Basal Cells Research Articles

Single-cell atlas of healthy vocal folds and cellular function in the endothelial-to-mesenchymal transition.

The vocal fold is an architecturally complex organ comprising a heterogeneous mixture of various layers of individual epithelial and mesenchymal cell lineages. Here we performed single-cell RNA sequencing profiling of 5836 cells from the vocal folds of adult Sprague-Dawley rats. Combined with immunostaining, we generated a spatial and transcriptional map of the vocal fold cells and characterized the subpopulations of epithelial cells, mesenchymal cells, endothelial cells, and immune cells. We also identified a novel epithelial-to-mesenchymal transition-associated epithelial cell subset that was mainly found in the basal epithelial layers. We further confirmed that this subset acts as intermediate cells with similar genetic features to epithelial-to-mesenchymal transition in head and neck squamous cell carcinoma. Finally, we present the complex intracellular communication network involved homeostasis using CellChat analysis. These studies define the cellular and molecular framework of the biology and pathology of the VF mucosa and reveal the functional importance of developmental pathways in pathological states in cancer.

Structural characteristics of the epidermis in marine and freshwater finless porpoises adapted to distinct osmotic environments

Abstract The Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis, Pilleri & Gihr, 1972; YFP) is an exclusively freshwater cetacean species inhabiting the Yangtze River and its connecting large lakes. As the primary line of defense in maintaining physiological equilibrium, the epidermis of the porpoise is expected to have undergone structural adaptations due to the shift from the marine to the freshwater environment. This study compared the microstructural and ultrastructural features of the epidermis of YFP and its marine counterpart, the East Asian finless porpoise (Neophocaena asiaeorientalis sunameri, Pilleri & Gihr, 1975; EAFP). Microscopic structural observations and statistical analyses of the epidermal thickness demonstrated no significant differences in the cell structure or distribution between the two porpoise species. However, the epidermis of the YFP contained more abundant stratum basale cells. The outermost lipid stratum corneum exhibited a thinner cell layer with wider neutral lipid droplets to resist the passive entry of water molecules in the hypotonic environment. In contrast, for the EAFP, a more uniformly arranged stratum basale in the epidermis led to denser keratin fibers and robust desmosomes within each epidermal layer at the ultrastructural level. This tight arrangement of cells can reduce transepidermal water loss (TEWL) in an environment with higher osmotic pressure. In conclusion, the 2 finless porpoise species appear to employ different epidermal mechanisms to adapt to their distinct osmotic environments. The YFP appears to possess a “lipid waterproofing” epidermal structure, while the EAFP possesses a “thick and compact water-retaining” epidermal structure to cope with potential water loss.

Culture Conditions Differentially Regulate the Inflammatory Niche and Cellular Phenotype of Tracheo-Bronchial Basal Stem Cells.

Human bronchial epithelial cells (HBECs) derived from the tracheo-bronchial regions of human airways provide an excellent in vitro model for studying pathological mechanisms and evaluating therapeutics in human airway cells. This cell population comprises a mixed population of basal cells (BCs), the predominant stem cell in airways capable of both self-renewal and functional differentiation. Despite their potential for regenerative medicine, BCs exhibit significant phenotypic variability in culture. To investigate how culture conditions influence BC phenotype and function, we expanded three independent BC isolates in three media, airway epithelial cell growth medium (AECGM), dual-SMAD inhibitor (DSI)-enriched AECGM, and Pneumacult Ex plus (PEx+). Extensive RNA sequencing, immune assays and electrical measurements revealed that PEx+ media significantly drove cell proliferation and a broad pro-inflammatory phenotype in BCs. In contrast, BCs expanded in AECGM, displayed increased expression of structural and extracellular matrix components at high passage. Whereas culture in AECGM increased expression of some cytokines at high passage, DSI suppressed inflammation altogether thus implicating TGF-β in BC inflammatory processes. Differentiation capacity declined with time in culture irrespective of expansion media except for PLUNC expressing secretory cells that were elevated at high passage in AECGM and PEx+ suggestive of an immune modulatory role of PLUNC in BCs. These findings underscore the profound impact of media conditions on inflammatory niche and function of in vitro expanded BCs. The broad pro-inflammatory phenotype driven by PEx+ media, in particular, should be considered in the development of cell-based models for airway diseases and therapeutic application.

Specific inhibitory effects of exogenous d-Aspartate on the proliferation of intestinal epithelial cells

d-amino acids have been actively examined since improved analytical techniques revealed their presence in animal bodies. Although D-Asp was identified in mammals earlier than D-Ser, research on D-Asp has lagged behind that on D-Ser, mainly because the target protein of D-Asp remains unknown. To date, the only reported functions of D-Asp are its roles in reproduction and suggested neuromodulatory functions. Since d-amino acids are also present in food, it is important to clarify their effects on gastrointestinal epithelial cells, which are always contacted after ingestion. Therefore, the present study examined the effects of d-amino acids on gastrointestinal tract basal cells. The effects of 11 types of amino acids (Ala, Arg, Asn, Asp, Gln, Glu, Leu, Lys, Pro, Ser, and Val) on the proliferation of three types of gastrointestinal epithelial cells (HGC-27, IEC-6, and Caco-2) were assessed. Although the proliferation of HGC-27 and Caco-2 was not affected by any of the 11 types of L- and d-amino acids, D-Asp inhibited the proliferation of IEC-6, derived from small intestinal epithelial cells, in concentration- and exposure time-dependent manners. The present study also examined uptake transporters, metabolic enzymes, and insulin signaling pathways; however, the mechanisms underlying the inhibitory effects of D-Asp on the proliferation of IEC-6 were not elucidated. A more detailed understanding of these mechanisms may lead to the development of pharmaceuticals as main drugs or formulation materials. Further studies are warranted on the physiological effects of d-amino acids, including D-Asp.

Derivation of human primary prostate epithelial cell lines by differentially targeting the CDKN2A locus along with expression of hTERT

Prostate cancer (PCa) is the most common cancer diagnosed in men worldwide and was the second leading cause of cancer-related deaths in US males in 2022. Prostate cancer also represents the second highest cancer mortality disparity between non-Hispanic blacks and whites. However, there is a relatively small number of prostate normal and cancer cell lines compared to other cancers. To identify the molecular basis of PCa progression, it is important to have prostate epithelial cell (PrEC) lines as karyotypically normal as possible. Our lab recently developed a novel methodology for the rapid and efficient immortalization of normal human PrEC that combines simultaneous CRISPR-directed inactivation of CDKN2A exon 2 (which directs expression of p16INK4A and p14ARF) and ectopic expression of an hTERT transgene. To optimize this methodology to generate immortalized lines with minimal genetic alterations, we sought to target exon 1α of the CDKN2A locus so that p16INK4A expression is ablated while the exons encoding p14ARF remains unaltered. Here we describe the establishment of two cell lines: one with the above-mentioned p16INK4A only loss, and a second line targeting both products in the CDKN2A locus. We characterize the potential lineage origin of these new cell lines along with our previously obtained clones, revealing distinct gene expression signatures. Based on the analyses of protein markers and RNA expression signatures, these cell lines are most closely related to a subpopulation of basal prostatic cells. Given the simplicity of this one-step methodology and the fact that it uses only the minimal genetic alterations necessary for immortalization, it should also be suitable for the establishment of cell lines from primary prostate tumor samples, an urgent need given the limited number of available prostate cancer cell lines.

Insights into epithelial-mesenchymal transition from cystic fibrosis rat models

BackgroundMolecular pathways contributing to Cystic Fibrosis pathogenesis remain poorly understood. Epithelial-mesenchymal transition (EMT) has been recently observed in CF lungs and certain CFTR mutation classes may be more susceptible than others. No investigations of EMT processes in CF animal models have been reported. AimThe aim of this study was to assess the expression of EMT-related markers in Phe508del and knockout (CFTR-KO) rat lung tissue and tracheal-derived basal epithelial stem cells, to determine whether CFTR dysfunction can produce an EMT state. MethodThe expression of EMT-related markers in lung tissue and cultured tracheal basal epithelial stem cells from wildtype (WT), Phe508del, and CFTR-KO rats were assessed using qPCR and Western blots. Cell responses were evaluated in the presence of Rho-associated protein kinase (ROCK) inhibitor Y27632, which blocks EMT-pathways, or after treatment with TGFβ1 to stimulate EMT. ResultsDifferent gene expression profiles were observed between Phe508del and CFTR-KO rat models compared to wild type. There was lower expression of type 1 collagen in KO lungs and primary cell cultures, while Phe508del lungs and cells had higher expression, particularly when treated with TGFβ1. The addition of Y27632 rescued changes in EMT related genes in Phe508del cells but not in KO cells. ConclusionOur findings show the first evidence of upregulated EMT pathways in the lungs and airway cells of any CF animal model. Differences in the regulation of the EMT genes and proteins in the Phe508del and CFTR-KO cells suggest that the signalling pathways underlying EMT are CFTR mutation dependent.

Paxbp1 Is Indispensable for the Maintenance of Epidermal Homeostasis

The mammalian epidermis is a structurally complex tissue that serves critical barrier functions, safeguarding the organism from the external milieu. The development of the epidermis is governed by sophisticated regulatory processes. However, the precise mechanism maintaining epidermal homeostasis remains incompletely elucidated. Recent studies have identified Paxbp1, an evolutionarily conserved protein, as being involved in the developmental regulation of various cells, tissues, and organs. Nonetheless, its role in skin development has not been explored. Here, we report that the targeted deletion of Paxbp1 in epidermal keratinocytes mediated by Keratin14-Cre leads to severe disruption in skin architecture. Mice deficient in Paxbp1 exhibited a substantially reduced epidermal thickness and pronounced separation at the dermo-epidermal junction upon birth. Mechanistically, we demonstrate that the absence of Paxbp1 hinders cellular proliferation, marked by a halt in cell cycle transition, suppressed gene expression of proliferation, and a compromised DNA replication pathway in basal keratinocytes, resulting in the thinning of the skin epidermis. Moreover, molecules and pathways associated with hemidesmosome assembly were impaired in Paxbp1-deficient keratinocytes, culminating in the detachment of the skin epidermal layer. Therefore, our study highlights an indispensable role of Paxbp1 in the maintenance of epidermal homeostasis.

Cell Culture Differentiation and Proliferation Conditions Influence the InVitro Regeneration of the Human Airway Epithelium.

The human airway mucociliary epithelium can be recapitulated invitro using primary cells cultured in an air-liquid interface (ALI), a reliable surrogate to perform pathophysiological studies. As tremendous variations exist among media used for ALI-cultured human airway epithelial cells, the aim of our study was to evaluate the impact of several media (BEGM, PneumaCult, Half & Half, and Clancy) on cell type distribution using single-cell RNA sequencing and imaging. Our work revealed the impact of these media on cell composition, gene expression profile, cell signaling, and epithelial morphology. We found higher proportions of multiciliated cells in PneumaCult-ALI and Half & Half, stronger EGF signaling from basal cells in BEGM-ALI, differential expression of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry factor ACE2, and distinct secretome transcripts depending on the media used. We also established that proliferation in PneumaCult-Ex Plus favored secretory cell fate, showing the key influence of proliferation media on late differentiation epithelial characteristics. Altogether, our data offer a comprehensive repertoire for evaluating the effects of culture conditions on airway epithelial differentiation and will aid in choosing the most relevant medium according to the processes to be investigated, such as cilia, mucus biology, or viral infection. We detail useful parameters that should be explored to document airway epithelial cell fate and morphology.

DNA-PKcs inhibition improves sequential gene insertion of the full-length CFTR cDNA in airway stem cells

Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Although many people with CF (pwCF) are treated using CFTR modulators, some are non-responsive due to their genotype or other uncharacterized reasons. Autologous airway stem cell therapies, in which the CFTR cDNA has been replaced, may enable a durable therapy for all pwCF. Previously, CRISPR-Cas9 with two AAVs was used to sequentially insert two halves of the CFTR cDNA and an enrichment cassette into the CFTR locus. However, the editing efficiency was <10% and required enrichment to restore CFTR function. Further improvement in gene insertion may enhance cell therapy production. To improve CFTR cDNA insertion in human airway basal stem cells (ABCs), we evaluated the use of the small molecules AZD7648 and ART558 which inhibit non-homologous end joining (NHEJ) and micro-homology mediated end joining (MMEJ). Adding AZD7648 alone improved gene insertion by 2-3-fold. Adding both ART558 and AZD7648 improved gene insertion but induced toxicity. ABCs edited in the presence of AZD7648 produced differentiated airway epithelial sheets with restored CFTR function after enrichment. Adding AZD7648 did not increase off-target editing. Further studies are necessary to validate if AZD7648 treatment enriches cells with oncogenic mutations.

Pathological Mechanisms Involved in Epidermolysis Bullosa Simplex: Current Knowledge and Therapeutic Perspectives.

Epidermolysis bullosa (EB) is a clinically and genetically heterogeneous group of mechanobullous diseases characterized by non-scarring blisters and erosions on the skin and mucous membranes upon mechanical trauma. The simplex form (EBS) is characterized by recurrent blister formation within the basal layer of the epidermis. It most often results from dominant mutations in the genes coding for keratin (K) 5 or 14 proteins (KRT5 and KRT14). A disruptive mutation in KRT5 or KRT14 will not only structurally impair the cytoskeleton, but it will also activate a cascade of biochemical mechanisms contributing to EBS. Skin lesions are painful and disfiguring and have a significant impact on life quality. Several gene expression studies were accomplished on mouse model and human keratinocytes to define the gene expression signature of EBS. Several key genes associated with EBS were identified as specific immunological mediators, keratins, and cell junction components. These data deepened the understanding of the EBS pathophysiology and revealed important functional biological processes, particularly inflammation. This review emphasizes the three EBS subtypes caused by dominant mutations on either KRT5 or KRT14 (localized, intermediate, and severe). It aims to summarize current knowledge about the EBS expression profiling pattern and predicted molecular mechanisms involved and to outline progress in therapy.

Human efferent ductules and epididymis display unique cell lineages with motile and primary cilia.

Previous research has illustrated the role of cilia as mechanical and sensory antennae in various organs within the mammalian male reproductive system across different developmental stages. Despite their significance in both organ development and homeostasis, primary cilia in the human male reproductive excurrent duct have been overlooked due to limited access to human specimens. This study aimed to characterize the unique cellular composition of human efferent and epididymal ducts, with a focus on their association with primary cilia. Human efferent ductules/epididymides from five donors aged 32-47 years, were obtained through our local organ transplant program. Cell lineage specificity and primary cilia features were examined by immunofluorescent staining and confocal microscopy in the efferent ductules and the distinct segments of the epididymis. The epithelium of the human efferent duct exhibited estrogen receptor-positive cells with primary cilia, FoxJ1-positive multiciliated cells with numerous motile cilia, and non-ciliated intraepithelial immune cells. Notably, intraluminal macrophages, identified by CD163/CD68 positivity, were observed to engage in sperm phagocytosis. In all three segments of the human epididymis, primary cilia were found on the surface of principal and basal cells. Our research indicates that the human efferent ductules create a distinct environment, characterized by the presence of two types of ciliated cells that are in contact with immune cells. The discovery of sensory primary cilia exposed on the surface of reabsorptive cells in the efferent ductules, as well as on basal and principal cells in the epididymis, lays the foundation for complementary functional studies. This research uncovers novel characteristics exclusive to human efferent ductules and epididymides, providing a basis for exploring innovative approaches to male contraception and infertility treatment.

SP-101, A Novel Adeno-Associated Virus Gene Therapy for the Treatment of Cystic Fibrosis, Mediates Functional Correction of Primary Human Airway Epithelia From Donors with Cystic Fibrosis.

Cystic fibrosis (CF) is caused by mutations in the gene encoding the CF transmembrane conductance regulator (CFTR) protein. Although CF affects multiple organs, lung disease is the main cause of morbidity and mortality, and gene therapy is expected to provide a mutation-agnostic option for treatment. SP-101 is a recombinant adeno-associated virus (AAV) gene therapy vector carrying a human CFTR minigene, hCFTRΔR, and is being investigated as an inhalation treatment for people with CF. To further understand SP-101 activity, in vitro studies were performed in human airway epithelia (HAE) derived from multiple CF and non-CF donors. SP-101 restored CFTR-mediated chloride conductance, measured via Ussing chamber assay, at a multiplicity of infection (MOI) as low as 5E2 in the presence of doxorubicin, a small molecule known to augment AAV transduction. Functional correction of CF HAE increased with increasing MOI and doxorubicin concentration and correlated with increasing cell-associated vector genomes and hCFTRΔR mRNA expression. Tropism studies using a fluorescent reporter vector and single-cell mRNA sequencing of SP-101-mediated hCFTRΔR mRNA demonstrated broad expression in all cell types after apical transduction, including secretory, ciliated, and basal cells. In summary, SP-101, particularly in combination with doxorubicin, shows promise for a novel CF treatment strategy and strongly supports continued development.

Eco-friendly synthesis of Co3O4 nanoparticles using Millettia pinnata towards increased anti-oxidant, anti-biofilm and cytocompatibility properties against biofilm producing bacteria and human pulmonary alveolar basal cells

Biofilm architecture is an emerging threat worldwide due to enhanced antibiotic resistance against existing antibiotics, thus heightening the need to discover the alternative therapies that can eradicate the biofilm colonization. Current study highlights the eco-friendly, non-toxic nature of Co3O4 nanoparticles (NPs) using Millettia pinnata (M. pinnata) leaves extract and their application in anti-oxidants, anti-bacterial, anti-biofilm and cytocompatibilities. The GC-MS and FT-IR spectral data suggested that phytochemical derivatives of M. pinnata leaf extract simultaneously contributed in bio-reduction process and capping agent of Co3O4 NPs, which might be reason for anti-oxidant activities. Further, XRD analysis confirms the polycrystalline behaviour of Co3O4 NPs having hexagonal crystal system. FESEM, HRTEM, EDAX and elemental mapping revealed the formation of agglomerated granular shape along with nanorods on the surface of Co3O4 NPs, which confirms the presence of carbon and oxygen. Additionally, the characteristic band and thermal decomposition of Raman and TGA spectral data confirmed the molecular structure of Co3O4 NPs. Subsequently, both the M. pinnata leaves extract and Co3O4 NPs were found to be excellent source of antioxidants and produced significant DPPH and hydroxyl scavenging activities. Furthermore, the sub-biofilm inhibitory concentration of biosynthesized Co3O4 NPs reduced the biofilm growth in S. aureus and K. pneumoniae by crystal violet assay at increasing concentration. In addition, the membrane integrity of biofilm damages and architecture destruction was evidently proved due to the biosynthesized Co3O4 NPs by confocal laser scanning electron microscope and scanning electron microscope observation. Finally, the cytotoxicity experiment results indicated, biosynthesized Co3O4 NPs has no toxicity against human pulmonary alveolar epithelial cells at maximum concentration. Accordingly, findings of this study supported the utility of safety and efficiency of biosynthesized Co3O4 NPs against biofilm producing bacteria, and deserve as promising future drug discovery target for various bacterial infections.

DERMOSCOPIC FEATURES OF BASAL CELL CARCINOMA: SYSTEMATIC REVIEW

Abstract Background: Basal cell carcinoma is one of the most common cancers worldwide, it is a locally destructive and slowly spreading tumor arising from the epidermis or hair follicles, in which the peripheral cells are similar to the cells of the basal layer of the epidermis and which rarely metastasize. Despite its slow growth and low potential for metastasis, basal cell carcinoma (BCC) is one of the most pressing problems in dermato-oncology due to its high incidence rates, variety of clinical forms and frequent relapses after any treatment methods. Currently, the most accessible non-invasive method for examining skin tumors is dermatoscopy. Aim: The purpose of this review is to highlight the dermoscopic features encountered in basal cell carcinoma and to outline the role of dermoscopy for diagnosis this cancer. Methodology: The systematic review included articles from Google Scholar, Medline, Scopus, Web Of Sciences, PubMed was conducted. For the search, the following keywords were included: dermoscopy of pigmented form of basal cell carcinoma; dermoscopy of nonpigmented form of basal cell carcinoma. Results: Conducted a preliminary search and reviewed 62 titles and abstracts in this review and 27 full-text articles were selected of high methodological quality. Dermoscopy is a non-invasive technique, which allows early diagnosis based on the presence of typical vascular structures, pigmented structures, and ulceration and the absence of specific melanocytic structures. However, the use of dermoscopic technique is limited by the lack of prognostically significant dermatoscopic criteria confirming the different morphological nature of tumor growth. Conclusion: The current systematic review demonstrated, based on the observed features, this method allows for differential diagnosis of malignant and benign, as well as melanocytic and non-melanocytic skin neoplasms, as well as diagnosis of tumors at the earliest stage of their development.

Establishing a Mouse Model of Surgical Vocal Fold Injury.

Animal models of vocal fold (VF) surgical injury and scar formation provide insight into the wound healing process. The purpose of this study was to establish an alternative model of surgical injury to the mouse VF using materials commonly available in most research laboratories or for purchase and to investigate wound healing of the epithelium (EP) and lamina propria (LP). Mice were anesthetized by isoflurane gas delivery and positioned on a platform so that the larynx could be observed using a laryngoscope and dissection microscope. Unilateral VF injury was created using a wire brush. Mice were euthanized and the larynx evaluated 1-, 3-, 5-, 7-, 14-, and 28-days following injury. Histological and immunofluorescent analysis was used to evaluate thickness of the EP, LP area, proliferative (Ki67+) and basal cells (p63+) in the EP, and collagen III content in the LP. The depth of injury reached the superficial thyroarytenoid muscle on Day 1. The thickness of the EP of the injured VF was increased on Days 3 and 5, and the LP area was increased on Days 3, 5, and 7 as compared with the uninjured VF. Ki67+ and p63+ cells were increased on Day 3 and collagen III content was increased on Days 5 and 28 as compared with the uninjured VF. We successfully established an alternative method of creating unilateral VF injury in the mouse. This method will be useful for future research regarding VF surgical injury and wound healing. N/A Laryngoscope, 2024.

First Report of Leaf Blight of Ginkgo biloba L. Caused by Neopestalotiopsis clavispora G.F. Atk. in Sichuan, China.

Ginkgo (Ginkgo biloba L.), the oldest existing tree species in the world, is an important ornamental and medicinal plant, widely planted in China. In October 2022, a new leaf blight disease was observed in Chengdu city (30°05'to 31°26'N, 102°54'to 104°53'E). Disease incidence averaged 82.5% across five foci. The typical symptomatology begins when leaf margins turn yellow and small black spots appear at the edge of the leaf, chlorotic areas turn brown, dry and deformed. Gradually, the necrotic lesions spreads to the middle of the leaf and eventually the whole leaf falls off. Infected tissues from ten leaves were cut into small pieces (5 × 5 mm); surface sterilized for 30 s in 3% sodium hypochlorite; 60 s in 75% ethanol; rinsed three times in sterile water; placed onto potato dextrose agar (PDA) amended with streptomycin sulfate (50 μg/mL); and incubated at 25°C for 3 to 8 days. A hyphae was removed from the edge of the fungal colony and placed onto potato dextrose agar (PDA) plates. After incubation at 25℃ with a 12-hour light/dark cycle for 8 days, the colony diameter reached 77.5 to 81.5 mm. Colonies grown on PDA were white, cotton, flocculent, undulating on the surface, dense in aerial hyphae and light yellow on the back. Black pycnidia formed superficially, scattered over the PDA, following two weeks of incubation. Pycnidia contained sticky black conidia. The spores were were spindle shaped, with five cells, and four septations measuring 20.9 to 34.8 µm × 6.8 to 8.8 µm (avg. 28.4 × 7.6 µm; n=40). The three median cells were versicolored, typically two dark brown cells and one light brown cell, whereas the basal and apical cells were hyaline. Conidia had a single basal appendage (2.87 to 4.1 µm long; n = 40) and two to three apical appendages (18.3 to 29.1 µm long; n = 40). Based on colony and conidial morphology, the isolate was identified as N. clavispora (Maharachchikumbura et al. 2014). The partial sequence of the internal transcribed spacer (ITS), β-tubulin gene (TUB2), and translation elongation factor subunit 1-a gene (TEF1) were amplified and sequenced using the universal primer pairs ITS1/ITS4(Zhang et al. 2022), BT2A/BT2B (Li Yuan et al. 2022), and EF1-526F/EF1-1567R (Maharachchikumbura et al. 2012), respectively. Sequences of representative isolate LQYX were deposited in GenBank (ITS: OQ152504, TUB: OQ168328, and TEF1: OQ168329). BLAST results indicated that the ITS, TUB, and TEF1-α sequences showed 99 to 100% identity with N. clavispora sequences at NCBI (GenBank MG729689, MG740735, and MG740758). Identification was confirmed by Bayesian inference using Mr. Bayes. Next, inoculations were conducted on leaves of ten G. biloba in the field to verify the pathogenicity of LQYX. Ten healthy leaves of each plant were surface sterilized with 75% ethanol, and the wound was rubbed out on the leaf edge on the sterilized sanding paper. A conidia suspension (1 × 107 ml-1) was sprayed on the leaves, aseptic water was used as the control, and the transparent plastic bag was used to maintain relative humidity. After 14 days (26 ℃, 14 hours light / 10 hours dark), the inoculated leaves had similar symptoms as the original diseased plants, whereas controls were asymptomatic. The N. clavispora was re-isolated from the infected leaves and identified by morphological characteristics and DNA sequence analysis. The pathogenicity test was repeated three times with similar results, confirming Koch's postulates. To our knowledge, this is the first report of leaf blight of G. biloba caused by N. clavispora in China, which has greatly affected the appearance of the city and should be further studied. This report can help identify this disease and further develop effective control measures.

Skin Absorbed Dose Coefficients for Human Legs from Beta Radiation as a Function of Height

External exposure to skin from beta-emitter radionuclides following severe reactor accidents or nuclear testing can result in beta burning and other health complications. The skin absorbed dose coefficient (SADC) measures the energy deposition into the skin during such accidents. The U.S. Environmental Protection Agency has published several reports to measure the possible energy deposition into the skin in such accidents. However, the most recent SADC published by Federal Guidance Report (FGR) 12 was computed only at one meter above the contaminated surface. Therefore, it was necessary to develop a model to estimate the absorbed dose coefficients for skin at different heights. In this manuscript, Geant4, a Monte Carlo simulator toolkit, was used to estimate the absorbed dose coefficients from electron sources located on the soil surface with energies ranging from 0.1 to 4 MeV. The energy deposited from primary electrons, secondary electrons, and photons in a 50 µm thick layer of epidermis tissue (Basal Cells Layer) located at a depth of 50 µm from the skin surface was estimated at several discrete heights of human leg phantom. More than 40% of the total energy deposited comes from secondary electrons and photons in energy sources of 0.1 and 0.2 MeV on average, but for higher energies, this percentage is less than 1%, which indicates primary electrons are the main source of the deposited energy in the skin. Furthermore, the results showed the energy deposited into skin closer to the ground was 50–100% higher than the previously estimated doses for 1 m above the ground. The results from Geant4 showed a great correlation (R2 = 0.972) with the FGR 12 data at one meter height, and they were aligned with the published values from FGR 12, which validated the simulation results. Therefore, the calculated dose coefficients for different energy sources and different heights could be used in radiation protection measurements.

Cowper Glands Identified in Prostate and Urethral Specimens: A Comprehensive Immunohistochemical Characterization and Potential Diagnostic Pitfall.

Cowper glands recognition remains one of the key histoanatomic benign mimics of prostatic adenocarcinoma. In most instances, these can be identified based on the dimorphic population of lobulated acini and duct(s). However, in the prostate biopsy setting with incomplete/distorted cores, this may not be immediately apparent and may warrant use of immunohistochemistry to argue against prostatic adenocarcinoma. Although immunohistochemical pitfalls in Cowper glands have been described, to our knowledge a comprehensive evaluation of both traditional and purportedly prostate-specific novel markers in Cowper glands has not been previously performed. Herein, we studied the clinicopathological and immunohistochemical features of 21 male patients (age range 39-81 years; mean = 63 years), including 15 prostate biopsies (7 of which also had prostate cancer in the same specimen set and 2 of which had both prostate cancer and Cowper glands in the same biopsy core). Immunohistochemistry showed the following results in Cowper glands: 100% positive for NKX3.1, 100% positive (basal cells) for both high molecular weight keratin and p63, 57% positive for PSAP, 25% positive for PSMA, 5% positive for AMACR, and 0% positive for PSA. In conclusion, for specimens lacking appreciable dimorphic morphology, caution should be rendered when using prostate-specific markers (PSA, PSAP, PSMA, and NKX3.1) as these can show considerable staining in Cowper glands and be a pitfall. Instead, findings from this cohort indicate relying on basal markers (high molecular weight keratin/p63; either individually or in a "cocktail" approach) and PSA are most useful in distinguishing Cowper glands (retained basal cell markers staining) from prostatic adenocarcinoma.

Changes of epidermal proteins immunolocalization in the corneous layer from embryonic to definitive avian beak

The process of keratinization and cornification in the developing beak has been studied through immunofluorescence and immunogold electron microscopy in chick and zebrafinch embryos. After the curved beak anlagen appears at the tip of the maxillar bone, 5–8 layers of embryonic epidermis are generated from the basal layer of the epidermis. These cells are weakly immunoabeled for IFKs (Intermediate Filament Keratins) and more intensely for scaffoldin, a protein of the EDC (Epidermal Differentiation Complex) involved in the soft keratinization of the embryonic epidermis. Immunolabeling for CBPs (Corneous Beta Proteins) is visible in the transitional embryonic layers that are temporarily generated between the embryonic and definitive beak epidermis. The electron microscope reveals that intermediate layers contain immunolabeled periderm granules for scaffoldin mixed with bundles of corneous material immunolabeled for CBPs. Intense CBPs labeling occurs in the compacting corneous bundles of beta-keratinocytes in the definitive beak while scaffolding labeling disappears. The embryonic epidermis is sloughed before hatching. Sox (Sulfhydryl Oxidase) immunolabeling reveals that the enzyme is almost absent in embryonic layers but is present in transitional and definitive beta-keratinocytes. This indicates the formation of cross-linked disulfide bonds in the definitive corneous layer of the beak. Some calcium precipitation, suggested from von Kossa staining, occurs in the corneous layers only on the 18th day of development in the chick, in preparation for hatching.

Dynamic regulation of tissue fluidity controls skin repair during wound healing

During wound healing, different pools of stem cells (SCs) contribute to skin repair. However, how SCs become activated and drive the tissue remodeling essential for skin repair is still poorly understood. Here, by developing a mouse model allowing lineage tracing and basal cell lineage ablation, we monitor SC fate and tissue dynamics during regeneration using confocal and intravital imaging. Analysis of basal cell rearrangements shows dynamic transitions from a solid-like homeostatic state to a fluid-like state allowing tissue remodeling during repair, as predicted by a minimal mathematical modeling of the spatiotemporal dynamics and fate behavior of basal cells. The basal cell layer progressively returns to a solid-like state with re-epithelialization. Bulk, single-cell RNA, and epigenetic profiling of SCs, together with functional experiments, uncover a common regenerative state regulated by the EGFR/AP1 axis activated during tissue fluidization that is essential for skin SC activation and tissue repair.