Some of these may indeed be true double positives, but my experience in sorting these only yields single positive events. If the duals do exist they are extremly rare. Jim Houston > ---------- > From: Mark Kukuruga > Sent: Thursday, November 8, 2001 11:17 AM > To: Cytometry Mailing List > Subject: RE: cd4 cd8 coexpression > > > . . . there's been quite a lot of discussion on this topic, and just as > many opinions > . . . you've hit on a point here, though . . . > Has anyone "sorted" these dual positives to further ID them? IF they're > real (not > doublets), they'll still be double positive after sorting. If they're > aggregates . . . > Also, if we're talking cell-cell coupling, this would be visible via > microscopic > evaluation of the sorted cells. > SO . . . ? > MAK. > > >>> Joseph Wright <JWright@niaid.nih.gov> 11/05/01 11:38AM >>> > I am working on mice, not humans, but I also see a small but consistent > population of CD4+/CD8+ cells in my current experiment. I am trying to > sort > this out, but it looks like there may be a bit of literature in animal > models to suggest that this is a real population of (perhaps) functioning > cells. I'm trying to gather this literature now, so I'd love to hear from > anyone who knows more about it than I do. > > Also, in J Virol's new issue there is an article which appears to be of > relevance, attached below. I've only seen this abstract, haven't yet got > the > article. > > best regards, > Joe Wright > Ghost Lab/ Laboratory of Cellular and Molecular Immunology > NIAID > ______________________________________ > J Virol 2001 Dec 1;75(23):11555-11564Activated Peripheral CD8 Lymphocytes > Express > CD4 In Vivo and Are Targets for > Infection by Human Immunodeficiency Virus Type 1.Imlach S, McBreen S, > Shirafuji T, > Leen C, Bell JE, Simmonds P.Laboratory for Clinical and Molecular > Virology, University > of Edinburgh, > Summerhall, Edinburgh EH9 1QH, United Kingdom.There is increasing evidence > that CD8 > lymphocytes may represent targets for > infection by human immunodeficiency virus type 1 (HIV-1) in vivo whose > destruction may contribute to the loss of immune function underlying AIDS. > HIV-1 may infect thymic precursor cells destined to become CD4 and CD8 > lymphocytes and contribute to the numerical decline in both subsets on > disease progression. There is also evidence for the induction of CD4 > expression and susceptibility to infection by HIV-1 of CD8 lymphocytes > activated in vitro. To investigate the relationship between CD8 activation > and infection by HIV-1 in vivo, activated subsets of CD8 lymphocytes in > peripheral blood mononuclear cells (PBMCs) of HIV-seropositive individuals > were investigated for CD4 expression and HIV infection. Activated CD8 > lymphocytes were identified by expression of CD69, CD71, and the human > leukocyte antigen (HLA) class II, the beta-chain of CD8, and the RO > isoform > of CD45. CD4(+) and CD4(-) CD8 lymphocytes, CD4 lymphocytes, other T > cells, > and non-T cells were purified using paramagnetic beads, and proviral > sequences were quantified by PCR using primers from the long terminal > repeat > region. Frequencies of activated CD8 lymphocytes were higher in > HIV-infected > study subjects than in seronegative controls, and they frequently > coexpressed CD4 (mean frequencies on CD69(+), CD71(+), and HLA class II(+) > cells of 23, 37, and 8%, respectively, compared with 1 to 2% for > nonactivated CD8 lymphocytes). The level of CD4 expression of the > double-positive population approached that of mature CD4 lymphocytes. That > CD4 expression renders CD8 cell susceptible to infection was indicated by > their high frequency of infection in vivo; infected CD4(+) CD8 lymphocytes > accounted for between 3 and 72% of the total proviral load in PBMCs from > five of the eight study subjects investigated, despite these cells > representing a small component of the PBMC population (<3%). Combined, > these > findings provide evidence that antigenic stimulation of CD8 lymphocytes in > vivo induces CD4 expression that renders them susceptible to HIV infection > and destruction. The specific targeting of responding CD8 lymphocytes may > provide a functional explanation for the previously observed impairment of > cytotoxic T-lymphocyte (CTL) function disproportionate to their numerical > decline in AIDS and for the deletion of specific clones of CTLs responding > to HIV antigens.PMID: 11689637 [PubMed - as supplied by publisher] > > > ---------- > > From: Howard.Gale@med.va.gov > > Sent: Friday, November 2, 2001 8:12 AM > > To: Cytometry Mailing List > > Subject: RE: cd4 cd8 coexpression > > > > > > CD4+/CD8+/CD3+ cells are 1-4% range of the lymphocyte population from > our > > HIV+ patients. Most of the cells are brightCD4+/dimCD8+ but all > > combinations > > of bright and dim are possible. We include these dual positive cells in > > both > > the CD3+/CD4+ and CD3+/CD8+ counts. What do other labs do with these > cells > > and why? > > > > -----Original Message----- > > From: Kenneth Ault [mailto:aultk@mmc.org] > > Sent: Wednesday, October 31, 2001 7:32 PM > > To: cyto-inbox > > Subject: Re: cd4 cd8 coexpression > > > > > > > > A phenomenon frequently forgotten is coincidence. If two cells enter > the > > observation > > volume at the same time they will be seen as one event with the > properties > > of both cells. > > This is a very common problem in my world (platelets) and should be > > considered as a > > possible explanation for any kind of unexpected dual expression of > > markers. > > It would > > be interesting to know if the frequency of CD4/CD8 doubles changes as > the > > particle > > flow rate changes (i.e change the sample pressure or dilute the > specimen.) > > > > Ken Ault > > > > > >
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