 5-3 beta-hydroxysteroid dehydrogenase (3 beta HSD), 17 beta-hydroxysteroid dehydrogenase, and testosterone 5 alpha-reductase by the conversion of [3H]dehydroepiandrosterone (DHA) to 3H-delta 4-androstenedione (AD), [3H]testosterone, and [3H]dihydrotestosterone. Homogenates of glands from bald (B) scalp had greater 3 beta HSD activity than homogenates of glands from hairy (H) scalp. After differential centrifugation, 3 beta HSD activity was found mainly in the microsomal and 105,000 X g supernatant fractions. Specific activity of the enzyme based on protein mass was highest in the microsomal fraction; however, the total 3 beta HSD activity in the 105,000 X g supernatent of B glands was significantly (p less than .01) greater than that of H glands. 3 beta HSD activity in sebaceous glands isolated from autopsy specimens did not differ from that of glands isolated from surgical specimens in apparent Km (0.13-0.14 microM), pH optima (8.0), or coenzyme requirement for NAD. Since substantial 3 beta HSD activity was present in the cytosol, and cytosol of B glands showed increased 3 beta HSD activity, the increased conversion of DHA to AD may be a critical step for androgenic action and may be responsible for excessive androgenicity in male-pattern baldness.

Apolipoprotein E epsilon 4, other risk factors, and course of Alzheimer's disease.

BACKGROUND: The epsilon 4 allele of apolipoprotein E (apoE epsilon 4) is associated with late-onset Alzheimer's disease (AD), but its relationship to various aspects of AD has become increasingly unclear. We studied the relationship of apoE genotype in AD to educational attainment, history of heart disease or head injury, age of onset, gender, severity of illness, depression, psychotic symptoms, rate of dementia progression, and time from initial evaluation to nursing home placement. METHODS: ApoE epsilon 4 genotype was determined for 97 clinically diagnosed AD patients and 61 neuropathologically confirmed cases of AD. RESULTS: Presence of one or more epsilon 4 alleles occurred in 66% of AD cases as compared with 27% in control subjects (allele frequency was .40 for AD, .15 for control subjects). Among AD subjects there was no significant relationship between epsilon 4 alleles and educational attainment, history of heart disease, head injury, age of onset, severity of illness, depression, history of depression, rate of dementia progression, or time to nursing home placement. Marginal correlations emerged between number of epsilon 4 alleles, and delusions (p = .05) and hallucinations (p = .05). There was a trend toward increased epsilon 4 homozygosity in patients with onset between ages 65 and 70 years. CONCLUSIONS: We did not find that individuals with one or two apoE epsilon 4 alleles differed significantly in clinical course of AD from those without epsilon 4 except for a trend toward increased psychotic symptoms in the group as a whole and an increase in epsilon 4 homozygosity in patients with reported symptom onset in the late 60s.

Event-related potential P300 in multiple sclerosis. Relation to magnetic resonance imaging and cognitive impairment.

Cerebral involvement in multiple sclerosis may result not only in sensory and motor symptoms but also in impaired mentation. We hypothesize that cognitive dysfunction occurs due to cortical deafferentation or disconnection arising from subcortical white-matter disease. We examined the P300 event-related potential in 31 patients with multiple sclerosis, correlating it with disease severity ratings based on magnetic resonance imaging signal intensity changes, cognitive dysfunction, and disability status. The patients with multiple sclerosis exhibited significantly prolonged P300 wave latencies compared with 32 control subjects. The P300 latency was strongly correlated with the presence of demyelinative brain lesions seen on magnetic resonance imaging scans and with cognitive impairment, but was only weakly associated with the Kurtzke disability status score, consistent with this scale primarily reflecting spinal rather than cerebral demyelination. Our study results support a relationship between subcortical white-matter lesions and cognitive impairment in multiple sclerosis.

Altered amyloid protein processing in platelets of patients with Alzheimer disease.

BACKGROUND: beta-Amyloid peptide, the core component of neuritic plaques in brain areas in patients with Alzheimer disease (AD), is 1 cleavage product of the beta-amyloid precursor protein (APP) in neurons and platelets. Alternate cleavage products of intact 140- to 150-kd APPs in platelets include nonamyloidogenic 120- to 130-kd and 110-kd isoforms. The possible differential significance of these 2 isoforms, structurally similar to protease nexin II, is unknown. OBJECTIVE: To determine whether the ratio of the 120- to 130-kd APP isoform to the 110-kd APP isoform as processed in platelets correlates with the presence of AD and/or the apolipoprotein E4 (ApoE4) allele, which is a major risk factor for AD. SETTING: The Alzheimer Disease Center at The University of Texas Southwestern Medical Center at Dallas. METHODS: The APP isoforms were quantitated with the use of 2 different Western blot detection methods in platelets from 15 patients with AD and 19 control subjects in whom genotyping of apolipoprotein E was performed. RESULTS: The mean ratio of the 120- to 130-kd APP isoform to the 110-kd APP isoform in the patients with AD was significantly lower than that of the control subjects (5.98 vs 7.64; P = .03 [method 1] and 5.98 vs 7.92; P = .01 [method 2]) after adjusting for age and the increased incidence of ApoE4 in patients with AD. The lower APP ratios were also associated with increased age and with the presence of an ApoE4 allele. CONCLUSIONS: The APP processing in platelets of patients with AD is different from that of control subjects. This difference, largely caused by factors other than the ApoE4 genotype, may reflect chronic platelet activation in patients with AD. The use of these data to estimate "AD risk," by using the APP isoform ratio, indicates an odds ratio of 1.75, suggesting possible utility as an adjunct in the diagnosis of AD. Moreover, these findings may relate to analogous alterations in APP processing that may occur in brain areas affected by AD.

Genetic factors for the development of Alzheimer disease in the Cherokee Indian.

OBJECTIVE: To study the relationship between the genetic degree of Cherokee ancestry, the apolipoprotein E *E4 (APOE*E4) allele type, and the development of Alzheimer disease (AD) in individuals from the Cherokee Nation who reside in northeastern Oklahoma. SETTING: Alzheimer disease center satellite clinic and university departments of neurology, psychiatry, and academic computing. DESIGN: Standardized dementia evaluations based on criteria from the National Institute of Neurological and Communicative Disorders and Stroke and the Alzheimer's Disease and Related Disorders Association were performed on 26 patients aged 65 years or older to establish a diagnosis of AD. Twenty-six control subjects were recruited and similarly assessed. The APOE allele type determinations were obtained on all patients and control subjects. Appropriate statistical analyses were used to compare the genetic degree of Cherokee ancestry, the APOE allele type, and the development of AD. RESULTS: The data indicated that as the genetic degree of Cherokee Indian ancestry increased, the representation of AD decreased. The 9 patients with AD with a greater than 50% genetic degree of Cherokee ancestry constituted 35% of the group with AD. The 17 remaining patients with AD who were less than 50% Cherokee constituted 65% of the group with AD. In contrast, 17 (65%) of the control subjects were more than 50% Cherokee; only 9 (35%) were less than 50% Cherokee. These percentages of AD were not changed by the *E4 allele. This inverse relationship between the genetic degree of Cherokee ancestry and AD, independent of the APOE*E4 allele status, diminished with increasing age, suggesting an age-related protective effect of being Cherokee. For a decrease of 10% in Cherokee ancestry, the odds of developing AD are estimated to be 9.00 times greater at age 65 years but only 1.34 times greater at age 80 years. CONCLUSIONS: A greater genetic degree of Cherokee ancestry reduces the risk of developing AD and, thus, seems protective. This protective genetic factor is independent of APOE allele type and diminishes with age.

Characterization of initiated cells in N-methylnitrosourea-induced carcinogenesis of the CNS in the adult rat.

Glial tumors may originate from the malignant transformation of multipotent glial progenitor cells, but tools to study malignant transformation leading to gliomas are limited by the lack of biological systems that represent early stages of this disease in adult animals. In order to characterize the initiated cells that give rise to gliomas, we have employed the N-methylnitrosourea (MNU) model for induction of brain tumors in adult rats (Rushing et al., 1998). Specifically, we have isolated and cultured transformed (premalignant) cells from normal-appearing brains of rats exposed to MNU for 10 weeks and from histologically abnormal brains of rats exposed to MNU for 15 weeks. We compared them with cells cultured from control animals under identical conditions. Cultured cells were classified according to their morphology, immunophenotype, karyotype, proliferation capacity, and tumorigenicity in athymic mice. Cultures from untreated normal rat brains grew as monolayers and had normal karyotypes (42 X,Y), epithelioid morphology, and slow proliferative capacity (doubling time > 120 h). In contrast, cultured cells from brains of MNU-exposed animals had karyotypes that ranged from normal to highly aneuploid. Aneuploid lines grew rapidly in multilayers (doubling time < 24 h), had differentiated astrocytic or oligodendroglial morphology and immunohistochemical staining profile, and yielded tumors in athymic mice. Initiated cells with minor chromosomal aberrations assumed mixed bipolar or tripolar morphologies in high density cultures, proliferated rapidly, but showed contact inhibition and failed to induce tumors when injected s.c. in athymic mice. In general, lines showing no evidence of chromosomal aberrations had the most epithelioid morphology, proliferated slowly (doubling time > 72 h), and retained strict contact growth inhibition. The presumed undifferentiated glial progenitor cells in culture from either control or MNU-treated rats variably expressed markers such as vimentin, nestin, and NG2 proteoglycan, and they weakly expressed the mature astrocytic or oligodendroglial markers glial fibrillary acidic protein or galactocerbroside, respectively. These cultures differentiated to bipolar-tripolar morphology with concomitant maturation to a GFAP+ or GalC+ phenotype upon exposure to secondary messengers such as dibutyryl-cyclic-AMP and/or growth factors such as basic fibrillary growth factor. Continuous stimulation with these messengers resulted in terminal differentiation and consequent death upon withdrawal of the stimulus. These results provide information that could lead to detailed characterization of initiated, premalignant cells in the adult brain and to a better understanding of glial carcinogenesis.

Glutamate transporter EAAT2 splice variants occur not only in ALS, but also in AD and controls.

OBJECTIVE: To ascertain the specificity of alternatively spliced mRNA variants of the astroglial glutamate transporter EAAT2 for ALS. BACKGROUND: An important hypothesis for ALS pathogenesis is that motor neuron injury may result from chronically elevated glutamate levels in the CNS. Supporting this idea are reports of decreased glutamate transport in ALS. This in turn has recently been suggested to be due to the presence of aberrant mRNA splice variants for EAAT2 in ALS. METHODS: Postmortem human brain tissue was obtained from different brain regions of patients with ALS, normal controls (NC), and patients with AD and Lewy body dementia (LB)-neurodegenerative diseases in which motor neurons are unaffected. Brain RNA was analyzed for EAAT2 isoforms using reverse transcription PCR and cDNA cloning/sequencing methods. RESULTS: Splice variants lacking exons 7 or 9 were present in ALS brain, as previously reported, but were also present in brains from NC, AD, and LB patients. PCR product sequence analyses from non-ALS brain show variant splicing identical to that reported for ALS. Quantitative PCR analysis shows that these isoforms may be somewhat more abundant in ALS than AD, LB, and NC brains. CONCLUSIONS: EAAT2 mRNA splice variants are found in the brains of NC and AD patients, as in ALS. The authors cannot exclude the possibility that quantitative changes in variant EAAT2 isoforms might relate directly, or indirectly, to ALS pathology. However, the qualitative presence of these "abnormal" EAAT2 splice variants does not appear to be sufficient to explain motor neuron degeneration in ALS.

HTLV-I-associated myelopathy in a Californian: diagnosis by reactivity to a viral recombinant antigen.

A male patient developed leg numbness and weakness, and bowel, bladder, and erectile dysfunction. Examination revealed an isolated thoracic myelopathy, with lower-extremity spasticity, decreased vibration and position sense, hyperreflexia, and Babinski's signs. Serum and CSF showed antibody reactivity to human T-cell lymphotropic virus type I or II (HTLV-I/II), suggesting HTLV-I-associated myelopathy. Antibody reactivity to a unique HTLV-I recombinant protein provided definitive diagnosis of HTLV-I infection.

Apoptosis and neurologic disease.

Many neurological disorders involve cell death. During development of the nervous system, cell death is a normal feature. Elimination of substantial numbers of initially generated cells enables useful pruning of "mismatched" or excessive cells produced by exuberance during the proliferative and migratory phases of development. Such cell death, occurring by "programmed" pathways, is termed apoptosis. In mature organisms, cells die in two major fashions, either by necrosis or apoptosis. In the adult nervous system, because there is little cell production during adulthood, there is little normal cell death. However, neurological disease is often associated with significant neural cell death. Acute disorders, occurring over minutes to hours, such as brain trauma, infarction, hemorrhage, or infection, prominently involve cell death, much of which is by necrosis. Chronic disorders, with relatively slow central nervous system degeneration, may occur over years or decades, but may involve cell losses. Such disorders include motor neuron diseases such as amyotrophic lateral sclerosis (ALS), cerebral dementing disorders such as Alzheimer's disease and frontotemporal dementia, and a variety of degenerative movement disorders including Parkinson's disease, Huntington's disease, and the inherited ataxias. There is evidence that the mechanism of neuronal cell death in these disorders may involve apoptosis. Direct conclusive evidence of apoptosis is scarce in these chronic disorders, because of the swiftness of cell death in relation to the slowness of the disease. Thus, at any particular time point of assessment, very few cells would be expected to be undergoing death. However, it is clearly of importance to define the type of cell death in these disorders. Of significance is that while treating the underlying causes of these conditions is an admirable goal, it may also be possible to develop productive therapies based on alleviating the process of cell death. This is particularly likely if this cell loss is through apoptosis, a programmed process for which the molecular cascade is increasingly understood. This article reviews our understanding of apoptosis in the nervous system, concentrating on its possible roles in chronic neurodegenerative disorders.

Parkinsonism during cyclosporine treatment.

Cyclosporine has been associated with various neurological side-effects including postural tremor, seizures, headaches, encephalopathy, cortical blindness, and visual hallucinations. We describe here two patients who developed parkinsonism, with rest tremor and bradykinesia, after receiving cyclosporin A following allogeneic bone marrow transplantation. The patients did not have pre-existing neurological disorders, and had not received significant amounts of dopamine-blocking drugs. One patient improved markedly with Sinemet (carbidopa-levodopa), while the other (who did not tolerate Sinemet) improved with decrease in cyclosporine dosage. The relation of the parkinsonian symptoms to cyclosporine therapy and lack of other evident causes for the symptoms, suggests that parkinsonism may be an occasional consequence of cyclosporine.

Enamel gene products during murine amelogenesis in vivo and in vitro.

Epithelial-mesenchymal interactions regulate determination and differentiation of amelogenesis. Our attention has focused on identification of ameloblast gene products, the regulation of enamel mRNA synthesis, and subsequent translation into enamel proteins in vivo and in vitro. Enamel proteins are the most abundant gene products synthesized in fully-differentiated ameloblasts. Our experimental strategy has been to isolate major proteins, produce antibodies, localize enamel protein antigens during tooth development in vivo as well as in vitro (using serumless, chemically-defined medium), develop an immunoprecipitation assay, isolate poly(A)-products in a cell-free translation system, and then initiate molecular cloning of the corresponding murine enamel gene(s). The major murine enamel mRNA appears to code for a predominant polypeptide of approximately 20,000 MW. Inner-enamel epithelial cells differentiate into ameloblasts, and synthesize and secrete enamel proteins within six d when cap-stage molar tooth organs are cultured in serumless, chemically-defined medium. The regulation of epithelial differentiation under these experimental conditions indicates that epithelial-mesenchymal interactions determine and maintian ameloblast differentiation in vitro.