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.

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.

[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.

[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).

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.

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.

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 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.

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.

MRI pattern of progressive multifocal leukoencephalopathy (PML) in AIDS. Pathological correlations.

Progressive multifocal leukoencephalopathy (PML) is a demyelinating disease which occurs in immunodepressed subjects and is particularly frequent in AIDS. Some authors having drawn attention to the protean aspect of the disease and claimed that AIDS may lose its basic characteristics and affect the grey matter as well as the white matter, we reviewed a series of 8 patients who had been biopsied and/or autopsied and had been examined at least once by MRI. In this series, contrary to what is regularly observed in toxoplasmic abscesses we did not find any lesion of the grey matter or any mass effect. On the other hand, we confirmed that PLM is not multifocal in all cases and that it course may be interrupted by prolonged remissions. The MRI criteria for PML therefore are reliable, provided multiple T2-weighted slices in coronal plane are performed, clearly showing the anatomy of the white fibres affected. However, it must be borne in mind that HIV-infected patients often have other associated brain pathologies, especially when the immune deficiency increases.

The rev gene product of the human immunodeficiency virus affects envelope-specific RNA localization.

By in situ hybridization analysis and immunoprecipitations following transfection of COS cells, we show that the Rev protein of the human immunodeficiency virus is necessary for envelope protein expression, which is correlated with the appearance in the cytoplasm of envelope-specific RNA. In the absence of cotransfection with a plasmid expressing Rev, envelope-specific RNA is retained in the nucleus. Several cis-acting sites in the envelope are involved, one of which is between nucleotides 7330 and 7735 and is required for the response to Rev. Other sequences (nucleotides 5797-7330 and 7735-7989) are involved in the apparent retention of the envelope-specific RNA in the nucleus in the absence of Rev and its response element. Because Rev affects the localization of envelope RNA both in the presence and in the absence of the normal splice sites on the RNA, the mechanism of Rev action is independent of splicing.

HIV-associated endometritis.

Using immunohistochemical staining, in situ hybridization and a combination of both, we demonstrate here the replication of HIV in the endometrial stroma. Infected cells do not belong to the T-lymphocyte lineage but rather to a monocyte-macrophage cell type. This report suggests a possible relationship between HIV infection and endometritis. Moreover, HIV replication in endometrial tissues could play a role in heterosexual and materno-fetal transmission.

Biological aspects and diagnosis procedures of materno-fetal transmission of HIV.

PIP: Since an estimated 1-2 million heterosexual women in Africa are infected with human immunodeficiency virus (HIV), maternal-fetal transmission of this virus is a serious issue. At this point, the only accurate data concern the high level of virus transmission from infected mothers to the newborn (at least 40%) and the severity of infection in infants. About 50% of infected infants develop acquired immunodeficiency syndrome, generally at 6 months of age, and die by the age of 3-4 years. There have been no studies large enough to determine the time of HIV transmission--antenatal, prepartum, or postpartum? Also unclear at this point are the consequences of pregnancy for the immunological status for women infected with HIV. Amniocentesis, cord blood punction, and trophoblastic biopsy are potential means of diagnosing HIV infection prenatally; however, there is a risk of infecting the fetus by the techniques themselves, a danger of false negative results and a likelihood that HIV infection can occur later in pregnancy after the tests have been completed.^ieng

AIDS subacute encephalitis. Identification of HIV-infected cells.

Human immunodeficiency virus (HIV) RNA and proteins were detected in the brains of several AIDS patients with subacute encephalitis, by in situ hybridization and immunohistology. The majority of infected cells were mononucleated and bore processes. Using single and double immunohistologic procedures, the authors identified these cells as macrophages. The majority of them had the phenotype of microglial cells (Leu-M3-, CD4-), others were labeled with markers of circulating macrophages (Leu-M3+, CD4+/-). The presence of HIV RNA and proteins in CD4- cells could be explained by depressed CD4 antigen expression, as a result of infection or macrophage tissue differentiation.

[Neuropathological study of 31 cases of acquired immunodeficiency syndrome]. / Etude neuropathologique de 31 cas de syndrome d'immuno-dépression acquise.

Post-mortem study of every patient who died from AIDS in Pitié-Salpêtrire Hospital from June 1984 to November 1985 was performed without regard to the presence of neurological signs and symptoms. Autopsy were performed in 31/48 cases. Patients had been hospitalized in the Departments of Parasitology-Infectious Disease (24 cases) Internal Medicine (4 cases) and Neurology (3 cases). In every case, formalin-fixed material from the brain and the spinal cord were embedded in paraffin (20 samples), stained with hematoxylin-eosin, PAS, Alcian blue, Giemsa, Grocott and Ziehl techniques and Bodian's silver impregnation along with Luxol fast blue, and, in celloïdin (8 samples), stained with hematoxylin-eosin and Loyez' impregnation. There were 30 men (27 caucasian, 1 egyptian, 1 haïtian, 1 senegalese) and one woman (congolese). Twenty eight (28) patients were homosexuals. AIDS was transfusion-associated in two cases. Neurologic complications revealed the disease in 2 cases. Eighteen (18) patients had neurological signs or symptoms before death. Age range at death was 22-58 (mean 38). Brain weight in AIDS (from 1150 gms to 1750 gms-mean 1428 gms) was not statistically different from the mean weight of 100 male patients in the same age range autopsied in the same laboratory during the identical period (mean 1427 gms, standard deviation: 23). Microscopic abnormalities were present in every brain examined. These included non-Hodgkin lymphoma (3 cases), opportunistic infections (21 cases: 13 toxoplasmosis, 4 cytomegalovirus encephalitis, 3 cryptococcal meningitis, 1 infection by mycobacterium avium-intracellulare), and subacute encephalitis (17 cases, 9 isolated, 8 associated with other disorders). The characteristic changes consisted of lympho-monocytic focal infiltrates (so-called microglial nodules) and mild lympho-monocytic perivascular cuffs in 10 cases. Typical giant cells were seen only in one case. Mild demyelinating changes were also seen in only one case. No spinal cord spongiosis, nor Progressive Multifocal Leukoencephalopathy was found. HIV localization was performed on frozen sections utilizing in situ hybridization techniques (2 cases) and immunohistologic techniques (5 cases). HIV, RNA and proteins, was detected in 2 cases with subacute encephalitis. Infected cells were labeled with macrophage markers, and rarely with T4 lymphocyte markers. Infected astrocytes (identified by anti-GFAP serum) or neurons (identified by anti-NSE serum) were never observed. No giant cells were seen in these two cases.

Positive Coombs test in acute leukaemia.

We present the results of the immunohaematological tests performed in an unselected group of patients with acute leukaemia at the time of diagnosis, and when possible the follow-up in remission and/or in relapse. Thirteen out of the 112 patients tested had a positive Coombs test at the time of diagnosis and, in six patients, the Coombs test became positive during remission. All the 19 positive Coombs test were of the complement type with, in 11 cases, an anti-I antibody eluted from the patient's RBC. There was no relationship between the FAB morphologic subtype, the presence of other immune abnormalities, the course of the leukaemia and the immunohaematological abnormalities. Several hypotheses on the possible relationships between the acute leukaemias and the immunohaematological abnormalities are discussed.