27 December 2017 : Animal Research
Isolation and Culture of Primary Mouse Retinal Pigment Epithelial (RPE) Cells with Rho-Kinase and TGFβR-1/ALK5 Inhibitor
Junhui Shen1ABCEF, Jianfeng He1BCD, Fang Wang1DEG*DOI: 10.12659/MSM.905569
Med Sci Monit 2017; 23:6132-6136
Abstract
BACKGROUND: Primary RPE cells could be a reliable model for representing in vivo status of RPE compared with cell lines. We present a protocol for in vitro isolation and culture of primary RPE cells from C57BL mice.
MATERIAL AND METHODS: We used C57BL mice ages 7 days to 4 months. The RPE layer was separated from the neural retina layer by digestion with 2% Dispase for 45 min and scraped off from the choroid after 25-min incubation in 37°C. Collected RPE sheets were gently pipetted up into smaller sheets. RPE sheets were transferred into well plates and cultured in vitro for 2 weeks. To inhibit epithelial-mesenchymal transition (EMT) of RPE cells, we used Y27632 and Repsox to treat cultured primary RPE cells.
RESULTS: RPE cells isolated from C57BL mice maintained pigmented and hexagonal morphology in culture. However, long-term in vitro culture lead to the periphery cells of a RPE sheet becoming mesenchymal-like cells. In contrast to the control group, Y27632 and Repsox, which are inhibitors of Rho-kinase or TGFβR-1/ALK5, promoted primary RPE cells to maintain epithelial-like morphology and eventually become confluent.
CONCLUSIONS: RPE cells isolated from C57BL mice could be a powerful cell model to study the biological function of RPE. Especially, C57BL mice with different defective genetic background resulting in ocular diseases, would expand the genome type of RPE cells. The method presented here could be an efficient and applicable technique to obtain large numbers of primary RPE cells that maintain some characteristics of in vivo RPE.
Keywords: primary cell culture, retinal pigment epithelium
Editorial
01 November 2024 : Editorial
Editorial: Artificial Intelligence (AI), Digital Image Analysis, and the Future of Cancer Diagnosis and PrognosisDOI: 10.12659/MSM.947038
Med Sci Monit 2024; 30:e947038
In Press
Clinical Research
Iliac Periosteal Bone Autografting vs Talus Non-Weight-Bearing Surgery in Hepple V Talus Osteochondral Inju...Med Sci Monit In Press; DOI: 10.12659/MSM.944912
Clinical Research
Impact of Different Orthodontic Appliances on Mandibular Condyle, Anterior Region of Mandibular, and Maxill...Med Sci Monit In Press; DOI: 10.12659/MSM.946265
Clinical Research
Risk Factors for Bone Cement Displacement After Percutaneous Kyphoplasty in Osteoporotic Vertebral Fracture...Med Sci Monit In Press; DOI: 10.12659/MSM.945884
Clinical Research
Adipose-Derived Mesenchymal Stem Cells from Arthritis Patients: Differential Modulation of CD4⁺ T Cell Acti...Med Sci Monit In Press; DOI: 10.12659/MSM.945273
Most Viewed Current Articles
17 Jan 2024 : Review article 6,407,835
Vaccination Guidelines for Pregnant Women: Addressing COVID-19 and the Omicron VariantDOI :10.12659/MSM.942799
Med Sci Monit 2024; 30:e942799
14 Dec 2022 : Clinical Research 1,909,112
Prevalence and Variability of Allergen-Specific Immunoglobulin E in Patients with Elevated Tryptase LevelsDOI :10.12659/MSM.937990
Med Sci Monit 2022; 28:e937990
16 May 2023 : Clinical Research 695,529
Electrophysiological Testing for an Auditory Processing Disorder and Reading Performance in 54 School Stude...DOI :10.12659/MSM.940387
Med Sci Monit 2023; 29:e940387
07 Jan 2022 : Meta-Analysis 260,637
Efficacy and Safety of Light Therapy as a Home Treatment for Motor and Non-Motor Symptoms of Parkinson Dise...DOI :10.12659/MSM.935074
Med Sci Monit 2022; 28:e935074