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. 2000 Jul 1;349(Pt 1):91–97. doi: 10.1042/0264-6021:3490091

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

W Zhang 1, C E Watson 1, C Liu 1, K J Williams 1, V P Werth 1
PMCID: PMC1221124  PMID: 10861215

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.

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Selected References

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  1. Agren U. M., Tammi M., Tammi R. Hydrocortisone regulation of hyaluronan metabolism in human skin organ culture. J Cell Physiol. 1995 Aug;164(2):240–248. doi: 10.1002/jcp.1041640204. [DOI] [PubMed] [Google Scholar]
  2. Akeson W. H., Amiel D., Abel M. F., Garfin S. R., Woo S. L. Effects of immobilization on joints. Clin Orthop Relat Res. 1987 Jun;(219):28–37. [PubMed] [Google Scholar]
  3. Autio P., Oikarinen A., Melkko J., Risteli J., Risteli L. Systemic glucocorticoids decrease the synthesis of type I and type III collagen in human skin in vivo, whereas isotretinoin treatment has little effect. Br J Dermatol. 1994 Nov;131(5):660–663. doi: 10.1111/j.1365-2133.1994.tb04978.x. [DOI] [PubMed] [Google Scholar]
  4. Bode-Lesniewska B., Dours-Zimmermann M. T., Odermatt B. F., Briner J., Heitz P. U., Zimmermann D. R. Distribution of the large aggregating proteoglycan versican in adult human tissues. J Histochem Cytochem. 1996 Apr;44(4):303–312. doi: 10.1177/44.4.8601689. [DOI] [PubMed] [Google Scholar]
  5. Brecht M., Mayer U., Schlosser E., Prehm P. Increased hyaluronate synthesis is required for fibroblast detachment and mitosis. Biochem J. 1986 Oct 15;239(2):445–450. doi: 10.1042/bj2390445. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brown T. J., Laurent U. B., Fraser J. R. Turnover of hyaluronan in synovial joints: elimination of labelled hyaluronan from the knee joint of the rabbit. Exp Physiol. 1991 Jan;76(1):125–134. doi: 10.1113/expphysiol.1991.sp003474. [DOI] [PubMed] [Google Scholar]
  7. Burnstein K. L., Jewell C. M., Cidlowski J. A. Human glucocorticoid receptor cDNA contains sequences sufficient for receptor down-regulation. J Biol Chem. 1990 May 5;265(13):7284–7291. [PubMed] [Google Scholar]
  8. Buttgereit F., Wehling M., Burmester G. R. A new hypothesis of modular glucocorticoid actions: steroid treatment of rheumatic diseases revisited. Arthritis Rheum. 1998 May;41(5):761–767. doi: 10.1002/1529-0131(199805)41:5<761::AID-ART2>3.0.CO;2-M. [DOI] [PubMed] [Google Scholar]
  9. Capewell S., Reynolds S., Shuttleworth D., Edwards C., Finlay A. Y. Purpura and dermal thinning associated with high dose inhaled corticosteroids. BMJ. 1990 Jun 16;300(6739):1548–1551. doi: 10.1136/bmj.300.6739.1548. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cato A. C., Wade E. Molecular mechanisms of anti-inflammatory action of glucocorticoids. Bioessays. 1996 May;18(5):371–378. doi: 10.1002/bies.950180507. [DOI] [PubMed] [Google Scholar]
  11. Ellis I. R., Schor S. L. Differential effects of TGF-beta1 on hyaluronan synthesis by fetal and adult skin fibroblasts: implications for cell migration and wound healing. Exp Cell Res. 1996 Nov 1;228(2):326–333. doi: 10.1006/excr.1996.0332. [DOI] [PubMed] [Google Scholar]
  12. Feinberg A. P., Vogelstein B. "A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity". Addendum. Anal Biochem. 1984 Feb;137(1):266–267. doi: 10.1016/0003-2697(84)90381-6. [DOI] [PubMed] [Google Scholar]
  13. Fraser J. R., Laurent T. C., Laurent U. B. Hyaluronan: its nature, distribution, functions and turnover. J Intern Med. 1997 Jul;242(1):27–33. doi: 10.1046/j.1365-2796.1997.00170.x. [DOI] [PubMed] [Google Scholar]
  14. Freund R. M., Siebert J. W., Cabrera R. C., Longaker M. T., Eidelman Y., Adzick N. S., Garg H. G. Serial quantitation of hyaluronan and sulfated glycosaminoglycans in fetal sheep skin. Biochem Mol Biol Int. 1993 Mar;29(4):773–783. [PubMed] [Google Scholar]
  15. Fülöp C., Salustri A., Hascall V. C. Coding sequence of a hyaluronan synthase homologue expressed during expansion of the mouse cumulus-oocyte complex. Arch Biochem Biophys. 1997 Jan 15;337(2):261–266. doi: 10.1006/abbi.1996.9793. [DOI] [PubMed] [Google Scholar]
  16. Genever P. G., Dickson I. R. Influence of vitamin D status on hyaluronan localization in bone. Bone. 1996 May;18(5):429–435. doi: 10.1016/8756-3282(96)00046-4. [DOI] [PubMed] [Google Scholar]
  17. Itano N., Kimata K. Molecular cloning of human hyaluronan synthase. Biochem Biophys Res Commun. 1996 May 24;222(3):816–820. doi: 10.1006/bbrc.1996.0827. [DOI] [PubMed] [Google Scholar]
  18. Kato Y., Gospodarowicz D. Stimulation by glucocorticoid of the synthesis of cartilage-matrix proteoglycans produced by rabbit costal chondrocytes in vitro. J Biol Chem. 1985 Feb 25;260(4):2364–2373. [PubMed] [Google Scholar]
  19. Kongtawelert P., Brooks P. M., Ghosh P. Pentosan polysulfate (Cartrophen) prevents the hydrocortisone induced loss of hyaluronic acid and proteoglycans from cartilage of rabbit joints as well as normalizes the keratan sulfate levels in their serum. J Rheumatol. 1989 Nov;16(11):1454–1459. [PubMed] [Google Scholar]
  20. Kosaki R., Watanabe K., Yamaguchi Y. Overproduction of hyaluronan by expression of the hyaluronan synthase Has2 enhances anchorage-independent growth and tumorigenicity. Cancer Res. 1999 Mar 1;59(5):1141–1145. [PubMed] [Google Scholar]
  21. Laurent T. C., Fraser J. R. Hyaluronan. FASEB J. 1992 Apr;6(7):2397–2404. [PubMed] [Google Scholar]
  22. Lavker R. M., Kaidbey K., Leyden J. J. Effects of topical ammonium lactate on cutaneous atrophy from a potent topical corticosteroid. J Am Acad Dermatol. 1992 Apr;26(4):535–544. doi: 10.1016/0190-9622(92)70076-r. [DOI] [PubMed] [Google Scholar]
  23. LeBaron R. G., Zimmermann D. R., Ruoslahti E. Hyaluronate binding properties of versican. J Biol Chem. 1992 May 15;267(14):10003–10010. [PubMed] [Google Scholar]
  24. McCrohon J. A., Nakhla S., Jessup W., Stanley K. K., Celermajer D. S. Estrogen and progesterone reduce lipid accumulation in human monocyte-derived macrophages: a sex-specific effect. Circulation. 1999 Dec 7;100(23):2319–2325. doi: 10.1161/01.cir.100.23.2319. [DOI] [PubMed] [Google Scholar]
  25. McDevitt C. A., Marcelino J., Tucker L. Interaction of intact type VI collagen with hyaluronan. FEBS Lett. 1991 Dec 9;294(3):167–170. doi: 10.1016/0014-5793(91)80660-u. [DOI] [PubMed] [Google Scholar]
  26. Meisler N., Keefer K. A., Ehrlich H. P., Yager D. R., Myers-Parrelli J., Cutroneo K. R. Dexamethasone abrogates the fibrogenic effect of transforming growth factor-beta in rat granuloma and granulation tissue fibroblasts. J Invest Dermatol. 1997 Mar;108(3):285–289. doi: 10.1111/1523-1747.ep12286461. [DOI] [PubMed] [Google Scholar]
  27. Melhus H., Wilén B., Kindmark A., Ljunghall S. Cloning of a novel retinoic acid-inducible mRNA from human osteoblast-like cells. Biochem Mol Biol Int. 1997 Dec;43(5):1145–1150. doi: 10.1080/15216549700204971. [DOI] [PubMed] [Google Scholar]
  28. Meyer L. J., Stern R. Age-dependent changes of hyaluronan in human skin. J Invest Dermatol. 1994 Mar;102(3):385–389. doi: 10.1111/1523-1747.ep12371800. [DOI] [PubMed] [Google Scholar]
  29. Mian N. Analysis of cell-growth-phase-related variations in hyaluronate synthase activity of isolated plasma-membrane fractions of cultured human skin fibroblasts. Biochem J. 1986 Jul 15;237(2):333–342. doi: 10.1042/bj2370333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Myers S. L. Suppression of hyaluronic acid synthesis in synovial organ cultures by corticosteroid suspensions. Arthritis Rheum. 1985 Nov;28(11):1275–1282. doi: 10.1002/art.1780281112. [DOI] [PubMed] [Google Scholar]
  31. Reid I. R. Glucocorticoid osteoporosis--mechanisms and management. Eur J Endocrinol. 1997 Sep;137(3):209–217. doi: 10.1530/eje.0.1370209. [DOI] [PubMed] [Google Scholar]
  32. Robey P. G., Fedarko N. S., Hefferan T. E., Bianco P., Vetter U. K., Grzesik W., Friedenstein A., Van der Pluijm G., Mintz K. P., Young M. F. Structure and molecular regulation of bone matrix proteins. J Bone Miner Res. 1993 Dec;8 (Suppl 2):S483–S487. doi: 10.1002/jbmr.5650081310. [DOI] [PubMed] [Google Scholar]
  33. Smith T. J. Glucocorticoid regulation of glycosaminoglycan synthesis in cultured human skin fibroblasts: evidence for a receptor-mediated mechanism involving effects on specific de novo protein synthesis. Metabolism. 1988 Feb;37(2):179–184. doi: 10.1016/s0026-0495(98)90015-4. [DOI] [PubMed] [Google Scholar]
  34. Spicer A. P., McDonald J. A. Characterization and molecular evolution of a vertebrate hyaluronan synthase gene family. J Biol Chem. 1998 Jan 23;273(4):1923–1932. doi: 10.1074/jbc.273.4.1923. [DOI] [PubMed] [Google Scholar]
  35. Spicer A. P., Olson J. S., McDonald J. A. Molecular cloning and characterization of a cDNA encoding the third putative mammalian hyaluronan synthase. J Biol Chem. 1997 Apr 4;272(14):8957–8961. doi: 10.1074/jbc.272.14.8957. [DOI] [PubMed] [Google Scholar]
  36. Spicer A. P., Seldin M. F., Olsen A. S., Brown N., Wells D. E., Doggett N. A., Itano N., Kimata K., Inazawa J., McDonald J. A. Chromosomal localization of the human and mouse hyaluronan synthase genes. Genomics. 1997 May 1;41(3):493–497. doi: 10.1006/geno.1997.4696. [DOI] [PubMed] [Google Scholar]
  37. Sugiyama Y., Shimada A., Sayo T., Sakai S., Inoue S. Putative hyaluronan synthase mRNA are expressed in mouse skin and TGF-beta upregulates their expression in cultured human skin cells. J Invest Dermatol. 1998 Feb;110(2):116–121. doi: 10.1046/j.1523-1747.1998.00093.x. [DOI] [PubMed] [Google Scholar]
  38. Suzuki M., Asplund T., Yamashita H., Heldin C. H., Heldin P. Stimulation of hyaluronan biosynthesis by platelet-derived growth factor-BB and transforming growth factor-beta 1 involves activation of protein kinase C. Biochem J. 1995 May 1;307(Pt 3):817–821. doi: 10.1042/bj3070817. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Tammi R., Paukkonen K., Wang C., Horsmanheimo M., Tammi M. Hyaluronan and CD44 in psoriatic skin. Intense staining for hyaluronan on dermal capillary loops and reduced expression of CD44 and hyaluronan in keratinocyte-leukocyte interfaces. Arch Dermatol Res. 1994;286(1):21–29. doi: 10.1007/BF00375839. [DOI] [PubMed] [Google Scholar]
  40. Tiedemann K., Malmström A., Westergren-Thorsson G. Cytokine regulation of proteoglycan production in fibroblasts: separate and synergistic effects. Matrix Biol. 1997 Mar;15(7):469–478. doi: 10.1016/s0945-053x(97)90020-2. [DOI] [PubMed] [Google Scholar]
  41. Tomkins G. M., Gelehrter T. D., Granner D., Martin D., Jr, Samuels H. H., Thompson E. B. Control of specific gene expression in higher organisms. Expression of mammalian genes may be controlled by repressors acting on the translation of messenger RNA. Science. 1969 Dec 19;166(3912):1474–1480. doi: 10.1126/science.166.3912.1474. [DOI] [PubMed] [Google Scholar]
  42. Tukey R. H., Okino S. T. Quantitation of related gene products by nuclear run-on and northern blot analysis. Methods Enzymol. 1991;206:284–290. doi: 10.1016/0076-6879(91)06098-n. [DOI] [PubMed] [Google Scholar]
  43. Uzuka M., Nakajima K., Ohta S., Mori Y. The mechanism of estrogen-induced increase in hyaluronic acid biosynthesis, with special reference to estrogen receptor in the mouse skin. Biochim Biophys Acta. 1980 Jan 17;627(2):199–206. doi: 10.1016/0304-4165(80)90321-9. [DOI] [PubMed] [Google Scholar]
  44. Watanabe K., Yamaguchi Y. Molecular identification of a putative human hyaluronan synthase. J Biol Chem. 1996 Sep 20;271(38):22945–22948. doi: 10.1074/jbc.271.38.22945. [DOI] [PubMed] [Google Scholar]
  45. Weigel P. H., Hascall V. C., Tammi M. Hyaluronan synthases. J Biol Chem. 1997 May 30;272(22):13997–14000. doi: 10.1074/jbc.272.22.13997. [DOI] [PubMed] [Google Scholar]
  46. Werth V. P., Kligman A. M., Shi X., Pagnoni A. Lack of correlation of skin thickness with bone density in patients receiving chronic glucocorticoid. Arch Dermatol Res. 1998 Jul;290(7):388–393. doi: 10.1007/s004030050322. [DOI] [PubMed] [Google Scholar]
  47. Werth V. P., Williams K. J., Fisher E. A., Bashir M., Rosenbloom J., Shi X. UVB irradiation alters cellular responses to cytokines: role in extracellular matrix gene expression. J Invest Dermatol. 1997 Mar;108(3):290–294. doi: 10.1111/1523-1747.ep12286462. [DOI] [PubMed] [Google Scholar]
  48. Yamada Y., Itano N., Zako M., Yoshida M., Lenas P., Niimi A., Ueda M., Kimata K. The gene structure and promoter sequence of mouse hyaluronan synthase 1. Biochem J. 1998 Mar 15;330(Pt 3):1223–1227. doi: 10.1042/bj3301223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Yoshikawa H., Tashiro H., Kurisu K. Effect of glucocorticoid on glycosaminoglycans synthesis in cultured mouse embryonic palatal mesenchymal cells. J Craniofac Genet Dev Biol. 1986;6(3):235–244. [PubMed] [Google Scholar]
  50. Zhu Y. Y., Schwartz R. J., Crow M. T. Phorbol esters selectively downregulate contractile protein gene expression in terminally differentiated myotubes through transcriptional repression and message destabilization. J Cell Biol. 1991 Nov;115(3):745–754. doi: 10.1083/jcb.115.3.745. [DOI] [PMC free article] [PubMed] [Google Scholar]

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