Despite playing a critical role in the development of naive T cells, the thymus is involuted with age. Whether a single age-associated defect or multiple aberrations contribute to thymic involution remains controversial. Here, we determined molecular aberrations in the thymocyte and epithelium compartments of the aging thymus. We demonstrated that total thymocyte numbers declined with a stepwise kinetics; clear demarcations occurred at 1.5, 3, 12 and 22 months of age. By quantitative PCR, a 2.4-fold reduction in the copies of signal joint TCR-excised circle (sjTREC)/10(5) thymocytes was first detected at 3 months; no further reduction observed thereafter. Nevertheless, the combined reductions in thymocyte numbers and sjTREC/10(5) cells caused a 7-fold decrease in sjTREC/thymus by 3 months, 21-fold by 18 months and 72-fold by 22 months as compared to 1 month. We showed aberration in expression of E2A, a transcription regulator critical for TCR beta rearrangement. While E2A expression declined 3-fold by 3 months and 18-fold by 7 months, expression of LMO2, a negative regulator of E2A activities, increased 5-fold by 18 months. Interestingly, expression of pre-T alpha and its transcriptional regulator HEB were not reduced with age. Furthermore, keratin-8 expression, specific for cortical thymic epithelium, declined 3-fold by 7 months and remained stable thereafter. In contrast, Foxn1 expression was reduced 3-fold by 3 months, 16-fold by 12 months and 37-fold by 18 months. IL-7 expression was not reduced until 7 months and reached 15-fold reduction by 22 months. Thus, the data demonstrate that thymic involution results not from a single defect, but culminates from an array of molecular aberrations in both the developing thymocytes and thymic epithelials.

The forkhead box n1 (Foxn1) transcription factor is essential for thymic organogenesis during embryonic development; however, a functional role of Foxn1 in the postnatal thymus is less well understood. We developed Foxn1 transgenic mice (Foxn1Tg), in which overexpression of Foxn1 is driven by the human keratin-14 promoter. Expression of the Foxn1 transgene increased the endogenous Foxn1 levels. In aged mice, overexpression of Foxn1 in the thymus attenuated the decline in thymocyte numbers, prevented the decline in frequency of early thymic progenitors, and generated a higher number of signal joint TCR excised circle. Histologic studies revealed that structural alterations associated with thymic involution were diminished in aged Foxn1 Tg.

The purpose of physiological cell death is the noninflammatory clearance of cells that have become inappropriate or nonfunctional. Consistent with this function, the recognition of apoptotic cells by professional phagocytes, including macrophages and dendritic cells, triggers a set of potent anti-inflammatory responses manifest on multiple levels. The immediate-early inhibition of proinflammatory cytokine gene transcription in the phagocyte is a proximate consequence of recognition of the apoptotic corpse, independent of subsequent engulfment and soluble factor involvement. Here, we show that recognition is linked to a characteristic signature of responses, including MAPK signaling events and the ablation of proinflammatory transcription and cytokine secretion. Specific recognition and response occurs without regard to the origin (species, tissue type, or suicidal stimulus) of the apoptotic cell and does not involve Toll-like receptor signaling. These features mark this as an innate immunity fundamentally distinct from the discrimination of "self" versus "other" considered to be the hallmark of conventional immunity. This profound unconventional innate immune discrimination of effete from live cells is as ubiquitous as apoptotic cell death itself, manifest by professional and nonprofessional phagocytes and nonphagocytic cell types alike. Innate apoptotic immunity provides an intrinsic anti-inflammatory circuit that attenuates proinflammatory responses dynamically and may act systemically as a powerful physiological regulator of immunity.

Although it is reported that B lymphopoiesis declines with age, the precursor stage(s) affected and the age of onset are ambiguous. Each progressive phase of B cell differentiation has distinct requirements; therefore, precise identification of the stage(s) that decline would yield insight into the age-related mechanisms affecting humoral immunity. We analysed the composition of B lineage cells of mice 1, 4, 12 and 24 months of age using flow cytometry. Numbers of prepro-B and pro-B cells were unchanged, and a profound decrease occurred only in the numbers of pre-B cells. This decrease occurred in two distinct phases: between 1 and 4 months and between 12 and 24 months. Notably, the numbers of newly formed B cells did not decline in parallel, suggesting that mechanisms are established to overcome the deficiency of pre-B cells. Since the age-related changes are limited to the pre-B cell stage, we hypothesized that the impairment acts at the pro-B to pre-B transition. We therefore evaluated whether the pro-B cells or the supporting stromal cells, which are necessary for normal progression of this stage, changed with age. The ability of pro-B cells to proliferate in the presence of stromal cells was reduced by 24 months of age, as was the ability of the stromal cells to support pro-B cell proliferation. In contrast, the ability to mature into IgM+ cells was unchanged. Thus, strategies that supplement the stromal environment may enhance B lymphopoiesis in aged animals.

Protective circulating Abs originate primarily from long-lived plasma cells in the bone marrow. However, the molecular and cellular basis of plasma cell longevity is unknown. We investigated the capacity of primary bone marrow-derived stromal cells to maintain plasma cell viability in vitro. Plasma cells purified from the bone marrow or lymph nodes died rapidly when plated in media, but a subpopulation of plasma cells survived and secreted high levels of Ab for up to 4 wk when cocultured with stromal cells. Ab secretion was inhibited by the addition of anti-very late Ag-4 to plasma cell/stromal cell cocultures indicating that direct interactions occur and are necessary between stromal cells and plasma cells. The addition of rIL-6 to plasma cells cultured in media alone partially relieved the sharp decline in Ab secretion observed in the absence of stromal cells. Moreover, when stromal cells from IL-6(-/-) mice were used in plasma cell/stromal cell cocultures, Ab levels decreased 80% after 7 days as compared with wild-type stromal cells. Further, IL-6 mRNA message was induced in stromal cells by coculture with plasma cells. These data indicate that bone marrow plasma cells are not intrinsically long-lived, but rather that plasma cells contact and modify bone marrow stromal cells to provide survival factors.