Eva del Valle

Immunohistochemical study of distribution of apolipoproteins E and D in human cerebral β amyloid deposits

Several molecules are known to be closely associated with amyloid deposits in human brain. Among these, apolipoproteins such as apolipoproteins E (apo E) and J (apo J) have been found in two neuropathological hallmarks of Alzheimers disease (AD) and cerebral amyloid angiopathy (CAA): senile plaques (SPs) and cerebrovascular amyloid. These apolipoproteins may be implicated in amyloid fibrillogenesis. Apo D is a multiligand-multifunctional glycoprotein present in SPs, as we previously reported. The aim of this work is to study the link between immunolocalization of apo E and apo D in AD and CAA brains. Both apolipoproteins were found in all types of SPs, but apo E was observed more often than apo D in mature plaques. Whereas apo E is always located overlapping the amyloid core, apo D seems to situate preferably around and near the amyloid. Immunohistochemistry revealed that these apolipoproteins behave differently in cerebral vessels. Apo E labeling in vessels appears mainly linked to amyloid deposits, whereas apo D shows a distribution almost opposite to that of apo E. This could be an indication of the different roles that each apolipoprotein plays in the pathogenesis of amyloid deposition.

Pattern of apolipoprotein D immunoreactivity in human brain

Presence of intracytoplasmatic apolipoprotein D (apo D), a lipophilic ligand transporter, was investigated in normal human brains between 20 and 55 years, using an anti-human apolipoprotein D antibody and extravidin-biotin-enhanced immunohistochemistry. Apo D immunoreactivity was found in neuroglial cells of white matter in all sampled brain regions studied but also in pial cells and perivascular cells. Immunoreactive neurons do not present a uniform pattern throughout the gray matter. The pons and the brainstem show a high immunoreactivity for apo D in several nuclei (olivary, arciforme, cuneado, raphe). In the cerebellum the immunoreactivity appears in some neurons of the Purkinje layer. Finally in the cerebral cortex apo D positive neurons were not observed. These results suggest that apo D role may vary depending of cellular synthesis or location.

Apolipoprotein D Expression in Human Brain Reactive Astrocytes

Astrocytosis is a hallmark of damage that frequently occurs during aging in human brain. Astrocytes proliferate in elderly subjects, becoming hypertrophic and highly immunoreactive for glial fibrillary acidic protein (GFAP). These cells are one type that actively responds in the repair and reorganization of damage to the neural parenchyma and are a source of several peptides and growth factors. One of these biomolecules is apolipo-protein D (apo D), a member of the lipocalin family implicated in the transport of small hydrophobic molecules. Although the role of apo D is unknown, increments in brain apo D expression have been observed in association with aging and with some types of neuropathology. We have found an overexpression of apo D mRNA in reactive astrocytes by in situ hybridization in combination with immunohistochemistry for apo D in normal aged human brains. The number of double-labeled cells varied according to the cerebral area and the gliosis grade. The possible significance of this increased synthesis of apo D in reactive astrocytes is discussed in relation to the role of apo D in aging and in glial function.

Gender differences in apolipoprotein D expression during aging and in Alzheimer disease

Apolipoprotein D (Apo D) is a lipocalin expressed in a wide variety of mammalian tissues. Different studies have shown that this protein is upregulated in the central nervous system (CNS) in several neuropathological conditions, after traumatic brain injury and in aging. The Apo D promoter shows 3 estrogen response elements and it has been shown that its expression is influenced by estrogens in breast cyst fluid. The aim of this work is to study the possible relationship between gender and Apo D expression in human hippocampus and in the entorhinal and frontal cortices during aging and Alzheimers disease (AD). We visualized Apo D immunohistochemically and then performed a quantification of the chromogen signal strength. Our findings show that Apo D expression is influenced by age, Braak stage, and sex. In most of the studied areas, Apo D expression is increased with age in women but not in men, and in AD progression in both genders. Apo D is always expressed by neurons with no signs of degeneration or death.

Aging and substitutive hormonal therapy influence in regional and subcellular distribution of ERα in female rat brain.

Estrogens are not only critical for sexual differentiation it is well-known for the role of 17β-estradiol (E2) in the adult brain modulating memory, learning, mood and acts as a neuroprotector. E2 exerts its actions through two classical receptors: estrogen receptor alpha (ERα) and estrogen receptor beta (ERβ). The distribution of both receptors changes from one brain area to another, E2 being able to modulate their expression. Among the classical features of aging in humans, we find cognitive impairment, dementia, memory loss, etc. As estrogen levels change with age, especially in females, it is important to know the effects of low E2 levels on ERα distribution; results from previous studies are controversial regarding this issue. In the present work, we have studied the effects of long-term E2 depletion as well as the ones of E2 treatment on ERα brain distribution of ovariectomized rats along aging in the diencephalon and in the telencephalon. We have found that ovariectomy causes downregulation and affects subcellular localization of ERα expression during aging, meanwhile prolonged estrogen treatment produces upregulation and overexpression of the receptor levels. Our results support the idea of the region-specific neuroprotection mechanisms mediated by estradiol.

Immunohistochemical Presence of Apolipoprotein D in Senile Plaques

Abstract Senile plaques are characteristic features of the neuropathology occurring in Alzheimers disease and normal aging brains. These plaques have extracellular amyloid deposits and a variety of amyloid associated molecules. Factors that influence amyloid deposits are incompletely understood, but some molecules likely play an important role. Some apolipoproteins (apolipoprotein E and J) are known to be closely associated with amyloid fibrillogenesis and deposition. To investigate the role of the apolipoproteins in amyloidosis, the localization of apolipoprotein D (apo D) in senile plaques was immunohistochemically studied. We report a novel finding: that polyclonal antibody to human apo D immunostained in all types of senile plaques found in normal aging and Alzheimers disease. This finding implicates apo D as a potentially relevant molecule in senile plaque pathogenesis. (The J Histotechnol 24:45, 2001) Submitted: September 6, 1999; Accepted with revisions January 31, 2000

Amyloid-β25-35 Induces Apolipoprotein D Synthesis and Growth Arrest in HT22 Hippocampal Cells

Apolipoprotein D (ApoD) is a secreted glycoprotein that is markedly induced in several pathological and stressful conditions in the nervous system. In the central nervous system, ApoD expression is upregulated during aging, after traumatic brain injury, and in several human neuropathologies such as Alzheimers disease (AD), where it is found associated with amyloid-β (Aβ) plaques. Recent studies have indicated that ApoD has an important function as a neuroprotective and antioxidant protein. The aim of this work is to study the effect of the peptide fragment Aβ25-35, which is believed to play a major role in the neurodegenerative process of AD, in ApoD expression in a mouse hippocampal cell line. In addition, we studied whether direct addition of exogenous human recombinant ApoD protein has neuroprotective effect against Aβ25-35 treatment on neuronal cells. Our results demonstrate that Aβ25-35 induces ApoD expression in hippocampal cells in response to stress-induced growth arrest. This observed relationship between Aβ and ApoD expression could explain the elevated levels of ApoD found in AD brain, where it may be a neuroprotective molecule in the course of AD, probably related to its lipid transport function or a direct antioxidant property. However, the addition of exogenous human recombinant ApoD does not exert any protective effect, most likely due to its major structural modifications.

Age-related changes of apolipoprotein D expression in female rat central nervous system with chronic estradiol treatment

Aging is associated with a reduction in metabolic functions, increased incidence of neurodegenerative diseases, and memory or cognitive dysfunction. With aging, a decrease in plasma estrogen levels, related to loss of gonadal function, occurs in females. Estrogens have neuroprotective effects and estradiol treatment improves some aspects of neuronal homeostasis affected by aging. In other way, recent studies show that apo D can play a neuroprotective role in some neuropathologies and during aging. The possible relation between estradiol treatment and the expression of apo D, during aging in the CNS, was investigated in female rats. Our results confirm an expression of apo D zone-dependent, in relation with aging, and an overexpression of apo D related to ovariectomy and estradiol treatment. This overexpression strengthens the idea that apo D plays a neuroprotective role in the CNS.

Microtubule interfering agents and KSP inhibitors induce the phosphorylation of the nuclear protein p54nrb, an event linked to G2/M arrest ☆

Microtubule interfering agents (MIAs) are anti-tumor drugs that inhibit microtubule dynamics, while kinesin spindle protein (KSP) inhibitors are substances that block the formation of the bipolar spindle during mitosis. All these compounds cause G2/M arrest and cell death. Using 2D-PAGE followed by Nano-LC-ESI-Q-ToF analysis, we found that MIAs such as vincristine (Oncovin) or paclitaxel (Taxol) and KSP inhibitors such as S-tritil-l-cysteine induce the phosphorylation of the nuclear protein p54(nrb) in HeLa cells. Furthermore, we demonstrate that cisplatin (Platinol), an anti-tumor drug that does not cause M arrest, does not induce this modification. We show that the G2/M arrest induced by MIAs is required for p54(nrb) phosphorylation. Finally, we demonstrate that CDK activity is required for MIA-induced phosphorylation of p54(nrb).

Could Apolipoprotein D be a Neuronal Marker of Necrobiosis

Abstract Apolipoprotein D (apo D) is a member of the lipocalin family that is involved in the transport of small hydrophobic ligands, although its physiological functions are yet well known. Using an anti-human apo D antibody and PAP immunocytochemistry on normal human cerebellum tissue, we found that a marked increase in apo D immunoreactivity occurs in human cerebellum during aging. Besides increased immunoreactivity levels in glial cells, we observed a major number of stained Purkinje cells. Morphometric studies were performed to study if quantitative deifferences exist between apo D positive and negative Purkinje cells. These 2 neuron populations presented the same morphological characteristics, and there were no structural evidences of cell injury or death. The data showed significant differences (p < 0.05) for mean cellular area and circular factors; neurons containing apo D were bigger and more rounded than neurons without apo D. These rescllts could be better understood if apo D were considered a marker of necrobiosis more than a marker of irreversible cell death. (The J Histotechnol 24:29, 2001) Submitted: May 15,2000; Accepted with revisions: December 1, 2000