Inhibition of the glucocorticoid-activating enzyme 11ß-hydroxysteroid dehydrogenase type 1 drives concurrent 11-oxygenated androgen excess.

Aldo-keto reductase 1C3 (AKR1C3) is a key enzyme in the activation of both classic and 11-oxygenated androgens. In adipose tissue, AKR1C3 is co-expressed with 11ß-hydroxysteroid dehydrogenase type 1 (HSD11B1), which catalyzes not only the local activation of glucocorticoids but also the inactivation of 11-oxygenated androgens, and thus has the potential to counteract AKR1C3. Using a combination of in vitro assays and in silico modeling we show that HSD11B1 attenuates the biosynthesis of the potent 11-oxygenated androgen, 11-ketotestosterone (11KT), by AKR1C3. Employing ex vivo incubations of human female adipose tissue samples we show that inhibition of HSD11B1 results in the increased peripheral biosynthesis of 11KT. Moreover, circulating 11KT increased 2-3 fold in individuals with type 2 diabetes after receiving the selective oral HSD11B1 inhibitor AZD4017 for 35 days, thus confirming that HSD11B1 inhibition results in systemic increases in 11KT concentrations. Our findings show that HSD11B1 protects against excess 11KT production by adipose tissue, a finding of particular significance when considering the evidence for adverse metabolic effects of androgens in women. Therefore, when targeting glucocorticoid activation by HSD11B1 inhibitor treatment in women, the consequently increased generation of 11KT may offset beneficial effects of decreased glucocorticoid activation.

Automated quantification of 3D wound morphology by machine learning and optical coherence tomography in type 2 diabetes.

Background: Driven by increased prevalence of type 2 diabetes and ageing populations, wounds affect millions of people each year, but monitoring and treatment remain limited. Glucocorticoid (stress hormones) activation by the enzyme 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) also impairs healing. We recently reported that 11ß-HSD1 inhibition with oral AZD4017 improves acute wound healing by manual 2D optical coherence tomography (OCT), although this method is subjective and labour-intensive. Objectives: Here, we aimed to develop an automated method of 3D OCT for rapid identification and quantification of multiple wound morphologies. Methods: We analysed 204 3D OCT scans of 3 mm punch biopsies representing 24 480 2D wound image frames. A u-net method was used for image segmentation into 4 key wound morphologies: early granulation tissue, late granulation tissue, neo-epidermis, and blood clot. U-net training was conducted with 0.2% of available frames, with a mini-batch accuracy of 86%. The trained model was applied to compare segment area (per frame) and volume (per scan) at days 2 and 7 post-wounding and in AZD4017 compared to placebo. Results: Automated OCT distinguished wound tissue morphologies, quantifying their volumetric transition during healing, and correlating with corresponding manual measurements. Further, AZD4017 improved epidermal re-epithelialisation (by manual OCT) with a corresponding trend towards increased neo-epidermis volume (by automated OCT). Conclusion: Machine learning and OCT can quantify wound healing for automated, non-invasive monitoring in real-time. This sensitive and reproducible new approach offers a step-change in wound healing research, paving the way for further development in chronic wounds.

The Effect of Endogenous Cushing Syndrome on All-cause and Cause-specific Mortality.

OBJECTIVE: We aimed to perform a systematic review and meta-analysis of all-cause and cause-specific mortality of patients with benign endogenous Cushing syndrome (CS). METHODS: The protocol was registered in PROSPERO (CRD42017067530). PubMed, EMBASE, CINHAL, Web of Science, and Cochrane Central searches were undertaken from inception to January 2021. Outcomes were the standardized mortality ratio (SMR), proportion, and cause of deaths. The I2 test, subgroup analysis, and meta-regression were used to assess heterogeneity across studies. RESULTS: SMR was reported in 14 articles including 3691 patients (13 Cushing disease [CD] and 7 adrenal CS [ACS] cohorts). Overall SMR was 3.0 (95% CI, 2.3-3.9; I2 = 80.5%) for all CS, 2.8 (95% CI, 2.1-3.7; I2 = 81.2%) for CD and 3.3 (95% CI, 0.5-6.6; I2 = 77.9%) for ACS. Proportion of deaths, reported in 87 articles including 19 181 CS patients (53 CD, 24 ACS, and 20 combined CS cohorts), was 0.05 (95% CI, 0.03-0.06) for all CS subtypes with meta-regression analysis revealing no differences between CS subtypes (P = .052). The proportion of deaths was 0.1 (10%) in articles published before 2000 and 0.03 (3%) in 2000 until the last search for CS (P < .001), CD (P < .001), and ACS (P = .01). The causes of death were atherosclerotic diseases and thromboembolism (43.4%), infection (12.7%), malignancy (10.6%), active disease (3.5%), adrenal insufficiency (3.0%), and suicide (2.2%). Despite improved outcomes in recent years, increased mortality from CS persists. The causes of death highlight the need to prevent and manage comorbidities in addition to treating hypercortisolism.

Non-invasive Approaches for the Diagnosis of Autoimmune/Autoinflammatory Skin Diseases-A Focus on Psoriasis and Lupus erythematosus.

The traditional diagnostic gold standard for inflammatory skin lesions of unclear etiology is dermato-histopathology. As this approach requires an invasive skin biopsy, biopsy processing and analysis by specialized histologists, it is a resource intensive approach requiring trained healthcare professionals. In many health care settings access to this diagnostic approach can be difficult and outside emergency cases will usually take several weeks. This scenario leads to delayed or inappropriate treatment given to patients. With dramatically increased sensitivity of a range of analysis systems including mass spectrometry, high sensitivity, multiplex ELISA based systems and PCR approaches we are now able to "measure" samples with unprecedented sensitivity and accuracy. Other important developments include the long-term monitoring of parameters using microneedle approaches and the improvement in imaging systems such as optical coherence tomography. In this review we will focus on recent achievements regarding measurements from non-invasive sampling, in particular from plucked hair and skin tape-strips which seem well suited for the diagnosis of lupus erythematosus and psoriatic inflammation, respectively. While these approaches will not replace clinical observation-they can contribute to improved subgroup diagnosis, stratified therapeutic approaches and have great potential for providing molecular and mechanistic insight in to inflammatory skin diseases.

Topical 11ß-Hydroxysteroid Dehydrogenase Type 1 Inhibition Corrects Cutaneous Features of Systemic Glucocorticoid Excess in Female Mice.

Glucocorticoid (GC) excess drives multiple cutaneous adverse effects, including skin thinning and poor wound healing. The ubiquitously expressed enzyme 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) activates mouse corticosterone from 11-dehydrocorticosterone (and human cortisol from cortisone). We previously demonstrated elevated 11ß-HSD1 activity during mouse wound healing, but the interplay between cutaneous 11ß-HSD1 and systemic GC excess is unexplored. Here, we examined effects of 11ß-HSD1 inhibition by carbenoxolone (CBX) in mice treated with corticosterone (CORT) or vehicle for 6 weeks. Mice were treated bidaily with topical CBX or vehicle (VEH) 7 days before wounding and during wound healing. CORT mice displayed skin thinning and impaired wound healing but also increased epidermal integrity. 11ß-HSD1 activity was elevated in unwounded CORT skin and was inhibited by CBX. CORT mice treated with CBX displayed 51%, 59%, and 100% normalization of wound healing, epidermal thickness, and epidermal integrity, respectively. Gene expression studies revealed normalization of interleukin 6, keratinocyte growth factor, collagen 1, collagen 3, matrix metalloproteinase 9, and tissue inhibitor of matrix metalloproteinase 4 by CBX during wound healing. Importantly, proinflammatory cytokine expression and resolution of inflammation were unaffected by 11ß-HSD1 inhibition. CBX did not regulate skin function or wound healing in the absence of CORT. Our findings demonstrate that 11ß-HSD1 inhibition can limit the cutaneous effects of GC excess, which may improve the safety profile of systemic steroids and the prognosis of chronic wounds.

The prediction and monitoring of toxicity associated with long-term systemic glucocorticoid therapy.

Glucocorticoids are often required for adequate control of inflammation in many serious inflammatory diseases; common indications for long-term treatment include polymyalgia rheumatica, giant cell arteritis, asthma and chronic obstructive pulmonary disease. Long-term glucocorticoid therapy is, however, associated with many adverse effects involving skin, gastro-intestinal, eye, skeletal muscle, bone, adrenal, cardio-metabolic and neuropsychiatric systems. This balance between benefits and risks of glucocorticoids is important for clinical practice and glucocorticoid-related adverse effects can significantly impair health-related quality of life. Understanding the nature and mechanisms of glucocorticoid-related adverse effects may inform how patients are monitored for toxicity and identify those groups, such as older people, that may need closer monitoring. For clinical trials in diseases commonly treated with glucocorticoids, standardised measurement of glucocorticoid-related adverse effects would facilitate future evidence synthesis and meta-analysis.

UVB induces epidermal 11ß-hydroxysteroid dehydrogenase type 1 activity in vivo.

Detrimental consequences of ultraviolet radiation (UVR) in skin include photoageing, immunosuppression and photocarcinogenesis, processes also significantly regulated by local glucocorticoid (GC) availability. In man, the enzyme 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) generates the active GC cortisol from cortisone (or corticosterone from 11-dehydrocorticosterone in rodents). 11ß-HSD1 oxo-reductase activity requires the cofactor NADPH, generated by hexose-6-phosphate dehydrogenase. We previously demonstrated increased 11ß-HSD1 levels in skin obtained from photoexposed versus photoprotected anatomical regions. However, the direct effect of UVR on 11ß-HSD1 expression remains to be elucidated. To investigate the cutaneous regulation of 11ß-HSD1 following UVR in vivo, the dorsal skin of female SKH1 mice was irradiated with 50, 100, 200 and 400 mJ/cm(2) UVB. Measurement of transepidermal water loss, 11ß-HSD1 activity, mRNA/protein expression and histological studies was taken at 1, 3 and 7 days postexposure. 11ß-HSD1 and hexose-6-phosphate dehydrogenase mRNA expression peaked 1 day postexposure to 400 mJ/cm(2) UVB before subsequently declining (days 3 and 7). Corresponding increases in 11ß-HSD1 protein and enzyme activity were observed 3 days postexposure coinciding with reduced GC receptor mRNA expression. Immunofluorescence studies revealed 11ß-HSD1 localization to hyperproliferative epidermal keratinocytes in UVB-exposed skin. 11ß-HSD1 expression and activity were also induced by 200 and 100 (but not 50) mJ/cm(2) UVB and correlated with increased transepidermal water loss (indicative of barrier disruption). UVB-induced 11ß-HSD1 activation represents a novel mechanism that may contribute to the regulation of cutaneous responses to UVR exposure.

Increased glucocorticoid activation during mouse skin wound healing.

Glucocorticoid (GC) excess inhibits wound healing causing increased patient discomfort and infection risk. 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) activates GCs (converting 11-dehydrocorticosterone to corticosterone in rodents) in many tissues including skin, where de novo steroidogenesis from cholesterol has also been reported. To examine the regulation of 11ß-HSD1 and steroidogenic enzyme expression during wound healing, 5 mm wounds were generated in female SKH1 mice and compared at days 0, 2, 4, 8, 14, and 21 relative to unwounded skin. 11ß-HSD1 expression (mRNA and protein) and enzyme activity were elevated at 2 and 4 days post-wounding, with 11ß-HSD1 localizing to infiltrating inflammatory cells. 11ß-HSD2 (GC-deactivating) mRNA expression and activity were undetectable. Although several steroidogenic enzymes displayed variable expression during healing, expression of the final enzyme required for the conversion of 11-deoxycorticosterone to corticosterone, 11ß-hydroxylase (CYP11B1), was lacking in unwounded skin and post-wounding. Consequently, 11-deoxycorticosterone was the principal progesterone metabolite in mouse skin before and after wounding. Our findings demonstrate that 11ß-HSD1 activates considerably more corticosterone than is generated de novo from progesterone in mouse skin and drives GC exposure during healing, demonstrating the basis for 11ß-HSD1 inhibitors to accelerate wound repair.

11ß-Hydroxysteroid dehydrogenase blockade prevents age-induced skin structure and function defects.

Glucocorticoid (GC) excess adversely affects skin integrity, inducing thinning and impaired wound healing. Aged skin, particularly that which has been photo-exposed, shares a similar phenotype. Previously, we demonstrated age-induced expression of the GC-activating enzyme 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) in cultured human dermal fibroblasts (HDFs). Here, we determined 11ß-HSD1 levels in human skin biopsies from young and older volunteers and examined the aged 11ß-HSD1 KO mouse skin phenotype. 11ß-HSD1 activity was elevated in aged human and mouse skin and in PE compared with donor-matched photo-protected human biopsies. Age-induced dermal atrophy with deranged collagen structural organization was prevented in 11ß-HSD1 KO mice, which also exhibited increased collagen density. We found that treatment of HDFs with physiological concentrations of cortisol inhibited rate-limiting steps in collagen biosynthesis and processing. Furthermore, topical 11ß-HSD1 inhibitor treatment accelerated healing of full-thickness mouse dorsal wounds, with improved healing also observed in aged 11ß-HSD1 KO mice. These findings suggest that elevated 11ß-HSD1 activity in aging skin leads to increased local GC generation, which may account for adverse changes occurring in the elderly, and 11ß-HSD1 inhibitors may be useful in the treatment of age-associated impairments in dermal integrity and wound healing.

Basin-scale distribution patterns of photosynthetic picoeukaryotes along an Atlantic Meridional Transect.

Photosynthetic picoeukaryotes (PPEs) of a size < 3 µm play a crucial role in oceanic primary production. However, little is known of the structure of the PPE community over large spatial scales. Here, we investigated the distribution of various PPE classes along an Atlantic Meridional Transect sampled in boreal autumn 2004 that encompasses a range of ocean provinces (gyres, upwelling, temperate regions), using dot blot hybridization technology targeting plastid 16S rRNA gene amplicons. Two algal classes, Prymnesiophyceae and Chrysophyceae, dominated the PPE community throughout the Atlantic Ocean, over a range of water masses presenting different trophic profiles. However, these classes showed strongly complementary distributions with Chrysophyceae dominating northern temperate waters, the southern gyre and equatorial regions, while prymnesiophytes dominated the northern gyre. Phylogenetic analyses using both plastid and nuclear rRNA genes revealed a high diversity among members of both classes, including sequences contained in lineages with no close cultured counterpart. Other PPE classes were less prevalent along the transect, with members of the Cryptophyceae, Pelagophyceae and Eustigmatophyceae essentially restricted to specific regions. Multivariate statistical analyses revealed strong relationships between the distribution patterns of some of these latter PPE classes and temperature, light intensity and nutrient concentrations. Cryptophyceae, for example, were mostly found in the upwelling region and associated with higher nutrient concentrations. However, the key classes of Prymnesiophyceae and Chrysophyceae were not strongly influenced by the variables measured. Although there appeared to be a positive relationship between Chrysophyceae distribution and light intensity, the complementary distributions of these classes could not be explained by the variables recorded and this requires further explanation.

Localization, age- and site-dependent expression, and regulation of 11ß-hydroxysteroid dehydrogenase type 1 in skin.

Glucocorticoids (GCs) are highly detrimental to skin integrity and function both when applied topically for anti-inflammatory treatments and during conditions of circulating excess, e.g., Cushing's syndrome. Within target tissues, GC availability is regulated at a prereceptor level, independently of systemic levels, by isozymes of 11ß-hydroxysteroid dehydrogenase (11ß-HSD) that interconvert active cortisol and inactive cortisone. Many of the adverse effects of GCs on skin are also reminiscent of the natural aging process. 11ß-HSD1 (which activates cortisol), but not 11ß-HSD2 (which inactivates cortisol), was expressed in epidermal keratinocytes and dermal fibroblasts in human skin and also in outer hair follicle root sheath cells in murine skin. 11ß-HSD1 activity was present ex vivo in both species and increased with age in human skin tissue explants. In primary human dermal fibroblasts (HDF) from both photoprotected and photoexposed sites, 11ß-HSD1 also increased with donor age. Additionally, photoexposed HDF displayed higher 11ß-HSD1 mRNA expression than donor-matched photoprotected HDF. GC treatment of HDF caused upregulation of 11ß-HSD1 mRNA levels independent of donor age or site. The age- and site-associated increase in dermal 11ß-HSD1, and the ensuing increased local GC activation, may contribute to the adverse changes in skin morphology and function associated with chronological aging and photoaging.

Steroid biomarkers and genetic studies reveal inactivating mutations in hexose-6-phosphate dehydrogenase in patients with cortisone reductase deficiency.

CONTEXT: Cortisone reductase deficiency (CRD) is characterized by a failure to regenerate cortisol from cortisone via 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1), resulting in increased cortisol clearance, activation of the hypothalamic-pituitary-axis (HPA) and ACTH-mediated adrenal androgen excess. 11beta-HSD1 oxoreductase activity requires the reduced nicotinamide adenine dinucleotide phosphate-generating enzyme hexose-6-phosphate dehydrogenase (H6PDH) within the endoplasmic reticulum. CRD manifests with hyperandrogenism resulting in hirsutism, oligo-amenorrhea, and infertility in females and premature pseudopuberty in males. Recent association studies have failed to corroborate findings that polymorphisms in the genes encoding H6PDH (R453Q) and 11beta-HSD1 (Intron 3 inserted adenine) interact to cause CRD. OBJECTIVE: Our objective was to reevaluate the genetics and steroid biochemistry of patients with CRD. DESIGN: We analyzed 24-h urine collection for steroid biomarkers by gas chromatography/mass spectrometry and sequenced the HSD11B1 and H6PD genes in our CRD cohort. PATIENTS: Patients included four cases presenting with hyperandrogenism and biochemical features clearly indicative of CRD. RESULTS: Gas chromatography/mass spectrometry identified steroid biomarkers that correlated with CRD in each case. Three cases were identified as homozygous (R109AfsX3, Y316X, and G359D) and one case identified as compound heterozygous (c.960G-->A and D620fsX3) for mutations in H6PD. No mutations affecting enzyme activity were identified in the HSD11B1 gene. Expression and activity assays demonstrate loss of function for all reported H6PDH mutations. CONCLUSIONS: CRD is caused by inactivating mutations in the H6PD gene, rendering the 11beta-HSD1 enzyme unable to operate as an oxoreductase, preventing local glucocorticoid regeneration. These data highlight the importance of the redox control of cortisol metabolism and the 11beta-HSD1-H6PDH pathway in regulating hypothalamic-pituitary-adrenal axis activity.

SW-ARRAY: a dynamic programming solution for the identification of copy-number changes in genomic DNA using array comparative genome hybridization data.

Comparative genome hybridization (CGH) to DNA microarrays (array CGH) is a technique capable of detecting deletions and duplications in genomes at high resolution. However, array CGH studies of the human genome noting false negative and false positive results using large insert clones as probes have raised important concerns regarding the suitability of this approach for clinical diagnostic applications. Here, we adapt the Smith-Waterman dynamic-programming algorithm to provide a sensitive and robust analytic approach (SW-ARRAY) for detecting copy-number changes in array CGH data. In a blind series of hybridizations to arrays consisting of the entire tiling path for the terminal 2 Mb of human chromosome 16p, the method identified all monosomies between 267 and 1567 kb with a high degree of statistical significance and accurately located the boundaries of deletions in the range 267-1052 kb. The approach is unique in offering both a nonparametric segmentation procedure and a nonparametric test of significance. It is scalable and well-suited to high resolution whole genome array CGH studies that use array probes derived from large insert clones as well as PCR products and oligonucleotides.