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Biomedical subjects

J Stein

Publications and source records attributed to J Stein.

At least 253 records · Page 14Linked to original sources
PubMed1994

Cytomegalovirus infection in transplant recipients. The role of tumor necrosis factor.

Human cytomegalovirus (CMV) infection is an important cause of morbidity and mortality in transplant recipients. CMV infection commonly results from the reactivation of a latent infection. Using a set of monoclonal anti-CMV antibodies, we found CMV antigen expression in peripheral blood mononuclear cells (PBMNC), particularly in monocytes, in 312 of 816 samples from 190 allograft recipients. The detection of CMV-IE antigens and CMV-IE DNA in PBMNC indicates that positive cells may represent truly infected cells. The relation between increased cytokine plasma levels (particularly following treatment by pan-T cell antibodies) and the appearance of CMV antigens in PBMNC suggests that cytokines may play an important role in the reversal of CMV latency. This hypothesis is supported by our finding that tumor necrosis factor-alpha (TNF) is able to stimulate the activity of the CMV-IE enhancer/promoter region in the human monocytic cell line, HL-60. The interleukins 1, 2, 3, 4, 6, 8 and 10; transforming growth factor-beta; interferongamma; and granulocyte/macrophage colony-stimulating factor did not show any enhancing effect on the CMV promoter activity. Thus, TNF-alpha seems to play a key role in regulating the balance between latency and reactivation of CMV infection. Inhibition of TNF-alpha release or action may be an alternative strategy for preventing CMV-associated morbidity in allograft recipients.

Adult↗
PubMed1994

DNase I hypersensitive sites in promoter elements associated with basal and vitamin D dependent transcription of the bone-specific osteocalcin gene.

Nuclease hypersensitive sites were mapped in the proximal promoter of the osteocalcin gene, which is expressed only in bone cells exhibiting the mature osteoblast phenotype. Nuclei from proliferating and confluent rat osteosarcoma (ROS) 17/2.8 cells were subjected to DNase I digestion, and hypersensitivity was assayed by the indirect end-labeling method, using osteocalcin gene probes. Hypersensitive sites were detected in two promoter domains: -590 to -390, which spans the vitamin D responsive element, and -170 to -70, which spans the TATA box and the CCAAT-containing OC box domain. Together, these elements regulate basal and vitamin D enhanced osteocalcin gene transcription. We observed a parallel relationship between the intensity of bands representing the hypersensitive sites and the extent to which the osteocalcin gene is transcribed. Both in confluent cultures and in response to vitamin D, when osteocalcin transcription was upregulated, the hypersensitive bands were significantly intensified. Additionally, the bands were decreased under conditions that downregulate osteocalcin gene transcription. A functional relationship between the presence of hypersensitive sites and osteocalcin gene transcription is further supported by the absence of hypersensitivity in nonosseous cells that do not express osteocalcin, although these proliferating cells exhibited hypersensitivity in a cell cycle regulated histone gene promoter. Our results suggest the involvement of chromatin structure in transcriptional responsiveness of the osteocalcin gene to physiologic modulation.

Animals↗
PubMed1994

Interferon alpha selectively affects expression of the human myeloid cell nuclear differentiation antigen in late stage cells in the monocytic but not the granulocytic lineage.

The human myeloid cell nuclear differentiation antigen (MNDA) is expressed constitutively in cells of the myeloid lineage, appearing in myeloblast cells in some cases of acute myeloid leukemia and consistently being detected in promyelocyte stage cells as well as in all later stage cells including peripheral blood monocytes and granulocytes. The human myeloid leukemia cell lines, HL-60, U937, and THP-1, express similar levels of immunochemically detectable MNDA. Although, the level of MNDA mRNA in primary monocytes is very low it was up-regulated at 6 h following the addition of interferon alpha. The effect of interferon alpha on the MNDA mRNA is also observed in the cell lines HL-60, U937, and THP-1. The MNDA mRNA level in primary granulocytes was unaffected by addition of interferon alpha and other agents including interferon gamma, endotoxin, poly (I).poly (C), and FMLP. The MNDA mRNA level in the myeloid cell lines was also unaffected by the latter four agents. Induction of differentiation in the myeloid cell lines with phorbol ester induces monocyte differentiation which was accompanied by a decrease in MNDA mRNA level. This reduced level of mRNA could then be elevated with subsequent interferon alpha treatment. The effects of phorbol ester on MNDA mRNA appeared to be associated with induced differentiation since inhibiting cell proliferation did not alter the level of MNDA mRNA and cell cycle variation in MNDA mRNA levels were not observed. The ability of interferon alpha to up-regulate MNDA mRNA in phorbol ester treated myeloid cell lines is consistent with the observations made in primary monocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, Differentiation, Myelomonocytic↗
PubMed1994

Expression of cell growth and bone phenotypic genes during the cell cycle of normal diploid osteoblasts and osteosarcoma cells.

Establishing regulatory mechanisms that mediate proliferation of osteoblasts while restricting expression of genes associated with mature bone cell phenotypic properties to post-proliferative cells is fundamental to understanding skeletal development. To gain insight into relationships between growth control and the developmental expression of genes during osteoblast differentiation, we have examined expression of three classes of genes during the cell cycle of normal diploid rat calvarial-derived osteoblasts and rat osteosarcoma cells (ROS 17/2.8): cell cycle and growth-related genes (e.g., histone), genes that encode major structural proteins (e.g., actin and vimentin), and genes related to the biosynthesis, organization, and mineralization of the bone extracellular matrix (e.g., alkaline phosphatase, collagen I, osteocalcin, and osteopontin). In normal diploid osteoblasts as well as in osteosarcoma cells we found that histone genes, required for cell progression, are selectively expressed during S phase. All other genes studied were constitutively expressed both at the transcriptional and posttranscriptional levels. Alkaline phosphatase, an integral membrane protein in both osteoblasts and osteosarcoma cells, exhibited only minimal changes in activity during the osteoblast and osteosarcoma cell cycles. Our findings clearly indicate that despite the loss of normal proliferation-differentiation interrelationships in osteosarcoma cells, cell cycle regulation or constitutive expression of growth and phenotypic genes is maintained.

Actins↗
PubMed1994

Acquisition of Borrelia burgdorferi by Ixodes ricinus ticks fed on the European hedgehog, Erinaceus europaeus L.

A hedgehog, Erinaceus europaeus, was found to be heavily infested with larval and nymphal Ixodes ricinus in a forest park in Co. Galway, Ireland. A large proportion of the ticks that engorged and detached were infected with the spirochaete, Borrelia burgdorferi, the causative agent of human Lyme borreliosis. The identity of these spirochaetes was confirmed by immunofluorescent assay with B. burgdorferi-specific monoclonal antibody and by polymerase chain reaction test and they were transmitted from the hedgehog to laboratory-reared ticks and from the ticks obtained from the hedgehog to gerbils (Meriones unguiculatus). The high infection rate of the larvae that fed on the hedgehog in comparison with unfed larvae from the same habitat was interpreted as strong evidence that this host species is reservoir competent. Since hedgehogs can evidently feed adult ticks as well as many immature stages, they may well have an important role in the ecology of Lyme borreliosis in some habitats.

Animals↗
PubMed1994

Simultaneous preparation of rabbit intestinal brush border and basolateral membrane vesicles.

A highly reproducible method is described for simultaneous preparation of the rabbit intestinal BBMV and BLMV. When BLMV were purified on continuous sorbitol gradient the final enrichment was over ten-fold for both Na+, K(+)-ATPase and K(+)-activated phosphatase. BBMV were separated using repeated precipitation with magnesium and EGTA. Marker enzymes of the brush border membrane also showed over ten-fold enrichment. There was no contamination with other cellular organelles. The characteristics of the preparations were checked by SDS gel electrophoresis, which showed distinct protein patterns for both membranes; monoclonal antibodies for villin reacted with the BBMV but not with BLMV, showing the absence of contamination of the BLMV fraction. The quality of the membranes, assessed by D-glucose transport, showed good functional properties for both BBMV and BLMV. Orientation of the vesicles was checked by permeabilizing with SDS buffered with BSA and showed that BLMV are oriented inside out and BBMV right side out. The model simulates normal transepithelial intestinal transport at the level of membrane vesicles and could be applied in transport studies in physiology and pharmacology.

Animals↗
PubMed1994

Effects of prostaglandins and indomethacin on gastric emptying in the rat.

Prostaglandins have been shown to stimulate contraction of gastrointestinal smooth muscle in vitro, and their synthesis has been reported in different gastrointestinal diseases. The aim of this study was to determine the different effects of PGs and indomethacin on gastric emptying in the rat in vivo, and to investigate their possible role as modulators of osmolarity induced changes in gastric emptying. Rats were fed via gastric tube with suspensions of various osmolarity containing different prostaglandins or indomethacin. Gastric emptying, measured by the quantity of food remaining in the stomach, was studied by a noninvasive technique using orally administered nonabsorbable marker 141Cerium. PGE2 given orally and its synthetic analogue 16-16-dimethyl PGE2 given subcutaneously caused significant delay of gastric emptying, while PGF2 alpha increased gastric emptying regardless of the administration route. Tube-feeding with high osmolarity suspension significantly reduced emptying rate of 141Cerium compared to feeding with low osmolarity suspension. Gastric emptying rate of the tracer was significantly higher in rats fed with low osmolarity test meal, in comparison to those fed with high osmolarity test meal. This effect was completely antagonized by indomethacin. Due to the known inhibitory effect of indomethacin on cyclooxygenase, the effects observed in the present study are also presumably mediated by the action of prostaglandins.

Animals↗
PubMed1994

Dynamic X-ray compensation for conformal radiotherapy by means of multi-leaf collimation.

The application of a multiple fixed field technique employing individually shaped and intensity-modulated beams makes it possible to produce dose distributions of high conformity even in the case of concave target volumes. With the technique presented here arbitrary intensity-modulated beams for the practical solution of the inverse problem can be generated. It is also possible to omit wedges, blocks and compensators in conventional radiotherapy. A continuous unidirectional sweep of independently computer-controlled leaves of a multi-leaf collimator is used to modulate the primary uniform beam. A new algorithm is introduced that computes the leaf trajectories. Also, a method is presented that accounts for leaf penumbra and transmission, which causes the generated fluence distribution to deviate from the desired fluence distribution. An optimization algorithm minimizing this deviation is described. The algorithm calculating the leaf trajectories, as well as the method considering penumbra and transmission and the successive optimization technique are used to calculate examples. Treatment times are calculated and compared to those needed when using compensators. A relationship between the treatment time and the maximum leaf speed is also deduced. To achieve good performance the maximum leaf speed should be no less than 20 mm/s.

Algorithms↗
PubMed1994

Rapid postabsorptive metabolism of nicotinic acid in rat small intestine may affect transport by metabolic trapping.

Postabsorptive metabolism of [3H]nicotinic acid by rat proximal jejunum was studied in vitro using the everted sac technique and in vivo by applying ligated loops. Metabolites were analyzed by HPLC with radiochemical detection. Within 60 min of incubation nicotinic acid was almost completely metabolized in vivo. Only 3.2% of the label could be detected in the original substrate nicotinic acid, whereas > 90% of the radioactivity present in the gut wall was nicotinamide. Further in vitro experiments revealed that conversion was rapid; after 10 min of incubation > 40% of the substrate was converted to other metabolites. From analysis of the serosal fluid, it was evident that only nicotinic acid and nicotinamide were transferred to the serosal compartment. Rapid conversion of nicotinic acid might affect the transport step itself by metabolic trapping, resulting in the maintenance of a substrate gradient necessary for passive absorption. In contrast, as examined by gradient centrifugation, the substrate in subcellular organelles is transferred only to a minor extent.

Animals↗
PubMed1994

'Alarming but very necessary': working with staff groups around the sexual abuse of adults with learning disabilities.

This study focused on the knowledge of front line staff, as individuals and in groups, in relation to the sexual abuse of adults with learning disabilities and was designed to inform staff training and policy initiatives and to explore gender dynamics. It showed that front line staff were alert to the possibility of sexual abuse but unclear about their roles and responsibilities making this an area of great concern to them. One third knew of people who had been abused but these had been dealt with haphazardly, leaving front line staff holding disproportionate anxiety on behalf of the service as a whole.

Adult↗
PubMed1994

What children see affects how they spell.

The authors predicted that children who have experienced frequent visual confusion of text during their development because of unstable binocular control may be prevented from discovering the rules and patterns of English orthography. The authors compared the spelling errors made by children who had unstable binocular vision with those of children who had stable binocular vision and the same spelling ability. The subjects were drawn from a clinical population of children referred because of suspected reading difficulties; though they were of mixed abilities, they were generally poor spellers and readers. Children with unstable binocular control made spelling errors which were more phonologically plausible than those made by the control group with normal binocular vision.

Child↗
PubMed1994

Tyrphostins inhibit secretagogue-induced 1,4,5-IP3 production and amylase release in pancreatic acini.

We examined the role of protein tyrosine kinase inhibitors (tyrphostins) in secretagogue-induced inositol 1,4,5-trisphosphate (1,4,5-IP3) production and amylase secretion in rat pancreatic acinar cells. The data show that various specific cell-permeant tyrphostins (methyl 2,5-dihydroxycinnamate, tyrphostin 25, and genistein) inhibited the cholecystokinin octapeptide-, carbachol-, and bombesin-induced 1,4,5-IP3 production and amylase release. In digitonin-permeabilized cells, tyrphostins decreased 1,4,5-IP3 accumulation and amylase release generated by directly stimulating G proteins with the weakly hydrolyzable GTP analogue guanosine 5'-O-(3-thiotriphosphate). Tyrphostins had no effect on vasoactive intestinal peptide-induced amylase secretion. In isolated pancreatic acinar membranes, cholecystokinin octapeptide caused a rapid increase in tyrosine phosphorylation of a synthetic peptide containing the 12-amino acid sequence around a tyrosine phosphorylation site in pp6osrc. These results provide evidence that tyrosine kinases are involved in the activation of phospholipase C by G protein-coupled receptors in pancreatic acinar cells.

Amylases↗
PubMed1994

Effect of a Rab3A effector domain-related peptide, CCK, and EGF in permeabilized pancreatic acini.

We report here that a synthetic peptide of the effector domain of the small-molecular-weight GTP-binding protein Rab3A (EDRab3AL) is a potent stimulator of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] production and amylase secretion in digitonin-permeabilized pancreatic acini. Moreover, the Rab3A effector domain peptide caused phosphatidylinositol 4,5-bisphosphate breakdown, indicating that the observed increase in Ins(1,4,5)P3 is due to stimulation of a phosphoinositide-specific phospholipase C (PLC). The dose-response curve for EDRab3AL-induced amylase release was biphasic, showing a maximum at 0.3 nM EDRab3AL and a decline at higher peptide concentrations. By contrast, the dose-response curve for EDRab3AL-induced Ins(1,4,5)P3 production was monophasic, showing stimulation with increasing EDRab3AL concentrations. A peptide of the effector domain of Rab1A, EDRab1AL, had no effect, indicating that the response to EDRab3AL is specific. Cholecystokinin octapeptide (CCK-8) and EDRab3AL had additive effects on the acinar Ins(1,4,5)P3 level. Epidermal growth factor (EGF), which has recently been shown to inhibit CCK-8-induced Ins(1,4,5)P3 production in pancreatic acinar cells, also decreased EDRab3AL-induced Ins(1,4,5)P3 production. These results suggest that EDRab3AL and CCK-8 act on the same EGF-inhibitable PLC by independent mechanisms. CCK-8 increased and EGF decreased amylase release in response to submaximal EDRab3AL concentrations. By contrast, at supramaximal EDRab3AL concentrations EGF increased and CCK-8 decreased EDRab3AL-stimulated amylase release. EDRab3AL had no effect in intact acini, indicating that the site of action of EDRab3AL is intracellular. We conclude that EDRab3AL regulates phosphoinositide-specific PLC activity and thereby amylase secretion in an analogous fashion to CCK-8, but from within the cell.(ABSTRACT TRUNCATED AT 250 WORDS)

Amylases↗
PubMed1994

Parathyroid-responsive modifications in the nuclear matrix of ROS 17/2.8 rat osteosarcoma cells.

PTH is a mediator of skeletal development and remodeling that influences gene expression in osteoblastic cells. It is well established that PTH modulates the activity of membrane-associated second messenger signal transduction pathways. In these studies we have addressed the potential contribution of components of cell structure to the integration of PTH-related regulatory signals that influence the expression of bone cell genes. Chronic treatment of ROS 17/2.8 rat osteosarcoma cells with PTH is accompanied by changes in gene expression that are at least in part transcriptionally controlled. To explore the involvement of nuclear architecture in PTH-responsive modifications in gene expression, we investigated changes in the nuclear matrix after PTH treatment. Consistent with a role for the nuclear matrix in determining spatial organization and topology of chromatin as well as in the localization and targeting of transcription factors, we observed PTH-associated changes in a 200-kilodalton nuclear matrix protein in response to PTH. A significant down-regulation of synthesis was observed when nuclear matrix proteins were resolved electrophoretically in two-dimensional gels. This protein was restricted to the nuclear matrix and was not detected in the chromatin or cytoskeletal cellular fractions. These alterations in nuclear matrix proteins that occur after PTH treatment in osteosarcoma cells were phenotype related. They did not occur in UMR-106 POL or H4 hepatoma cells. Our findings support a role for the nuclear matrix in transducing PTH-mediated regulatory signals to facilitate the extent to which genes in osteoblasts are transcribed.

Animals↗