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ABSTRACT:
PROVIDER: PRJNA930146 | ENA |
REPOSITORIES: ENA
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Project description:To study the development and composition of human ocular surface, we performed single cell (sc) RNA-Seq at key embryonic, fetal and adult stages and generated the first atlas of the corneal and conjunctival cell types from development to adulthood. The conjunctival epithelium is the first to be specified in the epithelial layer , followed by the corneal epithelium and the establishment of proliferative epithelial progenitors, which predate the formation of limbal niche by a few weeks. Bioinformatic comparison of adult cell clusters identified GPHA2,a novel cell-surface marker for quiescent limbal stem cells (qLSCs), whose function is to maintain qLSCs self-renewal. Combining scRNA- and ATAC-Seq analysis, we identified multiple upstream regulators for qLSCs and transit amplifying (TA) cells and demonstrated a close interaction between the immune cells and epithelial stem and progenitor cells in the cornea. Single cell RNA-Seq analysis indicated loss of qLSCs and acquisition of proliferative limbal epithelial progenitor markers during limbal epithelial cell expansion: this phenomenon was independent of culture method used. Extending the single cell analyses to Keratoconus, we were able to reveal activation of collagenase in the corneal stroma and a reduced pool of TA cells in the corneal epithelium as two key mechanistic changes underlying the disease phenotype. Our scRNA-Seq data comparisons of developing and adult cornea and conjunctiva provide a unique resource for defining pathways/genes that could lead to improvement in ex vivo expansion methods for cell based replacement therapies and better understanding and treatment of ocular surface disorders.
2021-04-19 | GSE155683 | GEO
Project description:Side population (SP) cells isolated from limbal and conjunctival epithelia derive from cells that are slow cycling in vivo, a known feature of tissue stem cells. The purpose of this study was to define the molecular signature of the conjunctival side population cells by global differential gene expression to identify markers and signaling pathways associated with this cell phenotype. Four overnight cultures of freshly isolated human conjunctival epithelial cells stained with Hoechst 3342 were cytometrically sorted into SP and non-SP cohorts. RNA was isolated and processed for microarray analysis using a commercial oligonucleotide array representing more than 55,000 transcripts derived from about 30,000 different genes. Selected microarray results were validated at the gene and protein levels by quantitative PCR, and immunological methods. Data mining methods were used to identify cellular processes relevant for stem cell function. Comparative analysis of transcripts expression based on expression levels and present/absent software calls across 4 replicate experiments identified 16993 expressed conjunctival epithelial transcripts including 10,266 unique known genes. Of those genes, 1254 and 363 were over expressed (> 2-fold ) or under expressed (< 0.5-fold), respectively, in the SP. The overexpressed set included genes coding for non-epithelial genes (e.g., CD62E/E-selectin and CD93), genes that have been associated with stem cell function in other cellular systems, including several homeodomain genes and genes whose over- or under-expression may underpin the stem cell slow cycling phenotype ( e.g., dual specificity phosphatases and cyclin kinases).
2009-10-30 | GSE12631 | GEO
Project description:We developed a compartmental model of the small intestinal epithelium that describes stem and progenitor cell proliferation and differentiation and cell migration onto the villus. The model includes a negative feedback loop from villus cells to regulate crypt proliferation and integrates heterogeneous epithelial-related processes, such as the transcriptional profile, citrulline kinetics and probability of diarrhea.
2023-09-04 | MODEL2212120003 | BioModels
Project description:Side population (SP) cells isolated from limbal and conjunctival epithelia derive from cells that are slow cycling in vivo, a known feature of tissue stem cells. The purpose of this study was to define the molecular signature of the conjunctival side population cells by global differential gene expression to identify markers and signaling pathways associated with this cell phenotype. Four overnight cultures of freshly isolated human conjunctival epithelial cells stained with Hoechst 3342 were cytometrically sorted into SP and non-SP cohorts. RNA was isolated and processed for microarray analysis using a commercial oligonucleotide array representing more than 55,000 transcripts derived from about 30,000 different genes. Selected microarray results were validated at the gene and protein levels by quantitative PCR, and immunological methods. Data mining methods were used to identify cellular processes relevant for stem cell function. Comparative analysis of transcripts expression based on expression levels and present/absent software calls across 4 replicate experiments identified 16993 expressed conjunctival epithelial transcripts including 10,266 unique known genes. Of those genes, 1254 and 363 were over expressed (> 2-fold ) or under expressed (< 0.5-fold), respectively, in the SP. The overexpressed set included genes coding for non-epithelial genes (e.g., CD62E/E-selectin and CD93), genes that have been associated with stem cell function in other cellular systems, including several homeodomain genes and genes whose over- or under-expression may underpin the stem cell slow cycling phenotype ( e.g., dual specificity phosphatases and cyclin kinases). Experiment Overall Design: Whole human conjunctivae from unidentifiable cadaver donors, aged 55 to 65, were obtained from the National Disease Research Interchange (NDRI, Philadelphia, PA) within 48 hours of collection. No donor details apart from age, sex, and cause of death were released. The use of human tissue in the study was in accordance with the provisions of the Declaration of Helsinki and sanctioned by the Institutional Review Board. Fresh rabbit tissue was obtained from local abattoirs within one hour of sacrifice. Unless stated otherwise all reagents were procured from Sigma (St. Louis, Mo). Experiment Overall Design: Eight microarray experiments using side population (Hoechst 33342-transporting cells; SP) or non side population (Hoechst 33342-stained cells; nSP) cells from four cadavers.
2009-11-11 | E-GEOD-12631 | biostudies-arrayexpress
Project description:The identification of a marker that is expressed in the conjunctival epithelium but not in the corneal epithelium has been a growing need. A more specific marker of limbal and conjunctival epithelia would be necessary to detect non-corneal epithelial cells on the corneal surface. To search for conjunctival specific marker(s), we first performed preferential gene profiling in the conjunctiva in direct comparison to that in the cornea using microarray technique.
2011-06-09 | GSE29402 | GEO
Project description:Although tropism of SARS-CoV-2 for respiratory tract epithelial cells is well established, an open question is whether the conjunctival epithelium is also a target for SARS-CoV-2. Conjunctival epithelial cells, which express viral entry receptors ACE2 and TMPRSS2, constitute the largest exposed epithelium of the ocular surface tissue, and may represent a relevant viral entry route. To address this question, we generated an organotypic air-liquid-interface model of conjunctival epithelium, composed of progenitor, basal and superficial epithelial cells and fibroblasts, which could be maintained successfully up to day 75 of differentiation. Using single cell RNA-Seq, with complementary imaging and virological assays, we observed that while all conjunctival cell types were permissive to SARS-CoV-2 genome expression, a productive infection did not ensure. The early innate immune response to SARS-CoV-2 infection in conjunctival cells was characterised by a robust NF-Kβ activity, alongside evidence of suppression of antiviral interferon signalling. Collectively these data enrich our understanding of SARS-CoV-2 infection at the human ocular surface, with potential implications for the design of preventive strategies such as personal protective equipment.
2022-06-06 | GSE191232 | GEO
Project description:Immortalized colonic epithelial progenitor cells express stem cell markers and differentiate in vitro
| PRJNA118233 | ENA
Project description:There is increasing evidence that breast and other cancers originate from and are maintained by a small fraction of stem/progenitor cells with self-renewal properties. Whether such cancer stem/progenitor cells originate from normal stem cells based on initiation of a de novo stem cell program, by reprogramming of a more differentiated cell type by oncogenic insults or both remains unresolved. A major hurdle in addressing these issues is lack of immortal human stem/progenitor cells that can be deliberately manipulated in vitro. We present evidence that normal and human telomerase reverse transcriptase (hTERT)-immortalized human mammary epithelial cells (hMECs) isolated and maintained in DFCI-1 medium retain a fraction with progenitor cell properties. These cells co-express basal, luminal and stem/progenitor cell markers. Clonal derivatives of progenitors co-expressing these markers fall into two distinct types: K5+/K19- (Type I) and K5+/K19+ (Type II). We show that both types of progenitor cells have self-renewal and differentiation ability. Through microarray analysis, we want to identify genes and pathways linked to human mammary epithelial stem/progenitor cell self-renewal and differentiation. Normal human mammary epithelial cells (hMECs) were isolated from Reduction Mammoplasty and immortalized by hTERT. Type I K5+/K19- and Type II K5+/K19+ cell colonies were isolated from hTERT-immortalized hMECs and cultured in MEGM medium for self-renewal and differentiation. Total RNA isolated from Type I, Type II, and differentiated Myoepithelial (Myo) cells were used on Affymetrix microarrays.
2010-07-29 | E-GEOD-22580 | biostudies-arrayexpress
Project description:The identification of a marker that is expressed in the conjunctival epithelium but not in the corneal epithelium has been a growing need. A more specific marker of limbal and conjunctival epithelia would be necessary to detect non-corneal epithelial cells on the corneal surface. To search for conjunctival specific marker(s), we first performed preferential gene profiling in the conjunctiva in direct comparison to that in the cornea using microarray technique. The study consisted of gene expression profiles comparing the cornea and conjunctiva.
2011-06-09 | E-GEOD-29402 | biostudies-arrayexpress
Project description:Genome-wide analysis of dihydrotestosterone (DHT) induced changes in gene expression in immortalized human conjunctival epithelial cells. Analysis of regulation of immortalized human conjunctival epithelial cells by dihydrotestosterone at gene expression level. The hypothesis tested in the present study was that the androgen-eye interaction in ocular surface epithelial cells like conjunctival cells is influenced by androgens through regulation of the expression of multiple genes. Results provide important information of the differential regulation of numerous genes in response to dihydrotestosterone incubation of immortalized human conjunctival epithelial cells. Total RNA was obtained from immortalized human conjunctival epithelial cells treated for 96 hours with 10 nM dihydrotestosterone (n=3) or vehicle (n=3). The RNA was then used with Illumina HumanHT-12 v3 Expression BeadChips to determine the effect of DHT on gene expression in an immortalized human conjunctival epithelial cell line developed in Dr. Rheinwald's laboratory [Rheinwald et al. MCB, 22 (14): 5157. (2002)] and charecterized in Dr. Ilene Gibson's laboratory [Gipson et al. IOVS, 44 (6): 2496. (2003)].
2011-03-31 | E-GEOD-18094 | biostudies-arrayexpress