Objective To determine if protein kinase Cα (PKCα) affects the migration of retinal pigment epithelium (RPE) cells. Methods In this experimental study, primary human RPE cells were obtained from normal human eyes (cornea donors). The human RPE cells were divided into four groups: control, thymeleatoxin, nonsilencing siRNA-PKCα, and siRNA-PKCα. PKCα mRNA was analyzed by reverse transcriptase-polymerase chain reaction. Cell migration was analyzed by wound healing, transwell chamber assays, and live-cell imaging. Independent samples t tests were used for comparison between pairs of the four groups. Results Human RPE cells were successfully cultured and identified in vitro. Microscopy and staining tests demonstrated that each cell culture consisted of a pure population of epithelial cells. Thymeleatoxin, an activator of PCK, significantly increased the mRNA levels of PKCα compared to the control group (t=3.712, P=0.034). siRNA-PKCα significantly reduced the mRNA levels of PKCα compared to the nonsilencing siRNA group (t=3.274, P=0.031). The wound healed faster in the thymeleatoxin group than in the control group (t=9.743, P＜0.001), and it healed slower in the siRNA-PKCα group compared to the nonsilencing siRNA group (t=3.643, P＜0.001). A similar tendency among the four groups was supported by cell number counts in the transwell chamber assays and cell migration distances in the 12-hour tracking test. Conclusion PKCα-mediated signal transduction plays a crucial role in RPE cell migration and may be used as a potential therapeutic target against RPE cell migration and proliferative vitreoretinopathy disease.
Tseng W, Cortez RT, Ramirez G, et al. Prevalence and risk factors for proliferative vitreoretinopathy in eyes with rhegmatogenous retinal detachment but no previous vitreoretinal surgery[J]. Am J Ophthalmol, 2004, 137(6): 1105- 1115. DOI: 10.1016/j.ajo.2004.02.008.
[2]
Pastor JC, Rojas J, Pastor-Idoate S, et al. Proliferative vitreoretinopathy: A new concept of disease pathogenesis and practical consequences[J]. Prog Retin Eye Res, 2016, 51: 125-155. DOI: 10.1016/j.preteyeres.2015.07.005.
[3]
Leiderman YI, Miller JW. Proliferative vitreoretinopathy: pathobiology and therapeutic targets[J]. Semin Ophthalmol, 2009, 24(2): 62-69. DOI: 10.1080/08820530902800082.
[4]
Hiscott P, Hagan S, Heathcote L, et al. Pathobiology of epiretinal and subretinal membranes: possible roles for the matricellular proteins thrombospondin 1 and osteonectin (SPARC)[J]. Eye (Lond), 2002, 16(4): 393-403. DOI: 10.1038/sj.eye.6700196.
Nagineni CN, Samuel W, Nagineni S, et al. Transforming growth factor-beta induces expression of vascular endothelial growth factor in human retinal pigment epithelial cells: involvement of mitogen-activated protein kinases[J]. J Cell Physiol, 2003, 197(3): 453-462. DOI: 10.1002/jcp.10378.
[8]
Gao Q, Tan J, Ma P, et al. PKC alpha affects cell cycle progression and proliferation in human RPE cells through the downregulation of p27kip1[J]. Mol Vis, 2009, 15: 2683-2695.
Chen Y, Tian Q. The role of protein kinase C epsilon in neural signal transduction and neurogenic diseases[J]. Front Med, 2011, 5(1): 70-76. DOI: 10.1007/s11684-011-0119-9.
[11]
Palma-Nicolás JP, López E, López-Colomé AM. Thrombin stimulates RPE cell motility by PKC-zeta- and NF-kappaB- dependent gene expression of MCP-1 and CINC-1/GRO chemokines[J]. J Cell Biochem, 2010, 110(4): 948-967. DOI: 10. 1002/jcb.22608.
[12]
Igumenova TI. Dynamics and membrane interactions of protein kinase C[J]. Biochemistry, 2015, 54(32): 4953-4968. DOI: 10.1021/ acs.biochem.5b00565.
[13]
Roivainen R, Messing RO. The phorbol derivatives thymeleatoxin and 12-deoxyphorbol-13-O-phenylacetate-10-acetate cause
translocation and down-regulation of multiple protein kinase C isozymes[J]. FEBS Lett, 1993, 319(1-2): 31-34.
[14]
Qiu S, Jiang Z, Huang Z, et al. Migration of retinal pigment epithelium cells is regulated by protein kinase Cα in vitro[J]. Invest Ophthalmol Vis Sci, 2013, 54(10): 7082-7090. DOI: 10. 1167/iovs.13-12099.
[15]
Zhao LJ, Xu H, Qu JW, et al. Modulation of drug resistance in ovarian cancer cells by inhibition of protein kinase C-alpha (PKC-α) with small interference RNA (siRNA) agents[J]. Asian Pac J Cancer Prev, 2012, 13(8): 3631-3636.
[16]
Schadlu R, Tehrani S, Shah GK, et al. Long-term follow-up results of ilm peeling during vitrectomy surgery for premacular fibrosis[J]. Retina, 2008, 28(6): 853-857. DOI: 10.1097/IAE.0b01 3e3181631962.
[17]
Hou H, Huffman K, Rios S, et al. A novel approach of daunorubicin application on formation of proliferative retinopathy using a porous silicon controlled delivery system: pharmacodynamics[J]. Invest Ophthalmol Vis Sci, 2015, 56(4): 2755-2763. DOI: 10.1167/iovs.15-16526.
浙公网安备 33030202001892号