Astrocyte cultures from human embryonic brain: Characterization and modulation of surface molecules by inflammatory cytokines

Abstract

Astrocyte-enriched cultures were established upon passaging of primary cultures from the myelencephalon and mesencephalon of 7-9-week-old human embryos. Immunocytochemical analysis showed that third-fourth passage cultures were composed of a highly enriched population of proliferating, epithelioid cells, up to 90% of which expressed glial fibrillary acidic protein (GFAP); no macrophages and very few fibroblasts (less than 2%) were present. GFAP expression and proliferation declined upon further culturing in serum-containing medium but could be transiently reinduced by growing the cells in a serum-free chemically defined medium. Large numbers of GFAP+ astrocytes were obtained from each embryo and could be stored frozen and recultured. Using flow cytometric analysis, human astrocyte cultures were examined for basal and cytokine [interferon-gamma (IFN-gamma), interleukin-1 beta (IL-1 beta), and tumor necrosis factor-alpha (TNF-alpha)]-induced expression of molecules that may be involved in astrocyte-T-lymphocyte interactions. Cultured human astrocytes spontaneously expressed major histocompatibility complex (MHC) class I antigens and variable levels of MHC class II; MHC class I levels were increased upon IFN-gamma and TNF-alpha treatment, whereas MHC class II antigens were induced on most of the astrocytes by IFN-gamma. Among the molecules involved in antigen-independent interactions between T lymphocytes and target cells, lymphocyte function-associated molecule-3 (LFA-3) was spontaneously expressed by most cultured human astrocytes, whereas intercellular adhesion molecule-1 (ICAM-1) was present at variable levels in non-stimulated astrocytes and was greatly induced by IFN-gamma, TNF-alpha, and IL-1 beta. In this study we also show that the above cytokines upregulate astroglial expression of adhesion molecules of the integrin family (VLA-1, VLA-2, and VLA-6) that may be involved in astrocyte-extracellular matrix interaction and play a role in the astrocyte reactive changes occurring at sites of brain injury and inflammation. The human astrocyte cultures developed here represent a useful in vitro model to further investigate mechanisms involved in bidirectional communication between central glia and cells of the immune system.

Human immunodeficiency virus coat protein gp120 inhibits the beta-adrenergic regulation of astroglial and microglial functions.

“…Finally, the elevation of cAMP induced by forskolin was unaffected by gpl20 ( Fig. 1) In the attempt to confirm the data described above with human cells, astrocyte-enriched cultures prepared from embryonic human brain (12) and four astroglioma cell lines were tested for expression of functional 8-adrenergic receptors by measuring the isoproterenol-induced cAMP formation. In six astrocyte preparations and in three astrogliomas, isoproterenol was ineffective (probably due to the absence of mature 13-adrenergic receptors, since forskolin did raise cAMP levels), whereas in one glioma cell line consisting of >95% GFAP-positive cells (kindly donated by E. Vigneti), 1 ,uM isoproterenol caused a 3-fold increase in cAMP that was 50%yo antagonized by 1 These results are qualitatively consistent with those obtained with the rat cultures.…”

“…25), and microglia (26) of AIDS patients with neurological involvement. TNF-a has been shown to be toxic for oligodendrocytes and myelin (27), to induce major histocompatibility complex antigen expression in astrocytes (12,28), to stimulate the astroglial expression and production of interleukin 6 (29,30) and ofcolony-stimulating factors (30,31), to stimulate astrocyte proliferation (32), to synergize with other inflammatory cytokines such as interferon y (12), to facilitate the passage of T cells across the blood-brain barrier (33), and to stimulate HIV replication in infected cells (see ref. 4).…”

Oncostatin M and the Interleukin-6 and Soluble Interleukin-6 Receptor Complex Regulate α1-Antichymotrypsin Expression in Human Cortical Astrocytes

“…We did not address specifically the question as to which of the components involved in LIF signaling was missing in astrocytes. However, considering that astrocytes produce both IL-6 and LIF (35,36) we speculate that LIF-R is a likely candidate. Contrary to our present observations with human astrocytes, IL-6 has been reported to upregulate nerve growth factor expression in mouse astrocytes (37).…”

The Brain Parenchyma Is Permissive for Full Antitumor CTL Effector Function, Even in the Absence of CD4 T Cells

“…(data not shown), one of the signs of reactive gliosis that accompanies many forms of CNS stress (47). Astrocytes in such an activated state can participate in immune reactions by up-regulation of MHC and adhesion molecules (44,48,49) and liberation of inflammatory chemokines and cytokines (45,46,50,51). Analysis of events even nearer to the day of implantation is difficult to interpret in a transplanted tumor model because of the inevitable physical trauma associated with implantation, but these issues of the initiating events for an antibrain tumor response are evidently important to address in future studies.…”