Abstract
Dupuytren's disease (DD) is a benign, fibroproliferative disease of the palmar fascia, with excessive extracellular matrix (ECM) deposition and over-production of cytokines and growth factors, resulting in digital fixed flexion contractures limiting hand function and patient quality of life. Surgical fasciectomy is the gold standard treatment but is invasive and has associated morbidity without limiting disease recurrence. Injectable Collagenase Clostridium histolyticum (CCH) - Xiaflex® - is a novel, nonsurgical option with clinically proven in vivo reduction of DD contractures but with limited in vitro data demonstrating its cellular and molecular effects. The aim of this study was to delineate the effects of CCH on primary fibroblasts isolated from DD and non-DD anatomical sites (using RTCA, LDH, WST-1, FACS, qRT-PCR, ELISA and In-Cell Quantitative Western Blotting) to compare the efficacy of varying concentrations of Xiaflex® against a reagent grade Collagenase, Collagenase A. Results demonstrated that DD nodule and cord fibroblasts had greater proliferation than those from fat and skin. Xiaflex® exposure resulted in dose- and time-dependent inhibition of cellular spreading, attachment and proliferation, with cellular recovery after enzyme removal. Unlike Collagenase A, Xiaflex® did not cause apoptosis. Collagen expression patterns were significantly (p<0.05) different in DD fibroblasts across anatomical sites - the highest levels of collagen I and III were detected in DD nodule, with DD cord and fat fibroblasts demonstrating a smaller increase in both collagen expression relative to DD skin. Xiaflex® significantly (p<0.05) down-regulated ECM components, cytokines and growth factors in a dose-dependent manner. An in vitro scratch wound assay model demonstrated that, at low concentrations, Xiaflex® enabled a faster fibroblast reparatory migration into the wound, whereas, at high concentrations, this process was significantly (p<0.05) inhibited. This is the first report elucidating potential mechanisms of action of Xiaflex® on Dupuytren fibroblasts, offering a greater insight and a better understanding of its effect in DD.
Highlights
Dupuytren’s Disease (DD) is a common, benign, fibroproliferative disorder affecting the palmar fascia of the hands, resulting in progressively disabling fixed flexion deformities of the digits [1,2,3,4]
The complex aetiological picture is mirrored by the incomplete multi-factorial pathophysiological model - Dupuytren’s disease (DD) manifests with a tripartite natural history of myofibroblast proliferation, cytokine- and mechanically-triggered actin microfilament contraction coupled with extracellular matrix (ECM) remodelling and nodule regression, leaving the inelastic, tendon-like cords which result in the classically-described digital flexion contractures [2,18,19,20,21]
The first pilot assessed the effect of XiaflexH or Collagenase A diluted in complete Dulbecco’s Modified Eagle Medium (cDMEM) to the desired concentrations
Summary
Dupuytren’s Disease (DD) is a common, benign, fibroproliferative disorder affecting the palmar fascia of the hands, resulting in progressively disabling fixed flexion deformities of the digits [1,2,3,4]. The CCH family, first discovered by Maclennon in 1953, comprises a group of matrix metalloproteases that digest the triple-helical structure of collagen under physiological conditions They have had an extensive history of laboratory utilisation but are becoming increasingly of interest in the therapeutic management of a variety of fibrotic disorders, to date most successfully in the amelioration of palpable DD cords [22,23,24]. They contain both class I and class II collagenolytic enzymes, which act to rapidly digest triple-helical collagens into small peptides [25,26]. To date, there have been no in vitro studies published demonstrating the effect of XiaflexH at the cellular or molecular level
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