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Biomedical subjects

J L Hudson

Publications and source records attributed to J L Hudson.

68 records · Page 4Linked to original sources
PubMed1982

A microsample collection device for electrostatically sorted cells or particles and its preparative use for biochemical analysis.

Biochemical analysis of proteins extracted from mammalian cell nuclei which were sorted according to DNA content was greatly simplified using a special collection device to recover sorted nuclei. The device was developed for use with a sorting flow cytometer to provide sufficient numbers of particles to achieve adequate sensitivity and resolution while minimizing sample degradation and loss. The design, evaluation and application of the collection device were described with emphasis on sample preparation. Nuclear populations G0 + G1, S, and G2 + M, were sorted form regenerating rat liver tissue preparations and collected separately in the special chambers. Nuclear proteins were extracted and analyzed by polyacrylamide gel isoelectric focusing or capillary isotachophoresis.

Animals↗
PubMed1981

The modulating effect of isoproterenol on DNA replication and protein synthesis. Synthesis patterns of the HMG proteins from electrostatically sorted salivary gland nuclei during the in vivo cell cycle.

Within 96 h after initial isoproterenol administration, DNA replication and cell cycling were activated, as reflected in the bimodal distribution of nuclear fluorescence determined by flow-microfluorometric techniques. A group of proteins, the cetyltrimethylammonium bromide extractable nuclear proteins (CTAB-proteins), isolated form electrostatically sorted nuclei of rat salivary glands, was shown by staining and autoradiography after two-dimensional electrophoresis to undergo differential synthesis during various phases of the in vivo cell cycle after isoproterenol administration. Stained chromatographs revealed quantitative differences in protein synthesis. Gel autoradiography was a more sensitive technique than staining for detecting nuclear protein synthesis during cell cycling. As observed in the autoradiographs of the CTAB-proteins, isoproterenol initiated two distinct periods of protein synthesis in the salivary gland cell cycle: one during the 2C population G0/G1), and one during the 4C population (G2/M). Protein synthesis after isoproterenol administration was much more dramatic in the 2C (isoproterenol) population, where five new spots were seen. There was less radioactive incorporation in the 4C (isoproterenol) population. Two spots 'a' and 'b' that demonstrate differential protein synthesis in stained gel chromatographs and gel autoradiographs were shown to have electrophoretic mobilities, molecular weights and amino acid compositions highly similar to those of HMG1 and HMG2, respectively. A positive correlation could also be drawn between quantitative levels of 'a' and 'b' and their levels of incorporation during cellular activity with HMG (high mobility group) proteins. For example protein 'b' (HMG2) was consistently more abundant in proliferating cell populations than in the quiescent ones. Autoradiographic patterns of the CTAB-proteins indicated that proteins 'a' and 'b' were synthesized during the G0/G1 phase of the cell cycle, as were the majority of CTAB-proteins.

Amino Acids↗
PubMed1980

The effect of isoproterenol on nuclear protein synthesis in electrostatically sorted rat hepatocytes.

Rat hepatocyte nuclei were electrostatically sorted and collected from specific regions of the cell cycle. Proteins were extracted from homogeneous populations of nuclei with cetyltrimethylammonium bromide and chromatographed by one- and two-dimensional acid-urea polyacrylamide gel electrophoresis. Small quantitative differences were observed in amido black stained two-dimensional patterns of the nuclear proteins of the G0 + G1 and G2 + M populations. However, comparison of the two-dimensional autoradiographs of 3H-leucine labeled nuclear proteins in these populations revealed dramatic differences. After isoproterenol administration, animals showed enhanced labeling of certain nuclear proteins in G2 + M populations, but no noticeable effect in protein labeling was observed in the G0 + G1 populations. Four days after the administration of isoproterenol, fluorescent histograms exhibited increased amounts of DNA in the G2 + M populations. Comparison of the total amino acid composition of two proteins from control animals revealed the distinct chemical contrast of these proteins. Both proteins possessed higher ratios of acidic to basic residues than anticipated for basic proteins. These procedures and findings should prove useful for further understanding of mechanisms of toxicant effects upon nuclear protein metabolism.

Amino Acids↗
PubMed1979

Differences in flow cytometry and 3H-thymidine analyses of perturbed human lymphocytes.

A calf thymocyte crude aqueous extract was tested for DNA synthesis inhibitory activity using phytohemagglutinin-stimulated human peripheral blood lymphocytes. Inhibition of DNA synthesis was assayed using tritiated thymidine and flow cytometry. Although the calf thymocyte crude extract inhibited tritiated thymidine incorporation by over 50%, only very slight changes in the flow cytometric analysis were observed. When dibutyryl-cyclic adenosine monophosphate was used as an inhibitor, a correlation in terms of the inhibition of tritiated thymidine to the inhibition by flow cytometry was observed.

Animals↗
PubMed1979

Nonparametric flow cytometry analysis.

A nonparametric statistical test for the analysis of flow cytometry derived histograms is presented. The method involves smoothing and translocation of data, area normalization, channel by channel determination of the mean and S.D., and use of Bayes' theorem for unknown histogram classification. With this statistical method, different sets of histograms from numerous biological systems can be compared.

Cells, Cultured↗
PubMed1979

Multiparameter kinetic analysis of killer cell initiation by using immune RNA.

By treating nonsensitized C57BL/6J spleen derived lymphocytes with EL-4 tumor cell directed xenogeneic extracted RNA we were able to monitor early changes in cellular DNA content by flow cytometric (FCM) analysis and 3H-thymidine uptake. These kinetic parameters were correlated with cell mediated cytotoxicity which appeared as early as 8 hr after activation as measured by release of chromium-51 from labeled EL-4 target cells. Flow cytometric analysis and 3H-thymidine uptake data shown peak S phase activity at 72 hr. Maximum cytotoxicity was observed at 48 hr. Cell cycle kinetic parameters were correlated with the appearance of cell mediated cytotoxicity.

Animals↗
PubMed1978

Inhibition of growth and guanylate cyclase activity of an undifferentiated prostate adenocarcinoma by an extract of the balsam pear (Momordica charantia abbreviata).

We have recently described the presence of a guanylate cyclase [GTP pyrophosphate-lyase (cyclizing), EC 4.6.1.2] inhibitor (GCI) in an aqueous extract of the balsam pear (Momordica charantia abbreviata). Because the guanylate cyclase-cyclic GMP system is though to be involved in cell growth, DNA and RNA synthesis, and possible malignant transformation, we examined the effect of the aqueous extract containing GCI on an undifferentiated adenocarcinoma of the rat prostate and concanavalin-A-stimulated [3H]thymidine incorporation into cultured splenic lymphocytes, a process thought to be mediated by cyclic GMP. The results demonstrate that the extract of the balsam pear blocks both the growth of the rat prostatic adencarcinoma in vitro and [3H]thymidine incorporation into DNA. DNA histograms from flow cytometry indicated that the extract containing GCI inhibited in the G2 + M phase of the cell cycle, a presumed locus of cyclic GMP effects. In addition, guanylate cyclase activity was significantly greater in the tumor than normal prostate tissue and was decreased by the extract containing GCI. Cyclic GMP levels in the tumor in culture wer also decreased by addition of the extract. It remains to be determined whether or not the anti-tumor agent and GCI are the same substance.

Adenocarcinoma↗
PubMed1978

Effect of methionine-loading on methyl group synthesis and activation in rat brain and liver.

Much greater increases in S-adenosylmethionine concentrations are observed in the liver in response to methionine-loading than in the brain due to differences in the methionine adenosyltransferase activities in these tissues. Liver methione adenosyltransferase exhibits a bimodal saturation curve with a nonlinear Line-weaver-Burk plot, indicating that high methionine concentrations are required for saturation. In the brain the methionine adenosyltransferase is saturated in vitro at a methionine concentration less than the normal physiological concentration. The increased S-adenosylmethionine concentrations in the livers of methionine-treated rats also account for the observed inhibition of N5,N10-methylenetetrahydrofolate reductase activity in this tissue. No inhibition of this enzyme is observed in the brain of methionine treated animals. Nor are S-adenosylmethionine concentrations increased significantly in brain. Serine hydroxymethyltransferase activity responds to methionine-loading by decreasing in brain and increasing in liver.

Animals↗
PubMed1977

High resolution proton magnetic resonance spectral characteristics of water, lipid, and protein signals from three mouse cell populations.

The high resolution proton magnetic resonance spectral properties of three C57BL/6J mouse cell populations (EL4 ascites tumor cells, normal spleen leukocytes, and normal erythrocytes) were investigated. Packed cell pellets were investigated at 100 MHz and 13.56 MHz. The 100-MHz spectra contained identifiable non-water proton resonances as well as the water resonance. The major non-water resonances from the EL 4 cells and normal leukocytes resembled resonances from lipid extracts, whereas the non-water resonances from erythrocytes resembled resonances from hemoglobin solutions. The reciprocals of the water proton spin-lattice relaxation times were roughly proportional to the total sample water content at both 100 and 13.56 MHz. The estimated slopes of these plots were frequency-dependent. The water proton relaxation rate generally increased with increasing total protein content of the packed cell pellets.

Animals↗
PubMed1975

Immunoprophylaxis and cytotoxic effector cells against EL 4 leukemia induced in syngeneic C57BL/6J mice by use of irradiated EL 4 cells.

Tumor-specific immunoprophylaxis was achieved in C57BL/6J mice against EL 4 leukosis cell challenge by sensitization of the syngeneic host with multiple ip injections of irradiated EL 4 cells. A minimal radiation dose was used to replication-block EL 4 cells before inoculation, as defined by dose-response analysis of irradiated EL 4 cells. Multiple ip injections of irradiated EL 4 cells stimulated development of significant, yet relatively low, levels of cytotoxic lymphoid activity (CLA) in lymphoid cells of the peritoneal exudate as measured by in vitro 51Cr-release cytotoxicity assays. The specific temporal and frequency dependencies of the inoculation regimen for achieving immunoprophylaxis indicated that, in addition to CLA, other, short-lived, immune processes were important in the tumor rejection. These observations showed the capacity of the C57BL/6J host for tumor-specific immune recognition and rejection of the syngeneic EL 4 leukemia. The tumor rejection could be elicited solely by inoculations of irradiated EL 4 cells and did not require exogenous amplifiers, such as immunoadjuvants, chemical modifiers, and/or allogeneic immune information transfer.

Animals↗
PubMedDate not supplied

A 16-channel automated rotometer system for reliable measurement of turning behavior in 6-hydroxydopamine lesioned and transplanted rats.

Unilateral 6-hydroxydopamine lesions of the nigrostriatal pathway in rats result in a massive dopamine (DA) denervation of the ipsilateral striatum. Such animals have proven extremely useful as a model for the study of Parkinson's disease, an idiopathic neurodegenerative disorder of humans. Extensive unilateral DA disruption leaves the rat relatively normal in motor behavior; however, the extent of the lesion can be documented by drug-induced rotational behavior. When given an injection of a dopamine agonist, such as apomorphine or D-amphetamine, a lesioned animal will manifest rotational behavior; the number of turns correlates with the degree of unilateral denervation. In order to identify, for various studies, large numbers of animals with specific levels of denervation, the necessity of an automated and reliable rotational counting system (rotometer) becomes obvious. We have developed such a device that allows up to 16 rats to be tested concurrently with one inexpensive computer. This system is more reliable than, and certainly preferable to, more tedious methods such as videotaping and subsequent manual analysis or various other mechanical systems. Plexiglass, formed into large bowls, serve as the rotometer chambers. We have designed a simple, inexpensive, and accurate counting head that can be manufactured from readily available parts and that is very sturdy and reliable. This, together with a thoracic harness, completes the rotometer assembly. The rotational data, from up to 16 separate channels, is collected by a single-chip microprocessor and sent on a serial line to an IBM-type or MacIntosh host computer. There, it is graphically displayed on line and subsequently saved to disk with a novel acquisition program.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenal Medulla↗