A bioengineered living cell construct activates metallothionein/zinc/MMP8 and inhibits TGFβ to stimulate remodeling of fibrotic venous leg ulcers

Abstract

Venous leg ulcers (VLU) represent a major clinical unmet need, impairing quality of life for millions worldwide. The bioengineered bilayered living cell construct (BLCC) is the only FDA‐approved therapy demonstrating efficacy in healing chronic VLU, yet its in vivo mechanisms of action are not well understood. Previously, we reported a BLCC‐mediated acute wounding response at the ulcer edge; in this study we elucidated the BLCC‐specific effects on the epidermis‐free ulcer bed. We conducted a randomized controlled clinical trial (http://clinicaltrials.gov NCT01327937) enrolling 30 subjects with nonhealing VLUs, and performed genotyping, genomic profiling, and functional analysis on wound bed biopsies obtained at baseline and 1 week after treatment with BLCC plus compression or compression therapy (control). The VLU bed transcriptome featured processes of chronic inflammation and was strikingly enriched for fibrotic/fibrogenic pathways and gene networks. BLCC application decreased expression of profibrotic TGFß1 gene targets and increased levels of TGFß inhibitor decorin. Surprisingly, BLCC upregulated metallothioneins and fibroblast‐derived MMP8 collagenase, and promoted endogenous release of MMP‐activating zinc to stimulate antifibrotic remodeling, a novel mechanism of cutaneous wound healing. By activating a remodeling program in the quiescent VLU bed, BLCC application shifts nonhealing to healing phenotype. As VLU bed fibrosis correlates with poor clinical healing, findings from this study identify the chronic VLU as a fibrotic skin disease and are first to support the development and application of antifibrotic therapies as a successful treatment approach.

miR-19a/b and miR-20a Promote Wound Healing by Regulating the Inflammatory Response of Keratinocytes

“…Compared with the human normal wounds at the inflammatory phase (NW1) or the proliferative phase (NW7), we found that the expression of miR-19a/b and miR-20a were significantly downregulated in all the three major types of chronic wounds, that is, VU, DFU, and PU, which may contribute to sustained inflammation and impaired healing there. Interestingly, recent studies have revealed beneficial effects of converting chronic wound microenvironment to a healing milieu similar to that in an acute wound (Stone et al, 2020(Stone et al, , 2017. In line with this, our study suggested that the replenishment of miR-19b and miR-20a in wounds with a deficiency of these miRs has therapeutic potential.…”

Gut–Skin Axis: Current Knowledge of the Interrelationship between Microbial Dysbiosis and Skin Conditions

“…In addition, several processes such as angiogenesis, ECM remodelling, and induction of stem cells are hindered. Chronic wounds were also continuously inflamed, as was demonstrated by several studies [308,[314][315][316][317][318]. However, the underlying cellular and molecular mechanisms of impaired wound healing are not yet fully understood.…”

Genomics of Human Fibrotic Diseases: Disordered Wound Healing Response

“…In venous leg ulcers (VLU) in particular, the wound bed is histopathologically characterized by disorganized ECM, marked fibrosis, and chronic inflammatory infiltrates, all of which contribute to impaired healing; in fact, increased presence of dense fibrosis and high mature collagen levels in VLUs correlate with poor healing outcomes in the clinical setting [ 74 ]. Using microarray profiling, our group reported enrichment of inflammatory response and fibrogenetic pathways in non-healing VLU [ 13 ], corresponding to the histological findings of disorganized ECM, fibrosis, and chronic inflammation [ 73 , 74 , 75 ]. Specifically, we observed enrichment of collagens and secreted matricellular proteins in conjunction with upregulation of fibronectin (FN1), tenascin (TNC), osteopontin (SPP1), connective tissue and hepatocyte growth factors (CTGF, HGF), and PAI-1 (SERPINE1).…”

“…We extended our findings in a randomized controlled post-marketing clinical trial examining the mechanism of action of a bioengineered bilayered cellular construct (BLCC), a commercial skin substitute with demonstrated efficacy in promoting VLU closure. BLCC application triggered an acute wound healing response at the ulcer edge to reverse chronic inflammation [ 76 ] while coordinately stimulating remodeling of the ulcer bed, as evidenced by decreased expression of profibrotic TGFß1 gene targets, increased levels of TGF-ß inhibitor decorin, and endogenous release of matrix metalloproteinase (MMP)-activating zinc to stimulate antifibrotic remodeling [ 13 ]. As such, the development and application of anti-fibrotic therapies represent a novel treatment approach for VLUs and other non-healing ulcers.…”

“…Dysregulation of these processes in any connective tissue-containing organ can cause excessive ECM deposition and tissue fibrosis, ultimately leading to loss of organ function [ 6 ]. As an example, robust yet short-lived wound healing response of injured oral epithelium can result in scarless healing [ 12 ], while a prolonged response with pro-fibrotic mediators can produce the non-healing bed of chronic wounds [ 8 , 13 ].…”