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.

Abstract-Rodents studies suggest that androgens are involved in sex-specific differences in blood pressure. In humans, there is no difference in blood pressure between boys and girls, but after puberty, blood pressure increases more in men than in women. We investigated androgen-dependent regulation of the ␣-subunit of the epithelial sodium channel (␣EnaC) in human kidney and in the human renal cell line immortalized human renal proximal tubular cell line (HKC-8).We used microarray technique to analyze androgen-dependent gene regulation and performed quantitative RT-PCR for verification. Promoter constructs for human ␣ENaC were used in transfection studies to analyze the regulation by testosterone. We investigated the in vivo effect of testosterone on ␣ENaC in a rat model and used the mouse collecting duct cell line M-1 for transepithelial electrophysiological measurements. The androgen receptor (AR) was expressed in male kidney and HKC-8 cells. ␣ENaC mRNA expression increased 2-to 3-fold after treatment with testosterone in HKC-8 cells. The induction by testosterone was completely blocked by adding the AR antagonist flutamide. Analysis of the ␣ENaC promoter sequence identified a putative AR response element (ARE) located 140 nucleotides upstream from the transcription start site. HKC-8 cell transfection studies showed that testosterone directly upregulated gene expression via this ARE. In vivo, testosterone treatment of orchiectomized rats resulted in an increased renal ␣ENaC mRNA expression. In testosterone-treated mouse M-1 cells, amiloride caused a significant stronger decrease in short circuit current than in control cells. These data show that ␣ENaC expression is directly regulated by androgens in vitro and in vivo and highlight a potential mechanism explaining the reported gender differences in blood pressure. Key Words: blood pressure Ⅲ gender Ⅲ kidney Ⅲ sodium channels A ndrogens are known to play an important role in renal tubular epithelial cell growth, hypertrophy, and erythropoetin production, and may be important determinants of sex-specific differences in blood pressure. 1 In spontaneously hypertensive rats, males have higher blood pressures than females, 2 an effect that is reversed by male castration. Furthermore, increases in blood pressure were observed in castrated male and female rats after the administration of testosterone, 2,3 whereas administration of the androgen receptor (AR) antagonist flutamide attenuated hypertension. 4 These studies strongly implicate androgens in the regulation of blood pressure in the rat, but the mechanisms for this are still unknown.In humans, there is no difference in blood pressure between boys and girls, but during and after puberty, boys show higher blood pressure than age-matched girls, 5,6 and men have a higher overall mean arterial pressure than women, 7 regardless of ethnic origin. 8 The higher blood pressure and stronger progression of hypertension in men is associated with a higher risk and mortality for cardiovascular diseases than in women. 9,10 Alt...

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.