Characterization of limbal explant sites: Optimization of stem cell outgrowth in in vitro culture

Autor: Wiwit Tantibhedhyangkul, Chawikan Boonwong, Naharuthai Inthasin, Patimaporn Wongprompitak, Methichit Wattanapanitch, Sutthicha Matamnan, Pattama Ekpo, Panotsom Ngowyutagon, Chareenun Chirapapaisan, Mongkol Uiprasertkul, Pinnita Prabhasawat
Jazyk: angličtina
Rok vydání: 2020
Předmět:
0301 basic medicine
Cell Transplantation
medicine.medical_treatment
Immunofluorescence
Cell
Cell Culture Techniques
Stem cell marker
Cornea
Corneal Transplantation
Tissue Culture Techniques
0302 clinical medicine
Animal Cells
Medicine and Health Sciences
Blood and Lymphatic System Procedures
Indirect Immunohistochemistry
Limbal stem cell
Multidisciplinary
Reverse Transcriptase Polymerase Chain Reaction
Stem Cells
Stem Cell Therapy
Ophthalmic Procedures
Epithelium
Corneal
Stem-cell therapy
Adult Stem Cells
medicine.anatomical_structure
Medicine
Anatomy
Cellular Types
Stem cell
Research Article
Ocular Anatomy
Science
Surgical and Invasive Medical Procedures
In Vitro Techniques
Limbus Corneae
Biology
Research and Analysis Methods
03 medical and health sciences
Ocular System
Cadaver
medicine
Humans
Amino Acid Sequence
Immunoassays
Immunohistochemistry Techniques
Cell Proliferation
Clinical Genetics
Transplantation
Tumor Suppressor Proteins
Biology and Life Sciences
Epithelial Cells
Cell Biology
Molecular biology
eye diseases
Histochemistry and Cytochemistry Techniques
030104 developmental biology
Immunologic Techniques
030221 ophthalmology & optometry
Eyes
sense organs
LCOR
Head
Biomarkers
Stem Cell Transplantation
Transcription Factors
Explant culture
Zdroj: PLoS ONE, Vol 15, Iss 5, p e0233075 (2020)
PLoS ONE
ISSN: 1932-6203
Popis: Simple limbal epithelial transplantation (SLET) and cultivated limbal epithelial transplantation (CLET) are proven techniques for treating limbal stem cell deficiency (LSCD). However, the precise regions that are most suitable for preparing explants for transplantation have not been identified conclusively. Accordingly, this in vitro study aimed at determining ideal sites to be selected for tissue harvest for limbal stem cell culture and transplantation. We evaluated cell outgrowth potential and the expression of stem cell markers in cultures from 48 limbal explants from five cadaveric donors. The limbal explants were generated from the three specific sites: Lcor (located innermost and adjacent to the cornea), Lm (middle limbus), and Lconj (located outermost adjacent to the conjunctiva). We found that explants from the Lconj and Lm sites exhibited higher growth potential than those from the Lcor site. Transcript encoding the stem cell marker and p63 isoform, ΔNp63, was detected in cells from Lm and Lconj explants; expression levels were slightly, though significantly (p-value < 0.05), higher in Lm than in Lconj, although expression of ΔNp63α protein was similar in cells from all explants. Differential expression of ATP-Binding Cassette Subfamily G Member 2 (ABCG2) did not reach statistical significance. Immunohistochemistry by indirect immunofluorescence analysis of limbus tissue revealed that the basal layer in explant tissue from Lconj and Lm contained markedly more stem cells than found in Lcor explant tissue; these findings correlate with a higher capacity for growth. Collectively, our findings suggest that explants from the Lconj and Lm sites should be selected for limbal cell expansion for both CLET and SLET procedures. These new insights may guide surgeons toward specific limbal sites that are most suitable for stem cell culture and transplantation and may ultimately improve treatment outcomes in the patients with LSCD.
Databáze: OpenAIRE
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