Human decellularized dermal matrix seeded with adipose-derived stem cells enhances wound healing in a murine model: Experimental study.

OBJECTIVE: Full-thickness cutaneous wounds treated with split-thickness skin grafts often result in unaesthetic and hypertrophic scars. Dermal substitutes are currently used together with skin grafts in a single treatment to reconstruct the dermal layer of the skin, resulting in improved quality of scars. Adipose-derived stem cells (ASCs) have been described to enhance wound healing through structural and humoral mechanisms. In this study, we investigate the compatibility of xenogen-free isolated human ASCs seeded on human acellular dermal matrix (Glyaderm®) in a murine immunodeficient wound model. METHODS: Adipose tissue was obtained from abdominal liposuction, and stromal cells were isolated mechanically and cultured xenogen-free in autologous plasma-supplemented medium. Glyaderm® discs were seeded with EGFP-transduced ASCs, and implanted on 8 mm full-thickness dorsal wounds in an immunodeficient murine model, in comparison to standard Glyaderm® discs. Re-epithelialization rate, granulation thickness and vascularity were assessed by histology on days 3, 7 and 12. Statistical analysis was conducted using the Wilcoxon signed-rank test. EGFP-staining allowed for tracking of the ASCs in vivo. Hypoxic culture of the ASCs was performed to evaluate cytokine production. RESULTS: ASCs were characterized with flowcytometric analysis and differentiation assay. EGFP-tranduction resulted in 95% positive cells after sorting. Re-epithelialization in the ASC-seeded Glyaderm® side was significantly increased, resulting in complete wound healing in 12 days. Granulation thickness and vascularization were significantly increased during early wound healing. EGFP-ASCs could be retrieved by immunohistochemistry in the granulation tissue in early wound healing, and lining vascular structures in later stages. CONCLUSION: Glyaderm® is an effective carrier to deliver ASCs in full-thickness wounds. ASC-seeded Glyaderm® significantly enhances wound healing compared to standard Glyaderm®. The results of this study encourage clinical trials for treatment of full-thickness skin defects. Furthermore, xenogen-free isolation and autologous plasma-augmented culture expansion of ASCs, combined with the existing clinical experience with Glyaderm®, aid in simplifying the necessary procedures in a GMP-laboratory setting.

Xenogen-free isolation and culture of human adipose mesenchymal stem cells.

BACKGROUND: Adipose-derived Stem Cells (ASCs) present great potential for reconstructive procedures. Currently, isolation by enzyme digestion and culturing using xenogenic substances remain the gold standard, impairing clinical use. METHODS: Abdominal lipo-aspirate and blood samples were obtained from healthy patients. A novel mechanical isolation method for ASCs was compared to (the standard) collagenase digestion. ASCs are examined by flowcytometry and multilineage differentiation assays. Cell cultures were performed without xenogenic or toxic substances, using autologous plasma extracted from peripheral blood. After eGFP-transfection, an in vivo differentiation assay was performed. RESULTS: Mechanical isolation is more successful in isolating CD34+/CD31-/CD45-/CD13+/CD73+/CD146- ASCs from lipo-aspirate than isolation via collagenase digestion (p <0 .05). ASCs display multilineage differentiation potential in vitro. Autologous plasma is a valid additive for ASCs culturing. eGFP-ASCs, retrieved after 3 months in vivo, differentiated in adipocytes and endothelial cells. CONCLUSION: A practical method for human ASC isolation and culturing from abdominal lipo-aspirate, without the addition of xenogenic substances, is described. The mechanical protocol is more successful than the current gold standard protocol of enzyme digestion. These results are important in the translation of laboratory-based cell cultures to clinical reconstructive and aesthetic applications.

Microsurgical Reconstruction of the Nasal Ala using a Composite Auricular Graft Based on the Superficial Temporal Vessels.

Full-thickness defects of the nasal ala can be challenging to reconstruct. The original texture, color and shape of this specific aesthetic unit requires careful planning of the surgical approach and technique in order to minimize donor-site morbidity and repetitive procedures. We describe the use of the chondrocutaneous composite auricular graft to -reconstruct a full-thickness defect of the ala of the nose with a successful and aesthetically pleasing outcome.

The serum estradiol concentration is the main determinant of the estradiol concentration in normal breast tissue.

BACKGROUND: The estrogen concentration has been determined in breast tissue, focusing largely on samples obtained from breast cancers. In this study, estradiol concentration was determined in normal breast tissue obtained from women undergoing esthetic, and oncoplastic surgery. METHODS: Normal breast tissue was obtained during 68 operations for esthetic or reconstructive indications in women with and without a history of breast cancer. Our study included six different groups of women. The first three groups were normal cycling women, women taking oral contraceptives, and normal postmenopausal women, all undergoing a bilateral esthetic breast reduction. The second three groups were premenopausal and postmenopausal women, with a history of breast cancer and currently taking tamoxifen treatment, or postmenopausal women currently taking an aromatase inhibitor, needing contra-lateral corrective esthetic surgery. FINDINGS: In the group of women without history of breast cancer, normal cycling women (n=24) presented a strong correlation (r=0.853; P<0.0001) between 17ß-estradiol concentration in serum (median: 29.7 pg/mL; IQR: 10.8-82.3 pg/mL) and in breast tissue (30.6 pg/g; IQR: 18.6-183.8 pg/g). Postmenopausal women had low 17ß-estradiol concentrations both in serum and breast tissue (r=0.813; P<0.0001, n=16). Women taking oral contraceptives (n=12) had low serum and breast tissue levels of estradiol (r=0.376; P=n.s.). Premenopausal women (n=6, mean age: 44.2 years) with a history of breast cancer and currently taking tamoxifen, had very high concentrations of 17ß-estradiol both in serum (277.9 pg/mL; IQR: 96.2-544.7 pg/mL) and in epithelial cells (251.9 pg/g; IQR: 115.0-426.5 pg/g) (r=0.803; P<0.001). Postmenopausal women taking tamoxifen (n=4, mean age: 48.3) had low concentrations of 17ß-estradiol in serum (7.0 pg/mL; IQR: 5.7-16.3 pg/mL) and in epithelial cells (14.6 pg/g IQR: 13.3-16.3 pg/g) (r=0.10; P=n.s.). The estradiol concentration in the breast of premenopausal women taking tamoxifen was 8.2 times higher that observed in the breast of normal cycling women, and 17.3 times higher that observed in postmenopausal women taking tamoxifen. Women taking adjuvant aromatase inhibitors had extremely low concentrations of 17ß-estradiol both in serum and in breast tissue. INTERPRETATION: This study shows that serum estradiol levels largely determine estrogen levels in normal breast tissue.

The role of the internal mammary vessels as recipient vessels in secondary and tertiary head and neck reconstruction.

BACKGROUND: Successful microsurgical free tissue transfer for head and neck reconstruction highly depends on the quality of the recipient vessels. In most cases, vessels near the site of resection are available; however, when the bilateral vascular network in the neck is compromised or inaccessible due to prior surgery and/or irradiation, alternatives have to be sought. METHODS: Secondary or tertiary head and neck reconstruction was performed using the internal mammary vessels (IMVs) as recipient vessels in seven patients who had undergone previous neck dissection and radiation therapy. Indications were: tracheal-oesophageal fistula or stenosis (n = 4), oesophageal-cutaneous fistula (n = 1), saliva fistula (n = 1) and oral cancer (n = 1). Free flaps used for reconstruction were radial forearm flap (FRFF) (n = 5), anterolateral thigh flap (ALT) (n = 3) and transverse rectus abdominis myocutaneous flap (TRAM) (n = 1). Within two patients an additional ALT flap was necessary for soft-tissue coverage and resurfacing of the neck. The IMVs were separately exposed in a standard fashion over the second or third rib. The pedicle of the flap was anastomosed anterograde and end-to-end to the recipient vessels in all cases. Mean pedicle length was 14.3 cm (11-20 cm), with a mean distance of 9.8 cm (7-13 cm) between the resection and recipient vessel site. RESULTS: All patients were tumour free at time of re-operation and no sign of radiation injury was observed in the recipient vessels. All flaps survived and all patients healed without major complications. Mean follow-up time was 18 months. Four patients died of local recurrence or distant metastases during follow-up. CONCLUSION: In the vessel-depleted neck, the IMVs are a reliable and easy accessible recipient area for microsurgical reconstruction of the head and neck. Surgical management and technique refinements for dissection of the vessels are discussed. In combination with free flaps with a long pedicle, especially perforator flaps, vein grafts are unnecessary and microsurgery can safely be performed outside the zone of injury.

Free DIEAP and SGAP flap breast reconstruction after abdominal/gluteal liposuction.

Perforator flaps are widely used in our unit for breast reconstruction. They provide ample tissue with minimal donor site morbidity together with long lasting aesthetic results. Increasing number of patients may have liposuction procedure which may jeopardise areas such as the abdomen and the buttock which are the donor sites for perforator-free flaps in breast reconstruction. Therefore, liposuction has been considered as a relative contraindication of raising perforator flaps. Six patients who had previous liposuction of the donor sites underwent autologous breast reconstruction with perforator-free flaps. Colour Duplex imaging was obtained in all cases preoperatively in order to evaluate the blood supply to the flap and to map the perforators. There were five deep inferior epigastric artery flaps (DIEP) and one superior gluteal artery perforator (SGAP) flap used. Total flap survival was obtained in all cases. Postoperative course was uneventful. Our results showed that raising perforator flaps after liposuction of the donor sites is possible. Preoperative radiological evaluation of the perforators is mandatory for such difficult cases.

A simple new technique for neo-umbilicoplasty.

This article describes a new technique for reconstruction of the umbilicus. The technique used simply originated from a few clinical cases in which we were confronted with an absent or destroyed umbilicus. Previously described techniques for neo-umbilicoplasty were unsatisfying or seemed too complex in our hands. We introduce this easy, safe and reproducible technique which in our experience resulted in a very acceptable newly formed umbilicus.

The compound thoracodorsal perforator flap in the treatment of combined soft-tissue defects of sole and dorsum of the foot.

Nine cases of massive soft-tissue loss of the foot were reconstructed by means of a compound (chimera) thoracodorsal artery perforator (TAP) flap, which reconstituted the different functional units (dorsum, heel, instep, weight-bearing surface). In each case, the flap consisted of a skin component isolated on its perforator in combination with a portion of latissimus dorsi muscle and/or serratus fascia, all pedicled on the thoracodorsal vessels. The pedicle length allows up to 4-6 cm of independent mobility of the skin island. The mobility of the various flap components allows the various functional units of the foot to be reconstructed without relying on multiple flaps or anastomoses. The pedicle length was sufficient to be able to perform the anastomosis out of the zone of injury. In some cases the skin island was harvested along with intercostal nerve branches, this gave us the potential to develop a sensate flap. The indications and advantages of this reconstructive method are discussed.

A new technique of microvascular suturing: the chopstick rest technique.

With the conventional techniques of tying knots during microvascular anastomosis or neural suturing, time may be lost due to various reasons. The loose end of the suture often falls down into the operative field and gets stuck to the surrounding tissues. In the process of retrieving the suture, the surrounding tissues can be picked up together with the suture. When the posterior wall technique [Br J Plast Surg 34 (1981) 47, Plast Reconstr Surg 69 (1982) 139, Microsurgery 8 (1987) 22, J Reconstr Microsurg 15 (1999) 321] is used, the loose end of the suture may be stuck to the backside of the vessel and may be hard to grab. In order to avoid those problems, a new way of tying a microsuture was developed. By avoiding contact of the loose end of the suture to the surrounding tissue at any point during tying, the microvascular anastomosis can be performed quicker and more efficiently.

The role of iodine in antisepsis and wound management: a reappraisal.

For more than 150 years, iodine has been used for the prevention of infection and for the treatment of wounds. Nowadays a large amount of published evidence is available and, although it is generally in support of the use of iodine product, it is confused by being a mixture of laboratory, animals and human studies, often using different preparations. This makes interpretation and comparison difficult. After new developments and publications, the role of iodine in antisepsis and in wound management needs to be reevaluated. We mainly focused our review on the following problems: the role of the newly developed formulations of iodine preparations, its antimicrobial activity, the possibility of impairing the wound healing process, the role of iodine in the problem of growing resistance against antibiotics and antiseptics. New formulations seem to keep the same clinical efficacy, avoiding the problem of toxicity; it seems that the antibacterial activity of iodine is superior compared to other products and, in contrast with antibiotics and other antiseptics, it seems to have no resistance problem. It seems that povidone-iodine has all the characteristics to become the first choice antiseptic in wound treatment.

The pyramidalis muscle free flap.

The pyramidalis muscle is introduced as a new small muscle free flap, with description of its anatomy, the surgical technique and the clinical results in five different cases in which this flap was used to treat small recalcitrant wounds in the foot/ankle region. The pyramidalis muscle can be an alternative option in selective cases to reduce donor site morbidity as compared with more traditional free flaps.