SUMMARYIn this paper we addressed the expression of the HIV co-receptors CXCR-4 and CCR-5 in human thymocytes by phenotypic, molecular and functional approaches. Cytofluorimetric analysis disclosed that CXCR-4 was constitutively expressed by freshly isolated thymocytes (~10 000 molecules/cell in about 30% of thymocytes); the receptor was endowed with functional activity, as it mediated polarization, migration and intracellular Ca 2+ increase in response to its ligand, SDF-1. On the contrary, CCR-5 expression in freshly isolated thymocytes was significantly lower (<4000 molecules/cell in less than 5% of the cells), and no functional response to CCR-5 agonists could be documented. Northern blot analysis of freshly isolated thymocytes showed high CXCR-4 mRNA levels, whereas the message for CCR-5 was barely detectable. On the other hand, a modest increase in the expression of CCR-5 was associated with in vitro thymocyte stimulation, and CCR-5 density at the cell surface attained CXCR-4 figures in most cases. None the less, no functional response to CCR-5 agonists could be documented in in vitro stimulated thymocytes. In vitro infection of thymocytes by CAT-expressing recombinant HIV bearing the envelope glycoproteins from different isolates showed that T-tropic strains, which use CXCR-4 as a co-receptor, were more efficient in infecting thymocytes than M-tropic strains, which preferentially use CCR-5. Altogether, these data indicate that expression of the major co-receptors involved in infection by M-tropic HIV strains is very poor in human thymocytes, and would suggest that thymocyte infection by M-tropic HIV strains may be a rare event in vivo.
To better characterize the cellular source of lymphotactin (XCL1), we compared XCL1 expression in different lymphocyte subsets by real-time PCR. XCL1 was constitutively expressed in both PBMC and CD4+ cells, but its expression was almost 2 log higher in CD8+ cells. In vitro activation was associated with a substantial increase in XCL1 expression in both PBMC and CD8+ cells, but not in CD4+ lymphocytes. The preferential expression of XCL1 in CD8+ cells was confirmed by measuring XCL1 production in culture supernatants, and a good correlation was found between figures obtained by real-time PCR and XCL1 contents. XCL1 expression was mostly confined to a CD3+CD8+ subset not expressing CD5, where XCL1 expression equaled that shown by γδ+ T cells. Compared with the CD5+ counterpart, CD3+CD8+CD5− cells, which did not express CD5 following in vitro activation, showed preferential expression of the αα form of CD8 and a lower expression of molecules associated with a noncommitted/naive phenotype, such as CD62L. CD3+CD8+CD5− cells also expressed higher levels of the XCL1 receptor; in addition, although not differing from CD3+CD8+CD5+ cells in terms of the expression of most α- and β-chemokines, they showed higher expression of CCL3/macrophage inflammatory protein-1α. These data show that TCR αβ-expressing lymphocytes that lack CD5 expression are a major XCL1 source, and that the contribution to its synthesis by different TCR αβ-expressing T cell subsets, namely CD4+ lymphocytes, is negligible. In addition, they point to the CD3+CD8+CD5− population as a particular T cell subset within the CD8+ compartment, whose functional properties deserve further attention.
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