Autologous Lipofilling Improves Clinical Outcome in Patients With Symptomatic Dermal Scars Through Induction of a Pro-Regenerative Immune Response

BACKGROUND: Autologous lipofilling is an emerging procedure to treat and possibly reverse dermal scars and to reduce scar-related pain, but its efficacy and mechanisms are poorly understood. OBJECTIVES: The aim of this study was to test the hypothesis that repeated lipografts reverse dermal scars by reinitiation of wound healing. METHODS: In a prospective, non-placebo-controlled clinical study, 27 adult patients with symptomatic scars were given 2 lipofilling treatments at 3-month intervals. As primary outcome, clinical effects were measured with the Patient and Observer Scar Assessment Scale (POSAS). Scar biopsies were taken before and after treatments to assess scar remodeling at a cellular level. RESULTS: Twenty patients completed the study. Patients’ scars improved after lipofilling. The total POSAS scores (combined patient and observer scores) decreased from 73.2  [14.7] points (mean [standard deviation]) pretreatment to 46.1 [14.0] and 32.3 [13.2] points after the first and second lipofilling treatment, respectively. Patient POSAS scores decreased from 37.3 [8.8] points to 27.2 [11.3] and 21.1 [11.4] points, whereas observer POSAS scores decreased from 35.9 [9.5] points to 18.9 [6.0] and 11.3 [4.5] points after the first and second treatment, respectively. After each lipofilling treatment, T lymphocytes, mast cells, and M2 macrophages had invaded scar tissue and were associated with increased vascularization. In addition, the scar-associated epidermis showed an increase in epidermal cell proliferation to levels similar to that normal in skin. Moreover, lipofilling treatment caused normalization of the extracellular matrix organization towards that of normal skin. CONCLUSIONS: Autologous lipofilling improves the clinical outcome of dermal scars through the induction of a pro-regenerative immune response, increased vascularization, and epidermal proliferation and remodeling of scar tissue extracellular matrix. LEVEL OF EVIDENCE: 4: [Image: see text

Human adipose tissue-derived stromal cells suppress TGF-β induced fibroblast differentiation of human dermal fibroblasts: implications for scar formation

Adipose tissue-derived stromal cells (ADSC) are multipotent stem cells which can be isolated and cultured from white adipose tissues throughout the body. In wound healing and regeneration of the skin, ADSC have already proven to be effective. However, it is not yet known if and how ADSC can prevent or reduce the formation of dermal scars. Scar formation is caused by persisting stimulation of fibroblasts by Transforming Growth Factor- β (TGF-β), which causes them to differentiate to myofibroblasts. The aim of this research is to investigate the influence of ADSC on TGF-β-mediated fibroblast-myofibroblast differentiation. We hypothesize that ADSC inhibit fibrotic remodeling by (1) repressing myofibroblast formation and (2) increasing matrix turnover. ADSC were isolated from human adipose tissue. ADSC conditioned medium (ADSC CM) was obtained by incubating ADSC with culture medium for 24h, thus collecting trophic factors from ADSC. Human dermal fibroblasts, adult type (HDFa) were incubated with ADSC CM or co-cultured with ADSC, in presence or absence of TGF-β1. After 4 days, HDFa were collected for gene transcript and immunoblot analysis of collagen I and III, matrix metalloproteinases (MMP), tissue inhibitors of metalloproteinases (TIMP) and SM22α, a mesenchymal marker. Functionally, contraction of HDFa was measured by means of a gel contraction assay. HDFa proliferation is up regulated by TGF-β1. ADSC CM brings proliferation back to control levels. ADSC CM reduces SM22α gene and protein expression in HDFa to 0.38±0.02 and 0.45±0.09 fold, respectively. After TGF-β1 stimulation, SM22α protein increased to 1.71±0.19 fold which was brought back to 1.11±0.16 fold by ADSC CM. Collagen I and collagen III mRNA expression to 0.49± 0.04 fold and 0.48± 0.06 fold. On protein level, collagen I is reduced to 0.51±0.13 fold. Furthermore, MMP-1, MMP-2, MMP-14 and TIMP-1 and TIMP-2 gene expression in HDFa was up regulated by ADSC CM or direct co-culture with ADSC. Collagen gel contraction by HDFa was significantly reduced after ADSC CM, irrespective of TGF-β1 stimulation. In this study we show ADSC inhibit fibrotic remodeling by repressing myofibroblast formation and by increasing matrix remodeling. In term, inhibition of fibroblast differentiation might lead to prevention or even reduction of dermal scars.

Bridging the gap: Adipose tissue-based therapy for dermal scarring

In dit proefschrift hebben we het gebruik van eigen vetweefsel als therapie voor littekens onderzocht. Vetweefsel kan door middel van liposuctie verkregen worden en ingespoten worden in en onder littekens van de huid. Dit wordt al lange tijd gebruikt in de plastische chirurgie als littekenbehandeling en lijkt verbetering van littekens te geven. Hiervoor ontbreekt echter nog sluitend wetenschappelijk bewijs. In onze patiëntenstudie hebben we vet ingespoten in littekens die klachten (bijvoorbeeld pijn) geven. Lipofilling leidt bij deze patiënten tot een vermindering van klachten, waarbij de littekens meer gaan lijken op normale huid. Ook vinden er microscopische veranderingen in het littekenweefsel plaats, die wijzen op regeneratie. Daarnaast hebben we in het laboratorium laten zien dat volwassen stromale (stam)cellen uit vetweefsel activatie van huidbindweefselcellen (huidfibroblasten) kan remmen: overmatige activatie van deze cellen zorgt voor littekenvorming. Groeifactoren van (stam)cellen uit vet zouden dus als therapie ter voorkoming van littekens kunnen dienen. Echter, het verkrijgen van deze stromale (stam)cellen uit vet is een tijdrovend proces. Daarom hebben we in dit proefschrift verschillende manieren onderzocht om voldoende stromale (stam)cellen uit vet te verkrijgen voor gebruik in patiënten. Als eerste hebben we stromale (stam)cellen uit vet in het laboratorium sneller laten groeien met behulp van extra groeifactoren uit menselijke bloedplaatjes. Daarnaast hebben we verschillende manieren vergeleken om tijdens operaties snel stromale (stam)cellen uit vet te verkrijgen. Hiervan testen we op het moment nog één van deze methoden in een klinische placebo-gecontroleerde patiëntenstudie, om te zien of inspuiten van de stromale (stam)celfractie de vorming van ontsierende littekens kan tegengaan. Concluderend, hebben we in dit proefschrift aangetoond dat lipofilling en stromale (stam)cellen uit vetweefsel kunnen bijdragen aan de behandeling van littekens

Adipose Tissue-Derived Stromal Cells Inhibit TGF-beta 1-Induced Differentiation of Human Dermal Fibroblasts and Keloid Scar-Derived Fibroblasts in a Paracrine Fashion

Background: Adipose tissue-derived stromal cells augment wound healing and skin regeneration. It is unknown whether and how they can also influence dermal scarring. The authors hypothesized that adipose tissue-derived stromal cells inhibit adverse differentiation of dermal fibroblasts induced by the pivotal factor in scarring, namely, transforming growth factor (TGF)-beta. Methods: TGF-beta 1-treated adult human dermal fibroblasts and keloid scar-derived fibroblasts were incubated with adipose tissue-derived stromal cell-conditioned medium and assessed for proliferation and differentiation, particularly the production of collagen, expression of SM22 alpha, and development of hypertrophy and contractility. Results: TGF-beta 1-induced proliferation of adult human dermal fibroblasts was abolished by adipose tissue-derived stromal cell-conditioned medium. Simultaneously, the medium reduced SM22a gene and protein expression of TGF beta 1-treated adult human dermal fibroblasts, and their contractility was reduced also. Furthermore, the medium strongly reduced transcription of collagen I and III genes and their corresponding proteins. In contrast, it tipped the balance of matrix turnover to degradation through stimulating gene expression of matrix metalloproteinase (MMP)-1, MMP-2, and MMP-14, whereas MMP-2 activity was up-regulated also. Even in end-stage myofibroblasts (i.e., keloid scar-derived fibroblasts), adipose tissue-derived stromal cell-conditioned medium suppressed TGF-beta 1-induced myofibroblast contraction and collagen III gene expression. Conclusion: The authors show that adipose tissue-derived stromal cells inhibit TGF-beta 1-induced adverse differentiation and function of adult human dermal fibroblasts and TGF-beta 1-induced contraction in keloid scar-derived fibroblasts, in a paracrine fashion

Platelet-Rich Plasma Influences Expansion and Paracrine Function of Adipose-Derived Stromal Cells in a Dose-Dependent Fashion

Background: Lipofilling is a treatment modality to restore tissue volume. Both platelet-rich plasma and adipose-derived stromal cells have been reported to augment the efficacy of lipofilling, yet results are not conclusive. The authors hypothesized that the variation reported in literature is caused by a dose-dependent influence of platelet-rich plasma on adipose-derived stromal cells. Methods: Whole blood (n = 3) was used to generate platelet-rich plasma, which was diluted with Dulbecco's Modified Eagle Medium to 15%, 5%, and 1.7%, with 15% platelet-poor plasma and 10% fetal calf serum as controls. Pooled adipose-derived stromal cells (n = 3) were cultured in these media. Gene expression was assessed, along with angiogenic sprouting of endothelial cells by conditioned medium and platelet-rich plasma. Results: platelet-rich plasma in culture medium affected the expression of genes in a dose-dependent manner. The 15% concentration stimulated proliferation almost eightfold. Mesenchymal markers were unaffected. Interestingly, expression of collagens type 1 and 3 increased at lower concentrations, whereas transforming growth factor-beta showed reduced expression in lower concentrations. Proangiogenic gene expression was unaltered or strongly reduced in a dose-dependent manner. platelet-rich plasma promoted endothelial sprouting and survival in a dose-dependent manner; however, conditioned medium from adipose-derived stromal cells exposed to platelet-rich plasma blocked endothelial sprouting capabilities. Conclusion: The dose-dependent influence of platelet-rich plasma on the therapeutic capacity of adipose-derived stromal cells conditioned medium in vitro warrants caution in clinical trials

Comparison of intraoperative procedures for isolation of clinical grade stromal vascular fraction for regenerative purposes: a systematic review

INTRODUCTION: Intraoperative application of stromal vascular fraction (SVF) of adipose tissue, requires a fast and efficient isolation procedure of adipose tissue. This review was performed to systematically assess and compare procedures currently used for the intraoperative isolation of cellular SVF (cSVF) and tissue SVF (tSVF) which still contains the extracellular matrix. METHODS: Pubmed, EMBASE and The Cochrane Central Register of controlled trials databases were searched for studies that compare procedures for intraoperative isolation of SVF (searched 28(th) of September, 2016). Outcomes of interest were cell yield, viability of cells, composition of SVF, duration, cost and procedure characteristics. Procedures were subdivided in procedures resulting in a cSVF or tSVF. RESULTS: Thirteen out of 3038 studies were included, evaluating eighteen intraoperative isolation procedures, were considered eligible. In general, cSVF and tSVF intraoperative isolation procedures had comparable cell yield, cell viability and SVF composition compared to a non-intraoperative (i.e. culture lab-based collagenase protocol) control group within the same studies. The majority of intraoperative isolation procedures are less time consuming than non-intraoperative control groups, however. CONCLUSION: Intraoperative isolation procedures are less time-consuming than non-intraoperative control group with similar cell yield, viability of cells and composition of SVF and therefore more suitable for use in the clinic. Nevertheless, none of the intraoperative isolation procedures could be designated as preferred procedure to isolate SVF

The power of fat and its adipose-derived stromal cells: Emerging concepts for fibrotic scar treatment

Lipofilling or lipografting is a novel and promising treatment method for reduction or prevention of dermal scars after injury. Ample anecdotal evidence from case reports supports the scar-reducing properties of adipose tissue grafts. However, only a few properly controlled and designed clinical trials have been conducted thus far on this topic. Also, the underlying mechanism by which lipofilling improves scar aspect and reduces neuropathic scar pain remains largely undiscovered. Adipose-derived stromal or stem cells (ADSC) are often described to be responsible for this therapeutic effect of lipofilling. We review the recent literature and discuss anticipated mechanisms that govern anti-scarring capacity of adipose tissue and its ADSC. Both clinical and animal studies clearly demonstrated that lipofilling and ADSC influence processes associated with wound healing, including extracellular matrix remodelling, angiogenesis and modulation of inflammation in dermal scars. However, randomized clinical trials, providing sufficient level of evidence for lipofilling and/or ADSC as an anti-scarring treatment, are lacking yet warranted in the near future. (c) 2017 The Authors Journal of Tissue Engineering and Regenerative Medicine Published by John Wiley & Sons Lt