Bacterial nanocellulose (BNC), synthesized by the bacterium Gluconacetobacter xylinus, is composed of highly hydrated fibrils (99 % water) with high mechanical strength. These exceptional material ...properties make BNC a novel biomaterial for many potential medical and tissue engineering applications. Recently, BNC with cellulose content of 15 % has been proposed as an implant material for auricular cartilage replacement, since it matches the mechanical requirements of human auricular cartilage. This study investigates the biocompatibility of BNC with increased cellulose content (17 %) to evaluate its response in vitro and in vivo. Cylindrical BNC structures (Ø48 × 20 mm) were produced, purified in a built-in house perfusion system, and compressed to increase the cellulose content in BNC hydrogels. The reduction of endotoxicity of the material was quantified by bacterial endotoxin analysis throughout the purification process. Afterward, the biocompatibility of the purified BNC hydrogels with cellulose content of 17 % was assessed in vitro and in vivo, according to standards set forth in ISO 10993. The endotoxin content in non-purified BNC (2,390 endotoxin units (EU)/ml) was reduced to 0.10 EU/ml after the purification process, level well below the endotoxin threshold set for medical devices. Furthermore, the biocompatibility tests demonstrated that densified BNC hydrogels are non-cytotoxic and cause a minimal foreign body response. In support with our previous findings, this study concludes that BNC with increased cellulose content of 17 % is a promising non-resorbable biomaterial for auricular cartilage tissue engineering, due to its similarity with auricular cartilage in terms of mechanical strength and host tissue response.
Objectives
Microvascular anastomoses in microvascular reconstructions induce rheological changes in the anastomosed vessels and are usually counteracted by anticoagulative medication. There is no ...regimen commonly agreed on. This study provides an easy to use anticoagulative regimen.
Patients and Methods
Consecutive cases of either anticoagulative regimen between 2013 and 2018 that underwent microvascular reconstruction in the head and neck area were included in this retrospective study, resulting in 400 cases in total. Two different anticoagulative regimens were applied to 200 patients in each group: (a) intraoperatively administered unfractionated 5000 I.U. high molecular weight heparin (HMWH) and postoperatively low molecular weight heparin (LMWH, Enoxaparin) 1 mg/kg/body weight postoperatively and (b) intraoperatively LMWH 0.5 mg/kg/body weight as well as 12 h later and 1 mg/kg/body weight postoperatively.
Results
The LMWH cohort showed fewer overall thromboembolic (8.5% vs. 11%; p = .40) and peripheral thrombotic events (1% vs. 3.5%; p = .18) and lung embolisms (3% vs. 4%; p = .59). The number of thromboses at the site of the anastomosis was equally distributed. In regard to flap‐specific complications, LMWH was associated with a positive effect, in particular with respect to total flap losses (5% vs. 7%; p = .40) and wound‐healing disorders (14.5% vs. 20%; p = .145).
Conclusion
Findings indicate that intra‐ and postoperatively administered LMWH as the only anticoagulative medication seems reliable in our clinical routine of head and neck free flap reconstructions.
The reconstruction of an auricle for congenital deformity or following trauma remains one of the greatest challenges in reconstructive surgery. Tissue-engineered (TE) three-dimensional (3D) cartilage ...constructs have proven to be a promising option, but problems remain with regard to cell vitality in large cell constructs. The supply of nutrients and oxygen is limited because cultured cartilage is not vascular integrated due to missing perichondrium. The consequence is necrosis and thus a loss of form stability. The micro-surgical implantation of an arteriovenous loop represents a reliable technology for neovascularization, and thus vascular integration, of three-dimensional (3D) cultivated cell constructs. Auricular cartilage biopsies were obtained from 15 rabbits and seeded in 3D scaffolds made from polycaprolactone-based polyurethane in the shape and size of a human auricle. These cartilage cell constructs were implanted subcutaneously into a skin flap (15 × 8 cm) and neovascularized by means of vascular loops implanted micro-surgically. They were then totally enhanced as 3D tissue and freely re-implanted in-situ through microsurgery. Neovascularization in the prefabricated flap and cultured cartilage construct was analyzed by microangiography. After explantation, the specimens were examined by histological and immunohistochemical methods. Cultivated 3D cartilage cell constructs with implanted vascular pedicle promoted the formation of engineered cartilaginous tissue within the scaffold in vivo. The auricles contained cartilage-specific extracellular matrix (ECM) components, such as GAGs and collagen even in the center oft the constructs. In contrast, in cultivated 3D cartilage cell constructs without vascular pedicle, ECM distribution was only detectable on the surface compared to constructs with vascular pedicle. We demonstrated, that the 3D flaps could be freely transplanted. On a microangiographic level it was evident that all the skin flaps and the implanted cultivated constructs were well neovascularized. The presented method is suggested as a promising alternative towards clinical application of engineered cartilaginous tissue for plastic and reconstructive surgery.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Allogenic graft material and tissue engineering have recently shown promising results for the improvement of both esthetic and functional outcomes in the treatment of large skin defects. We chose ...human amniotic membrane as a cellular scaffold in order to develop a skin substitute for later in vivo uses. Various methods of de-epithelialization of the human amniotic membrane were evaluated by histological analysis including hematoxylin–eosin and laminin staining, optic coherence tomography, and scanning electron microscopy with 0.25/0.02% trypsin/ethylenediaminetetraacetic acid treatment and mechanical cell removal showing an almost complete loss of the epithelium and a mainly intact basement membrane. Novel examination of human amniotic membrane by optic coherence tomography was feasible, but difficulties were experienced in handling and interpretation of the tissue as no comparable data exist. Subsequently, we developed an air–liquid interface cell culture to cultivate keratinocytes and fibroblasts on the de-epithelialized human amniotic membrane. We achieved a mostly keratinized surface on the epidermal side with a confluent fibroblast network on the chorion side.
Since its clinical implementation, microvascular surgery has depended on the continuous improvement of magnification tools. One of the more recent developments is a high-definition three-dimensional ...(3D) digital system (exoscope), which provides an alternative to the state-of-the-art operating microscopes. This study aimed to evaluate the advantages and disadvantages of this technology and compare it with its predecessor. The study included 14 surgeons with varying levels of experience, none of which had used a 3D optical system previously. Six of these surgeons performed five arterial and five venous anastomoses in the chicken thigh model with both the VITOM 3D exoscope-guided system and the Pentero operating microscope. These anastomoses were then evaluated for their quality and anastomosis time. The participants and the other eight surgeons, who had used the digital 3D camera system for microsurgical training exercises and vascular sutures, answered a questionnaire. The anastomosis time and number of complications were lower with the conventional microscope. Participants rated the image quality with the conventional microscope as higher, whereas the field of view and ergonomics were favorable in the digital 3D camera system. Exoscopes are optics suitable for performing simple microvascular procedures and are superior to classical microscopes ergonomically. Thus far, they are inferior to classical microscopes in terms of image quality and 3D imaging.
Vitrification of endothelial cells (MHECT-5) has not previously been compared with controlled slow freezing methods under standardized conditions. To identify the best cryopreservation technique, we ...evaluated vitrification and standardized controlled-rate -1°C/minute cell freezing in a -80°C freezer and tested four cryoprotective agents (CPA), namely dimethyl sulfoxide (DMSO), ethylene glycol (EG), propylene glycol (PG), and glycerol (GLY), and two media, namely Dulbecco's modified Eagle medium Ham's F-12 (DMEM)and K+-modified TiProtec (K+TiP), which is a high-potassium-containing medium. Numbers of viable cells in proliferation were evaluated by the CellTiter 96® AQueous One Solution Cell Proliferation Assay (Promega Corporation, Mannheim, Germany). To detect the exact frozen cell number per cryo vial, DNA content was measured by using Hoechst 33258 dye prior to analysis. Thus, results could be evaluated unconstrained by absolute cell number. Thawed cells were cultured in 25 cm2 cell culture flasks to confluence and examined daily by phase contrast imaging. With regard to cell recovery immediately after thawing, DMSO was the most suitable CPA combined with K+TiP in vitrification (99 ±0.5%) and with DMEM in slow freezing (92 ±1.6%). The most viable cells in proliferation after three days of culture were obtained in cells vitrificated by using GLY with K+TiP (308 ±34%) and PG with DMEM in slow freezing (280 ±27%).
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Free flap rescue using an extracorporeal perfusion device Fichter, Andreas M., MD, DDS; Ritschl, Lucas M., MD, DMD; Rau, Andrea, MD, DDS, PhD ...
Journal of cranio-maxillo-facial surgery,
12/2016, Letnik:
44, Številka:
12
Journal Article
Recenzirano
Summary The warm ischemia time of microvascular free flaps is limited. Incalculable events, such as lack of adequate recipient vessels or intraoperative medical emergencies, can lead to prolonged ...ischemia and potentially to flap loss. In this study, critically perfused ischemic or congested flaps were temporarily perfused with an extracorporeal perfusion system until anastomosis could be commenced. Temporary extracorporeal perfusion was performed in 8 radial forearm flaps for 147 ± 52 (range 77–237) minutes. Flap perfusion was assessed using Indocyanine Green fluorescence angiography and combined laser Doppler flowmetry and remission spectroscopy. Results were compared with those of 30 patients who underwent conventional reconstruction with radial forearm flaps. Flap survival, flap microcirculation, postoperative complications, and hospital stay did not differ between groups. We report successful free flap transfer after short-term extracorporeal perfusion for up to 4 hours in 8 patient cases. Temporary extracorporeal free flap perfusion reduces the warm ischemia time in emergency situations and can help to prevent flap failure in critically perfused or congested flaps. The trial is registered with ClinicalTrials.gov , number NCT02449525.
Systematic assessment of computed tomography (CT) scans and clinical symptoms is necessary to quickly indicate the correct treatment of zygomatico-orbital (ZMO) fractures. For this purpose, a ...clinical scoring system (=Clinical Score) was developed and correlated with CT scans to analyse its validity. Every operated, isolated, and unilateral ZMO fracture between January 2012 and December 2016 was screened retrospectively, including patient and treatment data. All available CT scans were analysed, and the grade of dislocation was measured for each case and plane. Four hundred and sixty-one cases were included and showed a median surgery time of 66.0 min (5.0−361.0) and a median postoperative hospital stay of three days (0−25). The distribution of gender, aetiologies and age groups was significantly different (each p = 0.001), and the aetiology had a significant influence on the Clinical Score (p = 0.038). The degree of dislocation in the coronary and sagittal planes correlated significantly with the Clinical Score with regard to the orbital involvement (p < 0.001, ρ = 0.566; p < 0.001, ρ = 0.609). The simple, quick, and easy-to-apply Clinical Score showed a significant correlation with the most important planes in CT scans as well as with the clinical course. It may facilitate fast risk stratification of the patient. However, the validity of the proposed score in determining indications must now be evaluated in a prospective setting, including both operated and non-operated fractures.
BACKGROUND:Human amniotic membranes (hAMs) have shown promising results in recent studies aimed at improving wound healing through several mechanisms. We wanted to investigate its properties as a ...scaffold by adding autologous cells to treat full-thickness skin defects and hypothesized that recultivated hAM would show an even improved wound healing by accelerating the epidermal closure of the wound.
METHODS:In an air–liquid cell culture, we cultivated autologous keratinocytes and fibroblasts on the hAM until a mostly keratinized surface was achieved. These hAM, de-epithelialized hAM, native hAM with remaining allogenous cells, and negative controls were compared in the treatment of circular 30 × 30 mm full-thickness skin defects in 4 groups of 6 rats with one wound each. We evaluated the wound contraction every 10 days until wound closure, the macroscopic scar appearance on the Vancouver Scar Scale and the qualitative histological properties of the scar regarding morphology and continuity of the basement membrane.
RESULTS:Rats treated with de-epithelialized hAM showed more extent wound contraction (P < 0.001) than the other 3 groups, which did not differ significantly compared with the control group (P > 0.05). Vancouver Scar Scale showed no significantly statistical differences between the 4 groups (P = 0.46). The scar structure of all rats showed similar morphologies, the only difference being the absence of a basement membrane in the negative controls compared with the groups treated with hAM.
CONCLUSION:The rats treated with hAM showed no improved wound healing but a tendency toward a more prominent basement membrane in the resulting scar.
A critical limitation in the cultivation of cartilage for tissue engineering is the dedifferentiation in chondrocytes, mainly during in vitro amplification. Despite many previous studies ...investigating the influence of various conditions, no data exist concerning the effects of hypothermia. Our aim has been to influence chondrocyte dedifferentiation in vitro by hypothermic conditions. Chondrocytes were isolated from cartilage biopsies and seeded in monolayer and in three-dimensional pellet-cultures. Each cell culture was either performed at 32.2°C or 37°C during amplification. Additionally, the influence of the redifferentiation of chondrocytes in three-dimensional cell culture was examined at 32.2°C and 37°C after amplification at 32.2°C or 37°C. An 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay was used to measure cell proliferation in monolayer, whereas the polymerase chain reaction and immunohistochemical and histological staining were used in three-dimensional pellet-cultures. Real-time polymerase chain reaction was employed to measure the relative expression of the target genes collagen II, collagen I, aggrecan and versican. Ratios were estimated between collagen II/collagen I and aggrecan/versican to evaluate differentiation. A higher value of these ratios indicated an advantageous status of differentiation. In monolayer, hypothermia at 32.2°C slowed down the proliferation rate of chondrocytes significantly, being up to two times lower at 32.2°C compared with culture at 37°C. Simultaneously, hypothermia in monolayer decelerated dedifferentiation. The ratio of aggrecan/versican was significantly higher at 32.2°C compared with that at 37°C. In three-dimensional pellet-culture, the chondrocytes redifferentiated at 32.2°C and at 37°C, and this process is more distinct at 37°C than at 32.2°C. Similar results were obtained for the ratios of collagen II/collagen I and aggrecan/versican and were supported by immunochemical and histological staining. Thus, hypothermic conditions for chondrocytes are mainly advantageous in monolayer culture. In three-dimensional pellet-culture, redifferentiation predominates at 37°C compared with at 32.2°C. In particular, the results from the monolayer cultures show potential in the avoidance of dedifferentiation.
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