Abstract |
Transplantation of cultivated human limbo-corneal epithelial (HLE) cells has been recognized as an effective stem cell (SC) therapy for treating corneal epithelial SC deficiency caused by burn or other diseases. With this technique, cryo-preserved human intact amniotic membrane (IAM) has been successfully used as a cell culture substrate and carrier, and is reported to preferentially preserve HLE stem/progenitor cells in vitro. However, little is known about what factors released by HLE cells are involved in the progenitor cell-preserving mechanism. Using proteomic method, we identified 13 proteins over-expressed by HLE cells cultured on IAM, which included heat shock protein 70-1 (Hsp70-1), Hsp-27, glutathione (GSH) S- transferase, annexin A2, galectin-7, and protein S100-A9. Increased Hsp70-1 expression was confirmed by Western blot and real-time PCR. The role of Hsp70-1 in promoting HLE cell survival was demonstrated by increased apoptosis index and increased cleaved poly ADP-ribose polymerase (CPARP) formation in hsp70-1-silenced, but not normal HLE cells after exposure to sublethal UVB irradiation or hydrogen peroxide. To understand the regulatory mechanism of Hsp70-1 expression in HLE cells, the role of transcription factor deltaNp63 (a well-recognized HLE stem cell; SC marker) was studied. We found that over-expression of deltaNp63α by plasmid vector induced a corresponding increase in Hsp70-1 protein production. Likewise, Hsp70-1 expression decreased in HLE cells after addition of deltaNp63α SiRNA. Immunoconfocal microscopy also revealed a paralleled expression of both proteins in corneal specimens. Thus, deltaNp63α-associated Hsp70-1 over-expression may promote HLE progenitor cell survival on IAM, possibly through the cytoprotective and anti-apoptotic effect of Hsp70-1.
|
Authors | David Hui-Kang Ma, Jui-Yang Lai, Szu-Tzu Yu, Jia-Ye Liu, Unique Yang, Hung-Chi Jesse Chen, Lung-Kung Yeh, Yi-Ju Ho, Grace Chang, Su-Fang Wang, Jan-Kan Chen, Ken-Kuo Lin |
Journal | Journal of cellular physiology
(J Cell Physiol)
Vol. 227
Issue 5
Pg. 2030-9
(May 2012)
ISSN: 1097-4652 [Electronic] United States |
PMID | 21751214
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
|
Copyright | Copyright © 2011 Wiley Periodicals, Inc. |
Chemical References |
- HSP70 Heat-Shock Proteins
- TP63 protein, human
- Transcription Factors
- Tumor Suppressor Proteins
- heat-shock protein 70.1
- Hydrogen Peroxide
|
Topics |
- Amnion
(cytology, metabolism)
- Cell Culture Techniques
- Cells, Cultured
- Electrophoresis, Gel, Two-Dimensional
- Epithelial Cells
(cytology, drug effects, physiology, radiation effects)
- Gene Silencing
- HSP70 Heat-Shock Proteins
(genetics, metabolism)
- Humans
- Hydrogen Peroxide
(pharmacology)
- Limbus Corneae
(cytology)
- Proteomics
(methods)
- Stem Cells
(cytology, physiology)
- Transcription Factors
(genetics, metabolism)
- Tumor Suppressor Proteins
(genetics, metabolism)
- Ultraviolet Rays
- Up-Regulation
|