Bioabsorbable bone fixation plates for X-ray imaging diagnosis by a radiopaque layer of barium sulfate and poly(lactic-co-glycolic acid).

Bone fixation systems made of biodegradable polymers are radiolucent, making post-operative diagnosis with X-ray imaging a challenge. In this study, to allow X-ray visibility, we separately prepared a radiopaque layer and attached it to a bioabsorbable bone plate approved for clinical use (Inion, Finland). We employed barium sulfate as a radiopaque material due to the high X-ray attenuation coefficient of barium (2.196 cm(2) /g). The radiopaque layer was composed of a fine powder of barium sulfate bound to a biodegradable material, poly(lactic-co-glycolic acid) (PLGA), to allow layer degradation similar to the original Inion bone plate. In this study, we varied the mass ratio of barium sulfate and PLGA in the layer between 3:1 w/w and 10:1 w/w to modulate the degree and longevity of X-ray visibility. All radiopaque plates herein were visible via X-ray, both in vitro and in vivo, for up to 40 days. For all layer types, the radio-opacity decreased with time due to the swelling and degradation of PLGA, and the change in the layer shape was more apparent for layers with a higher PLGA content. The radiopaque plates released, at most, 0.5 mg of barium sulfate every 2 days in a simulated in vitro environment, which did not appear to affect the cytotoxicity. The radiopaque plates also exhibited good biocompatibility, similar to that of the Inion plate. Therefore, we concluded that the barium sulfate-based, biodegradable plate prepared in this work has the potential to be used as a fixation device with both X-ray visibility and biocompatibility.

Functional properties of ultrasonically generated flaxseed oil-dairy emulsions.

This study reports on the functional properties of 7% flaxseed oil/milk emulsion obtained by sonication (OM) using 20 kHz ultrasound (US) at 176 W for 1-8 min in two different delivery formulae, viz., ready-to-drink (RTD) and lactic acid gel. The RTD emulsions showed no change in viscosity after sonication for up to 8 min followed by storage up to a minimum of 9 days at 4±2 °C. Similarly, the oxidative stability of the RTD emulsion was studied by measuring the conjugated diene hydroperoxides (CD). The CD was unaffected after 8 min of ultrasonic processing. The safety aspect of US processing was evaluated by measuring the formation of CD at different power levels. The functional properties of OM gels were evaluated by small and large scale deformation studies. The sonication process improved the gelation characteristics, viz., decreased gelation time, increased elastic nature, decreased syneresis and increased gel strength. The presence of finer sono-emulsified oil globules, stabilized by partially denatured whey proteins, contributed to the improvements in the gel structure in comparison to sonicated and unsonicated pasteurized homogenized skim milk (PHSM) gels. A sono-emulsification process of 5 min followed by gelation for about 11 min can produce gels of highest textural attibutes.

High-frequency (20 to 40 MHz) acoustic response of liquid-filled nanocapsules.

Liquid-core nanoparticles are promising candidates for targeted ultrasound-controlled therapy, but their acoustic detection remains challenging. High-frequency (20 to 40 MHz) tone burst sequences were implemented with a programmable ultrasound biomicroscope to characterize acoustic response from perfluorooctyl bromide-core nanoparticles with thick poly(lactide-coglycolide) (PLGA) shells. Radio-frequency signals were acquired from flowing solutions of nanoparticles with two different shell-thickness-to-particle-radius ratios, solid PLGA nanoparticles, and latex nanobeads (linear controls). Normalized fundamental (20 MHz) and second-harmonic power spectral density (PSD) increased with particle concentration and was highest for the thinnest shelled particles. The second- harmonic PSD was detectable from the nanoparticles for peak rarefactional pressures (PRP) from 0.97 to 2.01 MPa at 23 cycles and for tone bursts from 11 to 23 cycles at 2.01 MPa. Their second-harmonic¿to¿fundamental ratio increased as a function of PRP and number of cycles. Within the same PRP and cycle ranges, the second-harmonic¿to¿fundamental ratios from matched concentration solutions of latex nanobeads and solid PLGA nanoparticles was more weakly detectable but also increased with PRP and number of cycles. Nanoparticles were detectable under flow conditions in vitro using the contrast agent mode of a high-frequency commercial scanner. These results characterize linear acoustic response from the nanoparticles (20 to 40 MHz) and demonstrate potential for their highfrequency detection.

In vitro gene delivery with ultrasound-triggered polymer microbubbles.

In the work described here, gene delivery using polymer microbubbles triggered by ultrasound in vitro was investigated. The effects of pressure amplitude (0-2 MPa), center frequency (1-5 MHz), pulse length (3-12,000 µs), pulse repetition frequency (5-20,000 Hz) and exposure time (0-30 s) on transfection efficiency and cell viability were examined. The effects of radiation force, calcium ion concentration and timing of treatments were also examined. Cells were successfully transfected with pressure amplitudes as low as 250 kPa. Transfection was most efficient at lower frequencies and longer pulse lengths, with a transfection efficiency of 24.2 ± 2.0% achieved using a center frequency of 1 MHz, pressure amplitude of 1 MPa, pulse length of 12,000 µs and pulse repetition frequency of 5 Hz. Gene delivery was also affected by the extracellular calcium ion concentration and the timing of treatments.

In vitro effects on platelets irradiated with short-wave ultraviolet light without any additional photoactive reagent using the THERAFLEX UV-Platelets method.

BACKGROUND: A novel short-wave ultraviolet light (UVC) pathogen reduction technology (THERAFLEX UV-Platelets; MacoPharma, Mouvaux, France) without the need of any additional photoactive reagent has recently been evaluated for various bacteria and virus infectivity assays. The use of UVC alone has on the one hand been shown to reduce pathogens but may, on the other hand, have some impact on the platelet (PLT) quality. The purpose of this study was to determine the potential effects on PLT quality of pathogen inactivation treatment using the novel UVC method for PLT concentrates. STUDY DESIGN AND METHODS: Buffy-coat-derived PLTs suspended in SSP+ were irradiated with UVC light in plastic bags (MacoPharma) made of ethyl vinyl acetate, considered to be highly permeable to UVC light. The UVC-treated (test, n=8) as well as the untreated (reference, n=8) PLT units were stored in PLT storage bags composed of n-butyryl, tri n-hexyl citrate-plasticized polyvinyl chloride (MacoPharma) on a flat bed agitator for in vitro testing during 7 days of storage. RESULTS: No significant difference in PLT counts and lactate dehydrogenase between the groups was detected. During storage, glucose decreased more and lactate increased more in the test units. Statistically significant differences were found for glucose (P<0·01) and lactate (P<0·05) on day 7. ATP levels were higher (P<0·01 from day 5) in the reference units. With exception of day 7 (P<0·01 reference vs. test), hypotonic shock response reactivity was not different between groups. Extent of shape change was lower (P<0·01), and CD62P (P<0·05 day 5) was higher in the test units. CD42b and CD41/61 showed similar trends throughout storage, without any significant difference between the units. pH was maintained at >6·8 (day 7) and swirling remained at the highest level (score = 2) for all units throughout storage. CONCLUSION: Our results suggest that irradiation with UVC light has a slight impact on PLT in vitro quality and appears to be insignificant with regard to current in vitro standards.

Correlating quantitative MR measurements of standardized tumor lines with histological parameters and tumor control dose.

PURPOSE: To correlate non-invasively acquired radiobiologically relevant magnetic resonance (MR) parameters with functional histology and tumor control doses (TCD(50)). MATERIALS AND METHODS: The MR parameters relative perfusion, re-oxygenation and lactate (Lac) concentration from eight human xenograft squamous tumor lines were compared with the histologically acquired pimonidazole hypoxic fraction, the perfused vessel area and TCD(50). RESULTS: Good spatial correlation in the parameter maps could be observed between the pimonidazole staining and tumor regions, which can be reoxygenated when breathing carbogen. A strong positive correlation (R=0.74) was found between whole tumor pimonidazole hypoxic fraction and re-oxygenation, as one would expect. A good correlation was also observed between Lac concentration and re-oxygenation (R=0.71) and between TCD(50) and re-oxygenation (R=0.64), whereas Lac and TCD(50) showed a moderate relation (R=0.44). The in vivo measurement of relative perfusion could be validated to reflect the perfused vessel area (R=0.63). No correlation was detected between perfusion and re-oxygenation or TCD(50). CONCLUSIONS: Lac and re-oxygenation were shown to be pretreatment predictive markers independent from the pathophysiological changes induced during a fractionated course of radiotherapy. These parameters hold promise to be acquired non-invasively with results just a few minutes after measurement and to tailor radiotherapy to individual patterns of a tumor microenvironment.

Tissue healing during degradation of a long-lasting bioresorbable gamma-ray-sterilised poly(lactic acid) mesh in the rat: a 12-month study.

AIM: The purpose was to evaluate soft-tissue healing after poly(lactic acid) (PLA(94)) mesh implantation in a rat model. METHODS: Full-thickness abdominal wall defects were created in 108 Wistar rats, and reconstructed with 83 PLA(94) and 25 lightweight polypropylene (PPL) meshes. The meshes were previously gamma-ray sterilised with 25, 75 or 125 kGy to accelerate PLA(94) degradation. RESULTS: The inflammatory response in PLA(94) was significantly less pronounced and collagen organisation significantly better than in PPL. The higher the level of gamma-radiation, the higher the incidence of abdominal wall herniation (22.2, 31.3 and 52.6% with 25, 75 and 125 kGy, respectively). No herniation occurred in the PPL group. Tensile strength was dramatically reduced after gamma-ray-sterilised PLA(94) mesh implantation. CONCLUSION: The gamma-ray-sterilised PLA(94) mesh was poor in preventing abdominal wall hernia recurrences in a rat model.

Glycolytic metabolism and tumour response to fractionated irradiation.

BACKGROUND AND PURPOSE: To study whether pre-therapeutic lactate or pyruvate predict for tumour response to fractionated irradiation and to identify possible coherencies between intermediates of glycolysis and expression levels of selected proteins. MATERIALS AND METHODS: Concentrations of lactate, pyruvate, glucose and ATP were quantified via bioluminescence imaging in tumour xenografts derived from 10 human head and neck squamous cell carcinoma (HNSCC) lines. Tumours were irradiated with 30 fractions within 6 weeks. Expression levels of the selected proteins in tumours were measured at the mRNA and protein level. Tumour-infiltrating leucocytes were quantified after staining for CD45. RESULTS: Lactate but not pyruvate concentrations were significantly correlated with tumour response to fractionated irradiation. Lactate concentrations in vivo did not reflect lactate production rates in vitro. Metabolite concentrations did not correlate with GLUT1, PFK-L or LDH-A at the transcriptional or protein level. CD45-positive cell infiltration was low in the majority of tumours and did not correlate with lactate concentration. CONCLUSIONS: Our data support the hypothesis that the antioxidative capacity of lactate may contribute to radioresistance in malignant tumours. Non-invasive imaging of lactate to monitor radiation response and testing inhibitors of glycolysis to improve outcome after fractionated radiotherapy warrant further investigations.

Ultrasound triggered cell death in vitro with doxorubicin loaded poly lactic-acid contrast agents.

Traditional chemotherapy generally results in systemic toxicity, which also limits drug levels at the area of need. Two ultrasound contrast agents (UCA), with diameters between 1-2 microm in diameter and shell thicknesses of 100-200 nm, composed of poly lactic-acid (PLA), one loaded by surface adsorption and the other loaded by drug incorporation in the shell, were compared in vitro for potential use in cancer therapy. These poly lactic-acid (PLA) UCA platforms contain a gas core that in an ultrasound (US) field can cause the UCA to oscillate or rupture. Following a systemic injection of drug loaded UCA with external application of US focused at the area of interest, this platform could potentially increase drug toxicity at the area of need, while protecting healthy tissue through microencapsulation of the drug. In vitro toxicity in MDA-MB-231 breast cancer cells of the surface-adsorbed and shell-incorporated doxorubicin (Dox) loaded UCA were examined at 5 MHz insonation using a pulse repetition frequency of 100 Hz at varying pressure amplitudes. Both platforms resulted in equivalent cell death compared to free Dox and US when insonated at peak positive pressure amplitudes of 1.26 MPa and above. While no significant changes in cell death were seen for surface adsorbed Dox-UCA with or without insonation, cell death using the platform with Dox incorporated within the shell increased from 16.12% to 25.78% (p=0.0272), approaching double the potency of the platform when insonated at peak positive pressure amplitudes of 1.26 MPa and above. This mechanism is believed to be the result of UCA rupture at higher insonation pressure amplitudes, resulting in more exposed drug and shell surface area as well as increased cellular uptake of Dox containing polymer shell fragments. This study has shown that a polymer UCA with drug housed within the shell may be used for US-triggered cell death. US activation can be used to make a carrier significantly more potent once in the area of need.

The modification of PLA and PLGA using electron-beam radiation.

The degradable polymers polylactide (PLA) and polylactide-co-glycolide (PLGA) have found widespread use in modern medical practice. However, their slow degradation rates and tendency to lose strength before mass have caused problems. The aim of this study was to ascertain whether treatment with e-beam radiation could address these problems. Samples of PLA and PLGA were manufactured and placed in layered stacks, 8.1 mm deep, before exposure to 50 kGy of e-beam radiation from a 1.5 MeV accelerator. Gel permeation chromatography testing showed that the molecular weight of both materials was depth-dependent following irradiation, with samples nearest to the treated surface showing a reduced molecular weight. Samples deeper than 5.4 mm were unaffected. Computer modeling of the transmission of a 1.5 MeV e-beam in these materials corresponded well with these findings. An accelerated mass-loss study of the treated materials found that the samples nearest the irradiated surface initiated mass loss earlier, and at later stages showed an increased percentage mass loss. It was concluded that e-beam radiation could modify the degradation of bioabsorbable polymers to potentially improve their performance in medical devices, specifically for improved orthopedic fixation.

Synthesis of polylactic acid-polyglycolic acid blends using microwave radiation.

Degradation rates of a copolymeric PLGA can be controlled by varying the constituent amount in the copolymer. In the present study we have made an attempt to utilize microwave irradiation to blend PLLA and PGA in different concentrations. FTIR, NMR and DSC measurements clearly show the blending and cross-linking between the constituents.

Production of GMP-grade radioactive holmium loaded poly(L-lactic acid) microspheres for clinical application.

Radioactive holmium-166 loaded poly(L-lactic acid) microspheres are promising systems for the treatment of liver malignancies. The microspheres are loaded with holmium acetylacetonate (HoAcAc) and prepared by a solvent evaporation method. After preparation, the microspheres (Ho-PLLA-MS) are activated by neutron irradiation in a nuclear reactor. In this paper, the aspects of the production of a (relatively) large-scale GMP batch (4 g, suitable for treatment of 5-10 patients) of Ho-PLLA-MS are described. The critical steps of the Ho-PLLA-MS production process (sieving procedure, temperature control during evaporation and raw materials) were considered and the pharmaceutical quality of the microspheres was evaluated. The pharmaceutical characteristics (residual solvents, possible bacterial contaminations and endotoxins) of the produced Ho-PLLA-MS batches were in compliance with the requirements of the European Pharmacopoeia. Moreover, neutron irradiated Ho-PLLA-MS retained their morphological integrity and the holmium remained stably associated with the microspheres; it was observed that after 270h (10 times the half-life of Ho-166) only 0.3+/-0.1% of the loading was released from the microspheres in an aqueous solution. In conclusion, Ho-PLLA-MS which are produced as described in this paper, can be clinically applied, with respect to their pharmaceutical quality.

Encapsulation of osteoblast seeded microcarriers into injectable, photopolymerizable three-dimensional scaffolds based on d,l-lactide and epsilon-caprolactone.

UMR-106 seeded microcarriers were encapsulated into in situ, photopolymerizable three-dimensional scaffolds based on d,l-lactide and epsilon-caprolactone. UMR-106 and rat bone marrow cells proliferated and differentiated well on the microcarriers. The microcarriers were completely colonized after 14 days in culture. The viscous polymer paste allowed to mix the UMR-106 seeded microcarriers and gelatin (porosigen) properly. After the photopolymerization process, microcarriers and gelatin were evenly distributed throughout the scaffold. Gelatin was leached out within 7 h, and a porous scaffold was obtained. The microcarriers remained in the scaffold even after 7 days which demonstrates that they were well entrapped in the polymer. Increasing the amount of entrapped microcarriers (20-50%) leads to scaffolds with a reduced cross-linking. Hence, the microcarriers leached out. The encapsulated UMR-106 cells did not show pyknotic nuclei which demonstrates that the photopolymerization and handling the viscous polymer/gelatin/microcarrier paste is not detrimental for the cells.

Influence of electron-beam radiation on the hydrolytic degradation behaviour of poly(lactide-co-glycolide) (PLGA).

The purpose of this study is to examine the effect of electron-beam (e-beam) radiation on the hydrolytic degradation of poly(lactide-co-glycolide) (PLGA) films. PLGA films were irradiated and observed to undergo radiation-induced degradation through chain scission, as observed from a drop in its average molecular weight with radiation dose. Irradiated (5, 10 and 20 Mrad) and non-irradiated (0 Mrad) samples of PLGA were subsequently hydrolytically degraded in phosphate-buffered saline solution at 37.0 degrees C over a span of 12 weeks. It was observed that the natural logarithmic molecular weight (lnMn) of PLGA decreases linearly with hydrolytic degradation time. The rate of water uptake is higher for samples irradiated at higher radiation dose (e.g. 20 Mrad) and subsequently causing an earlier onset of mass loss. It is postulated that the increase in water uptake is due to the presence of more hydrophilic end groups, which results in the formation of microcavities because of an increase in osmotic pressure. A relationship between radiation dose and the rate of hydrolytic degradation of PLGA films, through its molecular weight was also established. This relationship allows a more accurate and precise control of the life span of PLGA through the use of e-beam radiation.

Degradation of poly(lactide-co-glycolide) (PLGA) and poly(L-lactide) (PLLA) by electron beam radiation.

This paper seeks to examine the effects of electron beam (e-beam) radiation on biodegradable polymers (PLGA and PLLA), and to understand their radiation-induced degradation mechanisms. PLGA (80:20) and PLLA polymer films were e-beam irradiated at doses from 2.5 to 50 Mrad and the degradation of these films were studied by measuring the changes in their molecular weights, FTIR spectra, thermal and morphological properties. The dominant effect of e-beam irradiation on both PLGA and PLLA is chain scission. Chain scission occurs first through scission of the polymer main chain, followed by hydrogen abstraction. Chain scission, though responsible for the reduction in the average molecular weight, Tc, Tg and Tm of both polymers, encourages crystallization in PLGA. PLLA also undergoes chain scission upon irradiation but to a lesser degree compared to PLGA. The higher crystallinity of PLLA is the key factor in its greater stability to e-beam radiation compared to PLGA. A linear relationship is also established between the decrease in molecular weight with respect to radiation dose.

Release of paclitaxel from polylactide-co-glycolide (PLGA) microparticles and discs under irradiation.

Paclitaxel is a promising anti-cancer drug as well as a radiosensitizer for chemotherapy and radiotherapy applications. Because of the poor solubility of paclitaxel in water and most pharmaceutical reagents, it is usually formulated with an adjuvant called Cremophor EL, which causes severe side effects. This work develops new dosage forms of paclitaxel for controlled release application, which do not require the adjuvant and, thus, can avoid its associated side effects. Paclitaxel was encapsulated into the PLGA matrix with various additives such as polyethylene glycol (PEG), isopropyl myristate (IPM) and d-alpha tocopheryl polyethylene glycol (Vitamin E TPGS). These additives were used to enhance the release rate of paclitaxel from the polymer matrix. Spray-drying and an hydraulic press were used to prepare paclitaxel-PLGA microspheres and discs. The microspheres and discs were given different irradiation doses to investigate their effects on the surface morphology (characterized by SEM, AFM and XPS) and in vitro release properties. There seems to be a small effect of the ionizing radiation on various formulations. Although the irradiation did not cause observable changes on the morphology of the polymer matrix, the release rate can be enhanced by a few per cent. It was found that PEG has the highest enhancement effect for release rate among all the additives investigated in this study.

Tissue welding with biodegradable polymer films-demonstration of acute strength reinforcement in vivo.

BACKGROUND AND OBJECTIVES: To demonstrate, in vivo, acute strength reinforcement benefits of polymer film patches. STUDY DESIGN/MATERIALS AND METHODS: Full thickness incisions created in a dorsal skin flap of Sprague-Dawley rats were closed by laser-tissue welding: albumin solder was topically applied to the incision on the dermal surface, and a poly(lactic-co-glycolic acid) (PLGA) polymer film placed on the solder as a patch (controls had no film). Breaking strength was tested acutely (15-20 minutes after sacrifice). RESULTS: The patched incisions were statistically stronger than the controls (ANOVA, P < 0.05). CONCLUSIONS: Polymer film patches may be a viable method to increase acute breaking strengths of welds using topically applied solder.

The effect of therapeutic irradiation on LactoSorb absorbable copolymer.

Bioabsorbable implants continue to gain popularity in providing temporary internal fixation due to their many advantages over metallic internal fixation. Coincident with the presence of internal fixation devices, it may be necessary to use radiotherapy to treat tumors. While metal implants can alter the distribution of the radiotherapy beam, bioabsorbable polymer implants are, essentially, tissue equivalent. This ionizing irradiation, in sufficiently high dose, can affect polymers through chain scission and cross-linking and accelerate the hydrolysis of absorbable polymers. However, little is known about the effects of therapeutic doses on such materials. This study exposed LactoSorb (Biomet, Inc., Warsaw, IN) absorbable copolymer to doses of x-ray irradiation in a clinically relevant manner, in vitro, with individual doses of 2 Gy administered five days per week for up to eight weeks, yielding a total cumulative dose of up to 80 Gy. Specimens were tested both mechanically and for inherent viscosity. Overall, the LactoSorb specimens withstood exposure to the irradiation exceedingly well, providing empirical evidence of the suitability of this material for temporary internal fixation when subsequent radiotherapy in the region is probable.

PLGA-based microparticles: elucidation of mechanisms and a new, simple mathematical model quantifying drug release.

The two major aims of this study were: (i) to elucidate the underlying release mechanisms from drug-loaded, erodible microparticles based on poly(lactic-co-glycolic acid) (PLGA) showing biphasic drug release behavior: an initial 'burst' effect, followed by a zero order release phase; and (ii) to develop a new, simple mathematical model that allows the quantitative description of the observed in vitro drug release patterns from this type of delivery system. PLGA-based microparticles offer various advantages, such as the possibility to control the resulting drug release rate accurately over prolonged periods of time, easiness of administration (e.g., by stereotaxic injection), good biocompatibility and complete erosion (avoiding the removal of empty remnants). Consequently, the practical importance of these advanced drug delivery systems is remarkably increasing. However, only little knowledge is yet available concerning the processes controlling the release rate of the drug out of these devices. Various chemical and physical phenomena are involved, rendering the identification of the crucial mechanisms and the mathematical description of the resulting drug release kinetics difficult. In the present study, different physicochemical characterization methods (e.g., DSC, SEM, SEC, particle size analysis) were used to monitor the changes occurring within anticancer drug-loaded PLGA microparticles upon exposure to phosphate buffer pH 7.4. Based on these experimental findings, the most important underlying drug release rate controlling mechanisms were identified and a new mathematical model was developed that allows the quantitative description of the resulting release patterns.

Resorbable polymer fibers for ligament augmentation.

Resorbable augmentation devices for cruciate ligament surgery have been developed to temporarily protect healing tendon grafts or sutured ligaments against high tensile loads during the postoperative healing period. Materials available at present [e.g., polydioxanone (PDS)] show a half-life tensile strength of only 4-6 weeks, whereas the process of revitalization and recovering of the transplanted tendon graft can take up to 12 months. Therefore, a device that provides gradually decreasing mechanical properties with a half-time strength of at least 6 months would be desirable. In order to obtain a suitable material, we investigated the degradation kinetics of a variety of different resorbable fibers made of poly(L-lactide) and poly(L-lactide-co-glycolide). The fiber materials differed in processing and treatment parameters like thermal posttreatment, irradiation, and fiber diameter. The fibers were degraded in vitro and were tested for mechanical properties and molecular weight at various time points up to 72 weeks. The half-time strength of the materials ranged between 5 and 64 weeks, depending on their treatment parameters. In contrast, the stiffness did not decrease adequately. However, an augmentation stiffness that does not change much versus time could not provide a gradual increase in graft load, which is important to stimulate the orientation of the collagenous tissue. Therefore, design of an augmentation construct braided out of more than one quickly degrading fiber materials is suggested. After the breakdown of the faster-degrading fiber components the stiffness would automatically decrease by the diminution of the load-carrying fiber volume.