We investigated the biological role of CC chemokines in the Th1-mediated pathogenesis of spontaneous type I diabetes in nonobese diabetic (NOD) mice. Whereas an elevated ratio of macrophage inflammatory protein-1α (MIP-1α):MIP-1β in the pancreas correlated with destructive insulitis and progression to diabetes in NOD mice, a decreased intrapancreatic MIP-1α:MIP-1β ratio was observed in nonobese diabetes-resistant (NOR) mice. IL-4 treatment, which prevents diabetes in NOD mice by polarizing intraislet Th2 responses, decreased CCR5 expression in islets and potentiated a high ratio of MIP-1β and monocyte chemotactic protein-1 (MCP-1): MIP-1α in the pancreas. Furthermore, NOD.MIP-1α−/− mice exhibited reduced destructive insulitis and were protected from diabetes. Neutralization of MIP-1α with specific Abs following transfer of diabetogenic T cells delayed the onset of diabetes in NOD.Scid recipients. These studies illustrate that the temporal expression of certain CC chemokines, particularly MIP-1α, and the CCR5 chemokine receptor in the pancreas is associated with the development of insulitis and spontaneous type I diabetes.

We examined the extravasation and subsequent migration and growth of murine mammary tumor cell lines (D2A1 and D2.OR) which differ in their metastatic ability in lung and liver, invasiveness in vitro and expression of the cysteine proteinase cathepsin L. In light of the differences in invasiveness and cathepsin L expression, we hypothesized that during hematogenous metastasis the two cell lines would differ primarily in their ability to extravasate. We used in vivo videomicroscopy of mouse liver and chick embryo chorioallantoic membrane to examine the process and timing of extravasation and subsequent steps in metastasis for these cell lines. In contrast to our expectations, no differences were found between the cell lines in either the timing or mechanism of extravasation, at least 95% of cells having extravasated by 3 days after injection. However, after extravasation, the more metastatic and invasive D2A1 cells showed a greater ability to migrate to sites which favor tumor growth and to replicate to form micrometastases. These studies point to post-extravasation events (migration and growth) as being critical in metastasis formation.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is expressed in different tissues and cells, including pancreas and lymphocytes, and can induce apoptosis in various tumor cells but not in most normal cells. The specific roles of TRAIL in health and disease remain unclear. Here we show by cDNA array analyses that TRAIL gene expression is upregulated in pancreatic islets during the development of autoimmune type 1 diabetes in nonobese diabetic (NOD) mice and in Min6 islet beta-cells activated by TNF-alpha + interferon-gamma. However, stimulation of freshly isolated pancreatic islets or Min6 cells with TRAIL did not induce their apoptosis. TRAIL blockade exacerbates the onset of type 1 diabetes in NOD.Scid recipients of transferred diabetogenic T-cells and in cyclophosphamide-treated NOD mice. TRAIL inhibits the proliferation of NOD diabetogenic T-cells by suppressing interleukin (IL)-2 production and cell cycle progression, and this inhibition can be rescued in the presence of exogenous IL-2. cDNA array and Western blot analyses indicate that TRAIL upregulates the expression of the cdk inhibitor p27(kip1). Our data suggest that TRAIL is an important immune regulator of the development of type 1 diabetes.

Activation-induced cell death (AICD) is a mechanism of peripheral T cell tolerance that depends upon an interaction between Fas and Fas ligand (FasL). Although c-Jun NH2-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) may be involved in apoptosis in various cell types, the mode of regulation of FasL expression during AICD in T cells by these two MAPKs is incompletely understood. To investigate the regulatory roles of these two MAPKs, we analyzed the kinetics of TCR-induced p38 MAPK and JNK activity and their regulation of FasL expression and AICD. We report that both JNK and p38 MAPK regulate AICD in T cells. Our data suggest a novel model of T cell AICD in which p38 MAPK acts early to initiate FasL expression and the Fas-mediated activation of caspases. Subsequently, caspases stimulate JNK to further upregulate FasL expression. Thus, p38 MAPK and downstream JNK converge to regulate FasL expression at different times after T cell receptor stimulation to elicit maximum AICD.

Escape of cancer cells from the circulation (extravasation) is thought to be a major rate-limiting step in metastasis, with few cells being able to extravasate. Furthermore, highly metastatic cells are believed to extravasate more readily than poorly metastatic cells. We assessed in vivo the extravasation ability of highly metastatic ras-transformed NIH 3T3 cells (PAP2) versus control nontumorigenic nontransformed NIH 3T3 cells and primary mouse embryo fibroblasts. Fluorescently labeled cells were injected intravenously into chicken embryo chorioallantoic membrane and analyzed by intravital videomicroscopy. The chorioallantoic membrane is an appropriate model for studying extravasation, since, at the embryonic stage used, the microvasculature exhibits a continuous basement membrane and adult permeability properties. The kinetics of extravasation were assessed by determining whether individual cells (n = 1481) were intravascular, extravascular, or in the process of extravasation, at 3, 6, and 24 h after injection. Contrary to expectations, our results showed that all three cell types extravasated with the same kinetics. By 24 h after injection >89% of observed cells had completed extravasation from the capillary plexus. After extravasation, individual fibroblasts of all cell types demonstrated preferential migration within the mesenchymal layer toward arterioles, not to venules or lymphatics. Thus in this model and for these cells, extravasation is independent of metastatic ability. This suggests that the ability to extravasate in vivo is not necessarily predictive of subsequent metastasis formation, and that postextravasation events may be key determinants in metastasis.Metastasis, the spread of cancer cells from a primary tumor to distant sites, is the major cause of death from cancer. Mortality results from the direct anatomical and physiological effects of metastases on other organ systems (e.g., brain and liver) or sometimes from complications associated with treatment (1, 2). The metastatic process is thought to include the following steps: detachment of cancer cells from the primary tumor, invasion of surrounding tissue, entrance into blood or lymphatic vessels (intravasation), transport to new sites, escape from the microvasculature (extravasation), invasion of target tissue, and growth of metastatic tumors (3-5). Cancer cells also must evade the immune system throughout the metastatic process.Extravasation is thought to be a major rate-limiting step in metastasis, with only a few cancer cells capable of degrading the basement membrane and extracellular matrix to escape from microvessels. Moreover, poorly metastatic cells are believed to extravasate less readily than highly metastatic cells (4, 6-8). Our recent results from intravital videomicroscopy have raised questions about these views. Using in vivo assays with a variety of cell types (including melanoma and mammary carcinoma) in chicken embryo chorioallantoic membrane (CAM) and mouse liver, we found that the majority of observed cancer cells...