Self-healing encapsulation and controlled release of vaccine antigens from PLGA microparticles delivered by microneedle patches.

There is an urgent need to reduce reliance on hypodermic injections for many vaccines to increase vaccination safety and coverage. Alternative approaches include controlled release formulations, which reduce dosing frequencies, and utilizing alternative delivery devices such as microneedle patches (MNPs). This work explores development of controlled release microparticles made of poly (lactic-co-glycolic acid) (PLGA) that stably encapsulate various antigens though aqueous active self-healing encapsulation (ASE). These microparticles are incorporated into rapid-dissolving MNPs for intradermal vaccination. PLGA microparticles containing Alhydrogel are loaded with antigens separate from microparticle fabrication using ASE. This avoids antigen expsoure to many stressors. The microparticles demonstrate bi-phasic release, with initial burst of soluble antigen, followed by delayed release of Alhydrogel-complexed antigen over approximately 2 months in vitro. For delivery, the microparticles are incorporated into MNPs designed with pedestals to extend functional microneedle length. These microneedles readily penetrate skin and rapidly dissolve to deposit microparticles intradermally. Microparticles remain in the tissue for extended residence, with MNP-induced micropores resealing readily. In animal models, these patches generate robust immune responses that are comparable to conventional administration techniques. This lays the framework for a versatile vaccine delivery system that could be self-applied with important logistical advantages over hypodermic injections.

Printing of small molecular medicines from the vapor phase.

There is growing need to develop efficient methods for early-stage drug discovery, continuous manufacturing of drug delivery vehicles, and ultra-precise dosing of high potency drugs. Here we demonstrate the use of solvent-free organic vapor jet printing to deposit nanostructured films of small molecular pharmaceutical ingredients, including caffeine, paracetamol, ibuprofen, tamoxifen, BAY 11-7082 and fluorescein, with accuracy on the scale of micrograms per square centimeter, onto glass, Tegaderm, Listerine tabs, and stainless steel microneedles. The printed films exhibit similar crystallographic order and chemistry as the original powders; controlled, order-of-magnitude enhancements of dissolution rate are observed relative to powder-form particles. In vitro treatment of breast and ovarian cancer cell cultures in aqueous media by tamoxifen and BAY 11-7082 films shows similar behavior to drugs pre-dissolved in dimethyl sulfoxide. The demonstrated precise printing of medicines as films, without the use of solvents, can accelerate drug screening and enable continuous manufacturing, while enhancing dosage accuracy.Traditional approaches used in the pharmaceutical industry are not precise or versatile enough for customized medicine formulation and manufacture. Here the authors produce a method to form coatings, with accurate dosages, as well as a means of closely controlling dissolution kinetics.

Self-healing of pores in PLGAs.

Self-healing of pores in Poly(lactic-co-glycolic acid)s (PLGA) plays an important role in the encapsulation and controlled release of drugs from PLGA microparticles. Despite the importance of this phenomenon, neither the mechanics of the deformation nor the material properties that control it have been fully studied. In this study, the material properties of PLGA have been characterized using mechanical tests, and a finite-element model has been developed to predict how pores heal. This model assumes that the healing process occurs by viscous flow resulting from the deviatoric stress field induced by the interaction between the surface curvature and the surface tension of the PLGA. The simulations, which incorporate measured material properties, show good agreement with experimental observations. However, annealing processes that occur over prolonged times increase the viscosity and slow the healing times of PLGA films at intermediate temperatures above the glass-transition temperature. These findings may be reasonably applied towards the prediction of healing processes in PLGA and in related biomaterials for important biomedical applications such as drug delivery.

The nature of peptide interactions with acid end-group PLGAs and facile aqueous-based microencapsulation of therapeutic peptides.

An important poorly understood phenomenon in controlled-release depots involves the strong interaction between common cationic peptides and low Mw free acid end-group poly(lactic-co-glycolic acids) (PLGAs) used to achieve continuous peptide release kinetics. The kinetics of peptide sorption to PLGA was examined by incubating peptide solutions of 0.2-4mM octreotide or leuprolide acetate salts in a 0.1M HEPES buffer, pH7.4, with polymer particles or films at 4-37°C for 24h. The extent of absorption/loading of peptides in PLGA particles/films was assayed by two-phase extraction and amino acid analysis. Confocal Raman microspectroscopy, stimulated Raman scattering (SRS) and laser scanning confocal imaging, and microtome sectioning techniques were used to examine peptide penetration into the polymer phase. The release of sorbed peptide from leuprolide-PLGA particles was evaluated both in vitro (PBST+0.02% sodium azide, 37°C) and in vivo (male Sprague-Dawley rats). We found that when the PLGA-COOH chains are sufficiently mobilized, therapeutic peptides not only bind at the surface, a common belief to date, but also can be internalized and distributed throughout the polymer phase at physiological temperature forming a salt with low-molecular weight PLGA-COOH. Importantly, absorption of leuprolide into low MW PLGA-COOH particles yielded ~17 wt.% leuprolide loading in the polymer (i.e., ~70% of PLGA-COOH acids occupied), and the absorbed peptide was released from the polymer for >2 weeks in a controlled fashion in vitro and as indicated by sustained testosterone suppression in male Sprague-Dawley rats. This new approach, which bypasses the traditional encapsulation method and associated production cost, opens up the potential for facile production of low-cost controlled-release injectable depots for leuprolide and related peptides.

Healing kinetics of microneedle-formed pores in PLGA films.

The spontaneous healing of aqueous pores in poly(D,L-lactic-co-glycolic acid) (PLGA) drug delivery systems has been identified to play a key role in terminating the burst release of large molecules, and to provide a means for novel aqueous-based microencapsulation. To examine healing of PLGA, pores were created of defined size and depth on the surface of thin PLGA films by stamping with blunt-tip microneedles. Pore dimensions on the micron-scale were relevant to surface pores of common PLGA microspheres and could be easily monitored by light microscopy. Most pores healed reproducibly at temperatures above the glass-transition temperature (T(g)) of the films, with healing times decreasing sharply with increasing temperature according to Williams-Landel-Ferry (WLF) behavior. It is suggested that healing is driven by high surface tension in the films and occurs through viscoelastic creep. Hydrated films healed at lower temperatures than dry films, consistent with a drop in Tg upon polymer hydration. Larger pores took longer to heal than smaller ones, while pores larger than 20 µm did not heal before significant polymer degradation occurred. Films of a less hydrophobic PLGA showed slower healing kinetics, attributed to a weaker surface tension driving force. Deeper pores showed signs of in-plane stress from spin-coating, and either ruptured or only partially healed when incubated wet and dry, respectively.

Microvascular cytomegalovirus endothelialitis of the lung: a possible cause of secondary pulmonary hypertension in a patient with AIDS.

Postmortem examination of the lungs of a patient with advanced AIDS who had developed pulmonary arterial hypertension late in the course of the illness demonstrated extensive cytomegalovirus (CMV) infection in endothelial cells of the lung microvasculature. Enlarged CMV-infected endothelial cells were present in virtually all histologic sections of the lungs, protruded into and compromised the lumens of the small vessels they lined, and were estimated by image cytometry of immunohistochemically stained sections to comprise 0.8% of the total lung tissue volume. Comparison with experimental microvascular embolization studies suggests that this amount of compromise of the microvascular luminal area of the lung is sufficient to elevate pulmonary arterial pressure significantly. Pathologic features in this case differed from both the plexogenic arteriopathy seen in previously reported cases of AIDS-associated primary pulmonary hypertension and the usual form of CMV pneumonitis in AIDS in which alveolar epithelial cells are the predominant site of infection.

The evaluation of compartmental syndromes using somatosensory evoked potentials in monkeys.

Somatosensory evoked potentials (SEPs) of the median, deep peroneal, and tibial nerves were recorded in experimentally induced compartmental syndromes in nine Macaca mulatta monkeys. A total of 17 trials were performed at the following mean pressure levels: 17, 30, 36, and 40 mm Hg. Evoked potentials were recorded using a four-channel evoked response system. Significant changes in SEP waveforms (greater than 3-millisecond increases in N1 or P1 latencies, or a 50% decrease in wave amplitudes in the presence of normal contralateral waveforms) were seen at pressures as low as 30 mm Hg as early as 45 minutes. Pressures of 35 and 40 mm Hg led to more pronounced abnormalities in SEP waveforms, with marked decreases in wave amplitudes and variability in the time to onset of these changes. Somatosensory evoked potentials are noninvasive, sensitive, and dynamic determinations of nerve function and may have clinical significance in the early detection of nerve dysfunction in compartment syndromes.

Onlay technique for occipitocervical fusion.

Twenty-eight occipitocervical fusions using the onlay technique were performed in 27 patients ranging in age from 13 to 77 years (average age, 47.6 years). The indications for fusion included neurologic involvement from atlantoaxial instability associated with superior migration of the odontoid and destructive changes at the occiput-C1-C2 articulation, causing pain unrelieved by conservative treatment. Preoperative diagnoses included rheumatoid arthritis, congenital anomalies, posttraumatic, failed C1-C2 fusions, ankylosing spondylitis, and tumor. A standard posterior exposure of occiput-C1-C2 was used, and iliac crest bone graft was placed over the area to be fused. Postoperative immobilization consisted of skull tong traction, minerva jacket, and halo apparatus. There were no neurologic complications, two superficial wound infections, and minor difficulties with halo loosening. There was one perioperative death. Primary fusion was obtained in 89% of patients at an average of 12.8 weeks. Occipitocervical fusion by the onlay technique is safe, requires no internal fixation, and has a high success rate when compared with other methods of obtaining fusion in the occipitocervical region.

Effect of C1-C2 rotation on canal size.

An anthropometric study of the atlas and axis was undertaken to determine spinal canal diameters, the degree of narrowing with rotation, and odontoid tilt. The mean sagittal diameter of the atlas was 30.1 mm and the mean coronal diameter was 28.8 mm. The mean sagittal diameter of the axis was 19.0 mm and the mean coronal diameter was 22.8 mm. Approximately 64 degrees of rotation occurred at the atlantoaxial complex before sufficient narrowing caused spinal cord compression in the average specimen. An average of 63 degrees of rotation was required to cause facet dislocation. Most specimens examined had a posterior odontoid tilt up to 45 degrees and a facet angle between 1 degree and 29 degrees. Bilateral facet dislocation occurred at 63 degrees. Spinal canal narrowing to 1 cm occurred at 64 degrees. This narrowing would probably not damage the cord, which averages 1 cm in diameter; however, if the rotary motion of C1 and C2 progressed, cord damage could occur. Facet angles and posterior odontoid tilts that are not within the normal range indicate pathological changes.

Angiographic demonstration of coronary sinus thrombosis: a potential consequence of trauma to the coronary sinus.

Abnormalities of the coronary sinus are rarely encountered. A case is presented demonstrating for the first time the angiographic appearance of coronary sinus thrombosis. This may have been the result of surgical trauma during mitral valve replacement or inadvertent cannulation of the coronary sinus during right heart catheterization or pacemaker insertion. Although the clinical significance of coronary sinus thrombosis is uncertain, obstruction of coronary sinus blood flow should not be deleterious because of multiple anastomoses between the coronary sinus system and the anterior cardiac veins. Difficulty in cannulating the coronary sinus for physiologic studies should suggest the possibility of coronary sinus thrombosis, especially in patients who have undergone mitral valve replacement. This may be confirmed by observing the venous phase of selective left coronary arteriography. Finally, coronary sinus thrombosis may be important as a source of pulmonary emboli. The prevalence of this serious complication requires further study.

Constant-pressure coronary artery perfusion during aortic valve operations.

We considered the theoretical differences between the normal relationships of coronary blood flow and perfusion pressure in the working heart and those obtained with continuous, steady-flow perfusion by a roller pump during aortic valve replacement. Steady pump perfusion should deliver less blood flow to the endocardium because: 1. For the same mean artery perfusion pressure, the average coronary blood flow is less with constant-flow pump perfusion. 2. With constant pump perfusion, pressure would be excessively high during systole, and during diastole it would be significantly lower than the mean perfusion pressure. Instantaneous pressure and flow were measured in the left coronary artery in 8 patients undergoing aortic valve replacement, employing either roller pump perfusion or a gravity flow system to provide a steady pressure source. Although we did not attempt to demonstrate improved endocardial flow, the mean left coronary flow was always greater with gravity perfusion (297 versus 153 ml/min), lending support to the theoretically proposed differences between the two perfusion methods.