Submitted on June 6, 2002
Accepted on December 16, 2002
Bryostatin-1 Attenuates TNF-Induced Epithelial Barrier Dysfunction: Role of novel PKC isozymes
James Yoo1, Anthony Nichols2, Jaekyung C. Song3, Joshua Mammen2, Isabel Calvo3, Roger Worrell2, John Cuppoletti2, Karl Matlin3, and Jeffrey B. Matthews3*
1 Department of Surgery, Beth Israel Deaconess Medical Center, Boston, MA, USA
2 Department of Surgery, University of Cincinnati Medical Center, Cincinnati, Ohio, USA
3 Department of Surgery, University of Cincinnati Medical Center, Cincinnati, Ohio, USA; Department of Surgery, Beth Israel Deaconess Medical Center, Boston, MA, USA
* To whom correspondence should be addressed. E-mail: jeffrey.matthews{at}uc.edu.
Tumor necrosis factor (TNF) increases epithelial permeability in many model systems. Protein kinase C (PKC) isozymes regulate epithelial barrier function and alter ligand-receptor interactions. We sought to define the impact of PKC on TNF-induced barrier dysfunction in T84 intestinal epithelia. TNF induced a dose- and time-dependent fall in transepithelial electrical resistance (TER) and an increase in 3H-mannitol flux. The TNF-induced fall in TER was not PKC-mediated, but was prevented by pre-treatment with bryostatin-1, a PKC agonist. Based on a pattern of sensitivity to pharmacologic inhibitors of PKC, this epithelial barrier preservation was mediated by novel PKC isozymes. Bryostatin-1 reduced TNF receptor (TNF-R1) surface availability, based on radio-labeled TNF binding and cell surface biotinylation assays, and increased TNF-R1 receptor shedding. The pattern of sensitivity to isozyme-selective PKC inhibitors suggested that these effects were mediated by activation of PKC
. In addition, after bryostatin-1 treatment, PKC
and TNF-R1 became associated based on mutual co-immunoprecipitation assay, which has been shown to lead to receptor desensitization in neutrophils. TNF-induced barrier dysfunction occurs independently of PKC, but selective modulation of novel PKC isozymes may regulate TNF-R1 signaling.
