Susceptibility of the domestic duck (Anas platyrhynchos) to experimental infection with Toxoplasma gondii oocysts.

A total of 28 domestic ducks were divided into seven groups of four ducks. Six groups were inoculated per os with 10(1), 10(2), 10(3), 10(4), 10(5) and 10(5.7) oocysts Toxoplasma gondii oocysts (K21 strain, which is avirulent for mice), and the remaining group was used as a control. Antibodies to T. gondii were detected in all ducks by the indirect fluorescence antibody test first on day 7 post-inoculation (p.i.). Antibody titres were found in the range of 1:20 to 1:640 depending on the infectious dose of the oocysts. From day 14 p.i. antibody titres increased to 1:80 to 1:20 480. Between days 14 and 28 p.i. (end of the experiment), antibody titres decreased in 14 ducks, remained the same in seven ducks, and continued to increase in three ducks. Bioassay in mice revealed T. gondii in the breast and leg muscles and the heart (100%, n=47), brain (91%, n=22), liver (54%, n=13) and stomach (46%, n=24). The infected ducks showed no clinical signs; however, the results of bioassay indicate that, compared with some gallinaceous birds, domestic ducks were relatively susceptible to T. gondii infection.     

Enzymatic digestion of adult human articular cartilage yields a small fraction of the total available cells

We investigated whether different protocols for the digestion of adult human articular cartilage influence the cell yield and capacity to attach and proliferate in culture dishes. Chondrocyte yields were expressed as a percentage of the total number of cells in the tissue, determined both histologically (using the dissector method) and biochemically (measuring the DNA content of tissue digests). Human cartilage specimens (n = 79) were digested using different protocols based on combinations of collagenase II (CGN), trypsin/EDTA, hyaluronidase, and tosyllysylchloromethane (TLCM). Yields of viable chondrocytes were the highest within a specific range of CGN concentrations and digestion times, but always < 22% of the total available cells. The combination of CGN with trypsin/EDTA or TLCM accelerated the digestion process but did not significantly increase cell yields. The percentage of viable cells that attached to culture dishes ranged 75-85% (< 19% of the total) and was reduced by TLCM. Doubling times of attached cells were comparable in all experimental groups. Our results indicate that chondrocyte yields and capacity to attach and proliferate are not highly sensitive to the specific isolation protocol used. However, typically used cartilage digestion protocols yield only a small fraction of the total available cells, possibly introducing an uncontrolled selection of certain chondrocyte subpopulations.     

Custom step wedge blocking using dynamic multileaf collimation for parametrial pelvic boost irradiation following brachytherapy for carcinoma of the cervix

Carcinoma of the cervix is typically treated with a combination of intracavitary brachytherapy and external beam radiation. The external beam dose is delivered with whole pelvis fields followed by split fields that protect midline organs at risk (bladder and rectum) while treating the parametria. Three approaches have been developed to shield midline structures: a simple rectangular block, a block customized to a single brachytherapy isodose line, and a step wedge filter constructed to conform to multiple brachytherapy isodose lines. A customized step wedge filter has the potential to produce a more homogeneous dose distribution but has not achieved widespread use due to labor intensive construction. We have developed a simple, novel method to produce a custom midline step wedge using dynamic multileaf collimation (dMLC). A comparison of film measurements in a phantom with the dose calculated by a commercial treatment planning system demonstrated agreement within 3% or 3 mm. The technique requires delivery times comparable to conventional techniques.     

Validation of target volume and position in respiratory gated CT planning and treatment

The capability of a commercial respiratory gating system based on video tracking of reflective markers to reduce motion-induced CT planning and treatment errors was evaluated. Spherical plastic shells (2.8-82 cm3), simulating the gross target volume (GTV), were placed in a water-filled body phantom that was moved sinusoidally along the longitudinal axis of the CT scanner and the accelerator for +/- 1 cm at 15-30 cycle/min. During gated CT imaging, the x-ray exposure was initiated by the gating system shortly before the end of expiration (so that the imaging time would be centered at the end of expiration); it was terminated by the scanner after completion of each slice. In nongated CT images, the target appeared distorted and often broken up. GTVs volume errors ranged 16%-110% in axial scans, and 7%-36% in spiral scans. In gated CT images, the spheres appeared 3 and 5 mm longer than their actual diameters (volume errors 2%-16%), at the respective respiration rates of 15 and 20 cycles/min. At 30 cycles/min the target appeared 1 cm longer, and volume error ranged 25%-53%. During treatment, gating kept the beam on for a duration equal to the CT acquisition time of 1 s/slice. The difference in positional errors between gated CT and portal films was 1 mm, regardless the size of residual motion errors. Because of the potential of suboptimal placement of the gating window between CT imaging and treatment, an extra 1.5-2.5 mm safety margin can be added regardless of the size of residual motion error. For respiratory rates > or = 30 cycles/min, the effectiveness of gating is limited by large residual motion in the 1 s CT acquisition time.     

Triglycidylamine crosslinking of porcine aortic valve cusps or bovine pericardium results in improved biocompatibility, biomechanics, and calcification resistance: chemical and biological mechanisms

We investigated a novel polyepoxide crosslinker that was hypothesized to confer both material stabilization and calcification resistance when used to prepare bioprosthetic heart valves. Triglycidylamine (TGA) was synthesized via reacting epichlorhydrin and NH(3). TGA was used to crosslink porcine aortic cusps, bovine pericardium, and type I collagen. Control materials were crosslinked with glutaraldehyde (Glut). TGA-pretreated materials had shrink temperatures comparable to Glut fixation. However, TGA crosslinking conferred significantly greater collagenase resistance than Glut pretreatment, and significantly improved biomechanical compliance. Sheep aortic valve interstitial cells grown on TGA-pretreated collagen did not calcify, whereas sheep aortic valve interstitial cells grown on control substrates calcified extensively. Rat subdermal implants (porcine aortic cusps/bovine pericardium) pretreated with TGA demonstrated significantly less calcification than Glut pretreated implants. Investigations of extracellular matrix proteins associated with calcification, matrix metalloproteinases (MMPs) 2 and 9, tenascin-C, and osteopontin, revealed that MMP-9 and tenascin-C demonstrated reduced expression both in vitro and in vivo with TGA crosslinking compared to controls, whereas osteopontin and MMP-2 expression were not affected. TGA pretreatment of heterograft biomaterials results in improved stability compared to Glut, confers biomechanical properties superior to Glut crosslinking, and demonstrates significant calcification resistance.     

Investigation of the morphology of cell clones, derived from the mammalian EBTr cell line and their susceptibility to vaccine avian poxvirus strains FK and Dessau

The ability for replication of vaccine avian pox viral strains FK and Dessau in cell clones, derived from the EBTr cell line, derived from embryonic bovine trachea, was studied. The derived seven cell clones showed different morphological characteristics and diverse sensitivity to both vaccine avian pox viral strains. Hence, the EBTr-derived cell clones could be used for cultivation, as well as for differentiation of vaccine avian pox viral strains. In addition, studies have been undertaken to elucidate the possible use of cultivated strains in these heterologous cell culture system's vaccine avian pox viral strains for biotechnology, as well as for solving problems, related to infection of people with avian viruses.     

Activated polyurethane modified with latent thiol groups

A novel type of modified polyurethane with pendant acetylthio groups (as a latent form of thiol groups) has been proposed for the use in surface modifications with various biomolecules. The polymer was prepared via a modified variant of low-temperature bromoalkylation of urethane hard segments followed by the reaction of pendant bromoalkyl groups with thiolacetic acid in mild conditions. The extent of modification with acetylthio groups can be made as high as 0.45 mmol/g. After deprotection of acetylthio groups and reaction of the resulting thiol groups with an excess of Ellman's reagent, 0.1 nmol/cm(2) of thiol-reactive 3-carboxy-4-nitrophenyldithio groups were detected on the surface of films cast from the modified polymer. A sensitive fluorescent probe--dansyl-L-cysteine was used for the quantification of thiol-reactive groups bound to the surface. The acetylthio-modified polyurethane is sufficiently stable to withstand conditions typical for the high-temperature processing (molding, extrusion) of polyurethanes.     

Dosimetric effect of respiration-gated beam on IMRT delivery

Intensity modulated radiation therapy (IMRT) with a dynamic multileaf collimator (DMLC) requires synchronization of DMLC leaf motion with dose delivery. A delay in DMLC communication is known to cause leaf lag and lead to dosimetric errors. The errors may be exacerbated by gated operation. The purpose of this study was to investigate the effect of leaf lag on the accuracy of doses delivered in gated IMRT. We first determined the effective leaf delay time by measuring the dose in a stationary phantom delivered by wedge-shaped fields. The wedge fields were generated by a DMLC at various dose rates. The so determined delay varied from 88.3 to 90.5 ms. The dosimetric effect of this delay on gated IMRT was studied by delivering wedge-shaped and clinical IMRT fields to moving and stationary phantoms at dose rates ranging from 100 to 600 MU/min, with and without gating. Respiratory motion was simulated by a linear sinusoidal motion of the phantom. An ionization chamber and films were employed for absolute dose and 2-D dose distribution measurements. Discrepancies between gated and nongated delivery to the stationary phantom were observed in both absolute dose and 2-D dose distribution measurements. These discrepancies increased monotonically with dose rate and frequency of beam interruptions, and could reach 3.7% of the total dose delivered to a 0.6 cm3 ion chamber. Isodose lines could be shifted by as much as 3 mm. The results are consistent with the explanation that beam hold-offs in gated delivery allowed the lagging leaves to catch up with the delivered monitor units each time that the beam was interrupted. Low dose rates, slow leaf speeds and low frequencies of beam interruptions reduce the effect of this delay-and-catch-up cycle. For gated IMRT it is therefore important to find a good balance between the conflicting requirements of rapid dose delivery and delivery accuracy.