Guoji Wang

Chemokines and receptors in HIV encephalitis

Background: Chemokines are involved in the migration of leukocytes and have been implicated in several inflammatory diseases of the central nervous system. Some of their receptors have been proposed to mediate HIV infection. Objective: To determine changes in chemokine and receptor expression in HIV encephalitis, and to determine whether upregulation leads to recruitment of infected monocytes across the blood‐brain barrier and participates in HIV neuropathology. Methods: Immunocytochemistry and double‐label immunofluorescent laser confocal microscopy was performed with antibodies to chemokines and their receptors on brain tissues from patients who died with or without HIV encephalitis. In vivo distribution was compared with in vitro cultures of human neuroglial cells. Results: The β‐chemokines monocyte chemotactic protein‐1, macrophage inflammatory protein‐1α, and RANTES were detected on brain macrophages. Their presence was associated with the histopathological signs of HIV encephalitis. The α‐chemokines IP‐10 (10 kDa inflammatory protein) and interleukin‐8 were expressed by astrocytes in all tissues, including controls. Presence of the CXC‐chemokine receptor (CXCR)‐4 was seen on brain macrophages/microglia, neurons, and astrocytes. CC‐Chemokine receptor (CCR)‐5 was detected only on macrophages/microglia. CCR‐3 and CCR‐1 were expressed by macrophages and endothelial cells. In vitro studies examining the presence of CCR‐3, CCR‐5, and CXCR‐4 on human brain cell cultures demonstrated abundant neuronal and microglial expression.Background:Chemokines are involved in the migration of leukocytes and have been implicated in several inflammatory diseases of the central nervous system. Some of their receptors have been proposed to mediate HIV infection.Objective:To determine changes in chemokine and receptor expression in HIV en

In vivo CHI3L1 (YKL-40) expression in astrocytes in acute and chronic neurological diseases

BackgroundCHI3L1 (YKL-40) is up-regulated in a variety of inflammatory conditions and cancers. We have previously reported elevated CHI3L1 concentration in the cerebrospinal fluid (CSF) of human and non-human primates with lentiviral encephalitis and using immunohistochemistry showed that CHI3L1 was associated with astrocytes.MethodsIn the current study CHI3L1 transcription and expression were evaluated in a variety of acute and chronic human neurological diseases.ResultsELISA revealed significant elevation of CHI3L1 in the CSF of multiple sclerosis (MS) patients as well as mild elevation with aging. In situ hybridization (ISH) showed CHI3L1 transcription mostly associated with reactive astrocytes, that was more pronounced in inflammatory conditions like lentiviral encephalitis and MS. Comparison of CHI3L1 expression in different stages of brain infarction showed that YKL40 was abundantly expressed in astrocytes during acute phases and diminished to low levels in chronic infarcts.ConclusionsTaken together, these findings demonstrate that CHI3L1 is induced in astrocytes in a variety of neurological diseases but that it is most abundantly associated with astrocytes in regions of inflammatory cells.

PK11195 labels activated microglia in Alzheimer's disease and in vivo in a mouse model using PET.

Activated microglia may promote neurodegeneration in Alzheimers disease (AD) and may also help in amyloid clearance in immunization therapies. In vivo imaging of activated microglia using positron emission tomography (PET) could assist in defining the role of activated microglia during AD progression and therapeutics. We hypothesized that PK11195, a ligand that binds activated microglia, could label these cells in postmortem AD tissues and in vivo in an animal model of AD using PET. [(3)H](R)-PK11195 binding was significantly higher in AD frontal cortex compared to controls and correlated mainly with the abundance of immunohistochemically labeled activated microglia. With age, the brains of APP/PS1 transgenic mice showed progressive increase in [(3)H](R)-PK11195 binding and [(11)C](R)-PK11195 retention in vivo assessed using microPET, which correlated with the histopathological abundance of activated microglia. These results suggest that PK11195 binding in AD postmortem tissue and transgenic mice in vivo correlates with the extent of microglial activation and may help define the role of activated microglia in the pathogenesis and treatment of AD.

Expression of HGF and cMet in the developing and adult brain

Hepatocyte growth factor (HGF) was recently recognized as a potential neurotrophic factor in the developing brain. We studied expression of HGF and its receptor using Northern blot analysis and in situ hybridization for mRNA and double immunofluorescent laser confocal microscopy. HGF and cMet messages were abundant in the hippocampus of both human and rat brains. In this region, both messages were localized in the neuronal layer. Segregation of HGF predominantly in the hippocampal CA3-4 and cMet in CA1 supports the hypothesis that HGF may mediate important neurotrophic functions in both developing and adult brains.

The Positron Emission Tomography Ligand DAA1106 Binds With High Affinity to Activated Microglia in Human Neurological Disorders

Chronic microglial activation is an important component of many neurological disorders, and imaging activated microglia in vivo will enable the detection and improved treatment of neuroinflammation. 1-(2-Chlorphenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinoline-carbox-amide (PK11195), a peripheral benzodiazepine receptor ligand, has been used to image neuroinflammation, but the extent to which PK11195 binding distinguishes activated microglia and reactive astrocytes is unclear. Moreover, PK11195 may lack sufficient sensitivity for detecting mild neuroinflammation. We hypothesized that N-(2,5-dimethoxybenzyl)-N-(4-fluoro-2-phenoxyphenyl) acetamide (DAA1106), a new ligand that binds specifically to peripheral benzodiazepine receptor, binds to activated microglia in human neurological diseases with higher affinity than does PK11195. We therefore compared the pharmacological binding properties of [3H](R)-PK11195 and [3H]DAA1106 in postmortem tissues from patients with cerebral infarcts, amyotrophic lateral sclerosis, Alzheimer disease, frontotemporal dementia, and multiple sclerosis (n = 10 each). In all diseases, [3H]DAA1106 showed a higher binding affinity as reflected by lower dissociation constant (KD) values than that of [3H](R)-PK11195. Moreover, specific binding of both ligands correlated with the presence of activated microglia identified by immunohistochemistry in situ. We conclude that 1) ligands that bind peripheral benzodiazepine receptor mainly label activated microglia in human neurological disorders and that 2) DAA1106 may possess binding characteristics superior to those of PK11195, which may be beneficial for in vivo positron emission tomography imaging.

HIV mediates a productive infection of the brain.

BACKGROUND Approximately one quarter of patients with AIDS develop severe cognitive deficits called HIV-associated dementia complex. There is some controversy regarding the importance of viral load and distribution in mediating this neurologic disease. OBJECTIVE Brain HIV proviral and RNA loads were compared to define the molecular nature of HIV infection of the brain. METHOD Neuropathologic examination was performed on brains from 10 autopsies of patients with AIDS that had short post-mortem intervals and no evidence of opportunistic infection. Viral DNA and RNA were extracted and quantified from multiple brain regions. These findings were compared with triple-label immunofluorescence for viral and cell markers. RESULTS Brains with histopathologic evidence of HIV encephalitis contained abundant HIV RNA and DNA. Regions without productive HIV infection showed minimal proviral load. By immunocytochemistry, only brain macrophages/microglia double labeled for viral proteins. CONCLUSIONS HIV mediates a productive infection of brain macrophages/microglia. There was no evidence supporting the hypothesis of substantial neuronal or macroglial infection, or evidence of substantial proviral burden prior to the development of productive infection.

A comparison of the high‐affinity peripheral benzodiazepine receptor ligands DAA1106 and (R)‐PK11195 in rat models of neuroinflammation: implications for PET imaging of microglial activation

Activated microglia are an important feature of many neurological diseases and can be imaged in vivo using 1‐(2‐chlorophenyl)‐N‐methyl‐N‐(1‐methylpropyl)‐3‐isoquinolinecarboxamide (PK11195), a ligand that binds the peripheral benzodiazepine receptor (PBR). N‐(2,5‐dimethoxybenzyl)‐N‐(5‐fluoro‐2‐phenoxyphenyl) acetamide (DAA1106) is a new PBR‐specific ligand that has been reported to bind to PBR with higher affinity compared with PK11195. We hypothesized that this high‐affinity binding of DAA1106 to PBR will enable better delineation of microglia in vivo using positron emission tomography. [3H]DAA1106 showed higher binding affinity compared with [3H](R)‐PK11195 in brain tissue derived from normal rats and the rats injected intrastriatally with 6‐hydroxydopamine or lipopolysaccharide at the site of the lesion. Immunohistochemistry combined with autoradiography in brain tissues as well as correlation analyses showed that increased [3H]DAA1106 binding corresponded mainly to activated microglia. Finally, ex vivo autoradiography and positron emission tomography imaging in vivo showed greater retention of [11C]DAA1106 compared with [11C](R)‐PK11195 in animals injected with either lipopolysaccaride or 6‐hydroxydopamine at the site of lesion. These results indicate that DAA1106 binds with higher affinity to microglia in rat models of neuroinflammation when compared with PK11195, suggesting that [11C]DAA1106 may represent a significant improvement over [11C](R)‐PK11195 for in vivo imaging of activated microglia in human neuroinflammatory disorders.

Induction of cell-cycle regulators in Simian immunodeficiency virus encephalitis

Neuronal degeneration associated with human immunodeficiency virus encephalitis has been attributed to neurotoxicity of signaling molecules secreted by activated, infected macrophages. We hypothesized that the barrage of signals present in the extracellular milieu of human immunodeficiency virus-infiltrated brain causes inappropriate activation of neuronal cell-cycle machinery. We examined the presence of three members of the cell-cycle control machinery: pRb, E2F1, and p53 in the simian immunodeficiency virus encephalitis (SIVE) model. Compared to noninfected and simian immunodeficiency virus-infected, nonencephalitic controls, we observed increased protein expression of E2F1 and p53 and aberrant cellular localization of E2F1 and pRb. In SIVE, E2F1 was abundant in the cytoplasm of neurons in both neurons and astrocytes proximal to SIVE pathology in the basal ganglia. pRb staining was nuclear and cytoplasmic in cortical neurons of SIVE cases. Antibodies to phosphorylated pRb also labeled the cytoplasm of cortical neurons. These data suggest that in SIVE, cell signaling results in phosphorylation of pRb which may result in subsequent alteration in E2F1 activity. As increased E2F1 and p53 activities have been linked to cell death, these data suggest that the neurodegeneration in SIVE could in part be because of changes in expression and activity of cell-cycle machinery.

Fatal fulminant pan-meningo-polioencephalitis due to West Nile virus.

We report a case of fatal fulminant West Nile virus (WNV) meningoencephalitis in an 87‐year‐old white male gardener. The Pennsylvania patient presented with a 3‐day history of flu‐like symptoms. His hospital course was gravely precipitous with onset of coma, ventilator dependence, loss of cortical and brainstem functions within ten days of admission. Acute serum and cerebrospinal fluid samples revealed elevated levels of WNV IgM antibodies by ELISA as well as elevated CSF white blood cells, protein and glucose. A complete autopsy revealed a multifocal lymphocytic myocarditis and severe chronic tubulointerstitial nephritis. Viral culture and PCR analysis of post‐mortem samples of the spleen, kidney and brain were positive for WNV. Histological sections from all regions of the brain and spinal cord demonstrated a severe, non‐necrotizing, subacute, polio‐meningoencephalitis. While both gray and white matter were inflamed, gray matter was much more severely involved. Many gray matter nuclei showed severe neuronal loss with residual dying neurons surrounded by activated microglia. Immunohistochemical stains revealed profuse infiltration of the meninges and cerebral parenchyma by CD8 T‐lymphocytes and perivascular B‐lympho‐cytes. Electron micrographs revealed diffuse intracellular and extracellular edema but no viral particles were identified. Immunohistochemical and immunofluorescent staining for WNV filled the cytoplasm of residual neurons. West Nile virus mediates a predominantly polioencephalitis secondary to direct infection of neurons.

Systemic distribution of west nile virus infection : Postmortem immunohistochemical study of six cases

Rare cases of West Nile virus (WNV)‐associated inflammation outside the central nervous system (CNS) have been reported. We evaluated the systemic distribution of WNV in postmortem tissues during encephalitis in six patients using immunohistochemistry. WNV antigens were detected in neurons of CNS (all 6 cases), kidney (4 cases), lungs (2 cases), pancreas (2 cases), thyroid (2 cases), intestine (2 cases), stomach (1 case), esophagus (1 case), bile duct (1 case), skin (1 case), prostate (1 case) and testis (1 case). In systemic organs epithelial cells were infected. In none of the six cases were viral antigens identified in hepatocytes, heart, adrenal gland, nerves, skeletal muscles, bone, vessels and fat. All cases in which viral antigens were identified in systemic organs in addition to CNS were severely immunocompromised transplant recipients. With the exception of testis and brain, most foci of infection were not associated with inflammation. While the absence of inflammation may in part be due to patient immunosuppression or to possible transient nature of any host response, compartmentalization of viral antigen to the luminal region of epithelial cells may sequester WNV from immune recognition. Comparison of our findings with previous reports suggests that patients with WNV encephalitis can have widespread systemic infection.