Expression of Granulocyte Colony Stimulating Factor and Its Receptor by Retinal Pigment Epithelial Cells: A Role in Maintaining Differentiation-Competent State

Abstract

Purpose: Granulocyte colony stimulating factor (GCSF) is a potent hematopoietic factor that stimulates the growth of neutrophil granulocyte precursors, and also regulates the differentiation and survival of neutrophils by inhibiting apoptosis. Incidentally, GCSF is also known to act as an endogenous ligand for brain cells, counteracting acute neuronal degeneration and contributing to long-term plasticity of progenitor cells after cerebral ischemia. Since GCSF was recently reported to be present in retinal ganglions, we examined its expression in retinal pigment epithelial (RPE) cells, which, together with retinal neurons, arise from the same underlying precursor cells.

Methods: We used reverse transcriptase polymerase chain reaction (PCR) to assay expression of GCSF and GCSF receptor (GCSFR) genes; immunostaining and flow cytometry to assay the presence of GCSFR on cell surfaces; bromodeoxyuridine (BrdU) incorporation measurement to monitor DNA synthesis; and 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay to monitor cell proliferation. The effect of GCSF on differentiation of RPE cells was assessed by immunocytochemistry to detect the presence of various marker proteins.

Results: The D407 RPE cells, as well as RPE derived from cadaver eyes, were found to express both GCSF and GCSFR. Despite the presence of the GCSF receptor, exogenously added GCSF did not result in any proliferation of these cells. We found that GCSF acts like a de-differentiating factor, maintaining RPE cells in the rounded form, and in a transdifferentiation-competent state.

Conclusions: The expression of GCSF and GCSFR by D407 RPE may be an important factor in RPE cell maintenance.

Keywords::

• Differentiation
• Granulocyte colony stimulating factor
• Granulocyte colony stimulating factor receptor
• Proliferation
• Retinal pigment epithelial cells

ACKNOWLEDGMENTS

The authors wish to thank Drs. Geeta Vemuganti, Kunal Ray, and Frans Cremers for the gift of the RPE cell line D407. Dr. Sanjay Bandyopadhyay and Zydus Research Centre are gratefully acknowledged for their gift of recombinant GCSF. MLG thanks the DST for funding the study, and SK thanks the ICMR for the senior research fellowship.

Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.