Glucocorticoids induce a near-total suppression of hyaluronan synthase mRNA in dermal fibroblasts and in osteoblasts: a molecular mechanism contributing to organ atrophy

Abstract

Glucocorticoid (GC) administration induces atrophy of skin, bone, and other organs, partly by reducing tissue content of glycosaminoglycans, particularly hyaluronic acid (HA). We took advantage of the recent cloning of the three human hyaluronan synthase (HAS) enzymes (HAS1, HAS2 and HAS3), to explore the molecular mechanisms of this side effect. Northern and slot blots performed on RNA extracted from cultured dermal fibroblasts and the MG-63 osteoblast-like osteosarcoma cell line indicated that HAS2 is the predominant HAS mRNA in these cells. Incubation of both cell types for 24 h in the presence of 10(-6) M dexamethasone (DEX) resulted in a striking 97--98% suppression of HAS2 mRNA levels. Time-course studies in fibroblasts demonstrated suppression of HAS2 mRNA to 28% of control by 1 h, and to 1.2% of control by 2 h, after addition of DEX. Dose-response studies in fibroblasts indicated that the majority of the suppressive effect required concentrations characteristic of cell-surface GC receptors, a point confirmed by persistent DEX-induced suppression in the presence of RU486, an antagonist of classic cytosolic steroid hormone receptors. Nuclear run-off experiments showed a 70% suppression of HAS2 gene transcription in nuclei from DEX-treated fibroblasts, which is unlikely to fully explain the rapid 50--80-fold reduction in message levels. Experiments with actinomycin D (AMD) demonstrated that the message half-life was 25 min in cells without DEX, whereas the combination of AMD with DEX dramatically increased the half-life of HAS2 mRNA, suggesting that DEX acts by inducing a short-lived destabilizer of the HAS2 message. Direct assessment of HAS2 mRNA stability by wash-out of incorporated uridine label established a half-life of 31 min in cells without DEX, which substantially shortened in the presence of DEX. In conclusion, GCs induce a rapid and sustained, near-total suppression of HAS2 message levels, mediated through substantial decreases in both gene transcription and message stability. These effects may contribute to the loss of HA in GC-treated organs.

Manipulation of Hyaluronan Synthase Expression in Prostate Adenocarcinoma Cells Alters Pericellular Matrix Retention and Adhesion to Bone Marrow Endothelial Cells

“…The most conspicuous HAS inhibitors reported to date include hydrocortisone (24), which suppresses HAS2 expression in mesothelial cells almost entirely, and dexamethasone, another glucocorticoid that rapidly down-regulates HAS2 expression in skin fibroblasts (30). The former study again reported no effect on HAS1 or HAS3 message levels, suggesting that HAS2 may be the most transiently regulated of the three genes.…”

Regulation of hyaluronan synthase-2 expression in human intestinal mesenchymal cells: mechanisms of interleukin-1β-mediated induction

“…HAS3 transcript levels are increased in response to IFN-␥ and decreased with TGF-␤ in cultured keratinocytes, whereas HAS1 and 2 levels were unchanged (49). In human dermal fibroblasts, basal transcript levels of HAS2 were significantly suppressed with the administration of glucocorticoids, therapeutic agents with pronounced side effects such as organ atrophy (62). When measuring HAS transcript levels in human fibroblast-like synoviocytes, TGF-␤ potently activated HAS1 but suppressed HAS3 in a dose-dependent manner (52).…”

“…Over the years, much work has focused on the interaction of hyaluronan with various extracellular matrix molecules such as aggrecan and versican (23). These interactions have proven critical for tissue volume changes and the creation and stability of cell-free spaces (36,41,42,55,59,62). Another area of interest is its interaction with a number of cell surface receptors, most notably CD44 (18).…”

Dietary Protein as a Potent Regulator of the Hyaluronan Synthase Gene in Rat Skin

“…The regulation of HAS genes in vivo by physiological factors such as nutrition has not previously been studied, and the present study is thus theˆrst attempt of this sort. It has been shown that human skinˆbroblasts abundantly expressed HAS1 and HAS2 mRNA, 12,27) whereas epidermal keratinocytes had a high HAS3 mRNA content. 13) Among them, the activity of HAS2 and HAS3 is thought to be important, but we found their mRNA levels to be very low.…”

“…Among the three members (HAS1, HAS2 and HAS3), HAS2 functions as the major producer of hyaluronan in the dermis, 11) whereas HAS3 is the one in the epidermis. 12,13) Studies using keratinocytes and dermalˆbroblasts have shown that these enzymes are regulated by some cytokines and glucocorticoids. However, their regulation in vivo has not yet been studied well.…”