Reticular Dermis Research Articles

Quantitative and anatomical imaging of dermal angiopathy by noninvasive photoacoustic microscopic biopsy.

The ability to noninvasively acquire the fine structure of deep tissues is highly valuable but remains a challenge. Here, a photoacoustic microscopic biopsy (PAMB) combined switchable spatial-scale optical excitation with single-element depth-resolved acoustic detection mode was developed, which effectively coordinated the spatial resolution and the penetration depth for visualizations of skin delamination and chromophore structures up to reticular dermis depth, with the lateral resolution from 1.5 to 104 μm and the axial resolution from 34 to 57 μm. The PAMB obtained anatomical imaging of the pigment distribution within the epidermis and the vascular patterns of the deep dermal tissue, enabling quantification of morphological abnormalities of angiopathy without the need for exogenous contrast agents. The features of healthy skin and scar skin, and the abnormal alteration of dermal vasculature in port wine stains (PWS) skin were first precisely displayed by PAMB-shown multi-layered imaging. Moreover, the quantitative vascular parameters evaluation of PWS were carried out by the detailed clinical PAMB data on 174 patients, which reveals distinct differences among different skin types. PAMB captured the PWS changes in capillary-loop depth, diameter, and vascular volume, making it possible to perform an objective clinical evaluation on the severity of PWS. All the results demonstrated the PAMB can provide vascular biopsy and new indexes deep into the dermal skin noninvasively, which should be meaningful to timely evaluate the pathological types and treatment response of skin diseases. This opens up a new perspective for label-free and non-invasive biopsies of dermal angiopathy.

Primary Anetoderma and Acquired Cutis Laxa Associated with Glomerulonephritis in a 37-year-old Filipino Male: A Case Report

A 37-year-old Filipino man presented with a 9-month history of sagging skin progressing cephalocaudally from the chin and neck to the axillae, side of the trunk, and pelvic area. This was followed by a 2-month history of increasing serum creatinine levels associated with periorbital and bipedal edema, generalized weakness, decreased appetite, vomiting, and headache. Subsequently, skin-colored, non-tender sac-like plaques appeared on the abdomen, inguinal, and intergluteal areas. Histopathology of the latter lesions showed increased spaces between collagen bundles in the dermis. Staining with Verhoeff-van Gieson revealed focal sparse elastic fibers in the papillary dermis compared to that of the reticular dermis consistent with anetoderma. Further work-up revealed normal ANA titer and low serum C3. Kidney biopsy showed IgG deposition in the tubular basement membrane and trace C3 deposition in the glomerular mesangium, giving a diagnosis of rapid progressive glomerulonephritis. On subsequent follow-up, the sac-like plaques became lax and presented as generalized wrinkling of the skin, raising the question whether cutis laxa and anetoderma are occurring in a spectrum instead as distinct entities. Based on the current review of literature, this is the first reported case of primary anetoderma co-occurring with cutis laxa in a patient with glomerulonephritis. Deposition of immunoglobulins along the elastic fibers could have activated the complement system, mediating the destruction of the elastic fibers, resulting to cutis laxa and anetoderma. This case also considers the possibility of anetoderma and type I acquired cutis laxa occurring either in a spectrum or as distinct diseases in a single patient. Further investigations may identify an ultrastructural pattern that can help differentiate the two entities.

Use of 816 Consecutive Burn Wound Biopsies to Inform a Histologic Algorithm for Burn Depth Categorization

Burn experts are only 77% accurate when subjectively assessing burn depth, leaving almost a quarter of patients to undergo unnecessary surgery or conversely suffer a delay in treatment. To aid clinicians in burn depth assessment (BDA), new technologies are being studied with machine learning algorithms calibrated to histologic standards. Our group has iteratively created a theoretical burn biopsy algorithm (BBA) based on histologic analysis, and subsequently informed it with the largest burn wound biopsy repository in the literature. Here, we sought to report that process. This was an IRB-approved, prospective, multicenter study. A BBA was created a priori and refined in an iterative manner. Patients with burn wounds assessed by burn experts as requiring excision and autograft underwent 4 mm biopsies procured every 25 cm2. Serial still photos were obtained at enrollment and at excision intraoperatively. Burn biopsies were histologically assessed for presence/absence of epidermis, papillary dermis, reticular dermis, and proportion of necrotic adnexal structures by a dermatopathologist using H&E with whole slide scanning. First degree and superficial second degree were considered to be burn wounds likely to have healed without surgery, while deep second- and third-degree burns were considered unlikely to heal by 21 days. Biopsy pathology results were correlated with still photos by five burn experts for consensus of final burn depth diagnosis. Sixty-six subjects were enrolled with 117 wounds and 816 biopsies. The BBA was used to categorize subjects' wounds into four categories: 7% of burns were categorized as first degree, 13% as superficial second degree, 43% as deep second degree, and 37% as third degree. Therefore, 20% of burn wounds were incorrectly judged as needing excision and grafting by the clinical team as per the BBA. As H&E is unable to assess the viability of papillary and reticular dermis, with time our team came to appreciate the greater importance of adnexal structure necrosis over dermal appearance in assessing healing potential. Our study demonstrates that a BBA with objective histologic criteria can be used to categorize BDA with clinical misclassification rates consistent with past literature. This study serves as the largest analysis of burn biopsies by modern day burn experts and the first to define histologic parameters for BDA.

28576 Pseudoxanthoma elasticum-like changes in panniculitides: A case series

Pseudoxanthoma elasticum (PXE) is a rare, inherited connective tissue disorder characterized by the accumulation of calcified elastic fibers. This mineralization can affect elastic fibers in the skin, eyes, cardiovascular tissue, and less commonly, the gastrointestinal tract, leading to various clinical presentations such as skin changes, vision problems, or claudication. Histopathologic features of PXE include thickened, calcified elastic fibers within the reticular dermis. There have been reported cases of inflammatory skin diseases that demonstrate identical or highly resemblant PXE histopathologic changes without clinical manifestations of PXE. We herein report a case series of four patients diagnosed with panniculitis associated with PXE-like changes without any clinical features of PXE. The clinical significance of these findings remain uncertain due to limited literature. In addition to reporting our institutional experience on four clinical cases, we will discuss the current understanding of the proposed pathogenesis of panniculitis with PXE features and encourage further investigation to elucidate the clinical significance of these findings.

4: ACTA2+ Cells Activation and Dermal Changes During Skin Adaptation to Mechanical Forces

Purpose:Tissue expansion (TE) is based on the skin’s exceptional ability to regenerate under mechanical stress and is widely used to repair skin defects. However, knowledge about molecular mechanisms involved in maintaining skin integrity and homeostasis is limited. The present study aims 1) to elucidate the role of myofibroblasts in the mechanism of adaptation to mechanical stress exerted by an inflated tissue expander, and 2) to describe morphological changes in collagen structure that could lead to the re-establishment of dermal tension.Methods:TE was performed on a porcine model. Each expander was placed subcutaneously over the ribs and two weeks later inflated with 30 cc of saline to induce subtle tension. After 1 day (acute stretch) and 7 days (sub-acute stretch) of expansion, the full-thickness skin biopsies were collected from the apex of the expander and control unexpanded skin (contralateral sites). Skin samples were fixed in formalin and embedded in paraffin for histological evaluation (Russell-Movat Pentachrome staining) or fixed in 4% PFA and embedded in OCT for immunohistochemistry staining (IF) of α-smooth muscle actin (α-SMA), a marker of myofibroblast. Area of fluorescent signal from α-SMA was calculated using ImageJ while collagen morphology was evaluated optically after staining.Results:We compared the presence of α-SMA fluorescence between control biopsies and expanded biopsies after 1-day and 7-days of stretch. The immunofluorescence staining revealed 2.32 times more α-SMA fluorescent staining in skin expanded for 1 day (p = 0.0065) and 2.19 times more α-SMA fluorescence in skin expanded for 7 days (p = 0.0047). The increase in number of α-SMA positive cells were mostly observed in the outermost 400µm of papillary dermis. Histological staining showed minimal collagen morphological changes in both the papillary and reticular dermis after 1-day of expansion. However, shortening of fibril length, increased density, and increased disorder were observed in the papillary dermis collagen after 7-days of subtle expansion.

Cutaneous mucinosis of infancy: a rare case of joint involvement

BackgroundPrimary cutaneous mucinosis are a heterogeneous group of diseases characterized by the deposition of glycosaminoglycans in the dermis and the follicles. These diseases are rare in children therefore their diagnosis and management are still challenging. Joint involvement has been reported in patients with secondary cutaneous mucinosis and, rarely, in primary mucinosis. We describe a case of Cutaneous Mucinosis of Infancy with joint involvement.Case presentationAn healthy 5-year-old boy showed acute arthritis of the left knee and left elbow confirmed by ultrasound. Laboratory tests were within normal range. Symptoms disappeared after a course of nonsteroid anti-inflammatory drugs. One year later, the knee swelling reappeared; juvenile idiopathic arthritis was diagnosed and intra-articular steroid injection was performed. Due to persistence of arthritis of the knee he was admitted to our hospital. On physical examination variable skin-colored lesions were observed, which had been in existence for over 2 years.We performed a skin biopsy that showed an interstitial mucine deposition in the reticular dermis. Cutaneous Mucinosis of Infancy was diagnosed.Discussion and conclusionsCutaneous Mucinosis of Infancy is a persistent dermatosis with benign prognosis and no treatment is generally required. Our case report is particularly interesting because it is the first in which joint involvement has been reported in CMI, a disorder that has so far been described as limited to skin involvement. Further studies will be necessary in order to clarify the pathogenesis of joint involvement in primary mucinosis.

A New Model for Specific Visualization of Skin Graft Neoangiogenesis Using Flt1-tdsRed BAC Transgenic Mice.

Neovascularization plays a critical role in skin graft survival. Up to date, the lack of specificity to solely track the newly sprouting blood vessels has remained a limiting factor in skin graft transplantation models. Therefore, the authors developed a new model by using Flt1-tdsRed BAC transgenic mice. Flt1 is a vascular endothelial growth factor receptor expressed by sprouting endothelial cells mediating neoangiogenesis. The authors determined whether this model reliably visualizes neovascularization by quantifying tdsRed fluorescence in the graft over 14 days. Cross-transplantation of two full-thickness 1 × 1-cm dorsal skin grafts was performed between 6- to 8-week-old male Flt1 mice and KSN/Slc nude mice (n = 5). The percentage of graft area occupied by tdsRed fluorescence in the central and lateral areas of the graft on days 3, 5, 9, and 14 was determined using confocal-laser scanning microscopy. Flt1+ endothelial cells migrating from the transgenic wound bed into the nude graft were first visible in the reticular dermis of the graft center on day 3 (0.5 ± 0.1; p < 0.05). Peak neovascularization was observed on day 9 in the lateral and central parts, increasing by 2- to 4-fold (4.6 ± 0.8 and 4.2 ± 0.9; p < 0.001). Notably, some limited neoangiogenesis was displayed within the Flt grafts on nude mice, particularly in the center. No neovascularization was observed from the wound margins. The ability of the Flt1-tdsRed transgenic mouse model to efficiently identify the origin of the skin-graft vasculature and visualize graft neovascularization over time suggests its potential utility for developing techniques that promote graft neovascularization.

In vitro beneficial effects of a flax extract on papillary fibroblasts define it as an anti-aging candidate

Objective: During aging, skin undergoes structural, cellular and molecular changes, which not only alter skin mechanical properties but also biological and physiological functions. Structurally the epidermis becomes thinner, the dermal epidermal junction flattens and the extra-cellular matrix component of the dermis is disorganized and degraded. The dermis is composed of two compartments: The Reticular dermis is the deepest and thickest part while the upper layer, the papillary dermis, which is much thinner and is in close contact with epidermis, plays an important role in the structure and function of the skin. We have recently shown that the papillary dermis was preferentially affected by skin aging because the activity of fibroblasts in this region was especially altered as a function of age. The purpose of this study was to investigate the capacity of a flax extract as anti-aging component. Method: We investigated the capacity of a flax extract to stimulate or restore the activity of papillary fibroblasts from young and old donors in cultured monolayers and in reconstructed skin. Several biological markers of extracellular matrix homeostasis and mechanical properties were investigated. Results: The tested flax extract seemed to improve parameters known to change with age: I/ In monolayers after treatment the number of aged fibroblasts increased II/ In reconstructed skin the flax extract appears to positively regulate some biological activities; particularly in aged fibroblasts where the deposition of laminin 5, fibrillin 1, procollagen I were increased in the dermis and the secretion of specific soluble factors like MMP1, MMP3 and KGF were regulated to levels similar to those observed in young fibroblasts III/ Mechanical properties were improved particularly for elastics parameters (R5, R2 and R7). Conclusion: The flax extract is a promising anti-aging compound. The treatment of aged papillary fibroblasts resulted in a return to a younger-like profile for some of the studied parameters.

Загоєння ран шкіри щурів старечого віку за умов хронічної гіперглікемії та при застосуванні збагаченої тромбоцитами плазми

Wound regeneration is a complex problem for modern healthcare, since the existing methods of treatment are not always effective, which necessitates the development of new promising methods. In this regard, today researchers are greatly interested in the possibility of using platelet-rich plasma in the treatment of patients with soft tissue injuries. The purpose of our research was to study the micro-, ultrastructural, planimetric and morphometric features of the skin reparative regeneration of old rats under the chronic hyperglycemia using platelet-rich plasma. Material and methods. The study was conducted on 30 white laboratory old rats: І group was a control group, ІІ group consisted of animals with chronic hyperglycemia of the body; ІІІ group included animals with chronic hyperglycemia of the body that were administered with platelet-rich plasma into the skin defect. The chronic hyperglycemia was simulated by the single intraperitoneal administration of streptozotocin (40 mg/kg) and nicotinic acid (1 mg/kg). The wound was formed by cutting out a piece of skin in the interscapular region. The animals with chronic hyperglycemia of the body ІІІ group were administered with autologous platelet-rich plasma into the skin defect. The studies were performed on the 14 days after trauma. The light microscopy was performed using an Olympus BH-2 microscope (Japan), ultramicroscopic study was performed using a JEM-1230 transmission electron microscope (JEOL, Japan), the planimetric analysis was performed using the morphometric program "Digimizer". The statistical analysis was performed using the SPSS-17 software package. Results and discussion. The obtained results showed that the process of skin regeneration in elderly rats with chronic hyperglycemia, which were injected with platelet-rich plasma, had a more favorable course than in animals that did not use the corrector. The wound area was reduced by 23.57 % (p = 0.654) than in rats with chronic hyperglycemia without correction and by 65.29 % (p = 0.857) larger than in the control group. It was shown that in animals with impaired carbohydrate metabolism, which received platelet-rich plasma, the beginning of the formation of the reticular and papillary layers dermis was observed. Conclusion. The vascular density of the microvasculature in animals with chronic hyperglycemia, which was injected with the corrector, was 22.89 % more (p = 0.246) than in animals simulated with chronic hyperglycemia without a corrector, but 15.69 % (p = 0.066) less than the control value

634 Single cell transcriptomics identifies a two way conversion program between dermal progenitors and adipocytes during skin development and regeneration

Dermal white adipose tissue (dWAT) is a unique layer of adipocytes and their fibroblast precursors within the reticular dermis of the skin, and recent studies have found that dWAT plays an essential role in skin development and regeneration. However, little is known how adipocytes are developed from their dermal progenitor and how adipogenesis is controlled during skin regeneration. Here, we performed single-cell RNA-sequencing (scRNA-seq) analyses of dermal fibroblasts (dFBs) isolated from mice at various post-developmental ages. Pseudotime analyses of these dFB clusters identified a Pdgfra+CD24hiThy1loSca1lo population as developmental progenitors for preadipocytes (pAD) and this progenitor population was highly abundant in neonatal skin early in life but declined in adulthood. In contrast, a Pdgfra+Sca1hiDPP4hi adult reticular interstitial (RI) adipocyte progenitor (AP) population that developed postnatally was found in adult skin. ScRNAseq analyses were then performed on dFBs from wounded skin to determine the cell fate and role of these adult RI-APs during skin regeneration. Pseudotime analysis and immunofluorescence analysis of wounded skin showed that RI-APs rapidly infiltrated the wound center and gave rise to pAD/AD and/or myofibroblasts during early stages of wound healing, contributing to dermal regeneration. Pathway analyses revealed that this two way conversion between dermal progenitors and pAD/AD was dynamically regulated by an interplay between the WNT, TGFβ and NFKB pathways. Together, our results have identified and defined the developmental pathways that shape adipogenesis from dermal progenitors during skin development and regeneration. These results provide insights into how defective adipocyte progenitor function and adipogenesis may be associated with defective wound healing associated with aging and diabetes and/or fibrotic diseases such as keloid scarring.

136 Use of 816 Consecutive Burn Wound Biopsies to Inform a Histologic Algorithm for Burn Depth Diagnosis

Abstract Introduction Burn experts are only 77% accurate when subjectively assessing burn depth, leaving almost a quarter of patients to undergo unnecessary surgery or conversely suffer a delay in treatment. To aid clinicians in burn depth assessment (BDA), new technologies are being studied with machine learning algorithms calibrated to histologic standards. Our group has iteratively created a theoretical burn biopsy algorithm (BBA) based on histologic analysis, and subsequently informed it with the largest burn wound biopsy repository in the literature. Here, we sought to report that process. Methods This was an IRB-approved, prospective, multicenter study. A BBA was created a priori and refined in an iterative manner, resulting in the current state of the algorithm seen in Figure 1. Patients with burn wounds assessed by burn experts as requiring excision and autograft underwent 4mm biopsies procured every 25cm2. Serial still photos were obtained at enrollment and at excision intraoperatively. Burn biopsies were histologically assessed for presence/absence of epidermis, papillary dermis, reticular dermis, and proportion of necrotic adnexal structures by a dermatopathologist using H&amp;E with whole slide scanning. First degree and superficial 2nd degree were considered to be burn wounds likely to have healed without surgery, while deep 2nd and 3rd degree burns were considered unlikely to heal by 21 days. Biopsy histopathology results were correlated with still photos by 3 burn experts for consensus of final burn depth diagnosis. Results Sixty-six subjects were enrolled with 117 wounds and 816 biopsies. The BBA was used to categorize 100% of subjects into 4 categories: 7% of burns were categorized as 1st degree, 13% as superficial 2nd degree, 43% as deep 2nd degree, and 37% as 3rd degree. Therefore 20% of burn wounds were incorrectly judged as needing excision and grafting by the clinical team. As H&amp;E is unable to assess the viability of papillary and reticular dermis, with time our team came to appreciate the greater importance of adnexal structure necrosis over dermal appearance in assessing healing potential. Conclusions Our study demonstrates that a BBA with objective histologic criteria can be used to categorize BDA with clinical misclassification rates consistent with past literature. This study serves as the largest analysis of burn biopsies by modern day burn experts and the first to define histologic parameters for BDA.

Classification and clinical evaluation of the types of angiokeratoma

Background: Angiokeratoma is a group of benign vascular telangiectasias in the superficial dermis associated with hyperkeratosis of the epidermis. There are different types of angiokeratoma and the color of the lesions—pink, red, dusky red, blue-red—depends on their age. Materials and Methods: Twenty-four patients with different varieties of angiokeratoma were collected from April 2013 through March 2020 and classified according to well-defined types. Each patient gave their formal consent after the nature of the disease was explained to them. Full medical history taking and clinical examination were performed on all patients. Skin biopsies were taken from 12 cases and processed for histopathology with H&amp;E stain. As for therapy, diathermy was used for the treatment of selected localized cases, especially those on the scrotum. Results: Twenty-four cases with different types of angiokeratoma were evaluated, 19 (79.16%) males and 5 (20.83%) females, with ages ranging from 13 to 25 years and a mean age of 19 years. The age of onset was most commonly around adolescence and early adulthood. The lesions were classified into the following: 11 (45.83%) cases of Fordyce, 5 (20.83%) cases solitary, 4 (16.66%) cases circumscriptum, 3 (12.5%) cases oral (on the tongue), and one (4.16%) corporis diffusum (Fabry disease). All patients displayed warty red, blue, or black papules, nodules, or plaques, with the exception of isolated angiokeratoma. Their histopathology revealed hyperkeratosis of the epidermis, which could have been mild or marked acanthosis, as seen in the form of pseudoepithelial hyperplasia with marked elongation of rete ridges. As a dermal change, markedly dilated blood vessels occupied the papillary and reticular dermis. Diathermy was an effective mode of therapy in selected patients with satisfactory cosmetic and therapeutic results. Conclusion: All types of angiokeratoma were recognized but the most common were angiokeratoma of Fordyce, mainly affecting male genitalia, followed by solitary angiokeratoma, affecting mainly the limbs, and angiokeratoma circumscriptum linearly along the limbs. Oral angiokeratoma of the tongue appeared in three patients. All these types have a characteristic presentation that is easy to recognize: warty dusky-red lesions.

What the pyogenic granuloma hides

A 10-year-old male was referred to our outpatient dermatological clinic complaining of a lesion with vascular characteristics on the right thigh of 2 months of evolution (Fig. 1A). The lesion was asymptomatic but bled with minimal trauma. Dermoscopic study revealed purplish and reddish lacunae with slight hyperkeratosis in the lower pole without melanocytic structures (Fig. 1B). The double rail sign was not appreciated (This sign comprises white lines that correspond to the histologic finding of fibrous septa that surround the lobules). Clinical diagnosis of pyogenic granuloma was stated, and a biopsy was performed. The histological report revealed a lesion made up of rounded cells and epithelioid to a lesser extent that affects papillary and reticular dermis (Fig.1C), ulcerated epidermis and significant inflammatory infiltrate and a high number of mitoses (9 mitoses/mm2) (Fig. 1D). Immunohistochemistry revealed Ki67 greater than 70% and HMB45 intensely +. What is your diagnosis and how to manage this case? (Answer on page ••) We concluded the diagnosis of Spitzoid Melanoma pT3b. A massive DNA and RNA sequencing study of genes associated with familial melanoma was performed, being negative. The case was discussed in the Multidisciplinary Committee and Sentinel Lymphatic Node Biopsy (SLNB) was performed with a negative result. Six months after the SLNB, the patient remains disease-free. Spitz nevus (SN) is a melanocytic neoplasm of epithelioid or spindle cells first described in 1910 by Darier and Civatte and characterised in 1948 by Spitz under the name of Juvenile Melanoma. It was described as a melanocytic tumour, present in childhood, with clinical and pathological characteristics similar to a melanoma but with a benign behaviour.1 Melanocytic neoplasms with a spitzoid morphology include a spectrum of progression between benign and malignant lesions. In recent decades, there has been a tendency to classify them into 3 types, sometimes difficult to distinguish from each other: conventional NS, atypical Spitz tumour (AST) and spitzoid melanoma (SM).2 SM is relatively rare in paediatric patients and has a favourable prognosis. Scientific evidence is supported by a retrospective series of patients who identify a low prevalence of death from this cause in the period 1949 to 2006 (25 fatal cases).3 The incidence of childhood cutaneous melanoma in the USA was 0.3–0.4% in patients less than 10 years old.4 From a clinical point of view, SM is usually presented as amelanotic lesions, which are frequently ulcerated with serous crusts. It is not uncommon for the clinician to raise the following as differential diagnoses: infantile haemangioma, pyogenic granuloma, xanthogranuloma or basal cell carcinoma (extremely rare in paediatric age). Therefore, the biopsy is mandatory. Concerning the complementary tests, histological examination remains the gold standard. Histopathology shows atypical melanocytic nests with fusiform or epithelioid morphology with a hyperchromatic and pleomorphic nucleus. The main immunohistochemical markers used to differentiate its malignant character are HMB45 and MIB-1. The use of genomic tests to assist in the diagnosis of SM has been expanded in recent years,5 helping us to understand and characterise it. Copy number abberations through FISH or CGH are increasingly been used to assist in its diagnosis. The main associated controversy arises in the positioning of the selective biopsy of the sentinel node in contrast to the adult population due to the high number of positive cases with good evolution. A series of 10 paediatric patients with SM treated by complete excision and without SLNB revealed, after a mean follow-up of 3 years, that all patients were disease free. In conclusión, MS is a rare disease that requires a different approach compared to adult melanoma. There is a propensity for SM to metastasize to local lymph nodes but not distant sites. Although there have been reports of late recurrence, we would like to emphasise that involvement with a multidisciplinary team is essential and an appropriate clinical long-term follow-up is important.6

T cell infiltration on local CpG-B delivery in early-stage melanoma is predominantly related to CLEC9A+CD141+ cDC1 and CD14+ antigen-presenting cell recruitment

BackgroundWe previously reported CpG-B injection at the primary tumor excision site prior to re-excision and sentinel node biopsy to result in immune activation of the sentinel lymph node (SLN), increased...

ABCB5+ dermal mesenchymal stromal cells with favorable skin homing and local immunomodulation for recessive dystrophic epidermolysis bullosa treatment.

Recessive dystrophic epidermolysis bullosa (RDEB) is a rare, incurable blistering skin disease caused by biallelic mutations in type VII collagen (C7). Advancements in treatment of RDEB have come from harnessing the immunomodulatory potential of mesenchymal stem cells (MSCs). Although human bone marrow-derived MSC (BM-MSC) trials in RDEB demonstrate improvement in clinical severity, the mechanisms of MSC migration to and persistence in injured skin and their contributions to wound healing are not completely understood. A unique subset of MSCs expressing ATP-binding cassette subfamily member 5 (ABCB5) resides in the reticular dermis and exhibits similar immunomodulatory characteristics to BM-MSCs. Our work aimed to test the hypothesis that skin-derived ABCB5+ dermal MSCs (DSCs) possess superior skin homing ability compared to BM-MSCs in immunodeficient NOD-scid IL2rgammanull (NSG) mice. Compared to BM-MSCs, peripherally injected ABCB5+ DSCs demonstrated superior homing and engraftment of wounds. Furthermore, ABCB5+ DSCs vs BM-MSCs cocultured with macrophages induced less anti-inflammatory interleukin-1 receptor antagonist (IL-1RA) production. RNA sequencing of ABCB5+ DSCs compared to BM-MSCs showed unique expression of major histocompatibility complex class II and Homeobox (Hox) genes, specifically HOXA3. Critical to inducing migration of endothelial and epithelial cells for wound repair, increased expression of HOXA3 may explain superior skin homing properties of ABCB5+ DSCs. Further discernment of the immunomodulatory mechanisms among MSC populations could have broader regenerative medicine implications beyond RDEB treatment.

Clinical assessment of the use of topical liquid diclofenac following laser microporation of cutaneous neurofibromas in individuals with neurofibromatosis type 1

BackgroundNeurofibromatosis type 1 (NF1) is an autosomal dominant genetic disorder with a prevalence of 1:3000 births and a wide variety of clinical manifestations. Cutaneous neurofibromas (cNF) are among the most common visible manifestations of NF1 and present a major clinical burden for patients. NF1 patients with cNF often report decreased quality of life, emotional well-being and physical comfort. Developing effective medical therapies for cNF has been identified as a priority for the majority of adults with NF1. MethodsThe study was an open, controlled and prospective proof-of-concept clinical trial. The topical treatment consisted of two steps: cNF microporation using a laser device followed by topical application of one drop of diclofenac 25 mg/mL on the surface of the cNF (T neurofibroma = treatment) or physiological saline (C neurofibroma = control) and reapplied twice daily for 3 days. Neurofibroma assessments included visual and dermatoscopy observations noting color and presence of necrosis, presence of flaccidity, measurements in two dimensions, photographs, and histopathology after excision. The primary efficacy variable was the presence of tissue necrosis. The primary safety variable was the occurrence of treatment-related adverse events. ResultsSix patients were included in the study. The treatment resulted in transitory topical changes (healing of the microporation grid with formation of scintillating tissue layer, hyperemia and desquamation), with no statistically significant variation in the dimensions of the T and C neurofibromas in relation to pretreatment measurements. There was no necrosis in the T or C neurofibromas. In the histopathological analysis, there was no significant difference in the distribution of chronic (lymphocytic) inflammatory infiltrate in the papillary reticular dermis (subepithelial), type of infiltrate (diffuse, perivascular, or both), presence of fibrosis, and presence of atrophy among the T and C neurofibromas. No adverse events attributable to the use of diclofenac were reported during the treatment period. ConclusionsTreatment did not result in significant alterations in terms of presence of tissue necrosis, size, or histopathological features in the T neurofibromas or in comparison to the C neurofibromas. Topical diclofenac with laser microporation was well-tolerated, with no adverse events attributable to diclofenac reported. Whether these observations are due to minimal systemic and neurofibroma exposure remain to be explored in dosage studies with larger patient groups. Trial registrationClinicalTrials.gov (NCT03090971) retrospectively registered March 27, 2017.

From SARS-CoV-2 hematogenous spreading to endothelial dysfunction: clinical-histopathological study of cutaneous signs of COVID-19

BackgroundTo date, very few studies on clinical-histopathological correlations of cutaneous disorders associated with COVID-19 have been conducted.Case presentationThe Case 1 was a 90-year-old man, who tested positive for SARS-CoV-2 from a nasopharyngeal swab. Two days later, he was hospitalized and after eleven days transferred to Intensive Care Unit. A chest CT showed bilateral ground-glass opacities. Just that day, an erythematous maculo-papular rash appeared on trunk, shoulders and neck, becoming purpuric after few days. Histological evaluations revealed a chronic superficial dermatitis with purpuric aspects. The superficial and papillary dermis appeared edematous, with a perivascular lympho-granulocytic infiltrate and erythrocytic extravasation. At intraepithelial level, spongiosis and a granulocyte infiltrate were detected. Arterioles, capillaries and post-capillary venules showed endothelial swelling and appeared ectatic. The patient was treated with hydroxychloroquine, azithromycin, lopinavir-ritonavir and tocilizumab. Regrettably, due to severe lung impairment, he died.The Case 2 was a 85-year-old man, admitted to Intensive Care Unit, where he was intubated. He had tested positive for SARS-CoV-2 from a nasopharyngeal swab two days before. A chest RX showed bilateral atypical pneumonia. After seven days, a cutaneous reddening involving trunk, upper limbs, neck and face developed, configuring a sub-erythroderma. Histological evaluations displayed edema in the papillary and superficial reticular dermis, and a perivascular lymphocytic infiltrate in the superficial dermis. The patient was treated with hydroxychloroquine, azithromycin, lopinavir-ritonavir and tocilizumab. Sub-erythroderma as well as respiratory symptoms gradually improved until healing.ConclusionsThe endothelial swelling detected in the Case 1 could be a morphological expression of SARS-CoV-2-induced endothelial dysfunction. We hypothesize that cutaneous damage could be initiated by endothelial dysfunction, caused by SARS-CoV-2 infection of endothelial cells or induced by immune system activation. The disruption of endothelial integrity could enhance microvascular permeability, extravasation of inflammatory cells and cytokines, with cutaneous injury. The Case 2 developed a sub-erythroderma associated with COVID-19, and a non-specific chronic dermatitis was detected at histological level. We speculate that a purpuric rash could represent the cutaneous sign of a more severe coagulopathy, as highlighted histologically by vascular abnormalities, while a sub-erythroderma could be expression of viral hematogenous spreading, inducing a non-specific chronic dermatitis.

A biphasic multilayer computational model of human skin

The present study investigates the layer-specific mechanical behavior of human skin. Motivated by skin’s histology, a biphasic model is proposed which differentiates between epidermis, papillary and reticular dermis, and hypodermis. Inverse analysis of ex vivo tensile and in vivo suction experiments yields mechanical parameters for each layer and predicts a stiff reticular dermis and successively softer papillary dermis, epidermis and hypodermis. Layer-specific analysis of simulations underlines the dominating role of the reticular dermis in tensile loading. Furthermore, it shows that the observed out-of-plane deflection in ex vivo tensile tests is a direct consequence of the layered structure of skin. In in vivo suction experiments, the softer upper layers strongly influence the mechanical response, whose dissipative part is determined by interstitial fluid redistribution within the tissue. Magnetic resonance imaging-based visualization of skin deformation in suction experiments confirms the deformation pattern predicted by the multilayer model, showing a consistent decrease in dermal thickness for large probe opening diameters.

Numerical Study on Factors Affecting the Induction of Apoptotic Temperatures of Tumor in the Multi-Layer Skin Structure Using Monte Carlo Method

The incidence of skin cancer is increasing with the recent increase in UV exposure. The treatment of skin cancer generally proceeds through an excision of the tumor area, which causes bleeding into the affected area and surrounding tissues, and there is a possibility that secondary infection may occur. Photothermal therapy is drawing attention as an alternative treatment to overcome this limitation. In this study, a numerical analysis was performed on skin cancer tumors located between the reticular dermis and the skin surface by applying the Monte Carlo method. The numerical analysis derives a quantitative correlation using an effective apoptosis ratio with respect to the intensity of the laser that produces the optimal photothermal therapy effect and the volume fraction of gold nanorods (GNRs) injected into a tumor. Through this study, it is confirmed that the optimal treatment effect exists for the depth and length of the various tumors, the intensity of the laser, and the volume fraction of GNRs to minimize the thermal damage to the surrounding normal tissues while maximizing the apoptosis in the tumor. It is expected that it can be used as an optimal condition for better treatment while performing photothermal therapy in the future.

The effect of TGFβRI inhibition on fibroblast heterogeneity in hypertrophic scar 2D in vitro models

In burn patients, wound healing is often accompanied by hypertrophic scarring (HTS), resulting in both functional and aesthetic problems. HTSs are characterized by abundant presence of myofibroblasts (MFs) residing in the dermis. HTS development and MF persistence is primarily regulated by TGF-β signalling. A promising method to target the transforming growth factor receptor I (TGFβRI; also known as activin-like kinase 5 (ALK5)) is by making use of exon skipping through antisense oligonucleotides. In HTS the distinguishing border between the papillary dermis and the reticular dermis is completely abrogated, thus exhibiting a one layered dermis containing a heterogenous fibroblast population, consisting of papillary fibroblasts (PFs), reticular fibroblasts (RFs) and MFs. It has been proposed that PFs, as opposed to RFs, exhibit anti-fibrotic properties. Currently, it is still unclear which fibroblast subtype is most affected by exon skipping treatment.Therefore, the aim of this study was to investigate the effect of TGFβRI inhibition by exon skipping in PF, RF and HTS fibroblast monocultures.Morphological analyses revealed the presence of a PF-like population after exon skipping in the different fibroblast cultures. This observation was further confirmed by the expression of genes specific for PFs, demonstrated by qPCR analyses. Further investigations on mRNA and protein level revealed that indeed MFs and to a lesser extent RFs are targeted by exon skipping. Furthermore, collagen gel contraction analysis showed that ALK5 exon skipping reduced TGF-β- induced contraction together with decreased alpha-smooth muscle actin expression levels.In conclusion, we show for the first time that exon skipping primarily targets pro-fibrotic fibroblasts. This could be a promising step towards reduced HTS development of burn tissue.