Geant4 Monte Carlo simulation study of the secondary radiation fields at the laser-driven ion source LION.

At the Center for Advanced Laser Applications (CALA), Garching, Germany, the LION (Laser-driven ION Acceleration) experiment is being commissioned, aiming at the production of laser-driven bunches of protons and light ions with multi-MeV energies and repetition frequency up to 1 Hz. A Geant4 Monte Carlo-based study of the secondary neutron and photon fields expected during LION's different commissioning phases is presented. Goal of this study is the characterization of the secondary radiation environment present inside and outside the LION cave. Three different primary proton spectra, taken from experimental results reported in the literature and representative of three different future stages of the LION's commissioning path are used. Together with protons, also electrons are emitted through laser-target interaction and are also responsible for the production of secondary radiation. For the electron component of the three source terms, a simplified exponential model is used. Moreover, in order to reduce the simulation complexity, a two-components simplified geometrical model of proton and electron sources is proposed. It has been found that the radiation environment inside the experimental cave is either dominated by photons or neutrons depending on the position in the room and the source term used. The higher the intensity of the source, the higher the neutron contribution to the total dose for all scored positions. Maximum neutron and photon ambient dose equivalent values normalized to 109 simulated incident primaries were calculated at the exit of the vacuum chamber, where values of about 85 nSv (109 primaries)-1 and 1.0 µSv (109 primaries)-1 were found.

Out-of-field doses for scanning proton radiotherapy of shallowly located paediatric tumours-a comparison of range shifter and 3D printed compensator.

The lowest possible energy of proton scanning beam in cyclotron proton therapy facilities is typically between 60 and 100 MeV. Treatment of superficial lesions requires a pre-absorber to deliver doses to shallower volumes. In most of the cases a range shifter (RS) is used, but as an alternative solution, a patient-specific 3D printed proton beam compensator (BC) can be applied. A BC enables further reduction of the air gap and consequently reduction of beam scattering. Such pre-absorbers are additional sources of secondary radiation. The aim of this work was the comparison of RS and BC with respect to out-of-field doses for a simulated treatment of superficial paediatric brain tumours. EURADOS WG9 performed comparative measurements of scattered radiation in the Proteus C-235 IBA facility (Cyclotron Centre Bronowice at the Institute of Nuclear Physics, CCB IFJ PAN, Kraków, Poland) using two anthropomorphic phantoms-5 and 10 yr old-for a superficial target in the brain. Both active detectors located inside the therapy room, and passive detectors placed inside the phantoms were used. Measurements were supplemented by Monte Carlo simulation of the radiation transport. For the applied 3D printed pre-absorbers, out-of-field doses from both secondary photons and neutrons were lower than for RS. Measurements with active environmental dosimeters at five positions inside the therapy room indicated that the RS/BC ratio of the out-of-field dose was also higher than one, with a maximum of 1.7. Photon dose inside phantoms leads to higher out-of-field doses for RS than BC to almost all organs with the highest RS/BC ratio 12.5 and 13.2 for breasts for 5 and 10 yr old phantoms, respectively. For organs closest to the isocentre such as the thyroid, neutron doses were lower for BC than RS due to neutrons moderation in the target volume, but for more distant organs like bladder-conversely-lower doses for RS than BC were observed. The use of 3D printed BC as the pre-absorber placed in the near vicinity of patient in the treatment of superficial tumours does not result in the increase of secondary radiation compared to the treatment with RS, placed far from the patient.

Stability of DNA in Purkinje cell nuclei of the mouse. An autoradiographic study.

Neurons of the mouse were labeled with [(3)H]thymidine during their prenatal period of proliferation. The (3)H activity of the Purkinje cell nuclei was then studied autoradiographically 8, 25, 55, and 90 days after birth. The measured grain number per nucleus decreased by about 14% between the 8th and 25th postnatal days and then remained constant up to 90 days. There was no significant decrease of the (3)H activity of the Purkinje cell nuclei after correction of the measured grain number per nucleus for increasing nuclear volume of the growing Purkinje cells and for the influence of [(3)H]beta self-absorption in the material of the sections. Injection of a high dose of [(3)H]thymidine into young adult mice did not result in (3)H labeling of either Purkinje or other neurons in other brain regions. The results agree with the concept of metabolic stability of nuclear DNA. "Metabolic" DNA could not be observed in these experiments.

Lesion evolution after gamma knife irradiation observed by magnetic resonance imaging.

PURPOSE: Our study is focused on the magnetic resonance imaging (MRI) observation of lesion development and hippocampus related functional impairments in rats after irradiation with a Leksell Gamma knife (LGK). MATERIALS AND METHODS: We exposed 32 three-month-old Long-Evans rats to various radiation doses (25 Gy, 50 Gy or 75 Gy). The rats were scanned by a 4.7 T magnetic resonance (MR) spectrometer at several timepoints (1 - 18 months) after irradiation. The lesion size was evaluated by manual segmentation; the animals were behaviorally tested in a Morris water maze and examined histologically. RESULTS: We found that a dose of 25 Gy induced no edema, necrosis or behavioral change. The response of the rats to higher doses was not uniform; the first occurrence of lesions in the rat brains irradiated with 50 and 75 Gy was detected six months post-irradiation. Functional impairment correlated well with the lesion size and histology. CONCLUSIONS: Rat brains showed the development of expanding delayed lesions after 50 or 75 Gy doses from the LGK during the first year after irradiation.

Altitude-dependent dose conversion coefficients in EPCARD.

Conversion coefficients that depend on altitude, cutoff rigidity and solar activity were developed and introduced in the European Program Package for the Calculation of Aviation Route Doses (EPCARD). A set of specially chosen long-distance flights were used to compare the new particle effective doses and ambient dose equivalents with those calculated using the previous averaged constant conversion coefficients. The data show very good agreement to each other. The dose differences for the chosen flights are <11%, for typical civil flight levels.

Long-term measurements of cosmic ray neutrons by means of a Bonner spectrometer at mountain altitudes - first results.

A Bonner multi-sphere spectrometer has been installed in 2005 at the Environmental Research Station 'Schneefernerhaus' (2660 m above sea level) on the Zugspitze mountain, Germany, to measure the energy spectrum of cosmic-ray neutrons at high altitudes continuously. The system can be used to investigate small temporal variations in the cosmic radiation intensity. For example, measurements were done during periods of 2 Forbush decreases of the cosmic radiation intensity in July and September 2005, respectively. The results were compared with those obtained by using neutron monitors, and neutron fluence spectra measured during these events are presented and discussed.

The influence of interleukin-1beta on gamma-glutamyl transpepidase activity in rat hippocampus.

Brain infections as well as peripheral challenges to the immune system lead to an increased production of interleukin-1beta (IL-1beta), a cytokine involved in leukocyte-mediated breakdown of the blood-brain barrier. The effects of IL-1beta have been reported to depend on whether the route of administration is systemic or intracerebral. Using 50-day-old male rats, we compared the effects of IL-1beta on brain gamma-glutamyl transpeptidase (GGT; an enzymatic marker of brain capillary endothelium) at 2, 24 and 96 h after either an intravenous (i.v.) injection of 5 microg IL-1beta or an intracerebroventricular (i.c.v. - lateral ventricle) infusion of 50 ng IL-1beta. When the i.v. route was used, the GGT activity underwent small but significant changes; decreasing in the hippocampus 2 h after the i.v. injection, increasing 24 h later and returning to control levels at 96 h. No significant changes in the hippocampal GGT activity were observed at 2 and 24 h following the i.c.v. infusion. The GGT activity in the hypothalamus remained unchanged regardless of the route of IL-1beta administrations. Similar changes in GGT activity were revealed histochemically. The labeling was found mainly in the capillary bed, the changes being most evident in the hippocampal stratum radiatum and stratum lacunosum-moleculare. A transient increase in GGT activity at 24 h, together with a less sharp delineation of GGT-stained vessels, may reflect IL-1beta induced increased turnover of glutathione and/or oxidative stress, that may in turn, be related to altered permeability of the blood-brain barrier in some neurological and mental disorders, including schizophrenia.

Up-regulation of gamma-glutamyl transpeptidase (GGT) activity in growth perturbed C6 astrocytes.

Activity of gamma-glutamyl transpeptidase (GGT) was studied in astrocyte-like C6 glial cells modulated in growth and maturation by different concentration of serum and dibutyryl cyclic AMP (Db-cAMP) supplement in culture medium. After reduction of serum concentration from 10% to 0.1%, the number of GGT positive cells determined histochemically increased 3.1 times and the GGT activity/mg protein in whole cell lysates was 5.1 times higher. In cultures with 0.1% serum + Db-cAMP, the histochemically and biochemically assayed GGT activity exceeded 5.1 and 7.9 times the values measured in control 10% serum cultures, respectively. The up-regulation of GGT was accompanied by an inhibition of proliferation, enhanced differentiation and hypertrophy of cells. In addition, the process of metabolic perturbation and/or cellular stress was revealed in these cultures by the (i) growth-support release followed by shrinkage and death of a small number of cells and (ii) higher oxidation of 2'7'dichlorofluorescein diacetate to its fluorescent form in the adherent/viable cells. The observed up-regulation of GGT is considered to primarily reflect increased metabolism of glutathione and/or the maintenance of the redox potential in cells stressed by sub-optimal concentration of serum and Db-cAMP supplement. The concomitant cellular hypertrophy and differentiation and their relationship to increased activity of GGT await further investigation. The study suggests that up-regulation of GGT can contribute to adaptation of astrocytic cells to metabolic and/or oxidative perturbances occurring under various pathological conditions, including radiation- and drug-induced toxicity.

Improving potato drought tolerance through the induction of long-term water stress memory.

Knowledge of drought tolerance in potato is limited and very little is known about stress memory in this crop. In the present study, long-term stress memory was tested on tuber yield and drought tolerance related traits in three potato varieties (Unica, Désirée and Sarnav) with contrasted yields under water restriction. Seed tubers produced by plants grown under non-restricted (non-primed tubers) and restricted (primed tubers) water conditions were sown and exposed to similar watering treatments. Tuber yield and leaf greenness of plants from primed and non-primed seeds as well as tuber carbon isotope discrimination (Δ(13)C) and antioxidant activity (AA) responses to watering treatments were compared. Higher tuber yield, both under non-restricted and restricted water regimes, was produced by primed Sarnav plants. The decrease of tuber yield and Δ(13)C with water restriction was lower in primed Unica plants. Long-term stress memory consequently appears to be highly genotype-dependent in potato. Its expression in plants originated from primed tubers and facing water restriction seems to be positively associated to the degree of inherent capability of the cultivar to yield under water restriction. However, other effects of priming appear to be genotype-independent as priming enhanced the tuber AA in response to water restriction in the three varieties.

Measurement of stray radiation within a scanning proton therapy facility: EURADOS WG9 intercomparison exercise of active dosimetry systems.

PURPOSE: To characterize stray radiation around the target volume in scanning proton therapy and study the performance of active neutron monitors. METHODS: Working Group 9 of the European Radiation Dosimetry Group (EURADOS WG9-Radiation protection in medicine) carried out a large measurement campaign at the Trento Centro di Protonterapia (Trento, Italy) in order to determine the neutron spectra near the patient using two extended-range Bonner sphere spectrometry (BSS) systems. In addition, the work focused on acknowledging the performance of different commercial active dosimetry systems when measuring neutron ambient dose equivalents, H(∗)(10), at several positions inside (8 positions) and outside (3 positions) the treatment room. Detectors included three TEPCs--tissue equivalent proportional counters (Hawk type from Far West Technology, Inc.) and six rem-counters (WENDI-II, LB 6411, RadEye™ NL, a regular and an extended-range NM2B). Meanwhile, the photon component of stray radiation was deduced from the low-lineal energy transfer part of TEPC spectra or measured using a Thermo Scientific™ FH-40G survey meter. Experiments involved a water tank phantom (60 × 30 × 30 cm(3)) representing the patient that was uniformly irradiated using a 3 mm spot diameter proton pencil beam with 10 cm modulation width, 19.95 cm distal beam range, and 10 × 10 cm(2) field size. RESULTS: Neutron spectrometry around the target volume showed two main components at the thermal and fast energy ranges. The study also revealed the large dependence of the energy distribution of neutrons, and consequently of out-of-field doses, on the primary beam direction (directional emission of intranuclear cascade neutrons) and energy (spectral composition of secondary neutrons). In addition, neutron mapping within the facility was conducted and showed the highest H(∗)(10) value of ∼ 51 µSv Gy(-1); this was measured at 1.15 m along the beam axis. H(∗)(10) values significantly decreased with distance and angular position with respect to beam axis falling below 2 nSv Gy(-1) at the entrance of the maze, at the door outside the room and below detection limit in the gantry control room, and at an adjacent room (<0.1 nSv Gy(-1)). Finally, the agreement on H(∗)(10) values between all detectors showed a direct dependence on neutron spectra at the measurement position. While conventional rem-counters (LB 6411, RadEye™ NL, NM2-458) underestimated the H(∗)(10) by up to a factor of 4, Hawk TEPCs and the WENDI-II range-extended detector were found to have good performance (within 20%) even at the highest neutron fluence and energy range. Meanwhile, secondary photon dose equivalents were found to be up to five times lower than neutrons; remaining nonetheless of concern to the patient. CONCLUSIONS: Extended-range BSS, TEPCs, and the WENDI-II enable accurate measurements of stray neutrons while other rem-counters are not appropriate considering the high-energy range of neutrons involved in proton therapy.

Ultrastructure of nuclei of cisplatin-treated C6 glioma cells undergoing apoptosis.

C6 glioma cells, treated with a cytostatic dose of cisplatin (1.66 x 10(-5) M) ceased dividing by 24 h and, most of them had undergone apoptosis by 72-96 h. The reactive cells were classified into 5 types (T-I to V), according to the ultrastructure of nuclei. At 4 h, 20.4% of cells (T-I) showed minute condensation and margination of chromatin. The nuclear envelope (NE) formed slim and deep invaginations consisting of the inner or both membranes. The later kind of NE invaginations often extended to the enlarged nucleoli and contained nucleolus-like material at its cytoplasmic side. Some nuclear pores were covered with a dome-shaped "cap" formed by fine filamentous material. The number of T-I cells increased to 53.3% by 72 h. In T-II cells, which appeared at 24 h, the chromatin was condensed into dense irregular masses separated from the NE by a lucent space with filamentous structures preventing complete margination of chromatin. Nucleoli of T-II cells were small and showed partial segregation of their components. The "capped" pores were absent in these apparently more damaged cells. From 24 h, cells with large and lobulated nuclei (T-III) started to increase in number and peaked at 72 h (6.6%). Except for some small lobules, the chromatin of T-III cells was moderately aggregated and the NE was well preserved. Typical apoptotic cells with highly condensed and marginated chromatin (T-IV) peaked at 48-72 h (2.4-4.8%). They appeared in 2 varieties, including cells with wrinkled nuclei with less condensed and incompletely marginated chromatin or more lobulated forms with highly condensed marginated chromatin suggesting their origin from T-II or T-III cells. T-IV cells, as well as their fragments, underwent phagocytosis and secondary necrosis (T-V cells, 48.6% at 96 h). Two alternative routes of nuclear changes leading to cisplatin-triggered apoptosis, as represented by the sequence T-I --> T-III --> T-IV/V or T-I --> T-II --> T-IV/V, may explain the initially less or more damaged cells. These alternatives, together with progressive recruitment of reactive cells, suggest intrapopulation differences in the sensitivity of cells or in the cell cycle perturbations induced by cisplatin. Except for the T-IV and T-V cells, observed alterations of cytoplasmic organelles, including mitochondria, were fewer than reported in previous studies on cisplatin.

Changes in cell cycle and chromatin distribution in C6 glioma cells treated by dibutyryl cyclic AMP.

C6 glioma cells in culture were treated with 1 mM dibutyryl cyclic AMP (Db-cAMP) for 5, 8, 24 and 72 h. The cells were labelled with [3H]-thymidine before either the end, or the beginning, of the Db-cAMP treatment. The cell cycle passage was monitored by the simultaneous determination of DNA content and DNA synthesis in propidium iodide stained autoradiograms. The data revealed an early (t less than or equal to 3-8 h) and moderate inhibitory effect of Db-cAMP on all phases of the cell cycle except mitosis; some cells (2%) were completely blocked in the S phase. Later (8 less than t less than 24-72 h), the cycling of a substantial part of the population became inhibited in G1 phase. Microdensitometric texture analysis of Feulgen-stained nuclei, performed 24 h after administration of Db-cAMP, showed a higher inhomogeneity of the DNA distribution in cell nuclei, caused by the condensation of a part of the chromatin. This may reflect either changes in genome expression taking part in the process of cAMP induced differentiation or transit of some cells into quiescent G0 or S0 phases.

An estimate of the number of cells arising by division in mouse cerebral hemispheres from age one to 12 months: an autoradiographic study of DNA synthesis.

Male CBA strain mice aged 1, 2, 4, 6, 8 and 12 months were injected with 5 muCi of 6-H3-thymidine per gm body weight and killed two hours after the injection. Incorporation of the isotope, as a measure of DNA synthesis and cell division, was studied in the cerebral cortex and corpus callosum by light-microscope autoradiography. DNA synthesis was found in a small number of nuclei of non-neuronal cells, both of ecto- and mesodermal origin. The percentage of labeled cells (Labeling Index, or LI) decreased in both regions exponentially with age. On the base of measured LI values, the total number of cells arising from age one to 12 months was calculated. As the reference value the number of cells in one month-old animals was taken. The calculated cell increment in the ectodermal population in the cortex amounted to 19.7-20.6% (average 20.2%) and to 196.0-425.0% (average 247.0%) in the corpus callosum (pairs of figures correspond to values calcualted from LI values measured in sagittal and frontal sections). When all labeled cells (LC) were taken into consideration (of both ecto- and mesodermal origin) the corresponding cell increment ranged from 25.0-30.5% (average 26.8%) in the cortex and from 205.0-454.8% (average 263.0%) in the corpus callosum. The number of all newly arising cells in the whole hemisphere (including both cerebral cortex and corpus calosum) calculated from mean LI values of all LC in sagittal and frontal sections amounted to 52.0%. It is suggested that the observed DNA synthesis and cell division, particularly in the corpus callosum and mesodermal cell groups of both regions studied, correspond mainly to renewal of cell populations. In the cerebral cortex accumulation of some newly arising cells in situ cannot be excluded.

Neurogenesis in the visual system of the rat. An autoradiographic investigation.

Rats of the BD III strain were injected with a single dose of 3H-thymidine on either the twelfth, fourteenth, sixteenth, eighteenth or twentieth day of gestation (ED 12. . . . .ED 20) or on the postnatal day one, three, or seven. Animals were killed at age 22 to 24 days. DNA synthesis, as an indicator of cell division, was studied in matrix precursors of nerve and glial cells in the visula centers, including the lateral geniculate body (LGB), the superior colliculus (SC) and the visual cortex (VC). It was found that proliferation of matrix precursors of nerve cells destined for all the regions studied was in progress on ED 12. In the subcortical regions (LGB, SC) this process was substantially more advanced than in the VC. The first neuroblasts appeared in the SC (ED 12) and only later (ED 14) in the LGB and VC. In comparison with the LGB, VC neuroblasts were quite rare on ED 14 and were present only in layer VI. They appeared more frequently in this region only after injection of isotope on ED 16. Matrix cell proliferation and nerve cell formation ceased in the LGB between ED 16 and ED 18. The number of labeled cells arising after injection of the isotope on ED 16 indicates that neurogenesis ceased somewhat earlier in the dorsal nucleus of the LGB than in the ventral. In the SC the last neurons arose between ED 18 and ED 20, and in the VC, with the possible exception of a few granular neurons (which may continue division into the first few days postnatally), proliferation continued until the end of gestation. The origin of neuroblasts initially followed a caudo-rostral gradient. Later, the times of neurogenesis in the regions studied overlapped significantly. This is clear, for example. on ED 16, when neurogenesis in the mesencephalic SC continued for about two days longer than in the more postral LGB, and coincided with that in the VC, especially in the deep layers. The end of neurogenesis in the LGB, especially in the ventral nucleus, coincided with the time of neurogenesis in the deep cortical layers. In the VC, and partly also in the SC, an inside-out pattern of proliferation and neuron formation was confirmed. The times of proliferation of precursor cells, with the exception of the very end of neurogenesis, substantially overlapped within both these regions. The degree of this overlapping, described in terms of Labeling Index values, decreased towards the end of the neurogenetic period. Division of neuroglial cell precursors, started as early as on ED 14 in/for subcortical centers (LGB, SC), but not until ED 18 in/for the VC. A few labeled endothelial-like cells were observed in all regions studied after isotope injection on ED 12.

Regional differences in protein and glycoprotein synthesis and their processing in the mouse brain as revealed by the incorporation of [3H]proline, N-6-[3H]acetyl-D-glucosamine and [3H]fucose.

The incorporation rate of [3H]fucose, N-6-[3H]acetyl-D-glucosamine and [3H]proline has been compared in five regions of the mouse brain on postnatal day 6. The olfactory bulbs and the cerebellum showed a prevalence of incorporation of [3H]fucose over other brain regions. Less expressed, but still well evident regional differences were observed in [3H]proline incorporation while the incorporation of N-6-[3H]acetyl-D-glucosamine was almost equal in all brain regions. The regional differences were also apparent after considering an actual pool of free isotopes in the individual regions. Gel electrophoresis of [3H]fucose-labelled membrane fraction showed that the higher incorporation of [3H]fucose in the olfactory bulbs is partly due to higher synthesis of low molecular weight glycoproteins, especially in the molecular range of 30,000. The data showed that the protein synthesis and fucosylation, and/or a fast transport of the corresponding molecules, varies more within the brain than the incorporation of N-6-[3H]acetyl-D-glucosamine and possibly also than the "core" part of glycan molecule synthesis.

Regional and cellular differences in fucosylation of glycomacromolecules in the mouse brain. A biochemical and autoradiographic study of early postnatal and adolescent animals.

Mice of CBA strain of both sex were injected with [3H]fucose at age 2, 6, 12 or 30 days and the incorporation was determined biochemically 45, 90 and 180 min later. Biochemical measurements of the whole brain (at 90 min postinjection interval) revealed a stepwise age decrease in the amount of incorporated isotope (per mg protein). The amount of [3H]fucose available in the brain has however been found to decrease with age while the degree of its utilization increases. Thus, after correction of the data on [3H]fucose incorporation for the pool of the free isotope, a transient increase in fucosylation rate appeared at postnatal day 12 while the differences among 2-, 6- and 30-day-old animals became negligible. Further, the synthesis of fucosylated soluble glycomacromolecules appeared relatively higher at postnatal day 2 than in older age groups. Examination of different brain regions revealed that the rate of fucosylation is relatively highest in the olfactory bulbs; this prevalence starts appearing with age and becomes most evident in 30-day-old mice. Autoradiography carried out with 6- and 12-day-old animals revealed that the incorporation of [3H]fucose into meningovascular apparatus and the choroid plexus is a few times higher than into brain parenchyma. The regional differences appeared to be similar to those indicated by the biochemical data. Cellular analysis has shown that the incorporation is relatively higher in the cytoplasm of large projecting neurons of the cerebellum, hippocampus and the olfactory bulbs; in the latter region high amounts of macromolecule-bound [3H]fucose also appeared in the synaptic glomeruli.(ABSTRACT TRUNCATED AT 250 WORDS)

DNA content in nerve-cell nucleus. A biochemical and cytophotometric study of the rat cerebrum.

The cerebral nuclei of 30-day-old rats were separated by a two-step gradient centrifugation into the fractions of large and small nuclei. The DNA content per nucleus was determined biochemically and cytophotometrically in these fractions as well as in the non-separated cerebellar and liver-cell nuclei used as reference cells. The DNA content of large and/or mainly neuronal, and the small and/or glial cell-enriched nuclei were 6.74 +/- 0.51 and 6.21 +/- 0.30 pg, respectively, and in the cerebellum 6.10 +/- 0.28 pg per nucleus. The DNA-content values determined cytophotometrically in Feulgen-stained samples of large and small bulk isolated cortical nuclei ranged within the diploid limits, indicated by a part of a liver-cell population measured in parallel. No evidence for the large scale existence of an "extra-DNA" reported earlier [Bregnard, Knuesel and Kuenzle (1975) Histochemie 43, 59-61; Bregnard, Kuenzle and Ruch (1977) Expl Cell Res. 107, 151-157; Kuenzle, Bregnard, Hübschner and Ruch (1978) Expl Cell Res. 113, 151-160] in cortical neurons has thus been obtained.

Tissue culture loading test with storage granules from animal models of neuronal ceroid-lipofuscinosis (Batten disease): testing their lysosomal degradability by normal and Batten cells.

Storage granules (SGs) from ovine and canine models of Batten disease were found to be easily phagocytosed by four cell types studied. The cell types tested were human fibroblasts and peripheral monocytes (control and from a late infantile Batten disease patient), rat C6 cell line, and neonatal cardiomyocytes. The phagocytosed SGs elicited an increase in acid phosphatase activity which was localized in the phagolysosome. After phagocytosis SGs were followed for various times ranging from 7 to 21 days and were found to be of unchanged density (phase contrast), autofluorescence, and ultrastructural appearance. These findings point to their undergradability, or very low degree of degradability, in phagolysosomes in both normal or Batten cultured cells. The Batten disease SGs are not toxic and did not cause any adverse affect on the host cells. Either the normal clearance rate from lysosomes is too slow to be measured by this technique or subunit c accumulation in lysosomes need not result from a primary lysosomal protease defect. Subunit c may aggregate, because of the lack of some normally preventive factor, resulting in a physical barrier to the degradation of this highly apolar molecule.

Molecular mechanisms of improved adhesion and growth of an endothelial cell line cultured on polystyrene implanted with fluorine ions.

Endothelial cells derived from the bovine pulmonary artery (line CPAE, CCL 209, American Tissue Culture Collection, Rockville, MD, USA) were cultured on pristine or fluorine ion-irradiated polystyrene (5 x 10(12) or 5 x 10(14) F ions/cm2, 150 keV). At 24-h post-seeding interval, the number of cells which adhered to the ion-modified polystyrene was significantly higher than on the unmodified material (+20 and +58% in cultures with the polystyrene irradiated by lower and higher ion doses, respectively). On day 7, the populations cultured on the irradiated substrates grew to higher densities, exceeding the controls at the lower and higher ion doses by 69 and 180%, respectively. The cells on ion-implanted samples were also larger (+70-95% and +90-99% at the lower and higher ion doses, respectively) and contained more protein (+16% at both ion doses). As was shown by ELISA, the polystyrene irradiated by the higher ion dose enhanced the expression of a cytoskeletal protein, vimentin (+65%) and protein of focal adhesion plaques, talin (+15%). The content of integrin alpha5beta1 (VLA-5), receptor for fibronectin, was increased at both lower and higher ion doses (+22 and +57%). In contrast to this, the content of ICAM-1 and vinculin was similar in cells grown on both pristine and ion-irradiated growth substrates. Moreover, the expression of VCAM-1 and ELAM-1 was lower by 11-14% in both ion dose groups. The present study has shown that ion implantation of polymers improves the adhesion and growth of endothelial cells without elevating the expression of immunoglobulin and selectin types of adhesion molecules. This surface modification should promote colonization of an artificial vascular prosthesis by endothelial cells and make it less vulnerable by immune system cells of the recipient.

A sex-related difference in the hypertrophic versus hyperplastic response of vascular smooth muscle cells to repeated passaging in culture.

Activation of growth of vascular smooth muscle cells (VSMC) in adults participates in pathogenesis of dysplastic diseases of the vascular system. In this study, we examined the impact of gender of rat donors on the degree of hyperplastic and hypertrophic responses of VSMC in cultures subjected to repeated passaging. The cells were derived from the outgrowth zone of explants of the thoracic aorta and were studied up to passage 45. Under these conditions, the cells undergo repeated growth stimulation by the serum growth factors mimicking some pathological situations in vivo. At lower passages (5-7), the cells from both sex donors did not differ significantly in their doubling time, maximum population density, protein content and ploidy. At higher passages (40-45), we found that the hyperplastic response, monitored by doubling time and BrdU-revealed DNA synthesis, was more intense in VSMC of male origin. In contrast, female-derived cells reacted by more prominent hypertrophic changes. The latter included a relatively higher increase in the volume and protein content of cells. As indicated by the DNA content histograms and chromosome numbers, these cells also showed a higher degree of passage-dependent polyploidization. In addition, the female-derived VSMC were found to be more effective in adhesion to the growth support evidenced by wider spreading and higher resistance of these cells to trypsin-mediated detachment as well as higher expression of some integrin and cytoskeletal molecules. These features could partly account for the slower proliferation and polyploidization of these cells. The results suggest that rat VSMC populations of male and female origin contain cells which are intrinsically different with respect to their capability of reacting to growth stimuli. The lower responsiveness of female-derived cells to growth stimuli may contribute to less frequent formation of hyperplastic vascular lesions in female organisms.