Selective changes in protein kinase C (PKC) isoform expression in rabbit corneal epithelium during wound healing. Inhibition of corneal epithelial repair by PKCalpha antisense.
Protein kinase C (PKC) isoforms display different sensitivities to modulators, tissue specificities and subcellular localizations. PKCalpha increases during rabbit corneal epithelial wound healing. Here we report differential expression of PKC isoforms in the cornea of rabbits at 1, 2, 4 and 8 days during re-epithelization. Cytosolic, membrane and detergent-insoluble fractions from epithelium were analysed by Western blot using monoclonal antibodies against the different PKC isoforms. We have identified PKCalpha, gamma, epsilon, mu and iota. PKCalpha and gamma were expressed only in the cytosolic fraction, with the expression of PKCalpha markedly increasing 4 days after injury. Corneas cultured in the presence of rabbit-specific PKCalpha antisense showed a greater than 50% inhibition of wound closure, compared to controls. The PKCepsilon and mu were expressed in the soluble, as well as in the membrane fraction. Additionally, 12% of PKCmu was found attached to the detergent insoluble fraction. The expression of the membrane-bound PKCepsilon and mu isoforms decreased between 1 and 2 days following injury. Only 10% of the PKCiota expressed in corneal epithelium was membrane bound, but between 4 and 8 days after de-epithelization, the expression in this fraction increased three-fold. Our results suggest that changes in the expression and distribution within the various fractions of selective isoforms of PKC after injury could be involved in events leading to wound healing and that PKCalpha is a key modulator in rabbit corneal wound repair.
Chandrasekher
G
G
Louisiana State University Medical Center, Department of Ophthalmology and Neuroscience Center of Excellence, 2020 Gravier St. , Suite D, New Orleans, LA, 70112, USA.
Bazan
N G
NG
Bazan
H E
HE
D000818:Animals; D015153:Blotting, Western; D002462:Cell Membrane / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D019573:Epithelium, Corneal / Q000201:enzymology*; D005260:Female; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D016376:Oligonucleotides, Antisense / Q000235:genetics; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D011817:Rabbits; D014945:Wound Healing / Q000502:physiology*
D007527:Isoenzymes; D016376:Oligonucleotides, Antisense; D011493:Protein Kinase C