Hits to bone cell nuclei from nonuniform radioactive labels

The stochastic aspects of -particle traversals through nuclei of bone-lining cells from nonuniform radioactive labels are worked out. Both the residence time of the target and the hit rate are considered random variables. It is shown that with any type of bone remodeling the fraction of cells not hit increases with increasing nonuniformity of the label concentration. Thus, a completely uniform concentration represents the most dangerous situation. A possible negative correlation between residence times and hit rates, observed in some experiments, tends to decrease the probability of -particle hits. As a practical application, the theory is applied to the International Commission on Radiological Protection model of the distribution of²³⁹ Pu in the human body. In the case of 50 years of chronic ingestion of 1 annual limit of intake (ALI) per year for class W and chronic inhalation of 1 ALI/year for class Y compounds, more than 19.4 and 8.5% of the nuclei of bone-lining cells are traversed by at least one -particle, respectively.     

Chips to hits

A report on the IBC USA International Microtechnology Conference 'Chips to hits', San Diego, USA, 27 October to 1 November 2001.     

The localization of 1,25-dihydroxycholecalciferol in bone cell nuclei of rachitic chicks

1. A simple technique has been developed to obtain subcellular fractions of chick bone. The method yielded 60-70% of total DNA in the nuclear debris fraction and 80-90% of total (14)C recovered in bone after a dose of radioactive vitamin D. 2. After a dose of [4-(14)C,1,2-(3)H(2)]cholecalciferol (0.5mug) was given to vitamin D-deficient chicks, the time-course of total (14)C radioactivity in the epiphysis, metaphysis and diaphysis of proximal tibiae was measured. The maximum concentrations were reached at 6h, corresponding to a similar peak of radioactivity in blood, decreasing until 24h and indicating the dependence on the circulating (14)C and on the blood supply of the three bone components. 3. The (14)C radioactivity of cholecalciferol and 25-hydroxycholecalciferol (expressed per mg of DNA) followed the pattern of incorporation of total (14)C radioactivity in all three bone components. The more polar metabolite fraction reached a peak of radioactivity at 6-9h and maintained its concentration over the 24h period studied in all anatomical bone components. 4. After a dose of [4-(14)C,1-(3)H]cholecalciferol (0.5mug) was given to vitamin D-deficient chicks, the subcellular distribution was studied. At 24h after dosing, the nuclear fraction contained 27% and the supernatant fraction had 67% of total (14)C recovered in the bone filtrate. When the (14)C in the residual bone fragments was included, the nuclear fraction contained up to 35% of the total radioactivity in the bone. 5. The subcellular distribution pattern of individual vitamin D metabolites indicated that the purified nuclear fraction concentrated the polar metabolite, which lost (3)H at C-1, so that 77% of the radioactivity could be accounted for by 1,25-dihydroxycholecalciferol. The supernatant fraction contained smaller amounts of 1,25-dihydroxycholecalciferol (9%), with 66% of 25-hydroxycholecalciferol forming the major metabolite, corresponding to its concentration found in blood at 24h. 6. The preferential accumulation of 1,25-dihydroxycholecalciferol in the nuclear fraction and the overall pattern of other metabolites, found previously in intestinal tissue, suggests a similar mechanism of action in bone to that postulated for the intestinal cell in calcium translocation.     

Symmetry analysis of cell nuclei

An algorithm is described that is used to analyze the two-dimensional spatial symmetry of cell nuclei. The method provides two symmetry features: the symmetry index (SI), which estimates the precise spatial symmetry of a given chromatin component, Cn, and the quadrant symmetry index (QSI), which estimates the number of quadrants being occupied by Cn. A previous analysis is used to show that age-related change in Malpighian tubule nuclei from the adult housefly is associated with significant alterations in the spatial symmetry of low-, medium-, and high-density chromatin components (LDC, MDC, HDC). This included a seven-fold increase in the spatial symmetry of HDC and a shift in the symmetry profile (from highest to lowest degree of symmetry) from LDC-MDC-HDC to MDC-LDC-HDC. The increased spatial symmetry of HDC suggests that it occurs at new nuclear sites as the fly ages and that these sites are distributed over approximately 60% of the chromosome population.     

On the statistical distribution of volumes of cell nuclei and on the dependence of volumes of cell nuclei and DNA amounts in the cell nuclei

Conclusion Based on the simplified model of the cell nucleus it has been shown that (V) is Gaussian. It follows also from this model the independence ofX andV. A number of further reasons for independence ofX andV is shown too.     

Fixation experiments on cell nuclei

Summary The addition of lanthanum salts to formalin improves nuclear fixation due to the irreversible precipitation of nucleic acids. Especially well preserved nuclei are obtained in smears prepared and dried at — 25° C., pre-fixed with formaldehyde vapors, and postfixed in buffered 5% formalin containing 1% lanthanum acetate or chloride.With this technique, the nuclei reveal a finer and more distinct chromatin pattern, are more intensively stained by hematoxylin and Feulgen reaction, and show no shrinkage. Feulgen-microspectrophotometry demonstrates a considerably higher content of DNA which is also more resistant to nuclease digestion or perchloric acid extraction as compared to nuclei which have been fixed in formalin alone.

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Zusammenfassung Die Zugabe von Lanthansalzen zu Formalin verbessert die Kernfixierung infolge einer irreversiblen Fällung der Nukleinsäuren. Morphologisch besonders gut erhaltene Kerne werden in Tupfpräparaten erzielt, die bei — 25° C hergestellt und getrocknet, zur Vorfixierung Formaldehyddämpfen ausgesetzt und mit gepuffertem 5%-Formol nachfixiert werden, das 1% Lanthanazetat oder -chlorid enthält.Nach Anwendung dieser Technik zeigen die Zellkerne ein feineres und distinkteres Chromatinmuster, sie sind stärker mit Hämatoxylin oder der Feulgen-Reaktion färbbar und lassen keine Schrumpfung erkennen. Das Fixations-Äquivalentbild scheint dem phasenkontrastmikroskopischen Erscheinungsbild lebender Zellkerne besser zu entsprechen. Die Mikrospektrophotometrie von Feulgen-Präparaten zeigt einen beträchtlich höheren Gehalt der individuellen Zellkerne an Desoxyribonukleinsäure. Verglichen mit einfacher Formolfixierung, bewirkt der Lanthanzusatz außerdem eine Erhöhung der DNS-Resistenz gegen Nukleasedigestion und Perchlorsäureextraktion.

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Supported by the General Research and Development Grant of the W. S. U. School of Medicine.     

Three-dimensional approaches to the residual structure of histone-depleted HeLa cell nuclei

Histone-depleted nuclei were prepared by high-salt extraction of interphase HeLa cell nuclei. A large amount of the nuclear DNA remained associated with a rapidly sedimenting residual nuclear structure including cytoplasmic (intermediate filament) and nuclear (matrix and lamina) proteins. Electron microscopy allowed detection in the insoluble structure of a residual nuclear envelope, nucleolar residues, and an intranuclear network whose correspondence with components of in situ fixed nuclei is discussed. Using three-dimensional electron microscopy, it is further demonstrated that the salt-insoluble structure remaining after histone depletion in 2 M NaCl is highly ordered. This is of the utmost importance when considering the roles reportedly ascribed to this structure in nuclear functions.