A 11β-Hydroxysteroid Dehydrogenase Type 1 (11β-HSD1) Inhibitor, 11b-0048, Effectively Suppresses the Expression of 11β-HSD1 Activated in Cultured Keratinocytes and in Diabetic Murine Skin

Abstract

Background

Elevated active glucocorticoids (GCs) are implicated in skin barrier dysfunction, notably in aging and diabetes. The enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) converts inactive GCs to active forms, potentially exacerbating this dysfunction.

Objective

We aimed to investigate the impact of a novel 11β-HSD1 inhibitor on skin inflammation using both in vitro and in vivo models.

Methods

To elucidate the efficacy of a new 11β-HSD1 inhibitor in mitigating skin inflammation induced by various triggers, including dexamethasone treatment, ultraviolet B irradiation, and high glucose levels, in cultured human keratinocytes and the db/db mice as a type 2 diabetes murine model. In cultured keratinocytes, we assessed the effects of the 11β-HSD1 inhibitor on cortisol levels, 11β-HSD1 expression, and cytokine production under conditions simulating inflammation. In db/db mice, we evaluated the inhibitor’s impact on skin barrier function, hemoglobin A1c (HbA1c) levels, corticosterone levels, 11β-HSD1 expression, and cytokine profiles following a 2-week treatment regimen.

Results

Our results demonstrated that both the novel 11β-HSD1 inhibitor and a known inhibitor reduced cortisol levels, 11β-HSD1 expression, and inflammatory cytokine production in cultured keratinocytes. In db/db mice, treatment with either inhibitor improved skin barrier function, lowered serum HbA1c levels, and decreased corticosterone, 11β-HSD1, and inflammatory cytokine expression.

Conclusion

A new 11β-HSD1 inhibitor, “11b-0048,” showed a significant inhibitory effect on the expression of 11β-HSD1 in keratinocytes activated by various conditions and diabetic skin.

INTRODUCTION

Glucocorticoids (GCs) are essential steroid hormones that regulate immune responses, epidermal differentiation, barrier homeostasis, and adaptation to environmental stress through activation of the glucocorticoid receptor (GR)¹. Owing to their potent anti-inflammatory properties, GCs are widely used in the treatment of inflammatory skin diseases²,³,⁴,⁵,⁶. Under acute and physiological conditions, GC signaling suppresses inflammatory responses primarily via GR-mediated repression of pro-inflammatory transcription factors, including nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) pathways⁷.

The hypothalamic-pituitary-adrenal (HPA) axis plays an important role in psychological or physical stress and inflammatory conditions. Although systemic GC levels are controlled by the HPA axis, the skin possesses an autonomous regulatory system that enables tissue-specific modulation of GC bioavailability¹,⁸,⁹,¹⁰,¹¹. A key component of this peripheral GC system is 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), which converts inactive glucocorticoids into their active forms within peripheral tissues⁸,⁹. In the skin, 11β-HSD1 is expressed in epidermal keratinocytes and dermal fibroblasts, allowing local amplification of GC signaling independently of circulating hormone levels¹²,¹³,¹⁴,¹⁵,¹⁶.

Transient activation of local GC metabolism is thought to serve a compensatory role during acute stress or inflammation¹⁷. However, increasing evidence suggests that persistent upregulation of 11β-HSD1 under chronic stress conditions may become maladaptive¹⁸,¹⁹. Sustained elevation of local GC levels has been associated with impaired keratinocyte differentiation, suppression of epidermal barrier components, delayed barrier recovery, and increased vulnerability to inflammatory stimuli²⁰. Importantly, these pathological features are not attributed to a direct pro-inflammatory action of GCs, but rather to dysregulated tissue-specific GC signaling¹⁷,¹⁸.

Chronic GC excess has been reported to disrupt epidermal barrier integrity by reducing structural proteins and lipids essential for stratum corneum (SC) function²¹. Barrier impairment subsequently facilitates secondary inflammatory activation through innate immune and stress-responsive pathways. Thus, inflammation observed under chronic GC exposure reflects an indirect consequence of barrier dysfunction and cellular stress, rather than canonical acute GC biology.

Aging skin and diabetic skin share common pathological features, including barrier fragility, delayed recovery following injury, and heightened susceptibility to inflammation²²,²³. Hyperglycemia has been shown to induce cellular stress responses and to upregulate 11β-HSD1 expression in peripheral tissues, including the skin. Sustained hyperglycemia-associated activation of 11β-HSD1 may therefore contribute to chronic local GC dysregulation and secondary inflammatory responses in diabetic skin²⁴,²⁵,²⁶.

Although dexamethasone is conventionally used as an anti-inflammatory agent under acute exposure conditions, prolonged GC exposure induces epidermal stress, barrier dysfunction, and dysregulated inflammatory signaling²⁰. Accordingly, dexamethasone-treated keratinocytes can be used as a model of chronic GC-induced stress, rather than as a classical acute inflammatory model.

We believe that the increase of 11β-HSD1 expression is an important cause of barrier abnormalities in diabetic skin or aged skin (Supplementary Fig. 1)²⁶. Therefore, treatment of 11β-HSD1 inhibitors will be necessary to restore the skin barrier abnormalities in aged or diabetic skin. We performed this study to elucidate the effect of a new 11β-HSD1 inhibitor, 11b-0048 (Fig. 1)²⁶, on the cultured keratinocytes and the type 2 diabetic murine skin.

Fig. 1. Structure of 11b-0048.

We performed this study to elucidate the effect of a new 11β-HSD1 inhibitor, 11b-0048. The patent for the 11b-0048, is in COSMAX^©.

11β-HSD: 11β-hydroxysteroid dehydrogenase type 1.

MATERIALS AND METHODS

11β-HSD1 inhibitors

We used “11b-0048,” 1-[1-(1-adamantylacetyl)pyrrolidin-3-yl]-3,5-dimethyl-1H-pyrazole (ChemBridge Corp., San Diego, CA, USA), a new material designed and screened by COSMAX^© (COSMAX BTI, Seongnam, Korea) entrusted to the Bioinformatics and Molecular Design Research Center. A proven 11β-HSD1 inhibitor, “385581” (Merck, Readington Township, NJ, USA) was used as a positive control. We compared the effects of both 11β-HSD1 inhibitors on human keratinocytes and the type 2 diabetic murine model.

Cell culture

Primary human epidermal keratinocytes (HEKs) were obtained from healthy human neonatal foreskin (American Type Culture Collection [ATCC], Manassas, VA, USA). HEKs (50,000 cells/mL) were cultured in EpiLife™ medium (Thermo Fisher Scientific, Waltham, MA, USA) supplemented with antibiotic-antimycotic and human keratinocyte growth supplement (Thermo Fisher Scientific) at 37°C under a 5% CO₂. The cells were seeded on type-1 collagen-coated plates, then cultured for 3 days before use in experiments.

Toxicity study

To evaluate the cellular toxicity of 11b-0048, we dissolved it in dimethyl sulfoxide (DMSO) and conducted a viability assessment using the 3-(4,5-dimethylthiazol-2-yl)-2,5-dimethyltetrazolium bromide assay. 11b-0048 was dissolved in DMSO at concentrations of 0.25, 0.5, 1, 5, and 10 μM, respectively. After treating HEKs, we compared their viability 72 hours later. The viability results were as follows; 98.2% for DMSO alone, 97.9% for 0.25 μM, 96.3% for 0.5 μM, 90.6% for 1 μM, 61.1% for 5 μM, and 26.7% for 10 μM (Supplementary Fig. 2). Based on these findings, we chose to use 1 μM of 11b-0048 for our experiments.

Activation of 11β-HSD1

After 3 days of cell seeding, we compared both 11β-HSD1 inhibitors in the HEKs inducing the activation of 11β-HSD1 with dexamethasone treatment, ultraviolet B (UVB) irradiation, and high glucose treatment. The selective 11β-HSD1 inhibitors, “385581” and “11b-0048,” were dissolved in DMSO for topical application at a concentration of 1 μM; HEKs were treated with dexamethasone 1 μM, or irradiated with UVB of 15 mJ/cm², and both inhibitors were treated. The hyperglycemic condition was set for D-glucose at a concentration of 26 mmol/L, and both inhibitors were treated. We measured supernatant for mRNAs and proteins of cortisol, 11β-HSD1, interleukin (IL)-1α, IL-1β, and IL-6 every 24 hours.

Preparation of animals

The Institutional Animal Care and Use Committee at Yonsei University Wonju College of Medicine approved all animal procedures (YWC-210409-1) and a waiver of informed consent was granted owing to the deidentified data used. As a type 2 diabetic model, 4-week-old db/db mice (Daehan Biolink, Eumseong, Korea) were introduced, and a high-fat diet was performed until 14 weeks of age to form a hyperglycemic model. For db/db mice, 9 mice for each group and 7 mice for wild-type were used. The selective 11β-HSD1 inhibitors, “385581” and “11b-0048” were dissolved in DMSO for topical application at a concentration of 10 μM. Each 100 μl of both 11β-HSD1 inhibitors solutions was applied on the back of 14-week-old mice twice a day for 14 days. The vehicle solution was applied to groups 1 and 4, “385581,” as a proven inhibitor, was applied to groups 2 and 5, and “11b-0048,” as a new inhibitor, is applied to groups 3 and 6 (Supplementary Fig. 3).

Measurement of skin barrier function

Skin barrier function was assessed as the basal trans-epidermal water loss (TEWL), SC hydration, and SC integrity (defined as changes in TEWL after 4 times of D-Squame^® disc tape stripping) of the dorsal skin of mice²⁵,²⁷,²⁸. The TEWL was quantified with a Tewameter TM 210 (Courage and Khazaka Electronic GmbH, Cologne, Germany). The calculation of recovery rate was as follows:

$$\frac{(\mathrm{T}\mathrm{E}\mathrm{W}\mathrm{L}\mathrm{a}\mathrm{t}0\mathrm{H}\mathrm{o}\mathrm{u}\mathrm{r}\mathrm{s}\mathrm{a}\mathrm{f}\mathrm{t}\mathrm{e}\mathrm{r}\mathrm{B}\mathrm{a}\mathrm{r}\mathrm{r}\mathrm{i}\mathrm{e}\mathrm{r}\mathrm{D}\mathrm{i}\mathrm{s}\mathrm{r}\mathrm{u}\mathrm{p}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}-\mathrm{T}\mathrm{E}\mathrm{W}\mathrm{L}\mathrm{a}\mathrm{t}4\mathrm{H}\mathrm{o}\mathrm{u}\mathrm{r}\mathrm{s}\mathrm{a}\mathrm{f}\mathrm{t}\mathrm{e}\mathrm{r}\mathrm{B}\mathrm{a}\mathrm{r}\mathrm{r}\mathrm{i}\mathrm{e}\mathrm{r}\mathrm{D}\mathrm{i}\mathrm{s}\mathrm{r}\mathrm{u}\mathrm{p}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n})}{(\mathrm{T}\mathrm{E}\mathrm{W}\mathrm{L}\mathrm{a}\mathrm{t}0\mathrm{H}\mathrm{o}\mathrm{u}\mathrm{r}\mathrm{s}\mathrm{a}\mathrm{f}\mathrm{t}\mathrm{e}\mathrm{r}\mathrm{B}\mathrm{a}\mathrm{r}\mathrm{r}\mathrm{i}\mathrm{e}\mathrm{r}\mathrm{D}\mathrm{i}\mathrm{s}\mathrm{r}\mathrm{u}\mathrm{p}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}-\mathrm{T}\mathrm{E}\mathrm{W}\mathrm{L}\mathrm{a}\mathrm{t}\mathrm{B}\mathrm{a}\mathrm{s}\mathrm{e}\mathrm{l}\mathrm{i}\mathrm{n}\mathrm{e})}\times 100.$$

SC hydration was measured using a Corneometer CM 850 (Courage and Khazaka Electronic GmbH). Skin surface pH was measured with a pH meter (WTW, Weilheim, Germany).

Enzyme-linked immunosorbent assay (ELISA)

The culture supernatant was centrifuged for 15 minutes at 1,000 ×g, 2°C–8°C. The amount of cortisol in the samples was measured using a Human Cortisol ELISA Kit (CSB-E05111h; Cusabio Technology LLC, Houston, TX, USA). According to the manufacturer’s protocol, the expression of 11β-HSD1 was measured using the Human HSD11B1/HSD1B ELISA Kit (LS-F11298; LSBio, Seattle, WA, USA). The amounts of cytokines were measured using a multiplex kit (LXSAHM; LABISKOMA, Seoul, Korea)²⁴.

In vivo, all experimental procedures were performed after 5% isoflurane inhalation anesthesia. Hemoglobin A1c (HbA1c) level was measured by ELISA (MBS2024955; MyBioSource, Inc., San Diego, CA, USA). SC was obtained by D-squame^® tape stripping and the dorsal skin of mice was obtained by biopsy. D-squame disc tapes (CuDerm, Dallas, TX, USA) were attached to mice back skin to collect SC. Four strips were used to extract proteins. Strips were placed in 500 μl of lysis buffer, followed by vortexing and overnight incubation at 4°C. The strip was used to measure the cortisol of whole SC. The amount of SC corticosterone was measured using a Cortisol ELISA Kit (CSB-E05113m; Cusabio Technology LLC). The expression of 11β-HSD1 in the epidermis and dermis was measured using the HSD11B1/HSD1B ELISA Kit (AE39390MO; Abebio, Wuhan, China), according to the manufacturer’s protocol. The amounts of cytokines were measured using a multiplex kit (LXSARM; LABISKOMA)¹⁸.

Quantitative real-time polymerase chain reaction (RT-PCR)

Total RNA was isolated from HEKs using QuantiTect Reverse Transcription Kits (Qiagen, Hilden, Germany). The product was reverse-transcribed into first-strand complementary DNA, then 11β-HSD1 expression was measured using QuantiFast SYBR Green PCR Master Mix (Qiagen) as described by the manufacturer. Messenger RNA was quantified by RT-PCR using a 7900HT FAST Real-Time PCR System (Applied Biosystems, Franklin Lakes, NJ, USA). Primer concentrations were first optimized to avoid non-specific binding of primers. After the PCR, dissociation curves were analyzed to verify the specificity of the amplification products. The mRNA was normalized using glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and the following sequence-specific primers were designed: GAPDH sense, 5′-TTGATTTTGGAGGGATCTCG-3′ and antisense, 5′-GAGTCAACGGATTTGGTCGT-3′; 11β-HSD1, sense: 5′-TCTCCTCTCTGGCTGGGAAAG-3′, antisense: 5′-GAACCCATCAAAGCAAACTTG-3′. The conditions for thermal cycling were as follows: denaturation for 5 minutes, followed by 45 cycles of amplification at 95°C for 15 seconds, annealing at 60°C for 30 seconds, and extension at 76°C for 30 seconds¹⁸,²⁴,²⁵.

Statistical analysis

The data are expressed as mean ± standard deviation. The unpaired Student’s t-tests and repeated-measures analysis of variance were performed to determine the level of significance of differences between the sample mean. Statistical analyses were performed using GraphPad Prism software (GraphPad Software Inc., San Diego, CA, USA). Values with p<0.05 were considered statistically significant.

RESULTS

Both 11β-HSD1 inhibitors decrease cortisol and 11β-HSD1 in normal HEKs

First, the expression of 11β-HSD1 was examined in HEKs that were treated with 1 μM of dexamethasone, 15 mJ UVB irradiation, and 26 mmol/L of D-glucose. Thereafter, cortisol and 11β-HSD1 levels in the cultured medium were measured at the 24-hours interval. After twenty-four hours after treatment, 11β-HSD1 protein and mRNA were significantly increased along with cortisol level in the cultured medium, and a time-dependent increase was observed (Fig. 2). The levels of cortisol and 11β-HSD1 in the cultured medium were higher when 15 mJ UVB irradiation was performed than 1 μM of dexamethasone or 26 mmol/L of D-glucose.

Fig. 2. Levels of cortisol and 11β-HSD1 in the cultured keratinocyte after the treatment with both 11β-HSD1 inhibitors.

In experiments measured at 24-hour intervals, time-dependent increases in the level of cortisol and 11β-HSD1 were observed in dexamethasone, ultraviolet radiation, and high glucose-treated conditions. (A) After additional treatment with a new 11β-HSD1 inhibitor and a proven inhibitor, cortisol levels in the culture medium decreased over time, and the new inhibitor showed a further decrease. (B, C) 11β-HSD1 protein level and mRNA in the culture medium decreased over time in both 11β-HSD1 inhibitor treatments but showed a further decrease in the new inhibitor.

11β-HSD1: 11β-hydroxysteroid dehydrogenase type 1, UVB: ultraviolet B.

^*p<0.05, ^**p<0.01.

After additional treatment with both 11β-HSD1 inhibitors, the level of cortisol in the culture medium decreased over time, and the new 11β-HSD1 inhibitor showed a further decrease (Fig. 2A). Both the protein and mRNA of 11β-HSD1 decreased when treated with 11β-HSD1 inhibitor and showed a further decrease when treated with the new inhibitor, “11b-0048” (Fig. 2B and C). Detailed values are provided in Supplementary Table 1.

Both 11β-HSD1 inhibitors decrease IL-1α, IL-1β, and IL-6 in HEKs

Inflammatory cytokines (IL-1α, IL-1β, and IL-6) were measured in HEKs that were treated with 1 μM of dexamethasone, 15 mJ UVB irradiation, and 26 mmol/L of D-glucose. Thereafter, cytokine levels in the cultured medium were measured at the 24-hour interval. After 24 hours after treatment, cytokine levels were significantly increased in the cultured medium, and a time-dependent increase was observed. The levels of cytokines in the cultured medium were higher when 15 mJ UVB irradiation was performed than 1 μM of dexamethasone or 26 mmol/L of D-glucose.

After additional treatment with both 11β-HSD1 inhibitors, the secretion of cytokines of HEKs decreased over time, and the new 11β-HSD1 inhibitor showed a further decrease (Fig. 3). Detailed values are provided in Supplementary Table 2.

Fig. 3. Cytokine levels in the cultured keratinocyte after the treatment with both 11β-HSD1 inhibitors.

In experiments measured at 24-hour intervals, a time-dependent increases in the levels of cytokines were observed in dexamethasone, ultraviolet radiation, and high glucose-treated conditions. The secretion of cytokines IL-1α, IL-1β, and IL-6 in the culture medium was also decreased in both 11β-HSD1 inhibitor treatments but showed a further decrease with the new inhibitor.

11β-HSD1: 11β-hydroxysteroid dehydrogenase type 1, IL: interleukin, UVB: ultraviolet B.

^*p<0.05, ^**p<0.01, ^***p<0.001.

Both 11β-HSD1 inhibitors decrease HbA1c and SC corticosterone in db/db mice

Following in vitro study, we examined whether these results were the same in mice. We applied vehicle, a proven inhibitor, and a new inhibitor twice a day to db/db mice and wild-type mice for 14 days. On the 14th day after application, blood samples from the mice orbit were collected and HbA1c level was measured. HbA1c was higher in db/db mice than in the wild type. After both inhibitors were applied to the skin, HbA1c was statistically significantly decreased (Fig. 4A).

Fig. 4. Levels of serum HbA1c, SC corticosterone and skin 11β-HSD1 in the diabetic murine model after the treatment with both 11β-HSD1 inhibitors.

When comparing the db/db mice and the wild-type mice, the db/db mice had higher levels of serum HbA1c, SC corticosterone, and 11β-HSD1. Both 11β-HSD1 inhibitors reduced not only HbA1c level but also SC corticosterone and 11β-HSD1 levels in the epidermis and dermis in diabetic mice.

HbA1c: hemoglobin A1c, SC: stratum corneum, 11β-HSD1: 11β-hydroxysteroid dehydrogenase type 1.

^*p<0.05, ^**p<0.01.

SC corticosterone was higher in db/db mice than in the wild type, and the protein and mRNA levels of 11β-HSD1 in the epidermis and dermis of skin samples obtained by skin biopsy were higher in db/db mice. Both inhibitors had significantly lower SC corticosterone than vehicle. However, SC corticosterone level did not differ between both inhibitors in db/db mice (14.01±2.00 vs. 13.32±3.54, p=0.368) and wild type (6.30±1.22 vs. 4.65±2.75, p=0.120) (Fig. 4B). Both 11β-HSD1 inhibitors decreased 11β-HSD1 protein and mRNA expression in the epidermis and dermis in both db/db mice and wild type (Fig. 4C-F). Detailed values are provided in Supplementary Table 3.

Both 11β-HSD1 inhibitors decrease epidermal cytokines in db/db mice

By skin biopsy, db/db mice and wild-type mice epidermis were obtained and inflammatory cytokine levels (IL-1α, IL-1β, tumor necrosis factor [TNF]-α) were measured. Cytokine levels were higher in db/db mice than in the wild type. When both inhibitors were applied, cytokines showed a decreasing trend, but there was no statistically significant difference (Fig. 5A-D). Detailed values are provided in Supplementary Table 4.

Fig. 5. Epidermal cytokine levels and parameters of skin barrier function in the diabetic murine epidermis after the treatment with both 11β-HSD1 inhibitors.

(A-D) When comparing the db/db mice and the wild-type mouse, db/db mouse had higher levels of the cytokine. In diabetic mice, the secretion of cytokines such as IL-1α, IL-1β, IL-6, and TNF-α was partially decreased in both 11β-HSD1 inhibitor treatments. (E-H) When comparing the db/db mice and the wild-type mice, skin barrier functions were poor in the db/db mice. When both 11β-HSD1 inhibitors were applied to the skin in diabetic mice, there was no difference in skin surface pH. SC hydration was decreased, basal TEWL was increased, and the barrier recovery rate accelerated compared to the vehicle.

11β-HSD1: 11β-hydroxysteroid dehydrogenase type 1, IL: interleukin, TNF: tumor necrosis factor, SC: stratum corneum, TEWL: trans-epidermal water loss.

^*p<0.05, ^**p<0.01, ^***p<0.001.

Both 11β-HSD1 inhibitors improved the skin barrier recovery in db/db mice

The skin barrier function was measured after 14 days of application. The db/db mice had higher skin surface pH and SC hydration compared to the wild type. In addition, the db/db mice had lower basal TEWL and barrier recovery rates after D-Squame^® disc tape stripping compared to the wild type.

There was no significant difference in skin surface pH between both inhibitors in db/db mice. Compared with the vehicle (7.37±0.03) in the wild type, the proven inhibitor decreased the pH (7.17±0.06, p=0.016), and the new inhibitor increased the pH (7.44±0.01, p=0.023) (Fig. 5E). SC hydration was decreased in both db/db mice and wild type. SC hydration was decreased in the new inhibitor than in the proven inhibitor (Fig. 5F). Basal TEWL was increased in both db/db mice and wild type, but the new inhibitor increased more than the proven inhibitor (Fig. 5G). The barrier recovery rate was increased in both db/db mice and wild type. In db/db mice, there was a statistically significant difference in recovery rates from proven inhibitors (30.38±19.57 vs. 93.91±12.00, p=0.024). However, in the wild type, the recovery rate increased more with the new inhibitor than with the proven inhibitor, but it was not statistically significant (40.61±9.56 vs. 60.10±10.59 vs. 104.94±35.32) (Fig. 5H). Detailed values are provided in Supplementary Table 4.

DISCUSSION

In the present study, we investigated the role of 11β-HSD1 in chronic stress-associated cutaneous inflammation and evaluated the therapeutic potential of the novel 11β-HSD1 inhibitor 11b-0048. Our findings demonstrate that chronic stress conditions—including prolonged GC exposure, ultraviolet irradiation, and hyperglycemia—are associated with upregulation of 11β-HSD1 and increased inflammatory responses in keratinocytes²⁹ and diabetic skin. Importantly, pharmacologic inhibition of 11β-HSD1 by 11b-0048 significantly attenuated these pathological changes both in vitro and in vivo. Based on prior patent data describing active-site–directed 11β-HSD1 inhibitors, 11b-0048 is presumed to act as a competitive inhibitor; however, definitive confirmation will require dedicated enzyme kinetic and structural analyses. 11b-0048 does not directly target GR, NF-κB, or MAPK signaling pathways; rather, its anti-inflammatory effects are mediated indirectly through inhibition of 11β-HSD1, resulting in reduced local GC activation and subsequent normalization of stress-associated inflammatory responses. This distinction is critical, as it reconciles our findings with canonical glucocorticoid biology, in which acute GC signaling is anti-inflammatory.

These findings support a conceptual framework in which chronic, tissue-specific dysregulation of GC metabolism, rather than GCs themselves, underlies inflammatory and barrier abnormalities in stressed skin²⁴. Sustained local GC excess has been reported to induce GR dysregulation or resistance, impair keratinocyte differentiation, and suppress the expression of barrier-forming proteins and lipids¹³. As epidermal barrier integrity deteriorates, the skin becomes increasingly susceptible to secondary inflammatory activation driven by cellular stress responses and innate immune signaling¹⁸. In this context, inflammation emerges as a downstream consequence of barrier dysfunction rather than a direct GC-driven process.

The role of 11β-HSD1 in inflammation has been debated, as its induction may initially represent a compensatory response aimed at limiting acute inflammation³⁰. However, our findings, together with accumulating evidence from genetic and pharmacological studies¹³, support the interpretation that persistent activation of 11β-HSD1 becomes pathogenic under chronic stress conditions. Inhibition of 11β-HSD1 has been shown to improve epidermal differentiation, enhance barrier recovery, and reduce inflammatory cytokine expression in aged, photo-damaged, and metabolically stressed skin¹³. Our results extend these observations by demonstrating that selective inhibition of 11β-HSD1 effectively attenuates inflammatory dysregulation in diabetic skin, a clinically relevant and therapeutically challenging condition.

The experimental models employed in this study further support the clinical relevance of our findings. While dexamethasone is classically used as an anti-inflammatory agent, prolonged exposure recapitulates pathological conditions characterized by sustained local GC excess, barrier impairment, and inflammatory dysregulation³¹. In this context, dexamethasone treatment serves as a model of chronic GC-induced stress rather than acute inflammation.

Hyperglycemia represents another clinically relevant condition associated with chronic 11β-HSD1 activation²⁶. In diabetic skin, increased 11β-HSD1 expression contributes to excessive local GC regeneration, impaired epidermal homeostasis, and secondary inflammation. Our findings also have important translational implications. Diabetic skin is characterized by impaired barrier function, delayed wound healing, and increased susceptibility to infection and inflammation³². Current therapeutic strategies primarily focus on glycemic control or symptomatic management of inflammation, with limited efficacy in restoring intrinsic skin homeostasis³². Targeting 11β-HSD1 offers a novel approach that addresses the upstream metabolic dysregulation underlying these pathological features²⁶. By attenuating excessive local GC activation, 11β-HSD1 inhibition may restore a more physiological balance between stress adaptation and inflammatory control²⁶.

Several limitations of this study should be acknowledged. We did not directly assess enzyme kinetics or structural binding interactions between 11b-0048 and 11β-HSD1, nor did we directly measure downstream signaling activities of NF-κB or MAPK. Additionally, while our data support a strong association between 11β-HSD1 inhibition and reduced inflammation, further studies will be required to delineate the precise molecular events linking GC dysregulation to barrier impairment and inflammatory activation. Future investigations incorporating genetic models, detailed biochemical analyses, and clinical studies will further refine the therapeutic potential of 11β-HSD1 inhibition.

In conclusion, this study demonstrates that sustained activation of 11β-HSD1 contributes to chronic cutaneous inflammation and barrier dysfunction through dysregulated local GC metabolism. Pharmacologic inhibition of 11β-HSD1 by 11b-0048 effectively mitigates these pathological processes in keratinocytes and diabetic skin. These findings highlight 11β-HSD1 as a promising therapeutic target for chronic inflammatory and metabolic skin disorders and provide a mechanistic rationale for the development of 11β-HSD1-directed therapies.

SUPPLEMENTARY MATERIALS

Supplementary Table 1

Levels of cortisol and 11β-HSD1 in the cultured keratinocyte after the treatment with both 11β-HSD1 inhibitors

Supplementary Table 2

Cytokine levels in the cultured keratinocyte after the treatment with both 11β-HSD1 inhibitors

Supplementary Table 3

Levels of serum HbA1c, SC corticosterone and skin 11β-HSD1 in the diabetic murine model after the treatment with both 11β-HSD1 inhibitors

Supplementary Table 4

Epidermal cytokine levels and parameters of skin barrier function in the diabetic murine epidermis after the treatment with both 11β-HSD1 inhibitors

Supplementary Fig. 1

Skin barrier disruption in diabetic skin.

Supplementary Fig. 2

Cellular toxicity study of 11b-0048.

Supplementary Fig. 3

Diabetic murine model study.