Boochoon Kieran S, Manarang Joseph C, Davis Joshua T, McDermott Alison M, Foster William J
The Department of Biology & Biochemistry, The University of Houston, Houston, TX, USA.
The College of Optometry, The University of Houston, Houston, TX, USA.
J Biomech. 2014 Sep 22;47(12):3237-40. doi: 10.1016/j.jbiomech.2014.06.021. Epub 2014 Jun 24.
To better understand if a complex process such as phagocytosis is influenced by substrate stiffness, we investigated the influence of substrate elastic modulus on phagocytosis in the retinal pigment epithelial (RPE) cell line ARPE-19. RPE cells lie on Bruch's membrane, directly under the retina, and phagocytose the shed photoreceptor outer segments. Bruch's membrane is known to increase in stiffness by an order of magnitude with age and thus, this study has potential relevance in explaining retinal changes in age-related macular degeneration. ARPE-19 cells were plated on laminin-coated polyacrylamide substrates of varying elastic modulus. After 14 days in culture, a solution of latex fluorescent beads suspended in PBS was placed in each well. After an incubation time of 4h, flow cytometry was performed to determine the number of cells that phagocytosed a bead. The number of ARPE-19 cells that phagocytosed a bead decreased continuously as a function of increasing substrate elastic modulus (p=0.0135), and this was found to be a linear relationship (slope=-0.03305 ± 0.01104, R2=0.4726 per 10,000 cells). Our results suggest that RPE cells display decreased phagocytosis when grown on firmer substrates, and thus, RPE cells in older eyes, in which Bruch's membrane is stiffer, may demonstrate decreased phagocytosis. Impaired phagocytosis by RPE cells may contribute to impaired metabolism of photoreceptor outer segments and to development of macular degeneration. Material stiffness may be a critical parameter in the development of neural therapies, including retinal prosthetics and stem cell therapies.
为了更好地理解诸如吞噬作用这样的复杂过程是否受底物硬度的影响，我们研究了底物弹性模量对视网膜色素上皮（RPE）细胞系ARPE - 19吞噬作用的影响。RPE细胞位于视网膜正下方的布鲁赫膜上，并吞噬脱落的光感受器外段。已知布鲁赫膜的硬度会随着年龄增长而增加一个数量级，因此，本研究对于解释年龄相关性黄斑变性中的视网膜变化具有潜在意义。将ARPE - 19细胞接种在具有不同弹性模量的层粘连蛋白包被的聚丙烯酰胺底物上。培养14天后，将悬浮于PBS中的乳胶荧光珠溶液加入每个孔中。孵育4小时后，进行流式细胞术以确定吞噬珠子的细胞数量。吞噬珠子的ARPE - 19细胞数量随着底物弹性模量的增加而持续减少（p = 0.0135），并且发现这是一种线性关系（斜率=-0.03305±0.01104，每10,000个细胞的R2 = 0.4726）。我们的结果表明，RPE细胞在较硬的底物上生长时吞噬作用降低，因此，在布鲁赫膜更硬的老年眼中的RPE细胞可能表现出吞噬作用降低。RPE细胞吞噬作用受损可能导致光感受器外段代谢受损以及黄斑变性的发展。材料硬度可能是神经治疗（包括视网膜假体和干细胞治疗）发展中的一个关键参数。
