A method for constructing vascularized muscle flap.

Abdominal wall reconstruction following extensive tissue loss is essential and can be achieved using autologous flaps. However, their use is limited due to their inadequate availability and due to post-operative donor site scarification. This work presents a step-by-step technique for fabrication of a vascularized muscle flap, to be applied in full-thickness abdominal wall defect reconstruction. Poly L-lactic acid/poly lactic-co-glycolic acid scaffolds, prepared using a salt leaching technique, were used as the supporting matrix in vitro for simultaneously seeded endothelial cells, fibroblasts and myoblasts. The cell-embedded graft was then implanted around femoral artery and vein vessels, which provided a central blood supply. Vascularization and perfusion were achieved by capillary sprouting from the main host vessel into the graft. A thick and vascularized tissue was formed within one week, and was then transferred as an autologous flap together with its main vessels, to a full-thickness abdominal wall defect. The flap remained viable after transfer and featured sufficient mechanical strength to support the abdominal viscera. Thus, this engineered muscle flap can be used as an alternative source for autologous flaps to reconstruct full-thickness abdominal wall defects.

Microcomputed Tomography-Based Analysis of Neovascularization within Bioengineered Vascularized Tissues.

In the field of tissue engineering, evaluating newly formed vascular networks is considered a fundamental step in deciphering the processes underlying tissue development. Several common modalities exist to study vessel network formation and function. However, a proper methodology that allows through three-dimensional visualization of neovessels in a reproducible manner is required. Here, we describe in-depth exploration, visualization, and analysis of vessels within newly formed tissues by utilizing a contrast agent perfusion protocol and high-resolution microcomputed tomography. Bioengineered constructs consisting of porous, biocompatible, and biodegradable scaffolds are loaded with cocultures of adipose-derived microvascular endothelial cells (HAMECs) and dental pulp stem cells (DPSCs) and implanted in a rat femoral bundle model. After 14 days of in vivo maturation, we performed the optimized perfusion protocol to allow host penetrating vascular visualization and assessment within neotissues. Following high-resolution microCT scanning of DPSC:HAMEC explants, we performed the volumetric and spatial analysis of neovasculature. Eventually, the process was repeated with a previously published coculture system for prevascularization based on adipose-derived mesenchymal stromal cells (MSCs) and HAMECs. Overall, our approach allows a comprehensive understanding of vessel organization during engraftment and development of neotissues.

Advances in the Use of Electrospun Nanofibrous Polymeric Matrix for Dermal Healing at the Donor Site After the Split-Thickness Skin Graft Excision: A Prospective, Randomized, Controlled, Open-Label, Multicenter Study.

Dressings used to manage donor site wounds (DSWs) have up to 40% of patients experiencing complications that may cause suboptimal scarring. We evaluated the efficacy and safety of a portable electrospun nanofibrous matrix that provides contactless management of DSWs compared with standard dressing techniques. This study included adult patients who underwent an excised split-thickness skin graft (STSG) with a DSW area of 10 to 200 cm2. Patients were allocated into two groups; ie, the nanofiber group managed with a nanofibrous polymer-based matrix, and the control group managed using the standard of care such as Jelonet® or Biatain® Ibu dressing. Primary outcomes were postoperative dermal healing efficacy assessed by Draize scores. The time to complete re-epithelialization was also recorded. Secondary outcomes included postoperative adverse events, pain, and infections during the first 21 days and extended 12-month follow-up. The itching and scarring were recorded during the extended follow-up (months 1, 3, 6, 9, and 12) using Numerical-Analogue-Score and Vancouver scores, respectively. The nanofiber and control groups included 21 and 20 patients, respectively. The Draize dermal irritation scores were significantly lower in the nanofiber vs control group (Z = -2.509; P = .028) on the first postoperative day but became similar afterward (Z ≥ -1.62; P ≥ .198). In addition, the average time to re-epithelialization was similar in the nanofiber (17.9 ± 4.4 days) and control group (18.3 ± 4.5 days; Z = -0.299; P = .764), so were postoperative adverse events, pain, and infection incidence, itching and scarring. The safety and efficacy of electrospun nanofibrous matrix are similar to standard wound care allowing its use as an alternative donor site dressing following the STSG excision.

Salivary monitoring related to major surgery.

BACKGROUND: Prolonged surgical procedures involving stress, extended general anaesthesia and a long pre-surgical fasting period may have systemic effects such as alterations in saliva flow rate and composition. These may compromise the patient's electrolytes and fluid balance and cause dehydration, systemic stress and oxidative changes. PATIENTS AND METHODS: Saliva was collected prior and following surgery from 20 patients and 20 control subjects. The saliva samples were analysed for flow rates and levels of the following: calcium (Ca), magnesium (Mg), total protein, albumin and lactate dehydrogenase (LDH), total antioxidant status (TAS), uric acid (UA), superoxide dismutase (SOD), carbonyls, matrix metalloproteinases (MMPs) -2, -3 and -9 and heat shock proteins (HSPs) 70 and 90. RESULTS: Salivary levels of Ca, Mg, protein, albumin and LDH were higher in post-surgical patients by 70% (P = 0·002), 88% (P = 0·0001), 120% (P = 0·13), 111% (P = 0·039) and 492% (P = 0·006) respectively than that in healthy controls. Salivary antioxidants in the surgical patients were higher while salivary carbonyls remained unchanged. Salivary TAS levels in pre- and post-surgical patients were higher by 63% (P = 0·001) and 85% (P = 0·0001) respectively, UA concentrations by 92% (P = 0·014) and 81% (P = 0·036) respectively and SOD values by 47% (P = 0·61) and 112% (P = 0·049) respectively. Salivary concentrations of MMP3 were higher in pre- and post-surgical patients by 23% (P = 0·067) and 30% (P = 0·044) respectively. CONCLUSIONS: Local salivary, oral and systemic-induced alterations should be prevented. Moreover, salivary collection and analysis may be a new, efficient tool in the monitoring of patients undergoing major surgery. Further related research is necessary.

Engineered Vascularized Muscle Flap.

One of the main factors limiting the thickness of a tissue construct and its consequential viability and applicability in vivo, is the control of oxygen supply to the cell microenvironment, as passive diffusion is limited to a very thin layer. Although various materials have been described to restore the integrity of full-thickness defects of the abdominal wall, no material has yet proved to be optimal, due to low graft vascularization, tissue rejection, infection, or inadequate mechanical properties. This protocol describes a means of engineering a fully vascularized flap, with a thickness relevant for muscle tissue reconstruction. Cell-embedded poly L-lactic acid/poly lactic-co-glycolic acid constructs are implanted around the mouse femoral artery and vein and maintained in vivo for a period of one or two weeks. The vascularized graft is then transferred as a flap towards a full thickness defect made in the abdomen. This technique replaces the need for autologous tissue sacrifications and may enable the use of in vitro engineered vascularized flaps in many surgical applications.

Adipose-derived endothelial and mesenchymal stem cells enhance vascular network formation on three-dimensional constructs in vitro.

BACKGROUND: Adipose-derived mesenchymal stem cells (MSCs) have been gaining fame mainly due to their vast clinical potential, simple isolation methods and minimal donor site morbidity. Adipose-derived MSCs and microvascular endothelial cells have been shown to bear angiogenic and vasculogenic capabilities. We hypothesized that co-culture of human adipose-derived MSCs with human adipose-derived microvascular endothelial cells (HAMECs) will serve as an effective cell pair to induce angiogenesis and vessel-like network formation in three-dimensional scaffolds in vitro. METHODS: HAMECs or human umbilical vein endothelial cells (HUVECs) were co-cultured on scaffolds with either MSCs or human neonatal dermal fibroblasts. Cells were immunofluorescently stained within the scaffolds at different time points post-seeding. Various analyses were performed to determine vessel length, complexity and degree of maturity. RESULTS: The HAMEC:MSC combination yielded the most organized and complex vascular elements within scaffolds, and in the shortest period of time, when compared to the other tested cell combinations. These differences were manifested by higher network complexity, more tube alignment and higher α-smooth muscle actin expression. Moreover, these generated microvessels further matured and developed during the 14-day incubation period within the three-dimensional microenvironment. CONCLUSIONS: These data demonstrate optimal vascular network formation upon co-culture of microvascular endothelial cells and adipose-derived MSCs in vitro and constitute a significant step in appreciation of the potential of microvascular endothelial cells and MSCs in different tissue engineering applications that can also be advantageous in in vivo studies.

Biodegradable soy wound dressings with controlled release of antibiotics: Results from a guinea pig burn model.

There is growing interest in the development of biodegradable materials from renewable biopolymers, such as soy protein, for biomedical applications. Soy protein is a major fraction of natural soybean and has the advantages of being economically competitive, biodegradable and biocompatible. It presents good water resistance as well as storage stability. In the current study, homogenous antibiotic-loaded soy protein films were cast from aqueous solutions. The antibiotic drug gentamicin was incorporated into the films in order to inhibit bacterial growth, and thus prevent or combat infection, upon its controlled release to the surrounding tissue. The current in vivo study of the dressing material in contaminated deep second-degree burn wounds in guinea pigs (n=20) demonstrated its ability to accelerate epithelialization with 71% epithelial coverage compared to an unloaded format of the soy material (62%) and a significant improved epithelial coverage as compared to the conventional dressing material (55%). Our new platform of antibiotic-eluting wound dressings is advantageous over currently used popular dressing materials that provide controlled release of silver ions, due to its gentamicin release profile, which is safer. Another advantage of our novel concept is that it is based on a biodegradable natural polymer and therefore does not require bandage changes and offers a potentially valuable and economic approach for treating burn-related infections.

Hybrid wound dressings with controlled release of antibiotics: Structure-release profile effects and in vivo study in a guinea pig burn model.

Over the last decades, wound dressings have evolved from a crude traditional gauze dressing to tissue-engineered scaffolds. Many types of wound dressing formats are commercially available or have been investigated. We developed and studied hybrid bilayer wound dressings which combine a drug-loaded porous poly(dl-lactic-co-glycolic acid) top layer with a spongy collagen sublayer. Such a structure is very promising because it combines the advantageous properties of both layers. The antibiotic drug gentamicin was incorporated into the top layer for preventing and/or defeating infections. In this study, we examined the effect of the top layer's structure on the gentamicin release profile and on the resulting in vivo wound healing. The latter was tested on a guinea pig burn model, compared to the neutral non-adherent dressing material Melolin® (Smith & Nephew) and Aquacel® Ag (ConvaTec). The release kinetics of gentamicin from the various studied formulations exhibited burst release values between 8% and 38%, followed by a drug elution rate that decreased with time and lasted for at least 7 weeks. The hybrid dressing, with relatively slow gentamicin release, enabled the highest degree of wound healing (28%), which is at least double that obtained by the other dressing formats (8-12%). It resulted in the lowest degree of wound contraction and a relatively low amount of inflammatory cells compared to the controls. This dressing was found to be superior to hybrid wound dressings with fast gentamicin release and to the neat hybrid dressing without drug release. Since this dressing exhibited promising results and does not require frequent bandage changes, it offers a potentially valuable concept for treating large infected burns.

Novel biodegradable composite wound dressings with controlled release of antibiotics: results in a guinea pig burn model.

Approximately 70% of all people with severe burns die from related infections despite advances in treatment regimens and the best efforts of nurses and doctors. Silver ion-eluting wound dressings are available for overcoming this problem. However, there are reports of deleterious effects of such dressings due to cellular toxicity that delays the healing process, and the dressing changes needed 1-2 times a day are uncomfortable for the patient and time consuming for the stuff. An alternative concept in wound dressing design that combines the advantages of occlusive dressings with biodegradability and intrinsic topical antibiotic treatment is described herewith. The new composite structure presented in this article is based on a polyglyconate mesh and a porous poly-(dl-lactic-co-glycolic acid) matrix loaded with gentamicin developed to provide controlled release of antibiotics for three weeks. In vivo evaluation of the dressing material in contaminated deep second degree burn wounds in guinea pigs (n=20) demonstrated its ability to accelerate epithelialization by 40% compared to an unloaded format of the material and a conventional dressing material. Wound contraction was reduced significantly, and a better quality scar tissue was formed. The current dressing material exhibits promising results, does not require frequent bandage changes, and offers a potentially valuable and economic approach to treating the life-threatening complication of burn-related infections.