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REDD1 (regulated in development and DNA damage response 1) dissociates therapeutic and adverse effects of topical steroids in skin

ABSTRACT: Even though cutaneous atrophy is the major adverse effect of topical glucocorticoids, its molecular mechanisms are poorly understood. We found that glucocorticoids strongly increased the expression of REDD1 (regulated in development and DNA damage response 1), a stress-inducible inhibitor of mTOR, in mouse and human epidermis. We found that REDD1 knockout animals are partially resistant to glucocorticoid-induced epidermal and subcutaneous adipose atrophy which correlated with the protection of CD34+ follicular epithelial stem cells as well as p63+ keratinocyte progenitors in REDD1 knockout epidermis during chronic steroid treatment. At the same time, REDD1 knockout did not affect anti- inflammatory effect of glucocorticoids, as evaluated by ear edema test. Expression profiling revealed that ~ 50% of the glucocorticoid receptor (GR) target genes were not activated in epidermis of REDD1 knockouts, however, the negative effect of glucocorticoids on gene expression was similar to that in w.t. animals. Overall, our findings reveal a novel pathway in GR activation by its target gene/protein REDD1; and indicate an important role of REDD1 in glucocorticoid-induced skin atrophy, and maintenance of the epidermis and subcutaneous adipose. In addition, our findings form the foundation for the development of safer topical glucocorticoid treatment regimens by combining this therapy with REDD1 inhibitors.

ORGANISM(S): Mus musculus

PROVIDER: GSE59151 | GEO | 2014/08/01

SECONDARY ACCESSION(S): PRJNA254515

REPOSITORIES: GEO

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Project description:Even though cutaneous atrophy is the major adverse effect of topical glucocorticoids, its molecular mechanisms are poorly understood. We found that glucocorticoids strongly increased the expression of REDD1 (regulated in development and DNA damage response 1), a stress-inducible inhibitor of mTOR, in mouse and human epidermis. We found that REDD1 knockout animals are partially resistant to glucocorticoid-induced epidermal and subcutaneous adipose atrophy which correlated with the protection of CD34+ follicular epithelial stem cells as well as p63+ keratinocyte progenitors in REDD1 knockout epidermis during chronic steroid treatment. At the same time, REDD1 knockout did not affect anti- inflammatory effect of glucocorticoids, as evaluated by ear edema test. Expression profiling revealed that ~ 50% of the glucocorticoid receptor (GR) target genes were not activated in epidermis of REDD1 knockouts, however, the negative effect of glucocorticoids on gene expression was similar to that in w.t. animals. Overall, our findings reveal a novel pathway in GR activation by its target gene/protein REDD1; and indicate an important role of REDD1 in glucocorticoid-induced skin atrophy, and maintenance of the epidermis and subcutaneous adipose. In addition, our findings form the foundation for the development of safer topical glucocorticoid treatment regimens by combining this therapy with REDD1 inhibitors. Seven wk old B6D2 females in the telogen stage of the hair cycle were shaved, and treated 3 days later. Glucocorticoid FA was applied topically (2 ug/animal) in 200 ul acetone to the back skin once or up to four applications every third day as described previously (Chebotaev et al., 2007b). Control animals were treated with acetone only. Skin was harvested 24 h after last FA application or 4-8 h after single FA application as indicated in Figure legends. To evaluate the anti-inflammatory effect of glucocorticoid FA in REDD1 KO and w.t. B6x129 mice, we used ear edema test (Schoepe et al., 2010; Schäcke et al., 2004; Park et al., 2006). Seven wk old female animals were pretreated with FA or vehicle applied to the back of the ear lobe one h before application of nonspecific contact irritant croton oil . Mice were sacrificed and ears were harvested nine hours after CO application, the time point at which we observed maximum ear edema in F1 C57Bl x 129 animals (data not shown). Four mm ear punch biopsies were immediately weighted to assess ear edema. Animals were injected i.p. with bromodeoxyuridine 1 hr before skin was harvested. Total RNA from murine epidermis and whole human skin was isolated with RiboPure kit (Ambion). Total RNA from murine s.c. adipose was isolated with RNeasy Lipid Tissue Kit (Qiagen). The RNA samples were treated with TURBOTM DNase (Ambion). Purity of RNA samples isolated from epidermis and s.c. fat was confirmed by the expression of adipose and keratinocyte cell markers.

Project description:Full title: Expression data from human primary subcutaneous preadipocytes treated with glucocorticoids prior to the initiation of differentiation. Preadipocytes are continuously exposed to glucocorticoids in situ due to both steroid present in the circulatory system as well as adipose tissue specific 11Î²HSD1 activity. While the effects of glucocorticoids during differentiation are well studied, the effect of exposure of preadipocytes to glucocorticoids prior to differentiation is unknown. We therefore treated confluent human primary preadipocytes drived from subcutaneous adipose tissue with the synthetic glucocorticoid dexamethasone for 48 hours prior to the initiation of differentiation and assessed what effect this had on their subsequent potential to differentiate. We found that pretreatment with glucocorticoids had a priming effect and resulted in increased differentiation of these preadipocytes. Furthermore, this treatment was additive to the effects of glucocorticoids during the initial phase of adipogenesis. Microarray analysis performed subsequent to the pretreatment with glucocorticoids (at the time point at which preadipocytes would have been induced to differentiate) identified glucocorticoid-responsive, candidate genes whose altered expression could mediate these effects. keywords: glucocorticoids, glucocorticoid receptor, preadipocytes, adipogenesis, human primary preadipocytes, subcutaneous, adipose tissue Experiment Overall Design: Human subcutaneous primary preadipocytes were purchased from Zen-Bio Inc. Preadipocytes from 5 female donors (average BMI 22.5Â±0.2kg/m2) were pooled prior to the initial seeding in T75 flasks. Cells were maintained at 5% CO2 in DMEM with 1.0g/L glucose, 20% calf serum, 100U/ml penicillin, 100mg/ml streptomycin and 50U/ml nystatin. Cells were expanded once prior to seeding in Nunc-brand 12-well dishes. Upon reaching confluence (24 h post-splitting), preadipocytes were stimulated with vehicle or 1uM dex for 48 hours in growth media containing 3% calf serum. Microarray analysis was performed on duplicate samples.

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REDD1 (regulated in development and DNA damage response 1) dissociates therapeutic and adverse effects of topical steroids in skin

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