ObjectivesSynovial fibroblasts and osteoblasts generate active glucocorticoids by means of the 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) enzyme. This activity increases in response to proinflammatory cytokines or glucocorticoids. During inflammatory arthritis synovium and bone are exposed to both these factors. This study hypothesised that glucocorticoids magnify the effects of inflammatory cytokines on local glucocorticoid production in both synovium and bone.MethodsThe effects of inflammatory cytokines (IL-1β/tumour necrosis factor alpha; TNFα) and glucocorticoids, alone or combined, were assessed on the expression and activity of 11β-HSD1 in primary synovial fibroblasts, primary human osteoblasts and MG-63 osteosarcoma cells. A range of other target genes and cell types were used to examine the specificity of effects. Functional consequences were assessed using IL-6 ELISA.ResultsIn synovial fibroblasts and osteoblasts, treatment with cytokines or glucocorticoids in isolation induced 11β-HSD1 expression and activity. However, in combination, 11β-HSD1 expression, activity and functional consequences were induced synergistically to a level not seen with isolated treatments. This effect was seen in normal skin fibroblasts but not foreskin fibroblasts or adipocytes and was only seen for the 11β-HSD1 gene. Synergistic induction had functional consequences on IL-6 production.ConclusionsCombined treatment with inflammatory cytokines and glucocorticoids synergistically induces 11β-HSD1 expression and activity in synovial fibroblasts and osteoblasts, providing a mechanism by which synovium and bone can interact to enhance anti-inflammatory responses by increasing localised glucocorticoid levels. However, the synergistic induction of 11β-HSD1 might also cause detrimental glucocorticoid accumulation in bone or surrounding tissues.

ObjectiveTissue glucocorticoid (GC) levels are regulated by the GC-activating enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). This enzyme is expressed in cells and tissues arising from mesenchymal stromal cells. Proinflammatory cytokines dramatically increase expression of 11β-HSD1 in stromal cells, an effect that has been implicated in inflammatory arthritis, osteoporosis, obesity, and myopathy. Additionally, GCs act synergistically with proinflammatory cytokines to further increase enzyme expression. The present study was undertaken to investigate the mechanisms underlying this regulation.MethodsGene reporter analysis, rapid amplification of complementary DNA ends (RACE), chemical inhibition experiments, and genetic disruption of intracellular signaling pathways in mouse embryonic fibroblasts (MEFs) were used to define the molecular mechanisms underlying the regulation of 11β-HSD1 expression.ResultsGene reporter, RACE, and chemical inhibitor studies demonstrated that the increase in 11β-HSD1 expression with tumor necrosis factor α (TNFα)/interleukin-1β (IL-1β) occurred via the proximal HSD11B1 gene promoter and depended on NF-κB signaling. These findings were confirmed using MEFs with targeted disruption of NF-κB signaling, in which RelA (p65) deletion prevented TNFα/IL-1β induction of 11β-HSD1. GC treatment did not prevent TNFα-induced NF-κB nuclear translocation. The synergistic enhancement of TNFα-induced 11β-HSD1 expression with GCs was reproduced by specific inhibitors of p38 MAPK. Inhibitor and gene deletion studies indicated that the effects of GCs on p38 MAPK activity occurred primarily through induction of dual-specificity phosphatase 1 expression.ConclusionThe mechanism by which stromal cell expression of 11β-HSD1 is regulated is novel and distinct from that in other tissues. These findings open new opportunities for development of therapeutic interventions aimed at inhibiting or stimulating local GC levels in cells of mesenchymal stromal lineage during inflammation.

T-cell immunity is important for controlling Kaposi sarcoma-associated herpesvirus (KSHV) diseases IntroductionKaposi sarcoma-associated herpesvirus (KSHV) is a lymphotropic human herpesvirus with oncogenic potential. KSHV infects endothelial and B cells where it can cause malignancies of these cell types, namely, Kaposi sarcoma (KS) and primary effusion lymphoma (PEL), respectively, 1 and it is associated with the B-cell pathology multicentric Castleman disease (MCD). The HIV epidemic has made these diseases significantly more prevalent, with KS being the most frequently reported HIV-associated malignancy.An important finding from studying KS patients either with HIV, or those immunosuppressed after solid organ transplantation, is that this malignancy can resolve on restoration of immune function through the administration of highly active antiretroviral therapy (HAART) 2 or relaxation of immunosuppression, 3 respectively. These findings and the increased incidence of PEL and MCD in HIV patients 4 suggest that T-cell immunity is critical for control of KSHV infection and disease. Potential immune targets in KS include the latent antigens, namely, the genome maintenance protein LANA that is expressed in all infected cells and malignancies, the viral cyclin vCyclin, the antiapoptotic multifunctional protein vFLIP, and Kaposin. PELs and infected cells in MCD also express at least 2 other proteins, the viral IL-6 and the immunomodulatory and antiapoptotic protein vIRF3.Relatively low T-cell responses to LANA and Kaposin have been described in healthy immunocompetent donors; 5,6 however, most studies have been undertaken using donors with a disrupted immune system or in disease settings, focusing on responses to LANA and Kaposin. Thus, HIV-infected patients on HAART who control KSHV have been found to make T-cell responses to LANA, MCD patients make responses comparable to HIV patients on HAART, but patients with KS disease have very weak or no detectable responses. [7][8][9][10][11] The administration of HAART to HIVassociated KS patients has been associated with the detection and increase in KSHV-specific responses over time. 2,12 Little is known, however, about the size of responses made to the potential tumor antigens vFLIP or vCyclin in healthy donors or patients with disease.How T-cell control is exercised over KSHV-associated malignancies is unclear, particularly in view of the immune evasion mechanisms used by the virus. Thus, LANA encodes an extensive acidic repeat sequence that inhibits efficient synthesis and proteasomal degradation of itself. 13 This strategy, also used by the Epstein-Barr virus (EBV) homolog EBNA1, 14 limits the supply of peptides for presentation to CD8 ϩ T cells. 15 KSHV-specific T-cell killing of infected endothelial cells has not been tested; however, such in vitro infected cells transiently express K5 16 that functions to down-regulate surface HLA class I and other costimulatory molecules. No studies have been performed on the ability of T cells to recognize KSHV-infected cells fro...