LOXL4, but not LOXL2, is the critical determinant of pathological collagen cross-linking and fibrosis in the lung

Abstract

Idiopathic pulmonary fibrosis is a progressive fibrotic disease characterized by excessive deposition of (myo)fibroblast produced collagen fibrils in alveolar areas of the lung. Lysyl oxidases (LOXs) have been proposed to be the central enzymes that catalyze the cross-linking of collagen fibers. Here, we report that, while its expression is increased in fibrotic lungs, genetic ablation of LOXL2 only leads to a modest reduction of pathological collagen cross-linking but not fibrosis in the lung. On the other hand, loss of another LOX family member, LOXL4, markedly disrupts pathological collagen cross-linking and fibrosis in the lung. Furthermore, knockout of both Loxl2 and Loxl4 does not offer any additive antifibrotic effects when compared to Loxl4 deletion only, as LOXL4 deficiency decreases the expression of other LOX family members including Loxl2 . On the basis of these results, we propose that LOXL4 is the main LOX activity underlying pathological collagen cross-linking and lung fibrosis.

Exploring Extracellular Matrix Crosslinking as a Therapeutic Approach to Fibrosis

“…High-performance liquid chromatography (HPLC) or ultra-performance LC (UPLC) is then used for separation, and mass spectrometry is used for quantification [ 52 , 53 ]. This type of method can be used to simultaneously detect multiple crosslinking modifications, both immature and mature [ 49 , 57 , 58 ]. For comparison across samples, normalization is often required.…”

“…Like TGs, LOX proteins bind to TGFβ, a key regulator of fibrosis [ 15 ]. LOX and LOXL2 are also regulated by hypoxia [ 15 ], which mechanistically explains the upregulation of LOX activity in fibrotic tissues where hypoxia is prevalent [ 1 , 3 , 49 ]. In a recent study by Brereton et al, the relationship between hypoxia-inducible factor (HIF) pathway and LOX activity was explored in depth.…”

“…In fibrosis, there is not only an observed increase in LOX expression and overall crosslinking, but also a change in proportions of different types of crosslinks [ 23 ]. Crosslinks derived from the hydroxylysine aldehyde pathway (deH-HLNL, deH-DHLNL, Pyr, and DPyr) have been shown to increase in fibrotic conditions [ 33 , 49 , 50 , 51 ]. These crosslinks not only alter ECM structure and biomechanical properties, but also change protein susceptibility to degradation by MMPs [ 14 , 23 ].…”

“…Jones et al showed differential gene expression of LOXL2 , LOXL3 , and LOXL4 , but not LOX and LOXL1 [ 50 ]. Brereton et al and Ma et al, however, showed an upregulation of all five LOX genes in IPF tissue [ 39 , 49 ]. Yet, another study by Tjin et al (2017) assessed two different IPF datasets and found that LOXL1 was upregulated in both, but LOXL2 was only upregulated in one [ 60 ].…”

“…They observed a strong increase in deH-DHLNL. A subtle change in Pyr was seen but was not significant when normalized by total collagen [ 49 ]. In summary, both crosslink enzyme expression and crosslinking modifications are dysregulated in lung fibrosis.…”