Smad7 Is Highly Expressed in Human Degenerative Discs and Participates in IL-1β-Induced Apoptosis of Rat AF Cells via the Mitochondria Pathway

Abstract

Background. Abnormal Smad7 expression can lead to apoptosis in different cell types. Previously, we found high expression of Smad7 in rat degenerative discs. However, the exact role of Smad7 in the apoptosis of disc cells and the possible underlying mechanism remain unclear. Methods. Degenerative and nondegenerative human lumbar intervertebral discs were collected from patients during operation. The expressions of SMAD7 mRNA and protein in the different components of these discs were measured with real-time PCR and Western blotting, respectively. Annulus fibrosus (AF) cells were isolated and cultivated from the discs of young healthy rats. Smad7 in the AF cells was overexpressed with adenovirus and knocked down with siRNA. IL-1β was used to induce apoptosis in the AF cells. Loss-and-gain cell function experiments were performed to show the effect of Smad7 on the apoptosis of AF cells. The function recovery experiments were performed to verify whether Smad7 regulates the apoptosis of AF cells through the mitochondria-mediated pathway. Results. Both the mRNA and protein expressions of Smad7 were significantly higher in the different components of human degenerative discs than in those of the nondegenerative discs. IL-1β stimulated apoptosis while upregulating the Smad7 expression in the AF cells in vitro. Overexpression of Smad7 in AF cells exaggerated the IL-1β-induced apoptosis in the cells while knockdown of Smad7 expression suppressed this apoptosis. With the exaggerated apoptosis in the AF cells with Smad7 overexpression, both active cleaved caspase-3 and cleaved caspase-9, the ratio of Bax/Bcl-2, and Cyt-c increased significantly. However, the inhibitor of caspase-9, Z-LEHD-FMK, significantly diminished the apoptosis in these cells. Conclusion. Smad7 is highly expressed in human degenerative discs and participates in IL-1β-induced apoptosis of rat AF cells via the mitochondria pathway. Smad7 may be a potential target for the prevention and treatment of degenerative disc disease.

Role of signal transduction pathways in IL‐1β‐induced apoptosis: Pathological and therapeutic aspects

“…The mature IL-1βthen binds to the type 1 IL-1 receptor (IL-1R1) to exert biological functions. In experiments related to IL-1β-induced apoptosis of nucleus pulposus cells 12 and intervertebral disc cells 13 changes in protein expression levels of Bcl-2, Bax, caspase-9, and caspase-3 can be detected, so it is inferred that IL-1β significantly increases the proapoptotic protein Bax and decreases the antiapoptotic protein Bcl-2. 6,14 The ratio of Bax to Bcl-2 affects the mitochondrial membrane potential leaving the mitochondrial permeability transition pore open and the mitochondria highly impaired.…”

Isoorientin ameliorates H2O2-induced apoptosis and oxidative stress in chondrocytes by regulating MAPK and PI3K/Akt pathways

“…Key proteins in the mitochondrion were quantified using western blot assays. As a result, mitochondrial proteins (Bax, Bcl-2, caspase-3, and cleaved caspase-3) were identified [ 26 ]. The H 2 O 2 -induced more Bax and more cleaved caspase-3 while less Bcl-2 in chondrocytes.…”