In silico subtraction approach reveals a close lineage relationship between follicular dendritic cells and BP3^hi stromal cells isolated from SCID mice

Abstract

Organization of the stromal compartments in secondary lymphoid tissue is a prerequisite for an efficient immune reaction. In particular, follicular dendritic cells (FDC) are pivotal for the activation and differentiation of B cells. To investigate the development of FDC, FDC together with tightly associated B cells (FDC networks) were micro-dissected from frozen tissue sections and follicular B cells were sorted by FACS. Using an in silico subtraction approach, gene expression of FDC was determined and compared with that of follicular stromal cells microdissected from the spleen of SCID mice. Nearly 90% of the FDC genes were expressed in follicular stromal cells of the SCID mouse, providing further evidence that FDC develop from the residual network of reticular cells. Thus, it suggests that rather minor modifications in the gene expression profile are sufficient for differentiation into mature FDC. The analysis of different immune-deficient mouse strains shows that a complex pattern of gene regulation controls the development of residual stromal cells into mature FDC. The in silico subtraction approach provides a molecular framework within which to determine the diverse roles of FDC in support of B cells and to investigate the differentiation of FDC from their mesenchymal precursor cells. [1][2][3][4]. The network of FDC is a micro-environment required for the survival of follicular B cells and is also a prerequisite for an efficient GC reaction. At the early stage of GC development FDC support B-cell proliferation, whereas at the later stages FDC have an important function in the selection and differentiation of high affinity B cells to memory and plasma cells [1,5].Although FDC are crucial for B-cell development, our knowledge of FDC transcriptional activity remains marginal. FDC are fragile cells and are tightly associated with B cells -properties that have thus far hampered the isolation of pure FDC populations [6][7]. To overcome these problems, FDC lines have been established, however, as these cells are maintained over several weeks in culture, their phenotype no longer reflects the in vivo situation [8][9][10][11][12][13]. A number of different approaches for the enrichment and gene expression analysis of FDC have been shown to be more representative of the in vivo situation [6,8,11].From a number of experiments, it is apparent that FDC are a highly specialized subset of reticular cells [14][15][16][17][18]. Using BP3, an ectoenzyme of the NAD glycohydrolase family, as a marker antigen for the splenic reticular network of stromal cells, one can distinguish the reticulum cells in the B-cell area from those in the T-cell zone. High expression of BP3 defines the follicle, the area to which B cells To identify molecular markers defining a developmental relationship between mature FDC and the BP3 hi reticular cells of SCID mice, gene expression profiles were determined. Using an in silico subtraction approach, we were able to identify a novel set of genes that showed specific expression in FDC. When gene ex...

Characterizing follicular dendritic cells: A progress report

“…Theoretically the microdissection approach should be superior to the LTbR decoys as it minimizes the risk of introducing uncontrollable, spurious effects that might be brought about by meddling with LTbR signaling. In practice, however, most of the markers described by Huber [30] were confirmed by Wilke et al [29]. In addition, Wilke and colleagues describe several novel transcripts particular to FDC -Cilp, Lrat, Ltbp3, Postn, and Parm1 -whose expression specificities were confirmed by histology.…”

“…In a report published in the current issue of EJI, Wilke et al [29] apply an interesting approach to identify potential FDC markers, which they term ''in silico subtraction''. This methodology has strengths and weaknesses, and is mostly dictated by the technical inability to isolate FDC in a pure state -whereas in any other context it might have been preferable to utilize ''in realitate subtraction''.…”

“…RNA expression profiles of the microdissected tissue and the isolated B cells are obtained, and the B-cell transcriptome subtracted numerically. Wilke et al [29] show a total of 575 FDC-enriched transcripts. Reassuringly, these include well-validated FDC markers such as Cxcl13, Bp3, Madcam-1, Vcam-1, and Prnp.…”

Follicular Dendritic Cells Emerge from Ubiquitous Perivascular Precursors

“…Although LTβR signaling is necessary for SLO development, early Mfge8 expression was LTβR-independent, suggesting the existence of additional LTi-dependent clues at this step of FDC ontogenesis. FDC precursors were postulated in the marginal sinus of TNFR1 −/− and SCID mice (Pasparakis et al, 2000; Wilke et al, 2010) and final FDC maturation requires not only LTi cells but also B cells (Fu et al, 1998; Tumanov et al, 2004). Thus, even though LTi cells share with B cells the expression of most TNF ligands, such as LTαβ and TNF (Kim et al, 2006), the factors provided by LTi cells are insufficient for terminal FDC differentiation.…”

Follicular Dendritic Cell-Specific Prion Protein (PrPc) Expression Alone Is Sufficient to Sustain Prion Infection in the Spleen

“…After peripheral exposure prions accumulate first upon follicular dendritic cells (FDC) as they make their journey from the site of infection to the CNS (a process termed, neuroinvasion ) [1]–[7]. FDC are a unique subset of stromal cells resident within the primary B cell follicles and germinal centres of lymphoid tissues [8]. Prion accumulation upon FDC is critical for efficient disease pathogenesis as in their absence neuroinvasion are impaired [1]–[4].…”