Cultivation of human corneal limbal stem cells in Mebiol gel--A thermo-reversible gelation polymer.
OBJECTIVE
Cultivated limbal stem cell transplantation is being used as a current treatment modality for limbal stem cell deficiency. However, use of allogenic biological material as substrate is associated with risks of transmission of certain diseases and allograft rejection. Therefore development of non-toxic biodegradable synthetic polymers is important. We undertook this study to evaluate the use of a synthetic polymer Mebiol gel as a substrate for the growth of limbal phenotype cells and cornea phenotype cells from limbal explants.


METHODS
Human cadaveric limbal explants cells were cultivated on Mebiol gel. The proliferative capacity of cultivated cells was analyzed with thymidine incorporation studies. Immunostaining for presumed limbal stem cell association markers and cornea differentiation markers was performed and confirmed with reverse transcription (RT-PCR).


RESULTS
The limbal explants underwent proliferation in vitro. The cultivated cells expressed the presumed limbal stem cell association markers (ABCG2 and p63), the transient amplifying cell markers (connexin 43, integrin alpha9) and the cornea differentiation marker (K3). RT PCR confirmed the immunohistochemical data.


CONCLUSIONS
Our findings showed that the synthetic polymer Mebiol gel was able to support limbal explant proliferation. The cultured cells expressed presumed limbal stem cell association markers, transient amplifying cells and cornea phenotype markers. Mebiol Gel can be used as a scaffold for growing limbal explants.
Sudha
B
B
Vision Research Foundation, Sankara Nethralaya, Chennai, India.
Madhavan
H N
HN
Sitalakshmi
G
G
Malathi
J
J
Krishnakumar
S
S
Mori
Y
Y
Yoshioka
H
H
Abraham
S
S
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000032:analysis; D018929:Cell Culture Techniques / Q000379:methods*; D002470:Cell Survival; D004268:DNA-Binding Proteins / Q000032:analysis; D005455:Fluorescent Antibody Technique; D005782:Gels; D006801:Humans; D007150:Immunohistochemistry; D039001:Integrin alpha Chains / Q000032:analysis; D016850:Limbus Corneae / Q000737:chemistry / Q000166:cytology*; D009363:Neoplasm Proteins / Q000032:analysis; D013234:Stem Cells / Q000166:cytology*; D015534:Trans-Activators / Q000032:analysis; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins / Q000032:analysis
C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D004268:DNA-Binding Proteins; D005782:Gels; D039001:Integrin alpha Chains; D009363:Neoplasm Proteins; C115051:TP63 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C084613:integrin alpha9