Spermatogonial stem cells (SSCs) are the foundation for spermatogenesis and, thus, preservation of a species. Because of stem cell rarity, studying their self-renewal is greatly facilitated by in vitro culture of enriched biologically active cell populations. A recently developed culture method identified glial cell line-derived neurotrophic factor (GDNF) as the essential growth factor that supports in vitro self-renewal of SSCs and results in an increase in their number. This system is a good model to study mechanisms of stem cell self-renewal because of the well defined culture conditions, enriched cell population, and available transplantation assay. By withdrawing and replacing GDNF in culture medium, we identified regulated expression of many genes by using microarray analysis. The expression levels of six of these genes were dramatically decreased by GDNF withdrawal and increased by GDNF replacement. To demonstrate the biological significance of the identified GDNF-regulated genes, we examined the importance of the most responsive of the six, bcl6b, a transcriptional repressor. By using siRNA to reduce transcript levels, Bcl6b was shown to be crucial for SSC maintenance in vitro. Moreover, evaluation of Bcl6b-null male testes revealed degeneration and͞or absence of active spermatogenesis in 24 ؎ 7% of seminiferous tubules. These data suggest that Bcl6b is an important molecule in SSC self-renewal and validate the biological relevance of the GDNF-regulated genes identified through microarray analysis. In addition, comparison of data generated in this study to other stem cell types suggests that self-renewal in SSCs is regulated by distinctly different molecular mechanisms.glial cell line-derived neurotrophic factor ͉ germ-line stem cell ͉ microarray ͉ mouse

ϩ T cells has three distinguishing characteristics that enable it to control viral infections more efficiently than does the primary response: immediate antiviral defense, more rapid production of new effector CD8 ϩ T cells, and the generation of greater numbers of effector CD8 ϩ T cells. The mechanisms involved in the first and second characteristics have been defined: the immediate antiviral response is mediated by ''effector'' memory CD8 ϩ T cells that are maintained by the cytokines IL-7 and IL-15 (1-3) and by CD4 ϩ T cells (4), and the accelerated production of new effector CD8 ϩ T cells reflects the relatively high frequency of ''central'' memory CD8 ϩ T cells that replicate in response to viral antigens (5). The mechanism underlying the capacity of the secondary response for generating larger numbers of effector CD8 ϩ T cells, however, remains unknown but presumably must be related to the factors that govern antigen-driven proliferation of CD8 ϩ T cells.

Clones of CD8+ T cells that have been selected in the primary response must have a mechanism by which they can continuously or intermittently generate new effector cells. Several years ago, this mechanism was proposed to involve a self-renewing, stem cell-like subset that could avoid the differentiating effects of interleukin-2 (IL-2). The model considered the stem cell subset to be contained within the central memory population of CD8+ T cells (T(CM)). This proposal was inconsistent with subsequent findings suggesting that all antigen-activated CD8+ T cells differentiated to effector cells (T(EFF)) during the primary response and that T(CM) developed during the memory phase by de-differentiating from effector memory cells (T(EM)). However, findings have since been reported that support the stem cell model. First, studies indicate that T(EM) do not serve as the precursors of T(CM). Second, transcriptional repressors of IL-2 signaling do enhance the memory response. Third, memory cells lacking effector functions and with a capacity to replicate in a secondary response develop in the absence of signaling through the IL-2/IL-15 receptor. Taken together, these findings suggest that antigen-activated CD8+ T cells with a stem cell-like capability for maintaining proliferative potential develop by an unknown IL-2-independent process. The challenge is now to identify this unknown pathway of clonal expansion.