Purification of circulating plasmacytoid dendritic cells using counterflow centrifugal elutriation and immunomagnetic beads

Background aimsPlasmacytoid dendritic cells (pDC) are a dendritic cell (DC) subset specialized in the production of high amounts of interferon (IFN) type I (IFN-α, -β) in response to viruses. They can be purified from peripheral blood mononuclear cells (PBMC), usually using magnetic bead sorting.MethodsIn this study, we set up a counterflow centrifugal elutriation (CCE) procedure to enrich pDC from PBMC. We first analyzed each CCE fraction for the presence of pDC using CD123 and BDCA-2 as markers. We then purified pDC using CCE and magnetic beads and verified that their functions were not affected by this procedure.ResultspDC were sorted by CCE into intermediate fractions between those containing lymphocytes and monocytes. The pDC frequency in these intermediate fractions was 3-fold that in PBMC. Using negative-magnetic bead sorting, starting with the same number of cells and beads, we obtained more than twice as many pDC from intermediate fractions as from PBMC. The phenotypes and IFN-α production capacities of sorted pDC from PBMC and from intermediate fractions were similar, both immediately after sorting and after stimulation with CpG-A oligodeoxynucleotides. In addition, we showed that intermediate fractions could be cryopreserved and that magnetic bead sorting could be performed with the same efficiency after thawing.ConclusionsAltogether, our results show that CCE can be used to enrich lymphocytes, monocytes and pDC from the same donor, without magnetic beads on their surface. Our method should be useful for the purification of these cells for experimental research and may also be adaptable for clinical use in immunotherapy.     

A model of erythropoiesis in adults with sufficient iron availability

In this paper we present a model for erythropoiesis under the basic assumption that sufficient iron availability is guaranteed. An extension of the model including a sub-model for the iron dynamics in the body is topic of present research efforts. The model gives excellent results for a number of important situations: recovery of the red blood cell mass after blood donation, adaptation of the number of red blood cells to changes in the altitude of residence and, most important, the reaction of the body to different administration regimens of erythropoiesis stimulating agents, as for instance in the case of pre-surgical administration of Epoetin-α. The simulation results concerning the last item show that choosing an appropriate administration regimen can reduce the total amount of the administered drug considerably. The core of the model consists of structured population equations for the different cell populations which are considered. A key feature of the model is the incorporation of neocytolysis.     

Algal Remodeling in a Ubiquitous Planktonic Photosymbiosis

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SOX10 mutations in chronic intestinal pseudo-obstruction suggest a complex physiopathological mechanism

The type IV Waardenburg syndrome (WS4), also referred to as Shah-Waardenburg syndrome or Waardenburg-Hirschsprung disease, is characterised by the association of Waardenburg features (WS, depigmentation and deafness) and the absence of enteric ganglia in the distal part of the intestine (Hirschsprung disease). Mutations in the EDN3, EDNRB, and SOX10 genes have been reported in this syndrome. Recently, a new SOX10 mutation was observed in a girl with a neural crest disorder without evidence of depigmentation, but with severe constipation due to a chronic intestinal pseudo-obstruction and persistence of enteric ganglia. To refine the nosology of WS, we studied patients with typical WS4 (including Hirschsprung disease) or with WS and intestinal pseudo-obstruction. We found three SOX10 mutations, one EDNRB and one EDN3 mutations in patients presenting with the classical form of WS4, and two SOX10 mutations in patients displaying chronic intestinal pseudo-obstruction and WS features. These results show that chronic intestinal pseudo-obstruction may be a manifestation associated with WS, and indicate that aganglionosis is not the only mechanism underlying the intestinal dysfunction of patients with SOX10 mutations.     

Hierarchy of effects of the MHC and T cell receptor beta-chain genes in susceptibility to Theiler's murine encephalomyelitis virus-induced demyelinating disease.

Intracerebral inoculation of susceptible mice with Theiler's murine encephalomyelitis virus induces a demyelinating disease that is similar to human multiple sclerosis. This murine model for human multiple sclerosis is apparently immune-mediated and the genes involved in the immune response influence the outcome of disease susceptibility as observed with human multiple sclerosis. These genes include the MHC and TCR genes. However, the functional relationships among these genes on the disease susceptibility has not yet been studied. In this study, we demonstrate that the effect of the H-2s genotype from susceptible SJL/J mice overrides the resistant effect of the BALB/c TCR beta-chain gene in CXJ recombinant-inbred and BALB.S congenic mice. These results strongly suggest the presence of a hierarchy of genes involved in the immune response in Theiler's murine encephalomyelitis virus-induced demyelinating disease.