Myeloperoxidase Inhibition Decreases the Expression of Collagen and Metallopeptidase in Mare Endometria under In Vitro Conditions

Abstract

Neutrophils can originate neutrophil extracellular traps (NETs). Myeloperoxidase (MPO) is a peroxidase found in NETs associated to equine endometrosis and can be inhibited by 4-aminobenzoic acid hydrazide (ABAH). Metallopeptidases (MMPs) participate in extracellular matrix stability and fibrosis development. The objectives of this in vitro work were to investigate, in explants of mare’s endometrium, (i) the ABAH capacity to inhibit MPO-induced collagen type I (COL1) expression; and (ii) the action of MPO and ABAH on the expression and gelatinolytic activity of MMP-2/-9. Explants retrieved from the endometrium of mares in follicular or mid-luteal phases were treated with MPO, ABAH, or their combination, for 24 or 48 h. The qPCR analysis measured the transcription of COL1A2, MMP2, and MMP9. Western blot and zymography were performed to evaluate COL1 protein relative abundance and gelatinolytic activity of MMP-2/-9, respectively. Myeloperoxidase elevated COL1 relative protein abundance at both treatment times in follicular phase (p < 0.05). The capacity of ABAH to inhibit MPO-induced COL1 was detected in follicular phase at 48 h (p < 0.05). The gelatinolytic activity of activated MMP-2 augmented in mid-luteal phase at 24 h after MPO treatment, but it was reduced with MPO+ABAH treatment. The activity of MMP-9 active form augmented in MPO-treated explants. However, this effect was inhibited by ABAH in the follicular phase at 48 h (p < 0.05). By inhibiting the pro-fibrotic effects of MPO, it might be possible to reduce the development of endometrosis. Metallopeptidase-2 might be involved in an acute response to MPO in the mid-luteal phase, while MMP-9 might be implicated in a prolonged exposition to MPO in the follicular phase.

Noscapine Acts as a Protease Inhibitor of In Vitro Elastase-Induced Collagen Deposition in Equine Endometrium

“…The proteases (ELA and CAT) and peroxidase (MPO) found in NETs were recently shown to induce COL1 expression in equine endometrium, suggesting NETs enzymes involvement in the pathophysiology of equine endometrial fibrosis [ 9 , 11 , 12 , 13 , 14 ]. However, not only equine endometrosis seems to be affected by profibrotic effects of NETs.…”

“…Endometrosis treatment has been challenging, with no effective treatment found so far. To make progress on endometrosis treatment, we have also shown that ELA, CAT or MPO fibrotic effect on the mare endometrium can be inhibited in vitro by sivelestat sodium salt, β-keto-phosphonic acid, or 4-aminobenzoic acid hydrazide, respectively [ 12 , 13 , 14 ]. These promising results are the latest advances in equine endometrosis putative treatment.…”

“…The role of some inflammatory mediators, such as prostaglandins, transforming growth factor β1, and interleukins have already been investigated [ 5 , 6 , 7 , 8 ]. Moreover, our recent work has been focused on the involvement of neutrophil extracellular traps (NETs) enzymes in the development of equine endometrosis, such as elastase (ELA), cathepsin G (CAT) and myeloperoxidase (MPO) [ 9 , 10 , 11 , 12 , 13 , 14 ]. At post-breeding endometritis, a fast physiological neutrophil influx to the uterus occurs [ 15 , 16 ] to eliminate the excess of spermatozoa, contaminating bacteria, and debris [ 17 , 18 ].…”

“…Afterwards, neutrophils release their DNA and cytoplasm enzymes to the extracellular environment in a way of enhancing the innate immune response [ 19 , 20 ]. Conversely, NETs enzymes may also drive a profibrotic reaction in mare endometrium that increases the endometrial production of collagen type I (COL1) [ 9 , 11 , 12 , 13 , 14 ].…”

“…Unfortunately, none of the proposed treatments revealed themselves as effective on endometrial fibrosis reduction, nor increased fertility rates. Interestingly, our latest in vitro studies in equine endometrial explants showed that selective inhibitors of NETs enzymes could reduce COL1 expression [ 9 , 11 , 12 , 13 , 14 ]. Sivelestat sodium salt, β-keto-phosphonic acid, and 4-aminobenzoic acid hydrazide are selective ELA, CAT and MPO inhibitors, respectively, which revealed capability of reducing the COL1 induced by these enzymes [ 11 , 12 , 13 , 14 ].…”