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Biomedical subjects

R Stein

Publications and source records attributed to R Stein.

At least 145 records · Page 8Linked to original sources
PubMed1995

In vitro and in vivo reactivity of an internalizing antibody, RS7, with human breast cancer.

RS7, a murine IgG1 antibody raised against human lung carcinoma, possesses pancarcinoma reactivity. The antigen defined by this antibody is present in tumors of the lung, stomach, bladder, breast, ovary, uterus, and prostate. Efficient targeting and therapy by radiolabeled RS7 has been demonstrated previously in animals bearing Calu-3 (an adenocarcinoma of the lung) xenografts. In this study, the efficiency of tumor targeting and the efficacy of therapy of this antibody in nude mice bearing the MDA-MB-468 human breast carcinoma were evaluated. The tumor:nontumor ratios of RS7 were 1.9-2.1 times higher than those for Ag8 (the control antibody) on day 14, except for the heart. These values were similar to that of RS7 in the Calu-3 xenograft model. Radioimmunotherapy using 250 microCi of 131I-labeled RS7 in animals bearing approximately 0.1 cm3 tumors (approximately 10 days old) caused the disappearance of tumors in 6 of 10 animals at 2 weeks postinjection. Tumors eventually disappeared from all animals, and animals remained tumor-free until the termination of the study (11 weeks of duration), except for one animal that developed a transient reappearance of tumor. The tumors in animals that received an equal dose of 131I-labeled Ag8, or unlabeled RS7 or Ag8, either were unchanged or continued to grow. Treatment that used 275 microCi of 131I-labeled RS7 in animals carrying established tumors (1 month old, approximately 0.2-0.3 cm3) showed that this antibody is effective in controlling the growth of this tumor. Tumors in the treatment group began to disappear between the second and third weeks after the injection of the radiolabeled antibody. Seven of 10 animals remained tumor free at 15 weeks after the injection. Tumors in animals that received an equal dose of control antibody persisted but grew at a slower pace compared to the untreated group. No systemic toxicity was observed.

Adenocarcinoma↗
PubMed1995

Reduction of insulin gene transcription in HIT-T15 beta cells chronically exposed to a supraphysiologic glucose concentration is associated with loss of STF-1 transcription factor expression.

Chronic exposure of HIT-T15 beta cells to elevated glucose concentrations leads to decreased insulin gene transcription. The reduction in expression is accompanied by diminished binding of a glucose-sensitive transcription factor (termed GSTF) that interacts with two (A+T)-rich elements within the 5' flanking control region of the insulin gene. In this study we examined whether GSTF corresponds to the recently cloned insulin gene transcription factor STF-1, a homeodomain protein whose expression is restricted to the nucleus of endodermal cells of the duodenum and pancreas. We found that an affinity-purified antibody recognizing STF-1 supershifted the GSTF activator complex formed from HIT-T15 extracts. In addition, we demonstrated a reduction in STF-1 mRNA and protein levels that closely correlated with the change in GSTF binding in HIT-T15 cells chronically cultured under supraphysiologic glucose concentrations. The reduction in STF-1 expression in these cells could be accounted for by a change in the rate of STF-1 gene transcription, suggesting a posttranscriptional control mechanism. In support of this hypothesis, no STF-1 mRNA accumulated in HIT-T15 cells passaged in 11.1 mM glucose. The only RNA species detected was a 6.4-kb STF-1 RNA species that hybridized with 5' and 3' STF-1-specific cDNA probes. We suggest that the 6.4-kb RNA represents an STF-1 mRNA precursor and that splicing of this RNA is defective in these cells. Overall, this study suggests that reduced expression of a key transcriptional regulatory factor, STF-1, contributes to the decrease in insulin gene transcription in HIT-T15 cells chronically cultured in supraphysiologic glucose concentration.

3T3 Cells↗
PubMed1995

The epithelial/carcinoma antigen EGP-1, recognized by monoclonal antibody RS7-3G11, is phosphorylated on serine 303.

RS7-3G11 is a murine monoclonal antibody (MAb) raised against human non-small-cell lung carcinoma, and is under clinical evaluation. The epithelial/carcinoma antigen EGP-1, defined by RS7-3G11, was isolated and purified to homogeneity from a cervical carcinoma cell line, ME180. EGP-1 is a glycoprotein with an average molecular mass of 47.8 kDa. Metabolic labeling of the antigen with 32P-orthophosphate and subsequent immunoprecipitation with RS7-3G11 showed that it is a phosphoprotein. Phosphoamino acid analysis of the in vivo phosphorylated EGP-1 revealed that the phosphorylation is on serine. In vitro analysis with purified antigen demonstrated that protein kinase C, and not protein kinase A, is involved in phosphorylating the antigen in vitro. In vitro analysis indicated a stoichiometry of phosphorylation of 0.54 mole of phosphate per mole of EGP-1. Phosphoamino acid analysis and phosphopeptide mapping of the antigen phosphorylated in vitro by protein kinase C showed that phosphorylation occurred on a serine residue, specifically on serine 303, located in the cytoplasmic domain of EGP-1. Treatment of ME180 cells with phorbol ester increased the phosphorylation of EGP-1. The biological function of EGP-1 remains to be elucidated. In this report we elucidate an involvement of protein kinase C in phosphorylating EGP-1, which may signify a role for this antigen in signal transduction across the cell membrane.

Amino Acid Sequence↗
PubMed1995

A human phosphatidylinositol 3-kinase complex related to the yeast Vps34p-Vps15p protein sorting system.

Phosphoinositide (PI) 3-kinases have been characterized as enzymes involved in receptor signal transduction in mammalian cells and in a complex which mediates protein trafficking in yeast. PI 3-kinases linked to receptors with intrinsic or associated tyrosine kinase activity are heterodimeric proteins, consisting of p85 adaptor and p110 catalytic subunits, which can generate the 3-phosphorylated forms of phosphatidylinositol (PtdIns), PtdIns4P and PtdIns(4,5)P2 as potential second messengers. Yeast Vps34p kinase, however, has a substrate specificity restricted to PtdIns and is a PtdIns 3-kinase. Here the molecular characterization of a new human PtdIns 3-kinase with extensive sequence homology to Vps34p is described. PtdIns 3-kinase does not associate with p85 and phosphorylates PtdIns, but not PtdIns4P or PtdIns(4,5)P2. In vivo PtdIns 3-kinase is in a complex with a cellular protein of 150 kDa, as detected by immunoprecipitation from human cells. Protein sequence analysis and cDNA cloning show that this 150 kDa protein is highly homologous to Vps15p, a 160 kDa protein serine/threonine kinase associated with yeast Vps34p. These results suggest that the major components of the yeast Vps intracellular trafficking complex are conserved in humans.

Amino Acid Sequence↗
PubMed1995

Effects of radiolabeling monoclonal antibodies with a residualizing iodine radiolabel on the accretion of radioisotope in tumors.

The effect of using a "residualizing" iodine radiolabel, dilactitol-iodotyramine, for radioimmunolocalization of antibodies to tumors was investigated. This tracer is designed to be lysosomally trapped after catabolism of the labeled antibody. mAbs RS7 and RS11 were used for in vivo and in vitro studies on the uptake and retention of radioisotope into tumor cells. Both are murine IgG1 mAbs with pancarcinoma reactivity, which react with integral membrane glycoproteins. mAb RS7 has been shown to be relatively rapidly catabolized by the antigen-bearing cell line Calu-3, whereas RS11 is catabolized more slowly in the same cells. An 111In- or 88Y-p-isothiocyanatobenzyl-diethylenetriamine pentaacetic acid conjugate was also tested because these radiometals are known to be lysosomally trapped, and iodination via chloramine T was used to provide a baseline. In vitro, a substantial increase in retention of the label by cells was observed when the dilactitol-tyramine DLT- or 111In-labeled mAbs were used, and the improvement gained by the use of these residualizing labels was greater with the use of the rapidly catabolized mAb (RS7) than it was with the more slowly catabolized mAb (RS11). In biodistribution studies in nude mice bearing Calu-3 tumor xenografts, a dramatic improvement in the tumor accretion of the radiolabel was seen with the use of the 131I-labeled DLT- or 88Y-labeled mAbs. For example, at day 7 the percentage of injected dose/g in the tumor was 5.54 +/- 1.47% (SD), 38.06 +/- 8.04%, and 43.18 +/- 19.50% for the conventionally iodinated, DLT- and 88Y-labeled RS7, respectively. Dosimetry calculations performed on the biodistribution data predict increases of approximately 8- and 4-fold in the absorbed dose to tumor with the use of 131I-labeled DLT- and 90Y-labeled mAbs, respectively, compared to the conventional 131I. In contrast to in vitro findings, these results were similar for both RS7 and RS11, suggesting that the use of DLT may be advantageous for most of the mAbs binding to the cell surface, including antibodies that are catabolized relatively slowly. The advantage of 131I-labeled DLT over 90Y is due to the longer physical half-life of the 131I.

Adenocarcinoma↗
PubMed1995

Pirenzepine-sensitive component of forelimb vascular resistance and heart rate in cats.

In vagotomized, midcollicular decerebrate, non-anesthetized cats, in which the preganglionic input to the stellate ganglion was intact and the right forelimb was autoperfused at constant flow, the muscarinic receptor antagonist pirenzepine (PZP, 50 micrograms/kg i.v.), which does not cross the blood-brain barrier, produced a decrease in forelimb perfusion pressure (FLPP), heart rate (HR) and systemic arterial pressure (SAP). Administration of the nicotinic receptor antagonist hexamethonium (30 mg/kg i.v.) after PZP produced a further, larger drop in FLPP, HR and SAP. The dose of PZP used blocked the increase in FLPP and HR evoked by the muscarinic receptor agonist McN-A-343, but not the increase evoked by the nicotinic receptor agonist DMPP, when these drugs were injected into the arterial supply of the ganglion. Following administration of phentolamine and propranolol (2 mg/kg i.v. of each) which caused bradycardia and forelimb vasodilation, PZP had no effect on FLPP and HR. This finding suggests that PZP decreases sympathetic tone in resistance vessels and in the heart. Pirenzepine did not depress the forelimb vasoconstriction and the cardioacceleration evoked by electrical stimulation of the postganglionic vertebral and inferior cardiac nerves, respectively, suggesting that PZP does not act at the neuro-effector junctions. On the other hand, PZP blocked the forelimb vasoconstriction and the cardioacceleration produced by stimulating the preganglionic input of the stellate ganglion in presence of hexamethonium (30 mg/kg i.v.), indicating the stellate ganglion as the likely site of action of the drug. These findings suggest that the ganglion cell firing that underlies the sympathetic tone of cardiovascular effector cells is generated, in part, by a muscarinic, PZP-sensitive, synaptic mechanism.

Animals↗
PubMed1995

Apoptosis induced by serum deprivation of PC12 cells is not preceded by growth arrest and can occur at each phase of the cell cycle.

Previous studies have shown that PC12 cells undergo apoptosis (programmed cell death) when deprived of serum. In the present study, we examined the relationship of this death process to the cell cycle. PC12 cell populations synchronized at different, specific phases of the cell cycle exhibit similar kinetics of cell death following deprivation of serum. Flow cytometry analysis was used to examine the levels of apoptotic death in these cell populations in relationship to their progression in the cell cycle during the course of serum deprivation. Such analysis revealed that the cells die during the G0-G1, S, and perhaps G2-M phases and at the G2 to G1 transition. These results, therefore, suggest that the death of synchronized, serum-deprived PC12 cells occurs throughout the cell cycle and is not dependent on growth arrest. Flow cytometry methodology (acridine orange staining), which determines the RNA content of cells in relationship to their position in the cell cycle, was used to address these questions in nonsynchronized cells. These experiments revealed that, upon serum deprivation, an immediate loss of RNA occurred from cells in G1, S, and G2-M phases. This loss is accompanied by a slower appearance of cells with degraded DNA content. These results show that cells from all phases of the cell cycle are damaged upon serum deprivation and thus suggest that the apoptotic cell death of nonsynchronized PC12 cells may occur from each phase of the cell cycle.

Animals↗
PubMed1995

NGF-dependent neurotrophic-like effects of AF102B, an M1 muscarinic agonist, in PC12M1 cells.

The non-selective muscarinic agonist oxotremorine induces atropine-sensitive neurite outgrowth in PC12 cells stably transfected with m1 muscarinic receptors. In contrast, AF102B, an M1-selective muscarinic agonist, mediated minimal neurite outgrowth in these cells. In the presence of nerve growth factor (NGF) however, it induced atropine-sensitive neurite outgrowth in almost half the cell population. AF102B mediated phosphoinositide hydrolysis, but unlike carbachol, it did not stimulate cyclic AMP accumulation in these cells. These signals were not affected by NGF, indicating that they were not directly responsible for the cholinergic neurotrophic-like response. Our observations suggest that AF102B may improve neuronal responsiveness to neurotrophic factors, and thus may provide another beneficial aspect for treating Alzheimer's disease.

Animals↗
PubMed1995

Cocaine-induced cerebrovascular impairment: challenges to neuropsychological assessment.

The patient who presents for evaluation and/or subsequent treatment of a neurological or psychiatric complaint accompanied by a positive history of substance abuse has generally received only modest attention in the clinical practice literature. Significantly more clinical attention has focused on the neurobehavioral sequelae of more pronounced brain insults, despite the rapid emergence of literature detailing psychopharmacologic-induced changes in brain-behavior functioning. This article describes recent clinical research findings related to the neuropsychology of cocaine use and associated issues of neurobiology and psychopharmacology. A description of strategies that have proven effective for assessing this population will be discussed. An emphasis on neurocognitive impairment that may precede as well as occur consequent to cocaine use are also examined. The literature reviewed here generally supports the conclusion that subgroups of cocaine abusing patients may demonstrate sustained brain perfusion anomalies and persistent neurocognitive deficits.

Cerebrovascular Disorders↗
PubMed1995

The health belief model and adherence with a community center-based, supervised coronary heart disease exercise program.

Some investigators have concluded that health beliefs do not influence the maintenance of coronary heart disease (CHD) exercise adherence. However, the beliefs tested have not been specific to CHD nor exercise. In addition, much of the research has been atheoretical. We conducted a retrospective study to explore the possible utility of the Health Belief Model (HBM) for explaining attendance at a supervised CHD exercise program, based in a community center. Two dimensions of the model, general health motivation and perceived severity of CHD, were associated with attendance in the theoretically predicted direction, while a third dimension, perceived benefits of exercise, was associated in a direction opposite that predicted by the model. The model as a whole accounted for 29% (adjusted R2) of the variance. This study provides some initial evidence that health beliefs are associated with CHD exercise adherence.

Attitude to Health↗
PubMed1995

New insights into proteasome function: from archaebacteria to drug development.

The proteasome is not simply a 'garbage disposal unit' but also has functions in the control of the cell cycle and immune responses. The structure of an archaebacterial proteasome has recently been determined to high resolution, and provides insight into the unusual mechanism of proteolytic cleavage by the proteasome.

Archaea↗
PubMed1995

Inhibition of drug-induced apoptosis by survival factors in PC12 cells.

Pheochromocytoma (PC12) cells have been shown to undergo apoptosis (programmed cell death) when deprived of serum and to be rescued by nerve growth factor, fibroblast growth factor, dibutyryl cyclic AMP, aurintricarboxylic acid, or exogenous expression of bcl-2. We show here that the cytotoxic drugs cycloheximide, actinomycin D, colchicine, and EGTA also induce apoptosis in PC12 cells. These findings prompted us to investigate whether apoptosis induced by these drugs involves similar pathways in each case, and whether the factors preventing the apoptotic death of serum-deprived PC12 cells can also protect the cells from apoptosis induced by the cytotoxic drugs. Nerve growth factor, dibutyryl cyclic AMP, and expression of bcl-2 inhibited apoptosis induced by all four cytotoxic drugs. Fibroblast growth factor inhibited apoptosis induced by EGTA or colchicine. Aurintricarboxylic acid inhibited apoptosis induced by EGTA. These results suggest that apoptosis induced by treatments with the various drugs is mediated by different initiating pathways, all of which converge into a final, common pathway. Nerve growth factor, dibutyryl cyclic AMP, and bcl-2 appear to affect the final common pathway, whereas fibroblast growth factor and aurincarboxylic acid appear to be more specific and affect only some of the pathways.

Animals↗
PubMed1995

Activation of muscarinic receptors inhibits apoptosis in PC12M1 cells.

Previous studies have shown that PC12 cells depend on growth factors for their survival. When deprived of growth factors, the cells undergo a dying process termed "apoptosis" (programed cell death). We show here that muscarinic agonists inhibited the apoptotic death of growth factor-deprived PC12M1 cells (PC12 cells stably expressing cloned m1 muscarinic acetylcholine receptors). This protective effect of the muscarinic agonists was observed in both proliferating and neuronal PC12M1 cells, was blocked by the muscarinic antagonist atropine, and was not observed in PC12 cells lacking m1 receptors. Muscarinic receptors therefore mediate inhibition of apoptosis in these cells. In addition to its effect on survival, the muscarinic agonist oxotremorine induced inhibition of DNA synthesis as well as growth arrest of exponentially growing PC12M1 cells at the S and G2/M phases of the cell cycle. Muscarinic receptors in these cells may therefore mediate inhibition of cell cycle progression.

Animals↗
PubMed1995

Electromyographic comparison of obturator nerve block to three-in-one block.

Obturator nerve block during spinal, epidural, or general anesthesia without muscle relaxants has been recommended for transurethral surgery to prevent thigh adductor muscle contractions during operative electrocautery. We investigated the effectiveness of direct obturator and 3-in-1 nerve motor blocks in 44 patients undergoing transurethral surgery during spinal anesthesia with isobaric bupivacaine. Patients were randomly assigned to receive 3-in-1 block with 40 mL (n = 13) or 50 mL (n = 11) of 1.5% lidocaine plus epinephrine, or direct obturator nerve block with 10 mL of 2% lidocaine plus epinephrine (n = 20). After both direct obturator and 3-in-1 blocks, compound muscle action potential (CMAP) testing of the obturator nerve was performed at 1-10-s intervals for 10 min. In patients given direct obturator nerve block (n = 20), CMAP amplitude decreased by 88.8 +/- 21% (mean +/- SD) from baseline. In contrast, 3-in-1 block reduced the evoked CMAP amplitude by 7.4 +/- 19% (P < 0.05). Peak lidocaine plasma levels of 1.6 +/- 0.2 micrograms/mL (range 1.0-2.8 micrograms/mL) were reached 60-90 min after the block in those patients receiving 50 mL of local anesthetic. The 3-in-1 technique fails to predictably result in effective motor block of the obturator nerve and thus may not prevent inadvertent thigh adductor muscle contractions during transurethral surgery. A direct approach to the obturator nerve is significantly more effective in producing motor block, and even when given in larger than recommended dosages it results in subtoxic peak plasma lidocaine concentrations.

Action Potentials↗
PubMed1995

c-jun inhibits insulin control element-mediated transcription by affecting the transactivation potential of the E2A gene products.

Pancreatic beta-cell-type-specific transcription of the insulin gene is principally controlled by trans-acting factors which influence insulin control element (ICE)-mediated expression. The ICE activator is composed, in part, of the basic helix-loop-helix proteins E12, E47, and E2-5 encoded by the E2A gene. Previous experiments showed that ICE activation in beta cells was repressed in vivo by the c-jun proto-oncogene (E. Henderson and R. Stein, Mol. Cell. Biol. 14:655-662, 1994). Here we focus on the mechanism by which c-Jun inhibits ICE-mediated activation. c-Jun was shown to specifically repress the transactivation potential of the E2A proteins. Thus, we found that the activity of GAL4:E2A fusion constructs was inhibited by c-Jun. The transrepression capabilities of c-Jun were detected only in pancreatic islet cell lines that contained a functional ICE activator. Repression of GAL4:E2A was mediated by the basic leucine zipper regions of c-Jun, which are also the essential regions of this protein necessary for controlling ICE activator-stimulated expression in vivo. The specific target of c-Jun repression was the transactivation domain (located between amino acids 345 and 408 in E12 and E47) conserved in E12, E47, and E2-5. In contrast, the activation domain unique to the E12 and E47 proteins (located between amino acids 1 and 99) was unresponsive to c-Jun. Our results indicate that c-Jun inhibits insulin gene transcription in beta cells by reducing the transactivation potential of the E2A proteins present in the ICE activator complex.

Adenoviridae↗
PubMed1995

Cardiac disease and nonorganic chest pain: factors leading to disability.

Research has shown that many chest pain patients, without coronary artery disease, may suffer from panic disorder, hypochondriasis, depression, and/or multiple phobias. Some patients with coronary artery disease may also suffer from these disorders and are often unable to return to previous activity. In spite of good prognosis for longevity and acceptable exercise test results, a large proportion of these patients continue to be disabled by chest pain and/or chronic cardiac fears and demand constant medical attention. This study examined the psychiatric and behavioral symptomatology that differentiated four groups of patients experiencing chest pain: the able (active/working patient) with and without coronary artery disease, as determined by exercise thallium-201 studies, and the disabled (inactive/nonworking patient) with and without coronary artery disease. The results of the study indicated that the inactive patients, both with and without heart disease, suffered from a host of debilitating psychiatric conditions.

Aged↗