Syncytium Formation in Cultured Human Lymphoid Tissue: Fusion of Implanted HIV Glycoprotein 120/41-Expressing Cells with Native CD4⁺Cells

Abstract

While glycoprotein gp120/41 clearly causes HIV-infected cells to form syncytia in monolayers and in suspension, there is unfortunately scant knowledge on syncytium formation in tissues. We implanted gp120/41-expressing cells labeled with fluorescent particles inside blocks of human lymphoid tissue kept in long-term histoculture. Observed by confocal microscopy, together with immunohistochemical and morphological analysis of implanted cells, more than one-third of these gp120/41-expressing cells fused with native CD4+ cells of the host tissue, yielding small (three to five nuclei) syncytia. Such widespread fusion of gp120/41-expressing cells in tissue in vitro, together with the finding of increased virulence of syncytium-inducing isolates of HIV, support the hypothesis that syncytium formation within lymph tissue of HIV-infected individuals contributes to AIDS pathogenesis. This system and the methods developed may provide a way to study HIV-infected cells inside the very tissue whose destruction may prevent immune system repopulation.

Replication of Aleutian mink disease parvovirus in mink lymph node histocultures

“…Taken together, these preliminary studies indicated that gel-supported histocultures derived from mink MLN remained viable and exhibited similar characteristics to those described for other species (Margolis et al, 1995 a ;Glushakova et al, 1997 ;Nilsson, 1971). Histocultures incubated for 120 h were stained with viability stain (ethidium homodimer-1 staining dead cells red, and calcein-AM staining living cells green) and analysed by confocal microscopy.…”

“…Histoculture system. The mink histoculture system was modelled on previously described examples (Margolis et al, 1995 a ;Glushakova et al, 1997 ;Li et al, 1991 ;Freeman & Hoffman, 1986 ;Nilsson, 1971 ;Sorour et al, 1975). ADV-negative mink were euthanized by cardiac puncture and the MLN was aseptically removed.…”

“…To determine viability, histocultures were analysed by confocal microscopy basically as described (Margolis et al, 1995 b). Histoculture blocks were harvested on successive days and incubated for 20 min at room temperature in PBS containing 4 µM ethidium homodimer-1 (which stains dead cells red) and 2 µM calcein-AM (which stains live cells green) (Molecular Probes), rinsed in PBS and analysed within 15 min.…”

“…In order to identify target cells for virus infection and to extend the analysis of the complex pathogenesis of AD, an ex vivo system would be an advantage. Recent studies describe the use of gel-supported histocultures for examining HIV pathogenesis (Margolis et al ., 1995 a ; Glushakova et al ., 1997 ). The advantage of the histoculture system compared to conventional culturing systems is a preservation of the microscopic three-dimensional cytoarchitecture in the cultured organ, even in very dynamic tissues such as secondary lymphoid organs (Nilsson, 1971 ).…”

Abortive HIV Infection Mediates CD4 T Cell Depletion and Inflammation in Human Lymphoid Tissue

“…HIV-1 virions (Holm et al, 2004; Jekle et al, 2003; Vlahakis et al, 2001), cell-mediated fusion (LaBonte et al, 2000; Margolis et al, 1995), and hemifusion (Garg et al, 2007) have been proposed to be involved in indirect killing. Therefore, the requirement for cell-cell interaction in indirect killing may be mediated either by effective delivery of HIV-1 virions or by cell-associated Env.…”