Nestor Torio-Padron

Negative pressure wound therapy reduces the ischaemia/reperfusion-associated inflammatory response in free muscle flaps☆

BACKGROUND We recently established negative pressure wound therapy (NPWT) as a safe postoperative care concept for free muscle flaps; however, the molecular effects of NPWT on free muscle flaps remain elusive. Here we investigated the effects of NPWT on pathological changes associated with ischaemia/reperfusion injury in free flap tissue. METHODS From July 2008 to September 2010, 30 patients receiving skin-grafted free muscle transfer for defect coverage were randomly assigned to two treatment groups: In one group the skin-grafted free flap was covered by a vacuum dressing (NPWT); in the second group, flaps were covered by conventional petroleum gauze dressings (conv). Biopsies were taken intra-operatively prior to clipping of the pedicle and on postoperative day 5. Samples were analysed by immunohistochemistry for infiltration of inflammatory cells, real-time polymerase chain reaction (RT-PCR) for the analysis of expression levels of interleukin-1β (IL-1β) and tumour necrosis factor (TNF)-alpha as markers of inflammation. Histological samples were also examined for interstitial oedema formation, and apoptosis was detected by a terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay. RESULTS NPWT leads to a significantly reduced tissue infiltration of CD68 + macrophages and reduced expression of the inflammatory cytokines IL-1β and TNFα. None of these parameters was significantly elevated in the pre-ischaemic biopsies. Furthermore, NPWT reduced the interstitial oedema formation and the number of apoptotic cells in free flap tissue. CONCLUSION NPWT of skin-grafted free muscle flaps leads to a reduced inflammatory response following ischaemia/reperfusion, resulting in reduced oedema formation improving the microcirculation and ultimately reduced tissue damage. We thereby deliver new insight into the effects of NPWT.

In vitro analysis of the interactions between preadipocytes and endothelial cells in a 3D fibrin matrix

The volume‐persistent survival of transplanted adipose tissue in vivo relies on early vascularization, due to an otherwise early induction of apoptosis of the centrally located cells. Thus, one way to enable the early formation of a capillary network resulting in a sufficient perfusion of the transplanted construct might be the co‐transplantation of autologous preadipocytes with endothelial cells. To investigate preadipocyte–endothelial cell interaction, three‐dimensional proliferation‐ and angiogenesis assays were performed in vitro. Proliferation rates of co‐cultured endothelial cells and preadipocytes suspended in a fibrin matrix were elucidated by Alamarblue assays. The spheroid angiogenesis model was applied for analyzing the effects of vascular endothelial cell growth factor (VEGF) and basic fibroblast growth factor (bFGF) (produced by preadipocytes) as well as the impact of cell‐cell interaction between preadipocytes and endothelial cells and fibrin matrix on endothelial cell migration. Preadipocytes proliferated in fibrin glue, whereas endothelial cells underwent apoptosis. By co‐culturing, both cell types demonstrated an increased proliferation rate. Preadipocytes provoked migration of endothelial cells. Blocking bFGF and/or VEGF led to a significant decrease of migration. Changes in fibrin structure were followed by migration of single cells instead of sprouting. An appropriate fibrin matrix as well as already differentiated endothelial cells are necessary for preadipocytes to develop their angiogenic activity via bFGF and VEGF.

Co‐culture of adipose‐derived stem cells and endothelial cells in fibrin induces angiogenesis and vasculogenesis in a chorioallantoic membrane model

Neovascularization of adipose tissue equivalents is a crucial step in successful adipose tissue engineering, since insufficient vascularization results in graft resorption in an in vivo situation. A possible cellular approach to overcome this limitation is the co‐implantation of adipose‐derived stem cells (ASCs) with endothelial cells to stimulate the formation of a vascular network. We investigated the potential of ASCs derived from human abdominal fat tissue co‐cultured with endothelial progenitor cells (EPCs) from human peripheral blood to stimulate neovascularization of fibrin constructs on the chorioallantoic membrane (CAM) of fertilized chicken eggs, in direct comparison to human umbilical vein endothelial cells (HUVECs). After 9 days of incubation, cell–fibrin constructs were explanted and histologically evaluated with respect to ingrowth of avian blood vessels into the construct and formation of human blood vessels by co‐implanted endothelial cells. When administered on the CAM, ASCs successfully guided host vasculature into the construct (angiogenesis) and guided formation of capillary‐like structures by co‐implanted human endothelial cells (vasculogenesis), with HUVECs being superior to EPCs, leading to a perfused avian and human capillary network within the fibrin construct. However, the results also showed that perfused human blood vessels were only observed near the CAM compared to unperfused capillary‐like structures near the top of the construct, indicating that perfusion of the cell–fibrin construct takes longer than 9 days. In conclusion, as blood vessel formation is an essential step during adipogenic differentiation, the data support our hypothesis that cellular communication between transplanted ASCs and endothelial cells is beneficial for vasculogenesis. Copyright

Monitoring molecular changes induced by ischemia/reperfusion in human free muscle flap tissue samples.

Background:Our current knowledge of the pathophysiological sequelae of ischemia or reperfusion (I/R) injury in free tissue transfer in reconstructive surgery is based on data obtained in animal experiments. In this study, we investigated the histologic and molecular changes after 11 free microsurgical muscle transfers in human muscle tissue. Methods:Biopsies of free muscle flap tissue were taken immediately before clipping of the pedicle and 5 days after ischemia and successful microanastomosis and restoration of the blood flow. Samples were analyzed histologically for edema formation and by immunohistochemistry for infiltration of inflammatory cells and angiogenesis. Expression levels of the inflammatory marker proteins interleukin-1&bgr; and tumor necrosis factor &agr; and of complement component 3 as a major mediator of I/R injury were analyzed by real-time polymerase chain reaction. A TUNEL (terminal desoxynucleotidyl transferase–mediated-dUTP-nick-end-labeling) assay was used to assess apoptosis levels within the human muscle tissue. Results:I/R injury leads to a significant up-regulation of inflammatory parameters, infiltration of inflammatory cells, and angiogenesis. Increased complement component 3 deposition and apoptosis of cells were accompanied by interstitial edema as indication for a pronounced postischemic inflammatory reaction within the muscle tissue after free tissue transfer. Conclusions:Our findings of molecular changes induced by I/R injury in human striated muscle tissue validate data obtained in animal models of I/R injury. The parameters and inflammatory patterns defined in this study will allow for the monitoring of the success of novel pharmaceutical strategies in the future and will help to transfer data obtained in animal work to the in vivo setting in human beings.

Correlation between complication rate and patient satisfaction in abdominoplasty.

Abdominoplasty has become one of the most frequently performed procedures for improving body contour. Numerous reports exist focusing on postoperative complications with even more studies suggesting technical modifications to decrease their occurrence. However, the impact of complications on patient satisfaction has not been sufficiently addressed to date. A retrospective analysis was performed investigating the occurrence of complications following abdominoplasty and their effect on patient satisfaction. Patient satisfaction was assessed by means of the client satisfaction questionnaire-8. From June 1994 to April 2004 a total of 139 patients underwent an abdominoplasty. Minor and major complications were encountered in 40 (28.8%) and 16 (11.5%) patients, respectively. The median client satisfaction questionnaire-8 score among those patients with and without complications was 29. Thus, it seems that postoperative complications after abdominoplasty do not negatively affect patient satisfaction.

Adipose precursor cells (preadipocytes) induce formation of new vessels in fibrin glue on the newly developed cylinder chorioallantoic membrane model (CAM)

Successful augmentation of soft tissues by transplantation of preadipocytes within a matrix requires the formation of a new capillary network with connection to the host vessel system. Particularly, cells located centrally within the transplanted cell‐matrix‐construct represent a population with a blood supply questionable for survival. We demonstrated that under in vivo conditions preadipocytes possess the ability to induce and support the vascularization of the implant presumably by expression of several growth factors, such as VEGF (vascular endothelial growth factor) and bFGF (basic fibroblast growth factor). Fertilized White‐Leghorn eggs were incubated under standardized conditions. Opening was performed at day three of incubation and preadipocytes with and without recombinant growth factors were transferred into a fibrin matrix and subsequently placed on the Chorioallantoic Membrane (CAM), respectively. Eight days later, the implanted constructs were explanted, histologically processed and vascularization evaluated by means of a computer‐assisted image analysis program. Matrices containing preadipocytes displayed a significantly higher density of vascularization, whereas in the control group (fibrin without preadipocytes) no vessel ingrowth was observed. Daily application of recombinant growth factors added to the medium did not positively influence vascularization of the implant. Our investigations demonstrate that preadipocytes possess a strong angiogenic potential to induce and support neovascularization of 3D‐fibrin matrices under in vivo conditions. Addition of recombinant growth factors did not result in any stimulatory effect. Neither did the application of fibrin alone demonstrate an angiogenic potential with regard to induction of vascularization.

Comparison between endothelial progenitor cells and human umbilical vein endothelial cells on neovascularization in an adipogenesis mouse model

Volume stability and growth of tissue engineered adipose tissue equivalents using adipose-derived stem cells (ASCs) rely strongly on angiogenesis and neovascularization to support the maintenance of cells. An attractive cellular approach is based on coimplantation of endothelial cells to create a vascular network. Endothelial progenitor cells (EPCs) are a promising cell population, since they can be easily isolated from autologous human peripheral blood and thus represent a clinically feasible option. We have previously shown in in vitro and semi-in vivo studies that ASCs exert beneficial effects on EPCs in terms of enhanced tube formation and formation of blood vessels, respectively. In this study, we investigated the in vivo effects of coimplantation on endothelial cell-mediated neovascularization and ASC-mediated adipose tissue formation. For this purpose, human ASCs and human EPCs (or HUVECs as direct comparison to EPCs) were suspended alone or in coculture in fibrin and subcutaneously injected into the back of athymic nude mice and explanted after 1, 3 or 6months. Our results show that monocultures of EPCs or HUVECs were not able to perform vasculogenesis and constructs exhibited complete resorption already after 1month. However, a remarkable difference between EPCs and HUVECs was detected when coimplanted with ASCs. While coimplanted HUVECs gave rise to a stable neovasculature which was characterized by perfusion with erythrocytes, coimplanted EPCs showed no ability to form vascular structures. In the case of HUVEC-derived neovasculature, coimplanted ASCs displayed perivascular properties by stabilizing these neovessels. However, formation of human adipose tissue was independent of coimplanted endothelial cells. Our results indicate that HUVECs are superior to EPCs in terms of promoting in vivo neovascularization and recruiting perivascular cells for vessel stabilization when coimplanted with ASCs.

Nipple Reconstruction: Evidence-Based Trials in the Plastic Surgical Literature

Although many technical descriptions of nipple reconstruction exist in the medical literature, insufficient evidence-based data are present about the outcome. This study aimed to identify randomized controlled trials (RCTs) and controlled clinical trials (CCTs) in the plastic surgical literature addressing nipple reconstruction, and to elucidate whether a hand search was superior to an extensive database search in retrieving all pertinent studies. The hand search included analysis of all “original articles” published in four of the leading plastic surgery journals from January 1990 to December 2005, with subsequent identification of RCTs and CCTs. Additionally, a computerized search was conducted including the following databases: PubMed, Web of Science, and Evidence-Based Medicine Reviews. From a total of 10,476 published original articles in four plastic surgery journals over a 16-year period, only one RCT was identified that addressed nipple reconstruction. The database search, however, retrieved two trials: the RCT identified by hand search and one CCT. The impact of nipple reconstruction is well described in the literature. However, it is astonishing that the plastic surgical literature lacks evidence-based trials addressing this issue. Clearly, more evidence-based trials are necessary to ensure that recommendations for a particular technique are based on solid scientific data.

Controlled Trials in Aesthetic Plastic Surgery: A 16-Year Analysis

Aesthetic surgery is one pillar of plastic surgery. Thus, not surprisingly, journals exist that focus predominantly on advances within this subspecialty. However, rarely has the process of systematic reviewing that identified randomized controlled trials (RCTs) and controlled clinical trials (CCTs) been conducted within this subspecialty. All original articles published in Aesthetic Plastic Surgery were analyzed to identify all RCTs and CCTs. The proportion of RCTs and CCTs in all original articles was determined, and the quality of reporting was assessed on the basis of established quality items. Additional parameters were investigated including reporting of statistically significant differences, type of institution, and country affiliation of the first author. Of the 1,048 original articles analyzed, 11 (1%) and 24 (2.3%) articles met the inclusion criteria for RCTs and CCTs, respectively. Only two studies were single blinded, whereas only one study reported on successful double blinding and appropriate allocation concealment. Notably, these trials were RCTs. Participant dropout was reported in one study. Statistically significant differences were reported in 18 trials, 6 of which were RCTs. The annual publication of RCTs has increased over the past 5 years. North America and Europe contributed a total of 28 controlled trials (80%). Controlled trials are being conducted in aesthetic surgery at a strikingly low rate. However, a recent increase in published RCTs reflects the recognition that performing outcome studies is pivotal in moving practice toward a foundation based on assessment by outcome. The quality of reporting, however, needs improvement.

Comparison of Pre-Adipocyte Yield, Growth and Differentiation Characteristics from Excised versus Aspirated Adipose Tissue

Adipose tissue precursor cells (pre-adipocytes) are part of a stromal vascular fraction that can be easily isolated from fat tissue. Adipose tissue can be harvested by 2 methods: aspiration and excision. We analyzed whether the pre-adipocyte yield, growth characteristics and ability to differentiate into mature adipose tissue are influenced by the type of harvesting procedure. Adipose tissue was simultaneously harvested from the abdomen by surgical excision or aspiration according to the Coleman procedure in 10 individuals. This permitted inter- and intra-individual comparisons. Cell viability and yield were determined directly after isolation of pre-adipocytes. The growth kinetics were investigated in culture. Furthermore, pre-adipocytes were cultured under adipogenic conditions to compare their differentiation potential. The number of viable pre-adipocytes was significantly higher after excision of adipose tissue compared to aspiration. The proliferation kinetic was not influenced by the type of harvesting. No differences were observed in the differentiation potential of the pre-adipocytes between both groups. Compared to excision, aspiration of adipose tissue negatively affects the yield of pre-adipocytes. However, growth characteristics and differentiation potential of viable cultured cells are not influenced by the type of surgical harvesting. Due to its reduced donor site morbidity, we conclude that aspiration of adipose tissue is a valid harvesting method for isolation of pre-adipocytes.