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Background: Reconstruction of extensive defects of the lower abdomen, penoscrotum, trochanter, groin and knee without using complex microsurgery is a reconstructive challenge. Pedicled anterolateral thigh (ALT) flaps offer many advantages over other regional flaps for this purpose, such as the large skin area and soft-tissue availability, a remarkable pedicle length, and possessing multiple components and reliability. We present our experience of using pedicled ALT flaps for repairing various defects., Methods: From September 2006 to December 2013, 42 pedicled ALT flaps were used in 41 patients for defects of the lower abdomen (three patients), trochanter (26 patients), penoscrotum (10 patients), groin (one patient) and knee (one patient). Twenty-eight were men and 13 were women, and their mean age was 70.5 years (range, 22-103 years). The characteristics of the patients' age, sex, cause, flap size, flap component, follow-up and donor sites were recorded., Results: The flap size ranged from 8 × 5 cm (40 cm 2 ) to 11 × 18 cm (198 cm 2 ). The length of the pedicle ranged from 9 to 16 cm, which was enough to reach the defect without tension. No surgery-related mortality occurred. In 34 flaps, donor sites were closed primarily and eight underwent split-skin grafting. Satisfactory coverage was achieved in all patients., Conclusion: Our experience has shown the wide arc of rotation, large skin replacement potential, multiple components and reliability of pedicled ALT flaps. They are technically simple to apply as myocutaneous/fasciocutaneous flaps with minimal donor site morbidity., (© 2015 Royal Australasian College of Surgeons.)

Zwitterionic poly(sulfobetaine acrylamide) (pSBAA)-based nanocomposite hydrogels impregnated with germicidal silver nanoparticles (AgNPs) were synthesized and implemented for the treatment of infected chronic wounds. The zwitterionic hydrogels exhibited excellent non-sticky properties and had reinforced mechanical properties by the addition of hectorite nanoclay and poly(ethylene glycol)dimethacrylate as physical and chemical crosslinkers, respectively. In addition, AgNPs were grown within the intercalated clay/polymer structure by in situ free radical reduction, as confirmed by UV-vis spectroscopy and transmission electron microscopy (TEM). The silver-containing pSBAA nanocomposite hydrogels (pSBAA/Ag) exhibited germicidal properties against Gram-positive S. epidermidis and Gram-negative P. aeruginosa. The zwitterionic hydrogels show higher water content than 2-hydroxyethyl methacrylate (pHEMA) hydrogels, owing to the strong hydration via ionic solvation. The negligible cytotoxicity of pSBAA/Ag hydrogels was assessed with human fibroblasts by the MTT assay. Moreover, the zwitterionic hydrogels demonstrated excellent resistance to the adsorption of bovine serum albumin (BSA). To evaluate the feasibility of the hydrogels for clinical application as wound dressings, we created infected diabetic rat models and compared with commercial wound dressings. The results show that pSBAA/Ag hydrogels did not adhere to the newly formed tissue, and were readily removed from the wounds after treatment for 3 days. Moreover, the healing recovery was evaluated by visual observation of infected dorsal wounds on rats with induction of diabetes by streptozotocin. The finding indicates complete healing with the pSBAA/Ag hydrogels after 15 days, faster than other dressings. A histological examination also proved that the zwitterionic hydrogels facilitated epithelialization and collagen distribution in the infected diabetic wounds. Consequently, these novel non-sticky and antimicrobial zwitterionic nanocomposite hydrogels can have high potential for the treatment of infected chronic wounds.

Introduction: Explosion injury is very common on the battlefield and is associated with major burn and inhalation injuries and subsequent high mortality and morbidity rates. Here we report six victims who suffered from explosion injuries caused by stun grenade; all were treated with extracorporeal membrane oxygenation (ECMO) as salvage therapy. This study was aimed to evaluate the indications and efficacy of ECMO in acute and critically ill major burn patients., Methods: This was a retrospective analysis of six patients from Tri-Service General Hospital, National Defense Medical Center in Taiwan. All suffered from major burns with 89.0±19.1% average of total body surface area over second degree (TBSA; range, 50-99%). ECMO was used due to inhalation injury in five patients and cardiogenic shock in one patient. The average interval to start ECMO was 26.5±19.0h (range, 14-63h). Venoarterial ECMO was used on in four patients due to unstable hemodynamic status, whereas venovenous ECMO was used in two patients for sustained hypoxemia., Results: All patients had rhabdomyolysis with acute renal failure. The average duration of ECMO was 169.6±180.9h (range, 27-401h). All patients developed coagulopathy and needed debridement surgery during ECMO support, and five underwent torso escharotomy due to inspiratory compromise. Only one patient whose second and third degree burns covered 50% TBSA was successfully weaned from ECMO and survived; he was discharged after 221 hospital days. All patients who died had second and third degree burns covering over 90% of their TBSA. Three patients died of multiple organ failure, one died of septic shock, and the other died of cardiogenic shock. Overall survival rate was 16.7%., Conclusions: In acute and critically ill major burn patients, ECMO could be considered as a salvage therapy, particularly in those with inhalation injury and burn-related acute respiratory distress syndrome. However, ECMO does not seem to provide benefits for circulatory support in those with hemodynamic compromise. The use of ECMO in these patients is still investigational, as our data provided no benefit in terms of the outcomes or survival, particularly in those with more than 90% TBSA burns., (Copyright © 2016 Elsevier Ltd and ISBI. All rights reserved.)

Massive bleeding is the leading cause of battlefield-related deaths and the second leading cause of deaths in civilian trauma centers. One of the challenges of managing severe wounds is the need to promote hemostasis as quickly as possible, which can be achieved by using hemostatic dressings. In this study, we fabricated 2 kinds of gelatin/polycaprolactone composites with 2 ratios of gelatin/polycaprolactone, 1:1 and 2:1 (GP11 and GP21, respectively). Scanning electron microscopy revealed that the GP11 composite exhibited rougher and more porous structure than the GP21 composite did. Furthermore, both composites showed similar biocompatibility as that of tissue culture polystyrene. Moreover, both GP composites tended to show a gradual decrease in contact angle to zero within 40 minutes. The in vitro blood plasma coagulation assay revealed that the prothrombin time was significantly longer for the GP composites than it was for the Quikclot composite, whereas the activated partial thromboplastin time of the GP11 composite was significantly shorter than that of the gauze. Furthermore, the GP11 had the largest platelet adsorption of all the composites. The in vivo coagulation test showed an obvious shortening of the bleeding time with the Quikclot and GP21 compared with gauze sample. In conclusion, the GP composites showed superior biocompatibility and hemostasis to the gauze and comparable effects with the Qickclot composite. Therefore, the GP composites have the potential for development as biodegradable surgical hemostatic agents.

Background: Acute compartment syndrome is a well-described surgical emergency that requires immediate diagnosis and operative intervention. Vascular access-associated compartment syndrome is rarely reported in haemodialysis patients. The purpose of this article is to document evidence that catheter-related puncture, which results in arteriovenous fistula injury in uraemia, may cause acute forearm compartment syndrome., Methods: Between September 2007 and September 2012, five consecutive patients presented to our section with tense swollen forearms with skin blistering, decreased hand sensation and reduced capillary return in the fingers. Their ages ranged from 65 to 81 years (mean 72.8 years). All of the patients underwent emergent exploration after the diagnosis of acute forearm compartment syndrome. The patients' details were reviewed., Results: The time interval between dialysis completion and return to the emergency department ranged from 6 to 9 h (mean 7.4 h). During operation, the bleeding was found to originate from the site of the fistula puncture and was repaired with 9-0 nylon suture under microscopy. After adequate wound care, a reconstructive procedure with a split-thickness skin graft was performed in all of the five patients. There was no vascular or neurological deficit of the forearm or hand within the mean follow-up period of 14.8 months (range 12-18 months)., Conclusions: In this series, we report five cases of forearm compartment syndrome in uraemia, secondary to bleeding from a catheter-related puncture of a haemodialysis access fistula. However, there is no case series that focuses upon this specific topic in the present literature. This problem deserves more attention., (© 2014 Royal Australasian College of Surgeons.)

Cell- and gene-based therapies have emerged as promising strategies for treating neurological diseases. The sources of neural stem cells are limited while the induced pluripotent stem (iPS) cells have risk of tumor formation. Here, we proposed the generation of self-renewable, multipotent, and neural lineage-related neural crest stem-like cells by chitosan substrate-mediated gene transfer of a single factor forkhead box D3 (FOXD3) for the use in neural repair. A simple, non-toxic, substrate-mediated method was applied to deliver the naked FOXD3 plasmid into human fibroblasts. The transfection of FOXD3 increased cell proliferation and up-regulated the neural crest marker genes (FOXD3, SOX2, and CD271), stemness marker genes (OCT4, NANOG, and SOX2), and neural lineage-related genes (Nestin, β-tubulin and GFAP). The expression levels of stemness marker genes and neural crest maker genes in the FOXD3-transfected fibroblasts were maintained until the fifth passage. The FOXD3 reprogrammed fibroblasts based on the new method significantly rescued the neural function of the impaired zebrafish. The chitosan substrate-mediated delivery of naked plasmid showed feasibility in reprogramming somatic cells. Particularly, the FOXD3 reprogrammed fibroblasts hold promise as an easily accessible cellular source with neural crest stem-like behavior for treating neural diseases in the future., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

In this study, zwitterionic poly(sulfobetaine acrylamide) (pSBAA) nanocomposite hydrogels were synthesized and implemented as effective chronic wound dressings. The hydrogels exhibited reinforced mechanical properties from added hectorite nanoclay as a physical crosslinker in the polymer chains. Due to the strong interaction with water molecules via ionic solvation, the hydration of the zwitterionic nanocomposite hydrogels was superior to the non-ionic 2-hydroxyethyl methacrylate (pHEMA) hydrogels, which interacts with water molecules via hydrogen bonding. The pSBAA nanocomposite hydrogels cytotoxicity was accessed with NIH-3T3 fibroblast by the MTT assay, the results indicated negligible cytotoxicity after incubation for three days. In addition, the zwitterionic hydrogels displayed evident resistance to adsorption of bovine serum albumin (BSA), NIH-3T3 fibroblast, and bacteria of gram positive S. epidermidis and gram negative P. aeruginosa. The need for antifouling properties in a wound dressing is because commercial dressings removal typically damaging newly formed tissues and colonization of microorganisms occurs on the dressings. For clinical applications as wound dressings, we created normal and diabetic wounds on mice and compared newly developed pSBAA nanocomposite hydrogels with commercial available products. We demonstrated that non-adhesive pSBAA nanocomposite hydrogels enabled ready wound surface removal. Moreover, the wound recovery was conducted with normal and diabetic wounds on rat dorsal by visual observation and showed a complete heal after 10 and 12 days, respectively. Moreover, the histological examination of mice skin confirmed that the zwitterionic hydrogels exhibited thorough re-epithelialization and total formation of new connective tissues in the normal and diabetic wounds after 10 and 12 days, respectively, which was faster than commercial dressings. Consequently, we demonstrated that the pSBAA nanocomposite can serve as an effective dressing for wound management.

Pyoderma gangrenosum (PG) is a neutrophilic dermatosis of unknown etiology characterized by an ulcerative skin condition and confirmed through a diagnosis of exclusion. Management usually consists of systemic drug therapy, such as corticosteroids, sulfones, or immunosuppressants, either alone or in combination. Long-term use of these medications often has untold side effects. Hyperbaric oxygen therapy (HBOT) has been shown effective in the treatment of PG, reducing pain and tempering the need for medication. A case is presented of a 54-year-old woman with diabetes, hypertension, and a peptic ulcer who presented with painful, purulent ulcers on her buttocks, hands, and lower extremities of 2 weeks' duration. She was ultimately diagnosed with PG and provided 20 mg/day of oral prednisone for 1 week, tapered to 10 mg/day in the next week and then stopped. In addition, she received 12 sessions of HBOT - she breathed in 100% oxygen under 2.5 atmospheres absolute pressure for 90 minutes over 2 weeks. Her wounds healed without scarring. This excellent outcome including good wound healing, decreased pain, and reduced doses of systemic corticosteroids warrants additional study of the adjunctive use of HBOT for PG.

Adipose-derived adult stem cells (ASCs) have gained much attention because of their multipotency and easy access. Here we describe a novel chitosan-based selection (CS) system instead of the conventional plastic adherence (PA) to obtain the primary ASCs. The minimal amount of adipose tissue for consistent isolation of ASCs is reduced from 10 mL to 5 mL. The selection is based on the specific interaction between cells and chitosan materials, which separate ASCs by forming spheroids during primary culture. The primary culture period was reduced from 4 days to one day and more ASCs (ten-fold expansion) were achieved in a week. The average duration for obtaining 1 × 10(7) cells takes about seven days from 5 mL of adipose tissue, compared to 14 days using the conventional PA method from 10 mL of adipose tissue. The replicative senescence of CS-ASCs is not evident until the fifteenth passage (vs. eighth for the PA-ASCs). The obtained ASCs (CS-ASCs) have less doubling time for the same passage of cells and show greater stemness than those obtained from the conventional PA method (PA-ASCs). Moreover, CS-ASCs undergo trilineage differentiation more effectively than PA-ASCs. The greater differentiation potential of CS-ASCs may be associated with the enrichment and maintenance of CD271 positive cells by chitosan selection of primary culture., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Introduction: Autologous skin transplantation is a common treatment for patients with full-thickness burns. Postoperative wound care is essential for skin graft donor and recipient sites, but traditional wound dressings such as cotton and gauze do not form an effective barrier to bacteria, and patients can feel uncomfortable when replacing dressings., Materials and Methods: The goal of this study was to evaluate the use of an antimicrobial spray dressing (JUC Spray Dressing, NMS Technologies Co Ltd, Nanjing, China), with respect to its antimicrobial efficiency and the degree of pain experienced by patients., Results: The authors found the antimicrobial spray can reduce pain during the recovery period, while providing equivalent antibacterial protection to the control treatment (AQUACEL Hydrofiber Wound Dressing, ConvaTec, Bridgewater, NJ) based on skin culture tests. The spray did not adversely affect the wound site recovery. No significant side effects were present during the treatment period., Conclusion: This antimicrobial spray could potentially be used in wound dressing applications.

Cartilage is exposed to compression forces during joint loading. Therefore, exogenous stimuli are frequently used in cartilage tissue engineering strategies to enhance chondrocyte differentiation and extracellular matrix (ECM) secretion. In this study, human adipose-derived stem cells were seeded on a gelatin/polycaprolactone scaffold to evaluate the histochemical and functional improvement of tissue-engineered cartilage after hyperbaric oxygen/air treatment in a rabbit articular defect model. Behavior tests showed beneficial effects on weight-bearing and rear leg-supporting capacities after treatment of tissue-engineered cartilage with 2.5 ATA oxygen or air. Moreover, positron emission tomography images and immunohistochemistry staining demonstrated hydroxyapatite formation and increased ECM synthesis, respectively, at the tissue-engineered cartilage graft site after high pressure oxygen/air treatment. Based on these results, we concluded that hyperbaric oxygen and air treatment can improve the quality of tissue-engineered cartilage in vivo by increasing the synthesis of ECM.

Soft tissue defects exposing the Achilles tendon are common in patients who have undergone trauma or in those with pressure ulcers associated with vascular diseases. The purpose of this article was to present our experience of 11 patients who underwent reconstruction of soft tissue defects of the Achilles tendon using bipedicled fasciocutaneous flaps. Between August 2008 and August 2012, 11 patients were admitted to our hospital, presenting with soft tissue defects overlying the Achilles tendon. After adequate debridement, the 11 patients underwent bipedicled fasciocutaneous flap placement to resurface the complex soft tissue defects and provide a gliding surface for the exposed Achilles tendon. The patients' age, comorbidity, etiology, defect size and location, wound culture, skin graft size, complications, surgery duration, and follow-up period were reviewed. The 11 fasciocutaneous bipedicled flaps survived completely, and the wounds healed satisfactorily at a mean follow-up period of 20.9 months (range, 6-48 months). Only 1 flap was complicated with wound dehiscence and superficial necrosis of its lateral edge, which healed conservatively. The donor sites were covered with split-thickness skin grafts and healed well without complications. The bipedicled fasciocutaneous flap is a reliable flap for coverage of defects overlying the Achilles tendon, especially in patients with vascular problems and/or elderly patients. The ease of handling, short operative time, and early recovery of mobilization function are of great benefit to patients. Thus, the bipedicled fasciocutaneous flap can be a valuable alternative for defect reconstructions overlying the Achilles tendon, with satisfactory results both functionally and cosmetically.

Background: Deep sternal incisional surgical site infection is a serious and potentially life-threatening complication after open heart surgery. Although a rare post-operative complication, the rates of post-operative morbidity and mortality are greater in patients who develop a deep sternal incisional surgical site infection than in those who do not., Methods: We evaluated retrospectively the results of patients who developed a deep sternal incisional surgical site infection who were treated with either a pectoralis major flap or delayed primary closure after previous negative-pressure wound therapy (NWPT). From July 2007 to July 2012, 25 patients had a deep sternal incisional surgical site infection after open heart surgery in the Departments of Plastic Surgery and Cardiac Surgery of the Tri-Service General Hospital Medical Center. Sternal refixation was not performed in our patients., Results: In 15 patients, a unilateral or bilateral pectoralis major advancement flap with a myocutaneous or muscle flap was used. In seven patients, delayed primary closure was performed after NPWT. One patient received a rectus abdominis myocutaneous flap and another received a free anterior lateral thigh flap. One patient died after developing nosocomial pneumonia with severe sepsis after debridement., Conclusions: In our series, no patient required sternal re-fixation. Our findings suggest that delayed primary closure and use of a unilateral or bilateral pectoralis major flap following NPWT for a deep sternal incisional surgical site infection are simple and quick methods for managing such difficult surgical incisions even if the deep sternal surgical site infection is located in the lower one-third of the sternum.

The structural evolution of three-dimensional spheroids self-assembled from two different types of cells on selective biomaterials is demonstrated in this study. The two types of cells involved in the self-assembly are human mesenchymal stem cells (MSCs) and endothelial progenitor cells (EPCs). When seeded in different population ratios, they can create a variety of cellular patterns on different biomaterial substrates. When the two populations are matched in initial numbers, they are self-assembled in co-spheroids with different morphologies (i.e. randomly mixed, bumped, or concentric spheroids). The morphologies are influenced by the specific cell-substrate interaction possibly through integrin signaling, as well as a substrate-dependent regulation of heterophilic cell-cell interaction possibly through Notch signaling. In particular, the self-assembled core-shell concentric spheroids from adipose-derived MSCs and EPCs show a greater angiogenic effect in vitro. This study reveals the possibility to modulate the self-assembled morphology as well as the effect of cocultured cells by changing the cell culture substratum., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Hypertrophic scarring is related to persistent activation of transforming growth factor-β (TGF-β)/Smad signaling. In the TGF-β/Smad signaling cascade, the TGF-β type I receptor (TGFBRI) phosphorylates Smad proteins to induce fibroblast proliferation and extracellular matrix deposition. In this study, we inhibited TGFBRI gene expression via TGFBRI small interfering RNA (siRNA) to reduce fibroblast proliferation and extracellular matrix deposition. Our results demonstrate that downregulating TGFBRI expression in cultured human hypertrophic scar fibroblasts significantly suppressed cell proliferation and reduced type I collagen, type III collagen, fibronectin, and connective tissue growth factor (CTGF) mRNA, and type I collagen and fibronectin protein expression. In addition, we applied TGFBRI siRNA to wound granulation tissue in a rabbit model of hypertrophic scarring. Downregulating TGFBRI expression reduced wound scarring, the extracellular matrix deposition of scar tissue, and decreased CTGF and α-smooth muscle actin mRNA expression in vivo. These results suggest that TGFBRI siRNA could be applied clinically to prevent hypertrophic scarring.

This study investigated the profiling of polycyclic aromatic hydrocarbon- (PAH-) induced genotoxicity in cell lines and zebrafish. Each type of cells displayed different proportionality of apoptosis. Mitochondrial DNA (mtDNA) copy number was dramatically elevated after 5-day treatment of fluoranthene and pyrene. The notable deregulated proteins for PAHs exposure were displayed as follows: lamin-A/C isoform 3 and annexin A1 for benzopyrene; lamin-A/C isoform 3 and DNA topoisomerase 2-alpha for pentacene; poly[ADP-ribose] polymerase 1 (PARP-1) for fluoranthene; and talin-1 and DNA topoisomerase 2-alpha for pyrene. Among them, lamin-A/C isoform 3 and PARP-1 were further confirmed using mRNA and protein expression study. Obvious morphological abnormalities including curved backbone and cardiomegaly in zebrafish were observed in the 54 hpf with more than 400 nM of benzopyrene. In conclusion, the change of mitochondrial genome (increased mtDNA copy number) was closely associated with PAH exposure in cell lines and mesenchymal stem cells. Lamin-A/C isoform 3, talin-1, and annexin A1 were identified as universal biomarkers for PAHs exposure. Zebrafish, specifically at embryo stage, showed suitable in vivo model for monitoring PAHs exposure to hematopoietic tissue and other organs.

Background: Microsurgical free flap has gained the popularity over pedicle flap nowadays in the reconstruction of head and neck. However, pedicled flaps remain a promising alternative and have a remarkable position in selected patients. This review study aimed to determine the reliability and versatility of the lower trapezius musculocutaneous flap for reconstructing complex defects in the head and neck., Methods: Between 1993 and 2012, 22 male and 10 female patients underwent a total of 32 lower trapezius flap reconstructions for complex defects that included neoplasm extirpation (n = 21), radionecrosis (n = 6), dehisced laminectomy (n = 2), pressure sore (n = 2), and necrotizing fasciitis (n = 1). The most common site of defect was the perioral region, followed by the neck, posterior skull, back, temporal region, shoulder, and the upper arm. Flap design was based on the defect size and location, as well as the ability to close the donor site primarily and to preserve muscle function. Outcome has been evaluated by the hospital course, postoperative morbidity, mortality, resultant cosmetics, and function at donor and recipient sites., Results: Stable wound coverage with total flap survival was achieved in 30 (93.75%) patients, 2 patients had partial flap necrosis which required flap reinsertion and skin graft coverage. All donor sites were closed primarily. Seroma developed in 2 (6.25%) patients, which were solved by needle aspiration. All muscle function was preserved. Eight patients died of their primary disease., Conclusions: For selected patients who have advanced stage cancer, surgical sequelae after free flap surgery, unable to tolerate microsurgery, or special defect location, pedicled lower trapezius musculocutaneous flap provides efficient and effective reconstruction for complex defects especially in the head and neck.

Ascorbigen (ABG) is the predominant indole-derived compound from Brassica vegetables. In this study, we attempted to evaluate the effects of ABG on hair growth. To this end, we examined the proliferation of isolated human dermal papilla (DP) cells and keratinocytes after incubation in various concentrations (0-1.25 mM) of ABG. Furthermore, hair shaft regrowth was monitored in a mouse model of chemotherapy-induced alopecia (CIA), and hematoxylin and eosin staining was performed for histological analyses. We found that 1.25 mM ABG induced a 1.2-fold increase in the growth of DP cells, but not keratinocytes. However, ABG did not exert significant protective effects against CIA in the mouse model. These findings suggest that ABG may not be able to counteract CIA and that further investigation of the therapeutic potential of ABG in disease models is required., (Copyright © 2013 John Wiley & Sons, Ltd.)

Embryonic stem cells (ESCs) are pluripotent cells that can differentiate into various cell types, including keratinocyte-like cells, within suitable microniches. In this study, we aimed to investigate the effects of culture media, cell coculture, and a tissue-engineering biocomposite on the differentiation of mouse ESCs (MESCs) into keratinocyte-like cells and applied these cells to a surgical skin wound model. MESCs from BALB/c mice (ESC26GJ), which were transfected using pCX-EGFP expressing green fluorescence, were used to track MESC-derived keratinocytes. Weak expression of the keratinocyte early marker Cytokeratin 14 (CK-14) was observed up to 12 days when MESCs were cultured in a keratinocyte culture medium on tissue culture plastic and on a gelatin/collagen/polycaprolactone (GCP) biocomposite. MESCs cocultured with human keratinocyte cells (HKCs) also expressed CK-14, but did not express CK-14 when cocultured with human fibroblast cells (HFCs). Furthermore, CK-14 expression was observed when MESCs were cocultured by seeding HKCs or HFCs on the same or opposite side of the GCP biocomposite. The highest CK-14 expression was observed by seeding MESCs and HKCs on the same side of the GCP composite and with HFCs on the opposite side. To verify the effectiveness of wound healing in vivo, adipose-derived stem cells were applied to treat surgical wounds in nude mice. An obvious epidermis multilayer and better collagen deposition during wound healing were observed, as assessed by Masson staining. This study demonstrated the potential of keratinocyte-like differentiation from mesenchymal stem cells for use in promoting wound closure and skin regeneration.

The reconstruction of complex defects in the abdominal wall after wound infection or trauma can be challenging. In this article, a superficial inferior epigastric artery flap, a tensor fascia lata flap, and an anterolateral thigh flap used for 3 different abdominal wall reconstructions are described. The authors conclude that different specific abdominal wall defects can be successfully reconstructed using different pedicled flaps in simple and effective single-stage reconstructions.

Complete circumferential degloving injury of the digits usually results in a large cutaneous defect with tendinous structure and bone and joint exposure. When revascularization is not possible, a thin and adequately sized flap is required to resurface the defect, restore finger function, and prevent amputation. In this report, we present our experience with reconstruction of the entire circumferential degloving injury of the digits using free fasciocutaneous flaps. Between February 2006 and January 2011, 9 male patients with circumferential degloving injury of 9 digits underwent reconstruction using free fasciocutaneous flap transfer with the posterior interosseous artery flap, medial sural artery flap, anteromedial thigh flap, or radial forearm flap. The average flap size was 14.2 × 6.9 cm. Donor sites were closed primarily or covered with split-thickness skin graft. All flaps survived completely and the donor sites healed without complications. The mean follow-up period was 34.8 months. A maximum Kapandji score (10/10) was seen in 2 cases with crushed thumbs. All patients could achieve good key pinch and grasping functions. All skin flaps showed acceptable static 2-point discrimination and adequate protective sensation. Patient satisfaction for resurfaced digits averaged 9 on a 10-points visual analogic scale. In conclusion, the free fasciocutaneous flaps used were thin and did not interfere with finger movements. The patient's finger formed a smooth contour and acceptable functional results were obtained after reconstruction. This method may be a valuable alternative for reconstruction of entire circumferential avulsion injury of the digits., (Copyright © 2012 Wiley Periodicals, Inc.)

Introduction: The promotion of wound healing using dermal substitutes has become increasingly widespread, but the outcomes of substitute-assisted healing remain functionally deficient. Adipose-derived stem cells (ASCs) have been investigated widely in regenerative medicine and tissue engineering, and they have the potential to enhance wound healing. In this study, we focused on investigating the effects and mechanism of ASCs combined with an acellular dermal matrix (ADM) to treat full-thickness cutaneous wounds in a murine model., Methods: The ADM was prepared from the dorsal skin of nude mice by decellularization by treatment with trypsin followed by Triton X-100. The human ASCs were isolated and cultured from abdominal lipoaspirate. We created a rounded, 8-mm, full-thickness cutaneous wound in nude mice and divided the mice into the following 4 groups: silicon sheet cover only, silicon sheet with spreading ASCs, ADM only, and ASCs seeded on ADM. The granulation thickness was evaluated by histology after 7 days. Further comparisons between the ADM only and ASC-seeded ADM groups were undertaken by assessing the reepithelialization ratio and blood vessel density at postoperative days 9 and 14. Statistical analyses were conducted using Student 2-tailed t test. Immunofluorescent histology and ASC labeling were also performed to identify possible mechanisms., Results: The ADM was successfully prepared, and the cytometry analysis and differentiation assay provided the characterization of the human ASCs. A marked improvement in granulation thickness was detected in the ADM-ASC group in comparison with other 3 groups. A significantly increased rate of reepithelialization in the ADM-ASC group (80 ± 6%) compared to the ADM only group (60 ± 7%) was noted on postoperative day 9. The blood vessel density was evidently increased in the ADM-ASC group (7.79 ± 0.40 vessels per field) compared to the ADM only group (5.66 ± 0.23 vessels) on day 14. Cell tracking experiments demonstrated that labeled ASCs were colocalized with staining for VEGF or endothelial cell maker vWF after the transplantation of ADM-ASCs on postoperative day 14., Conclusions: Adipose-derived stem cells seeded on an ADM can enhance wound healing, promote angiogenesis, and contribute to newly formed vasculature, and VEGF-expressing ASCs can be detected after transplantation. This model could be used to improve the other clinical applications of ASCs and to decipher the detailed mechanism by which ASCs interact with wound tissue.

There is an urgent need to develop tissue-engineered cartilage for patients experiencing joint malfunction due to insufficient self-repairing capacity of articular cartilage. The aim of this research was to explore the effect of hyperbaric oxygen and air on tissue-engineered cartilage formation from human adipose-derived stem cells seeding on the gelatin/polycaprolactone biocomposites. The results of histological analyses indicate that under hyperbaric oxygen and air stimulation, the cell number of chondrocytes in cartilage matrix was not significantly increased, but the 1,9-dimethylmethylene blue assay showed that the glycosaminoglycans syntheses markedly increased compared to the control group. In quantification real-time polymerase chain reaction results, the chondrogenic-specific gene expression of SOX9, aggrecan, and COL2A1 were compared respectively. Within the limitation of this study, it was concluded that 2.5 atmosphere absolute oxygen and air may provide a stress environment to help cartilage tissue engineering development.

Purpose: Firewalking is a common Taoist cleansing ceremony in Taiwan, but burns associated with the practice have rarely been reported. We analyzed the patients with plantar burns from one firewalking ceremony., Materials and Methods: In one firewalking ceremony, 12 Taoist disciples suffered from contact burns to the soles of their feet while walking over burning coals. Eight of them had at least second-degree burns over areas larger than 1% of their total body surface areas (TBSAs). The age, sex, medical history, date of injury, time taken to traverse the fire pit, depth and TBSA of the burns, treatment, length of stay, and outcome were recorded and analyzed., Results: Deep, disseminated second- to third-degree burns were noted and healing took as long as three weeks in some patients. Because disseminated hypertrophic scars form after burns, the soles involved regain much of their tensile strength while walking. The patients experienced only a few difficulties in their daily lives three months after injury., Conclusion: From our experience treating patients with deep disseminated second- to third-degree plantar burns caused by firewalking, we conclude that they should be treated conservatively, with secondary healing rather than a skin graft., (Copyright © 2011 Elsevier Ltd and ISBI. All rights reserved.)

Unlabelled: Taiwan's skin banking program was initially set up to provide a ready source of cadaveric skin for patients with severe burns. However, human cadaveric skin may offer a useful alternative to conventional dressings in other wounds as well., Methods: In this retrospective review, cadaveric skin transplantation was used as temporary coverage in 145 patients with chronic ulcers, diabetic foot ulcers (DFU), necrotizing fasciitis, and acute traumatic wounds. Sex, age, number of debridements, and number of cadaveric skin transplantations were analyzed using statistical methods., Results: After clinical determination of engraftment 1 week after cadaveric skin allograft, skin samples harvested for histology in 15 cases revealed migration of epithelia from patient's skin to the surface of cadaver skin and the presence of granulation tissue in the base of the cadaver skin. All wounds exhibited good wound-bed preparation after cadaveric skin transplantation, and could eventually be resurfaced with a skin autograft., Conclusion: Human cadaveric skin, in addition to being the mainstay in burn therapy, is a good biological dressing for chronic ulcers, DFUs, necrotizing fasciitis, and acute traumatic wounds. .

Botryomycosis is an uncommon chronic suppurative granulomatous bacterial infection that can affect the skin and viscera. Clinically, lesions typically consist of small tender nodules from which draining sinuses may develop to expel a purulent discharge. Histopathological features include characteristic aggregation of microorganisms (grain) within the inflammatory infiltrate. The commonest causative organisms are Staphylococcus aureus and Pseudomonas aeruginosa, of others. Botryomycosis resulting from Prevotella melaninogenica has not been reported previously. We report the case of a middle-aged patient with botryomycosis presenting as nasal cutaneous fistulas caused by P. melaninogenica, which was successfully treated with surgical intervention combined with systemic antibiotic treatment., (© The Author(s). CED © 2011 British Association of Dermatologists.)

Poly(ADP-ribose) polymerase-1 (PARP1) plays critical roles in the regulation of DNA repair. Accordingly, small molecule inhibitors of PARP are being developed as agents that could modulate the activity of genotoxic chemotherapy, such as topoisomerase I poisons. In this study we evaluated the ability of the PARP inhibitor veliparib to enhance the cytotoxicity of the topoisomerase I poisons topotecan and camptothecin (CPT). Veliparib increased the cell cycle and cytotoxic effects of topotecan in multiple cell line models. Importantly, this sensitization occurred at veliparib concentrations far below those required to substantially inhibit poly(ADP-ribose) polymer synthesis and at least an order of magnitude lower than those involved in selective killing of homologous recombination-deficient cells. Further studies demonstrated that veliparib enhanced the effects of CPT in wild-type mouse embryonic fibroblasts (MEFs) but not Parp1(-/-) MEFs, confirming that PARP1 is the critical target for this sensitization. Importantly, parental and Parp1(-/-) MEFs had indistinguishable CPT sensitivities, ruling out models in which PARP1 catalytic activity plays a role in protecting cells from topoisomerase I poisons. To the contrary, cells were sensitized to CPT in a veliparib-independent manner upon transfection with PARP1 E988K, which lacks catalytic activity, or the isolated PARP1 DNA binding domain. These results are consistent with a model in which small molecule inhibitors convert PARP1 into a protein that potentiates the effects of topoisomerase I poisons by binding to damaged DNA and preventing its normal repair.

Background: Large or extensive gouty tophi on the feet can cause functional impairment, drainage sinus, and infected necrosis, finally resulting in complex soft-tissue defects with tendon, joint, bone, nerve, and vessel exposure. Reconstruction of complex soft-tissue defects of the foot is still challenging. The purpose of this report was to review the outcomes of free-flap reconstructive surgery for treating the metatarsal joint defects of the feet caused by chronic tophaceous gout., Methods: Ten patients who had large tophus masses (>5 cm) and ulceration on the feet were admitted to our hospital between September 2006 and September 2010. Six patients underwent free-flap reconstruction after debridement to resurface the circumferential wound, protect the underlying structures, and provide a gliding surface for exposed tendons. The patients' age, sex, comorbidities, location and size of the defects, reconstructive procedures, surgical outcomes, complications, follow-ups, and recurrence of tophaceous gout were reviewed and recorded., Results: The mean patient age was 49.8 years (range, 36-72 years). The average skin defect size was 92.2 cm(2) . Five patients were treated using free anterolateral thigh flaps, and 1, using a free medial sural flap. These free flaps were safely raised and showed excellent functional and cosmetic results, with a mean follow-up of 31.7 months (range, 7-50 months)., Conclusion: Chronic tophaceous gout can cause severe skin infection and necrosis, even resulting in deformity or sepsis if left untreated. Surgical debridement is inevitable in patients with extensive wounds. We reconstructed the large, ulcerative skin and soft-tissue defects on the dorsum of the foot by performing free-flap reconstruction after adequate debridement and achieved good functional and cosmetic results., (Copyright © 2011 Wiley Periodicals, Inc.)

Perineal wound complications following abdominoperineal resection (APR) are still frequent and most troublesome complications. We report the case of a 79-year-old male found to have the huge precoccygeal defect with infection after APR for rectal carcinoma. Before surgery, the patient received a complete course of chemoradiation therapy to treat for downgrade staging of the rectal malignancy. Extensive debridement of the perianal wound was performed for three times, followed by perianal reconstruction and packing and augmentation of the precoccygeal dead space with free latissimus dorsi (LD) muscle flap. Although persisted wound infection was still observed after reconstruction, the patient still led a good result after one time of further debridement and split-thickness skin graft. We selected free LD muscle flap to fill and seal off the large pelvic dead space without the needs to change the jackknife position of the patient after debridement. To the best of our knowledge, this is the first case reported in the literature with the radiation-associated perianal wound infection after APR reconstructed successfully by free LD muscle flap., (Copyright © 2011 Wiley-Liss, Inc.)

Background: To provide an alternative choice for covering trochanteric pressure sores, we report on a modified pedicle anterolateral thigh (ALT) myocutaneous flap based on the descending branch of the lateral circumflex femoral artery., Methods: From August 2007 to January 2010, 20 consecutive patients (10 men and 10 women) underwent 21 pedicled ALT myocutaneous flaps for reconstruction of trochanteric pressure sores. The flap was designed and elevated, resembling the ALT perforator flap including part of the vastus lateralis muscle but without skeletonisation of the perforators., Results: The mean age of patients was 79.4 years (range: 46-103). The mean follow-up period was 13.9 months (range: 3-32). The flaps were 8-21 cm long and 5-11 cm wide. All flaps healed without major complications. All donor sites were closed primarily without skin grafting and showed good aesthetic results. No recurrence was observed., Conclusions: This modified design of pedicled ALT myocutaneous flap without skeletonisation of perforators is a reliable and easily harvested flap for reconstruction of trochanteric pressure sores with limited morbidity., (Crown Copyright © 2010. Published by Elsevier Ltd. All rights reserved.)

Background: Surgical wound infection is a common complication, which increases the hospital stay and costs after surgery for head and neck cancer. In this study, we evaluated the effect of Triclosan-coated sutures on surgical wounds and analyzed the risk factors for wound infections in head and neck cancer surgery., Patients and Methods: From January 2007 to December 2009, 253 consecutive patients underwent wide excision of a head or neck cancer and reconstructive procedures. All patient data were collected prospectively. Of these, 241 patients were included in this study, divided into two groups. The Triclosan group contained 112 patients, whose surgical wounds were closed with Triclosan-coated sutures (Vicryl Plus). The control group included the remaining 129 patients, whose surgical wounds were closed with conventional Vicryl sutures. We conducted a retrospective, multivariate analysis to determine independent risk factors for the cervical wound infection., Results: The cervical wound infection rate was 14.9% (17/112) in the Triclosan group and 14.7% (19/129) in the control group, and these rates were not significantly different. Tumour stage and delayed intra-oral flap healing were independent risk factors for cervical wound infection., Conclusions: In this preliminary study, Triclosan-coated Vicryl sutures did not reduce the infection rate of cervical wounds after head or neck cancer surgery. The effectiveness of this suture material in head and neck cancer surgery should be considered with caution., (Crown Copyright © 2011. Published by Elsevier Ltd. All rights reserved.)

Background: The anterolateral thigh (ALT) flap is a frequent choice for free flap transfer in head and neck cancer reconstruction because of its versatility. Preoperative mapping of the perforator pedicles of an ALT flap is still a challenge because of variations in vasculature. Although computed tomographic angiography (CTA) is used increasingly to evaluate the peripheral vasculature, the use of this method for evaluating the perforators of an ALT flap has not been described in detail., Methods: From September 2008 to March 2009, 32 patients underwent preoperative CTA before free ALT flap transfer for head and neck cancer reconstruction. The perforators were marked on a 64-section multidetector CT image for each patient. The preoperatively mapped perforators were compared with the actual intraoperative findings. Flap success rates and associated morbidity and complications were recorded., Results: Preoperative CTA identified major variations in perforators. Eighty-four were found by preoperative CTA; 64 of these were mapped to be explored during the operation, and 13 additional perforators were identified during surgery. The accuracy rate of identifying the branching origin of the ALT perforators was 98% (63/64). All of the ALT flaps survived except for one with necrosis (survival rate 97%). There was no donor site morbidity., Conclusions: Preoperative mapping of perforators by CTA proved valuable in free ALT flap transfer and shortened the operation time significantly. This modality provides useful information for head and neck cancer reconstruction in difficult cases, especially in patients with large or through-and-through defects that might need multiple perforators in flap design., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

The development and characterization of an enhanced composite skin substitute based on collagen and poly(epsilon-caprolactone) are reported. Considering the features of excellent biocompatibility, easy-manipulated property and exempt from cross-linking related toxicity observed in the 1:20 biocomposites, skin substitutes were developed by seeding human single-donor keratinocytes and fibroblasts alone on both sides of the 1:20 biocomposite to allow for separation of two cell types and preserving cell signals transmission via micro-pores with a porosity of 28.8 +/- 16.1 microm. The bi-layered skin substitute exhibited both differentiated epidermis and fibrous dermis in vitro. Less Keratinocyte Growth Factor production was measured in the co-cultured skin model compared to fibroblast alone condition indicating a favorable microenvironment for epidermal homeostasis. Moreover, fast wound closure, epidermal differentiation, and abundant dermal collagen deposition were observed in composite skin in vivo. In summary, the beneficial characteristics of the new skin substitutes exploited the potential for pharmaceutical screening and clinical application.

The farnesyltransferase inhibitor tipifarnib exhibits modest activity against acute myelogenous leukemia. To build on these results, we examined the effect of combining tipifarnib with other agents. Tipifarnib inhibited signaling downstream of the farnesylated small G protein Rheb and synergistically enhanced etoposide-induced antiproliferative effects in lymphohematopoietic cell lines and acute myelogenous leukemia isolates. We subsequently conducted a phase 1 trial of tipifarnib plus etoposide in adults over 70 years of age who were not candidates for conventional therapy. A total of 84 patients (median age, 77 years) received 224 cycles of oral tipifarnib (300-600 mg twice daily for 14 or 21 days) plus oral etoposide (100-200 mg daily on days 1-3 and 8-10). Dose-limiting toxicities occurred with 21-day tipifarnib. Complete remissions were achieved in 16 of 54 (30%) receiving 14-day tipifarnib versus 5 of 30 (17%) receiving 21-day tipifarnib. Complete remissions occurred in 50% of two 14-day tipifarnib cohorts: 3A (tipifarnib 600, etoposide 100) and 8A (tipifarnib 400, etoposide 200). In vivo, tipifarnib plus etoposide decreased ribosomal S6 protein phosphorylation and increased histone H2AX phosphorylation and apoptosis. Tipifarnib plus etoposide is a promising orally bioavailable regimen that warrants further evaluation in elderly adults who are not candidates for conventional induction chemotherapy. These clinical studies are registered at www.clinicaltrials.gov as #NCT00112853.

Background and Objective: This study aimed to evaluate the expression and bioactivities of endothelin-1 (ET-1) in gingiva during cyclosporine A (CsA) treatment., Material and Methods: After establishing edentulous ridges, experimental rats were fed 30 mg/kg/day CsA while control animals received mineral oil for 4 weeks, after which a reverse transcription-polymerase chain reaction (RT-PCR) and/or immunohistochemistry was used to examine the expression of ET-1, its receptors, proliferating cell nuclear antigen (PCNA) and inducible nitric oxide synthase (iNOS) in gingivae. The roles of the endothelin receptors A and B (ET(A) and ET(B)) in CsA-enhanced expression of PCNA and iNOS were examined in cultured human gingival fibroblasts pretreated with receptor antagonists, by immunocytochemistry and RT-PCR, respectively., Results: The mRNA expression of ET-1, ET(A) and ET(B), as well as of PCNA and iNOS, was significantly greater in edentulous gingiva that received CsA compared with control gingiva. Immunohistochemistry revealed more cells positively stained for ET-1 and its receptors in the tissues of CsA-treated rats than in those of control rats. In fibroblast cultures, enhanced mRNA expression of ET-1, ET(A) and ET(B) was observed after CsA treatment at the concentrations of 10 and 100 ng/mL. Cyclosporine A-enhanced PCNA expression was somewhat reduced by blockade of ET(A), but not ET(B), whereas iNOS expression was somewhat reduced by blockade of ET(B)., Conclusion: Based on the present findings, we suggest that: (1) CsA upregulates the gingival expression of ET-1 and its receptors; and (2) ET(A) and ET(B) have different bioactivities, ET(A) being involved in cell proliferation and ET(B) being associated with iNOS expression.

Although agents that inhibit DNA synthesis are widely used in the treatment of cancer, the optimal method for combining such agents and the mechanism of their synergy is poorly understood. The present study examined the effects of combining gemcitabine (2',2'-difluoro 2'-deoxycytidine) and 7-ethyl-10-hydroxycamptothecin (SN-38; the active metabolite of irinotecan), two S-phaseselective agents that individually have broad antitumor activity, in human cancer cells in vitro. Colony-forming assays revealed that simultaneous treatment of Ovcar-5 ovarian cancer cells or BxPC-3 pancreatic cancer cells with gemcitabine and SN-38 resulted in antagonistic effects. In contrast, sequential treatment with these two agents in either order resulted in synergistic anti-proliferative effects, although the mechanism of synergy varied with the sequence. In particular, SN-38 arrested cells in S phase, enhanced the accumulation of gemcitabine metabolites, and diminished checkpoint kinase 1, thereby sensitizing cells in the SN-38 --> gemcitabine sequence. Gemcitabine treatment followed by removal allowed prolonged progression through S phase, contributing to synergy of the gemcitabine --> SN-38 sequence. These results collectively suggest that S-phase-selective agents might exhibit more cytotoxicity when administered sequentially rather than simultaneously.

Human DNA topoisomerase I (topo I) is an essential mammalian enzyme that regulates DNA supercoiling during transcription and replication. In addition, topo I is specifically targeted by the anticancer compound camptothecin and its derivatives. Previous studies have indicated that topo I is a phosphoprotein and that phosphorylation stimulates its DNA relaxation activity. The locations of most topo I phosphorylation sites have not been identified, preventing a more detailed examination of this modification. To address this issue, mass spectrometry was used to identify four topo I residues that are phosphorylated in intact cells: Ser(10), Ser(21), Ser(112), and Ser(394). Immunoblotting using anti-phosphoepitope antibodies demonstrated that these sites are phosphorylated during mitosis. In vitro kinase assays demonstrated that Ser(10) can be phosphorylated by casein kinase II, Ser(21) can be phosphorylated by protein kinase Calpha, and Ser(112) and Ser(394) can be phosphorylated by Cdk1. When wild type topo I was pulled down from mitotic cells and dephosphorylated with alkaline phosphatase, topo I activity decreased 2-fold. Likewise, topo I polypeptide with all four phosphorylation sites mutated to alanine exhibited 2-fold lower DNA relaxation activity than wild type topo I after isolation from mitotic cells. Further mutational analysis demonstrated that Ser(21) phosphorylation was responsible for this change. Consistent with these results, wild type topo I (but not S21A topo I) exhibited increased sensitivity to camptothecin-induced trapping on DNA during mitosis. Collectively these results indicate that topo I is phosphorylated during mitosis at multiple sites, one of which enhances DNA relaxation activity in vitro and interaction with DNA in cells.

Accurate estimates of the surface area of burns are important for initial fluid resuscitation and prognosis of burn victims. The area of the surface of the open hand is often used to estimate the area of a burn. Although it is accepted that hand surface area is generally equal to 1% of total body surface area among Caucasians, this ratio has not been verified for Chinese people. We used two-dimensional projection to estimate the hand surface areas of Chinese adults. The area of the palm was 0.44% of total body surface area among men and 0.42% among women. The area of the hand, including that of the thumb and fingers, was 0.76% of total body surface area among men and 0.73% among women. We conclude that the ratio between hand and total body surface area among Chinese adults differs from the ratio among Caucasians, and suggest an adjustment of the ratio for use with Chinese people.

Background: The radial forearm flap is frequently considered the first choice for tongue reconstruction, but the disadvantages of donor site morbidity are well known. The search for another thin skin flap as an alternative has led to the application of the medial sural artery perforator flap., Methods: We used 12 medial sural artery perforator flaps to reconstruct tongue and floor of mouth following cancer ablation. We paid attention to the major perforator (vein >or= 1 mm) as the vascular relay., Results: Most flaps were raised with a single perforator. The size of the skin paddle varied from 9 cm x 5 cm to 14 cm x 12 cm. The mean thickness of the flap was 5.2 mm. We reexplored 1 patient for venous insufficiency and could not salvage the flap., Conclusions: The thin medial sural artery perforator flap permits high accuracy of tongue restoration and reduces the morbidity at the donor site., ((c) 2007 Wiley Periodicals, Inc. Head Neck, 2008.)

Previous studies have suggested that Mcl-1, an antiapoptotic Bcl-2 homolog that does not exhibit appreciable affinity for the caspase 8-generated C-terminal Bid fragment (tBid), diminishes sensitivity to tumor necrosis factor-alpha-related apoptosis-inducing ligand (TRAIL). This study was performed to determine the mechanism by which Mcl-1 confers TRAIL resistance and to evaluate methods for overcoming this resistance. Affinity purification/immunoblotting assays using K562 human leukemia cells, which contain Mcl-1 and Bcl-x(L) as the predominant antiapoptotic Bcl-2 homologs, demonstrated that TRAIL treatment resulted in binding of tBid to Bcl-x(L) but not Mcl-1. In contrast, TRAIL caused increased binding between Mcl-1 and Bak that was diminished by treatment with the caspase 8 inhibitor N-(N(alpha)-acetylisoleucylglutamylthreonyl) aspartic acid (O-methyl ester)-fluoromethyl ketone (IETD(OMe)-fmk) or the c-Jun N-terminal kinase inhibitor SP600125. In addition, TRAIL caused increased binding of Bim and Puma to Mcl-1 that was inhibited by IETD(OMe)-fmk but not SP600125. Further experiments demonstrated that down-regulation of Mcl-1 by short hairpin RNA or the kinase inhibitor sorafenib increased TRAIL-induced Bak activation and death ligand-induced apoptosis in a wide variety of neoplastic cell lines as well as clinical acute myelogenous leukemia specimens. Collectively, these observations not only suggest a model in which Mcl-1 confers TRAIL resistance by serving as a buffer for Bak, Bim, and Puma, but also identify sorafenib as a potential modulator of TRAIL sensitivity.

An activating point mutation in Janus kinase 2 (JAK2 V617F) was recently identified in myelofibrosis with myeloid metaplasia (MMM). To further elucidate the pathogenic significance, we examined the JAK2 mutation burden, phosphorylation of JAK2 substrates and neutrophil apoptotic resistance. Immunoblotting revealed phosphorylation of signal transducer and activator of transcription-3 (STAT3) in all four JAK2 with high V617F mutant allele burden and seven of eight with intermediate mutant allele burden, but only one of eight with wild-type JAK2 (P<0.001). In contrast, STAT5 phosphorylation was undetectable in patient MMM neutrophils; and phosphorylation of Akt and extracellular signal-regulated kinases (ERKs) failed to correlate with JAK2 mutation status. Apoptosis was lower in MMM neutrophils (median 41% apoptotic cells, n=50) compared to controls (median 66%, n=9) or other myeloproliferative disorder patients (median 53%, n=11; P=0.002). Apoptotic resistance in MMM correlated with anemia (P=0.01) and the JAK2-V617F (P=0.01). Indeed, apoptotic resistance was greatest in MMM neutrophils with high mutant allele burden (median 22% apoptosis, n=5) than with intermediate burden (median 39%, n=23) or wild-type JAK2 (median 47%, n=22; P=0.008). These results suggest that mutant JAK2 contributes to MMM pathogenesis by constitutively phosphorylating STAT3 and diminishing myeloid cell apoptosis.

Previous studies demonstrated that ataxia telangiectasia mutated- and Rad3-related (ATR) kinase and its downstream target checkpoint kinase 1 (Chk1) facilitate survival of cells treated with nucleoside analogs and other replication inhibitors. Recent results also demonstrated that Chk1 is depleted when cells are treated with heat shock protein 90 (Hsp90) inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG). The present study examined the effects of 17-AAG and its major metabolite, 17-aminogeldanamycin (17-AG), on Chk1 levels and cellular responses to cytarabine in human acute myelogenous leukemia (AML) cell lines and clinical isolates. Cytarabine, at concentrations as low as 30 nM, caused activating phosphorylation of Chk1, loss of the phosphatase Cdc25A, and S-phase slowing. Conversely, treatment with 100 to 300 nM 17-AAG for 24 hours caused Chk1 depletion that was accompanied by diminished cytarabine-induced S-phase accumulation, decreased Cdc25A degradation, and enhanced cytotoxicity as measured by inhibition of colony formation and induction of apoptosis. Additional studies demonstrated that small inhibitory RNA (siRNA) depletion of Chk1 also sensitized cells to cytarabine, whereas disruption of the phosphatidylinositol 3-kinase (PI3k) signaling pathway, which is also blocked by Hsp90 inhibition, did not. Collectively, these results suggest that treatment with 17-AAG might represent a means of reversing checkpoint-mediated cytarabine resistance in AML.

Mantle-cell lymphoma (MCL) is a unique subtype of B-cell non-Hodgkin lymphoma (NHL) that behaves aggressively and remains incurable. In order to understand the pathogenesis of MCL and design new therapies, it is important to accurately analyze molecular changes in pathways dysregulated in MCL. We used antibody microarrays to compare patterns of protein expression between CD19(+) purified B lymphocytes from normal tonsil and 7 cases of histologically confirmed MCL. Protein overexpression was defined as a higher than 1.3-fold or 2-fold increase in at least 67% of tumor samples compared with normal B-cell control. Of the polypeptides, 77 were overexpressed using the higher than 1.3-fold cutoff, and 13 were overexpressed using the 2-fold cutoff. These included cell cycle regulators (regulator of chromosome condensation 1 [RCC1], murine double minute 2 [MDM2]), a kinase (citron Rho-interacting kinase [CRIK]), chaperone proteins (heat shock 90-kDa protein [Hsp90], Hsp10), and phosphatase regulators (A-kinase anchor protein 1 [AKAP149], protein phosphatase 5 [PP5], and inhibitor 2). The elevated expression of some of these polypeptides was confirmed by immunoblotting and immunohistochemistry, whereas elevated expression of others could not be confirmed, illustrating the importance of confirmatory studies. This study describes a novel technique that identifies proteins dysregulated in MCL.

Tissue engineering of skin based on collagen:PCL biocomposites using a designed co-culture system is reported. The collagen:PCL biocomposites having collagen:PCL (w/w) ratios of 1:4, 1:8, and 1:20 have been proven to be biocompatible materials to support both adult normal human epidermal Keratinocyte (NHEK) and mouse 3T3 fibroblast growth in cell culture, respectively, by Dai, Coombes, et al. in 2004. Films of collagen:PCL biocomposites were prepared using non-crosslinking method by impregnation of lyophilized collagen mats with PCL/dichloromethane solutions followed by solvent evaporation. To mimic the dermal/epidermal structure of skin, the 1:20 collagen:PCL biocomposites were selected for a feasibility study of a designed co-culture technique that would subsequently be used for preparing fibroblast/biocomposite/keratinocyte skin models. A 55.3% increase in cell number was measured in the designed co-culture system when fibroblasts were seeded on both sides of a biocomposite film compared with cell culture on one surface of the biocomposite in the feasibility study. The co-culture of human keratinocytes and 3T3 fibroblasts on each side of the membrane was therefore studied using the same co-culture system by growing keratinocytes on the top surface of membrane for 3 days and 3T3 fibroblasts underneath the membrane for 6 days. Scanning electron microscopy (SEM) and immunohistochemistry assay revealed good cell attachment and proliferation of both human keratinocytes and 3T3 fibroblasts with these two types of cells isolated well on each side of the membrane. Using a modified co-culture technique, a co-cultured skin model presenting a confluent epidermal sheet on one side of the biocomposite film and fibroblasts populated on the other side of the film was developed successfully in co-culture system for 28 days under investigations by SEM and immunohistochemistry assay. Thus, the design of a co-culture system based on 1:20 (w/w) collagen:PCL biocomposite membranes for preparation of a bi-layered skin model with differentiated epidermal sheet was proven in principle. The approach to skin modeling reported here may find application in tissue engineering and screening of new pharmaceuticals.

Agents that target topoisomerase I are widely utilized to treat human cancer. Previous studies have indicated that both the ataxia telangiectasia mutated (ATM)/checkpoint kinase (Chk) 2 and ATM- and Rad 3-related (ATR)/Chk1 checkpoint pathways are activated after treatment with these agents. The relative contributions of these two pathways to survival of cells after treatment with topoisomerase I poisons are currently unknown. To address this issue, we assessed the roles of ATR, Chk1, ATM, and Chk2 in cells treated with the topoisomerase I poisons camptothecin and 7-ethyl-10-hydroxycamptothecin (SN-38), the active metabolite of irinotecan. Colony forming assays demonstrated that down-regulation of ATR or Chk1 sensitized cells to SN-38 and camptothecin. In contrast, ATM and Chk2 had minimal effect of sensitivity to SN-38 or camptothecin. Additional experiments demonstrated that the Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin, which down-regulates Chk1, also sensitized a variety of human carcinoma cell lines to SN-38. Collectively, these results show that the ATR/Chk1 pathway plays a predominant role in the response to topoisomerase I inhibitors in carcinoma cells and identify a potential approach for enhancing the efficacy of these drugs.

The preparation and characterisation of collagen:PCL composites for manufacture of tissue engineered skin substitutes and models are reported. Films having collagen:PCL (w/w) ratios of 1:4, 1:8 and 1:20 were prepared by impregnation of lyophilised collagen mats by PCL solutions followed by solvent evaporation. In vitro assays of collagen release and residual collagen content revealed an expected inverse relationship between the collagen release rate and the content of synthetic polymer in the composite that may be exploited for controlled presentation and release of biopharmaceuticals such as growth factors. DSC analysis revealed the characteristic melting point of PCL at around 60 degrees C and a tendency for the collagen component, at high loading, to impede crystallinity development within the PCL phase. The preparation of fibroblast/composite constructs was investigated using cell culture as a first stage in mimicking the dermal/epidermal structure of skin. Fibroblasts were found to attach and proliferate on all the composites investigated reaching a maximum of 2 x 10(5)/cm(2) on 1:20 collagen:PCL materials at day 8 with cell numbers declining thereafter. Keratinocyte growth rates were similar on all types of collagen:PCL materials investigated reaching a maximum of 6.6 x 10(4)/cm(2) at day 6. The results revealed that composite films of collagen and PCL are favourable substrates for growth of fibroblasts and keratinocytes and may find utility for skin repair.

Phosphorus burns are a rarely encountered chemical burn, typically occurring in battle, industrial accidents, or from fireworks. Death may result even with minimal burn areas. Early recognition of affected areas and adequate resuscitation is crucial. Amongst our 2765 admissions between 1984 and 1998, 326 patients had chemical burns. Seven admissions were the result of phosphorus burns. Our treatment protocol comprises 1% copper sulfate solution for neutralization and identification of phosphorus particles, copious normal saline irrigation, keeping wounds moist with saline-soaked thick pads even during transportation, prompt debridement of affected areas, porcine skin coverage or skin grafts for acute wound management, as well as intensive monitoring of electrolytes and cardiac function in our burns center. Intravenous calcium gluconate is mandatory for correction of hypocalcemia. Of the seven, one patient died from inhalation injury and the others were scheduled for sequential surgical procedures for functional and cosmetic recovery. Cooling affected areas with tap water or normal saline, prompt removal of phosphorus particles with mechanical debridement, intensive monitoring, and maintenance of electrolyte balance are critical steps in initial management. Fluid resuscitation can be adjusted according to urine output. Early excision and skin autografts summarize our phosphorus burn treatment protocol.

Over the last half century, advances in treatment have changed the principal cause of death in burn patients from burn shock and wound sepsis to pulmonary sepsis, of which inhalation injury has always played a key role in morbidity and mortality. Even though Navar et al., Am. J. Surg. 1985;150:716-720 have noted that patients with inhalation injury had a mean fluid requirement of 5.8 ml/kg/% burn to achieve resuscitation from early burn shock, while patients without inhalation injury required only a mean fluid of 4.0 ml/kg/% burn, to achieve successful resuscitation in inhalation injured patients with minimum but adequate fluid has always been a challenge. To further define the difference of early fluid therapy between inhalation and noninhalation in extensively burned patients, a retrospective analysis was carried out in the Tri-Service General Hospital. Sixty-two flame burned patients (aged from 16 to 81 years-old with a mean age of 33.2+/-15.1 years: with burn size ranging from 30% to 98% TBSA with a mean burn size of 60.5+/-22.7%; 26 with inhalation injury; noninhalation 36) were reviewed during a 5-year period. The Parkland formula is the initial choice of fluid regimen with 4 ml/kg/% burn and the amount of replacement is monitored by urine output and is titrated to maintain urine output between 0.5 and 1.0 ml/kg/h. The mean amounts of fluid requirements of both inhalation and noninhalation burned patients were 3.1 +/- 1.0 and 2.3+/-0.8 ml/kg/% burn respectively (p < 0.05). Our study showed less fluid requirement for both inhalation and noninhalation injured patients in comparison with the Navar study and Parkland predictions in the first 24 h postburn. Furthermore, the inhalation injured patients definitely required volumes of fluid in excess of those required in noninhalation injured cases.