In vitro differentiation of human amniotic epithelial stem cells into keratinocytes regulated by OPN3
Shuqi Jin
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorWei Zhang
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorWen Zeng
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorYulei Zhang
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorJianglong Feng
Department of Pathology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorYu Wang
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorHuanhuan Luo
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorTing Liu
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorCorresponding Author
Hongguang Lu
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Correspondence
Hongguang Lu, Department of Dermatology, Affiliated Hospital of Guizhou Medical University. No.28 Guiyi Street, Guiyang, Guizhou, 550 001, China.
Email: [email protected]; [email protected]
Search for more papers by this authorShuqi Jin
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorWei Zhang
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorWen Zeng
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorYulei Zhang
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorJianglong Feng
Department of Pathology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorYu Wang
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorHuanhuan Luo
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorTing Liu
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Search for more papers by this authorCorresponding Author
Hongguang Lu
Department of Dermatology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
Correspondence
Hongguang Lu, Department of Dermatology, Affiliated Hospital of Guizhou Medical University. No.28 Guiyi Street, Guiyang, Guizhou, 550 001, China.
Email: [email protected]; [email protected]
Search for more papers by this authorShuqi Jin, Wei Zhang, and Wen Zeng contributed equally.
Abstract
Human amniotic epithelial stem cells (hAESCs) are regarded as potential alternatives to keratinocytes (KCs) used for skin wound healing. Light is an alternative approach for inducing stem cell differentiation. Opsins (OPNs), a family of light-sensitive, G protein-coupled receptors, play a multitude of light-dependent and light-independent functions in extraocular tissues. However, it remains unclear whether the light sensitivity and function of OPNs are involved in light-induced differentiation of hAESCs to KCs. Herein, we determine the role of OPNs in differentiation of hAESCs into KCs through cell and molecular biology approaches in vitro. It is shown that mRNA expression of OPN3 in the amniotic membrane and hAESCs was higher than the other four primary OPNs by RT-qPCR analysis. Changes in OPN3 gene expression had a significant impact on cell proliferation, stemness and differentiation capability of hAESCs. Furthermore, we found a significant upregulation of OPN3, KRT5 and KRT14 with hAESCs treated at 3 × 33 J/cm2 irradiation from blue-light LED. Taken together, these results suggest that OPN3 acts as a positive regulator of differentiation of hAESCs into KCs. This study provides a novel insight into photosensitive OPNs associated with photobiomodulation(PBM)-induced differentiation in stem cells.
CONFLICT OF INTEREST STATEMENT
The authors declare no conflicts of interest that pertain to this work.
Open Research
DATA AVAILABILITY STATEMENT
The datasets used and/or analysed during the current study are available from the corresponding author upon reasonable request.
Supporting Information
| Filename | Description |
|---|---|
| exd15007-sup-0001-AppendixS1.docxWord 2007 document , 71.3 KB |
Appendix S1. Materials and methods. |
| exd15007-sup-0002-FigureS1.tifTIFF image, 4.4 MB |
Figure S1. Characterization of hAESCs. Representative photomicrograph of adherent hAESCs with spindle shapes on plastic cell culture dish. Flow cytometry analysis of CD90, CD105, CD34, HLA-DR and Pan-CK expression in hAESCs. The blue lines represent the isotype control, and the red lines represent the level of surface markers. |
| exd15007-sup-0003-FigureS2.tifTIFF image, 3.2 MB |
Figure S2. Silence and overexpression of opsin3 in hAESCs (A) hAESCs were transfected with small interfering siRNA directed against OPN3 or control. The efficiency of siRNA was determined by qPCR analysis. Opsin 3 mRNA levels were normalized with GAPDH levels (n = 3 independent experiments). Statistical significance was determined by T-tests. *p < 0.05, **p < 0.01, ***p < 0.001 versus the control group. (B) The efficiency of siRNA was determined by Western blot analysis. (n = 3 independent experiments). GAPDH as a loading control. (C) hAESCs were transfected with a lentiviral vector containing the full-length DNA sequence of human OPN3 (LVOPN3), or a lentiviral vector containing an empty control sequence (LVNC). The transfection of OPN3-suppressed lentivirus was observed under the fluorescence microscope. (D) RT-qPCR was used to analyse the transfection efficiency of the OPN3 gene. (n = 3 independent experiments). Statistical significance was determined by T-tests. *p < 0.05, **p < 0.01, ***p < 0.001 versus the control group. (E) WB was used to analyse the transfection efficiency of the OPN3 protein levels. GAPDH was used as a loading control. |
Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.
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