Abstract
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.
Highlights
With the exception of pathogens introduced during intravenous drug use, the first obstacle that a pathogen faces when invading an organism is an epithelial cell layer
The effect of seminal plasma (SP) on CACO-2 epithelial barrier function was evaluated by both Rt, as well as by measurement of the transepithelial diffusion of 14C-D-mannitol (Jm) across the same cell layers
The SP-induced phosphorylation of Erk was observed to have a rapid time course with pErk levels maximal at 15 mins after SP exposure, and declining to control levels by 4 hrs. This current study shows that in the CACO-2 epithelial model: 1) SP at fixed dilutions can have an enhancing effect on epithelial barrier properties when presented to the basolateral surface; 2) the effect is variable across individual SP samples, with certain samples producing a significant barrier-enhancing effect, while others produce a significant barrier-compromising doi:10.1371/journal.pone.0170306.g006
Summary
With the exception of pathogens introduced during intravenous drug use, the first obstacle that a pathogen faces when invading an organism is an epithelial cell layer. A pathogen can cross an epithelial barrier by invading an epithelial cell across its apical membrane and exiting the cell across its basolateral membrane, or by apically invading an epithelial cell, killing the cell, thereby creating opportunity for systemic invasion by more pathogens, or by infecting cells “poking through” the barrier, such as dendritic cells, which can “carry” the virus across the barrier.[7,8,9,10] certain pathogens have developed a different approach by directly binding to TJ components, such as specific claudin proteins, thereby enabling invasion of the epithelial cell, and inducing junctional leakiness This highly refined mechanism has been used by both bacterial and viral pathogens as seen, for example, in the etiology of Clostridium perfringens and hepatitis C.[11, 12] There are, other pathogens that enter the epithelial cell via interaction with non-junctional membrane proteins but still compromise the epithelial barrier by giving rise to junctional leakiness, a mechanism seemingly used by HIV and HPV.[13,14,15] A recent study by Abdulhaqq et al.[16] indicated that chronic unprotected sex is associated with changes in the female reproductive tract that may make HIV infection less likely. We focus here on SP contact with the basolateral surface of an epithelial layer, a situation that would occur whenever luminally presented SP comes into contact with an epithelium that has been damaged mechanically or by inflammation/infection
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