Search PubMed⌕ Search

Biomedical subjects

C Nicolini

Publications and source records attributed to C Nicolini.

At least 91 records · Page 5Linked to original sources
PubMed1980

FPi analysis. II. Use of the method to monitor the in vivo kinetics of cell populations perturbed by hydroxyurea.

In a previous report, we described a new method called FPi analysis to analyze time sequences of DNA histograms taken from a perturbed population of cells. In this paper we utilize the method to analyze the in vivo kinetic response of bone marrow and of lung metastases of the B16 tumor to various chemotherapeutic agents. We show that the technique allows useful kinetic data to be obtained with minimal processing of the raw histograms, thus allowing fast analysis of the data. We also show that, in order to monitor the kinetic response of living tissues, it is essential to collect multiparameter distributions; to monitor only the one dimensional fluorescence histogram can give rise to misleading results. Using these multiparameter histograms, we are also able to monitor the growth fraction of the lung metastases during treatment, allowing discrimination between cell synchrony and cell recruitment from the resting compartment.

Animals↗
PubMed1979

Morphometric analysis of B2cAMP induced reverse transformation in synchronized CHO cells.

Synchronized tranformed and reverse-transformed (by 10(-3) M B2cAMP) CHO-K1 cells, growing adherent to plastic, are characterized by means of geometric and densitometric parameters at the level of both the entire cell and of the nuclei at various time intervals after selective miotic detachment. Transformed and reverse-transformed cells triple-stained with Feulgen, Napthol Yellow S, and periodic acid-Schiff appeared very similar in terms of integrated optical density (IOD), related to either polysaccharides, protein, or DNA amount. On the other hand, a shift from a polygonal to a spindle-shaped morphology is a accompanied by a significant decrease in both form factor and average optical density (AOD) of intact cell and nuclei, which are the most conspicuous measured changes caused by B2cAMP, in addition to a lengthening of the cell cycle duration. In both control and treated cells, important and parallel cell-cycle-dependent modulations of geometric and densitometric parameters are also observed, for both the cytoplasmic (i.e., cell morphometry) and DNA space (i e., nuclear morphometry). Specifically, the modulation in nulear morphometry during G1, S, G2, and M phases confirms previous findings on synchronized HeLa cells. The optical density threshold-dependence of geometric parameters shows that, while becoming fusiform, the cytoplasm of reverse-transformed cells had a particularly low optical density precisely in the polar area. Utilization of such an approach in the development of an objective morphological classification of all cell lines grown as monolayers "in vitro" is also discussed.

Animals↗
PubMed1979

Mass action and acridine orange staining: static and flow cytofluorometry.

We present results involving an approach to acridine orange staining of intact cells based on basic physicochemical considerations. We show by static microfluorometry of several in vitro and in vivo cell lines that the important parameters for such staining are the molar ratio (Formula: see text), and molar concentration of acridine orange. Differential nuclear DNA and cytoplasmic RNA staining are totally controlled by these two parameters. We show this by a physicochemical model of cell-dye interaction. Finally, we use the method to study the growth parameters of complex in vivo cell populations by automated multiparameter flow microfluorometry. We have explored also, both by static and flow systems, the effect on AO-cell staining of various cell pretreatments such as Triton X-100 and chelating agents.

Acridine Orange↗
PubMed1979

Cytofluorometry of electromagnetically controlled cell dedifferentiation.

Cellular morphology changes, which appear related to dedifferentiation (despecialization), have been produced in vitro in the nucleated red blood cell of the frog. This has been achieved by controlled alterations in the electrochemical environment of these living cells, both by a selective modification of the ionic concentrations of an isotonic amphibian Ringer solution, and by the electromagnetic induction of pulsating current having specific waveform parameters. Laser flow microfluorometry shows that the modified Ringer solution is able, per se, to partially trigger the process in the same time interval that certain induced current waveforms can significantly affect the number of cells in the so-called dedifferentiated state. It has also been found that, for a given waveform, the repetition rate appears to have a significant effect on the rate of cell change. Preliminary automated image analysis of cell smears suggests that dedifferentiated and normal cells have the same integrated optical density but different nuclear areas. In conclusion, it appears that, after the initial electrochemical trigger, the early stage of the process, when the cells move from a state of specialized function to one of less specific activity, is the unfolding of their chromatin supercoil, not involving DNA synthesis. Then cytofluorometry allowed us to identify, for the first time, fundamental modifications which occur in the cell nucleus under electromagnetic exposure.

Acridine Orange↗
PubMed1979

Flow cytometric prescreening of cervical smears.

One-parameter (nuclear DNA) and two-parameter (nuclear DNA and protein or cellular light scatter) measurements of cervical smears were performed using an ICP 11 and a cytofluorograf 4800 respectively. A total of about 1000 cases was analyzed. For the estimation of nuclear DNA alone two fluorochromes were tested (ethidium bromide (EB) and mithramycin (MMC)) combined with three different methods of cell preparation. For the two-parameter measurements cells were double stained with EB and fluorescein isothiocyanate (FITC). Red fluorescence (EB) versus green fluorescence (FITC) or red fluorescence versus scatter were recorded. A computer analysis of the one-parameter histograms was performed using discriminant analysis and the results were compared with the cytodiagnosis of microscopic specimens stained with the Papanicolaou technique. The error rates of the flow cytometric (FCM) data were as follows: (a) standard EB staining, 11% false negative, 26% false positive, 6% unsatisfactory results; (b) pepsination of vital cells and EB staining, 12% false negative, 14% false positive and 4% unsatisfactory results; (c) MMC staining, 10% false negative, 65% false positive and 5% unsatisfactory results. Our two-parameter measurements prove that, as confirmed by cell sorting, red fluorescence versus scatter allows separation of at least three subpopulations in most analyzed samples: (a) anucleated cells; (b) leukocytes; and (c) intermediate and superficial cells.

Cell Nucleus↗
PubMed1979

Early detection of micrometastases via flow microfluorimetry.

We show that by measuring the joint distribution of fluorescent and scattered light in a suspension of lung taken from the C57B1/6J mouse, we are able to discriminate in vivo between the lung metastases of the B16 melanoma and the normal lung tissue. We can in such a way detect metastatic cells in a very early stage of growth and also obtain growth curves for the metastatic population. We analyze the sensitivity of the method of detection and speculate how it might be used in aiding in human diagnosis.

Animals↗
PubMed1978

Size dependence of cell cycle parameters for lung metastases from B16 melanoma.

Data presented show that the cell cycle time of the lung metastases from a B16 melanoma tumor growing in the foot pad of C57 BL/6J mice increases as the size of the metastases increases. This increased cell phase durations of the small metastases being closer to those of the primary tumor. The growth fractions of the primary tumor and of the lung metastases of different sizes are analyzed.

Animals↗
PubMed1977

Nuclear morphometry during the cell cycle.

Directly measured and derived geometric and densitometric parameters were obtained by means of the automated image analyzer Quantimet 720-D in Feulgen-stained HeLa cells synchronized by selective mitotic detachment. These data indicate substantial alteration of nuclear morphology during the entire cell cycle, even during the G1 and G2 phases, and the late G1-early S and late S-G2 transitions.

Cell Division↗
PubMed1977

Objective identification of cell cycle phases and subphases by automated image analysis.

Frequency distributions of integrated optical density, perimeter, projection, area, form factor, average optical density, and mean dispersion path of nuclear images of Feulgen-stained HeLa S3 cells were obtained by automated image analysis at the base threshold of 0.04 OD. The mean values and standard deviations of these geometric parameters were then computed versus increasing values of threshold (0.08--0.32 OD). There is clear evidence of differential chromatin dispersion and convolution during the cycle of synchronized HeLa S3 cells at different times after selective mitotic detachment. The combination of average OD, form factor, and mean dispersion path at base threshold with the threshold dependence of nuclear morphometric parameters permits objective identification of cell cycle phases and their subphases, by characterizing variations in chromatin geometry within and between phases, regardless of whether DNA content remains constant (early G1, middle G1, late G1), varies only slightly (late G1-early S or late S-G2 transitions), or varies significantly (early S-middle S).

Autoanalysis↗
PubMed1977

Crypt cell isolation in the small intestine of the mouse.

A successful method has been developed for isolating viable single cell suspensions of intestinal crypt cells from the small bowel of the mouse. The lumen of the intestine was perfused with a 0.2% trypsin solution that dissociated the lining epithelial cells. Crypt cell isolation, which proves to be extremely critical, occurred under optimal mechanical and chemical configurations about 75 min after the beginning of the procedure.

Animals↗
PubMed1977

Experimental results on mammalian cells growing in vitro in deuterated medium for neutron-scattering studies.

SV-40 virus-transfromed human diploid fibroblasts (2RA) were grown in a monolayer on plastic Petri dishes in an aqueous medium deuterated to different concentrations of deuterium oxide: 10, 20, 30, up to 60%. The cells must be acclimatized to concentrations higher than 20% D2O by stepping them from a lower initial concentration during their exponential growth. The increase of cell doubling time with increasing deuterium concentrations seems to correlate, at least at 20% D20, with an initial period of suspended cell growth (lag-phase), and is qualitatively similar to that previously reported for Escherichia coli.

Cell Division↗
PubMed1976

DNA structure in sheared and unsheared chromatin.

Shearing chromatin, by either sonication or vortex homogenization, introduces significant structural artifacts. These may be detected by the anomalously large increase in the number of ethidium bromide binding sites and the large alteration of the circular dichroism spectra of chromatin. Structural alterations are also suggested by the disappearance of differential light scattering after shearing.

Binding Sites↗
PubMed1976

Chromatin structure and function in proliferating cells.

The conclusions that we would like to draw from this review are the following: (a) Chromatin structure and function are exceedingly sensitive to changes in the proliferative state of a cell. Differences can be detected between cells in mitosis, G1 and S, and even between G0 and G1 cells. (b)These differences are very unlikely to be artifactual, since similar changes can also be demonstrated in intact nuclei. (c) Some of these differences can be abolished by extraction of chromatins with low concentrations of salt. (d) Differences between chromatins of normal and neoplastic cells can also be detected, but they are largely related to differences in the extent of cell proliferation. (e) A number of laboratories have been very busy in trying to elucidate chromatin structure with different technologies. Sometimes a change in a macromolecule cause by a physiological stimulus can tell us as much about its structure as a thousand instruments. The changes occuring in chromatin of proliferating cells could perhaps be profitably used to know more about chromatin structure.

Animals↗
PubMed1976

Thermal denaturation of sheared and unsheared chromatin by absorption and circular dichroism measurements.

Thermal denaturation of chromatin is observed by simultaneously monitoring absorption and circular dichroism at 276 nm as functions of temperature. Either observation indicates that sheared chromatins shows less thermal stability than native chromatin. The temperature-dependent ellipticities at 276 nm of these chromatins show features not seen in the absorption curves: the ellipticity of unsheared chromatin increases with temperature, while this increase is abolished or greatly reduced in the same chromatin after shearing. After its first thermal transition (prior to the helix-coli transition) the unsheared chromatin achieves the same ellipticity as sheared chromatin.

Chromatin↗