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Biomedical subjects

J A Miller

Publications and source records attributed to J A Miller.

At least 109 records · Page 6Linked to original sources

The enhancement of muscimol-stimulated 36C1 influx by the antispastic 5-aryl-3-(alkylsulfonyl)-4H-1,2,4-triazole (MDL 27,531) in rat brain membrane vesicles.

The antispastic triazole, 4-methyl-3-methylsulphonyl-5-phenyl-4H-1,2,4-triazole (MDL 27,531) was tested in glycine- and muscimol-stimulated 36Cl- influx into brain membrane preparations. MDL 27,531 (100 nM) had no effect on glycine- (100 nM-400 microM) stimulated 36Cl- influx in brain stem tissue; on the other hand, MDL 27,531 (10 nM-10 microM) enhanced muscimol- (1 microM) stimulated 36Cl- influx in cerebellar but not cortical membranes. In the presence of the benzodiazepine (BZD) antagonist, flumazenil (10 microM), MDL 27,531 inhibited muscimol-stimulated flux. These data suggest a novel interaction of this triazole with subtypes of the gamma-aminobutyric acid (GABA)A receptor complex and the BZD receptor.

Animals↗

Chemoprotective properties of chlorophyllin against vinyl carbamate, p-nitrophenyl vinyl ether and their electrophilic epoxides.

Chlorophyllin (CHL), a water-soluble sodium and copper derivative of chlorophyll, has been shown to be a strong antimutagen in several test systems, but its mechanism of antimutagenic action is largely unknown. In the present study, we have found the protective properties of CHL against vinyl carbamate, p-nitrophenyl vinyl ether and their electrophilic epoxides. CHL exhibited dose-related inhibition of his+ reversion in Salmonella typhimurium TA 1535 induced by these mutagens. Formation of DNA adducts from vinyl carbamate epoxide (VCO) and 2'-(4-nitrophenoxy)oxirane (NPO) was also markedly attenuated in the presence of CHL. Oral administration of CHL prior to the topical application of each of the above carcinogens resulted in significant reduction in both incidence and multiplicity of skin tumors in mice. The effective protection by CHL against VCO and NPO suggest that its formation of inactive complexes with these carcinogens is mediated by mechanisms other than pi-pi interactions.

Animals↗

Purification and identification of brain-derived neurotrophic factor from human serum.

Brain-derived neurotrophic factor (BDNF), a 27-kDa noncovalently linked homodimer with subunits of approximately 13.5 kDa as viewed by SDS-PAGE, is thought to be primarily produced in the central nervous system. We report here the isolation of BDNF from pooled normal human sera, using a two-step purification process followed by SDS-PAGE, transfer to a polyvinylidene difluoride membrane, and subsequent identification of the protein by sequence analysis of the appropriate band(s) from the membrane. The level of BDNF in pooled human sera was estimated to be approximately 15 ng/ml as determined by an enzyme-linked immunosorbant assay. The average for six individuals was 18.9 +/- 5.7 ng/ml. There is an approximately 200-fold increase in the levels of BDNF in serum relative to plasma. Results from experiments using differential centrifugation suggest that the source of this increase is due to release from platelets. The presence of high levels of BDNF in serum suggests a role for this neurotrophin either in nerve repair at sites of injured tissue or in nonneuronal functions.

Amino Acid Sequence↗

Apoptotic DNA fragmentation in the rat cerebral cortex induced by permanent middle cerebral artery occlusion.

Recent investigations have demonstrated internucleosomal DNA fragmentation in ischemic neuronal tissue. This type of fragmentation is characteristic of programmed cell death or apoptosis and suggests that neuronal death in stroke may be more complex than simple necrotic death. The present experiments provide a detailed examination of the regional localization and time course for apoptotic DNA fragmentation in the cerebral cortex following focal cerebral ischemia. Spontaneously hypertensive rats were subjected to permanent right middle cerebral artery occlusion and the cerebral cortices were examined for evidence of DNA fragmentation using electrophoretic, flow cytometric, and histological approaches. An electrophoretic examination of cortical DNA at 24 h after the occlusion indicated that the majority of nucleosomal ladders were in the transition zone or penumbra and the core of the infarction, with no fragmentation apparent in the contralateral normal cortex. A flow cytometric analysis of DNA fragmentation in intact cells revealed a similar pattern, with increased fragmentation observed in ischemic cortex vs. the contralateral cortex. Saggital sections taken 1.5 mm lateral to midline were collected from animals at 1, 4, and 24 h after the infarction and DNA fragmentation was examined histologically by terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL) staining. Quantitative analysis of these sections indicated that DNA fragmentation can be observed in the anterior and central area of the infarctions as soon as 1 h after the occlusion and that the extent and magnitude of the fragmentation increases at 4 and 24 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Chemoprotective effects of capsaicin and diallyl sulfide against mutagenesis or tumorigenesis by vinyl carbamate and N-nitrosodimethylamine.

Capsaicin (trans-8-methyl-N-vanillyl-6-nonenamide) is a major pungent and irritating ingredient of hot chilli peppers, which are frequently consumed as spices. This dietary phytochemical has been found to interact with microsomal xenobiotic metabolizing enzymes in rodents. Capsaicin and its saturated analog dihydrocapsaicin (trans-8-methyl-N-vanillyl-6-nonanamide) have been proposed to inactivate cytochrome P-450 HE1 by irreversibly binding to the active sites of the enzyme. Besides cytochrome P-450 HE1, other isoforms of the P-450 superfamily were also reported to be inhibited by capsaicin. The inhibition by capsaicin of microsomal monooxygenases involved in carcinogen activation implies its chemopreventive potential. As part of a program to investigate chemoprotective properties of capsaicin we initially determined the effect of capsaicin on vinyl carbamate (VC)- and N-nitrosodimethylamine (NDMA)-induced mutagenesis in Salmonella typhimurium TA100. Capsaicin (0.42 mM) attenuated the bacterial mutagenicity of VC and NDMA by 50% and 42% respectively. Diallyl sulfide, a thioether found in garlic with selective P-450 HE1 inhibitory activity, also lessened the mutagenicity of the above carcinogens in a concentration-dependent manner. The suppression of VC- and NDMA-induced mutagenesis by capsaicin and diallyl sulfide correlated with their inhibition of P-450 IIE1-mediated p-nitrophenol hydroxylation and NDMA N-demethylation. Pretreatment of female ICR mice with a topical dose of capsaicin lowered the average number of VC-induced skin tumors by 62% at 22 weeks after promotion. A similar degree of protection was attained with oral administration of diallyl sulfide before carcinogen treatment. The results of this study suggest that capsaicin and diallyl sulfide suppress VC- and NDMA-induced mutagenesis or tumorigenesis in part through inhibition of the cytochrome P-450 IIE1 isoform responsible for activation of these carcinogens.

Allyl Compounds↗

Measurement of mRNA levels in tumor xenografts with quantitative autoradiography and in situ hybridization.

In situ methodologies allow qualitative and semi-quantitative analysis of spatial gene expression in whole organisms or tissues. We have applied quantitative autoradiography to in situ hybridizations of sections from human breast tumor xenografts to measure mRNA levels for ornithine decarboxylase, estrogen receptor, transforming growth factor alpha, and glyceraldehyde-3-phosphate dehydrogenase. Comparisons of control and tamoxifen-treated animals show significant decreases in MCF-7 tumor estrogen receptor mRNA levels in the drug-treated animals. Combining quantitative autoradiography with in situ hybridization allows measurement of absolute rather than relative mRNA levels for genes of interest, and to monitor effector-induced changes in these mRNAs in vivo.

Animals↗

Dealing with complaints--are we doing well?

Investigates the views of complainants and how they felt that their complaints had been handled. Complainants from a three-month period were sent a nine-question questionnaire to which the response rate was 52.3 per cent. Shows that, on the whole, complaint-response times were being met; the complaints system was seen as user friendly; and overall, complaints were satisfied with the way in which their complaints were handled.

Consumer Behavior↗

Sympathetic vasoconstrictive responses to high- and low-sodium diets in diabetic and normal subjects.

Uncomplicated insulin-dependent diabetes mellitus (IDDM) is associated with a suppressed reflex response to sympathetic nervous system (SNS) stimulation and an enhanced pressor response to catecholamines. This study examined the SNS in subjects with IDDM (duration < 5 yr, n = 9) to determine the responsible mechanism within the cardiopulmonary baroreflex arc and the role played by extracellular fluid volume (ECFV) expansion. The reflex arc was tested by examining the plasma norepinephrine (PNE) and forearm vascular (FVR) responses to 60 min of cardiopulmonary baroreceptor unloading by lower body negative pressure (LBNP) at -15 mmHg. The effector limb was tested by measuring the PNE, FVR, and mean arterial pressure (MAP) response to the cold pressor test (CPT). The postganglionic neuroeffector junction was tested by measuring the venoconstrictive response to local norepinephrine infusion. ECFV was varied by altering dietary sodium. In IDDM subjects on a 250 mmol sodium diet, PNE and FVR responses to LBNP (delta PNE = 0.15 +/- 0.05; delta FVR = 4.3 +/- 1.2) were attenuated compared with controls (delta PNE = 0.36 +/- 0.23; delta FVR = 11.56 +/- 2.72). MAP and FVR responses to the CPT were intact (delta MAP = 9.74 +/- 1.9; delta FVR = 7.02 +/- 3.11) compared with controls (delta MAP = 10.74 +/- 2.69; delta FVR = 8.26 +/- 2.94), but the PNE response was attenuated. The peripheral vasculature was hyperresponsive to norepinephrine infusion in IDDM subjects [mean effective dose (ED50) = 57 +/- 10 ng/min] compared with controls (ED50 = 133 +/- 30 ng/min). Sodium restriction (20 mmol/day) normalized the FVR response to LBNP and the venous response to norepinephrine infusion. The PNE response both to LBNP and the CPT remained attenuated.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

Safety of hypertensive hypervolemic therapy with phenylephrine in the treatment of delayed ischemic deficits after subarachnoid hemorrhage.

BACKGROUND AND PURPOSE: Hypertensive hypervolemic therapy has been shown to reverse delayed ischemic deficits after aneurysmal subarachnoid hemorrhage. Concern has been raised about systemic complications of therapy, including pulmonary edema and myocardial ischemia, especially when high doses of vasopressors are used. Patients in whom delayed ischemic deficits were treated with hypervolemia and phenylephrine were prospectively evaluated for signs of systemic toxicity. METHODS: Twenty-four consecutive patients treated with hypertensive hypervolemic therapy after aneurysmal subarachnoid hemorrhage were studied. Sixty-seven percent had underlying cardiac disease, vascular disease, or hypertension. No patient was excluded because of age or preexisting cardiac disease. Patients were closely monitored for signs of congestive heart failure (physical examination, chest x-ray films, arterial blood gases, cardiac index, pulmonary artery wedge pressure, and oxygen requirement). Indicators of cardiac ischemia and other extracerebral toxicity that were monitored included cardiac enzymes, electrocardiograms, serum creatinine, electrolyte and lactic acid levels, gastrointestinal motility, and urine output. RESULTS: Volume expansion and phenylephrine infusion produced an increase in several hemodynamic parameters including pulmonary artery wedge pressure, which rose 28% (13 +/- 3.6 to 16 +/- 1.9 mm Hg), mean arterial blood pressure, which rose 25% (99 +/- 12.5 to 123 +/- 11.4 mm Hg), and systemic vascular resistance, which rose 46% (1234 +/- 294 to 1739 +/- 315 dyne.s-1.cm-5); however, there was no change in cardiac index (3.9 +/- 0.9 to 4.0 +/- 0.6 L.min-1.m-2). There were no clinically significant episodes of pulmonary edema requiring a change in vasopressor therapy and no myocardial infarctions. Phenylephrine was stopped in only one patient (incidence, 4%; 95% confidence interval, 0% to 12%), who developed an exacerbation of his preexisting bradycardia. There was no evidence of noncardiac organ system toxicity. Eighty-eight percent of the patients exhibited neurological improvement. CONCLUSIONS: Hypertensive hypervolemic therapy with the use of high-dose phenylephrine can be administered with acceptable systemic toxicity, even in patients with previous cardiac disease, provided that close monitoring is performed. To minimize risk, aggressive treatment should probably be reserved for patients with signs of delayed ischemia rather than administered prophylactically.

Adult↗

Comparison of thallium-201 and gallium-67 citrate scintigraphy in the diagnosis of pulmonary disease.

Gallium is presently used in the diagnosis of inflammatory, granulomatous, and neoplastic lung disease despite its many logistical problems. In contrast, thallium-201 scintigraphy, which was initially developed for myocardial imaging, offers the principal advantage of immediate imaging and diagnosis although it had not been investigated previously for use in pulmonary disease. In this study, thallium and gallium were prospectively compared with each other for the diagnosis of a variety of lung diseases. The overall concordance rate was 75%. Thallium was found to be significantly more sensitive than gallium for the entire group studied (N = 51, p < 0.006). In a subset of patients in whom a firm diagnosis was established, thallium was also found to be more sensitive, 86% vs 64%. The greater sensitivity of thallium-201 and its several inherent advantages suggest that thallium-201 should replace gallium-67 citrate as the radioisotope of choice for nuclear imaging of the chest.

Adolescent↗

Severe thrombocytopenia following intraarterial papaverine administration for treatment of vasospasm.

Selective intraarterial infusion of papaverine is used in the treatment of symptomatic cerebral vasospasm. The authors report two episodes of severe thrombocytopenia in a patient that were related to intraarterial administration of papaverine. A 70-year-old man with a right internal carotid artery aneurysm underwent craniotomy and aneurysm clipping. He became lethargic 8 days after the hemorrhage occurred. Cerebral angiography revealed moderate vasospasm. In addition to hypervolemic-hypertensive therapy, the patient was treated on two occasions with intraarterial administration of papaverine. Within 24 hours of both treatments he developed severe thrombocytopenia. On one occasion epistaxis requiring transfusion of blood products occurred. Laboratory data support the diagnosis of immune-mediated papaverine-induced thrombocytopenia. The authors conclude that intraarterial administration of papaverine for treatment of vasospasm can be associated with severe, rapidly reversible thrombocytopenia.

Aged↗

Regulation of ornithine decarboxylase mRNA levels in human breast cancer cells: pattern of expression and involvement of core enhancer promoter element.

Ornithine decarboxylase (ODC) expression is increased by growth factors and is obligatory for progression through the cell cycle in a wide variety of cell types. In this study, a variant human ODC cDNA was identified, sequenced, and used to probe mRNA levels in human breast tumor cell lines and xenografts. ODC mRNA was elevated about 3-fold in estrogen receptor-negative (ER-) tumors (MDA-MB-231) when compared with ER-positive (ER+) tumors (MCF-7), as assessed by quantitative autoradiographic analysis of in situ hybridization experiments. The pattern of ODC mRNA in MDA-MB-231 (ER-) xenografts was polarized to the extreme periphery of the tumor, whereas the distribution of ODC mRNA was more evenly distributed in MCF-7 (ER+) xenografts. This correlates with hematoxylin and eosin staining patterns, suggesting that ER+ and ER- xenografts have a differential dependence on host vasculature for growth factor supply. ODC mRNA was elevated 5-fold in MDA-MB-231 cells versus MCF-7 cells when analyzed in cell culture. These relative mRNA levels correlate with increased levels of "core" enhancer binding nuclear proteins in MDA-MB-231 cells over that detected in MCF-7 cells.

Animals↗

Formation of heterodimers from three neurotrophins, nerve growth factor, neurotrophin-3, and brain-derived neurotrophic factor.

Three neurotrophic factors, brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and nerve growth factor (NGF) form noncovalent homodimers in solution. Since they are highly homologous proteins, it seemed probable that two monomers of these proteins might associate together to form a heterodimer. This was tested by denaturing the two different proteins together in 6 M guanidine HCl and refolding them in phosphate-buffered saline. When the refolded mixture of BDNF and NT-3 was subjected to Mono S cation exchange chromatography, a new peak was observed eluting between NT-3 and BDNF, which accounted for about 30% of the protein used. This new protein species migrated as a single band upon native gel electrophoresis with mobility between that of the NT-3 homodimer and the BDNF homodimer, indicating that a complex had been formed. Sedimentation equilibrium data show that the dissociation constant of this heterodimer is < 3 x 10(-10) M. The heterodimer was stable upon incubation at 37 degrees C in phosphate-buffered saline over 11 days. Having determined that the heterodimer is highly stable, it was subjected to various biological assays. Autophosphorylation assay using TrkB receptor showed that the heterodimer is indistinguishable from the BDNF or NT-3 homodimer in the ability to induce phosphorylation of the receptor. It was also indistinguishable from the homodimers in the neurotrophic activity using chick dorsal root ganglion explant. In the sympathetic neuron survival assay, the heterodimer behaved more similarly to NT-3, whereas in the dopamine uptake assay, it was intermediate between the two homodimers. In addition, the heterodimer was shown to be retrogradely transported in the dorsal root ganglion neurons. A heterodimer between NGF and BDNF is formed but much less effectively than the NT-3.BDNF heterodimer, and it is not stable even at 4 degrees C. These results indicate that BDNF and NT-3 have an intersubunit contact surface for dimerization resembling each other's but different from the contact surface of NGF.

3T3 Cells↗

Recent studies on the metabolic activation of chemical carcinogens.

Xenobiotics continue to be found that are metabolized in mammals to electrophilic reactants that form covalently bound adducts to cellular DNA leading to tumor formation. This discussion will be limited to the following two topics: sulfonation in chemical carcinogenesis and endogenous etheno adducts in DNA.

Animals↗