In vitro cell fusion between CD4(+) and HIV-1 Env(+) T cells generates a diversity of syncytia varying in total number, size and cellular content.

Syncytia formation in HIV infections is driven by the virus fusion-active molecules (Env) interacting with membrane components of hosts cells. HIV-syncytia are usually interpreted as pathogenic entities and although they may potentially vary in size, numbers and types of constituent cells, little is known about the extent and significance of their diversity. Here, we describe numerically the cell population dynamics and the diversity of syncytia produced in the in vitro cell-fusion between two Jurkat T cell lines, one CD4(+) and the other Env(+). Cell-fusion partners were differentially stained with the lipophilic DiI and DiO, or with the cytoplasmic CMFDA and CMTMR tracers and syncytia showing double fluorescence were counted in a flow cytometer. The total number of syncytia formed, their size, cellular complexity and ratio of CD4(+)/Env(+) cells recruited, varied significantly in relation with time of reaction and initial proportions of fusion partners. The considerable structural diversity of syncytia formed, in so limited an in vitro cell fusion reaction, suggests that a greater heterogeneity may be formed in the natural course of disease. Identification of the main determinants of syncytia diversity allows for a detailed study of the relation between the syncytia structure and function.

Comparative toxicity of oleic acid and linoleic acid on Jurkat cells.

BACKGROUND: Lipid emulsions for parenteral nutrition commercially available are mainly composed of long-chain triacylglycerol containing a high proportion of alpha-6 polyunsaturated fatty acids or alpha-9 monounsaturated fatty acids. The immunological impact of such therapy is particularly important because parenteral and enteral diets are often administered to critical ill patients. The comparative toxicity of oleic acid and linoleic acid on Jurkat cells, a human T lymphocyte cell line, and the type of cell death induced by these fatty acids were determined. METHODS: Cell death was investigated by cytometry: decrease in cell volume, increase of granularity, DNA fragmentation, phosphatidylserine externalization, mitochondrial depolarization, lipid accumulation; by fluorescence microscopy: chromatin condensation and acridine orange/ethidium bromide assay; and by RT-PCR: mRNA expression of apoptotic genes. RESULTS: Evidence is presented herein that oleic acid is much less toxic to Jurkat cells than linoleic acid. Both fatty acids promote apoptosis and necrosis of these cells. The mechanism of cell death induced by these fatty acids seem to involve with mitochondrial depolarization, lipid accumulation and the levels of C-MYC and P53 mRNA expression. CONCLUSION: Therefore, oleic acid may offer an immunological less harmful alternative to linoleic acid for parenteral and enteral diets preparation.

The MAP kinases are differently utilized by CD28 and CD2 adhesion pathways in superantigen-activated Jurkat T cells.

To mimic the two-signal requirements for T cell activation mediated by ligands, we exposed the superantigens SEA or SEE (signal 1) to T cells incubated with HLA-DR/LFA-3 or HLA-DR/B7-1-CHO transfected cells (signal 2). LFA-3 costimulation was able to induce T cell proliferation as well as IFN-gamma and IL-4 production at similar levels as in cells induced by B7-1. Analysis of the CD28RE of the IL-2 promoter showed specific transcription factor recruitment at the CD28RE element upon induction by B7-1/SEE. Further functional studies with an IL-2 enhancer-promoter carrying either wild type or mutated versions of the CD28RE site revealed that this element is necessary for full activation upon B7-1 costimulation. While both CD28/B7-1 and CD2/LFA-3 costimulation resulted in the up-regulation of IL-4 and IFN-gamma promoters, IL-2 promoter activity and production of IL-2 were only seen after B7-1 costimulation. However, contrary to what has been previously proposed, we show that costimulation with either B7-1 or LFA-3 further enhanced the ERK-2 activity and strongly activated the p38 MAPK pathway, but only B7-1 costimulation induced high levels of JNK-1 activity. These data suggest that the differential effect of CD28 vs. CD2 can be related to the difference in the ability of the two pathways to induce JNK-1 activity.