cAMP regulates 11β‑hydroxysteroid dehydrogenase‑2 and Sp1 expression in MLO‑Y4/MC3T3‑E1 cells

Abstract

11β-hydroxysteroid dehydrogenase-2 (11β-HSD2) is one of the key enzymes in glucocorticoid metabolism, which can inactivate local corticosterone and regulate the level of active glucocorticoid in tissues. The expression of 11β-HSD2 and its regulatory pathway serve an important role in the apoptosis of steroid induced osteonecrosis of the femoral head (SANFH). The present study aimed to identify the regulatory effects of cAMP on the expression of Sp1 transcription factor (Sp1) and 11β-HSD2 in osteocytes at the cellular level. Murine long bone osteocyte Y4 (MLO-Y4) clone cells and mouse embryo osteoblast-like (MC3T3-E1) cells were cultured in vitro with adenylate cyclase activator or inhibitor (forskolin and SQ22536, respectively) to investigate the effects of alterations to intracellular cAMP levels. mRNA and protein expression levels of Sp1 and 11β-HSD2 were detected by reverse transcription-quantitative PCR and western blotting, respectively. Compared with the negative control group, the mRNA and protein expression levels of Sp1 were significantly increased in the activation group, whereas Sp1 expression levels were significantly decreased in the inhibition group. Similarly, compared with the negative control group, the mRNA and protein expression levels of 11β-HSD2 were significantly increased in the activator group, but significantly decreased in the inhibitor group. The aforementioned results indicated that intracellular cAMP levels significantly regulated the expression of Sp1 and 11β-HSD2 in mouse osteocytes and osteoblasts. Therefore, the present study suggested a potential therapeutic strategy for the prevention of osteonecrosis of the femoral head.

Gαs-Coupled CGRP Receptor Signaling Axis from the Trigeminal Ganglion Neuron to Odontoblast Negatively Regulates Dentin Mineralization

“…A high-K + solution (91 mM NaCl, 50 mM KCl, 2.5 mM CaCl 2 , 0.5 mM MgCl 2 , 10 mM HEPES, 10 mM glucose, and 12 mM NaHCO 3 ; pH = 7.4) was used to discern TG neurons from glial cells by the activation of depolarization-induced increases in the concentration of intracellular free Ca 2+ in the neurons. An AC activator, forskolin (FSK) [ 14 , 15 ]; an AC inhibitor, SQ22536 [ 16 , 17 ]; a non-selective CGRP receptor agonist, CGRP (rat) [ 18 ]; and a non-selective CGRP receptor antagonist, BIBN 4096 [ 19 ], were obtained from R&D Systems, Inc. (Minneapolis, MN, USA). Stock solutions of FSK, SQ22536, and BIBN 4096 were prepared in dimethyl sulfoxide using ultrapure water (Millipore, MA, USA).…”

Associations Between Aldosterone-Renin-Ratio and Bone Parameters Derived from Peripheral Quantitative Computed Tomography and Impact Microindentation in Men

“…In particular, mineralocorticoid receptors are present in bone tissue [ 5 ] and could affect bone through several possible mechanisms [ 6 , 7 ]. Osteoblasts and osteocytes express 11β-hydroxysteroid dehydrogenase-2, an enzyme that inactivates cortisol and allows aldosterone to function [ 8 ]. Circulating cortisol concentrations are 1000 times higher than circulating aldosterone and in tissues that do not express 11β-hydroxysteroid dehydrogenase-2, mineralocorticoid receptors bind almost exclusively to cortisol.…”