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Putative stem cell markers in limbal epithelial cells cultured on intact & denuded human amniotic membrane. 2008

Balasubramanian Sudha, and Guruswamy Sitalakshmi, and Geetha Krishnan Iyer, and Subramanian Krishnakumar
L&T Department of Ocular Pathology, Vision Research Foundation, Chennai, India.

OBJECTIVE The ocular surface is an ideal region to study the epithelial stem cell (SC) biology because of the unique spatial arrangement of stem cells and transient amplifying cells. A major challenge in corneal SC biology is the ability to identify SC in vitro and in situ, and one of the major controversies in the field relates to reliable SC markers. This study was carried out to evaluate and compare the expression of the stem cell associated marker: ABCG2, keratinocyte stem cell marker: p63 and corneal differentiation markers: Cnx43 and K3/K12 on limbal explants cultured on human amniotic membrane (HAM) with intact epithelium and HAM denuded of its epithelium. METHODS Human limbal biopsies obtained from the cadaveric donor eyes were used in this study. The cells were cultured over the HAM with intact and denuded epithelium. Reverse transcriptase PCR, immunohistochemistry, Western blotting for ABCG2, P63, Cnx43 and K3/K12 were done. RESULTS The limbal epithelial cells cultured over intact HAM expressed the stem cell associated markers (ABCG2, p63) and showed reduced expression of the differentiation markers (Cnx43 and K3/K12) when compared to limbal epithelial cells cultured over denuded HAM, which expressed more differentiation markers at the end of three weeks. BrdU label retaining cells were observed in the limbal epithelial cells cultured over HAM with epihelium only. CONCLUSIONS Our results showed that the intact HAM supported the growth of limbal epithelial cells expressing stem cell associated markers, and allowing little differentiation of the limbal cells to cornea phenotype. Further studies are needed to understand the properties of the amniotic epithelium that retains the stemness in the cultured limbal stem cells.

UI MeSH Term Description Entries
D009363 Neoplasm Proteins Proteins whose abnormal expression (gain or loss) are associated with the development, growth, or progression of NEOPLASMS. Some neoplasm proteins are tumor antigens (ANTIGENS, NEOPLASM), i.e. they induce an immune reaction to their tumor. Many neoplasm proteins have been characterized and are used as tumor markers (BIOMARKERS, TUMOR) when they are detectable in cells and body fluids as monitors for the presence or growth of tumors. Abnormal expression of ONCOGENE PROTEINS is involved in neoplastic transformation, whereas the loss of expression of TUMOR SUPPRESSOR PROTEINS is involved with the loss of growth control and progression of the neoplasm. Proteins, Neoplasm
D001973 Bromodeoxyuridine A nucleoside that substitutes for thymidine in DNA and thus acts as an antimetabolite. It causes breaks in chromosomes and has been proposed as an antiviral and antineoplastic agent. It has been given orphan drug status for use in the treatment of primary brain tumors. BUdR,BrdU,Bromouracil Deoxyriboside,Broxuridine,5-Bromo-2'-deoxyuridine,5-Bromodeoxyuridine,NSC-38297,5 Bromo 2' deoxyuridine,5 Bromodeoxyuridine,Deoxyriboside, Bromouracil
D003315 Cornea The transparent anterior portion of the fibrous coat of the eye consisting of five layers: stratified squamous CORNEAL EPITHELIUM; BOWMAN MEMBRANE; CORNEAL STROMA; DESCEMET MEMBRANE; and mesenchymal CORNEAL ENDOTHELIUM. It serves as the first refracting medium of the eye. It is structurally continuous with the SCLERA, avascular, receiving its nourishment by permeation through spaces between the lamellae, and is innervated by the ophthalmic division of the TRIGEMINAL NERVE via the ciliary nerves and those of the surrounding conjunctiva which together form plexuses. (Cline et al., Dictionary of Visual Science, 4th ed) Corneas
D004847 Epithelial Cells Cells that line the inner and outer surfaces of the body by forming cellular layers (EPITHELIUM) or masses. Epithelial cells lining the SKIN; the MOUTH; the NOSE; and the ANAL CANAL derive from ectoderm; those lining the RESPIRATORY SYSTEM and the DIGESTIVE SYSTEM derive from endoderm; others (CARDIOVASCULAR SYSTEM and LYMPHATIC SYSTEM) derive from mesoderm. Epithelial cells can be classified mainly by cell shape and function into squamous, glandular and transitional epithelial cells. Adenomatous Epithelial Cells,Columnar Glandular Epithelial Cells,Cuboidal Glandular Epithelial Cells,Glandular Epithelial Cells,Squamous Cells,Squamous Epithelial Cells,Transitional Epithelial Cells,Adenomatous Epithelial Cell,Cell, Adenomatous Epithelial,Cell, Epithelial,Cell, Glandular Epithelial,Cell, Squamous,Cell, Squamous Epithelial,Cell, Transitional Epithelial,Cells, Adenomatous Epithelial,Cells, Epithelial,Cells, Glandular Epithelial,Cells, Squamous,Cells, Squamous Epithelial,Cells, Transitional Epithelial,Epithelial Cell,Epithelial Cell, Adenomatous,Epithelial Cell, Glandular,Epithelial Cell, Squamous,Epithelial Cell, Transitional,Epithelial Cells, Adenomatous,Epithelial Cells, Glandular,Epithelial Cells, Squamous,Epithelial Cells, Transitional,Glandular Epithelial Cell,Squamous Cell,Squamous Epithelial Cell,Transitional Epithelial Cell
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000070997 ATP Binding Cassette Transporter, Subfamily G, Member 2 ATP-binding cassette transporter, sub-family G protein that functions as a high capacity UREA exporter, transporter of STEROLS, and in the absorption and efflux of many drugs. Its efflux activity for ANTINEOPLASTIC AGENTS contributes to DRUG RESISTANCE. It functions as a homodimer and is expressed by cells in a variety of organs, as well as by NEOPLASTIC STEM CELLS. ABCG2 Protein,ABCG2 Transporter,ATP Binding Cassette Transporter, Sub-Family G, Member 2,CD338 Antigen
D000650 Amnion The innermost membranous sac that surrounds and protects the developing embryo which is bathed in the AMNIOTIC FLUID. Amnion cells are secretory EPITHELIAL CELLS and contribute to the amniotic fluid. Amniotic Membrane,Amnions,Amniotic Membranes,Membrane, Amniotic,Membranes, Amniotic
D000943 Antigens, Differentiation Antigens expressed primarily on the membranes of living cells during sequential stages of maturation and differentiation. As immunologic markers they have high organ and tissue specificity and are useful as probes in studies of normal cell development as well as neoplastic transformation. Differentiation Antigen,Differentiation Antigens,Differentiation Antigens, Hairy Cell Leukemia,Differentiation Marker,Differentiation Markers,Leu Antigen,Leu Antigens,Marker Antigen,Marker Antigens,Markers, Differentiation,Antigen, Differentiation,Antigen, Leu,Antigen, Marker,Antigens, Leu,Antigens, Marker,Marker, Differentiation
D013234 Stem Cells Relatively undifferentiated cells that retain the ability to divide and proliferate throughout postnatal life to provide progenitor cells that can differentiate into specialized cells. Colony-Forming Units,Mother Cells,Progenitor Cells,Colony-Forming Unit,Cell, Mother,Cell, Progenitor,Cell, Stem,Cells, Mother,Cells, Progenitor,Cells, Stem,Colony Forming Unit,Colony Forming Units,Mother Cell,Progenitor Cell,Stem Cell
D015153 Blotting, Western Identification of proteins or peptides that have been electrophoretically separated by blot transferring from the electrophoresis gel to strips of nitrocellulose paper, followed by labeling with antibody probes. Immunoblotting, Western,Western Blotting,Western Immunoblotting,Blot, Western,Immunoblot, Western,Western Blot,Western Immunoblot,Blots, Western,Blottings, Western,Immunoblots, Western,Immunoblottings, Western,Western Blots,Western Blottings,Western Immunoblots,Western Immunoblottings

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