Inhibition of interleukin-1β-stimulated collagenase and stromelysin expression in human tendon fibroblasts by epigallocatechin gallate ester

The medicinal benefits of green tea (Camellia sinensis) consumption have been attributed to bioavailable polyphenols, notably epigallocatechin gallate (EGCG). We have assessed the effects of EGCG and its non-esterified counterpart EGC on the expression of the collagenases, matrix metalloproteinases (MMP)-1 and -13, and the stromelysin, MMP-3, in human tendon-derived fibroblasts. Interleukin (IL)-1ß increased MMP-1, -3 and -13 mRNA and output at least 30-fold. EGCG reduced this stimulation, by 20–30% at 2.5 µM and more than 80% at 25 µM, and had a smaller effect on MMP-2 mRNA expression, which was not stimulated by IL-1ß. In all experiments EGCG was at least 10-fold more potent than EGC. EGCG reduced the stimulation of p54 JNK/SAPK phosphorylation by IL-1ß but did not affect p38 MAPK phosphorylation, the degradation of I?B or the activating phosphorylation of NF?B. We conclude that EGCG reduces the IL-1-stimulated expression of both collagenase and stromelysin mRNA species, an effect which may be mediated by inhibition of the JNK/SAPK pathway. Taken together with previous reports of EGCG effects on the expression and/or activity of gelatinases and aggrecanases, our results underline the importance of extracellular matrix breakdown as a potential target for the actions of green tea polyphenols

The Regulation of Aggrecanase ADAMTS-4 Expression in Human Achilles Tendon and tendon-Derived Cells

Several members of the ADAMTS (A Disintegrin And Metalloproteinase with ThromboSpondin motifs) family have been identified as aggrecanases, whose substrates include versican, the principal large proteoglycan in the tendon extracellular matrix. We have characterized the expression of ADAMTS-4 in human Achilles tendon and tendon-derived cells. ADAMTS-4 mRNA levels were higher in ruptured tendon compared with normal tendon or chronic painful tendinopathy. In tissue extracts probed by Western blotting, mature ADAMTS-4 (68 kDa) was detected only in ruptured tendons, while processed ADAMTS-4 (53 kDa) was detected also in chronic painful tendinopathy and in normal tendon. In cultured Achilles tendon cells, transforming growth factor-ß (TGF-ß) stimulated ADAMTS-4 mRNA expression (typically 20-fold after 24 h), while interleukin-1 induced a smaller, shorter-term stimulation which synergised markedly with that induced by TGF-ß. Increased levels of immunoreactive proteins consistent with mature and processed forms of ADAMTS-4 were detected in TGF-ß-stimulated cells. ADAMTS-4 mRNA was expressed at higher levels by tendon cells in collagen gels than in monolayer cultures. In contrast, the expression of ADAMTS-1 and -5 mRNA was lower in collagen gels compared with monolayers, and these mRNA showed smaller or opposite responses to growth factors and cytokines compared with that of ADAMTS-4 mRNA. We conclude that both ADAMTS-4 mRNA and ADAMTS-4 protein processing may be differentially regulated in normal and damaged tendons and that both the matrix environment and growth factors such as TGF-ß are potentially important factors controlling ADAMTS aggrecanase activities in tendon pathology

Matrix metalloproteinase activities and their relationship with collagen remodelling in tendon pathology

Our aim was to correlate the activity of matrix metalloproteinases (MMPs) with denaturation and the turnover of collagen in normal and pathological human tendons. MMPs were extracted from ruptured supraspinatus tendons (n=10), macroscopically normal (‘control’) supraspinatus tendons (n=29) and normal short head of biceps brachii tendons (n=24). Enzyme activity was measured using fluorogenic substrates selective for MMP-1, MMP-3 and enzymes with gelatinolytic activity (MMP-2, MMP-9 and MMP-13). Collagen denaturation was determined by a-chymotrypsin digestion. Protein turnover was determined by measuring the percentage of d-aspartic acid (% d-Asp). Zymography was conducted to identity specific gelatinases. MMP-1 activity was higher in ruptured supraspinatus compared to control supraspinatus and normal biceps brachii tendons (70.9, 26.4 and 11.5 fmol/mg tendon, respectively;

Expression profiling of metalloproteinases and tissue inhibitors of metalloproteinases in normal and degenerate human achilles tendon

To profile the messenger RNA (mRNA) expression for the 23 known genes of matrix metalloproteinases (MMPs), 19 genes of ADAMTS, 4 genes of tissue inhibitors of metalloproteinases (TIMPs), and ADAM genes 8, 10, 12, and 17 in normal, painful, and ruptured Achilles tendons. Tendon samples were obtained from cadavers or from patients undergoing surgical procedures to treat chronic painful tendinopathy or ruptured tendon. Total RNA was extracted and mRNA expression was analyzed by quantitative real-time reverse transcription–polymerase chain reaction, normalized to 18S ribosomal RNA. In comparing expression of all genes, the normal, painful, and ruptured Achilles tendon groups each had a distinct mRNA expression signature. Three mRNA were not detected and 14 showed no significant difference in expression levels between the groups. Statistically significant (P < 0.05) differences in mRNA expression, when adjusted for age, included lower levels of MMPs 3 and 10 and TIMP-3 and higher levels of ADAM-12 and MMP-23 in painful compared with normal tendons, and lower levels of MMPs 3 and 7 and TIMPs 2, 3, and 4 and higher levels of ADAMs 8 and 12, MMPs 1, 9, 19, and 25, and TIMP-1 in ruptured compared with normal tendons. The distinct mRNA profile of each tendon group suggests differences in extracellular proteolytic activity, which would affect the production and remodeling of the tendon extracellular matrix. Some proteolytic activities are implicated in the maintenance of normal tendon, while chronically painful tendons and ruptured tendons are shown to be distinct groups. These data will provide a foundation for further study of the role and activity of many of these enzymes that underlie the pathologic processes in the tendon

Immune‐mediated inflammatory diseases (IMIDs) and biologic therapy: a medical revolution

Targeted biologic therapies have revolutionised treatment of immune‐mediated inflammatory diseases (IMIDs) due to their efficacy, speed of onset and tolerability. The discovery that clinically unrelated conditions, such as rheumatoid arthritis and Crohn's disease, share similar immune dysregulation has led to a shift in the management of IMIDs from one of organ‐based symptom relief to mechanism‐based treatment. The fact that anticytokine therapy has been effective in treating multiple orphan inflammatory conditions confirms the IMID paradigm. In this review we examine the biologic agents currently licensed for use in the US and Europe: infliximab, etanercept, adalimumab, rituximab, abatacept, anakinra, alefacept and efalizumab. We also discuss the rationale behind the management of IMIDs using rheumatoid arthritis, Crohn's disease, psoriasis and psoriatic arthritis as examples. For the medical profession, IMID represents a breakthrough in the way pathology is classified. In this burgeoning era of biologic therapy the prospect of complete disease remission is conceivable

Changes in matrix protein biochemistry and the expression of mRNA encoding matrix proteins and metalloproteinases in posterior tibialis tendinopathy

OBJECTIVES: Adult-acquired flat foot secondary to a dysfunctional posterior tibialis tendon (PTT) is often treated by surgical transfer of the flexor digitorum longus tendon (FDLT). In this study, the authors compared normal PTT, stage II dysfunctional PTT and replacement FDLT, aiming to define changes in collagen modification, glycosaminoglycan (GAG) and the expression of matrix and metalloproteinase mRNA. METHODS: Normal PTTs were obtained from patients with no history of tendon problems. Samples of dysfunctional PTT and replacement FDLT tissue were obtained from patients undergoing surgical reconstruction. Tissue samples were analysed for total collagen and GAG, pentosidine and collagen cross-links. Total RNA was assayed for mRNA encoding matrix proteins and metalloproteinases, using real-time reverse transcription PCR. Differences between clinical groups were assessed using non-parametric statistics. RESULTS: Dysfunctional PTT contained higher levels of GAG and lower levels of pentosidine than normal PTT or FDLT. In contrast, collagen in FDLT contained fewer ketoimine and more aldimine cross-links than either normal or dysfunctional PTT. mRNA encoding types I and III collagens, aggrecan, biglycan, matrix metalloproteinase (MMP)-2, -13 and -23, and a disintegrin and metalloproteinase (ADAM)-12L each showed increased levels in dysfunctional PTT compared with either normal PTT or (except MMP-13) FDLT. In contrast, MMP-3 and ADAM with thrombospondin domain (ADAMTS)-5 mRNA were lower in both dysfunctional PTT and FDLT than in normal PTT, while ADAMTS-1 mRNA was lower in dysfunctional PTT than in FDLT. CONCLUSIONS: Stage II dysfunctional PTT shows biochemical and molecular changes consistent with a chronic remodelling of the extracellular matrix, rather than rupture, while the replacement FDLT resembles normal PTT in many, but not all, parameters