ABSTRACT
Mammalian epidermal stem cells maintain homeostasis of the skin epidermis and contribute to its regeneration throughout adult life. While 2D mouse epidermal stem cell cultures have been established decades ago, a long-term, feeder cell- and serum-free culture system recapitulating murine epidermal architecture has not been available. Here we describe an epidermal organoid culture system that allows long-term, genetically stable expansion of adult epidermal stem cells. Our epidermal expansion media combines atypically high calcium concentrations, activation of cAMP, FGF, and R-spondin signaling with inhibition of bone morphogenetic protein (BMP) signaling. Organoids are established robustly from adult mouse skin and expand over at least 6 mo, while maintaining the basal-apical organization of the mouse interfollicular epidermis. The system represents a powerful tool to study epidermal homeostasis and disease in vitro.
Subject(s)
Adult Stem Cells/physiology , Cell Culture Techniques/methods , Epidermis/physiology , Keratinocytes/physiology , Organoids/physiology , Animals , Cell Differentiation , Cell Proliferation , Cells, Cultured , Gene Knock-In Techniques , Mice , Mice, Inbred C57BL , Mice, Transgenic , Receptors, G-Protein-Coupled/genetics , Time FactorsABSTRACT
Organoids are self-organizing 3D structures grown from stem cells that recapitulate essential aspects of organ structure and function. Here, we describe a method to establish long-term-expanding human airway organoids from broncho-alveolar resections or lavage material. The pseudostratified airway organoids consist of basal cells, functional multi-ciliated cells, mucus-producing secretory cells, and CC10-secreting club cells. Airway organoids derived from cystic fibrosis (CF) patients allow assessment of CFTR function in an organoid swelling assay. Organoids established from lung cancer resections and metastasis biopsies retain tumor histopathology as well as cancer gene mutations and are amenable to drug screening. Respiratory syncytial virus (RSV) infection recapitulates central disease features, dramatically increases organoid cell motility via the non-structural viral NS2 protein, and preferentially recruits neutrophils upon co-culturing. We conclude that human airway organoids represent versatile models for the in vitro study of hereditary, malignant, and infectious pulmonary disease.
Subject(s)
Carcinoma, Non-Small-Cell Lung/pathology , Cystic Fibrosis/pathology , Epithelial Cells/pathology , Organ Culture Techniques/methods , Organoids/pathology , Respiratory Syncytial Virus Infections/pathology , Respiratory System/pathology , Animals , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/metabolism , Cells, Cultured , Cystic Fibrosis/metabolism , Cystic Fibrosis Transmembrane Conductance Regulator/metabolism , Disease Models, Animal , Drug Screening Assays, Antitumor , Epithelial Cells/metabolism , Female , Humans , Lung Neoplasms/drug therapy , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Male , Mice , Mice, Inbred NOD , Mice, SCID , Organoids/metabolism , Respiratory Syncytial Virus Infections/virology , Respiratory Syncytial Viruses/isolation & purification , Respiratory System/metabolism , Xenograft Model Antitumor AssaysABSTRACT
BACKGROUND: This study was purposed to evaluate palatal width, height, and height index at various stages of dentition in Iranian children and adolescent with normal occlusion. METHODS: This cross-sectional study was conducted on a random sample of 237 children (45% male and 55% female, aged 5-18 years) with normal occlusion selected from kindergartens and elementary and high schools in Hamadan, Iran. The subjects were clinically examined and classified based on dentition to primary (21.5%), mixed (21.9%), and permanent (56.5%) stages. Dental casts were obtained from all subjects. Palatal width (inter-molar and -canine distances), and height (at molar and canine areas) were measured on the casts by Korkhaus' compass and digital caliper. Palatal height index was calculated for each dentition stage. Data were analyzed by SPSS 15 using one-way ANOVA followed by Tukey's post hoc test and t- test (p < 0.05). RESULTS: Palatal inter-molar and -canine width values were increased from primary to permanent dentition. Palatal height and palatal height index in mixed dentition were significantly lower than those in primary and permanent dentition. Palatal width at inter-molar and -canine distances was significantly higher in males than females. There was no significant difference in palatal height and palatal height index at molar area between males and females. However, palatal height and palatal height index at canine area were significantly higher in males compared to females. CONCLUSION: These findings showed that palatal width increased from primary to permanent stage. Palatal height and palatal height index decrease from primary to mixed dentation, then increase from mixed to permanent dentition.
