Endotoxin-induced changes of type VII collagen– cleaving matrix metalloproteinases in lamellar tissue of extracorporeally perfused equine limbs

Abstract

OBJECTIVE To investigate the effect of lipopolysaccharide (LPS) on type VII collagen- cleaving matrix metalloproteinases (MMPs) in the lamellar tissue of extracorporeally perfused equine limbs. SAMPLE 10 right forelimbs and 3 left forelimbs collected from 10 adult horses after slaughter at a licensed abattoir. PROCEDURES Extracorporeal perfusion of the isolated equine limbs was performed for 10 hours under physiologic conditions (control-perfused limbs; n = 5) and with the addition of 80 ng of LPS/L of perfusate (LPS-perfused limbs; 5). Lamellar tissue specimens were then collected from the dorsal aspect of the hooves. Additionally, corresponding control specimens were collected from the 3 nonperfused left forelimbs. Immunohistochemical analysis was performed on paraffin-embedded tissue blocks with antibodies against total (latent and active) MMP-1, MMP-2, MMP-8, and MMP-9 as well as antibody against active MMP-9. Intensity of immunohistochemical staining was scored, and stain distribution in the lamellar tissue was noted. RESULTS Staining intensity of total and active MMP-9 was significantly increased in LPS-perfused versus control-perfused limbs. No such difference was identified for MMP-1, MMP-2, and MMP-8. CONCLUSIONS AND CLINICAL RELEVANCE Of the 4 MMPs that are capable of degrading type VII collagen, MMP-9 was the only one for which production increased in the lamellar tissue of isolated equine limbs perfused with versus without a clinically relevant concentration of LPS. These results suggested that MMP-9 may be involved in initiation of pathological changes in lamellar tissue in endotoxin-induced laminitis, whereas MMP-1, MMP-2, and MMP-8 may be less relevant.

Endotoxin‐induced changes in expression of cyclooxygenase isoforms in the lamellar tissue of extracorporeally haemoperfused equine limbs

“…The LPS-induced increase in COX-2 which was observed in the present study may also be responsible for the increased expression of MMP-9 in the LPS-exposed lamellar tissue (Patan-Zugaj et al, 2018), because prostaglandin E 2 , a metabolite of COX-2, promotes the production of various MMPs including MMP-9 (Shankavaram, DeWitt, Funk, Sage, & Wahl, 1997). Specifically, MMP-9 is thought to play an important role in the degradation and structural failure of the dermo-epidermal basement membrane during SIRS-induced laminitis (Medina-Torres et al, 2011;Loftus et al, 2009;Pollitt, Pass, & Pollitt, 1998;Riggs et al, 2009).…”

“…In previous studies using this technique, it was shown that direct exposure of the lamellar tissue to LPS leads to changes in haemodynamics with vasodilation of laminar vessels and formation of microthrombi (Patan-Zugaj et al, 2012;Patan-Zugaj, Gauff, Plendl, & Licka, 2014). Additionally, LPS exposure induced inflammatory reaction in the lamellar tissue with intra-and extravascular accumulation of leucocytes, increased expression and activation of MMP-9 and damage of the dermo-epidermal basement membrane (Patan-Zugaj, Gauff, Egerbacher, & Licka, 2018;Patan-Zugaj et al, 2014). These events are similar to the pathological changes seen in lamellar tissue of horses with naturally acquired or experimentally induced SIRSrelated laminitis (French & Pollitt, 2004;Loftus, Johnson, Belknap, Pettigrew, & Black, 2009;Nourian et al, 2007); they also occur as first steps in the angiogenesis process (Alkim et al, 2015).…”

The Equine Hoof: Laminitis, Progenitor (Stem) Cells, and Therapy Development

“…42 The enzymes can also cause excessive degradation of key proteins that stabilize the epidermaldermal interface like laminin when tissue homeostasis is disrupted by increased neutrophil MMP (eg, MMP2 and MMP9) production in the presence of pro-inflammatory cytokines (eg, interleukin [IL] 1b and 8). [43][44][45][46] While it is clear that coordination of tissue production and degradation is disrupted by inflammatory mediators, the exact contributions of and consequences to resident progenitor cells have yet to be determined.…”

“…[57][58][59] Use of endotoxin to cause laminitis is controversial since lipopolysaccharide administration to normal horses tends to create signs of clinical sepsis and decrease digital perfusion, but it does not appear to cause laminitis in otherwise normal horses. 46,[60][61][62] Carbohydrate overload with corn starch and wood cellulose flour or black walnut extract (BWE) cause microstructural changes in the hoof which resemble those of naturally occurring laminitis (Figure 5). 57,63 Oligofructose derived from chicory root and hyperinsulinemia are also used to create laminitis.…”