Lydia Lee

Augmented induction of CD4+CD25+ Treg using monoclonal antibodies to CD200R

Background. CD200 is a transmembrane protein delivering immunoregulatory signals after engagement of CD200R. A family of CD200Rs exist (CD200R1–4) with different tissue expression and functional activity. In the presence of anti-CD200R2/3 monoclonal antibodies (mAbs), bone-marrow cells cultured in the presence of (interleukin [IL]-4+granulocyte-macrophage colony-stimulating factor) differentiate into dendritic cells (DCs), which induce CD4+CD25+Treg. The effect of these mAbs on Treg induced in anti-CD3 activated thymocyte cultures is unknown. Methods. BL/6 bone-marrow cells were cultured with GMC-SF and IL-4 in the presence/absence of anti-CD200Rs to generate DCs that induced Treg in C3H lymph-node T cells. Treg were also induced in anti–CD3/CD28-activated C3H thymocytes. Treg activity was assayed by (1) suppression of mixed leukocyte culture (MLC) in cultures using C3H stimulator spleen cells and BL/6 stimulator cells, and (2) by expression of the transcription factor, Foxp3. Results. Addition of anti-CD200R2/3 mAbs (but not anti-CD200R1) to bone-marrow cultures led to generation of DCs that induced a CD4+CD25+ (Treg) population inhibiting MLCs (C3H cells stimulated with C57BL/6 cells) in a cytotoxic T–lymphocyte-associated antigen (CTLA)-4 and transforming growth factor (TGF)–β-dependent manner. Anti-CD200R2, but not anti-R1/R3, augmented induction of Foxp3-expressing Treg from anti-CD3/CD28 activated thymocytes. Suppression in MLCs by anti-CD200R1 mAbs was dependent on IL-10 and TGF-β. Conclusions. Unlike anti-CD200R1, anti-CD200R2 both promotes development of DCs with capacity to induce Treg and directly augments thymocyte production of Treg.

Ecology of danger-dependent cytokine-boosted spontaneous abortion in the CBA x DBA/2 mouse model. I. Synergistic effect of LPS and (TNF-alpha + IFN-gamma) on pregnancy loss.

Problem:  Previous data have shown ‘danger’ signals, such as bacterial lipopolysaccharide (LPS) acting via toll‐like (tlr) receptors are required for early pregnancy failure in several murine abortion models. Indeed, the abortion rate increased in the CBA × DBA/2 model after a gestation day (gd) 7.5 injection of tumour necrosis factor (TNF)‐α + interferon (IFN)‐γ only if the LPS‐tlr signalling pathway was intact. High rates of cytokine‐boosted abortion >80% loss can be achieved in certain animal colonies, that have a high endogenous (spontaneous) rate of resorption (30–50%). A specific role for LPS has been postulated to determine both the endogenous and cytokine‐boosted losses.

Induction of tolerance-inducing antigen-presenting cells in bone marrow cultures in vitro using monoclonal antibodies to CD200R.

CD200 to CD200R interactions produce immunoregulation. We investigated whether the expression of CD200R on dendritic cell (DC) precursors affects their developmental fate. C57BL/6 bone marrow (BM) cells were cultured in vitro in the presence of (interleukin-4 + granulocyte-macrophage colony-stimulating activity) to generate allostimulatory DCs, which were in turn used to induce cytotoxic T-lymphocyte and cytokine production after culture with C3H responder spleen cells. Some marrow cultures included anti-CD200R antibodies. The inclusion of monoclonal antibodies in different isoforms of CD200R in the BM culture led to a generation of cells (tolerogenic DCs) that were unable to produce allostimulation in vitro with responder cells. Cells taken from these latter mixed leukocyte cultures (MLCs) now contained CD4+CD25+ cells able to inhibit the antigen-specific MLC response of fresh C3H responder cells to stimulation with C57BL/6 cells, but not stimulation with BALB/c cells. Tolerogenic DCs, infused in vivo into mice receiving C57BL/6 skin grafts, produced antigen-specific decreased rejection of BL/6 allografts, not BALB/c allografts, compared with mice receiving control DCs (generated from BM in the absence of anti-CD200R). The induction of CD4+CD25+ suppressor cells in MLCs using tolerogenic DCs from the initial BM cultures could be overcome by using limiting numbers of tolerogenic DCs and an excess of allostimulatory DCs derived from BM cultures maintained in the absence of anti-CD200R. These data indicate that anti-CD200R biases stem cells in BM toward the development of suppressive antigen-presenting cells, which can induce CD4+CD25+ regulatory T cells. Tolerogenic DCs have the potential to modify graft acceptance in vivo.

The use of smartphones in general and internal medicine units: A boon or a bane to the promotion of interprofessional collaboration?

Effective communication and coordination are critical components for improving collaborative care delivery among different healthcare providers who work in mobile and time-pressured environments. Increasingly, healthcare providers are exploring alternative communication technologies to help bridge the temporal and spatial issues that are often inherent in the clinical communication conundrum. Our study examined perceptions of General Internal Medicine (GIM) staff on the usage of Smartphone devices and a Webpaging system, which were implemented on the inpatient GIM units at two teaching hospitals in North America. An exploratory case study approach was employed and in-depth interviews with 31 clinicians were conducted. This data-set serves as a subset and prelude to a larger research study that examined and compared the impacts of different types of communication technologies used in five teaching hospitals. Findings from our study indicate that the use of Smartphone technology was well received among clinicians. Specifically, healthcare professionals valued the use of emails when communicating nonurgent issues and the availability of the phone function that enabled access to clinicians especially in urgent situations. Dissatisfaction, however, was expressed over the suitability of these smartphone features in different communication contexts as well as discrepancies between clinicians over the appropriate use of the communication modes. Future interventions in communication technology should take into considerations how communication mediums and situational contexts (e.g. urgent and nonurgent patient issues) impact interprofessional interactions.

An Interaction between CD200 and Monoclonal Antibody Agonists to CD200R2 in Development of Dendritic Cells That Preferentially Induce Populations of CD4+CD25+ T Regulatory Cells

In previous studies we reported that while interaction between the relatively ubiquitously expressed molecule CD200 and one of its receptors, CD200R1, resulted in direct suppression of alloreactivity, engagement of alternate receptors led instead to altered differentiation of dendritic cells (DCs) from marrow precursors, which could in turn foster development of Foxp3+ regulatory T cells. We have explored this effect of engagement of alternate receptors by using a monoclonal agonist Ab to CD200R2 and investigating expression of TLRs on DCs induced in vivo and in vitro after CD200 stimulation in mice in which the gene encoding CD200R1 was deleted. CD200 stimulation was achieved by using either a soluble form of CD200 (CD200Fc) or overexpression of CD200 as a doxycycline-inducible transgene. Although broadly similar effects were seen, consistent with the hypothesis that triggering of CD200R2 does produce DCs with a characteristic TLR repertoire, there are subtle differences in suppression of alloreactivity achieved by CD200 delivered in these two manners, which is consistent with a complexity of CD200:CD200R engagement not previously appreciated.

A role for altered TLR gene expression in association with increased expression of CD200R in the induction of mucosal tissue CD4+ Treg in aged mice following gavage with a liver extract along with intramuscular monophosphoryl lipid A (MPLA) injection

Previous studies showed a fetal sheep liver extract (FSLE), in association with LPS, injected into aged (>20 months) mice reversed the altered polarization (increased IL-4 and IL-10 with decreased IL-2 and IFN-gamma) in cytokine production seen from ConA stimulated lymphoid cells of those mice. Aged mice show a >60% decline in numbers and suppressive function of both CD4(+)CD25(+)Foxp3(+)Treg and so-called Tr3 (CD4(+)TGFbeta(+)). Their number/function is restored to levels seen in control (8-week-old) mice by FSLE. We have reported at length on the ability of a novel pair of immunoregulatory molecules, members of the TREM family, namely CD200:CD200R, to control development of dendritic cells (DCs) which themselves regulate production of Foxp3(+) Treg. The latter express a distinct subset of TLRs which control their function. We report that a feature of the altered Treg expression following combined treatment with FSLE and monophosphoryl lipid A, MPLA (a bioactive component of lipid A of LPS) is the altered gene expression both of distinct subsets of TLRs and of CD200Rs. We speculate that this may represent one of the mechanisms by which FSLE and MPLA alter immunity in aged mice.