Myofibroblast Distribution in Dupuytren's Cords: Correlation With Digital Contracture

doi:10.1016/j.jhsa.2009.08.005

The Journal of Hand Surgery

Volume 34, Issue 10, December 2009, Pages 1785-1794

https://doi.org/10.1016/j.jhsa.2009.08.005 Get rights and content

Purpose

Dupuytren's tissue has typically been described as being composed of myofibroblast-rich palmar nodules and relatively acellular tendon-like cords. We aimed to determine myofibroblast distribution (alpha-smooth muscle actin [α-SMA] positive cells) within Dupuytren's tissue and to correlate histologically defined α-SMA-positive nodules with digital contracture and recurrent disease.

Methods

One hundred and three digital Dupuytren's cords (72 fasciectomy, 31 dermofasciectomy) were stained with anti–α-SMA antibody. The presence of α-SMA–positive nodules, their surface area, and α-SMA–positive cells were quantified throughout excised Dupuytren's tissue. Clinical data on diathesis, flexion deformity, and previous surgeries were collected.

Results

Cords were nodular (66%) or non-nodular (34%). Nodular cords contained 1 (55%), 2 (33%), or 3 or more nodules (12%) composed of localized collections of cells. The mean total nodule surface area was 23 mm² (range, 1.3–105 mm²). Nodules contained the highest number of α-SMA–positive cells (mean 97%, 2374 cells/mm²) compared to peri-nodular areas (mean 32%, 763 cells/mm²), and more distant cord (mean 8%, 495 cells/mm²). Non-nodular cords contained 9% to 17% α-SMA–positive cells (mean 475–663 cells/mm²), with higher numbers distally. There was greater digital contracture in patients with non-nodular cords. Thirty-six of 38 proximal interphalangeal (PIP) joint–marked samples had a nodule that co-localized with the PIP joint. Nodule size did not correlate with flexion deformity or with primary or recurrent disease.

Conclusions

We found that two thirds of digital cords were nodular. Nodules were hypercellular, the majority being α-SMA–positive cells. Nodules varied in size and co-localized with the PIP joint. Cord was relatively cellular throughout; a proportion of these cells were α-SMA–positive and cells aligned with collagen fibers. Non-nodular cords correlated with significantly greater digital flexion contracture. We propose that cells in nodular cords contract and deposit extracellular matrix components. The matrix is then remodeled in shortened configuration, and as fixed flexion deformity develops, stress shielding eventually leads to myofibroblast apoptosis, and cord becomes less cellular.

Section snippets

Patient samples

In total, 103 digital cords were excised from 80 patients. The surgery was performed by 2 surgeons (J.N. and D.D.) between 2006 and 2008. Patients were recruited to the study after providing written consent. Specimens were collected from 2 sites: London and Edinburgh. The mean age in both groups was 64 years (range, 33–88 y) with an overall ratio of men to women of 3:1. Digital cords were mainly from ulna-sided disease (62, little finger; 30, ring finger; 7, middle finger; 2, index finger; and

Nodularity

Overall, two thirds of all samples were nodular. The size and number of nodules were extremely variable, with marked heterogeneity between samples (Fig. 2). Of the nodular samples, 55% had 1 nodule, 33% had 2 discrete nodules, and 12% had 3 or more nodules. Of the samples from London, 90% were nodular, compared to 55% from Edinburgh. The mean total nodular surface area was 23 mm² (range, 1.3–105 mm²), and the mean total cord surface area was 143 mm² (range, 20–388 mm²). Nodular cords were seen

Discussion

The comparison of Dupuytren's samples from Edinburgh and London highlighted differences between patients from the 2 sites. Of the 103 digital cords analyzed, the majority were from Edinburgh (n = 74), and the remainder from London (n = 29). In London, 90% of patients were men, the digits tended to be less flexed (mean flexion deformity, 63° ± 26°), and 3 out of every 4 patients presented with primary disease. Most of the patients (26/29) were treated by fasciectomy, and the remaining 3 by

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Normal human dermal fibroblasts (NHDFs) and DMFs from Dupuytren patients were isolated and cocultured with ASCs and PRP either alone or together. Adipocytes were detected by Oil Red O and perilipin staining. DMFs and NHDFs were transplanted into the forepaws of rats (Rowett Nude [rnu/rnu]) and treated with saline, PRP+ASCs, or collagenase Clostridium histolyticum (clinical comparison) 2 months later. After 2 weeks, the tissue was harvested and subjected to Masson trichrome staining, and collagen I and III and alpha-smooth muscle actin detection by immunohistochemistry.
Myofibroblasts transform into adipocytes upon coculture with PRP+ASCs. DMFs show increased alpha-smooth muscle actin expression in vivo compared with NHDFs, which is significantly decreased after PRP+ASCs and collagenase Clostridium histolyticum treatments. DMFs induce collagen I and III expressions in rat paws compared with NHDFs, with a type III to I ratio increase. Treatment with PRP+ASC reduced the ratio, but collagenase Clostridium histolyticum did not.
Treating DMFs with PRP+ASCs provides factors that induce myofibroblast to adipocyte transformation. This treatment reduces the contractile phenotype and fibrosis markers in vivo. Future studies should detail the mechanism of this conversion.
The combination of PRP and ASCs to induce the differentiation of DMFs into adipocytes may serve to limit surgery to a percutaneous contracture release and biological injection, rather than a moderate or radical fasciectomy, and reduce the recurrence of Dupuytren contracture.
Dupuytren's disease: a localised and accessible human fibrotic disorder
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Citation Excerpt :
As the fixed flexion deformity develops, myofibroblast numbers decrease, perhaps through apoptosis, and the cords become less cellular. The formation and shortening of the cord by the myofibroblasts are necessary for the development of flexion contractures that ultimately compromise hand function [32]. The predominant matrix proteins in DD are collagens, and the subtypes reported include collagen types I, III, V, and VI [16,63,64].
We review the biology of Dupuytren's disease (DD), a common localised fibrotic disorder of the hand. The disease develops through a complex interplay of genetic and environmental factors, and epigenetic signalling. The early-stage disease nodules comprise a complex milieu of stromal and immune cells which interact to promote disease development. Recently, inhibition of tumour necrosis factor (TNF) locally resulted in softening and a decrease in nodule size, potentially controlling disease progression. Unlike fibrotic disorders of the visceral organs, the easy access to tissue in DD patients enables dissection of the cellular landscape and molecular signalling pathways. In addition, the study of DD may have wider benefits in enhancing our understanding of less-accessible fibrotic tissues.
Anti-tumour necrosis factor therapy for early-stage Dupuytren's disease (RIDD): a phase 2b, randomised, double-blind, placebo-controlled trial
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Dupuytren's disease is a common fibrotic condition that causes the fingers to flex irreversibly into the palm. Treatments for late-stage disease all have limitations, and there is no approved treatment for early-stage disease. We identified tumour necrosis factor as a therapeutic target in Dupuytren's disease, and in a dose ranging trial found 40 mg adalimumab in 0·4 mL to be most efficacious. Here we aimed to assess the effects of intranodular injection of adalimumab in early-stage disease.
In this phase 2b, randomised, double-blind, placebo-controlled trial adults with early-stage Dupuytren's disease and an established clinically distinct nodule with a clear history of progression in the preceding 6 months were recruited from two clinical centres in the UK and were randomly assigned 1:1 to receive four injections of adalimumab or saline every 3 months. Participants and assessors were masked. The primary outcome was nodule hardness measured with a durometer at 12 months. Data were analysed by linear mixed effects regression models in the intention-to-treat population with multiple imputation for missing primary outcome data. The trial is registered at the ISRCTN registry, ISRCTN 27786905 and is complete.
Between Feb 17, 2017, and Jan 11, 2019, 284 participants were screened in the UK and 140 were enrolled. 47 (34%) participants were female and 93 (66%) were male. Mean age of participants was 59·7 years (SD 10·0). Primary outcome data were available from 113 participants. Nodule hardness was lower (−4·6 AU [95% CI −7·1 to −2·2], p=0·0002) in the adalimumab compared with the saline group at 12 months. There were no related serious adverse events; the most common adverse events were minor injection site reactions.
Intranodular injections of adalimumab in participants with early-stage Dupuytren's disease resulted in softening and reduction in size of the nodules. Longer follow-up would be required to assess the effect of tumour necrosis factor inhibition on disease progression, extension deficit and hand function.
Health Innovation Challenge Fund (Wellcome Trust, Department of Health) and 180 Life Sciences.
Stem cell research in tissue engineering and translational medicine
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The utilization of stem cells for applications in a variety of fields related to regenerative medicine and tissue engineering is an exciting research topic. An assortment of mesenchymal stem cells (MSCs), including adipose-derived stem cells (ASCs) and bone marrow-derived stem cells (BMSCs), provide an ideal source for tissue engineering applications due to the absence of ethical concerns, their high availability, and the increasing number of methods for their isolation and expansion. ASCs and BMSCs possess the same multipotency, but substituting adipose tissue for bone marrow, as a source of stem cells for regenerative medicine, is greatly advantageous. ASCs are simple to procure via a method that is less invasive compared to BMSCs. In addition, the percentage of cells from adipose tissue with multipotency is high, mass culture is simple, and the cells function in a manner that is similar, and often better, than BMSCs. Thus human ASCs hold great potential in the field of regenerative medicine. In this chapter, we examine the utilization of ASCs for cartilage tissue engineering, to combat fibrosis/scarring, to improve fat grafting techniques, and further explore utilizing the ASC secretome as a cell-free modality for tissue engineering. Altogether, the regenerative potential of ASCs for tissue engineering is clear, and ultimately, further studies on these topics will provide innovative mechanisms for alleviating some of the limitations that are currently faced in the tissue engineering arena.
Epidermal Paracrine Signals May Regulate Dupuytren Contracture Myofibroblasts
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Dupuytren’s disease (DD) is a common, progressive fibroproliferative disease affecting the palmar fascia of the hands, causing fingers to irreversibly flex toward the palm with significant loss of function. Surgical treatments are limited; therefore, effective new therapies for DD are urgently required. To identify the key cellular and molecular pathways driving DD, we employed single-cell RNA sequencing, profiling the transcriptomes of 35,250 human single cells from DD, nonpathogenic fascia, and healthy dermis. We identify a DD-specific population of pathogenic PDPN⁺/FAP⁺ mesenchymal cells displaying an elevated expression of fibrillar collagens and profibrogenic genes. In silico trajectory analysis reveals resident fibroblasts to be the source of this pathogenic population. To resolve the processes governing DD progression, genes differentially expressed during fibroblast differentiation were identified, including upregulated TNFRSF12A and transcription factor SCX. Knockdown of SCX and blockade of TNFRSF12A inhibited the proliferation and altered the profibrotic gene expression of cultured human FAP⁺ mesenchymal cells, demonstrating a functional role for these genes in DD. The power of single-cell RNA sequencing is utilized to identify the major pathogenic mesenchymal subpopulations driving DD and the key molecular pathways regulating the DD-specific myofibroblast phenotype. Using this precision medicine approach, inhibition of TNFRSF12A has shown potential clinical utility in the treatment of DD.

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: This work has been funded by grant awards from The Healing Foundation in association with the British Society for Surgery of the Hand, the Hammersmith Hospitals Trustees' Research Committee, and the Kennedy Institute of Rheumatology Trustees, Imperial College London. We are also grateful for support from the NIHR Biomedical Research Centre funding scheme.

: No benefits in any form have been received or will be received related directly or indirectly to the subject of this article.

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Scientific articleMyofibroblast Distribution in Dupuytren's Cords: Correlation With Digital Contracture

Purpose

Methods

Results

Conclusions

Section snippets

Patient samples

Nodularity

Discussion

J Hand Surg

Tissue Cell

Hand

J Hand Surg

J Hand Surg

J Hand Surg

J Hand Surg

J Hand Surg

Hand Clin

J Invest Dermatol

Eur J Cell Biol

J Hand Surg

J Hand Surg

J Hand Surg

J Surg Res

J Hand Surg

Br J Plast Surg

Scientific article
Myofibroblast Distribution in Dupuytren's Cords: Correlation With Digital Contracture