Extracellular matrix remodeling in experimental intervertebral disc degeneration

Abstract

Objective: To evaluate the remodeling of the extracellular matrix in intervertebral disc degeneration through the experimental model of intervertebral disc degeneration.

Methods: The model of disc degeneration induction, using needle 20G and 360° rotation, was applied for 30 seconds between the 6(th)/7(th), and 8(th)/9(th) coccygeal vertebrae of Wistar rats. The intermediary level, between the 7(th) and 8(th) vertebrae, was taken as control, not being subjected puncture. The distribution of the extracellular matrix components involved in the remodeling and inflammation process, such as proteoglycans (aggrecan, decorin, biglycan), growth factors (TGFβ), heparanase isoforms (HPSE1, HPSE2), metaloprotesasis-9 (MMP9) and interleukins (IL-6, IL-10) was analyzed during the post-injury period (15 to 30 days) and in the control group (discs collected immediately after the puncture, day zero). On the 15(th) day, acute phase of the disease, a reduced expression of extracellular matrix components had been observed, whilst there were no differences in the interleukins expression. At 30 days, the molecules followed a very similar pattern of expression in the control group (not affected by disc degeneration).

Results: The results show that during the acute phase significant alterations in the extracellular matrix components occur and in the late phase intervertebral disc returns to a profile similar to noninvolved tissue, probably due to extensive remodeling process of the extracellular matrix that is capable of regenerating the damaged tissue.

Conclusion: : The experimental model used demonstrated the occurrence of significant changes in the extracellular matrix during the period analyzed after induction of intervertebral disc degeneration. Laboratory investigation.

Keywords: Interleukins; Intervertebral disc degeneration; Proteoglycans; Rats, Wistar.

Figure 1. Immunohistochemistry reactions for analysis of expression of proteoglycans and TGF-β. (A, B and C), tissues marked with anti-aggrecan antibody 4F4; (D, E and F), tissues marked with primary antibody anti-decorin N-15; (G, H and I), Tissues marked with primary antibody anti-biglycan L-15; (J, L and M), tissues marked with primary antibody anti-TGFβ1(sc-146).

Figure 2. The values indicate the digital quantification of the immunohistochemistry reactions of the proximal and distal intervertebral discs of rats, after 0 days; 15 days and 30 days of degeneration. The values indicate the index of expression (IE), obtained by digital quantification after immunohistochemistry reactions.

Figure 3. Immunohistochemistry reactions for analysis of expression of heparanase isoforms (HPSE1 and HPSE2) and metaloproteinase-9 (MMP-9). (A, B and C), tissues marked with primary antibody anti-HPSE1 (HPA1 L-19); (D, E and F), tissues marked with primary antibody anti-HPSE2 (HPA2 C-17); (G,H and I), tissues marked with primary antibody anti-MMP-9 (H-129).

Figure 4. The values indicate the digital quantification of the immunohistochemistry reactions of the proximal and distal intervertebral discs of rats, after 0 days; 15 days and 30 days of degeneration. The values indicate the index of expression (IE), obtained by digital quantification after immunohistochemistry reactions.

Figure 5. Immunohistochemistry reactions for analysis of the expression of interleukins (IL-6 and IL-10). (A, B and C), tissues marked with primary antibody anti-interleukin-6 (sc-130326); (D, E and F), tissues marked with primary antibody anti-interleukin-10 (H-160).

Figure 6. The values indicate the digital quantification of the immunohistochemistry reactions of the distal and proximal intervertebral discs of rats, after 0 days; 15 days and 30 days of degeneration. The values indicate the index of expression (IE), obtained by digital quantification after immunohistochemistry reactions.