Zinc reduces epithelial barrier compromise induced by human seminal plasma.

Human semen has the potential to modulate the epithelial mucosal tissues it contacts, as seminal plasma (SP) is recognized to contain both pro- and anti-barrier components, yet its effects on epithelial barrier function are largely unknown. We addressed the role of human SP when exposed to the basal-lateral epithelial surface, a situation that would occur clinically with prior mechanical or disease-related injury of the human epithelial mucosal cell layers in contact with semen. The action of SP on claudins-2, -4, -5, and -7 expression, as well as on a target epithelium whose basolateral surface has been made accessible to SP, showed upregulation of claudins-4 and -5 in CACO-2 human epithelial cell layers, despite broad variance in SP-induced modulation of transepithelial electrical resistance and mannitol permeability. Upregulation of claudin-2 by SP also exhibited such variance by SP sample. We characterize individual effects on CACO-2 barrier function of nine factors known to be present abundantly in seminal plasma (zinc, EGF, citrate, spermine, fructose, urea, TGF, histone, inflammatory cytokines) to establish that zinc, spermine and fructose had significant potential to raise CACO-2 transepithelial resistance, whereas inflammatory cytokines and EGF decreased this measure of barrier function. The role of zinc as a dominant factor in determining higher levels of transepithelial resistance and lower levels of paracellular leak were confirmed by zinc chelation and exogenous zinc addition. As expected, SP presentation to the basolateral cell surface also caused a very dramatic yet transient elevation of pErk levels. Results suggest that increased zinc content in SP can compete against the barrier-compromising effect of negative modulators in SP when SP gains access to that epithelium's basolateral surface. Prophylactic elevation of zinc in an epithelial cell layer prior to contact by SP may help to protect an epithelial barrier from invasion by SP-containing STD microbial pathogens such as HPV or HIV.

Drug delivery of zinc to Barrett's metaplasia by oral administration to Barrett's esophagus patients.

BACKGROUND: Delivery of a pharmacologically effective drug dosage to a target tissue is critical. Barrett's epithelia are a unique challenge for drug delivery of orally administered zinc due to rapid transit down the esophageal lumen, incomplete absorptive differentiation of these epithelia, and the use of proton-pump inhibitor drugs abrogating intestinal uptake of supplemental zinc. METHODS: Barrett's esophagus patients were administered oral zinc gluconate (26 mg zinc twice daily) for 14 days prior to biopsy procurement. Barrett's biopsies were analyzed for total zinc content by atomic absorption spectroscopy and by western immunoblot for cellular proteins known to be regulated by zinc. RESULTS: Cellular levels of both the Znt-1 transport protein and the alpha isoform of PKC were over 50% lower in the zinc treatment group. CONCLUSION: Oral zinc administration can result in effective delivery of zinc to Barrett's epithelia with resulting effects on intracellular signal transduction.

Zinc enhancement of LLC-PK(1) renal epithelial barrier function.

BACKGROUND AND AIMS: Earlier work by our group and others has documented improvement of epithelial barrier function in human gastrointestinal models. Here we tested zinc's ability to improve a renal epithelial model. Our aim was to compare the functional and structural effects of zinc on the tight junctional (TJ) complexes of these two very distinct epithelial cell types. Zinc's ability to achieve barrier enhancement in very different epithelial cell types by action upon distinct molecular targets in each epithelial model may suggest a fundamental general role for supplemental zinc in epithelial barrier improvement throughout the body. METHODS: Cell layers were exposed to 50 or 100 µM zinc on both cell surfaces for 48 h followed by measurement of transepithelial electrical resistance (Rt) and transepithelial (14)C-mannitol flux (Jm). TJ proteins in cell layers were analyzed by Western immunoblot. RESULTS AND CONCLUSIONS: Zinc supplementation improved the basal TJ barrier function of LLC-PK1 renal cell layers, exemplified by increased Rt and decreased Jm. These zinc-induced changes were also accompanied by decreased NaCl dilution potentials. Of the tight junctional proteins that were tested (occludin, claudins 1, 2, 3, 4, and 5, and tricellulin), we did not observe a zinc-induced change in abundance of any of them, in detergent-soluble fractions of lysates of confluent differentiated cell layers. However, examination of cytosolic fractions showed concentration-dependent increases in the levels of claudins -2 and -4 in this compartment as a result of supplemental zinc. The effects of supplemental zinc on the tight junctional complexes and barrier properties of this renal epithelial model are contrasted with zinc effects on the CACO-2 gastrointestinal model.

Zinc supplementation modifies tight junctions and alters barrier function of CACO-2 human intestinal epithelial layers.

BACKGROUND: Zinc deficiency is known to result in epithelial barrier leak in the GI tract. Precise effects of zinc on epithelial tight junctions (TJs) are only beginning to be described and understood. Along with nutritional regimens like methionine-restriction and compounds such as berberine, quercetin, indole, glutamine and rapamycin, zinc has the potential to function as a TJ modifier and selective enhancer of epithelial barrier function. AIMS: The purpose of this study was to determine the effects of zinc-supplementation on the TJs of a well-studied in vitro GI model, CACO-2 cells. METHODS: Barrier function was assessed electrophysiologically by measuring transepithelial electrical resistance (Rt), and radiochemically, by measuring transepithelial (paracellular) diffusion of 14C-D-mannitol and 14C-polyethyleneglycol. TJ composition was studied by Western immunoblot analyses of occludin, tricellulin and claudins-1 to -5 and -7. RESULTS: Fifty- and 100-µM zinc concentrations (control medium is 2 µM) significantly increase Rt but simultaneously increase paracellular leak to D-mannitol. Claudins 2 and 7 are downregulated in total cell lysates, while occludin, tricellulin and claudins-1, -3, -4 and -5 are unchanged. Claudins-2 and -7 as well as tricellulin exhibit decreased cytosolic content as a result of zinc supplementation. CONCLUSIONS: Zinc alters CACO-2 TJ composition and modifies TJ barrier function selectively. Zinc is one of a growing number of "nutraceutical" substances capable of enhancing epithelial barrier function, and may find use in countering TJ leakiness induced in various disease states.

Omeprazole induces gastric transmucosal permeability to the peptide bradykinin.

AIM: To investigate omeprazole-induced transepithelial gastric leak and its effects on the permeability of the peptides bradykinin and oxytocin. METHODS: Rat gastric corpus tissue was isolated and mounted in an Ussing chamber apparatus to evaluate the permeability of (3)H-bradykinin, (3)H-oxytocin, and (14)C-EDTA in the presence or absence of omeprazole. Thin-layer chromatography was performed to identify any metabolic breakdown products of the peptides resulting from permeation through the gastric tissue, and thereby calculate the true flux of the peptide. RESULTS: The flux rate of intact (3)H-bradykinin increased substantially after omeprazole addition (109.5%) compared to the DMSO vehicle control (14%). No corresponding change in flux of intact (3)H-oxytocin was observed under the same conditions (11.9% and 6.4% in the DMSO- and omeprazole-treated conditions, respectively). After exposure to omeprazole, the flux rate of (14)C-EDTA also increased dramatically (122.3%) compared to the DMSO condition (36.3%). CONCLUSION: The omeprazole-induced gastric leak allows for transmucosal permeability to charged molecules as well as non-electrolytes. This induced leak will allow certain peptides to permeate.