N-terminal heterogeneity of parenchymal and cerebrovascular Abeta deposits

The goals of this study were twofold: to determine whether species differences in Abeta N-terminal heterogeneity explain the absence of neuritic plaques in the aged dog and aged bear in contrast to the human; and to compare Abeta N-terminal isoforms in parenchymal vs cerebrovascular Abeta (CVA) deposits in each of the species, and in individuals with Alzheimer disease (AD) vs nondemented individuals. N-terminal heterogeneity can affect the aggregation, toxicity, and stability of Abeta. The human, polar bear, and dog brain share an identical Abeta amino acid sequence. Tissues were immunostained using affinity-purified polyclonal antibodies specific for the L-aspartate residue of Abeta at position one (AbetaN1[D]), D-aspartate at N1 (AbetaN1[rD]), and pyroglutamate at N3 (AbetaN3[pE]) and p3, a peptide beginning with leucine at N17 (AbetaN17[L]). The results demonstrate that each Abeta N-terminal isoform can be present in diffuse plaques and CVA deposits in AD brain, nondemented human, and the examined aged animal models. Though each Abeta N-terminal isoform was present in diffuse plaques, the average amyloid burden of each isoform was highest in AD vs polar bear and dog (beagle) brain. Moreover, the ratio of AbetaN3(pE) (an isoform that is resistant to degradation by most aminopeptidases) vs AbetaN17(L)-x (the potentially nonamyloidogenic p3 fragment) was greatest in the human brain when compared with aged dog or polar bear. Neuritic plaques in AD brain typically immunostained with antibodies against AbetaN1(D) and AbetaN3(pE), but not AbetaN17(L) or AbetaN1(rD). Neuritic deposits in nondemented individuals with atherosclerotic and vascular hypertensive changes could be identified with AbetaN1(D), AbetaN3(pE), and AbetaN1(rD). The presence of AbetaN1(rD) in neuritic plaques in nondemented individuals with atherosclerosis or hypertension, but not in AD, suggests a different evolution of the plaques in the two conditions. AbetaN1(rD) was usually absent in human CVA, except in AD cases with atherosclerotic and vascular hypertensive changes. Together, the results demonstrate that diffuse plaques, neuritic plaques, and CVA deposits are each associated with distinct profiles of Abeta N-terminal isoforms.     

Astrocyte-derived monocyte-chemoattractant protein-1 directs the transmigration of leukocytes across a model of the human blood-brain barrier

The migration of leukocytes across the blood-brain barrier (BBB) into the central nervous system is critical in the pathogenesis of central nervous system inflammatory diseases. The production of chemokines, such as monocyte-chemoattractant protein-1 (MCP-1), by endothelial cells (EC) and astrocytes may initiate and amplify this process. Using a coculture of human EC and astrocytes to model the BBB, we demonstrated that exogenous MCP-1 induces the transmigration of monocytes in a dose-dependent manner. TNF-alpha, IFN-gamma, or IL-1beta treatment of cocultures also induced significant migration of monocytes that correlates with the induction of MCP-1 protein. TGF-beta, previously shown to induce MCP-1 expression in astrocytes, but not in EC, caused migration of monocytes across cocultures, but not across EC grown alone. Monocytes and lymphocytes transmigrated across cytokine-treated cocultures in greater numbers than across EC alone. Astrocytes were the main source of cytokine-induced MCP-1, supporting a role for astrocytes in facilitating leukocyte transmigration. A blocking Ab to MCP-1 inhibited MCP-1- and cytokine-induced transmigration of monocytes by 85-90%. Cytokine treatment of cocultures also resulted in the transmigration of activated, CD69-positive lymphocytes. The MCP-1-mediated transmigration of monocytes across cocultures was blocked using an Ab to ICAM-1 and inhibited by 55% using an Ab to E-selectin. These data suggest a central role for astrocyte-derived MCP-1 in directing the migration of monocytes and lymphocytes across the BBB.     

Dermatofibrosarcoma protuberans and the Bednar tumor: treatment in the pediatric population

Dermatofibrosarcoma protuberans (DFSP) is a low-intermediate-grade cutaneous sarcoma that has a marked propensity for local recurrence after excision. The Bednar variant of this tumor is even less common and is distinguished histologically by the dispersal of melanin containing cells in an otherwise typical DFSP. Both are considered to be tumors of the third and forth decades of life, but both DFSP and the Bednar variant have been described in children. Until this report of a congenital Bednar tumor, only the DFSP has also been described in the neonate. The histopathology and surgical management of DFSP and Bednar tumors are outlined with emphasis on reported experience in the pediatric population. The surgical management of these lesions in children is based on numerous series in adults and the limited pediatric experience. The recommended treatment is wide excision with 3-cm margins of visibly uninvolved tissue and inclusion of superficial fascia.     

Glucose transporter (GLUT-4) is targeted to secretory granules in rat atrial cardiomyocytes

The insulin-responsive glucose transporter GLUT-4 is found in muscle and fat cells in the trans-Golgi reticulum (TGR) and in an intracellular tubulovesicular compartment, from where it undergoes insulin-dependent movement to the cell surface. To examine the relationship between these GLUT-4-containing compartments and the regulated secretory pathway we have localized GLUT-4 in atrial cardiomyocytes. This cell type secretes an antihypertensive hormone, referred to as the atrial natriuretic factor (ANF), in response to elevated blood pressure. We show that GLUT-4 is targeted in the atrial cell to the TGR and a tubulo-vesicular compartment, which is morphologically and functionally indistinguishable from the intracellular GLUT-4 compartment found in other types of myocytes and in fat cells, and in addition to the ANF secretory granules. Forming ANF granules are present throughout all Golgi cisternae but only become GLUT4 positive in the TGR. The inability of cyclohexamide treatment to effect the TGR localization of GLUT-4 indicates that GLUT-4 enters the ANF secretory granules at the TGR via the recycling pathway and not via the biosynthetic pathway. These data suggest that a large proportion of GLUT-4 must recycle via the TGR in insulin-sensitive cells. It will be important to determine if this is the pathway by which the insulin-regulatable tubulo-vesicular compartment is formed.     

Correlates and consequences of workplace violence

A random telephone survey was used to interview 598 employees about instances, if any, when they had been physically attacked or threatened in the workplace. Demographic and workplace characteristics are analyzed as correlates of these forms of violence. The only characteristic consistently associated with higher risk is a work schedule that included nighttime hours. However, although few strong or consistent correlates of workplace violence were found, there were clear negative consequences associated with being victimized at work: lower job satisfaction, greater job stress, increased considerations of job change, and an increased likelihood of bringing mace, a gun, or another weapon to work.     

Structural features of thyroid hormone response elements that increase susceptibility to inhibition by an RTH mutant thyroid hormone receptor

The chicken lysozyme silencer F2 (F2) thyroid hormone response element (TRE) contains an unusual everted palindromic arrangement, has a high affinity for thyroid hormone receptor (TR) homodimers, and is especially sensitive to dominant negative inhibition by, the T3 resistance (RTH) mutant TR beta P453H. We used various TREs and TR mutations to determine the mechanisms for this sensitivity. Changing the F2 orientation from an everted palindrome to a direct repeat with a 4-bp gap (DR+4) (F2-DR) decreased the sensitivity to inhibition at high T3 concentrations, while a loss of this sensitivity occurred with a palindromic arrangement of these same half-sites. F2 contains the dinucleotide TG 5' to each consensus half-site conforming to the optimal TR-binding octamer, YRRGGTCA. A T to A change in position 1 of both F2 half-sites markedly reduced T3-induction, yet only slightly reduced TR homodimer or TR-retinoid X receptor (RXR) heterodimer binding. The TR beta ninth heptad mutation, L428R, prevents TR heterodimerization with RXR and eliminates the inhibitory effect of the P453H mutant TR on the F2-DR, but not the F2 element. Structural features of a TRE that favor strong TR binding of both TR homodimers and TR-RXR heterodimers containing the mutant TR, such as the everted palindromic conformation or the optimal TR-binding consensus octamer, enhance the sensitivity of a TRE to inhibition by the mutant TR. Thus, both half-site orientation and sequence contribute to the sensitivity of a given TRE to dominant negative inhibition by a mutant TR.     

Par-4 is a mediator of neuronal degeneration associated with the pathogenesis of Alzheimer disease

Prostate apoptosis response-4 (Par-4) is a protein containing both a leucine zipper and a death domain that was isolated by differential screening for genes upregulated in prostate cancer cells undergoing apoptosis. Par-4 is expressed in the nervous system, where its function is unknown. In Alzheimer disease (AD), neurons may die by apoptosis, and amyloid beta-protein (A beta) may play a role in this. We report here that Par-4 expression is increased in vulnerable neurons in AD brain and is induced in cultured neurons undergoing apoptosis. Blockade of Par-4 expression or function prevented neuronal apoptosis induced by Ab and trophic factor withdrawal. Par-4 expression was enhanced, and mitochondrial dysfunction and apoptosis exacerbated, in cells expressing presenilin-1 mutations associated with early-onset inherited AD.     

Conservative prediction of time to Clostridium botulinum toxin formation for use with time-temperature indicators to ensure the safety of foods

Integrating-type time-temperature indicators (TTIs) may be utilized to warn food processors and consumers about storage conditions that may have rendered a food potentially hazardous. As an example of how integrated TTIs could be manufactured to emulate an infinite set of time-temperature situations, a set of conditions which have supported C. botulinum growth and toxin production was compiled. The time-temperature curve representing conservative times required for toxin formation was constructed with data from literature relating to toxin formation as a function of temperature in any media or food product. This set of critical time-temperature data is fit by a conservative empirical relationship that can be used to predict combinations of incubation times and storage temperatures that represent a potential health risk from C. botulinum in foods. A TTI could be constructed to indicate deviation from such a given set of conditions to bring attention to foods that may have been exposed to potentially hazardous temperatures with respect to C. botulinum toxin formation.