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To investigate the role of antigen receptor-mediated interactions in lymphomagenesis we have analyzed the influence of αβTCR-mediated selection on the development of spontaneous thymic lymphomas, which appear with a high (up to 50%) frequency in mice expressing a transgenic TCR specific for the male antigen (HY) in the context of H-2Db molecules. To this end we compared the kinetics and the incidence of thymic lymphomas developing in females and males with selecting (H-2b ) and non-selecting (H-2k ) MHC molecules. The kinetics of development of thymic lymphomas was similar in positively selecting (H-2b females) and non-selecting (H-2k females and males) environments but significantly slower (P , 0.01) in the negatively selecting environment (H-2b male). Injection of lymphoma cells derived from a H-2b female into the thymus of a H-2b male resulted in strong, antigen-specific inhibition of growth, indicating that the slower kinetics of lymphomagenesis in H-2b males could be due, at least partially, to the sensitivity of oncogenically transformed thymocytes to TCR-mediated negative selection. Phenotypic and functional analysis of lymphoma cells indicated that they originated from the stage of pre-TCR-dependent transition of immature CD4- CD8- to CD4+CD8+ thymocytes, which in H-2b females and males developed into tumors under different environmental pressures. These results failed to provide convincing evidence for the role of positive selection but provided a strong indication that self antigeninduced negative selection, in addition to its well established role in self tolerance, can occasionally act as a tumor surveillance mechanism by eliminating or suppressing growth of thymocytes undergoing oncogenic transformation. [ABSTRACT FROM AUTHOR]

Abstract: The orphan nuclear receptors Nur77 and Nurr1 are the members of the Nur77 family of transcription factors. We demonstrate that transcription of the Nur77 family genes was upregulated in PC12 cells following incubation with Ca2+ ionophore as well as cyclic AMP (cAMP) analog. On the other hand, cAMP analog induced strong increase, while Ca2+ ionophore induced weak increase in the transactivation activity of Nur77. We found that Nur77 and Nurr1 proteins were expressed in the nucleus following stimulation with cAMP analog but not after stimulation with Ca2+ ionophore. However, expression of Nur77 protein was increased in the cytoplasm of cells treated with Ca2+ ionophore. In conclusion, our results suggest that cAMP-induced and Ca2+-induced processes may differentially regulate activity of Nur77 at the level of translocation of Nur77 protein from the cytoplasm into the nucleus. [Copyright &y& Elsevier]

Our previous reports showed that thymic lymphomas arising in TCR transgenic mice are resistant to Ca2+-mediated apoptosis. Here we show that induction of apoptosis in thymic lymphomas involves a process that is cAMP-mediated and which depends on the activation of protein kinase A (PKA) despite the lower level of PKA type I in these lymphomas compared to thymocytes. Further, we show that treatment of the lymphomas with HA1004, a serine/threonine protein kinase inhibitor, restores their susceptibility to ionomycin-induced apoptosis. Results indicate that HA1004-induced restoration of sensitivity to ionomycin proceeds through a PKA-independent mechanism. Moreover, activation of PKA instead of its inhibition induces apoptosis of lymphoma cells. [ABSTRACT FROM AUTHOR]

Background: The anti-angiogenic activity of thrombospondin-1 (TSP-1) is suppressed in cancer cells, fact which has generated considerable interest in generating the respective therapeutic mimetics. These efforts were almost exclusively centered on peptides targeting CD36, the known TSP-1 receptor. Since the effects of these agents were less dramatic than those of full-length. TSP-1 questions could be raised about the cancer-related roles of additional TSP-1 domains and receptors, such as CD47, which bind the C-terminal sequences of this protein., Materials and Methods: The MDFB6 and B6ras cell lines were treated with the anti-CD47 antibody or C-terminal TSP-1 peptide 4N1K and cell viability was monitored using Aqueous Non-Radioactive Cell Proliferation Assay, DNA fragmentation, caspase-3 activation, and cell membrane depolarization assays. The cells were also stained with acridine orange and the anti-LC3 antibody to detect autophagy. B6ras tumours were generated in SCID-NOD mice and tumour responses to injections of the 4N1K peptide were recorded over time., Results: We demonstrated a selective loss of viability of RAS-transformed cells upon ligation of the CD47 receptor. Affected cells did not exhibit hallmarks of apoptosis, but instead were stained with acridine orange and exhibited a punctuate pattern of immunoreactivity for LC3, both features of autophagy. The 4N1K peptide administration also caused a modest but specific and significant tumour growth inhibition in vivo., Conclusion: Our study offers an additional mechanism whereby TSP-1 affects tumour cells directly, in an angiogenesis-independent and CD47-mediated manner. This is consistent with near complete loss of TSP-1 expression in RAS-transformed cells and may open new avenues for therapies involving TSP-1 peptides, in a subset of tumours.

Background: Bcl-2/adenovirus E1B 19-kDa protein-interacting protein 3 (BNIP3) is a key regulator of cell death/autophagy and can act as an effector of a necrosis-like, atypical death program. It was implicated in execution of cell death induced by cluster of differentiation 47 (CD47). Despite the postulated role of BNIP3 in the regulation of survival of cancer cells, the influence of oncogenic transformation on BNIP3 expression is unclear., Materials and Methods: The influence of oncogenic transformation on expression of BNIP3 was studied using H-ras-transformed cells. The consequences of BNIP3 expression for sensitivity to CD47-mediated cell death were assessed using tetrazolium salt-based assay., Results: Here, the enforced and endogenous expression of Ras coincided with the up-regulation of BNIP3 across a wide spectrum of cancer cells, providing the first experimental evidence that BNIP3 is a regulatory target of H-Ras. This indicated that merely the introduction of a single oncogene may result in the up-regulation of BNIP3. The consequences of CD47 ligation strongly depended on the BNIP3 presence, which in turn correlated with Ras expression. Interestingly, the indirect effect of that phenomenon was the selective sensitivity of Ras-transformed cells to CD47-mediated cell death.

We have previously described that thymic lymphomas from anti HY-TCR transgenic mice were resistant to TCR-induced (calcium-mediated) apoptosis but sensitive to induction of apoptosis by etoposide. A defect of apoptosis in these cells is located downstream of the nuclear receptor Nur77 induction and upstream from the execution of apoptosis. Furthermore, in contrast to the normal thymocytes, the lymphoma cells do not express the Fas receptor on the cell surface. Here we show that Fas ligand (FasL) induction is also abrogated in thymic lymphomas treated with ionomycin but not with etoposide, which still induced the expression of FasL and Fas receptor as well as apoptosis. It suggested a specific inhibition of the calcium-mediated signaling pathway leading to induction of the FasL expression and apoptosis of lymphomas. Sensitivity to ionomycin-induced apoptosis could be restored by FK506 treatment, which also abolished the abrogation of induction of the FasL expression. Moreover, induction of FasL was always accompanied by increase in expression of Fas receptor. These results indicate that FK506, the agent broadly used as immuno-suppressant, can sensitize resistant tumor cells to induction of apoptosis and it could be considered as a potential agent in the combined therapy of thymic lymphomas. In addition, it appeared that the Fas/FasL system can successfully serve as a molecular target for tumor therapy, even for tumor cells initially lacking expression of Fas.