Jens Husemann

Adult mouse astrocytes degrade amyloid-β in vitro and in situ

Alzheimer disease (AD) is a progressive neurodegenerative disorder characterized by excessive deposition of amyloid-β (Aβ) peptides in the brain. One of the earliest neuropathological changes in AD is the accumulation of astrocytes at sites of Aβ deposition, but the cause or significance of this cellular response is unclear. Here we show that cultured adult mouse astrocytes migrate in response to monocyte chemoattractant protein-1 (MCP-1), a chemokine present in AD lesions, and cease migration upon interaction with immobilized Aβ1–42. We also show that astrocytes bind and degrade Aβ1–42. Astrocytes plated on Aβ-laden brain sections from a mouse model of AD associate with the Aβ deposits and reduce overall Aβ levels in these sections. Our results suggest a novel mechanism for the accumulation of astrocytes around Aβ deposits, indicate a direct role for astrocytes in degradation of Aβ and implicate deficits in astroglial clearance of Aβ in the pathogenesis of AD. Treatments that increase removal of Aβ by astrocytes may therefore be a critical mechanism to reduce the neurodegeneration associated with AD.

Scavenger receptors in neurobiology and neuropathology: Their role on microglia and other cells of the nervous system

Scavenger receptor class A (SR‐A, CD204), scavenger receptor‐BI (SR‐BI), and CD36 are cell surface proteins that mediate cell adhesion to, and endocytosis of, various native and pathologically modified substances, and participate in intracellular signaling, lipid metabolism, and host defense against bacterial pathogens. Microglia, Mato cells, astrocytes, cerebral microvascular endothelial cells, cerebral arterial smooth muscle cells, and retinal pigment epithelial cells express one or more of these SR. Expression of SR‐A and SR‐BI by microglia is developmentally regulated. Neonatal microglia express SR‐A and SR‐BI, while microglia in normal mouse and human adult brain express neither. Astrocytes in adult brain express SR‐BI. In Alzheimers disease, microglial expression of SR‐A is increased. Such findings, and evidence that SR‐A and SR‐BI mediate adhesion and endocytosis of fibrillar β‐amyloid by microglia and astrocytes, respectively, and that SR‐A, SR‐BI, and CD36 participate in secretion of reactive oxygen species by microglia, suggest roles for these receptors in homeostasis and neuropathology. GLIA 40:195–205, 2002.

CD36, a class B scavenger receptor, is expressed on microglia in Alzheimer's disease brains and can mediate production of reactive oxygen species in response to β-amyloid fibrils

A pathological hallmark of Alzheimers disease is the senile plaque, composed of beta-amyloid fibrils, microglia, astrocytes, and dystrophic neurites. We reported previously that class A scavenger receptors mediate adhesion of microglia and macrophages to beta-amyloid fibrils and oxidized low-density lipoprotein (oxLDL)-coated surfaces. We also showed that CD36, a class B scavenger receptor and an oxLDL receptor, promotes H(2)O(2) secretion by macrophages adherent to oxLDL-coated surfaces. Whether CD36 is expressed on microglia, and whether it plays a role in secretion of H(2)O(2) by microglia interacting with fibrillar beta-amyloid is not known. Using fluorescence-activated cell sorting analysis and immunohistochemistry, we found that CD36 is expressed on human fetal microglia, and N9-immortalized mouse microglia. We also found that CD36 is expressed on microglia and on vascular endothelial cells in the brains of Alzheimers disease patients. Bowes human melanoma cells, which normally do not express CD36, gained the ability to specifically bind to surfaces coated with fibrillar beta-amyloid when transfected with a cDNA encoding human CD36, suggesting that CD36 is a receptor for fibrillar beta-amyloid. Furthermore, two different monoclonal antibodies to CD36 inhibited H(2)O(2) production by N9 microglia and human macrophages adherent to fibrillar beta-amyloid by approximately 50%. Our data identify a role for CD36 in fibrillar beta-amyloid-induced H(2)O(2) production by microglia, and imply that CD36 can mediate binding to fibrillar beta-amyloid. We propose that similar to their role in the interaction of macrophages with oxLDL, class A scavenger receptors and CD36 play complimentary roles in the interactions of microglia with fibrillar beta-amyloid.

Scavenger receptor class B type I (SR-BI) mediates adhesion of neonatal murine microglia to fibrillar β-amyloid

Class A scavenger receptors (SR-A) mediate microglial interaction with fibrillar beta-amyloid (fAbeta). We report here that neonatal microglia from SR-A knockout mice (SR-A-/-) adhere to surface-bound fAbeta, and produce reactive oxygen species (ROS) as efficiently as wildtype microglia; that both wildtype and SR-A-/- microglia express SR-BI; that antibodies against SR-BI do not affect adhesion or ROS production by wildtype microglia, but inhibit adhesion and ROS production of SR-A-/- microglia to immobilized fAbeta by approximately 40%. Adhesion to fAbeta-coated surfaces, and uptake of fAbeta by both wildtype and SR-A-/- microglia was almost completely inhibited by incubation with fucoidan. Thus SR-BI and SR-A mediate similar effector functions in neonatal microglia, which suggests that SR-BI plays as important a role as SR-A, and can maintain the wildtype phenotype in SR-A-/- microglia.

Expression of scavenger receptor class B, type I, by astrocytes and vascular smooth muscle cells in normal adult mouse and human brain and in Alzheimer's disease brain.

In Alzheimers disease (AD), fibrillar beta-amyloid protein (fAbeta) accumulates in the walls of cerebral vessels associated with vascular smooth muscle cells (SMCs), endothelium, and pericytes, and with microglia and astrocytes in plaques in the brain parenchyma. Scavenger receptor class A (SR-A) and class B, type I (SR-BI) mediate binding and ingestion of fAbeta by cultured human fetal microglia, microglia from newborn mice, and by cultured SMCs. Our findings that SR-BI participates in the adhesion of cultured microglia from newborn SR-A knock-out mice to fAbeta-coated surfaces, and that microglia secrete reactive oxygen species when they adhere to these surfaces prompted us to explore expression of SR-BI in vivo. We report here that astrocytes and SMCs in normal adult mouse and human brains and in AD brains express SR-BI. In contrast, microglia in normal adult mouse and human brains and in AD brains do not express SR-BI. These findings indicate that SR-BI may mediate interactions between astrocytes or SMCs and fAbeta, but not of microglia and fAbeta, in AD, and that expression of SR-BI by rodent microglia is developmentally regulated. They suggest that SR-BI expression also is developmentally regulated in human microglia.

Statin Inhibition of Fc Receptor–Mediated Phagocytosis by Macrophages Is Modulated by Cell Activation and Cholesterol

Objectives—An inflammatory response to altered lipoproteins that accumulate in the arterial wall is a major component of the pathogenesis of atherosclerosis. Statins reduce plasma levels of low-density lipoprotein (LDL) and are effective treatments for atherosclerosis. It is hypothesized that they also modulate inflammation. The aim of this study was to examine whether lovastatin inhibits macrophage inflammatory processes and clarify its mechanism of action. Methods and Results—We examined the effects of statins on phagocytosis of antibody-coated red blood cells by cultured human monocytes and mouse peritoneal macrophages. Lovastatin, simvastatin, and zaragozic acid, a squalene synthase inhibitor, blocked Fc receptor–mediated phagocytosis by cultured human monocytes and mouse peritoneal macrophages. The inhibitory effect of lovastatin on Fc receptor–mediated phagocytosis was prevented completely by addition of mevalonate, farnesyl pyrophosphate, LDL, or cholesterol to the culture medium. The inhibitory effect of zaragozic acid was reversed by addition of LDL, but not by the addition of geranylgeranyl pyrophosphate, to the medium. In addition, the effect of lovastatin on phagocytosis is a function of cell activation because treatment of cells with tumor necrosis factor-&agr; or lipopolysaccharide prevented inhibition of phagocytosis by lovastatin. Conclusions—The inhibition of Fc receptor–mediated phagocytosis of lovastatin is related to its effect on cholesterol biosynthesis rather than its effect on the formation of isoprenoids.

Acute Cardiac Allograft Rejection in Nitric Oxide Synthase-2−/− and Nitric Oxide Synthase-2+/+ Mice Effects of Cellular Chimeras on Myocardial Inflammation and Cardiomyocyte Damage and Apoptosis

Background—The contribution of nitric oxide synthase (NOS)-2 to myocardial inflammation and cardiomyocyte necrosis and apoptosis during allograft rejection was investigated through heterotopic cardiac transplantation in mice. Methods and Results—In the first experiments, hearts from C3H donor mice were transplanted into NOS-2−/− and NOS-2+/+ C57BL/6J.129J recipients. A second series of experiments included NOS-2−/− donor hearts transplanted into NOS-2−/− recipients and wild-type NOS-2+/+ donor hearts transplanted into wild-type NOS-2+/+ recipients. (All donors were C57BL/6J and recipients were C57BL/6J.129J.) In the first series of experiments, no significant differences were observed in allograft survival, rejection score, total number of apoptotic nuclei (TUNEL), total number of apoptotic cardiomyocytes, or graft NOS-2 mRNA and protein. Positive NOS-2 immunostaining occurred in endothelial cells and cardiomyocytes in the allografts; the inflammatory infiltrate was NOS-2 positive only when recipients were NOS-2+/+. In the second series of experiments, cardiac allograft survival was significantly increased in the NOS-2−/− mice (26±13 versus 17±8 days, P <0.05), along with significant reductions in inflammatory infiltrate, rejection score, and total number of apoptotic nuclei (23.5±9.5 versus 56.4±15.3, P <0.01) and of apoptotic cardiomyocytes (2.9±1.6 versus 6.9±2.7, P <0.05). No NOS-2 or nitrotyrosine, a marker of peroxynitrite exposure, was detected in NOS-2−/− allografts transplanted into NOS-2−/− recipients. ConclusionsThe data suggest that NO derived from NOS-2 contributes to the inflammatory response and to cardiomyocyte damage and apoptosis during acute cardiac allograft rejection.

AbstractPoster presentation: Tuesday posterP3-287: Effects of lovastatin on secretion of MCP-1 and nitric oxide by cultured adult mouse astrocytes stimulated with IL-1B or TNF-A

rhesus monkeys and the effects on fA toxicity and recruitment of microglia examined. To determine the possible mechanism of microglia contribution to fA toxicity, generation of reactive oxygen species (ROS) in response to fA was investigated in microglia isolated from aged rhesus cortex. Results: fA resulted in significantly larger stereologically determined volume of damage in aged rhesus cortex when compared with injections of vehicle. Co-injection of MIF with fA or delivery of MIF via Alzet pump resulted in a dose-dependent reduction in the number of microglia and the volume of damage caused by fA determined using unbiased stereological methods. Microglia isolated from the aged rhesus cortex displayed robust production of ROS following stimulation with fA . Significantly, MIF resulted in a dose dependent reduction in the production of ROS by aged rhesus microglia following fA stimulation. Conclusions: These observations indicate that fA , which is deposited in plaques in Alzheimer’s disease, is likely to exert toxic effects on aging neurons in the primate. Microglia appear to make a major contribution to the age-related vulnerability of neurons to fA toxicity in the primate brain, most likely through enhanced production of reactive oxygen species. The absence of microglia activation following low dose fA injections in the young rhesus and inhibition of toxicity of such injections in the aged rhesus following MIF treatment suggest that aged primate microglia may display an enhanced response to fA stimulation.

P4-421: Effects of small anionic sulfates and sulfonates on amyloid-beta/heparan sulfate proteoglycan (HSPG) interaction in an in vitro model of amyloid deposition

cannabinoid receptor agonist, WIN-55212-2, on microglia activation induced by a chronic intracerebroventricular lipopolysaccharide (LPS) infusion and its possible mechanisms of action. Methods: Daily injections of WIN-55212-2 (0.5 or 1.0 mg/kg, i.p.) were given to three months old male rats implanted into the 4 ventricle with a chronic indwelling cannula connected to an osmotic minipump containing LPS (250 ng/hr) or vehicle. During the third week of treatment, the rats were behaviorally tested using the Morris water pool task. Results: Animals treated with WIN demonstrated a significant anti-inflammatory effect represented by a reduction in microglia activation. LPS-infused rats treated with WIN-55212-2 did not demonstrate impaired performance in the Morris water pool task as compared to the LPS-infused rats without WIN-55212-2. We are currently investigating the endocannabinoid receptor subtype that is responsible for this anti-inflammatory action and whether specific regions of the brain are more vulnerable to the consequences of the inflammation. Conclusions: These results highlight the potential benefits of pharmacological manipulation of endocannabinoid receptor within the brain and may guide the design of more effective therapies for AD.