Cellular and Molecular Mechanisms of Hypertrophy of Ligamentum Flavum

Abstract

Hypertrophy of the ligamentum flavum (HLF) is a common contributor to lumbar spinal stenosis (LSS). Fibrosis is a core pathological factor of HLF resulting in degenerative LSS and associated low back pain. Although progress has been made in HLF research, the specific molecular mechanisms that promote HLF remain to be defined. The molecular factors involved in the onset of HLF include increases in inflammatory cytokines such as transforming growth factor (TGF)-β, matrix metalloproteinases, and pro-fibrotic growth factors. In this review, we discuss the current understanding of the mechanisms involved in HLF with a particular emphasis on aging and mechanical stress. We also discuss in detail how several pathomechanisms such as fibrosis, proliferation and apoptosis, macrophage infiltration, and autophagy, in addition to several molecular pathways involving TGF-β1, mitogen-activated protein kinase (MAPKs), and nuclear factor-κB (NF-κB) signaling, PI3K/AKT signaling, Wnt signaling, micro-RNAs, extracellular matrix proteins, reactive oxygen species (ROS), etc. are involved in fibrosis leading to HLF. We also present a summary of the current advancements in preclinical animal models for HLF research. In addition, we update the current and potential therapeutic targets/agents against HLF. An improved understanding of the molecular processes behind HLF and a novel animal model are key to developing effective LSS prevention and treatment strategies.

Keywords: TGF-β; aging; fibrosis; hypertrophy; ligamentum flavum; low back pain; lumbar spinal stenosis.

Figure 1

General outlook of lumbar spinal stenosis (LSS) and hypertrophy of ligamentum flavum (HLF). (A) HLF along with hypertrophy of facet joint and intervertebral disc bulging, or degeneration contributes to narrowing of the spinal canal causing lumbar spinal stenosis (LSS). (B) The reported stress, signaling molecules, and consequences of fibrosis and HLF that cause LSS are shown. Mechanical stress in addition to aging and associated inflammatory or oxidative stress initiates the fibrotic changes mediated through myofibroblast differentiation, inflammation, proliferation, and inhibition of apoptosis regulated through molecular signaling such as transforming growth factor (TGF)-β, other cytokines and growth factors, reactive oxygen species (ROS), non-coding RNAs, etc. These signaling events cause the accumulation of extracellular matrix (ECM) and fibrosis leading to development of HLF. Please refer to text for detailed explanations. (Figure created with BioRender.com, accessed on 1 August 2024).

Figure 2

Proposed signaling pathways involved in the pathogenesis of hypertrophy of ligamentum flavum (HLF) based on reported findings. Mechanical stress/aging/microinjuries are the major factors inducing HLF through several signaling pathways leading to increase in inflammation, proliferation and ECM deposition and reduction in apoptosis thereby causing HLF. Transforming growth factor (TGF)-β1 signaling through TGF-β receptor (TGFR) is one of the main signaling events involved. Mechanical stress activates the inflammatory reaction at site of injury attracting macrophages, which along with endothelial cells and fibroblast releases the TGF-β1. TGF-β1 triggers the downstream signaling via TGFR to activate SMAD2 and SMAD3, which are then responsible for transcription of pro-inflammatory, pro-fibrotic markers such as interleukin (IL)-6, TGF-β1, matrix metallopeptidases (MMPs), Collagen I (COLI), COLIII, alpha-smooth muscle actin (α-SMA), etc. These factors further amplify the fibrotic process through cytokine receptor-mediated signaling to induce fibrosis and aid the pathogenesis of HLF. Specifically, α-SMA is involved in the myofibroblast differentiation of fibroblasts to increase the ECM components and fibrosis. Epidermal growth factor (EGF) also mediates the SMAD3-mediated signaling through the EGF receptor (EGFR). Increased level of connective tissue growth factor (CTGF) and periostin through TGF-β1 signaling induces fibrotic events by mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NF-κB) activation, respectively. In addition, periostin functions via the integrin receptor. Increased level of angiopoietin line 2 (ANGPTL2) during the HLF also activates Integrin α4β5-mediated NF-κB signaling. Furthermore, cytokine receptor-like factor 1 (CRLF) induced by TGF-β1 or IL-1β activates ERK MAPK to induce the transcription of pro-fibrotic genes. Accumulation of reactive oxygen species (ROS) occurs due to oxidative stress, inflammatory stress, or mechanical stress. An increase in ROS activates MAPK or NF-kB or phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of the rapamycin (mTOR) signaling to induce fibrotic changes. Degradation of antioxidant protein nuclear factor erythroid 2-related factor 2 (NRF2) via SMURF1 increases the oxidative stress, further amplifying the fibrotic process. Also, increased ROS is associated with telomere shortening and DNA damage in HLF. In addition, PI3K/AKT/mTOR signaling is also activated by insulin-like growth factor-1 (IGF) (through IGFR), lysophosphatidic acid (LPA) (through LPAR), and A disintegrin and metalloproteinase 10 (ADAM10). Activation of Hedgehog-Gli1 signaling by Wnt1-inducible-signaling pathway protein 1 (WISP1) induces the expression of pro-fibrotic genes. Thrombospondin-1 (THBS1) activates TGF-β1-induced pro-fibrotic signaling. Further, decorin (DCN) and clusterin (CLN), which are induced by mechanical stress, are involved in negative regulation of TGF-β1-induced fibrotic signaling. For clarity, only some selected signaling pathways are included in this figure. Please refer to text for details. (Figure created with BioRender.com, accessed on 1 August 2024).