Benoit Hendrickx

Multipotent adult progenitor cells sustain function of ischemic limbs in mice.

Despite progress in cardiovascular research, a cure for peripheral vascular disease has not been found. We compared the vascularization and tissue regeneration potential of murine and human undifferentiated multipotent adult progenitor cells (mMAPC-U and hMAPC-U), murine MAPC-derived vascular progenitors (mMAPC-VP), and unselected murine BM cells (mBMCs) in mice with moderate limb ischemia, reminiscent of intermittent claudication in human patients. mMAPC-U durably restored blood flow and muscle function and stimulated muscle regeneration, by direct and trophic contribution to vascular and skeletal muscle growth. This was in contrast to mBMCs and mMAPC-VP, which did not affect muscle regeneration and provided only limited and transient improvement. Moreover, mBMCs participated in a sustained inflammatory response in the lower limb, associated with progressive deterioration in muscle function. Importantly, mMAPC-U and hMAPC-U also remedied vascular and muscular deficiency in severe limb ischemia, representative of critical limb ischemia in humans. Thus, unlike BMCs or vascular-committed progenitors, undifferentiated multipotent adult progenitor cells offer the potential to durably repair ischemic damage in peripheral vascular disease patients.

Integration of Blood Outgrowth Endothelial Cells in Dermal Fibroblast Sheets Promotes Full Thickness Wound Healing

Vascularization is the cornerstone of wound healing. We introduced human blood outgrowth endothelial cells (hBOEC) in a self‐assembled human dermal fibroblast sheet (hDFS), intended as a tissue‐engineered dermal substitute with inherent vascular potential. hBOEC were functionally and molecularly different from early endothelial progenitor cells and human umbilical vein endothelial cells (HUVEC). hBOEC alone, unlike HUVEC, efficiently revascularized and re‐oxygenated the wound bed, both by active incorporation into new vessels and by trophic stimulation of host angiogenesis in a dose‐dependent manner. Furthermore, hBOEC alone, but not HUVEC, accelerated epithelial coverage and matrix organization of the wound bed. In addition, integration of hBOEC in hDFS not only further improved vascularization, epithelial coverage and matrix organization but also prevented excessive wound contraction. In vitro analyses with hBOEC, fibroblasts and keratinocytes revealed that these effects were both due to growth factor crosstalk and to short cutting hypoxia. Among multiple growth factors secreted by hBOEC, placental growth factor mediated at least in part the beneficial effects on keratinocyte migration and proliferation. Overall, this combined tissue engineering approach paves the way for clinical development of a fully autologous vascularized dermal substitute for patients with large skin defects that do not heal properly. STEM CELLS 2010;28:1165–1177

Regulable vascular endothelial growth factor165 overexpression by ex vivo expanded keratinocyte cultures promotes matrix formation, angiogenesis, and healing in porcine full-thickness wounds.

The intricate wound repair process involves the interplay of numerous cells and proteins. Using a porcine full-thickness wound (FTW) healing model, we hypothesized that the ex vivo gene transfer of vascular endothelial growth factor (VEGF)-transfected basal keratinocyte (KC) cell suspensions may generate cross-talk and induce matrix formation, angiogenesis, and accelerated healing. Moreover, to regulate overexpression of isoform 165 of VEGF and its effect on healing, we introduced a tetracycline (TC)-inducible gene switch in the expression plasmid. Autologous basal KCs were cultivated from the porcine donor and transfected using cationic liposomes. A dose-response curve was established to determine optimal activation of the gene switch by TC. In vivo, FTWs were treated with VEGF-transfected KCs and controls. Wound fluids were collected daily and examined using enzyme-linked immunosorbent assay. Biopsies were evaluated using hematoxylin and eosin and immunostaining for fibronectin, CD144, and lectin BS-1. In vitro, highest regulable VEGF165-expression was obtained with 1 microg/mL of TCs. In vivo, after induction of the gene switch by adding 1 microg/mL of TCs to the FTW, we obtained upregulated VEGF165 levels and enhanced fibronectin deposition and found more endothelial cell tubular formations and higher rates of reepithelialization than in controls. This ex vivo gene transfer model may serve as a platform for vascular induction in full-thickness tissue repair.

A Plasma-Based Biomatrix Mixed with Endothelial Progenitor Cells and Keratinocytes Promotes Matrix Formation, Angiogenesis, and Reepithelialization in Full-Thickness Wounds

In search of an autologous vascularized skin substitute, we treated full-thickness wounds (FTWs) with autologous platelet-rich plasma gel (APG) in which we embedded endothelial progenitor cells (EPCs) and basal cell keratinocytes (KCs). We cultivated autologous KCs in low-serum conditions and expanded autologous EPCs from venous blood. FTWs (n = 55) were created on the backs of four pigs, covered with wound chambers, and randomly assigned to the following treatments: (1) APG, (2) APG + KCs, (3) APG + EPCs, (4) APG + KCs + EPCs, and (5) saline. All wounds were biopsied to measure neovascularization (lectin Bandeiraea Simplicifolia-1 (BS-1), alpha smooth muscle actin [alphaSMA], and membrane type 1 matrix metalloproteinase (MT1-MMP)), matrix deposition (fibronectin, collagen type I/III, and alphavbeta3), and reepithelialization. Wound fluids were analyzed for protein expression. All APG-treated wounds showed more vascular structures (p < 0.001), and the addition of EPCs further improved neovascularization, as confirmed by higher lectin, alphaSMA, and MT1-MMP. APG groups had higher collagen I/III (p < 0.05), alphavbeta3, and fibronectin content (p < 0.001), and they exhibited higher concentrations of platelet-derived growth factor subunit bb, basic fibroblast growth factor, hepatocyte growth factor, insulin growth factor-1, transforming growth factor-beta1 and -beta3, matrix metalloproteinase-1 and -z9, and tissue-inhibiting matrix metalloproteinase-1 and -2. Applying APG + KCs resulted in the highest reepithelialization rates (p < 0.001). No differences were found for wound contraction by planimetry. In this porcine FTW model, APG acts as a supportive biomatrix that, along with the embedded cells, improves extracellular matrix organization, promotes angiogenesis, and accelerates reepithelialization.

Lumbar Artery Perforator Flap: An Anatomical Study Using Multidetector Computed Tomographic Scan and Surgical Pearls for Breast Reconstruction.

Background: Breast reconstruction with the lumbar artery perforator flap is indicated in patients with unfavorable abdominal donor site. In addition to their clinical experience with lumbar artery perforator free flap breast reconstruction, the authors present an anatomical study of the origin and course of the perforators. Methods: Images of multidetector computerized tomography scans were used to visualize the location of the dominant lumbar artery perforator in 20 patients. The medical files of the authors’ patients who underwent lumbar artery perforator flap breast reconstruction were also analyzed. Results: Multidetector computed tomographic imaging in 20 female patients with a mean age of 47 years revealed an equal number of dominant perforators (10 left and 10 right); 60 percent were third lumbar artery perforators, 30 percent were fourth, and the remaining were second. The dominant perforators were mainly located 42.6 mm from the y axis at their origin at the transverse process, and 69.5 mm when emerging in the subcutaneous tissue. Six patients had nine successful lumbar artery perforator flaps for breast reconstruction. Average operative time was 270 minutes. Due to shortness of pedicle and mismatching between diameter of lumbar artery and internal mammary artery, vascular bypass (harvested from the deep inferior epigastric vessels) was required in 50 percent of the cases. The major complication at the donor site was seroma (80 percent). Conclusions: The lumbar artery perforator has a constant anatomical location. The free lumbar artery perforator flap provides an ample amount of tissue for breast reconstruction; however, its major disadvantages are the small artery diameter, shortness of the pedicle, and high seroma rate at the donor site. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

Breast Cancer–related Lymphedema: Quality of Life after Lymph Node Transfer

Background: Breast cancer–related lymphedema affects multiple aspects of patients’ daily lives. The main aim of this study was to assess the impact of vascularized lymph node transfer on the quality of life in patients with lymphedema. Methods: Between 2007 and 2012, 25 female patients with breast cancer–related lymphedema underwent vascularized lymph node transfer. In 22 cases, the patients underwent a simultaneous deep inferior epigastric artery perforator flap breast reconstruction based on the superficial circumflex iliac artery. The influence on quality of life was evaluated using the Upper Limb Lymphedema-27 questionnaire, which includes physical, psychological, and social dimensions. The authors also investigated risk factors for lymphedema, such as body mass index, smoking, age, and time between start of lymphedema and vascularized lymph node transfer, and their impact on quality of life. Results: Twenty-one patients (84 percent) had an improvement of quality of life after vascularized lymph node transfer. The mean physical, psychological, and social scores were significantly improved postoperatively (p < 0.001). Risk factors for the development of lymphedema did not influence quality of life among patients with breast cancer–related lymphedema. Skin infections disappeared in 50 percent of the cases. Eleven patients (44 percent) discontinued compression therapy at a mean postoperative time interval of 29 months (range, 8 to 64 months). In the other patients (56 percent), the average frequency of compression therapy decreased from three sessions to one session per week. Conclusion: Vascularized lymph node transfer significantly improves quality of life among patients with breast cancer–related lymphedema. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

Monsplasty for Women After Massive Weight Loss

Level of Evidence V This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of

The vascularized groin lymph node flap (VGLN): Anatomical study and flap planning using multi‐detector CT scanner. The golden triangle for flap harvesting

Introduction: A growing number of surgeons perform lymph node transfers for the treatment of lymphedema. When harvesting a vascularized lymph node groin flap (VGLNF) one of the major concerns is the potential risk of iatrogenic lymphedema of the donor‐site. This article helps understanding of the lymph node distribution of the groin in order to minimize this risk.

Cell and Gene Transfer Strategies for Vascularization During Skin Wound Healing

Adequate vascularization is pivotal to skin wound healing. Therefore, designing efficient revascularization strategies based on the mechanisms behind electromechanical stimulation of wound vascularization would be beneficial to the growing number of patients in need of improved wound healing. Recent attention has centered on applying gene/protein transfer and cell differentiation/transplantation approaches to stimulate and mimic the molecular events occurring during wound revascularization. Although both gene/protein transfer and cell differentiation/transplantation are faced with important challenges, researchers have made tremendous advances and shown both strategies to be a promising approach. In this chapter, we give an overview of the myriad of molecular players involved in neovascularization. We also discuss the molecular mechanisms of neovascularization during wound healing and provide an in-depth review on neovascular strategies and techniques for wound healing and tissue-engineered skin equivalents.

Multipotent Adult Progenitor Cells Support Lymphatic Regeneration at Multiple Anatomical Levels during Wound Healing and Lymphedema

Lymphatic capillary growth is an integral part of wound healing, yet, the combined effectiveness of stem/progenitor cells on lymphatic and blood vascular regeneration in wounds needs further exploration. Stem/progenitor cell transplantation also emerged as an approach to cure lymphedema, a condition caused by lymphatic system deficiency. While lymphedema treatment requires lymphatic system restoration from the capillary to the collector level, it remains undetermined whether stem/progenitor cells support a complex regenerative response across the entire anatomical spectrum of the system. Here, we demonstrate that, although multipotent adult progenitor cells (MAPCs) showed potential to differentiate down the lymphatic endothelial lineage, they mainly trophically supported lymphatic endothelial cell behaviour in vitro. In vivo, MAPC transplantation supported blood vessel and lymphatic capillary growth in wounds and restored lymph drainage across skin flaps by stimulating capillary and pre-collector vessel regeneration. Finally, human MAPCs mediated survival and functional reconnection of transplanted lymph nodes to the host lymphatic network by improving their (lymph)vascular supply and restoring collector vessels. Thus, MAPC transplantation represents a promising remedy for lymphatic system restoration at different anatomical levels and hence an appealing treatment for lymphedema. Furthermore, its combined efficacy on lymphatic and blood vascular growth is an important asset for wound healing.