David Askew

Mouse microglial cell lines differing in constitutive and interferon-γ-inducible antigen-presenting activities for naive and memory CD4+ and CD8+ T cells

We developed a panel of non-virus transformed cell lines derived from individual microglial precursors residing in the brains of normal mice. These colony stimulating factor-1-dependent cell lines are B7-1+ (CD80), Mac-1+, Mac-2+, Mac-3+, CD45+, MHC class I+, colony stimulating factor-1 receptor+, and they ingest antibody-coated particles. However, the cell lines differ in their expression of B7-2 (CD86), F4/80, Ly-6C and MHC class II molecules. They also differ in their ability to constitutively process and present antigens to naive CD4+ and CD8+ T cells, memory CD4+ and CD8+, and in the manner by which interferon gamma modulates their antigen-presenting activities. These cell lines should be valuable as models for studies on the immunobiology of the microglia.

Alloantigen presentation to naive CD8+ T cells by mouse microglia: Evidence for a distinct phenotype based on expression of surface-associated and soluble costimulatory molecules

We previously showed that approximately one‐third of mouse primary microglial clones derived from individual precursor cells residing in normal brain constitutively present alloantigens (alloAgs) to naive CD8?? T cells (Moore et al.: J Neuroimmunol 41:203, 1992). To understand the basis for this alloAg presenting (alloAgP) activity, we developed a panel of microglial cell lines that were characterized by patterns of alloAgP activity similar to that of the primary clones. Flow cytometric analysis revealed that microglia with and without alloAgP activity expressed similar levels of major histocompatibility complex class I molecules; however, CD80 (B7‐1) and CD86 (B7‐2) expression was primarily restricted to the alloAgP?? cell lines. Monoclonal antibody (Mab) to CD80 only partially blocked the proliferative response of allogeneic CD8?? T cells cocultured with the presenting cell lines, whereas Mab to CD86 completely inhibited the response, indicating a significant role for this molecule in T‐cell activation. Using an immunoassay, recombinant mouse cytokines, cytokine‐specific Mabs, and the reverse transcriptase‐polymerase chain reaction to detect specific cytokine mRNAs, we found the synthesis of interleukin (IL)‐1α, IL‐6, IL‐12, and tumor necrosis factor‐α (TNF‐α) to be restricted to the alloAgP?? cell lines. Costimulatory roles were then identified for these molecules. We conclude that the ability to present alloAg is a property of a subset of microglia that constitutively express CD86 and secrete costimulatory cytokines that promote the expansion of the alloAg‐stimulated CD8?? T cells.

Regulation of Apoptosis in Interleukin-3-Dependent Myeloid Progenitor Cells

Apoptosis plays an important role in the regulation of hematopoiesis by deleting growth factor-dependent progenitors following physiologic reductions in hemopoietins.1,2 To examine signalling pathways which may influence apoptosis in hematopoietic progenitors we have utilized the Interleukin-3 (IL-3) dependent 32Dcl3 murine myeloid cell line. 32D cells are absolutely dependent upon IL-3 for proliferation and survival,3,4 and have no propensity for developing factor-independent growth. In the absence of IL-3, 32D cells rapidly arrest in the Gl phase of the cell cycle and eventually undergo apoptosis.5

Mechanisms of IL-3 Regulated Growth and Transformation of Hematopoietic Cells

The hematopoietic system is derived from the progeny of relatively few pluripotential stem cells which differentiate along either the lymphoid or the myeloid lineages (Joyner, 1983; Williams et al., 1984; Keller et al., 1985; Lemischka et al., 1986). Interleukin 3 (IL-3) is a T-Cell derived, 28 kd glycoprotein hematopoietic growth factor which has been shown to support the proliferation and differentiation of early hematopoietic stem cells in vitro and is speculated to have a comparable function in vivo (Weinstein et al., 1986; Ihle, 1988b). Although considerable information is available concerning the mechanisms by which IL-3 regulates the growth of hematopoietic cells, the genes and the mechanisms involved in the control of differentiation of committed hematopoietic progenitors are largely unknown.