A Synthetic Protocell-Based Heparin Scavenger.

Heparin is a commonly applied blood anticoagulant agent in clinical use. After treatment, excess heparin needs to be removed to circumvent side effects and recover the blood-clotting cascade. Most existing heparin antidotes rely on direct heparin binding and complexation, yet selective compartmentalization and sequestration of heparin would be beneficial for safety and efficiency. However, such systems have remained elusive. Herein, a semipermeable protein-based microcompartment (proteinosome) is loaded with a highly positively charged chitosan derivative, which can induce electrostatics-driven internalization of anionic guest molecules inside the compartment. Chitosan-loaded proteinosomes are subsequently employed to capture heparin, and an excellent heparin-scavenging performance is demonstrated under physiologically relevant conditions. Both the highly positive scavenger and the polyelectrolyte complex are confined and shielded by the protein compartment in a time-dependent manner. Moreover, selective heparin-scavenging behavior over serum albumin is realized through adjusting the localized scavenger or surrounding salt concentrations at application-relevant circumstances. In vitro studies reveal that the cytotoxicity of the cationic scavenger and the produced polyelectrolyte complex is reduced by protocell shielding. Therefore, the proteinosome-based systems may present a novel polyelectrolyte-scavenging method for biomedical applications.

Optically Controlled Construction of Three-Dimensional Protein Arrays.

Protein crystallization is an important tool for structural biology and nanostructure preparation. Here, we report on kinetic pathway-dependent protein crystals that are controlled by light. Photo-responsive crystallites are obtained by complexing the model proteins with cationic azobenzene dyes. The crystalline state is readily switched to a dispersed phase under ultraviolet light and restored by subsequent visible-light illumination. The switching can be reversibly repeated for multiple cycles without noticeable structure deterioration. Importantly, the photo-treatment not only significantly increases the crystallinity, but creates crystallites at conditions where no ordered lattices are observed upon directly mixing the components. Further control over the azobenzene isomerization kinetics produces protein single crystals of up to ≈50 µm. This approach offers an intriguing method to fabricate metamaterials and study optically controlled crystallization.

Environment-Dependent Stability and Mechanical Properties of DNA Origami Six-Helix Bundles with Different Crossover Spacings.

The internal design of DNA nanostructures defines how they behave in different environmental conditions, such as endonuclease-rich or low-Mg2+ solutions. Notably, the inter-helical crossovers that form the core of such DNA objects have a major impact on their mechanical properties and stability. Importantly, crossover design can be used to optimize DNA nanostructures for target applications, especially when developing them for biomedical environments. To elucidate this, two otherwise identical DNA origami designs are presented that have a different number of staple crossovers between neighboring helices, spaced at 42- and 21- basepair (bp) intervals, respectively. The behavior of these structures is then compared in various buffer conditions, as well as when they are exposed to enzymatic digestion by DNase I. The results show that an increased number of crossovers significantly improves the nuclease resistance of the DNA origami by making it less accessible to digestion enzymes but simultaneously lowers its stability under Mg2+ -free conditions by reducing the malleability of the structures. Therefore, these results represent an important step toward rational, application-specific DNA nanostructure design.

Characterization of Reversibly Immortalized Calvarial Mesenchymal Progenitor Cells.

BACKGROUND: Bone morphogenetic proteins (BMPs) play a sentinel role in osteoblastic differentiation, and their implementation into clinical practice can revolutionize cranial reconstruction. Preliminary data suggest a therapeutic role of adenoviral gene delivery of BMPs in murine calvarial defect healing. Poor transgene expression inherent in direct adenoviral therapy prompted investigation of cell-based strategies. OBJECTIVE: To isolate and immortalize calvarial cells as a potential progenitor source for osseous tissue engineering. MATERIALS AND METHODS: Cells were isolated from murine skulls, cultured, and transduced with a retroviral vector bearing the loxP-flanked SV40 large T antigen. Immortalized calvarial cells (iCALs) were evaluated via light microscopy, immunohistochemistry, and flow cytometry to determine whether the immortalization process altered cell morphology or progenitor cell profile. Immortalized calvarial cells were then infected with adenoviral vectors encoding BMP-2 or GFP and assessed for early and late stages of osteogenic differentiation. RESULTS: Immortalization of calvarial cells did not alter cell morphology as demonstrated by phase contrast microscopy. Mesenchymal progenitor cell markers CD166, CD73, CD44, and CD105 were detected at varying levels in both primary cells and iCALs. Significant elevations in alkaline phosphatase activity, osteocalcin mRNA transcription, and matrix mineralization were detected in BMP-2 treated iCALs compared with GFP-treated cells. Gross and histological analyses revealed ectopic bone production from treated cells compared with controls in an in vivo stem cell implantation assay. CONCLUSION: We have established an immortalized osteoprogenitor cell line from juvenile calvarial cells that retain a progenitor cell phenotype and can successfully undergo osteogenic differentiation upon BMP-2 stimulation. These cells provide a valuable platform to investigate the molecular mechanisms underlying intramembranous bone formation and to screen for factors/small molecules that can facilitate the healing of osseous defects in the craniofacial skeleton.

Unusual sequela from a pencil stab wound reveals a retained graphite foreign body.

Penetrating pencil-tip injuries are common among children and usually resolve without long-term sequelae. However, failure to detect and remove embedded pencil fragments can result in increased morbidity or misdiagnoses of other, more serious, conditions. We report on the case of a 10-year-old boy stabbed with a pencil on his right chin. Initial treatment in the emergency department included irrigation and closure of the laceration. Following suture removal, the patient returned to the emergency department (with bright-purple drainage from the wound site). Radiographic evaluation led to the discovery of an embedded foreign body requiring surgical removal.

Pre-mastectomy sentinel lymph node biopsy: a strategy to enhance outcomes in immediate breast reconstruction.

The pre-mastectomy sentinel lymph node biopsy (PM-SLNB) is a technique that provides knowledge regarding nodal status prior to mastectomy. Because radiation exposure is associated with poor outcomes in breast reconstruction and reconstructed breasts can interfere with the planning and delivery of radiation therapy (RT), information regarding nodal status has important implications for patients who desire immediate breast reconstruction. This study explores the safety and utility of PM-SLNB as part of the treatment strategy for breast cancer patients desiring immediate reconstruction. We reviewed the charts of adult patients (≥18 years old) who underwent PM-SLNB from January 2004 to January 2011 at our institution. PM-SLNB was offered to patients with stage I or IIa, clinically and/or radiographically node-negative breast cancer who desired immediate breast reconstruction following mastectomy. PM-SLNB was also offered to patients with ductal carcinoma in situ if features concerning for invasive carcinoma were present. Ninety-one patients underwent PM-SLNB of 94 axillae. PM-SLNB was positive in 25.5% of breasts (n = 24). Nineteen node-positive patients (79.2%) have undergone or planning to undergo delayed reconstruction at our institution. Seventeen of these 19 node-positive patients (89.5%) have received adjuvant RT. Two patients (10.5%) elected against RT despite our recommendation for it. No biopsy-positive patient underwent immediate reconstruction or suffered a radiation-induced complication with their breast reconstruction. There were two minor complications associated with PM-SLNB, both in node-negative patients. This study demonstrates the utility of PM-SLNB in providing information regarding nodal status, and therefore the need for adjuvant RT, prior to mastectomy. This knowledge can be used to appropriately counsel patients regarding optimal timing of breast reconstruction.

DNA-origami-directed virus capsid polymorphism.

Viral capsids can adopt various geometries, most iconically characterized by icosahedral or helical symmetries. Importantly, precise control over the size and shape of virus capsids would have advantages in the development of new vaccines and delivery systems. However, current tools to direct the assembly process in a programmable manner are exceedingly elusive. Here we introduce a modular approach by demonstrating DNA-origami-directed polymorphism of single-protein subunit capsids. We achieve control over the capsid shape, size and topology by employing user-defined DNA origami nanostructures as binding and assembly platforms, which are efficiently encapsulated within the capsid. Furthermore, the obtained viral capsid coatings can shield the encapsulated DNA origami from degradation. Our approach is, moreover, not limited to a single type of capsomers and can also be applied to RNA-DNA origami structures to pave way for next-generation cargo protection and targeting strategies.

The transconjunctival deep-plane midface lift: a 9-year experience working under the muscle.

BACKGROUND: Rejuvenation of the midface with a natural-looking, safe, and long-lasting result is a challenge in aesthetic surgery. The ideal approach should be easy to perform, with minimal risk and significant benefit. The combination of transconjunctival exposure and preservation of the periosteum may result in lower morbidity than traditional midface rejuvenation. OBJECTIVES: The authors present their 9-year experience with the transconjunctival deep-plane midface lift (TDML) and discuss the benefits and limitations of this procedure relative to traditional approaches. METHODS: A retrospective review (2000-2009) was conducted of 124 consecutive patients treated by the same surgeon (JWF) with the TDML approach. The technique combines transconjunctival and preperiosteal dissection under direct vision. Collected data included patient demographics, operative technique, concomitant procedures, and postoperative results. Complications and revisions were reviewed to assess safety and long-term efficacy. RESULTS: Patients included 97 women and 27 men, with a minimum of 13 months of postoperative follow-up (median, 56 months). The mean operating time for upper blepharoplasty with TDML was less than 150 minutes. No significant complications occurred. One patient required reexcision of redundant lower eyelid skin, and another patient underwent secondary excision via traditional midface lift. CONCLUSIONS: The TDML procedure is safe and effective. The technique is readily applicable and more "forgiving" than the traditional midface lift. With proper patient selection, the limited soft-tissue dissection reduces surgical morbidity. Patients with excessive skin redundancy or festoons should be treated with more traditional techniques.

Optically Responsive Protein Coating of DNA Origami for Triggered Antigen Targeting.

DNA nanostructures have emerged as modular building blocks in several research fields including biomedicine and nanofabrication. Their proneness to degradation in various environments has led to the development of a variety of nature-inspired protection strategies. Coating of DNA origami nanostructures with proteins can circumvent degradation and alter their properties. Here, we have used a single-chain variable antibody fragment and serum albumin to construct positively charged and stimuli-responsive protein-dendron conjugates, which were complexed with DNA origami through electrostatic interactions. Using a stepwise assembly approach, the coated nanostructures were studied for their interaction with the corresponding antigen in fluorescence-based immunoassays. The results suggest that the antibody-antigen interaction can be disturbed by the addition of the bulky serum albumin. However, this effect is fully reversible upon irradiation of the structures with an optical stimulus. This leads to a selective dissociation of the serum albumin from the nanostructure due to cleavage of a photolabile group integrated in the dendron structure, exposing the antibody fragment and enabling triggered binding to the antigen, demonstrating that serum albumin can be considered as an externally controlled "camouflaging" agent. The presented stimuli-responsive complexation approach is highly versatile regarding the choice of protein components and could, therefore, find use in DNA origami protection, targeting, and delivery as well as their spatiotemporal control.

Prospective Cancer Therapies Using Stimuli-Responsive DNA Nanostructures.

Nanostructures based on DNA self-assembly present an innovative way to address the increasing need for target-specific delivery of therapeutic molecules. Currently, most of the chemotherapeutics being used in clinical practice have undesired and exceedingly high off-target toxicity. This is a challenge in particular for small molecules, and hence, developing robust and effective methods to lower these side effects and enhance the antitumor activity is of paramount importance. Prospectively, these issues could be tackled with the help of DNA nanotechnology, which provides a route for the fabrication of custom, biocompatible, and multimodal structures, which can, to some extent, resist nuclease degradation and survive in the cellular environment. Similar to widely employed liposomal products, the DNA nanostructures (DNs) are loaded with selected drugs, and then by employing a specific stimulus, the payload can be released at its target region. This review explores several strategies and triggers to achieve targeted delivery of DNs. Notably, different modalities are explained through which DNs can interact with their respective targets as well as how structural changes triggered by external stimuli can be used to achieve the display or release of the cargo. Furthermore, the prospects and challenges of this technology are highlighted.

Latissimus dorsi/rib intercostal perforator myo-osseocutaneous free flap reconstruction in composite defects of the scalp: case series and review of literature.

Adequate coverage of complex, composite scalp defects in previously radiated, infected, or otherwise compromised tissue represents a challenge in reconstructive surgery. To provide wound closure with bony protection to the brain, improve cranial contour, and prevent or seal cerebrospinal fluid (CSF) leaks, composite free tissue transfer is a reliable and safe option. We report our experience with the latissimus dorsi/rib intercostal perforator myo-osseocutaneous free flap in the reconstruction of bony and soft tissue defects of the cranium and overlying scalp. The surgical technique, design, and outcomes of the latissimus dorsi/rib intercostal perforator myo-osseocutaneous free flap reconstruction in five patients with cranial defects between 2003 and 2007 were retrospectively evaluated. Patient characteristics, defect size, underlying cause, reconstructive details, and complications were analyzed. All patients (age 43 to 81) had composite defects ranging from 36 to 750 cm2 (mean size 230 cm2) for the bony component and from 16 to 400 cm2 (mean size 170 cm2) for the soft tissue defect. All patients had a history of prior or current infection of the affected area, and two patients had a CSF leak. Defects were due to malignancy and infection (n = 2), infiltrative cutaneous mucormycosis with osteomyelitis (n = 1), and hemorrhagic stroke requiring craniectomy (n = 2), complicated by infection and failed cranioplasty in one patient and continuous CSF leak in the other. The latissimus dorsi composite free flap consisting of skin, muscle, and vascularized rib can successfully cover large complex cranial defects, provide skeletal support, improve contour, and significantly enhance functional outcome with limited donor site morbidity.

Wide local excision of perianal Paget's disease with gluteal flap reconstruction: an interdisciplinary approach.

Perianal Paget's disease (PPD) is an extremely rare condition characterized as intraepithelial adenocarcinoma of unclear etiology. It can be either primary or secondary. The disease typically presents as an eczema-like, erythematous, and painful skin lesion that is associated with pruritus. It is usually misdiagnosed as a common anorectal problem. Surgical excision is the preferred treatment of PPD, with the specific technique being dependent upon disease invasiveness. The treatment may involve reconstructive surgery. A 61-year-old female with a history of rectal pain and intermittent pruritus for the past two years presented with large painful lesions in her perianal area including the anal verge, diagnosed as primary PPD. After excluding other malignancies elsewhere, a laparoscopic ileostomy followed by a wide local excision (WLE) of the PPD was performed by a colorectal team. Reconstruction of the defect with gluteal advancement flaps was performed by the plastic surgeon. The patient recovered uneventfully. Her surgical site showed healing without flap compromise, widely open anal opening, and full sphincter control at the three-month follow-up exam. The patient returned to normal function after ileostomy closure. WLE with bilateral V-Y gluteal flap advancement is a feasible treatment for primary PPD.

"NACsomes": A new classification system of the blood supply to the nipple areola complex (NAC) based on diagnostic breast MRI exams.

BACKGROUND: Breast MRIs have become increasingly common in breast cancer work-up. Previously obtained breast MRIs may facilitate oncoplastic surgery by delineating the blood supply to the nipple-areola complex (NAC). The aim of this study was to identify and classify the in vivo blood supply to the NAC using breast MRI exams. METHODS: Breast MRIs obtained over a one-year period were retrospectively reviewed. Patients with negative MRI findings (BI-RADS category 1) were included; patients with diagnoses of breast cancer or previous breast surgery were excluded. Twenty-six patients were evaluated. Dominant blood supply was determined by maximum filling at 70 s post-contrast. Blood supply to the NAC was classified into five anatomic zones: medial (type I), lateral (type II), central (type III), inferior (type IV) and superior (type V). RESULTS: Patient age ranged from 33 to 70 years. Fifty-two breasts were evaluated and 80 source vessels were identified (37 right, 43 left). Twenty-eight breasts had type I only blood supply, 22 breasts had multi-zone blood supply (type I + II, n = 20; type I + III n = 2), one breast had type II only blood supply, and a single breast had type III only blood supply. Anatomic symmetry was observed in 96% of patients. CONCLUSION: This study utilized MRI to evaluate in vivo vascular anatomy of the NAC, classify NAC perfusion ("NACsomes"), and assess vascular symmetry between breasts. Superomedial source vessels supplying the NAC were predominant. Preoperatively defining NAC blood supply may aid planning for oncoplastic procedures.

Impaired vascular reactivity of isolated rat middle cerebral artery after cortical spreading depression in vivo.

Cortical spreading depression (CSD) is accompanied by hyperemia followed by long-lasting hypoperfusion and impaired cerebrovascular reactivity. The authors show that vasodilation to extraluminal acidosis (pH 7.0) and increased concentrations of extraluminal potassium (12, 20, 40 mmol/L) was significantly reduced in isolated rat middle cerebral arteries after CSD in vivo before the artery was isolated, compared with sham-operated controls. Application of 80-mmol/L potassium induced vasoconstriction after CSD. Therefore, the impairment of vascular reactivity after CSD in vivo occurs, at least in part, at the vascular level itself.

A ten-year review of myelodysplastic defect management and use of a novel closure technique with V-Y crescentic rotation advancement flaps.

The reconstructive goals for myelodysplastic defects are to provide a multilayered, tension-free and well-vascularized closure to prevent cerebrospinal fluid leakage, wound infection or breakdown and to optimize neurologic outcomes. We reviewed our ten-year experience with myelodysplastic defects and our preferred technique for large defects utilizing paraspinous flaps followed by V-Y crescentic rotation advancement flaps. A retrospective chart review was performed on all myelodysplastic defects closed at the University of Chicago Medicine from 2002 to 2012. Twenty-three patients were treated: eight were closed using V-Y crescentic rotation advancement flaps, eight primarily, two with transposition flaps and five with bilateral latissimus dorsi and gluteus maximus myocutaneous flaps. Patient defect characteristics, reconstructive details, follow up time, and wound complications were analyzed. The primary closure group included eight patients. There was one minor complication and two major complications that required debridement and plastic surgery consultation in this group. The transposition group included two patients and had no wound healing issues. The latissimus and gluteus myocutaneous group included five patients and had one minor wound healing issues. The V-Y crescentic group included eight patients. There were four minor wound breakdowns in the lateral donor sites and one major wound complication involving a CSF leak, meningitis and wound breakdown that required debridement. The groups were stratified by size, <5 cm and >5 cm, and further analyzed. Bilateral V-Y crescentic rotation advancement flap is a useful option when confronted with large myelodysplastic defects. It provides a multilayer, tension-free wound closure and spares the gluteus maximus and latissimus dorsi muscle groups.

CASE REPORT Case Report and Review of the Literature: Deep Inferior Epigastric Perforator Flap for Breast Reconstruction After Abdominal Recontouring.

OBJECTIVE: The report herein presents a case of a 49-year-old woman with left breast cancer who presented seeking immediate autologous reconstruction. Surgical history included an abdominal hysterectomy and an abdominal contouring procedure. This is a first description of a deep inferior epigastric perforator flap after abdominal wall manipulation of this magnitude. METHODS: Computed tomographic angiography identified patent medial row perforators. Doppler confirmed the location of the perforators. The flap was designed with the inferior incision at the previous lower abdominal scar. Laser-assisted indocyanine green imaging confirmed adequate flap perfusion on the basis of a single left deep inferior epigastric perforator. RESULTS: The flap was harvested on one perforator and anastomosed to the internal mammary system. The postoperative course was complicated by venous anastomosis kinking, requiring revision, but otherwise unremarkable. CONCLUSION: Computed tomographic angiography confirmed presence of perforators, communication with the deep inferior epigastric system, and location acceptable for flap design. Laser-assisted indocyanine green angiography facilitated perforator selection and provided intraoperative assessment of flap perfusion. Utilization of these modalities allowed safe completion of an operation considered contraindicated by conventional algorithms and highlights their role in complex perforator flap reconstruction.

Facilitating harvest of the serratus fascial flap with ultrasonic dissection.

OBJECTIVES: Upper extremity reconstruction presents a functional and aesthetic challenge in plastic surgery. Exposure of vital structures often requires vascularized soft tissue coverage to achieve primary wound healing and optimize functional results. Specifically, the serratus fascial flap may satisfy the functional and cosmetic requirements for small- to medium-sized soft tissue defects of the upper extremity with limited donor site morbidity. We describe our technique of serratus fascial flap harvest, using the Harmonic SYNERGY curved blade (Ethicon Endo-Surgery, Cincinnati, Ohio). MATERIAL AND METHODS: A 21-year-old, right-hand-dominant, male carpenter and martial arts expert was involved in a motorcycle collision and sustained a left-hand dorsal degloving injury and extensor tendon rupture. Soft tissue reconstruction was performed with a serratus fascial free flap, immediate split-thickness skin graft, and palmaris longus tendon grafts. The flap was harvested with the Harmonic blade, which utilizes ultrasonic energy translated into mechanical energy, thereby allowing dissection and hemostasis simultaneously. RESULTS: Flap elevation proceeded facilely using the Harmonic curved blade. The patient had no postoperative complications involving his flap or donor site. The closed suction drain in the donor site was removed on postoperative day 3, and the patient was discharged on postoperative day 10. The patient is doing well at 4 months follow-up. CONCLUSION: The Harmonic blade may assist in the dissection of the serratus fascial flap by aiding with hemostasis and minimizing surrounding tissue damage. This may reduce flap damage associated with harvesting techniques as well as decrease donor site seroma formation.

Split lateral iliac crest chimera flap: utility of the ascending branch of the lateral femoral circumflex vessels.

BACKGROUND: Complex head and neck reconstruction often requires multiple tissue components to restore form and function to the traumatized area. Here, the authors describe the split lateral iliac crest chimera flap and demonstrate the utility of the ascending branch of the lateral femoral circumflex system to provide vascularized bone for complex head and neck reconstruction. METHODS: A retrospective case series analysis was performed for patients undergoing complex head and neck reconstruction utilizing the split lateral iliac crest chimera flap to provide vascularized bone and soft tissue. The blood supply to the lateral iliac crest was via the ascending branch of the lateral femoral circumflex system, and the soft tissue was supplied by the transverse and descending branches of the circumflex system. RESULTS: Four patients with advanced recurrent head and neck cancer undergoing split lateral iliac crest chimera reconstruction between November of 2007 and April of 2009 were included. Three patients required reconstruction of segmental mandibulectomy defects, and one required reconstruction of a maxillectomy defect. All components of the chimeric flaps in each patient survived. Bone vascularity was confirmed with triphasic bone scans within the first week postoperatively in two patients. CONCLUSIONS: The split lateral iliac crest chimera flap employs distinct branches of the lateral femoral circumflex system to supply the split lateral iliac crest and soft tissue of the thigh, each isolated on separate vascular leashes connected to a common source vessel. Through inclusion of a vascularized bone component, the flap extends the versatility of the lateral femoral circumflex flap for complex head and neck reconstruction requiring both hard-tissue and soft-tissue replacement.

Reconstruction of scalp and forehead defects.

Reconstruction of scalp and forehead defects is a complex field with a broad variety of reconstructive options. The thought process and techniques used for reconstruction of scalp and forehead defects are the subject of this article.

Pedicled thoracodorsal artery perforator flap in breast reconstruction: clinical experience.

BACKGROUND: The thoracodorsal artery perforator (TDAP) flap has been described for reconstruction of the head and neck, trunk and extremities. Yet, its use as a pedicled flap in breast reconstruction has not gained wide popularity and has not been widely documented, especially not for complete breast reconstruction or in combination with expanders or permanent implants. The authors present their clinical experience with the thoracodorsal artery perforator flap in breast reconstruction. METHODS: From February 2007 to February 2009, eighteen patients had breast reconstruction utilizing a TDAP flap. Retrospective analyzes of patient characteristics, breast history, clinical indications, complications and outcomes were performed. The follow-up period ranged from 1 to 17 months. RESULTS: Eleven patients had complete breast reconstruction using a TDAP flap with simultaneous insertion of an expander or implant. Four cases were partial reconstruction to gain additional volume after previous breast reconstruction and the 3 other cases were reconstruction after lumpectomy. All flaps survived. Two case required evacuation of hematoma. One case had late extrusion of the expander after expansion in the previously irradiated tissue, requiring expander removal. There were no donor site complications. CONCLUSIONS: The TDAP flap has proven to be a reliable flap with minimal donor site morbidity. Patients who had radiation treatment prior to reconstruction with pronounced radiated chest skin changes might still benefit from additional tissue from the LD muscle.