Normal Skin Research Articles

Design and In Vitro Evaluation of Fluorescent MOF-Core CaCO₃-PEI-FA Shell Nanoparticles for Targeted Therapy of Laryngeal Cancer Cells.

Laryngeal cancer, a common malignant respiratory tumor, is primarily treated through surgery. However, challenges such as recurrence, metastasis, and drug resistance persist. In recent years, multifunctional drug delivery systems (DDS) based on nanoparticles have shown great potential in improving drug loading and release. We developed a biocompatible core-shell nanoparticle system with a zinc-based metal-organic framework (MOF) as the core, named CP1. The shell, composed of polyethyleneimine (PEI), folic acid, and calcium carbonate, forms a composite called CaCO3-PEI-FA. This system enhances biocompatibility and increases the efficacy of biomedical applications. Encapsulating CP1 within the CaCO3-PEI-FA shell allows for the targeted delivery of the anticancer drug doxorubicin (DOX) to laryngeal cancer cells (Hep-2), resulting in the CaCO3-PEI-FA@CP1@DOX system. The CaCO3-PEI-FA composite exhibits strong fluorescence with a peak around 350nm, confirming successful synthesis and demonstrating its potential as a bioimaging probe. Importantly, the nanoparticle system without DOX showed low toxicity to normal human skin fibroblasts (HSF). In vitro cytology experiments revealed a 38% inhibition rate of Hep-2 cells after 24h, highlighting the nanocomposite's significant potential in inhibiting laryngeal cancer cell proliferation and inducing apoptosis, underscoring its promise in targeted laryngeal cancer therapy.

Biological effects in normal human fibroblasts following chronic and acute irradiation with both low- and high-LET radiation.

Radiobiological studies at low dose rates allow us to improve our knowledge of the mechanisms by which radiation exerts its effects on biological systems following chronic exposures. Moreover, these studies can complement available epidemiological data on the biological effects of low doses and dose rates of ionizing radiation. Very few studies have simultaneously compared the biological effects of low- and high-LET radiations at the same dose rate for chronic irradiation. We compared, for the first time in the same experiment, the effects of chronic (dose rates as low as ~18 and 5 mGy/h) and acute irradiations on clonogenicity and micronucleus formation in AG1522 normal human skin fibroblasts in the confluent state exposed to doses of low- and high-LET radiation (gamma rays and alpha particles) to investigate any differences due to the different radiation quality and different dose rate (in the dose range 0.006-0.9 Gy for alpha particles and 0.4-2.3 Gy for gamma rays). As expected, alpha particles were more effective than gamma rays at inducing cytogenetic damage and reduced clonogenic cell survival. For gamma rays, the cytogenetic damage and the reduction of clonogenic cell survival were greater when the dose was delivered acutely instead of chronically. Instead, for the alpha particles, at the same dose, we found equal cytogenetic damage and reduction of clonogenic cell survival for both chronic and acute exposure (except for the highest doses of 0.4 and 0.9 Gy, where cytogenetic damage is greater at a low dose rate). The results of this study may have an impact on space and terrestrial radioprotection of humans at low doses and low dose rates, on biodosimetry, and on the use of ionizing radiation in medicine. These results also provide insights into understanding damage induction and cell reaction mechanisms following chronic exposure (at dose rates as low as 18 and 5 mGy/h) to low- and high-LET radiation.

LesionNet: an automated approach for skin lesion classification using SIFT features with customized convolutional neural network.

Accurate detection of skin lesions through computer-aided diagnosis has emerged as a critical advancement in dermatology, addressing the inefficiencies and errors inherent in manual visual analysis. Despite the promise of automated diagnostic approaches, challenges such as image size variability, hair artifacts, color inconsistencies, ruler markers, low contrast, lesion dimension differences, and gel bubbles must be overcome. Researchers have made significant strides in binary classification problems, particularly in distinguishing melanocytic lesions from normal skin conditions. Leveraging the "MNIST HAM10000" dataset from the International Skin Image Collaboration, this study integrates Scale-Invariant Feature Transform (SIFT) features with a custom convolutional neural network model called LesionNet. The experimental results reveal the model's robustness, achieving an impressive accuracy of 99.28%. This high accuracy underscores the effectiveness of combining feature extraction techniques with advanced neural network models in enhancing the precision of skin lesion detection.

In vivo CRISPRi screen identified lncRNA portfolio crucial for cutaneous squamous cell carcinoma tumor growth.

Cutaneous squamous cell carcinoma (cSCC) accounts for 20% of all skin cancer deaths globally, making it the second-highest subtype of skin cancer. The prevalence of cSCC in humans, as well as the poor capacity for an efficient prognosis, highlights the need to uncover alternative actors and mechanisms at the foundation of skin cancer development. Significant advances have been made to better understand some key factors in cSCC progression. However, little is known about the role of noncoding RNAs, particularly of a specific category called long noncoding RNA (lncRNA). By performing pseudobulk analysis of single-cell sequencing data from normal and cSCC human skin tissues, we determined a global portfolio of lncRNAs specifically expressed in keratinocyte subpopulations. Integration of CRISPR interference screens in vitro and the xenograft model identified several lncRNAs impacting the growth of cSCC cancer lines both in vitro and in vivo. Among these, we further validated LINC00704 and LINC01116 as proliferation-regulating lncRNAs in cSCC lines and potential biomarkers of cSCC progression. Taken together, our study provides a comprehensive signature of lncRNAs with roles in regulating cSCC progression.

Streptomyces fradiae Mitigates the Impact of Potato Virus Y by Inducing Systemic Resistance in Two Egyptian Potato (Solanum tuberosum L.) Cultivars

In this study, the impact of culture media filtrate of QD3 actinobacterial isolate on two potato cultivars, Spunta and Diamond, infected with potato virus Y (PVY) was investigated. Various parameters, including infection percentage, PVY virus infectivity, disease severity scoring, PVY optical density, photosynthetic and defense-related biochemical markers, enzymatic profiling, phenolic compounds, proline content, salicylic acid levels, and growth and yield parameters, were assessed to elucidate the potential of the QD3 actinobacterial isolate culture filtrate in mitigating PVY-induced damage. The physiological and biochemical characteristics of the QD3 actinobacterial isolate, including its salinity tolerance, pH preferences, and metabolic traits, were investigated. Molecular identification via 16S rRNA gene sequencing confirmed its classification as Streptomyces fradiae QD3, and it was deposited in GenBank with the gene accession number MN160630. Distinct responses between Spunta and Diamond cultivars, with Spunta displaying greater resistance to PVY infection. Notably, pre-infection foliar application of the QD3 filtrate significantly reduced disease symptoms and virus infection in both cultivars. For post-PVY infection, the QD3 filtrate effectively mitigated disease severity and the PVY optical density. Furthermore, the QD3 filtrate positively influenced photosynthetic pigments, enzymatic antioxidant activities, and key biochemical components associated with plant defense mechanisms. Gas chromatography‒mass spectrometry (GC‒MS) analysis revealed palmitic acid (hexadecanoic acid, methyl ester) and oleic acid (9-octadecanoic acid, methyl ester) as the most prominent compounds, with retention times of 23.23 min and 26.41 min, representing 53.27% and 23.25%, respectively, of the total peak area as primary unsaturated fatty acids and demonstrating antiviral effects against plant viruses. Cytotoxicity assays on normal human skin fibroblasts (HSFs) revealed the safety of QD3 metabolites, with low discernible toxicity at high concentrations, reinforcing their potential as safe and effective interventions. The phytotoxicity results indicate that all the seeds presented high germination rates of approximately 95–98%, suggesting that the treatment conditions had no phytotoxic effect on the Brassica oleracea (broccoli) seeds, Lactuca sativa (lettuce) seeds, and Eruca sativa (arugula or rocket) seeds. Overall, the results of this study suggest that the S. fradiae filtrate has promising anti-PVY properties, influencing various physiological, biochemical, and molecular aspects in potato cultivars. These findings provide valuable insights into potential strategies for managing PVY infections in potato crops, emphasizing the importance of Streptomyces-derived interventions in enhancing plant health and crop protection.

Multimodal Photoacoustic/Elastography Imaging for the Detection of Acute Radiation Dermatitis in Breast Radiation Therapy

Purpose: This study evaluates whether photoacoustic (PA), sound touch elastography (STE), and viscoelasticity (VE) can distinguish between normal and abnormal post-radiotherapy breast skin and compare these methods with RTOG criteria at a 20 Gy threshold. Methods: Patients meeting inclusion and exclusion criteria underwent PA, STE, and VE on the same day. Collected data included radiation dose, molecular type, RTOG, Fitzpatrick skin type, pathology, neoadjuvant chemotherapy status, TNM classification, surgical procedures, primary breast cancer location, BMI, and age. A sample of 41 patients was determined using a two-sample t-test. Statistical tools like T-tests, variance analysis, rank sum tests, and chi-square tests, along with random forest analysis and ROC curves, were used to evaluate the radiation dose effects. Results: Data from 66 patients showed significant differences in parameters such as dermis and subcutaneous tissue oxygen saturation, dermal thickness, and skin elasticity (p-values < 0.05). However, minimum values of oxygen saturation and some photoacoustic measures were not significantly different. Notably, at a 20 Gy radiation threshold, significant variations in oxygen saturation, abnormal dermal thickness, and skin STE and VE were observed, proving more accurate than RTOG grading. Conclusion: The study confirms the effectiveness of PA and elastography imaging in differentiating normal from abnormal breast tissue and assessing radiation-induced changes, highlighting the potential of these imaging techniques.

Antibacterial and wound healing potential of biosynthesized zinc oxide nanoparticles against carbapenem-resistant Acinetobacter baumannii: an in vitro and in vivo study

Carbapenem-resistant Acinetobacter baumannii denotes a significant menace to public health, and it mandates an urgent development of new effective medications. Here, we aimed to estimate the efficiency of the zinc oxide nanoparticles (ZnO NP) biosynthesized from Arthrospira maxima (Spirulina) both in vitro and in vivo. Carbapenem-resistant A. baumannii isolates were collected, identified, tested for their antibiotic susceptibility, and then subjected to PCR to detect carbapenemase-producing genes. The most predominant carbapenemase resistance gene was blaKPC. The biosynthesized ZnO NP were characterized using UV, FTIR, XRD, SEM, and TEM. The prepared ZnO NP was then tested against A. baumannii isolates to determine the minimum inhibitory concentration (MIC), which ranged from 250 to 1000 μg/ml. Burn wound was persuaded in twenty rats and inoculated with carbapenem-resistant A. baumannii isolate. Rats were allocated into four groups: a negative control group, a positive control group treated with topical 0.9% saline, a test treatment group that received topical ZnO NP, and a standard treatment group. All groups received treatment for 15 consecutive days and then euthanized. Skin samples were harvested and then subjected to histopathological and immunochemical investigations. ZnO NP revealed a comparable antibacterial activity to colistin as it revealed a lower level of fibrosis, mature surface epithelization with keratinization, and restoration of the normal skin architecture. In addition, it significantly decreased the immunoreactivity of the studied inflammatory markers. Thus, ZnO NP synthesized by A. maxima could be considered a promising, safe, and biocompatible alternative to traditional antibiotics in the therapy of carbapenem-resistant A. baumannii infections.

Case Report: Alpha6 Integrin Disorder Presenting in Childhood with Nail Dysplasia and Onycholysis But No History of Fragile or Bullous Skin Changes

AbstractWe report a 7-year-old girl born with pyloric atresia but without congenital epidermolysis bullosa or skin fragility. Nail dysplasia developed at age 8 months and throughout childhood she suffered from onycholysis and mild nail hypertrophy. Whole-exome sequencing demonstrated biallelic mutations in alpha6 integrin (ITGA6): p. Q139* and R153W. ITGA6 normally forms a protein heterodimer with beta4 integrin (ITGB4), and this dimer participates in anchoring the basal skin cells to the extracellular matrix. Biallelic mutations in each gene are well known to cause epidermolysis bullosa and pyloric atresia. However, this child had ostensibly normal skin without any evidence of skin fragility. In a literature search, we identified 11 cases involving ITGA6 mutations, and all had epidermolysis skin changes. Thus, this case adds to the reported phenotype of ITGA6 disease since it is the first to show absence of an epidermolysis bullosa phenotype in the setting of pyloric atresia and nail dysplasia.

Application of Modified Skin Stretching for Soft Tissue Defect Reconstruction in the Ankle and Foot: A Retrospective Report.

The failure rate of foot and ankle soft tissue defect reconstruction with flap is relatively high, often posing a significant burden on patients. The aim of this study is to explore the effectiveness of repeated stretch sutures in repairing skin and soft tissue defects of the ankle and foot. Twenty-three patients with ankle and foot skin and soft tissue defects were retrospectively analyzed between February 2016 and February 2019. Sutures were repeatedly stretched every 3-5 days. Local skin grafting was performed if necessary after wound surfaces disappeared or exposed tendons and bones were covered by soft tissue. Wound healing time, postoperative healing area, Vancouver Scar Assessment Scale, sensation, and function of the new skin were evaluated. Healing time was 17-35 (24.43 ± 5.29) days. Ten patients wholly healed, and 13 healed by approximately 70.08% ± 6.59%. The Vancouver Scar Assessment Scale average score was 2.83 ± 1.19 points, of which 15 cases were excellent (0-3 points) and 8 cases were good (4-7 points). The sensation and function of the new skin after repair were equivalent to those of normal skin after the last follow-up. Applying repeated tension sutures on the skin and soft defects of the ankle and foot reduced the skin graft area and decreased complex high-risk surgical flaps' use and transplantation area.

The Anti-proliferative Effect, Apoptotic Induction, and Cell Cycle Arrest of Tetra Halo Ruthenate Nanocomposites in Different Human Cancer Cell Lines.

Chemotherapy is the most common cancer treatment, and metallic anticancer compounds have generated increasing amounts of interest since the discovery of cisplatin. More recently, scientists have focused on ruthenium-based compounds as alternatives for platinum compounds, which seem like ideal therapeutic anticancer alternatives to platinum derivatives. The present study aims to assess whether one or more of three Ruthenium-based nanocomposites, namely Ru+Lysine+CTAB (RCTL), Ru+CTAB (RCT), and Ru+Lysine (RL) exhibit pronounced anti-proliferative properties against different cancer cells. Three Ruthenium nanocomposites have been synthesized by standard chemical methods and characterized by Dynamic light scattering (DLS) and Transmission electron microscopy (TEM). The cytotoxic effect of the three composites has been evaluated by MTT in-vitro assay for different human cancer cell lines, namely MCF7, HepG2, A549, and PC3 versus normal human skin cell line (BJ1). The molecular underlying mechanisms of cytotoxicity have been assessed via qRT-PCR for pro-apoptotic makers P53 and Casp-3, and anti-apoptotic marker Bcl-2 as well as flow cytometric analysis of the cell cycle. Among the 3 nanocomposites, RCTL gave the best sensitivity and cytotoxicity especially on HepG2 with IC50 0.55 µg/ml but was still toxic on normal cell line with dose <12.5 µg/ml. RCTL and RCT nanocomposites have demonstrated a significant increase in the expression of P53 and Casp-3 markers versus untreated controls, but a significant reduction in the expression of Bcl-2. There was a direct correlation between the cytotoxic effect and the degree of apoptosis in the different cancer cell lines. The present study has also proved cell cycle arrest at G2-M and pre-G1 phases under the effect of IC50 of RCTL and RCT nanocomposites in different cancer lines with the best effect being achieved in HepG2 cells. Ruthenium nanocomposites seem to open a new avenue in cancer therapy.

Early Effects of Porcine Placental Extracts and Stem Cell-Derived Exosomes on Aging Stress in Skin Cells.

The initial efficacy of placental extracts (Pla-Exts) and human mesenchymal stem-cell-derived exosomes (hMSC-Exo) against aging-induced stress in human dermal fibroblasts (HDFs) was examined. The effect of Pla-Ext alone, hMSC-Exo alone, the combined effect of Pla-Ext and hMSC-Exo, and the effect of hMSC-Exo (Pla/MSC-Exo) recovered from cultures with Pla-Ext added to hMSC were verified using collagen, elastin, and hyaluronic acid synthase mRNA levels for each effect. Cells were subjected to photoaging (UV radiation), glycation (glycation end-product stimulation), and oxidation (H2O2 stimulation) as HDF stressors. Pla-Ext did not significantly affect normal skin fibroblasts with respect to intracellular parameters; however, a pro-proliferative effect was observed. Pla-Ext induced resistance to several stresses in skin fibroblasts (UV irradiation, glycation stimulation, H2O2 stimulation) and inhibited reactive oxygen species accumulation following H2O2 stimulation. Although the effects of hMSC-Exo alone or the combination of hMSC-Exo and Pla-Ext are unknown, pretreated hMSC-Exo stimulated with Pla-Ext showed changes that conferred resistance to aging stress. This suggests that Pla-Ext supplementation may cause some changes in the surface molecules or hMSC-Exo content (e.g., microRNA). In skin cells, the direct action of Pla-Ext and exosomes secreted from cultured hMSCs pretreated with Pla-Ext (Pla/MSC-Exo) also conferred resistance to early aging stress.

PTB Regulates Keloid Fibroblast Migration and Proliferation Through Autophagy.

Keloid disease is a chronic fibroproliferative disease that occurs after tissue injury, and the currently available treatments are unsatisfactory. We aimed to explore the level of autophagy in keloid fibroblasts (KFbs) and adjacent normal fibroblasts (NFbs). In addition, whether polypyrimidine tract-binding protein (PTB) regulates the biological functions of KFbs via autophagy was also investigated. The morphology of fibroblasts in normal skin and keloids was observed transmission electron microscopy. We silenced PTB with PTB-specific siRNA to determine whether PTB-regulated KFb proliferation. Acridine orange and LysoTracker Red staining was performed to label acidic compartments. Interestingly, when autophagy was inhibited by wortmannin, the PTB knockdown-mediated decrease in KFb migration and proliferation was abolished, while the collagen I and III levels were not altered; these results indicated that PTB regulated the migration and proliferation of KFbs via autophagy, while collagen synthesis occurred independently of PTB regulation. Many activities related to the survival and function of KFbs are controlled by PTB. Transmission electron microscopy revealed more autophagosomes and autolysosomes in KFbs than in NFbs. PTB induced autophagy in KFbs, as demonstrated by the significantly greater number of autophagosomes in KFbs after PTB knockdown, which was revealed by acridine orange and LysoTracker staining. Our study is the first to show that PTB regulates the migration and proliferation of KFbs via autophagy and that PTB regulates collagen synthesis in KFbs in an autophagy-independent manner. This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Assessment of the Antioxidant and Photoprotective Properties of Cornus mas L. Extracts on HDF, HaCaT and A375 Cells Exposed to UVA Radiation.

The influence of UV radiation on skin discoloration, skin aging and the development of skin cancer is widely known. As a part of this study, the effect of extracts from three varieties of Cornus mas L. (C. mas L.) on skin cells exposed to UVA radiation was assessed. The analyses were performed on both normal and cancer skin cells. For this purpose, the potential photoprotective effects of the obtained extracts (aqueous and ethanolic) was assessed by performing two cytotoxicity tests (Alamar blue and Neutral red). Additionally, the antioxidant capacity was compared using three different assays. The 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) probe was used to evaluate the intracellular level of free radicals in cells exposed to the simultaneous action of UVA radiation and dogwood extracts. Additionally, the ability to inhibit excessive pigmentation was determined by assessing the inhibition of melanin formation and tyrosinase activity. The obtained results confirmed the strong antioxidant properties of dogwood extracts and their photoprotective effect on normal skin cells. The ability to inhibit the viability of melanoma cells was also observed. Additionally, a reduction in oxidative stress in skin cells exposed to UVA radiation and a strong inhibition of melanin formation and tyrosinase activity have been demonstrated. This study shows that dogwood extract could be a valuable cosmetic raw material that can play both a photoprotective and antihyperpigmentation role in cosmetic preparations.

The assessment of the phototoxic potential of drugs forming complexes with melanin - Screening in vitro studies using normal skin cells with varying pigmentation irradiated by a sunlight simulator

Phototoxic reactions are among the most common skin-related adverse effects induced by drugs. It is believed that the binding of chemicals to melanin biopolymers is a significant factor influencing skin toxicity. The formation of drug-melanin complexes can lead to the accumulation of drugs or their photodegradation products in pigmented cells, potentially affecting phototoxic reactions. Current procedures for assessing the phototoxic potential of drugs are based on tests using immortalized mouse fibroblasts.This study aimed to assess the phototoxic potential of selected drugs that form complexes with melanin (chloroquine, chlorpromazine, doxycycline) using human melanocytes with varying degrees of pigmentation. Parallel research was conducted on human dermal fibroblasts. To induce phototoxicity, cell cultures were irradiated using a sunlight simulator (5 J/cm2 for UVA spectrum). To account for the process of drug accumulation, two experimental models with different incubation times of cells with drugs before irradiation were used. The photo-irritation factor (PIF) was calculated based on NRU and WST-1 screening tests. Additionally, cell viability was examined cytometrically, and analyses of the cell cycle and reduced glutathione levels were conducted.The results indicated that drugs binding with melanin exhibited different levels of cytotoxicity and phototoxicity towards fibroblasts and melanocytes. These observed differences impact the values of PIF, potentially complicating the interpretation of the studies. Additional analyses, such as examining cell subpopulations in the sub-G1 phase and determining the level of reduced glutathione, can enhance the assessment of the phototoxicity of drugs on pigmented cells.

Characterization and Physiological Differences of Two Primary Cultures of Human Normal and Hypertrophic Scar Dermal Fibroblasts: A Pilot Study.

Background/Objectives: Dermal fibroblasts (DFs) are key participants in skin hypertrophic scarring, and their properties are being studied to identify the molecular and cellular mechanisms underlying the pathogenesis of skin scarring. Methods: In the present work, we performed a comparative analysis of DFs isolated from normal skin (normal dermal fibroblasts, NDFs), and hypertrophic scar skin (hypertrophic scar fibroblasts, HTSFs). The fibroblasts were karyotyped and phenotyped, and experiments on growth rate, wound healing, and single-cell motility were conducted. Results: Comparative analysis revealed a minor karyotype difference between cells. However, HTSFs are characterized by higher proliferation level and motility compared to NDFs. These significant differences may be associated with quantitative and qualitative differences in the cell secretome. A proteomic comparison of NDF and HTSF found that differences were associated with metabolic proteins reflecting physiological differences between the two cells lines. Numerous unique proteins were found only in the vesicular phase of vHTSFs. Some proteins involved in cell proliferation (protein-glutamine gamma-glutamyltransferase K) and cell motility (catenin delta-1), which regulate gene transcription and the activity of Rho family GTPases and downstream cytoskeletal dynamics, were identified. A number of proteins which potentially play a role in fibrosis and inflammation (mucin-5B, CD97, adhesion G protein-coupled receptor E2, antileukoproteinase, protein S100-A8 and S100-A9, protein caspase recruitment domain-containing protein 14) were detected in vHTSFs. Conclusions: A comparative analysis of primary cell cultures revealed their various properties, especially in the cell secretome. These proteins may be considered promising target molecules for developing treatment or prevention strategies for pathological skin scarring.

RETRACTION: MiR-625-5p/PKM2 Negatively Regulates Melanoma Glycolysis State.

H. Zhang, C. Feng, M. Zhang, A. Zeng, L. Si, N. Yu, and M. Bai, "MiR-625-5p/PKM2 Negatively Regulates Melanoma Glycolysis State," Journal of Cellular Biochemistry 120, no. 3 (2019): 2964-2972, https://doi.org/10.1002/jcb.26917. The above article, published online on 30 November 2018 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the authors; the journal Editor-in-Chief, Christian Behl; and Wiley Periodicals LLC. The authors requested a retraction of this manuscript stating that different cell lines were wrongly labelled and that the conclusions of this manuscript needed to be re-evaluated. Further investigation by the publisher revealed that the article mentions use of the non-verifiable/unknown normal skin cells HFE. Additional flaws and inconsistencies between results presented and experimental methods described were identified. Furthermore, the experimental methods were found to lack sufficient details or have unavailable supporting data, making the experiments not comprehensible nor reproducible to readers. Accordingly, the editors consider the conclusions of this article to be invalid.

7640 The first description of an MC4R variant in a patient with Kallmann syndrome and obesity

Abstract Disclosure: A.A. Aslam: None. S. Lim: None. R. Willemsen: None. K. Koysombat: None. M. Young: None. W.S. Dhillo: None. A. Abbara: None. S. Howard: None. E.F. Gevers: None. Introduction: Pathogenic MC4R gene variants result in hyperphagia and early onset obesity but puberty is not usually affected. We describe an MC4R variant in a patient with Kallmann syndrome and obesity. A 16 year old male with repaired Tetralogy of Fallot, anosmia, autism and anxiety, was referred with obesity and delayed puberty. His birth weight was 3360 g (-0.26 SDS). Height was -1.31 SDS, BMI 30.7 kg/m2. He had high arched palate, normal skin and hair colour, mild hypertelorism and upward slanting palpebral fissures. Pubertal staging was A2P1G1 5ml testes bilaterally. Results: LHRH test showed borderline low peaks (LH 5.0 U/L, FSH 2.6 U/L) and inhibin B 108 pg/mL (25-325 pg/mL). MRI brain showed hypoplastic olfactory bulbs, absent olfactory sulci and normal pituitary anatomy. CGH microarray, karyotype and the UK hypogonadism gene panel were normal. He underwent whole genome sequencing in the Genetic Factors Affecting the Timing of Puberty Study; no abnormalities in 52 known genes associated with GnRH deficiency were found but a previously described heterozygous variant was detected, MC4R c.542G&amp;gt;A, p.Gly181Asp, classified pathogenic and absent in control databases. Management: He was commenced on Testosterone but showed progression of testicular size to 10 mL and testosterone was stopped but required restarting. Repeat investigations off treatment, aged 22 years (height 178 cm, BMI 42 kg/m2, testes 15 mL): inhibin B 66 pg/mL, testosterone 2 nmol/L, baseline LH 2.4 U/L, stimulated peak 24.9 U/L, baseline FSH 1.2 U/L, stimulated peak 4.2 U/L. Cortisol and IGF1 were in the normal range. He also developed autoimmune hypothyroidism with raised TPO antibodies (TSH 24.7 mU/L, FT4 5.6 pmol/L). Discussion: Previous in vitro work has shown complete loss of function of MC4R p.Gly181Asp due to reduced cell surface expression and reduced binding to a-MSH. Heterozygous p.Gly181Asp variants have been described in several children/adults with obesity; but hypogonadism in heterozygous carriers has not. Homozygous MC4R p.Gly181Asp was found in a male with obesity and partial HH thought to be due to abnormal GnRH production but with normal olfactory bulbs. We are the first to describe a heterozygous MC4R p.Gly181Asp variant in a patient with partial hypogonadism and anosmia with hypoplastic bulbs in addition to obesity. Interaction between POMC-MC-leptin circuits and Kisspeptin-GnRH circuits is recognised although not completely understood. Previous work has shown that a subset of kisspeptin neurons express MC4R and that KNDy neurones receive synaptic input from POMC neurons. In addition, inhibition of MC4R/MC3R results in delayed onset of puberty in rats. Our results support a role for MC4R in GnRH secretion and potentially olfactory /GnRH neuronal migration. Assessment of spontaneous gonadotrophin production and response of gonadotrophin production to kisspeptin stimulation is currently in progress. Presentation: 6/1/2024

8283 Diabetes Mellitus in Patients with Hemochromatosis- A Diagnostic and Therapeutic Challenge

Abstract Disclosure: U. Rafat: None. M. Siddiqui: None. J.L. Gilden: Other; Self; Novartis Pharmaceuticals. A. Moid: Other; Self; Novartis Pharmaceuticals. Background: Primary Hemochromatosis is a genetic condition most commonly due to two sets of mutations in the Human homeostatic iron regular Protein (HFE) gene. HFE modulates the expression of hepcidin, which regulates intestinal iron absorption, plasma iron concentration and distribution of iron in tissues. Hemochromatosis patients are at high risk for developing diabetes mellitus (DM) due to iron overload in the pancreas with damage to Beta cells and impaired insulin secretory capacity. Hemochromatosis is often called Bronze Diabetes due to brown discoloration of the skin from accumulation of iron, and is characterized by hepatic fibrosis, fatigue, arthralgias, cirrhosis, DM, arthropathy, pigmentation and cardiomyopathy. Case: We present a Caucasian male with homozygous hereditary hemochromatosis, diagnosed by liver biopsy at age 40 due to transaminitis and Ferritin level of 1500 ng/mL and Genetic testing- 2 copies of HFE gene pathogenic variant (C282Y). He was later diagnosed with DM age 51, started on Metformin and referred to Endocrine clinic with abdominal bloating, and polyarticular arthralgias. Physical exam=Vital Signs normal (WT. 231lbs, BMI 34), with splenomegaly, but normal musculoskeletal exam and skin without bronze discoloration. Both parents had T2DM. Lab- Hb a1c 8.7% (0.0-5.6), Iron saturation 71% (10-50), Ferritin 66.3 ng/mL (26-388), C-Peptide 3.30 ng/ml (0.81-3.85), with glucose level of 119 mg/dL, and negative IA-2 &amp;lt;5.4 U/mL, negative Islet cell antibody, GAD 65 &amp;lt;5 IU/mL (nl &amp;lt;5), Insulin Autoantibody &amp;lt;0.4 U/mL (nl &amp;lt;0.4), and Zinc Transporter 8 antibodies &amp;lt;10 U/mL (nl &amp;lt;15). Liver tests- AST 39 U/L(10-37), ALT 77 U/L (10-65), ALP 89 U/L (45-117), total bilirubin 1.1 mg/dL(0.2-1.2). Hormonal evaluation included TSH 1.2 uIU/mL (0.55-4.78), ACTH 19 pg/mL (0-47), AM Cortisol 16.28 ug/d (5.30-22.50) IGF1 158 ng/mL (50-317) FSH 9.6 mIU/mL(l.40-18.10), LH 5.09 mIU/mL (1.50-9.30), Prolactin 7.40 ng/mL(2.10-17.70), total testosterone 374 ng/dL(250-1100), free testosterone 66.2 pg/mL (35.0-155.0). Echocardiogram-left ventricular hypertrophy and thick right ventricular wall with concerns for iron overload. Hba1c improved to 6.6% on Metformin. Fructosamine 351umol/L (151-300). In the absence of advanced cardiac, hepatic and other organ involvement, and presence of elevated C-peptide and high BMI, this patient is being managed as T2DM, with low threshold to initiate insulin therapy considering possibility of concomitant Bronze DM. Conclusion: It is important to determine type of DM, so appropriate management options can be devised. While insulin is recommended for management in Bronze Diabetes, insulin sensitizers, such as metformin and/or GLP 1 agonists can be used in type 2 DM. One must also consider therapy based upon the risk of cancer and acute pancreatitis in the setting of pancreatic iron overload, when considering antihyperglycemic therapies. Presentation: 6/2/2024

Epidermal RORα Maintains Barrier Integrity and Prevents Allergic Inflammation by Regulating Late Differentiation and Lipid Metabolism.

The skin epidermis provides a barrier that is imperative for preventing transepidermal water loss (TEWL) and protecting against environmental stimuli. The underlying molecular mechanisms for regulating barrier functions and sustaining its integrity remain unclear. RORα is a nuclear receptor highly expressed in the epidermis of normal skin. Clinical studies showed that the epidermal RORα expression is significantly reduced in the lesions of multiple inflammatory skin diseases. In this study, we investigate the central roles of RORα in stabilizing skin barrier function using mice with an epidermis-specific Rora gene deletion (RoraEKO). While lacking spontaneous skin lesions or dermatitis, RoraEKO mice exhibited an elevated TEWL rate and skin characteristics of barrier dysfunction. Immunostaining and Western blot analysis revealed low levels of cornified envelope proteins in the RoraEKO epidermis, suggesting disturbed late epidermal differentiation. In addition, an RNA-seq analysis showed the altered expression of genes related to "keratinization" and "lipid metabolism" in RORα deficient epidermis. A lipidomic analysis further uncovered an aberrant ceramide composition in the RoraEKO epidermis. Importantly, epidermal Rora ablation greatly exaggerated percutaneous allergic inflammatory responses to oxazolone in an allergic contact dermatitis (ACD) mouse model. Our results substantiate the essence of epidermal RORα in maintaining late keratinocyte differentiation and normal barrier function while suppressing cutaneous inflammation.

Immunotherapy may be more appropriate for ERBB2 low-expressing extramammary paget’s disease patients: a prognosis analysis and exploration of targeted therapy and immunotherapy of extramammary paget’s disease patients

Extramammary Paget’s disease (EMPD) is a rare cutaneous malignancy characterized by its uncertain etiology and metastatic potential. Surgery remains the first-line clinical treatment for EMPD, but the efficacy of radiotherapy and chemotherapy remains to be fully evaluated, and new therapies for EMPD are urgently needed. In this study, we initially screened 815 EMPD patients in the Surveillance, Epidemiology, and End Results (SEER) database and analyzed their clinical features and prognostic factors. Using the dataset from the Genome Sequence Archive (GSA) database, we subsequently conducted weighted gene coexpression network analysis (WGCNA), gene set enrichment analysis (GSEA), gene set variation analysis (GSVA), and immune infiltration analyses, grouping the samples based on EMPD disease status and the levels of ERBB2 expression. The prognostic analysis based on the SEER database identified increased age at diagnosis, distant metastasis, and receipt of radiotherapy as independent risk factors for EMPD. Moreover, our results indicated that patients who received chemotherapy had worse prognoses than those who did not, highlighting the urgent need for novel treatment approaches for EMPD. Functional analysis of the GSA-derived dataset revealed that EMPD tissues were significantly enriched in immune-related pathways compared with normal skin tissues. Compared with those with high ERBB2 expression, tissues with low ERBB2 expression displayed greater immunogenicity and enrichment of immune pathways, particularly those related to B cells. These findings suggest that patients with low ERBB2 expression are likely to benefit from immunotherapy, especially B-cell-related immunotherapy.