Obtaining Lissamine Green 1% Solution for Clinical Use.

PURPOSE: With new compounding pharmacy laws, the only economically feasible approach to using lissamine is through dye-impregnated strips. This research aims to determine the concentration of lissamine that can be obtained using a single commercially available lissamine strip. With the optimal vital staining requiring 1% concentration of lissamine, we sought to obtain this concentration using supplies in an ordinary ophthalmology clinic. METHODS: A standard curve was generated using compounded lissamine green 1% solution. Serial dilutions were made with 3 different diluents and measured using a spectrophotometer at a wavelength of 633 µm. Combinations of the number of strips, amount of solvent, and absorption time were performed to obtain a 1% solution. Cost analyses were performed to select the most economical method. RESULTS: Single lissamine strips wetted with any of the diluents produced 0.17% ± 0.05% (95% confidence interval) lissamine solution, a 5-fold weaker concentration than the optimal for vital staining. Combinations of 4 strips in 200 µL (4 drops) for 1 minute and 2 strips in 200 µL for 5 minutes were found to reach concentrations of 1%. Cost analysis showed that the 2 strip/4 drops/5 minutes method costs $0.67 and the 4 strips/4 drops/1 minute method $1.27. CONCLUSIONS: Use of a single lissamine strip leads to suboptimal concentrations for vital staining. With only the addition of disposable microcentrifuge tubes to the clinical setting, ophthalmologists can make 1% solutions of lissamine. This solution is both more economical and in compliance with both state and national compounding laws.

Correlation between papilledema grade and diffusion-weighted magnetic resonance imaging in idiopathic intracranial hypertension.

BACKGROUND: To explore the relationship between diffusion-weighted magnetic resonance imaging (DWI) hyperintensity of the optic nerve head (ONH) and papilledema grade in patients with idiopathic intracranial hypertension (IIH). METHODS: A retrospective chart review was conducted of patients with definitively diagnosed IIH by clinical examination and visual field (VF) analysis who underwent orbital magnetic resonance imaging (MRI) within 4 weeks of diagnosis. A neuroradiologist masked to the diagnosis assessed the results of DWI for each eye independently and graded the signal intensity of the ONH into none, mild, and prominent categories. DWI grading was compared with papilledema grade and visual field mean deviation (VFMD) by Spearman rank correlation analysis and t-tests. RESULTS: Forty-two patients were included in the study. A statistically significant difference (P = 0.0195) was found between papilledema grade and patients with prominent DWI findings (n = 16; mean papilledema grade 3.75 ± 1.25) vs mild or no ONH hyperintensity (n = 26; mean papilledema grade 2.79 ± 1.24) at the time of initial diagnosis. DWI hyperintensity of the ONH at diagnosis was also found to be significantly correlated with the degree of papilledema at follow-up (ρ = 0.39, P = 0.0183) but not with VFMD. CONCLUSIONS: We found a significant correlation between the severity of papilledema and ONH hyperintensity on DWI in patients with IIH but not with VF loss or other visual parameters. These findings may offer insight into the pathophysiology of papilledema in IIH and provide a surrogate marker for the presence and severity of papilledema.

Collagen IV-modified scaffolds improve islet survival and function and reduce time to euglycemia.

Islet transplantation on extracellular matrix (ECM) protein-modified biodegradable microporous poly(lactide-co-glycolide) scaffolds is a potential curative treatment for type 1 diabetes mellitus (T1DM). Collagen IV-modified scaffolds, relative to control scaffolds, significantly decreased the time required to restore euglycemia from 17 to 3 days. We investigated the processes by which collagen IV-modified scaffolds enhanced islet function and mediated early restoration of euglycemia post-transplantation. We characterized the effect of collagen IV-modified scaffolds on islet survival, metabolism, and insulin secretion in vitro and early- and intermediate-term islet mass and vascular density post-transplantation and correlated these with early restoration of euglycemia in a syngeneic mouse model. Control scaffolds maintained native islet morphologies and architectures as well as collagen IV-modified scaffolds in vivo. The islet size and vascular density increased, while ß-cell proliferation decreased from day 16 to 113 post-transplantation. Collagen IV-modified scaffolds promoted islet cell viability and decreased early-stage apoptosis in islet cells in vitro-phenomena that coincided with enhanced islet metabolic function and glucose-stimulated insulin secretion. These findings suggest that collagen IV-modified scaffolds promote the early restoration of euglycemia post-transplantation by enhancing islet metabolism and glucose-stimulated insulin secretion. These studies of ECM proteins, in particular collagen IV, and islet function provide key insights for the engineering of a microenvironment that would serve as a platform for enhancing islet transplantation as a viable clinical therapy for T1DM.

Asymptomatic intracardiac mass in a 14-year-old girl with granulomatosis with polyangiitis: Case report.

Granulomatosis with polyangiitis (GPA; Wegener's granulomatosis) is a systemic necrotizing vasculitis of unknown etiology that commonly involves the upper airways, lungs, and kidneys. Cardiac involvement with an intracardiac mass is an exceedingly rare manifestation of this disease, especially in the pediatric population where, to our knowledge, only one article exists to date that has described such a finding. In this report, we present the case of an adolescent female who initially presented with renal failure and an intracardiac mass. Subsequent work-up led to a diagnosis of granulomatosis with polyangiitis (GPA). Cardiac manifestations in pediatric GPA are not common; however, they may be more prevalent than reported given recent adult literature and concern for clinically silent abnormalities.

Phosphatidylserine immobilization of lentivirus for localized gene transfer.

Localized and efficient gene transfer can be promoted by exploiting the interaction between the vector and biomaterial. Regulation of the vector-material interaction was investigated by capitalizing on the binding between lentivirus and phosphatidylserine (PS), a component of the plasma membrane. PS was incorporated into microspheres composed of the copolymers of lactide and glycolide (PLG) using an emulsion process. Increasing the weight ratio of PS to PLG led to a greater incorporation of PS. Lentivirus, but not adenovirus, associated with PS-PLG microspheres, and binding was specific to PS relative to PLG alone or PLG modified with phosphatidylcholine. Immobilized lentivirus produced large numbers of transduced cells, and increased transgene expression relative to virus alone. Microspheres were subsequently formed into porous tissue engineering scaffolds, with retention of lentivirus binding. Lentivirus immobilization resulted in long-term and localized expression within a subcutaneously implanted scaffold. Microspheres were also formed into multiple channel bridges for implantation into the spinal cord. Lentivirus delivery from the bridge produced maximal expression at the implant and a gradient of expression rostrally and caudally. This specific binding of lentiviral vectors to biomaterial scaffolds may provide a versatile tool for numerous applications in regenerative medicine or within model systems that investigate tissue development.

Lentivirus delivery by adsorption to tissue engineering scaffolds.

Biomaterial scaffolds capable of localized gene delivery are being investigated for numerous regenerative medicine applications and as model systems for fundamental studies of tissue formation. In this manuscript, we investigate the delivery of lentivirus from a tissue engineering scaffold using a surface immobilization strategy. Poly(lactide-co-glycolide) (PLG) was employed as the biomaterial for delivery, which has been widely used for a number of tissue engineering applications. The virus was immobilized by freezing and subsequent lyophilization of the virus with the scaffold. The presence of sucrose during freezing and lyophilization maintained the activity of the lentivirus, and was similar to an adenovirus control. Collagen and fibronectin were investigated for their ability to enhance surface immobilization. Fibronectin modestly increased binding and transduction of the adenovirus, yet did not significantly impact the lentivirus delivery. Most of the immobilized lentivirus was released from the scaffold within 24 h. In vivo implantation of the scaffolds yielded transgene expression that persisted for at least 4 weeks. These findings indicate the potential for delivering lentivirus from tissue engineering scaffolds using a surface immobilization strategy. To our knowledge, this report is the first to investigate lentivirus delivery from porous tissue engineering scaffolds. Delivery of lentiviral vectors from PLG scaffolds could provide an efficient and versatile gene delivery system for use with in vitro and in vivo models of tissue formation, and ultimately for therapeutic applications.

Abdominal wall reconstruction: lessons learned from 200 "components separation" procedures.

OBJECTIVES: To determine the efficacy and describe the evolution of the "components separation" technique for abdominal wall repair in 200 consecutive patients. DESIGN: Retrospective medical record review. SETTING: Northwestern Memorial Hospital, Chicago, Illinois. PATIENTS: Two hundred consecutive patients who underwent ventral hernia repair using the components separation technique. INTERVENTIONS: Biological and permanent meshes were used in select patients to augment the repair of the midline fascial closure but were not used as "bridging" materials. MAIN OUTCOME MEASURES: Hernia recurrence rates and major and minor complication rates for the overall series and for the different techniques. RESULTS: Primary components separation (n = 158) yielded a 22.8% recurrence rate. Closure of the midline tissues with augmentation of the repair using an acellular cadaveric dermis underlay (n = 18) had a 33.3% recurrence rate requiring a second operation, whereas intra-abdominal soft polypropylene mesh (n = 18) had 0% recurrence (P = .04). Elevated body mass index was a significant risk factor predicting hernia recurrence (P = .003). Contamination (P = .04) and enterocutaneous fistula (P = .02) at the time of surgery were associated with increased major complications, whereas body mass index (P = .01) and diabetes mellitus (P = .04) were associated with increased minor complications. CONCLUSIONS: Large complex hernias can be reliably repaired using the components separation technique despite the presence of open wounds, the need for bowel surgery, and numerous comorbidities. The long-term strength of the hernia repair is not augmented by acellular cadaveric dermis but seems to be improved with soft polypropylene mesh.

Soft polypropylene mesh, but not cadaveric dermis, significantly improves outcomes in midline hernia repairs using the components separation technique.

BACKGROUND: The search continues for the "ideal" repair of the midline ventral hernia, and the components separation technique has a low, but still concerning, hernia recurrence rate. The authors hypothesize that adding prosthetic or bioprosthetic meshes to the midline closure during components separation would reduce recurrence rates with minimal added morbidity. METHODS: Over a 3-year period, patients had a components separation procedure where either acellular cadaveric dermis (n = 26) or soft polypropylene mesh (n = 28) was used as an intraperitoneal underlay for reinforcement of the midline repair, but not as a "bridging material." In 36 operations, the mesh or cadaveric dermis was placed at the time of the components separation, and in the remaining cases (n = 18), the underlay was used to treat a recurrence after components separation. RESULTS: Cadaveric dermis was associated with a 46 percent "true" recurrence rate that required reoperation (mean follow-up, 17.3 months), whereas soft polypropylene mesh had a significantly lower recurrence rate of 11 percent (p = 0.0057) during a follow-up period of 16 months. Because of a higher incidence of concomitant bowel surgery and contamination in the cadaveric dermis group, additional subset analysis of uncontaminated cases was performed, demonstrating a 61 percent recurrence rate for cadaveric dermis compared with 12 percent for soft polypropylene (p = 0.0017). No significant differences in major and minor complications were seen between groups. CONCLUSION: Soft polypropylene mesh, but not acellular dermis, demonstrates acceptably low complication and hernia recurrence rates when used as a reinforcement of the midline ventral hernia closure in conjunction with components separation.

Extracellular matrix protein-coated scaffolds promote the reversal of diabetes after extrahepatic islet transplantation.

BACKGROUND: The survival and function of transplanted pancreatic islets is limited, owing in part to disruption of islet-matrix attachments during the isolation procedure. Using polymer scaffolds as a platform for islet transplantation, we investigated the hypothesis that replacement of key extracellular matrix components known to surround islets in vivo would improve graft function at an extrahepatic implantation site. METHODS: Microporous polymer scaffolds fabricated from copolymers of lactide and glycolide were adsorbed with collagen IV, fibronectin, laminin-332 or serum proteins before seeding with 125 mouse islets. Islet-seeded scaffolds were then implanted onto the epididymal fat pad of syngeneic mice with streptozotocin-induced diabetes. Nonfasting glucose levels, weight gain, response to glucose challenges, and histology were used to assess graft function for 10 months after transplantation. RESULTS: Mice transplanted with islets seeded onto scaffolds adsorbed with collagen IV achieved euglycemia fastest and their response to glucose challenge was similar to normal mice. Fibronectin and laminin similarly promoted euglycemia, yet required more time than collagen IV and less time than serum. Histopathological assessment of retrieved grafts demonstrated that coating scaffolds with specific extracellular matrix proteins increased total islet area in the sections and vessel density within the transplanted islets, relative to controls. CONCLUSIONS: Extracellular matrix proteins adsorbed to microporous scaffolds can enhance the function of transplanted islets, with collagen IV maximizing graft function relative to the other proteins tested. These scaffolds enable the creation of well-defined microenvironments that promote graft efficacy at extrahepatic sites.

Inductive tissue engineering with protein and DNA-releasing scaffolds.

Cellular differentiation, organization, proliferation and apoptosis are determined by a combination of an intrinsic genetic program, matrix/substrate interactions, and extracellular cues received from the local microenvironment. These molecular cues come in the form of soluble (e.g. cytokines) and insoluble (e.g. ECM proteins) factors, as well as signals from surrounding cells that can promote specific cellular processes leading to tissue formation or regeneration. Recent developments in the field of tissue engineering have employed biomaterials to present these cues, providing powerful tools to investigate the cellular processes involved in tissue development, or to devise therapeutic strategies based on cell replacement or tissue regeneration. These inductive scaffolds utilize natural and/or synthetic biomaterials fabricated into three-dimensional structures. This review summarizes the use of scaffolds in the dual role of structural support for cell growth and vehicle for controlled release of tissue inductive factors, or DNA encoding for these factors. The confluence of molecular and cell biology, materials science and engineering provides the tools to create controllable microenvironments that mimic natural developmental processes and direct tissue formation for experimental and therapeutic applications.