Expression and role of connective tissue growth factor in painful disc fibrosis and degeneration
From: Spine. 2009 Mar 1;34(5):E178-82
Low back pain is a common clinical symptom, which will affect majority of the population at some time during the course of their lives. Although the causes of the low back pain remain unclear at present, theoretically, each structural component of the lumbar spine that is innervated, such as the vertebrae, intervertebral discs, facet joints, muscles and ligaments, may be the origin of low back pain. The degree of disc degeneration and the incidence of low back pain increase in parallel with age, suggesting that disc degeneration may be the main cause of low back pain. However, disc degeneration is common in patients without low back pain, particularly in those aged older than 50 years. Disc degeneration usually appears in magnetic resonance imaging (MRI) T2-weighted images as a decline in signal intensity, i.e., the so-called black disc. Both a normal aging disc and a pathologically painful disc appear as an area of decreased signal intensity on T2-weighted images. MRI cannot differentiate between these 2 disease entities. Many studies have reported regarding the histologic changes and the possible mechanisms underlying normal age-related disc degeneration. However, the main histologic changes and the exact molecular mechanisms underlying the painful pathologic disc remain unknown.
Recent researches have shown that connective tissue growth factor is the downstream effector mediated by transforming growth factor-β1, and is closely associated with the regulation of cell proliferation and differentiation and the fibrosis of tissues and organs, and can induce the in vivo expression of the gene involved with fibroblast extracellular matrix composition. This article reports the histologic findings of surgically excised specimens from patients with discogenic low back pain and studies the connective tissue growth factor expression and distribution in the disc, using immunohistochemical staining, with the aim of investigating the histologic characteristics of the painful pathologic disc, and exploring the role of connective tissue growth factor in painful disc degeneration.
Connective tissue growth factor was first discovered in 1991 by Bradham et al in a conditioned culture medium of human umbilical vein endothelial cells. It is a cysteine-rich secreting peptide encoded by immediate early genes. Connective tissue growth factor is synthesized and secreted by fibroblasts, smooth muscle cells, and endothelial cells. Many studies have proved that connective tissue growth factor, the putative downstream efficient response element of transforming growth factor-β1, only mediates a part of the biologic effects of transforming growth factor-β1, i.e., stimulation of fibroblast proliferation, and promotion of tissue and organ fibrosis. These effects have been confirmed in studies regarding a variety of fibrosis diseases such as renal fibrosis and liver cirrhosis. Connective tissue growth factor promotes the proliferation and activation of fibroblasts by activating signaling pathways. It stimulates the synthesis of Type I and III collagen via the respondent elements of the precollagen α2 gene. Further, it is involved in the production and accumulation of extracellular matrices. Because of these, it promotes the occurrence and development of fibrosis diseases.
Disc degeneration is a very complicated biologic process. Previous views on disc degeneration and the mechanism underlying it were mainly based on histologic and biochemical studies, using human disc herniation specimens from surgery, animal models of aging, and degenerative discs. With the development and popularization of lumbar fusion, a greater number of painful pathologic disc specimens can be obtained, which are beneficial for studies regarding the pathogenesis of painful disc degeneration. Based on our present histologic studies, we found that the composition and structure of painful disc differed from those of nonpainful degenerative disc. Specifically, normal fibroblasts in the anulus fibrosus were replaced by cartilage-like cells. The anulus fibrosus lamellar structure was disordered and fractured. The normal highly hydrated gelatin-like nucleus pulposus, whose matrices showed obvious fibrosis, and cartilage-like cells, were completely replaced with fibroblasts, was substituted by fibrous tissues. The histologic changes in the nucleus pulposus were divided into 3 major types: obvious fibrosis, vascular invasion, and inflammatory granulation tissue formation. In our previous studies, we found that the characteristic change in painful pathologic discs was the formation of inflammatory vascular granulation tissues with extensive innervation along the tears in the posterior anulus fibrosus, along with mass expression of some growth factors such as basic fibroblast growth factor and transforming growth factor-β1. Vascular granulation tissue was not formed in aging discs, and only a few growth factors were expressed. Our studies suggested that these pathologic processes were associated with anulus fibrosus injury and subsequent repair processes, unlike normal asymptomatic disc degeneration. Moreover, we conclusively determined from this study that connective tissue growth factor expression remains silent in normal discs, is weakly positive in asymptomatic degenerative discs, and is significantly increased in painful discs, suggesting that connective tissue growth factor is closely related to disc fibrosis and subsequent degeneration processes.
In the repair processes of injuries and fibrosis diseases, extracellular matrix deposition and angiogenesis often appear synchronously. Connective tissue growth factor in chicken embryo chorioallantoic membranes can induce mild angiogenesis. Connective tissue growth factor injected in the backs of rats can also promote angiogenesis. Aggrandized mCTGF protein stimulates adhesion and migration of human dermal microvascular endothelial primary cells and enhances their survival rate by inhibiting apoptosis. Recombinant connective tissue growth factor has also induced proliferation, adhesion, migration, and remodeling of bovine aortic endothelial cells. Painful disc fibrosis, vascular invasion, and granulation tissue formation might be closely related to the role of connective tissue growth factor in stimulating fibroblast proliferation and inducing angiogenesis.
Growth factors control cell proliferation or differentiation in vitro and mediate interaction between cells in vivo. They control not only the development and growth of the organism, the regenesis, and healing of tissue injuries, but also may facilitate abnormal changes in tissues. The author’s study showed that there were strong connective tissue growth factor expressions in painful disc, only weak expressions in asymptomatic degenerative disc, and no expression in normal control disc. Disc tissues are different from other tissues because they comprise the largest avascular tissue. In other tissues, injury healing proceeds from the inside to the outside. On the contrary, healing in disc tissues proceeds from the outside to inside, since only the outer anulus fibrosus and longitudinal ligaments of discs possess vascular distribution. When the anulus fibrosus is lacerated or injured, vascular tissues can only gradually develop from the outer to the inner anulus fibrosus. Endothelial cells migrating into discs form the principal parts of a new capillary vessel. With the help of various growth factors, endothelial cells migrating into the avascular disc tissues differentiate, proliferate, and gradually form complicated capillary networks. The endothelial cell itself can also produce plasminogen activators and collagenases, facilitating its migration. This study suggests that as anulus fibrosus injuries stimulated local vascular inflammatory reactions, cells in inflammatory regions produce a large number of growth factors such as basic fibroblast growth factor, transforming growth factor-β1, and connective tissue growth factor. These growth factors acted in the intervertebral disc cells which separated from the circulatory system and promoted disc cell differentiation, proliferation, and large-scale extracellular matrix synthesis, via signal transduction. This may be the main cause of disc fibrosis and degeneration.
The relatively simple role of connective tissue growth factor, which was mainly expressed in mesenchymal cells, was primarily confined to connective tissues. Compared with the upstream control component transforming growth factor-β1, the biologic effects of connective tissue growth factor were more limited in fibrosis. Therefore, the authors speculated that blocking connective tissue growth factor functions may provide a novel strategy for biologic treatment of degenerative discs.
Interestingly, an article in Nan Fang Yi Ke Da Xue Xue Bao. 2009 Feb;29(2):316-8, Overexpression of transforming growth factor-beta1 in degenerative ligamentum flavum, indicates; Degenerative ligamentum flavum shows hypertrophy and fibrosis, and transforming growth factor -beta1 overexpression may be associated with in the development and progression of ligamentum flavum degeneration in the lumbar spine.
To summarize:
Painful discs showed chronic inflammatory reaction with blood vessel infiltration in varying degrees. The anulus fibrosus had lost its normal lamellar architecture, and instead, disorganization, disruption, and crossed fusion were observed. Normal fibroblasts were replaced by chondrocytes in the anulus fibrosus.
The nucleus pulposus showed marked fibrosis, blood vessel infiltration, and inflammatory granulation tissue formation.
Immunohistochemical staining demonstrated strong connective tissue growth factor expression in the painful discs, weak expression in the asymptomatic degenerative disc, and no expression in the control discs.
