Chronic Low Back Pain in Young Adults: Pathophysiological Aspects of Neuroinflammation and Degeneration

Abstract

Degenerative disc disease (DDD) is a major cause of chronic low back pain (LBP), yet the molecular mechanisms driving disc degeneration and pain remain poorly understood. This study analyzed intervertebral disc (IVD) tissue from 36 young patients (median age = 36.00 [31.00, 42.50] years) with herniated discs and LBP, alongside healthy controls, to investigate changes in the extracellular matrix (ECM) and neurochemical alterations. Disc degeneration was assessed using MRI (Pfirrmann grading) and histology (Sive's criteria). Histochemical and immunohistochemical methods were used to evaluate aggrecan content, calcification, and the expression of nerve growth factor (NGF), substance P (SP), and S-100 protein. MRI findings included Pfirrmann grades V (30.55%), IV (61.11%), III (5.56%), and II (2.78%). Severe histological degeneration (10-12 points) was observed in three patients. Aggrecan depletion correlated with longer pain duration (r = 0.449, p = 0.031). NGF expression was significantly elevated in degenerated discs (p = 0.0287) and strongly correlated with SP (r = 0.785, p = 5.268 × 10^-9). Free nerve endings were identified in 5 cases. ECM calcification, present in 36.1% of patients, was significantly associated with radiculopathy (r = 0.664, p = 0.005). The observed co-localization of NGF and SP suggests a synergistic role in pain development. These results indicate that in young individuals, aggrecan loss, neurochemical imbalance, and ECM calcification are key contributors to DDD and chronic LBP.

Keywords: NGF; S-100; aggrecan; back pain; calcification; degenerative disc disease; immunohistochemistry; substance P; young age.

Figure 1

Calcification in intervertebral disc (IVD) tissue in patients with varying degrees of IVD degeneration. (A). Hematoxylin-eosin staining. (B). Trichrome staining: left-nucleus pulposus (NP); right-transition from annulus fibrosus to vertebral endplate. (C). Alizarin red staining: upper-NP; lower-annulus fibrosus (left) and hypertrophied chondrocyte clusters in the NP (right). Scale bar for (A–C): 100 µm. (D). Individual calcifications in IVD tissue stained with alizarin red, imaged using a histology scanner. Scale bar for top images: 50 µm; for bottom images: 15 µm. Comparison of grayscale intensity distribution between calcified areas and IVD extracellular matrix (ECM) using the 0–255 scale in ImageJ 1.52u, where 0 corresponds to pure black and 255 to pure white.

Figure 2

Extracellular matrix degradation in degenerative disc disease (DDD): reduced aggrecan content, erosive changes, and fissures. (A). Immunofluorescence staining in nucleus pulposus (NP) regions of DDD and control samples: aggrecan (green), nuclei counterstained with DAPI (blue). (B). Safranin O/Fast Green FCF staining. Safranin O stains cartilage glycosaminoglycans pink to red, while Fast Green FCF counterstains the background bluish green; nuclei appear purple to black. Left: transition zone between NP and annulus fibrosus (AF). Center and right: NP region. (C). Safranin O staining of degenerated NP with chondrocyte clusters. (D). Vertebral endplate in DDD patient; blue staining reflects degeneration. Black-stained chondrocytes on the right may point to pigment accumulation due to cell stress or apoptosis. Scale bar: 100 µm.

Figure 3

Immunohistochemistry (IHC) of NGF, S-100, and Substance P (SP) in NP cells of degenerative disc disease (DDD) patients and controls. IHC was performed using DAB chromogen and Gill’s hematoxylin counterstain. (A). Statistical comparison (Mann–Whitney U test). (B). NGF IHC. (C). S-100 IHC. Arrows indicate nerve ingrowth in patient tissue. (D). SP IHC. Scale bar: 50 µm. *: p < 0.05, **: p < 0.01. ECM—extracellular matrix, NP—nucleus pulposus.

Figure 4

Triple immunofluorescence for S-100 (green), NGF (red), and Substance P (SP, violet) in intervertebral disc (IVD) tissue. (a–g): Samples from degenerative disc disease (DDD) patients. Images show a presumptive mast cell (a) and a lymphocyte (b). A nerve fiber is shown growing near a chondrocyte, with NGF localized nearby; the chondrocyte’s fluorescence is restricted to its nucleus (c). A nerve fiber lacking SP expression (d). Nerve fibers in topographical proximity to SP (e–g). (h–l): Control samples. Non-specific staining was observed in areas of tissue densification, particularly for S-100. Only a few chondrocytes demonstrated visible NGF and SP staining (h–j). Scale bar: 5 µm.

Figure 5

Correlation between clinical-imaging findings and histochemical/immunohistochemical markers (Spearman’s correlation coefficient). Note: FCF—Fast Green FCF stain; SP—substance P; LBP—low back pain; OA—osteoarthritis; SMS—spinal motion segment. Statistical significance in the correlation matrix is denoted as p ≤ 0.05 (*), p < 0.01 (**), and p < 0.001 (***); lack of an asterisk denotes a non-significant correlation (p > 0.05).