ArticleMolecular vision2015
Phenotypes of trypsin- and collagenase-prepared bovine corneal endothelial cells in the presence of a selective Rho kinase inhibitor, Y-27632.
Article in Molecular vision, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 4 papers.
What it found
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Who cites it
4 citing papers in PubMed, 10 citations in OpenAlex.
- Comparative culture of human corneal endothelial cells following treatment with human platelet lysate/fibrin hydrogel versus Y-27632 ROCK inhibitor: in vitro and ex vivo study.International ophthalmology · 2022Article
- Isolation efficiency of collagenase and EDTA for the culture of corneal endothelial cells.Molecular vision · 2022Article
- P2Y12 regulates microglia activation and excitatory synaptic transmission in spinal lamina II neurons during neuropathic pain in rodents.Cell death & disease · 2019Article
- Potential Effect of Human Platelet Lysate onJournal of ophthalmic & vision researchArticle
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Authors and funding
6 authors at 3 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
purposeTo optimize isolation of viable bovine corneal endothelial cells (BCECs), we evaluated the effectiveness of various preparation protocols. This entailed comparing the effects of collagenase A and trypsin in the presence and absence of a Rho kinase inhibitor, Y-27632, on proliferation and tight junctional and cytoskeletal integrity during their expansion.
methods5-bromo-2'-deoxyuridine (BrdU) incorporation evaluated cell proliferation. Western blot analysis evaluated F-actin, zonule occludin, and ZO-1 associated nucleic acid binding protein (ZONAB) and RhoA expression. Rho A pulldown assay evaluated Rho A activity.
resultsIn the trypsin (TrypLE)-prepared BCECs, BrdU incorporation decreased whereas nuclear ZONAB expression increased and became stable from day 3 to 7. In contrast, in the collagenase-A-prepared BCECs, we observed preserved ZO-1 integrity, invariant nuclear ZONAB expression, and dense cortical F-actin expression, and BrdU incorporation was invariant from days 1 to 7. Y-27632 did not increase BrdU incorporation and nuclear ZONAB expression in the TrypLE-prepared and the collagenase-A-prepared BCECs. Moreover, Y-27632 increased irregular cellular morphology and downregulated the expression of ZO-1 in the collagenase-A-prepared BCECs from days 1 to 7. Y-27632 inhibited RhoA activation irrespective of whether the cells were isolated with trypsin or collagenase A.
conclusionsIt is preferable to isolate BCECs with collagenase A and expand them without Y-27632. With this protocol, proliferative activity and tight junctional and cytoskeletal integrity are better preserved than if trypsin is used in the presence or absence of Y-27632.
Indexed as
Identifiers
26097378PMC4455892W2309315017What OpenQuestion holds
Registered trials
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