Endocan is a novel chondroitin sulfate/dermatan sulfate proteoglycan that promotes hepatocyte growth factor/scatter factor mitogenic activity.

Proteoglycans that modulate the activities of growth factors, chemokines, and coagulation factors regulate in turn the vascular endothelium with respect to processes such as inflammation, hemostasis, and angiogenesis. Endothelial cell-specific molecule-1 is mainly expressed by endothelial cells and regulated by pro-inflammatory cytokines (Lassalle, P., Molet, S., Janin, A., Heyden, J. V., Tavernier, J., Fiers, W., Devos, R., and Tonnel, A. B. (1996) J. Biol. Chem. 271, 20458-20464). We demonstrate that this molecule is secreted as a soluble dermatan sulfate (DS) proteoglycan. This proteoglycan represents the major form either secreted by cell lines or circulating in the human bloodstream. Because this proteoglycan is specifically secreted by endothelial cells, we propose to name it endocan. The glycosaminoglycan component of endocan consists of a single DS chain covalently attached to serine 137. Endocan dose-dependently increased the hepatocyte growth factor/scatter factor (HGF/SF)-mediated proliferation of human embryonic kidney cells, whereas the nonglycanated form of endocan did not. Moreover, DS chains purified from endocan mimicked the endocan-mediated increase of cell proliferation in the presence of HGF/SF. Overall, our results demonstrate that endocan is a novel soluble dermatan sulfate proteoglycan produced by endothelial cells. Endocan regulates HGF/SF-mediated mitogenic activity and may support the function of HGF/SF not only in embryogenesis and tissue repair after injury but also in tumor progression.

The leukocyte integrin alpha D beta 2 binds VCAM-1: evidence for a binding interface between I domain and VCAM-1.

The trafficking of leukocytes through tissues is supported by an interaction between the beta 2 (CD18) integrins CD11a/CD18 (LFA-1) and CD11b/CD18 (Mac-1) and their ligand ICAM-1. The most recently identified and fourth member of the beta 2 integrins, alpha D beta 2, selectively binds ICAM-3 and does not appear to bind ICAM-1. We have reported recently that alpha D beta 2 can support eosinophil adhesion to VCAM-1. Here we demonstrate that expression of alpha D beta 2 in a lymphoid cell that does not express alpha 4 integrins confers efficient binding to VCAM-1. In addition, a soluble form of alpha D beta 2 binds VCAM-1 with greater efficiency relative to ICAM-3. The I domain of alpha D contains a binding site for VCAM-1 since recombinant alpha D I domain binds specifically to VCAM-1. In addition, alpha D mAb that block cellular binding to VCAM-1 bind the alpha D I domain. Using VCAM-1 mutants we have determined that the binding site on VCAM-1 for alpha D beta 2 overlaps with that of alpha 4++ integrins. Substitution of VCAM-1 aspartate at position 40, D40, within the conserved integrin binding site, diminishes binding to alpha D beta 2 and abrogates binding to the alpha D I domain. The corresponding integrin binding site residue in ICAM-3 is also essential to alpha D beta 2 binding. Finally, we demonstrate that alpha D beta 2 can support lymphoid cell adhesion to VCAM-1 under flow conditions at levels equivalent to those mediated by alpha 4 beta 1. These results indicate that VCAM-1 can bind to an I domain and that the binding of alpha D beta 2 to VCAM-1 may contribute to the trafficking of a subpopulation of leukocytes that express alpha D beta 2.

ICAM-3 and E-selectin endothelial cell expression differentiate two phases of angiogenesis in infantile hemangiomas.

Cellular adhesion molecules are newly identified mediators of angiogenesis. Infantile hemangiomas, characterized in the early stages by a proliferation of poorly differentiated vessels followed in the late stages by a vascular differentiation and regression of the tumor, represent an interesting model to study angiogenesis. We studied by immunohistochemistry the distribution of HLA-DR and three adhesion molecules ICAM-3, E-selectin and VCAM-1 on endothelial cells in different stages of vessel differentiation in infantile hemangiomas. We found high levels of ICAM-3 expression on proliferating vessels, while its expression was low or undetectable on well differentiated vessels. A different set of E-selectin antibodies showed a more heterogenous pattern of distribution and VCAM-1 antigens were found in both proliferating and differentiated vessels. HLA-DR expression on endothelial cells was inversely correlated to the vascular differentiation. Our results are consistent with the hypothesis that ICAM-3 plays a role in the early stages of vessel formation. Our results also suggest that variation of E-selectin and HLA-DR expression may be related either to vessel differentiation or may reflect the acquisition of an activated endothelial cell status.

Intercellular adhesion molecule-3 on endothelial cells. Expression in tumors but not in inflammatory responses.

Intercellular adhesion molecule-3 (ICAM-3) was identified as the third counter-receptor for lymphocyte function-associated antigen-1. ICAM-3 is absent on endothelial cells in normal tissues but found on endothelial cells in lymphomas. Here, we examined ICAM-3 expression on vascular endothelial cells in lymphomas, nonlymphoid malignancies, benign tumors, and inflammatory diseases. We compared the expression of ICAM-3 on endothelial cells with the severity of inflammatory infiltrates and with the presence of E-selectin and VCAM-1. We found that ICAM-3 expression on endothelial cells was high on both benign and malignant tumors whereas it was low in inflammatory diseases. In contrast to E-selectin, ICAM-3 expression on endothelial cells was not correlated to the severity of inflammatory infiltrates. In hemangiomas, we showed by Northern blot analysis and immunocytochemistry that ICAM-3 expression was induced and that it was localized in immature areas that sustain the early stages of angiogenesis. Therefore, expression of ICAM-3 on blood vessels does not seem to play a role in the recruitment of leukocytes during inflammation but rather is correlated with angiogenesis and tumor development.

Expression of ICAM-R (ICAM-3), a novel counter-receptor for LFA-1, in rheumatoid and nonrheumatoid synovium. Comparison with other adhesion molecules.

OBJECTIVE: To study the distribution of intercellular adhesion molecule receptor (ICAM-R, or ICAM-3), a novel ligand for the leukointegrin lymphocyte function-associated antigen 1 (LFA-1), in normal and rheumatoid synovial membranes and to compare this with the distribution of ICAM-1, ICAM-2, vascular cell adhesion molecule 1 (VCAM-1), and endothelial leukocyte adhesion molecule 1 (ELAM-1). METHODS: We performed immunohistochemical analyses of frozen sections of normal and rheumatoid synovial tissue using monoclonal antibodies to the molecules examined. RESULTS: ICAM-1 staining was detectable on the vascular endothelium and the synovial lining cells of both normal and rheumatoid synovial membranes. A variable proportion of lymphocytes infiltrating rheumatoid tissues expressed ICAM-1, ICAM-2 staining was demonstrable in the vascular endothelium of both normal and inflamed tissues, the latter demonstrating a significantly higher proportion of positive vessels. ELAM-1 staining was not detectable in normal synovial membranes but was seen on the endothelium of a limited number of rheumatoid synovial vessels, usually close to the synovial lining cell layer. VCAM-1 staining was intense in both normal and rheumatoid synovial lining cells, but vascular staining was weak in both. In contrast, ICAM-R staining was not detected in association with any synovial blood vessels, but was widely expressed by lymphocytes and macrophages. Cells of the lining layer did not stain for ICAM-R. CONCLUSION: Although ICAM-R is a ligand for LFA-1 and shares considerable sequence homology with ICAM-1 and ICAM-2, it does not appear to be expressed by the endothelium of normal or inflamed synovial vessels. Intense expression of ICAM-R by rheumatoid synovial lymphocytes and macrophages suggests that it may play a role in processes requiring cell-cell contact, such as antigen presentation and homotypic aggregation.

[Cerebral lymphoma in AIDS: clinical study and clinicopathological correlations]. / Les lymphomes cérébraux du SIDA: étude clinique et corrélations clinico-pathologiques.

We report the natural history of 17 brain lymphomas (11 primary, 6 disseminated) from a post-mortem series of 130 patients with AIDS. Primary lymphomas appeared lately in the course of AIDS. They were often associated with a severe T-cell immunodepression and with more frequent opportunistic disorders than disseminated lymphomas. Associated Kaposi's sarcomas were surprisingly frequent. All patients presented with neurological manifestations. Heterogeneous features were seen at CT examination. The CSF was abnormal in 12/13 cases, with an increase of protein contents and secretion of immunoglobulins; it contained activated lymphocytes in 5/6 cases of disseminated lymphomas, and malignant cells in only one case. Cellular density never exceeded 8/mm3 for primary lymphomas, and the lymphocytes were considered normal. The pre-mortem diagnosis of cerebral lymphomas was made in five patients, with a time lapse of 1 to 7 months between the first neurological symptoms and death, and of 5 to 30 days between the diagnosis and death. Cerebral biopsy was diagnostic in 4 cases of primary cerebral lymphomas. In only 1/6 patients with disseminated lymphomas, the diagnosis had been made when the patient was still alive, based on CSF and bone marrow lymphomatous infiltrations. The diagnosis of cerebral lymphoma (7 primary, 5 disseminated) was post-mortem in 12 cases. It was made only at microscopic examination in 2/12 cases of primary lymphomas. The histopathological study frequently showed a multicentric involvement, and always an immunoblastic cell type with plasmablastic differentiation and frequent medium size cells. Marked gliosis and significant necrosis were often observed. Neuropathological lesions associated with HIV-1 infection (toxoplasmosis, CMV and HIV-1 encephalitis) were seen in 8 cases with primary lymphomas.

HIV-1-associated cognitive/motor complex: absence of neuronal loss in the cerebral neocortex.

We performed a postmortem morphometric study in six AIDS patients and six controls to determine if a neocortical neuronal loss occurs in HIV-1-associated cognitive/motor complex. Patients were selected during a prospective study including psychometric evaluation and neuroimaging, and none had focal lesions. Two had HIV-1-associated myelopathy with mild cognitive impairment, and four had HIV-1-associated dementia complex. Planimetry did not show any cerebral atrophy. Cortical thickness, mean neuronal size, and mean neuronal densities in Brodmann's areas 4, 9, and 40 were not statistically different in patients and controls. There were no significant changes in neuronal densities of columnar and laminar samples, indicating that there was neither global nor selective neuronal loss. HIV-1-associated cognitive/motor complex is not necessarily related to neocortical neuronal loss, but could be due to subcortical lesions or metabolic dysfunction.

[A neuropathological study of 10 HIV-infected children]. / Etude neuropathologique de 10 enfants infectés par le VIH.

In 10 children infected by HIV at birth, who presented early and severe immunodeficiency encephalopathy, the lesions observed in the central nervous system were different from those found in adults. Using standard neuropathological techniques, the main abnormalities were white matter palor and atrophy, pyramidal tract demyelination, moderate perivascular inflammation, numerous calcifications of blood vessels in basal ganglia, white matter and occasional in the cortex, few opportunistic infections including cytomegalovirus ventriculitis, polymorphonuclear microabcessses and aspergillus abscesses; no toxoplasma was detected. An 18-month-old girl presented with an angiocentric lymphoproliferative disorder in central and peripheral nervous system and muscle, with predominance of B cells. In most cases, low levels of HIV replication were detected in brain tissue, as demonstrated by the presence of only few microglial nodules and giant cells, feeble detection of HIV p24 and p17 antigens by immunocytochemistry, in situ hybridization of HIV DNA and RNA and polymerase chain reaction, despite severe clinical encephalopathy. Zidovudine did not improve any patient. In children with severe AIDS encephalopathy, HIV might not be directly implicated in the central nervous system lesions: an intermediate factor such as cytokines or another toxic substance secreted by activated macrophages and/or lymphocytes, could induce severe lesions in the central nervous system and minor pathology of the peripheral nervous system and muscles.

Cloning and characterization of a new intercellular adhesion molecule ICAM-R.

The human intercellular adhesion molecules ICAM-1, ICAM-2 and their counter-receptors, the beta 2 or leukointegrins, mediate a variety of homotypic and heterotypic leukocyte and endothelial cell-cell adhesions central to immunocompetence. It has been found that cell-cell adhesion which is dependent on expression of the leukocyte function-associated antigen LFA-1 is not always blocked completely by antibodies raised against ICAM-1 and ICAM-2. Other leukointegrin ligands therefore probably exist, such as a glycoprotein of M(r) 124K that binds LFA-1 and has been designated ICAM-3 on the basis of this function. We have molecularly cloned a new member of the ICAM family, ICAM-R, which is related to ICAM-1 and ICAM-2. The complementary DNA encoding ICAM-R is 1,781 base pairs long and the protein has five extracellular immunoglobulin-family type domains. The mature cell-surface form of the ICAM-R protein has an M(r) which varies from 116 to 140K in a cell type-specific fashion. Overall identities in protein sequence with ICAM-1 and ICAM-2 are 48% and 31% respectively, with the degree of similarity varying between individual domains. The high level of expression of ICAM-R on resting leukocytes of all lineages and its lack of expression on either resting or cytokine-activated endothelial cells indicates a pattern of expression distinct from ICAM-1 and ICAM-2. In common with ICAM-1 and ICAM-2, ICAM-R is a ligand for the beta 2-integrin CD11a/LFA-1 (CD18).

Detection of human immunodeficiency virus type 2 in brain tissue.

Infection due to human immunodeficiency virus (HIV) type 2 is believed to cause a clinical picture similar to that of HIV-1, although extensive data are not available. In 2 patients with West African exposure and neurologic symptoms, HIV-2 was detected in the central nervous system using DNA and RNA polymerase chain reaction, in situ hybridization, and immunohistology. In the first patient, the neurologic disease was most likely due to productive infection with HIV-2. In the second, a combination of neuropathologic abnormalities (including the presence of HIV-2) explained the clinical features. Thus HIV-2, like HIV-1, can be readily detected in brain tissue in patients with neurologic abnormalities, although the exact role of HIV-2 in pathogenesis of AIDS-associated neurologic disease requires further study.

Low levels of human immunodeficiency virus replication in the brain tissue of children with severe acquired immunodeficiency syndrome encephalopathy.

The authors examined the autopsy brain samples of nine children infected with human immunodeficiency virus (HIV) at birth by histology, immunologic staining, and in situ hybridization. Surprisingly, although seven of these children presented with typical AIDS encephalopathy, the authors could detect a multifocal HIV infection in the brains of only three of these patients. The authors could not detect any significant HIV replication in the brain of four other children despite severe neurologic disease. However, HIV DNA was detected by polymerase chain reaction (PCR) in the central nervous system (CNS) of all patients. In addition, the authors found associated lesions in the brains of three of these four patients. This study shows that severe AIDS encephalopathy exists in children and therefore might exist in adults with few signs or without any signs of HIV replication or inflammation in the CNS. Understanding the pathogenesis of this neurologic disease and the kinetics of HIV replication in brain tissue of children with AIDS encephalopathy is essential to determine the best therapeutic strategy.

Clinical and virological aspects of HIV2 infection in rhesus monkeys.

To establish an animal model of AIDS, two different "wild" or "adapted" HIV2 Rod and Eho strains were cultivated on monkey cells from different species (baboons, cynomolgus, Rhesus monkeys). Five different available strains were then injected both by intravenous (i.v.) and intracerebral (i.c.) route into ten Rhesus monkeys. Seven animals seroconverted between days 13 and 230. Reverse transcriptase activity in the lymphocyte culture supernatants was detectable in six of the seven animals that seroconverted, and in one animal that remained seronegative. Lymphopenia and a decrease in the CD4+ cell counts were observed in eight animals. One animal, inoculated with HIV2-Rod "wild type," developed a severe cachexia, with dyspnea, and associated neurological symptoms 150 days after inoculation. This animal was sacrificed on day 220. Pathological examination showed typical lesions of actinomycetes infection in the lungs and in the meninges. Another monkey had significant weight loss associated with lymphadenopathies and pancytopenia. These results suggest that in vivo replication of HIV2 in Rhesus monkeys may induce clinical symptoms of immune deficiency. This method is reproducible and may provide a good model for AIDS.

Early viral replication in the brain of SIV-infected rhesus monkeys.

To investigate the mechanism of simian immunodeficiency virus (SIV) entry into the central nervous system (CNS) and the initial events leading to neuropathogenesis, SIV replication was studied by in situ hybridization in the CNS of 5 Rhesus macaques at 7 days, 1, 2, and 3 months after SIV intravenous inoculation. CNS infection was found to be a frequent and early event, as SIV was detected in the CNS of all the animals studied and as early as 7 days postinoculation. At the earliest stage, the infection localized mainly to perivascular cells. Using combined immunohistochemistry and in situ hybridization, infected cells were shown to express the CD68 marker, suggesting that infected mononuclear phagocytes crossing the blood-brain barrier represent the main source of virus in the CNS. Early viral replication coincided with neuropathologic changes, consisting in gliosis, perivascular infiltrates and rare glial nodules. Immunophenotyping of brain tissue showed that increased macrophage infiltration, microglial reactivity and MHC class II induction occurred within the first week of infection, indicating a possible immunopathologic mechanism in early CNS pathogenesis.

HIV proteins absent from placentas of 75 HIV-1-positive women studied by immunohistochemistry.

Recent epidemiological and virological data suggest that the incidence of maternofetal transmission of HIV-1 infection is between 20 and 30%. The available evidence points to a possible role of peri- and postnatal contamination, but the isolation of HIV from fetuses shows that transplacental transmission also occurs. We attempted to detect, by means of an immunohistochemical method, HIV proteins in frozen placentas from 75 HIV-1-positive women (30 at term, 45 induced abortions). In addition, in situ hybridization using HIV-specific probes was performed in three cases. Neither HIV proteins nor nucleic acid sequences were detected, but CD4+ mononuclear cells were present in the chorion and villi, regardless of the clinical and biological status of the mother (particularly in the nine cases in which the infants were infected). There are several possible mechanisms involving the placenta in the maternofetal transmission of HIV, including active transport of the HIV-immunoglobulin G complex via Fc receptors on trophoblastic cells, passive transplacental passage of HIV during a viraemic episode, the passage of infected maternal cells, and infection of the placenta itself. The methods we used could not rule out the presence of HIV DNA provirus within the genome of placental cells. In any event, immunohistochemical detection of HIV proteins in the placenta is not a technique suitable for the prenatal diagnosis of HIV infection or for identifying newborns likely to develop HIV infection.

AIDS encephalopathy and tropism of HIV for brain monocytes/macrophages and microglial cells.

HIV induces severe dementia in about 20% of adult AIDS patients. In children HIV-infected at birth, the incidence of specific neurological complications is still higher since severe encephalopathy occurs in almost all children who develop an early and severe immunosuppression. In all cases, the brain monocytes/macrophages and the microglial cells are the only cells which replicate HIV in the central nervous system (CNS) of these patients, and the appearance of neurological symptoms seems induced by an interaction between HIV-infected macrophages with neurons and glial cells. AIDS encephalopathy is related to two properties of HIV: to the viral tropism for monocytes/macrophages/microglial cells, which allow the brain infection, and to HIV tropism for CD4+ lymphocytes responsible for the appearance of immunosuppression, which trigger viral dissemination in the CNS. However, childhood encephalopathy is not always associated with HIV replication in the CNS at the time of death, and mild dementia in HIV-infected adults were described without signs of HIV replication in autopsy CNS samples. Those findings suggest that persistent, productive viral infection is not required for the development of HIV encephalopathy. Therefore, if the relationship between HIV CNS infection and AIDS encephalopathy in adults and children is clearly demonstrated, the pathogenesis of the neurological disease and the kinetics of HIV replication in the CNS are unclear. In addition, the very high incidence of AIDS encephalopathy in children could be related to HIV infection of microglia which is differentiating in fetal or newborn brain.

Skeletal muscle involvement in human immunodeficiency virus infection.

In addition to muscle changes due to peripheral nervous system involvement, primary myopathic changes associated with the human immunodeficiency virus (HIV) have also been described. We studied seven cases: two had developed an acquired immunodeficiency syndrome (AIDS), four had seroconverted to HIV but were otherwise asymptomatic, one was HIV seronegative when the biopsy was performed and one was biopsied twice. Besides the HIV no other infectious agent was detected. Muscle biopsies showed: (a) muscle fiber necrosis and regeneration; (b) inflammatory changes with moderate perivascular infiltration; and (c) unusual myofibrillary disorganization. Immunocytochemical techniques using anti-HIV monoclonal antibodies showed the presence of the virus in one biopsy. HIV-RNA was detected by in situ hybridization in the same biopsy. With both techniques the HIV was detected in isolated mononuclear cells in the muscle endomysium and not within the muscle fibers. Muscle involvement associated with HIV infection may be related, at least in some cases, to the presence of the virus in interstitial cells.

[AIDS dementia]. / La démence du SIDA.

Human immunodeficiency virus type I (HIV), the etiology of AIDS is also the cause of a primary infection of the central nervous system. The AIDS Dementia Complex is a common and important cause of morbidity in patients in advanced stages of infection. Subacute encephalitis demonstrating, the neuropathogenicity of HIV 1 constitute an human model for slow virus encephalitis.

Severe encephalitis resulting from coinfections with HIV and JC virus.

We observed 3 cases of progressive multifocal leukoencephalopathy (PML) among frozen CNS samples obtained at autopsy from 102 adult AIDS patients. In 2 patients, PML was associated with severe HIV encephalitis. In those 2 cases, the areas of extensive JC-induced demyelination were massively infiltrated by HIV infected macrophages/microglial cells with evidence for localized increase of HIV encephalitis in PML lesions. Using immunohistochemistry and in situ hybridization, we demonstrated that each virus infects, in a latent or productive fashion, different CNS cell populations. Therefore, the extension of HIV encephalitis could not be related to an intracellular transactivation of 1 virus by the other. However, the results are consistent with dissemination of viral infection by the recruitment of HIV-infected macrophages to damaged areas of the brain. This phenomenon might be generalized to other pathogens that are frequently associated with HIV CNS infection. Early detection and treatment of opportunistic CNS lesions could be important to prevent extension of HIV encephalitis.