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PubMed1982

"Hidden" cytotoxic antibodies that react with allogeneic cultured fetal and tumor cells contained in soluble immune complexes from normal human sera.

Hidden complement-dependent cytotoxins were demonstrated in normal donor sera after removing anti-immunoglobulins that normally block the ability of these immunoglobulin Gs to react with surface antigens on tumor cells. The blocking antibodies had certain properties of anti-idiotypes. Immunoglobulin G from Cohn Fraction II, after removing these anti-immunoglobulins, was cytotoxic for cultured fetal cells and for malignant melanoma, breast, lung, colon, and other tumor cells maintained either in tissue culture or by serial passage in nude mice. The cytotoxins were not adsorbed by extracts from normal lymphoid, liver, skin, or red blood cells. These results suggest that a heterogeneous group of natural antibodies reactive with antigens expressed on a variety of neoplastic and fetal cells circulate in normal donor sera as part of a soluble immune complex, together with a blocking anti-immunoglobulin.

Animals↗
PubMed1994

Liver regeneration after partial hepatectomy in the rat. Sequential events monitored by 31P-nuclear magnetic resonance spectroscopy and biochemical studies.

BACKGROUND: Little data exist wherein both the 31P nuclear magnetic resonance (NMR) signals and biochemical changes associated with hepatic regeneration after a 70% hepatic resection have been assessed simultaneously. EXPERIMENTAL DESIGN: Two groups of rats were used: one group underwent a 70% partial hepatectomy and the second underwent a sham operation. Both groups were followed sequentially for 192 hours by in vivo serial 31P-NMR spectroscopy of the liver and its phospholipid extracts. Liver injury and function were assessed by biochemical means. RESULTS: After surgery, a significant reduction in ATP and an increase in the phosphomonoester signal for the hepatectomized animals were noted as compared with the controls (p < 0.05). The phosphodiester content of the liver in the hepatectomized rats declined to nonmeasurable amounts in vivo. The nadir of ATP occurred 72 hours after surgery. The area of the phosphomonoester relative to an external reference of methylenediphosphonic acid peak increased steadily over the first 96 hours, whereas that of the area ratio of the inorganic phosphate/methylenediphosphonic increased over the first 72 hours posthepatectomy. The intracellular pH declined sharply in the first 3 days, followed by a gradual recovery over the next 5 days. Little change in the intracellular pH was observed for the control animals. A significant increase in the area of the phosphorylethanolamine relative to an internal reference of methylenediphosphonic and a reduction in the glycerophosphorylethanolamine and glycerophosphorylcholine peaks were noted during the first four post-hepatectomy days as measured by 31P-NMR of perchloric acid liver extracts (p < 0.05). CONCLUSIONS: It has been found that a concerted reduction in the intracellular ATP and intracellular pH coupled with an increase in inorganic phosphate and high levels of phosphorylethanolamine occur as a result of hepatic regeneration and the physiologic changes induced. These data demonstrated that a coordinated pattern of biochemical changes occur with and after hepatic regeneration. Moreover, NMR spectroscopy demonstrates an increase in phosphomonoesters and a decline in phosphodiesters during hepatic regeneration. These measures and, more specifically, the ratio of these two lipid classes may provide a biochemical snapshot of the regeneration status of the liver.

Adenosine Triphosphate↗
PubMed1958

The role of carbon dioxide as an essential nutrient for six permanent strains of fibroblasts.

The results of these studies demonstrate that carbon dioxide is required for the growth and maintenance of strains of fibroblasts derived from human tissues, strains FS4-705 and U12-705, from mouse tissue, strain L-705, and from rabbit tissues, strains RM3-56, RS1-56, and RT-56 in a chemically defined medium containing phosphite buffer in place of bicarbonate and supplemented with dialyzed serum and dialyzed embryo extract. Under these conditions, the cells fail to proliferate at a significant rate and begin to degenerate within 5 to 10 days when the flasks are not stoppered. Sufficient carbon dioxide is produced by the cells to promote growth as indicated by the fact that maximal proliferation is obtained in the same phosphite media when stoppered flasks are employed. With the exception of RS1-56, all the remaining strains tested can be propagated serially in open flasks containing phosphite medium prepared with whole serum and embryo extract. The rate of growth under these conditions, however, is only one-half to one-third that obtained in stoppered flasks containing phosphite medium or the conventional bicarbonate medium.

Animals↗
PubMed2002

Accurate diagnosis of occlusal carious lesions--a stereo microscope evaluation of clinical diagnosis.

This study was undertaken to validate the caries status of 214 teeth by serial sectioning and microscopy after caries diagnosis using four methods. Two hundred and fourteen extracted human teeth with varying degrees of caries were mounted in the jaws of nine training manikins. All tooth surfaces were examined and recorded for caries by four dentists using bitewing radiographs, fibre-optic transillumination (FOTI), mirror alone and a mirror and sharp probe on two separate occasions. Thereafter the teeth were serially sectioned and assessed microscopically for depth of caries lesion on a graded score of 0-7. This report assessed the diagnostic outcome of 2,183 observations for occlusal surfaces. Sound diagnoses predominated over unsound until caries was present in the inner half of dentine. Specificity was between 90% and 95% and sensitivity 26% and 50% depending on which diagnostic method was used and where the sound/unsound threshold was set. Negative and positive predictive values were similarly influenced and varied between 53% and 80% and 73% and 90%, respectively. Probit analysis showed no significant differences (P < 0.05) between examiners and diagnostic methods. Diagnosis of occlusal caries undertaken in an in vitro simulated clinical situation is inaccurate until the caries lesion extends deep into the dentine no matter which of the four methods was used.

Chi-Square Distribution↗
PubMed1970

Induction of papillomas in rabbits with nucleic acid extracts from Vx7 carcinomas.

Nucleic acid extracts from transplantable carcinomas Vx7 and Vx2, long maintained in domestic rabbits, were assayed for their ability to produce papillomas in animals of this kind. The Vx7 had been serially transferred 111 times when this was attempted and Vx2 was in its 203rd generation. The nucleic acid extracts from the Vx7 carcinomas consistently yielded papillomas whereas those from Vx2 completely failed to do so. The tumorigenicity of the Vx7 extracts was slight and regression of the induced papillomas often took place, as happens not infrequently to the growths caused by nucleic acid extracts obtained directly from papillomas. Malignant conversion, a common event in tumors induced by Shope papilloma virus, was also observed to occur among papillomas which were induced by the nucleic acid extracts from Vx7 carcinomas, and which kept persisting over a year on the skin of experimental animals.

Animals↗
PubMed2001

Detection of bacteria in environmental samples by direct PCR without DNA extraction.

Cultured cells and environmental samples were used directly in PCRs without the isolation of DNA. Serial dilution was used to eliminate the inhibitory effect of materials in natural samples. Primers specific for pmoA, which encodes a subunit of the particulate methane monooxygenase, were used to detect and quantify methanotrophic bacteria by direct most probable number PCR. Phototrophic bacteria were detected in environmental samples by direct PCR with primers specific for pufM, and members of the bacterial domain were detected with primers for 16S rDNA. Direct PCR provides a rapid, simple, and sensitive methodfor detecting and quantifying bacteria in environmental samples. Detection of methanotrophic bacteria can be applied to monitoring bioremediation.

Bacteria↗
PubMed1999

A Three-Dimensional Finite Element Scheme to Investigate the Apparent Mechanical Properties of Trabecular Bone.

A computational technique is described for investigating the apparent mechanical properties of trabecular bone based on tissue geometry obtained from the marching cubes volume rendering scheme. Using this scheme, a 3D representation of the trabecular bone was extracted from two-dimensional cross-sections of the tissue originating from a quantitative serial sectioning procedure. Surface information consists of node coordinates and polygon connectivity in a 3D space. A custom, adaptive mesh generation technique using a normal offset was used to prepare 3D finite element volume meshes (4-node tetrahedral elements) of variable mesh density from the extracted surface geometry. Nine target mesh resolutions (32 µm to 107 µm) were examined for a (1.5 mmx 1.5 mmx 2 mm) volume of trabecular bone. A mesh density of 50,000 elements/mm(3) of bone tissue was found to be adequate for convergence of apparent (bulk) modulus for 1% uniaxial compression. For this convergent case, the maximum local normal compressive tissue stress was 400 MPa which was six hundred-fold greater than the computed apparent stress. Variation in the apparent modulus was less than 5% when Poisson's ratio values were varied between 0.1 and 0.4. Poisson's ratio values greater than 0.4 had a more marked effect on the apparent modulus. Based upon these results, approximately 1 million, 4-node tetrahedral elements are required to analyze a continuum scale model of trabecular bone (5 mm cube).

Journal Article↗
PubMed2003

Liquid chromatography/mass spectrometry of malonyl-ginsenosides in the authentication of ginseng.

Different negative ion electrospray (ES) source conditions are required to concentrate the ion current in [M-H](-) for malonylated and non-malonylated ginsenosides. However, both can be ionised optimally in a single liquid chromatography/mass spectrometry (LC/MS) analysis by employing switchable voltages in the post-source ion optics of a quadrupole ion trap mass spectrometer. Coupled with automatic MS/MS scanning and post-acquisition neutral loss data analysis, this method provides a means of profiling the malonylated and acetylated ginsenosides in ginseng extracts. Analyses revealed numerous malonylated ginsenosides that could be partially characterised by serial MS/MS experiments. The ratio of mRb(1) to other isomeric forms present and to mRb(2) and mRc appears to show consistent differences among Panax ginseng (Asian ginseng), P. quinquefolius (American ginseng) and P. notoginseng (Sanchi ginseng). The ratio of malonylated to non-malonylated ginsenosides is reduced in the red form of Asian ginseng compared with the white form and there is a concomitant increase in the levels of the corresponding acetylated ginsenosides. The ability to analyse malonylated ginsenosides is an important contribution to the range of chemical characteristics that can be used to authenticate the different species of ginseng and will assist in quality control and standardisation.

Chromatography, Liquid↗
PubMed1999

Lymph node extraction of radiopaque nanoparticulates in the rabbit as measured in vivo with CT.

RATIONALE AND OBJECTIVES: The purpose of this study was to estimate in vivo extraction of lymphographic material in the popliteal node of the rabbit. MATERIALS AND METHODS: Serial quantitative computed tomography (CT) of target tissues in four legs of two rabbits was performed after subcutaneous injection of an improved lymphographic contrast agent. Massage was used as a lymphotrophic intervention. RESULTS: At 15 minutes, the mean change in Hounsfield units measured 815 in the popliteal node, 219 in afferent lymphatic vessels, and 127 in efferent lymphatic vessels. The nodal extraction of nanoparticulates from the lymph was approximately 55%. Nodal massage allowed the amount of nanoparticulate remaining in sinusoidal lymph to be estimated. CONCLUSION: Functional CT performed with timed studies, proper radiopaque materials, and physiologic interventions can depict in vivo lymphatic physiology under minimally invasive conditions.

Animals↗
PubMed1980

Immunochemical differences between angiotensin I-forming enzymes in man.

1. Human plasma, amniotic fluid and acidified amniotic fluid were incubated at pH 5.5 with the same concentrations of human plasma renin substrate and rat plasma renin substrate. They produced three to eight times more angiotensin I with human than with rat renin substrate. By contrast, human brain extracts generated 20 times more angiotensin I when incubated with rat plasma renin substrate than with human plasma renin substrate. 2. Serial dilutions of anti-(human renin) antibody inhibited, in a dose-dependent manner, the production of angiotension I when plasma, amniotic fluid and brain extracts were incubated with human plasma renin substrate. They also inhibited the production of angiotensin I when plasma and amniotic fluid were incubated with rat plasma renin substrate. They were ineffective on the angiotensin I generation by human brain extracts acting on rat plasma renin substrate. 3. Affinity chromatography on an haemoglobin-Sepharose gel separated the fraction of brain extract acting on human renin substrate and inhibited by anti-(human renin) antiserum; this was not retained on the gel at pH 3.3. Part of the angiotensin I-forming activity detected by rat renin substrate hydrolysis was not retained on the gel and part was eluted at pH 8.5. These angiotensin I-forming activities did not hydrolyse human renin substrate, and were not neutralized by anti-(human renin) antibody. 4. These results demonstrate that a renin, immunochemically identical with renal, plasma amd amniotic fluid renin, is present in the human brain. Other angiotensin I-forming activity, acting on an heterologous substrate at a more acidic pH, is also present in human brain.

Angiotensin I↗
PubMed1994

Event-related potentials indicate information extraction in a comparative judgement task.

Event-related potentials (ERPs) were recorded from normal subjects in a serial paired-comparison task with the spoken digits one to five. For each digit, the subject decided whether it was smaller or larger than the preceding one. We examined to what extent ERP components are associated with the probability information each digit provides with respect to the subsequent response. Shortest reaction time (RT) and highest accuracy was obtained for the two end terms one and five. The N1 and P2 components of the ERP were attenuated for the two end terms. Three late positivities were found: early P3, P3b, and slow wave. P3b increased in amplitude the more information the eliciting digit provided with respect to the response to the subsequent digit. Moreover, a negative correlation was found between the P3b amplitude associated with information extraction and the RTs to the subsequent digit. During comparative judgement, subjects process the relative position of a stimulus within a stimulus set to extract information about the probability of a forthcoming response, and this process is reflected in P3b amplitude.

Adult↗
PubMed1990

Age-related changes in the response of chick lens cells during long-term culture to insulin, cyclic AMP, retinoic acid and a bovine retinal extract.

We have reported that 1-day-old post-hatch chick lens epithelial cells lose the capacity for lentoid body formation and delta-crystallin expression during long-term serial subculture, although they continue to synthesize, but not to accumulate, alpha- and beta-crystallins, even in cells with a transformed phenotype. Here we present evidence that dedifferentiation may reflect an age-related change in the capacity for response to regulatory signals. We have tested the capacity of these cells in serial subcultures to respond to agencies which affect lens cell growth and differentiation in primary culture: retinoic acid (RA), insulin, cAMP and bovine retinal extract (BRE). Secondary cultures responded only to RA and BRE, by an increase in lentoid formation and by alpha- and beta-accumulation, while RA also restored delta-crystallin expression. Later cultures showed no such responses. The results suggest that the process of lens cell dedifferentiation may, at first, be reversible but later becomes irreversible, despite the continuing persistence of low levels of crystallin expression.

Animals↗
PubMedDate not supplied

True versus immunoreactive angiotensin II in human plasma.

To measure specifically angiotensin-(1-8)octapeptide, peptides were extracted from 2 ml of plasma by reversible adsorption to bonded-phase silica. The angiotensin-(1-8)octapeptide was then isolated by isocratic reversed-phase high-performance liquid chromatography and quantified by radioimmunoassay. The extraction recovery of 125I-angiotensin II added to 2 ml of plasma was 99 +/- 2% (mean +/- SD). The overall recovery of 5, 10, and 20 fmol unlabeled angiotensin II added to 1 ml of plasma was 80 +/- 10%. The coefficient of variation for within-assay precision was 0.06 and for between-assay precision 0.13. The detection limit was 0.4 fmol/ml. Buffer and plasma blanks were below the detection limit. Normal subjects on a free diet in supine position averaged 4.2 +/- 1.7 fmol/ml angiotensin-(1-8)octapeptide. Furosemide (40 mg p.o.) and standing increased these values to 22 +/- 7.6 fmol/ml. In four volunteers, immunoreactive "angiotensin II" (more or less angiotensin-like material) was measured serially before and after converting-enzyme inhibition (Hoe 498) with conventional Dowex extraction. At peak inhibition, plasma immunoreactive "angiotensin II" levels decreased by only 44%. In contrast, angiotensin-(1-8)-octapeptide isolated by high-performance liquid chromatography completely disappeared. In hypertensive patients receiving long-term treatment with enalapril, plasma levels of angiotensin-(1-8)octapeptide fell from 2.7 +/- 0.9 to 0.9 +/- 0.3 fmol/ml (mean +/- SEM) 2 hours after the morning dose, whereas levels of immunoreactive "angiotensin II" were not significantly changed. We found that this sensitive method specifically measured angiotensin-(1-8)octapeptide and demonstrated that true angiotensin II virtually disappears during converting-enzyme inhibition.

Adult↗
PubMed2005

Solving mysteries of DNA replication and frog cloning.

Compared to sperm nuclei, nuclei from adult somatic cells replicate inefficiently in frog egg extract. In this issue of Cell, Lemaitre et al. (2005) show that pre-exposure of erythrocyte nuclei to a mitotic extract removes this difference, reorganizes the chromatin into shorter loops, and allows replication at much shorter intervals along the DNA. Remarkably, these observations also explain an old mystery of why serial nuclear transplantation was so successful for cloning frogs.

Animals↗
PubMed2004

Interferon-alpha-inducible proteins are novel autoantigens in murine lupus.

OBJECTIVE: To investigate the spectrum of B cell autoimmunity in the recently described anti-CD1-autoreactive T cell receptor (TCR)-transgenic murine lupus-like (CD1 lupus-like) model. METHODS: Lethally irradiated BALB/c/nu/nu mice were injected intravenously with donor BALB/c bone marrow and spleen cells expressing TCRalpha and TCRbeta transgenes that recognize CD1d. Sera from adoptive host animals that developed lupus (i.e., CD1 lupus mice) were collected at serial time points and analyzed by Western blotting and immunoprecipitation, using protein extracts prepared from NIH3T3 mouse fibroblasts and EL-4 lymphocytes, respectively. Sera obtained from older animals in several models of spontaneous lupus (NZB/NZW, MRL++, and MRL/lpr mice), unmanipulated BALB/c/nu/nu mice, and normal BALB/c mice were used as controls. RESULTS: Analyses demonstrated that the prominent targets of autoantibodies in the CD1 lupus-like model are interferon-alpha (IFNalpha)-inducible antigens. Biochemical and serologic characterizations identified one antigen as belonging to the interferon-inducible 202 (Ifi202) subfamily of proteins within the Ifi200 family, and a second antigen as a member of the 70-kd heat-shock protein family. Autoantibodies directed against these antigens were rapidly produced at an early stage of disease. Anti-p50 autoantibodies were present in sera from 7 (78%) of 9 CD1 lupus mice that developed severe kidney disease. CONCLUSION: IFNalpha-inducible proteins represent a novel class of autoantigens in murine lupus, and the findings suggest additional roles for IFNalpha in this disease. Since Ifi202 autoantigens are encoded by the murine non-major histocompatibility complex lupus-susceptibility gene locus Ifi202, these data provide a link between recent advances in lupus genetics and the formation of autoantibodies.

Animals↗
PubMed2002

Kinetics of fluorescence expression in nonhuman primates transplanted with GFP retrovirus-modified CD34 cells.

Downregulation and loss of proviral expression have been demonstrated to occur in a variety of in vitro studies and in mouse models. Here we evaluated the kinetics of proviral expression after transplantation in a competitive repopulating model in the baboon. Transgene persistence and green fluorescent protein (GFP) expression in peripheral blood leukocytes (PBL) were analyzed in four animals by semiquantitative PCR and flow cytometry for up to 80 weeks (range 17-80). All animals were transplanted with cells transduced with EGFP or EYFP reporters driven by Moloney murine leukemia virus (MoMuLV) or a modified promoter/enhancer, (MND) respectively. Simultaneous dual-color analysis of fluorescence levels in granulocyte and lymphocyte subsets following hematopoietic reconstitution demonstrated progressive loss of fluorescence intensity occurring predominantly early after transplant in cells transduced with both retrovirus backbones and at serial time points. In addition, we carried out PCR analysis of DNA extracted from sorted EGFP(-)/EYFP(-) cells and confirmed the presence of cells genetically marked by either vector in this population, indicating the persistence of cells that have downregulated or lost retroviral gene expression. In comparison to mouse studies, however, we did not detect substantial differences between MND and MoMuLV backbones.

Animals↗
PubMed1982

Transplantation of enzyme-treated teeth in the monkey Macaca fascicularis.

Homologous tooth transplants evoke immunogenic rejection against antigens within donor periodontal ligaments cells and ground substance. Eight maxillary incisors of 6 juvenile animals were extracted, crowns and pulps removed by gutta percha; the roots were incubated serially in enzyme solutions and glutaraldehyde at 37 degrees C for 5 h to destroy cells, glycoproteins and proteoglycans. Seven control teeth from the same 6 animals were incubated in saline at 37 degrees C for 15 min. All teeth were then speedily transplanted to other animals which were killed at 6 wk or 3 months. Radiographs and histologic sections revealed extensive resorption and inflammation of all control teeth. Enzyme-treated teeth showed less inflammation with resorption mostly on the surface of the cut coronal dentine. A loose fibrous attachment usually formed on most aspects of the root surface with deposition of new cellular cementum within the donor ligament. Inflammation, where present, may have resulted from incomplete removal of periodontal ligament cells, ground substance, or from salivary bacterial contamination.

Animals↗
PubMed1988

Distribution and characterization of neuropeptide Y in the brain of an elasmobranch fish.

Using a specific antiserum raised against synthetic neuropeptide Y (NPY), the distribution of immunoreactivity in the brain and pituitary of the elasmobranch fish Scyliorhinus canicula has been examined with the indirect fluorescence and the peroxidase-antiperoxidase methods. The highest density of NPY-immunoreactive neurons was found in the basal telencephalon and in the hypothalamus. Numerous NPY-containing perikarya were located in the entopeduncular and the preoptic nuclei, in the nucleus lobi lateralis and in the nucleus lateralis tuberis. NPY-immunopositive fibers were observed throughout the fish brain. In particular, dense networks of fibers were present in the entopeduncular and the habenular nuclei, in the nucleus tuberculi posterioris and in the lateral lobes. Scattered fibers were observed in all other parts of the brain except in the cerebellum where no NPY-immunoreactive material could be detected. A plexus of NPY-immunoreactive fibers arising from the preoptic neurosecretory complex appeared to run through the basal hypothalamus and the pituitary stalk. These fibers terminated in the intermediate lobe of the pituitary, suggesting that NPY may be involved in the control of melanotropin secretion. The NPY-immunoreactive material localized in the brain and pituitary was characterized by combining high-performance liquid chromatography (HPLC) analysis and radioimmunological detection. Brain and pituitary extracts showed a good cross-reactivity to the NPY antiserum, but serial dilutions of tissue samples did not completely parallel the standard curve. HPLC analysis resolved two major forms of immunoreactive NPY in the hypothalamus while the pars intermedia contained only authentic NPY. The widespread distribution of NPY neurons in the fish brain and pituitary suggests the involvement of NPY in a variety of physiological functions, including the neuroendocrine control of the pituitary.

Animals↗