INTRODUCTION:

Intervertebral disc (IVD) degeneration can cause chronic back pain and disability and has no available drug treatment till date. Growth differentiation factor (GDF) regulates IVD homeostasis and upregulates the production of healthy nucleus pulposus (NP) cells in degenerated tissues during the repair of disc injury. Moreover, atelocollagen gel (AC) is a cell-supportive scaffold for sustained drug release. Therefore, we hypothesized that GDF6 predominantly inhibits NP cell catabolites in vivo and examined its efficacy, using AC as a carrier, on the structural integrity of the body surface in a rat tail puncture model.

METHODS:

A previously established rat tail annular puncture model for IVD degeneration was used. Continuous disc histological degeneration scale and levels of extracellular matrix components in the rat tails (Co8-11, random order) were compared across the four experimental groups. Group C was the control group, and Groups P, A, and G comprised rats whose tail discs were punctured to a depth of 5 mm using a 20G needle with a handmade stopper, based on the approximate vertical distance from the skin to the center of the NP. The needle was rotated 360° and held in position for 30 s. After puncture, the tail discs of rats in Group A and G were injected with 2 µl AC and 20 µg GDF6, respectively. Catabolic factors, such as tumor necrosis factor-α(TNF-α) and interleukin-6 (IL-6), were evaluated using immunofluorescence on days 14 and 28 after the puncture. Two-way analysis of variance and Tukey post-hoc test were used to compare the degree of alteration between the groups. Statistical significance was set at p<0.05.

RESULTS:

In the control group, TNF-α-positive cells were rarely observed at both time points (C: day 14, 16.2±3.4%; day 28, 16.3±2.9%), and IL-6-positive cells were abundant at both time points (C: day 14, 15.0±4.8%; day 28, 16.7±3.4%). There was a significant increase in the number of TNF-α-positive cells in Groups P and A on day 14 after the puncture, with no significant difference in the count between Groups G and C (P: 28.0±6.0%, p=0.016; A: 27.2±7.0%, p=0.021; G: 23.5±5.3%, p=0.196). IL-6-positive cells showed a similar trend (P: 27.9±7.0%, p=0.02; A: 27.9±7.8%, p=0.02; G: 22.6 ± 5.1%, p=0.256). On day 28 after the puncture, TNF-α-positive cells (P: 71.8±7.4%, p<0.001; A: 71.4±7.9%, p<0.001; G: 33.3±10.5%, p=0. 017) and collagen IL-6-positive cells (P: 68.0±7.7%, p<0.001; A: 67.3±9.6%, p<0. 001, G: 35.7±10.3%, p=0.022) were significantly more in Groups P, A, and G than in Group C. However, this increase was less in Group G than in Groups P (TNF-α: p<0.001, IL-6: p=0.001) and A (TNF-α: p=0.004, IL-6: p=0.003).

DISCUSSION:

A disc degeneration model was created by percutaneously inserting a needle into the middle of the rat tail; however, administration of GDF6 using AC as a carrier prevented the progression of IVD degeneration. Therefore, the protective effect of GDF6 administered through AC on disc NP cells may inhibit IVD degeneration.