Dupuytren's Disease: An Overview
One of our major research interests is Dupuytren's contracture or disease (DD), a benign fibro-proliferative disease of the hand (palmar fascia) (Fig. 1)
Figure 1. Clinical presentation of DD
Contraction of disease cords - collagen-rich scar-like tissue that develop in defined palmar fascia bands - results in fixed, flexed position of the fingers (ring finger in this case). Surgical ressection of the disease tissue followed by extensive physiotherapy to restore full-range of motion is currently the only accepted form of treatment.
- Dupuytren's disease (DD) is studied for several reasons:
- First and foremost, the current lack of knowledge about the biochemical mechanisms of Dupuytren's contracture coupled with the limited therapeutic intervention available (i.e. surgery) emphasizes the need for a better understanding of the basic biology of DD.
- Secondly, DD is associated with other diseases and biological processes that makes it worthy of further study. For example, DD is closely related to other fibro-proliferative disorders, such as Peyronie's disease, aggressive fibromatosis (desmoid tumors), fibrosarcoma, breast fibromatosis, and hypertrophic scars. Moreover, recent epidemiological studies have shown an increase in total mortality and cancer mortality rates in patients with established DD, even after adjustment for age, smoking, and other possible confounding factors.
- Lastly, DD and wound granulation tissue share many biochemical and morphologic characteristics, which has led to the hypothesis that DD may be analogous to a localized, exaggerated wound healing response.
Dupuytren's Disease: Cell Biology
Immunocytochemistry of primary cell cultures reveals distinct morphological and biochemical differences between patient-matched disease and normal fascia cell lines. As shown in Figure 1, disease cells are actively producing fibronectin (Fn), an important extracellular matrix (ECM) protein. By comparison, control cells seem to produce less Fn that is largely comprised of shorter polymerized filaments. These Fn filaments also appear to be more closely associated with the cell. In addition, the disease cells appear to have a more extensive network of intracellular filamentous actin (stress-fibres), a major cytoskeletal component that is involved in cell motility and contraction. These differences in F-actin are also apparent when the cells are cultured in three-dimensions (3D), using a Fibroblast Populated Collagen Lattice (FPCL) assay (Figure 2)
Primary cell lines were derived from surgical tissue - disease (right image) and adjacent, patient-matched normal palmar fascia (left image). Cells (passages 3-6) were plated in 6-well plates (Costar, Corning NY, USA) containing glass coverslips and allowed to adhere overnight. Cells were washed with PBS and then fixed with 4% paraformaldehyde for 1hr at room temperature (RT). Cells were then permeabilized, blocked overnight (PBS + 2% donkey serum, 4 °C), washed (PBS) and then incubated with an anti-fibronectin antibodies (1:1000, clone IST-4, Sigma, St. Louis MO, USA) for 1hr at 4°C. Cells were incubated with Cy3-conjugated secondary antibodies (Jackson Immunoresearch, West Grove PA, USA) for 45 min at RT, washed 2x with PBS, incubated for 20 min. at RT with the nucleic acid stain DAPI (Molecular Probes, Eugene OH, USA), and finally stained for 20 min. at RT with Oregon Green Phalloidin (Molecular Probes, Eugene OH, USA). Coverslips were then briefly washed in PBS and mounted onto glass slides using DAKO faramount (DAKO Diagnostics, Carpinteria CA, USA). Digital images were acquired on a Nikon eclipse TE-200 inverted fluorescent microscope using a Photometrics series 300 cooled CCD camera, and deconvolved using softWoRx (v 2.5) software (Applied Precision Inc., Issaquah WA, USA).
Figure 2. Immunofluorescence of FPCL cultures of DD cells.
Primary cultures derived from disease and normal fascia tissue were cultured in three-dimensional collagen lattices, or FPCLs(see FPCL description), for 2 days. Cells were fixed and stained with Oregon green phalloidin and DAPI to label filamentous actin and the nucleus, respectively. Note the prominent stress fibres in the disease cell cultures. Digital images were acquired on a Nikon eclipse TE-200 inverted fluorescent microscope (10x objective) using a Photometrics series 300 cooled CCD camera, and deconvolved using softWoRx (v 2.5) software (Applied Precision Inc., Issaquah WA, USA).