Dosimetric effects of polyethylene glycol surface coatings on gold nanoparticle radiosensitization.

One of the main appeals of using gold nanoparticles (GNPs) as radiosensitizers is that their surface coatings can be altered to manipulate their pharmacokinetic properties. However, Monte Carlo studies of GNP dosimetry tend to neglect these coatings, potentially changing the dosimetric results. This study quantifies the dosimetric effects of including a polyethylene glycol (PEG) surface coating on GNPs over both nanoscopic and microscopic ranges. Two dosimetric scales were explored using PENELOPE Monte Carlo simulations. In microscopic simulations, 500-1000 GNPs, with and without coatings, were placed in cavities of side lengths 0.8-4 µm, and the reduction of dose deposited to surrounding medium within these volumes due to the coating was quantified. Including PEG surface coatings of up to 20 nm thickness resulted in reductions of up to 7.5%, 4.0%, and 2.0% for GNP diameters of 10, 20, and 50 nm, respectively. Nanoscopic simulations observed the dose falloff in the first 500 nm surrounding a single GNP both with and without surface coatings of various thicknesses. Over the first 500 nm surrounding a single GNP, the presence of a PEG surface coating reduced dose by 5-26%, 8-28%, 8-30%, and 8-34% for 2, 10, 20, and 50 nm diameter GNPs, respectively, for various energies and coating thicknesses. Reductions in dose enhancement due to the inclusion of a GNP surface coating are non-negligible and should be taken into consideration when investigating GNP dose enhancement. Further studies should be carried out to investigate the biological effects of these coatings.

Optimization of photon beam energies in gold nanoparticle enhanced arc radiation therapy using Monte Carlo methods.

As a recent area of development in radiation therapy, gold nanoparticle (GNP) enhanced radiation therapy has shown potential to increase tumour dose while maintaining acceptable levels of healthy tissue toxicity. In this study, the effect of varying photon beam energy in GNP enhanced arc radiation therapy (GEART) is quantified through the introduction of a dose scoring metric, and GEART is compared to a conventional radiotherapy treatment. The PENELOPE Monte Carlo code was used to model several simple phantoms consisting of a spherical tumour containing GNPs (concentration: 15 mg Au g-1 tumour, 0.8 mg Au g-1 normal tissue) in a cylinder of tissue. Several monoenergetic photon beams, with energies ranging from 20 keV to 6 MeV, as well as 100, 200, and 300 kVp spectral beams, were used to irradiate the tumour in a 360° arc treatment. A dose metric was then used to compare tumour and tissue doses from GEART treatments to a similar treatment from a 6 MV spectrum. This was also performed on a simulated brain tumour using patient computed tomography data. GEART treatments showed potential over the 6 MV treatment for many of the simulated geometries, delivering up to 88% higher mean dose to the tumour for a constant tissue dose, with the effect greatest near a source energy of 50 keV. This effect is also seen with the inclusion of bone in a brain treatment, with a 14% increase in mean tumour dose over 6 MV, while still maintaining acceptable levels of dose to the bone and brain.

A method for converting dose-to-medium to dose-to-tissue in Monte Carlo studies of gold nanoparticle-enhanced radiotherapy.

Gold nanoparticles (GNPs) have shown potential in recent years as a means of therapeutic dose enhancement in radiation therapy. However, a major challenge in moving towards clinical implementation is the exact characterisation of the dose enhancement they provide. Monte Carlo studies attempt to explore this property, but they often face computational limitations when examining macroscopic scenarios. In this study, a method of converting dose from macroscopic simulations, where the medium is defined as a mixture containing both gold and tissue components, to a mean dose-to-tissue on a microscopic scale was established. Monte Carlo simulations were run for both explicitly-modeled GNPs in tissue and a homogeneous mixture of tissue and gold. A dose ratio was obtained for the conversion of dose scored in a mixture medium to dose-to-tissue in each case. Dose ratios varied from 0.69 to 1.04 for photon sources and 0.97 to 1.03 for electron sources. The dose ratio is highly dependent on the source energy as well as GNP diameter and concentration, though this effect is less pronounced for electron sources. By appropriately weighting the monoenergetic dose ratios obtained, the dose ratio for any arbitrary spectrum can be determined. This allows complex scenarios to be modeled accurately without explicitly simulating each individual GNP.

Inhibition of E rosetting and of OKT 11 in patients with oral dysplasia and neoplasia.

A group of patients with oral dysplastic or early neoplastic lesions has been followed for 6 years to examine the change in ALS (antilymphocyte serum) induced 25% E rosette inhibition during the course of their disease. Control patients with other oral problems were examined similarly. The E rosette inhibition titer differences seen in our previous studies between control and diseased patients do not appear to persist with a longitudinal examination of these patients. While at early examination the patients exhibit higher inhibition titers than controls, later analysis shows that they seem to return to normal. To improve methodology, inhibition of OKT 11 uptake was substituted for E rosette inhibition and it is clear that both methods measure the same phenomenon. Thus, the former is more accurate and quantitative than the latter and will be used in future work. Continuation of these studies with the examination of more patients will be of value in assessing possible T lymphocyte changes in patients with oral lesions.

Monoclonal antibodies to human osteosarcoma-associated antigen(s).

Monoclonal antibodies (MoAbs) against human osteosarcoma cells were obtained by the production and cloning of hybrids resulting from the fusion of mouse myeloma cells P3 X 63Ag8.653 with spleen cells from partially purified, osteosarcoma-associated antigen (OSAA)-immunized BALB/c mice. OSAAs were isolated from the spent culture medium of a human osteosarcoma cell line (TE-85). Five hybrid clones were established and designated as OSA1, OSA2, OSA3, OSA4, and OSA5. OSA1 and OSA2 had similar activity. All 5 MoAbs reacted strongly with most osteosarcoma cell lines and with all osteosarcoma tissues tested but not with 10 tumor cell lines and 2 tumor tissues from other cancers. OSA3, OSA4, and OSA5 cross-reacted with a fibrosarcoma cell line, a colon cell line, and fibrosarcoma, respectively, as well as with a melanoma cell line. None of the MoAbs were reactive with activated normal human peripheral blood mononuclear cells (PBMC). Immunoprecipitation of membrane protein isolated from LM cells and TE-85 cells with the MoAbs OSA1 and OSA2 conjugated with Staphylococcus aureus yielded a molecule with molecular weight of approximately 92,000. No detectable membrane protein was precipitated when 125I-labeled membrane protein from pooled activated human PBMC and tumor cells of other histologic types were used in the immunoprecipitation.

Mononuclear cell phenotypes in patients with psoriasis.

Mononuclear blood cells from 19 patients with psoriasis were stained with a panel of monoclonal antibodies which detect total T lymphocytes, their helper/inducer and suppressor/cytotoxic subsets and Ia-positive lymphocytes and monocytes. Fluorescence-positive cells were enumerated by cell flow cytometry. In these patients, percentages of all mononuclear cell studies were within reference range with few, sporadic exceptions. Statistical analysis of data showed no significant differences between patient values and reference values from a panel of 100 normal subjects.

Defect of autologous mixed lymphocyte reaction and interleukin-2 in aged individuals.

The proliferative responsiveness of T cells of aged individuals is known to be depressed in both autologous mixed lymphocyte reaction (AMLR) and in PHA-stimulated cultures. In the present study we confirm previous results and also report decreased IL-2 and normal IFN-gamma production (PHA-induced) in aged subjects as compared to young normals. In addition, similar percentages of T lymphocytes expressing surface IL-2 receptors both in the peripheral blood and after different stimulations, i.e. AMLR and PHA, were detected in young and aged individuals. The addition of exogenous IL-2 induces a sharp increase of spontaneous and AMLR proliferation in young individuals, whereas the increase is only slight in aged subjects. The experiments reported herein suggest that in general the T cell proliferation in AMLR is not completely dependent on the presence of IL-2 in the cultures and that aged subjects are probably defective in the production of other factor(s) presumably involved in AMLR proliferation, since the addition of exogenous IL-2 does not produce T-cell proliferation comparable to normal young subjects. The possible meanings of these experimental evidences in AMLR and in the defective immune responses of aged subjects are discussed.

Interaction of human T lymphocytes with E. coli lipopolysaccharide (LPS).

We have examined the interaction of fluorescein isothiocyanate (FITC) labeled E. coli lipopolysaccharide (LPS) with human T lymphocytes by means of cell flow cytometry, in an effort to establish the type and the conditions of this interaction. Our results indicate that the interaction of LPS with human T lymphocytes is optimal at 37 degrees C. This interaction is rapid and reaches the maximum after 5 minutes and is followed by endocytosis. Our results show that sodium azide and trypsin treatment does not affect LPS- human T cell interaction, suggesting a nonspecific lipid-lipid interaction.

Flow cytometric analysis of bursal, thymic, and splenic cells from normal and cyclophosphamide-treated embryos.

Cell flow cytometry was used to assess the life cycle of embryonic lymphocytes. Cells were prepared for flow cytometry by fixation for 30 min in a final concentration of 70% ethyl alcohol, treated with ribonuclease (50 to 70 Kunitz/ml) and incubated for 30 min. in propidium iodide. Our data demonstrated that bursal lymphocytes from 20-day embryos (DE) exhibited significantly fewer cells in pre-DNA (deoxyribonucleic acid) and more cells in DNA synthesis (S) than thymic cells from 20 DE, and thymic lymphocytes from 16 DE expressed a more active S than bursal cells. Lymphocytes were separated from granulocytes by a Ficoll-hypaque double density gradient. The DNA cycle of bursal lymphocytes was more easily disrupted with cyclophosphamide (Cy) than the DNA cycle of thymic lymphocytes. However, unfractionated splenic cells from embryos treated with Cy revealed a greater percentage of cells in S and post-DNA synthesis and mitosis than splenic cells from control embryos. Because the splenic cells of Cy-treated embryos were predominantly immature granulocytes, it was concluded that Cy stimulated myelopoiesis in the spleen.

Cell flow cytometry of fixed and unfixed bursal and thymic cells.

Analysis in a cell flow cytometer (Ortho Spectrum III, Ortho Inst.) of single cell suspensions from the bursa and thymus of 20-day embryos revealed two distinct cell clusters. The two clusters were less apparent after fixation of the cells in paraformaldehyde and assumed a comet-like appearance at 30 min fixation in ethyl alcohol (EA). The G (postmitotic), S [deoxyribonucleic acid (DNA) synthesis], and G2/M (premitotic and mitotic) phases of the life cycle were visualized in two cell flow cytometers (Ortho Spectrum III and FACS IV, Bectin Dickinson) after treating the cells with EA, ribonuclease (RNase), and propidium iodide (PI, a fluorescent dye). Bursal cell suspensions exposed to the EA-RNase-PI protocol and stored for 3 weeks in phosphate-buffered saline showed minor changes in the G1 coefficient of variation, G1, and S percentages, but marked changes in these parameters occurred after 6 weeks of storage. Thymic cells treated in a similar fashion could not be maintained for 3 weeks. Bursal and thymic cells may remain in the EA for one day and possibly as long as 7 days prior to preparing them for DNA life cycle analysis. Paraformaldehyde was not a satisfactory cell fixative for assessing a cell's DNA life cycle.

The use of Ficoll-Hypaque double density gradients in the separation of avian granulocytes from other cell types for the purpose of cell flow cytometric analysis.

Avian peripheral blood and embryonic spleen cells were prepared for cell flow cytometry. The Ortho Spectrum III was the flow cytometer used in these experiments. The major objectives were to identify the location of lymphocytes and granulocytes in the cytogram displayed by flow cytometry, to develop a technique which would allow the collection of granulocytes relatively free of other cell types and to characterize the cell cycle within these cell populations. The cytogram of fresh avian cells developed in the Ortho Spectrum III revealed three characteristic cell clusters. Peripheral blood or embryonic spleen cells were separated on a Ficoll-Hypaque double density gradients into two distinct layers and a pellet. Light microscopic examination revealed the top layer of cells to be primarily lymphocytes while the middle layer of cells was granulocytes. Presentation of the cells from these layers to the Ortho Spectrum III revealed that granulocytes made up Cluster 3 while lymphocytes were included in the other clusters. The Ortho Spectrum III was employed to determine the presence of G1 (pre-DNA synthesis), S (DNA synthesis), and G2/M (post-DNA synthesis and mitosis) phases of cells in Clusters 1 and 2 and Cluster 3. While all the cells from peripheral blood were in G1, the embryonic spleen revealed cells in G1, S and G2/M in both Clusters 1 and 2 and Cluster 3.