Fumiko Suenaga

Strength of inflammatory stimuli during the early expansion phase plays a crucial role in the effector versus memory cell fate decision of CD8(+) T cells. But it is not known how early lymphocyte distribution after infection has an impact on this process. We demonstrate that the chemokine receptor CXCR3 is involved in promoting CD8(+) T cell commitment to an effector fate rather than a memory fate by regulating T cell recruitment to an antigen/inflammation site. After systemic viral or bacterial infection, the contraction of CXCR3(-/-) antigen-specific CD8(+) T cells is significantly attenuated, resulting in massive accumulation of fully functional memory CD8(+) T cells. Early after infection, CXCR3(-/-) antigen-specific CD8(+) T cells fail to cluster at the marginal zone in the spleen where inflammatory cytokines such as IL-12 and IFN-α are abundant, thus receiving relatively weak inflammatory stimuli. Consequently, CXCR3(-/-) CD8(+) T cells exhibit transient expression of CD25 and preferentially differentiate into memory precursor effector cells as compared with wild-type CD8(+) T cells. This series of events has important implications for development of vaccination strategies to generate increased numbers of antigen-specific memory CD8(+) T cells via inhibition of CXCR3-mediated T cell migration to inflamed microenvironments.

We previously demonstrated that VEGF and its receptors were expressed in human herniated discs (HD). TNF-alpha induced VEGF, resulting in neovascularization of disc tissues in a model of HD. The goal of the current research was to investigate the precise role of TNF-alpha-induced VEGF and the mechanism of angiogenesis in disc tissues. We performed ELISAs, Western blots, and immunohistological examinations to assess the role of TNF-alpha-induced VEGF using organ disc cultures with wild type, TNF receptor 1-null (TNF-RI(null)), or TNF receptor 2-null (TNF-RII(null)) mice. VEGF induction was inhibited when we used TNF-RI(null)-derived disc tissues. NF-kappaB pathway inhibitors also strongly suppressed VEGF induction. Thus, TNF-alpha induced VEGF expression in disc cells primarily through the NF-kappaB pathway. In addition, VEGF immunoreactivity was detected predominantly in annulus fibrosus cells and increased after TNF-alpha stimulation. TNF-alpha treatment also resulted in CD31 expression on endothelial cells and formation of an anastomosing network. In contrast, angiogenic activity was strongly inhibited in the presence of NF-kappaB inhibitors or anti-VEGF antibody. Our data show angiogenesis activity in disc tissues is regulated by VEGF and the NF-kappaB pathway, both of which are induced by TNF-alpha. The level of angiogenic activity in disc tissues was closely related to aging. Because neovascularization of HD is indispensable for HD resorption, the prognosis of HD and the rate of the resorption process in patients may vary as a function of the patient's age.

OBJECTIVE: To determine whether thymic stromal lymphopoietin (TSLP) plays a role in the resorption of herniated disc tissue. METHODS: The expression of TSLP messenger RNA (mRNA) and protein in mouse intervertebral disc cells was assessed by quantitative real-time polymerase chain reaction, enzyme-linked immunosorbent assay (ELISA), and immunohistochemical analysis. The ability of mouse intervertebral disc cells to respond to TSLP stimulation was examined by Western blot analysis, ELISA, and protein array analysis. Intracellular signaling pathways involved in TSLP signaling in mouse intervertebral disc cells were investigated using several chemical inhibitors. The role of TSLP in macrophage migration into the intervertebral disc was assessed by in vitro migration assay. Finally, TSLP expression in clinical specimens derived from patients with a herniated disc was examined by immunohistochemistry. RESULTS: Mouse intervertebral disc cells expressed TSLP mRNA and protein upon stimulation with NF-kappaB-activating ligands such as tumor necrosis factor alpha. In addition, the mouse intervertebral disc cells expressed the TSLP receptor and produced monocyte chemotactic protein 1 (MCP-1; CCL2) and macrophage colony-stimulating factor in response to TSLP stimulation. Both anulus fibrosus and nucleus pulposus intervertebral disc cells expressed MCP-1 upon TSLP stimulation, which was mediated via the phosphatidylinositol 3-kinase/Akt pathway. Consistently, the supernatants of TSLP-activated intervertebral disc cultures had the capacity to induce macrophage migration in an MCP-1-dependent manner. Finally, TSLP and MCP-1 were coexpressed in human herniated disc specimens in which macrophage infiltration into the tissue was observed. CONCLUSION: TSLP induced by NF-kappaB-activating ligands in intervertebral discs may contribute to the recruitment of macrophages to the intervertebral disc by stimulating MCP-1 production and may be involved in the resorption of herniated disc tissue.