title: HC-HA/PTX3 Purified From Amniotic Membrane Promotes BMP Signaling in Limbal Niche Cells to Maintain Quiescence of Limbal Epithelial Progenitor/Stem Cells.
abstract: To explore how limbal niche cells (LNCs) may control quiescence, self-renewal, and corneal epithelial lineage commitment/differentiation of limbal epithelial progenitor/stem cells (LEPCs), we have established an in vitro sphere assay by reunion between the two cell types in three-dimensional Matrigel. The resultant sphere exhibits inhibition of corneal epithelial lineage commitment/differentiation and marked clonal growth of LEPCs, of which the latter is correlated with activation of canonical Wnt signaling. Herein, we have created a similar reunion assay in immobilized heavy chain-hyaluronic acid/pentraxin 3 (HC-HA/PTX3), which is purified from amniotic membrane (AM) and consists of a complex formed by hyaluronic covalently linked to heavy chain 1 of inter-α-inhibitor and noncovalently linked to pentraxin 3. The resultant spheres exhibited similar suppression of corneal epithelial lineage commitment/differentiation but upregulation of quiescence markers including nuclear translocation of Bmi-1, and negligible clonal growth of LEPCs. This outcome was correlated with the suppression of canonical Wnt but activation of noncanonical (Planar cell polarity) Wnt signaling as well as BMP signaling in both LEPCs and LNCs. The activation of BMP signaling in LNCs was pivotal because nuclear translocation of pSmad1/5/8 was prohibited in hLEPCs when reunioned with mLNCs of conditionally deleted Bmpr1a;Acvr1(DCKO) mice. Furthermore, ablation of BMP signaling in LEPCs led to upregulation of cell cycle genes, downregulation of Bmi-1, nuclear exclusion of phosphorylated Bmi-1, and marked promotion of the clonal growth of LEPCs. Hence, HC-HA/PTX3 uniquely upregulates BMP signaling in LNCs which leads to BMP signaling in LEPCs to achieve quiescence, helping explain how AM transplantation is clinically useful to be used as a matrix for ex vivo expansion of LEPCs and to treat corneal blindness caused by limbal stem cells deficiency.
authors.affiliation: R&D Department, TissueTech, Inc., Miami, Florida, USA.
authors.forename: Szu-Yu
authors.initials: SY
authors.lastname: Chen
authors.affiliation: Ocular Surface Research & Education Foundation, Miami, Florida, USA.|Department of Ophthalmology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Bo
authors.initials: B
authors.lastname: Han
authors.affiliation: R&D Department, TissueTech, Inc., Miami, Florida, USA.
authors.forename: Ying-Ting
authors.initials: YT
authors.lastname: Zhu
authors.affiliation: R&D Department, TissueTech, Inc., Miami, Florida, USA.
authors.forename: Megha
authors.initials: M
authors.lastname: Mahabole
authors.affiliation: Department of Ophthalmology and Visual Sciences, Washington University, St. Louis, Missouri, USA.
authors.forename: Jie
authors.initials: J
authors.lastname: Huang
authors.affiliation: Department of Ophthalmology and Visual Sciences, Washington University, St. Louis, Missouri, USA.
authors.forename: David C
authors.initials: DC
authors.lastname: Beebe
authors.affiliation: R&D Department, TissueTech, Inc., Miami, Florida, USA.|Ocular Surface Research & Education Foundation, Miami, Florida, USA.
authors.forename: Scheffer C G
authors.initials: SC
authors.lastname: Tseng
mesh_terms: D016475:3T3 Cells; D000650:Amnion / Q000378:metabolism*; D000818:Animals; D019485:Bone Morphogenetic Proteins / Q000096:biosynthesis*; D002097:C-Reactive Protein / Q000096:biosynthesis* / Q000302:isolation & purification; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000378:metabolism*; D006801:Humans; D006820:Hyaluronic Acid / Q000096:biosynthesis* / Q000302:isolation & purification; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis / Q000302:isolation & purification; D051379:Mice; D008822:Mice, Transgenic; D000683:Serum Amyloid P-Component / Q000096:biosynthesis* / Q000302:isolation & purification; D015398:Signal Transduction / Q000502:physiology; D055153:Stem Cell Niche / Q000502:physiology*; D013234:Stem Cells / Q000378:metabolism; D000099153:Pentraxins
chemical_list: D019485:Bone Morphogenetic Proteins; D002097:C-Reactive Protein; D006820:Hyaluronic Acid; D007143:Immunoglobulin Heavy Chains; D000683:Serum Amyloid P-Component; D000099153:Pentraxins
keywords: Amniotic membrane; BMP; HC-HA/PTX3; Limbus; Niche cell; Planar cell polarity; Quiescence; Self-renewal; Signaling; Stem cell niche; Stem cells; Wnt