Effect of cell growth supporting surface on phagocytosis

Girish K Srivastava 1, 2, David Rodriguez-Crespo 1, J. Carlos Pastor 1, 2
1. Instituto Universitario de Oftalmobiologia Aplicada (IOBA), Universidad de Valladolid, Valladolid, Spain.
2. Centro en Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Spain.

Phagocytosis is involved in several functions including nutrient uptake, immune response, inflammation, tissue homeostasis, cellular apoptotic bodies and debris elimination. Its dysfunction has impacts on normal functioning of an organism. It is involved in several diseases including the AMD pathogenesis, and treatment approaches. Mostly an initial proof of concept is tested in a laboratory conditions. Cell and tissue cultures need proper surfaces to grow and maintain them in laboratory conditions. The study aims to evaluate effect of two surfaces; tissue culture plate polystyrene and glass coverslip, on phagocytosis performed by RPE cells growing on each surface. Fresh RPE cells and ARPE-19 cell line were grown in corresponding cell culture conditions. Phagocytosis in fresh RPE cell cultures was detected and recorded. ARPE-19 cells were incubated for 4 and 24 hours with 0.2 and 1 μl red fluorescent latex beads, and then images were taken to measure the average numbers of cells, cells performing no phagocytosis and phagocytosis and engulfed red fluorescent latex beads. Results showed that RPE cell membrane was forming blebs releasing black dots like particles as cell debris in cell culture medium. Subsequently cytoplasmic membrane extensions were formed which had engulfed black dots like particles. ARPE-19 cells are a good cellular model for performing initial in vitro test in comparison to fresh RPE cells. There was no significant difference between numbers of ARPE-19 cells growing on polystyrene and glass surface performing latex bead phagocytosis. Both had engulfed red fluorescent latex beads; however, numbers of engulfed beads was significantly higher for glass surface than polystyrene surface. Results support the hypothesis that surface of cell culture plates affects the cellular phagocytosis mechanism which play a critical role in the functions in many cases such as in vivo retina. These findings are very crucial when it comes to transfer a treatment approach based on phagocytosis to the clinics and that the materials and the conditions of the experiments can have very important consequences.    
Keywords: Retinal pigment epithelial cells, phagocytosis, cell growth supporting surface, latex beads  


Bhat, S., and Kumar, A. (2013). Biomaterials and bioengineering tomorrow’s healthcare. Biomatter 3.

Dąbrowska, A.M., and Skopiński, P. (2017). Stem cells in regenerative medicine – from laboratory to clinical application – the eye. Cent.-Eur. J. Immunol. 42, 173–180.

Haruta, M., Sasai, Y., Kawasaki, H., Amemiya, K., Ooto, S., Kitada, M., Suemori, H., Nakatsuji, N., Ide, C., Honda, Y., et al. (2004). In vitro and in vivo characterization of pigment epithelial cells differentiated from primate embryonic stem cells. Invest. Ophthalmol. Vis. Sci. 45, 1020–1025.

Kevany, B.M., and Palczewski, K. (2010). Phagocytosis of Retinal Rod and Cone Photoreceptors. Physiol. Bethesda Md 25, 8–15.

Kocaoglu, O.P., Liu, Z., Zhang, F., Kurokawa, K., Jonnal, R.S., and Miller, D.T. (2016). Photoreceptor disc shedding in the living human eye. Biomed. Opt. Express 7, 4554–4568.

Li, W. (2013). Phagocyte dysfunction, tissue aging and degeneration. Ageing Res. Rev. 12, 1005–1012.

Lu, L., Yaszemski, M.J., and Mikos, A.G. (2001). Retinal pigment epithelium engineering using synthetic biodegradable polymers. Biomaterials 22, 3345–3355.

Mao, Y., and Finnemann, S.C. (2013). Analysis of Photoreceptor Outer Segment Phagocytosis by RPE Cells in Culture. Methods Mol. Biol. Clifton NJ 935, 285–295.

Nadig, R.R. (2009). Stem cell therapy – Hype or hope? A review. J. Conserv. Dent. JCD 12, 131–138.

Nguyen-Legros, J., and Hicks, D. (2000). Renewal of photoreceptor outer segments and their phagocytosis by the retinal pigment epithelium. Int. Rev. Cytol. 196, 245–313.

Phelan, K., and May, K.M. (2016). Basic Techniques in Mammalian Cell Tissue Culture. Curr. Protoc. Toxicol. 70, A.3B.1-A.3B.22.

Rabinovitch, M. (1995). Professional and non-professional phagocytes: an introduction. Trends Cell Biol. 5, 85–87.

Rodriguez-Crespo, D., Di Lauro, S., Singh, A.K., Garcia-Gutierrez, M.T., Garrosa, M., Pastor, J.C., Fernandez-Bueno, I., and Srivastava, G.K. (2014). Triple-layered mixed co-culture model of RPE cells with neuroretina for evaluating the neuroprotective effects of adipose-MSCs. Cell Tissue Res. 358, 705–716.

Rodríguez-Rodero, S., Fernández-Morera, J.L., Menéndez-Torre, E., Calvanese, V., Fernández, A.F., and Fraga, M.F. (2011). Aging Genetics and Aging. Aging Dis. 2, 186–195.

Rosales, C., and Uribe-Querol, E. (2017). Phagocytosis: A Fundamental Process in Immunity. BioMed Res. Int. 2017.

Sparrrow, J.R., Hicks, D., and Hamel, C.P. (2010). The Retinal Pigment Epithelium in Health and Disease. Curr. Mol. Med. 10, 802–823.

Srivastava, G.K., Reinoso, R., Singh, A.K., Fernandez-Bueno, I., Martino, M., Garcia-Gutierrez, M.T., Pastor, J.C., and Corell, A. (2013). Flow cytometry assessment of the purity of human retinal pigment epithelial primary cell cultures. J. Immunol. Methods 389, 61–68.

Strunnikova, N., Zhang, C., Teichberg, D., Cousins, S.W., Baffi, J., Becker, K.G., and Csaky, K.G. (2004). Survival of Retinal Pigment Epithelium after Exposure to Prolonged Oxidative Injury: A Detailed Gene Expression and Cellular Analysis. Invest. Ophthalmol. Vis. Sci. 45, 3767–3777.

Tian, B., Al-Moujahed, A., Bouzika, P., Hu, Y., Notomi, S., Tsoka, P., Miller, J.W., Lin, H., and Vavvas, D.G. (2017). Atorvastatin Promotes Phagocytosis and Attenuates Pro-Inflammatory Response in Human Retinal Pigment Epithelial Cells. Sci. Rep. 7.

Trounson, A., and McDonald, C. (2015). Stem Cell Therapies in Clinical Trials: Progress and Challenges. Cell Stem Cell 17, 11–22.

Ms. Ref. No.: A0201006     RA  
Submitted: 26/07/2018
Published Online: 15/01/2020

How to cite this paper