Oral Presentation 50th International Society for the Study of the Lumbar Spine Annual Meeting 2024

INTRODUCTION

Low back pain (LBP) is a widespread ailment affecting approximately 11.9% of the global population. Inflammation-associated processes impair the function of nucleus pulposus cells (NPCs) and promote angiogenesis and sensory nerve fiber growth into the disrupted fibrous ring of the nucleus pulposus, contributing to mechanical and chemical sensitization. p300, a 300 KDa acetyltransferase, and its counterpart CREB-binding protein (CBP) plays a crucial role in gene transcription, inflammation, organ development, and tumor formation. This study aims to investigate the role of p300-IL-4/IL-1β in IVDD and pain and elucidate the underlying molecular mechanism. 

METHODS

Human intervertebral disc samples were collected and graded based on the Pfirrmann grading scheme. A rat model of IVDD was established through posterior disc puncture, and various treatments were administered. Human NPCs were isolated and cultured, followed by assessments including qRT-PCR, Western blotting, immunofluorescence, and histopathological analyses. The mechanical and thermal hyperalgesia were assessed using von Frey hair tests and the plantar test, respectively. Chromatin immunoprecipitation (ChIP) and ChIP-qPCR were performed, and lentivirus transduction was used for p300 knockdown. 

RESULTS

We investigated the role of CBP/p300 in IVDD. Degenerated nucleus pulposus (NP) tissues from IVDD patients exhibited a significant downregulation of p300 at both nucleic acid and protein levels, while CBP levels remained unchanged. Manipulating p300 expression in NPCs regulated matrix production, with p300 overexpression enhancing aggrecan and collagen II expression and knockdown leading to increased matrix-degrading proteins. In a rat IVDD model, activating p300 function with CTPB alleviated IVDD and pain, while inhibiting p300 with C646 exacerbated IVDD and pain. Transcriptome analysis implicated IL-4 as a downstream effector of p300, and IL-4 demonstrated protective effects against IVDD and pain. Mechanistically, p300 was found to acetylate H3K27 at the IL-4 promoter, promoting its expression.

DISCUSSION

The novel findings of this study suggest that the p300-IL-4/IL-1β axis may represent a promising therapeutic target for IVDD and LBP. These results propose a hypothesis that early in the inflammatory phase, NPCs may initiate a self-protective mechanism through p300, aiding itself in preserving a favorable metabolic microenvironment. This mechanism is primarily mediated by the expression of IL-4, which has the capacity to counteract IL-1β, rather than TNF-α.

One figure of the manuscript

IL-4 has a therapeutic effect on IVDD and pain behavior. (A-B) Morphological staining and quantitative analysis of the effect of IL-4 and dupilumab on disc degeneration in vivo. IHC staining of TNF-α and IL-1β positive cells in sham, puncture, puncture + IL-4 and puncture + IL-4 + dupilumab group. Scale bar: 100 μm. n = 8 per group. (C-D) Quantitative analysis of TNF-α and IL-1β positive cells in (A). (E-F) IHC staining and quantitative analysis of CGRP in DRGs. Scale bar: 100 μm. n = 8 per group. (G-H) The 50% withdrawal threshold and thermal withdrawal threshold in puncture model of sham, puncture, puncture + IL-4 and puncture + IL-4 + dupilumab group. (I) Schematic diagram illustrating p300/IL-4 alleviate IVDD and pain by inhibiting the inflammatory microenvironment.