Actin filament localization and distribution in the young adult mouse cornea: a correlative immunofluorescent and cytochemical study

Higbee, Russell G.; Hazlett, L. D.

doi:10.1016/0014-4835(83)90003-9

Cited by 10 publications

(5 citation statements)

References 15 publications

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“…Furthermore, Ki67 immunofluorescence staining and qRT-PCR analysis of proliferation related gene expression levels confirmed that 6C treatment promoted cell proliferation ( Figures 1F,G ). Fibrillar actin (F-actin) and tight junctional ZO-1 proteins play important roles in sustaining both cell adhesion and cell-cell tight junctional connectivity ( Higbee and Hazlett, 1983 ; Ryeom et al, 2000 ). When the cells were treated with 6C in vitro , F-actin immunostaining appeared tight and regular, but its pattern in the other four treatment groups was less well defined and more irregular ( Figure 1H ).…”

Section: Resultsmentioning

confidence: 99%

Novel Cell Culture Paradigm Prolongs Mouse Corneal Epithelial Cell Proliferative Activity in vitro and in vivo

Wang

Jin

et al. 2021

Front. Cell Dev. Biol.

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It has been a long-standing challenge to obtain from cell cultures adequate amounts of mouse corneal epithelial cells (mCEC) to perform transplantation surgery. This limitation is attributable to the passage dependent declines in their proliferative activity. We describe here development of a novel 6C medium that contains six different modulators of different signaling pathways, which control proliferative mCEC activity. Its usage shortens the time and effort required to obtain epithelial sheets for hastening healing of an epithelial wound in an experimental animal model. This serum-free 6C medium contains:Y27632, forskolin, SB431542, DAPT, IWP-2, LDN-193189 and also DermaLife K keratinocyte calcium. Their inclusion inhibits rises in four specific markers of epithelial mesenchymal transdifferentiation:ZEB1/2, Snail, β-catenin and α-SMA. This medium is applied in a feeder-free air-lifted system to obtain sufficient populations of epithelial progenitor cells whose procurement is facilitated due to suppression of progenitor epithelial cell transdifferentiation into epithelial-mesenchymal cells. Diminution of this decline in transdifferentiation was confirmed based on the invariance of P63, K14, Pax6, and K12 gene expression levels. This cell culture technique is expected to facilitate ex vivo characterization of mechanisms underlying cell fate determination. Furthermore, its implementation will improve yields of progenitor mouse corneal epithelial cells, which increases the likelihood of using these cells as a source to generate epithelial sheets for performing transplantation surgery to treat limbal stem cell deficiency in a clinical setting. In addition, the novel insight obtainable from such studies is expected to improve the outcomes of corneal regenerative medicine.

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Section: Resultsmentioning

confidence: 99%

Novel Cell Culture Paradigm Prolongs Mouse Corneal Epithelial Cell Proliferative Activity in vitro and in vivo

Wang

Jin

et al. 2021

Front. Cell Dev. Biol.

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“…Antibody to actin was prepared and characterized as described previously (4,5). Antibodies to myosin heavy chain used for light microscopic immunofluorescence were kindly provided by Harry Maisel (Department of Anatomy/Cell Biology, Wayne State University, Detroit, MI).…”

Section: Antibody Characterizationmentioning

confidence: 99%

“…Antibodies to myosin heavy chain used for light microscopic immunofluorescence were kindly provided by Harry Maisel (Department of Anatomy/Cell Biology, Wayne State University, Detroit, MI). Myosin antiserum was characterized and found to be specific by indirect immunofluorescence and immunoblot methods in the same manner as the anti-actin antiserum (4,5). Calmodulin antibody was a kind gift from Michael Welsh (Department of Anatomy, University of Michigan, Ann Arbor, MI).…”

Section: Antibody Characterizationmentioning

confidence: 99%

“…Eyes to be used for culture, were processed as described below. Eyes were removed from culture at 4,8,12,16,20 and 24 hr intervals and either processed for immunofluorescence as previously described (4,5,7) or stained with 1% sodium fluorescein, rinsed with PBS, and examined and photographed under shortwave ultraviolet illumination for wound closure rate analysis. A Wild M8 stereomicroscope (Wild/Leitz, Inc., Rockleigh, NJ) equipped with a 35 mm camera was used for photography.…”

Section: Corneal Epithelial Woundingmentioning

confidence: 99%

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Topical Ocular Anesthetics Affect Epithelial Cytoskeletal Proteins of Wounded Cornea

Higbee¹,

Hazlett²

1989

Journal of Ocular Pharmacology and Therapeutics

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The purpose of this study was to characterize by immunofluorescent microscopy, the cytoskeletal proteins actin and myosin and the protein calmodulin (CaM) in trephined, n-heptanol treated murine corneal epithelium during in vitro organ culture before and following topical ocular anesthetic application. The study has shown that corneal epithelial wounds exposed to anesthetic agents fail to close, even after 24 hr of culture. Failure of the wound to close was accompanied by a general decrease in immunofluorescence for actin, myosin and CaM following application of the anesthetics.

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“…Numerous reports indicate that actin or actinlike proteins are a constituent of many nonmuscle cells (8,14) such as those of the corneal epithelium (11) and ocular lens (15) as well as blood platelets (1), lymphocytes (5), and synaptosomes (6,12). Additionally, there have been sporadic reports on the presence of actinlike proteins in lower eucaryotes and procaryotes (2,3,7,(19)(20)(21).…”

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confidence: 99%

Actinlike material in Pseudomonas aeruginosa

et al. 1985

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Actinlike material was obtained from disrupted Pseudomonas aeruginosa cells by a modification of the method of Hancock and Nikaido (J. Bacteriol. 136:381-390) for isolating outer membrane vesicles. Pelleted membranes were dissolved in Laemmli sample buffer and electrophoresed in parallel lanes with purified rabbit skeletal muscle actin. The bacterial preparation migrated similarly to rabbit skeletal muscle actin. A doublet band was detectable by affinity-purified antiactin antibody in a passive transfer immunoblot. Molecular weight of the actinlike protein doublet was 60,000 to 63,000 as determined by linear regression analysis of Bio-Rad low-molecular-weight standards run on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Transmission electron microscopy of the actinlike material in high salt concentrations revealed 10-to 14-nm filaments of various lengths. Despite its ability to form filaments and to react with a polyclonal rabbit antiactin antibody, the bacterial filaments did not bind the S-1 fragment of heavy meromyosin.

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Actin filament localization and distribution in the young adult mouse cornea: a correlative immunofluorescent and cytochemical study

Cited by 10 publications

References 15 publications

Novel Cell Culture Paradigm Prolongs Mouse Corneal Epithelial Cell Proliferative Activity in vitro and in vivo

Novel Cell Culture Paradigm Prolongs Mouse Corneal Epithelial Cell Proliferative Activity in vitro and in vivo

Topical Ocular Anesthetics Affect Epithelial Cytoskeletal Proteins of Wounded Cornea

Actinlike material in Pseudomonas aeruginosa

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