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Biomedical subjects

R T Miller

Publications and source records attributed to R T Miller.

At least 91 records · Page 5Linked to original sources
PubMed1991

Structure of the 5'-flanking regulatory region and gene for the human growth factor-activatable Na/H exchanger NHE-1.

The amiloride-sensitive, growth factor-activatable Na/H exchanger (NHE-1) is a ubiquitous mammalian protein that is involved in the regulation of intracellular pH and cell volume. We have determined the intron/exon boundaries and the transcription initiation sites and have characterized a portion of the 5'-flanking regulatory region of the human NHE-1 gene. The Na/H exchanger gene spans approximately 70 kilobases. The coding region is divided into 12 exons and 11 introns, one of which is 41.5 kilobases in length. The first exon contains the entire 5'-noncoding region, which is 786 bases long, and 352 bases of the coding sequence. Primer extension identified two discrete start sites for RNA polymerase. 1377 bases of the 5'-regulatory region were sequenced. The promoter/enhancer region is characterized by a TATA box, four GC boxes, two CAAT boxes, five CACCC boxes, three Ap-1 sites, a cyclic AMP response element, and four partial glucocorticoid response elements. Promoter activities of a 313- and a 1441-base pair fragment containing the TATA box were demonstrated by their ability to direct chloramphenicol acetyltransferase expression when transiently expressed in fibroblasts.

Animals↗
PubMed1991

Skin pustules and nodules caused by Actinomyces viscosus in a horse.

A 12-year-old Arabian stallion was evaluated for a refractory skin problem of 2 years' duration. Skin nodules and pustules, which would rupture, had developed over the right lumbar muscles. Anaerobic culturing from the pustules yielded Actinomyces viscosus, and histologic examination of biopsy specimens revealed globular eosinophilic structures. Concomitant treatment with isoniazid (8 mg/kg of body weight, q 24 h for 8 weeks), trimethoprim-sulfadiazine (30 mg/kg, q 24 h for 8 weeks), and sodium iodide solution (66 mg/kg, every 1, 2, or 4 weeks, for 32 weeks) resolved the condition.

Actinomycosis↗
PubMed1991

Multitumor "sausage" blocks in immunohistochemistry. Simplified method of preparation, practical uses, and roles in quality assurance.

This report describes a simplified method for preparing multitumor sausage blocks (MTSBs) for use in immunohistochemical procedures. Rather than relying on previously processed paraffin blocks as a source of material, this procedure involves procuring tissue at the time of gross specimen examination. The tissue is processed along with routine surgical pathologic material, and the paraffinized samples are placed in "storage cassettes" for easy cataloging and storage. Thin strips are cut from the tissue in the "storage cassettes" and combined by dripping liquid paraffin onto them while they are rolled between the thumbs and forefingers, somewhat like making a cigarette. This results in a tissue "log." Transverse sections of the "log" are embedded in paraffin blocks and used as MTSBs. Practical uses of MTSBs are discussed, and their role in quality assurance is stressed.

Histological Techniques↗
PubMed1991

Differential regulation of Na/H antiporter by acid in renal epithelial cells and fibroblasts.

Increased Na/H antiporter activity has been demonstrated after in vivo chronic metabolic acidosis as well as in vitro acid preincubation of cultured rabbit renal tubule cells. To study the underlying molecular mechanisms of this adaptive increase in Na/H antiporter activity, the present studies examined the effect of low pH media on Na/H antiporter activity and mRNA abundance in cultured renal tubule cells. Na/H antiporter activity was increased by 60% in a mouse renal cortical tubule cell line (MCT), and by 90% in an opossum kidney cell line (OKP) after 24 h of preincubation in acid (low [HCO3]) media. The ethylisopropylamiloride sensitivity of the Na/H antiporters were different in these two cell lines (MCT IC50 = 65 nM; OKP IC50 = 4.5 microM). In MCT cells, Na/H antiporter mRNA abundance measured by RNA blots increased by two- to fivefold after 24 h in low [HCO3] media. Na/H antiporter mRNA abundance was also increased in MCT cells with high CO2 preincubation as well as in rat renal cortex with in vivo chronic acid feeding. In contrast to renal epithelia, acid preincubation of NIH 3T3 fibroblasts led to suppression of Na/H antiporter activity. RNA blots of 3T3 fibroblasts revealed the same size Na/H antiporter transcript as in MCT cells. However, Na/H antiporter mRNA levels were suppressed by acid preincubation. These studies demonstrate differential regulation of Na/H antiporter activity and mRNA abundance in renal epithelial cells and fibroblasts in response to an acidotic environment.

Acidosis↗
PubMed1991

Chronic adaptations in proximal tubular H/HCO3 transporters.

Chronic acidosis, chronic K deficiency, and chronic hyperfiltration all lead to similar parallel increases in Na/H antiporter and Na/3HCO3 cotransporter activities. These effects persist when the transporters are removed from the inducing environment, and thus represent memory effects. The effect of acidosis on the Na/H antiporter can be reproduced by preincubating cultured proximal tubule cells chronically in acid media, suggesting that low extracellular fluid pH is an important signal in acidosis. The specific extracellular signals in chronic K deficiency and chronic hyperfiltration have not been elucidated. The effect of acid preincubation on Na/H antiporter activity in cultured proximal tubule cells is dependent on protein synthesis and is associated with increased abundance of mRNA for the Na/H antiporter. The cell signalling pathways which mediate these effects have not been determined. However, the common response in chronic acidosis, chronic K deficiency, and chronic hyperfiltration to the demand for increased H transport, suggests that a common signalling pathway will be found in all these conditions.

Adaptation, Physiological↗
PubMed1990

Improved avidin-biotin immunoperoxidase method for terminal deoxyribonucleotidyl transferase and immunophenotypic characterization of blood cells.

This report describes a simple avidin-biotin immunoperoxidase method for detection of cell surface antigens and terminal deoxyribonucleotidyl transferase (TdT) in blood, bone marrow, and body fluids. It employs enhanced sensitivity avidin-biotin complex (ABC) reagents and includes a step for quenching of endogenous peroxidase activity that does not diminish antigenic reactivity, allowing its application to specimens (such as blood or bone marrow films) that have significant levels of endogenous peroxidase. This quenching step is performed after the secondary (biotinylated) antibody incubation, which also allows for maximum sensitivity. Examples of results obtained with this method are illustrated, and advantages over other immunocytochemical methods are discussed.

Antigens, CD↗
PubMed1989

Mutations in the GTP-binding site of GS alpha alter stimulation of adenylyl cyclase.

Mutational replacements of specific residues in the GTP-binding pocket of the 21-kDa ras proteins (p21ras) reduce their GTPase activity. To test the possibility that the cognate regions of G protein alpha chains participate in GTP binding and hydrolysis, we compared signaling functions of normal and mutated alpha chains (termed alpha s) of Gs, the stimulatory regulator of adenylyl cyclase. alpha s chains were expressed in an alpha s-deficient S49 mouse lymphoma cell line, cyc-. alpha s in which leucine replaces glutamine 227 (corresponding to glutamine 61 of p21ras) constitutively activates adenylyl cyclase and reduces the kcat for GTP hydrolysis more than 100-fold. There is a smaller reduction in GTPase activity in another mutant in which valine replaces glycine 49 (corresponding to glycine 12 of p21ras). This mutant alpha s is a poor activator of adenylyl cyclase. Moreover, the glycine 49 protein, unlike normal alpha s, is not protected against tryptic cleavage by hydrolysis resistant GTP analogs; this finding suggests impairment of the mutant protein's ability to attain the active (GTP-bound) conformation. We conclude that alpha s residues near glutamine 227 and glycine 49 participate in binding and hydrolysis of GTP, although the GTP binding regions of alpha s and p21ras are not identical.

Adenylyl Cyclases↗
PubMed1988

Right-sided Staphylococcus aureus endocarditis in intravenous drug abusers: two-week combination therapy.

STUDY OBJECTIVE: To determine the efficacy of short-course combination regimens for selected cases of Staphylococcus aureus endocarditis in intravenous drug abusers. DESIGN: Open study of nafcillin and tobramycin or vancomycin and tobramycin administered for 2 weeks with no further therapy. SETTING: County hospital. PATIENTS: Consecutive sample of 53 intravenous drug abusers with relatively uncomplicated right-sided S. aureus endocarditis, defined by clinical and echocardiographic criteria, and without renal insufficiency, extrapulmonary metastatic infectious complications requiring prolonged therapy or surgery for cure, meningitis, methicillin-resistant organism, aortic or mitral valve involvement, or pregnancy. INTERVENTIONS: Nafcillin, 1.5 g intravenously every 4 hours, plus tobramycin, 1 mg/kg body weight intravenously every 8 hours, administered for 2 weeks. Vancomycin, 30 mg/kg per day intravenously, in two or three divided doses, was used instead of nafcillin for patients allergic to penicillin. MEASUREMENTS AND MAIN RESULTS: Forty-seven of 50 patients (94%; 95% CI, 87 to 99+) treated with the nafcillin and tobramycin combination were cured. Only 1 of 3 patients treated with vancomycin plus tobramycin (33%, 95% CI, 2 to 86) was cured. CONCLUSIONS: Selected patients with S. aureus endocarditis can be treated safely and effectively with a 2-week course of nafcillin plus tobramycin. Only one of three patients treated with vancomycin plus tobramycin was cured, but three patients are too few to define with confidence the efficacy of this regimen.

Adult↗
PubMed1988

A mutation that prevents GTP-dependent activation of the alpha chain of Gs.

Membrane-bound G proteins carry information from receptors on the outside of cells to effector proteins inside cells. The alpha subunits of these heterotrimeric proteins bind and hydrolyse GTP and control the specificity of interactions with receptor and effector elements. Signalling by G proteins involves a cycle in which the inactive alpha beta gamma-GDP complex dissociates to produce alpha*-GTP, which is capable of activating the effector enzyme or ion channel; the alpha*-GTP complex hydrolyses bound GTP and reassociates with beta gamma to form the inactive complex. We have characterized a mutation that interrupts this GTP-driven cycle in alpha s, the alpha-chain of Gs, the G protein that stimulates adenylyl cyclase. The mutation converts a glycine to an alanine residue in the presumed GDP-binding domain of alpha s. The location and biochemical consequences of this mutation suggest a common mechanism by which binding of GTP or ATP may induce changes in the conformation of a number of nucleoside triphosphate binding proteins.

Adenosine Triphosphate↗
PubMed1988

Carboxyl terminal domain of Gs alpha specifies coupling of receptors to stimulation of adenylyl cyclase.

The alpha subunits of Gs and Gi link different sets of hormone receptors to stimulation and inhibition, respectively, of adenylyl cyclase. A chimeric alpha i/alpha s cDNA was constructed that encodes a polypeptide composed of the amino terminal 60% of an alpha i chain and the carboxyl terminal 40% of alpha s. The cDNA was introduced via a retroviral vector into S49 cyc- cells, which lack endogenous alpha s. Although less than half of the hybrid alpha chain is derived from alpha s, its ability to mediate beta-adrenoceptor stimulation of adenylyl cyclase matched that of the normal alpha s polypeptide expressed from the same retroviral vector in cyc- cells. This result indicates that carboxyl terminal amino acid sequences of alpha s contain the structural features that are required for specificity of interactions with the effector enzyme, adenylyl cyclase, as well as with the hormone receptor.

Adenylyl Cyclases↗
PubMed1988

Adenocarcinoid tumor of appendix presenting as unilateral Krukenberg tumor.

We report a case of adenocarcinoid tumor of the appendix that presented initially as a unilateral Krukenberg tumor (a signet ring cell mucinous adenocarcinoma with prominent cellular stroma). The primary tumor in the appendix was discovered 10 months later at the time of a "second look" laparotomy. The ovarian metastasis showed both goblet cell elements and tubular formations with numerous argyrophilic cells, indicating that both components of these tumors may metastasize, a finding at variance with the conclusions of some authors who suggest that only the mucinous component may metastasize. Theories of histogenesis of these tumors are discussed, and 12 previously reported cases presenting as Krukenberg tumors (all bilateral) are reviewed. Because the primary tumor in the appendix may be small and easily missed, appendectomy is recommended in all patients with Krukenberg tumors when another primary site cannot be identified at the time of surgery.

Adenocarcinoma↗
PubMed1988

Mutations probe structure and function of G-protein alpha chains.

The molecular genetic approach has just begun to provide hints of answers to some of the questions posed at the outset of this paper. We have some idea of which portions of the alpha chain of Gs interact with receptors and effectors, and we guess that the same is true of corresponding regions of other alpha chains. We have a tantalizing hint that points to a key region of alpha s that is necessary for the conformational change induced by binding GTP, and the vague outline of a hypothesis regarding the mechanism by which receptors release GDP from its binding site on the alpha chain. The alpha chain region (domain) that interacts with beta gamma remains unknown. The tenuous quality of all these hints and hypotheses is obvious, and at least in the short term, frustrating. Even the present level of our understanding, however, is impressive in comparison with what was known of G-protein function only 5 years ago. Now we can pose much more precise questions and hope that a combination of the molecular genetic approach with biophysical probes of structure will provide satisfying answers.

Animals↗
PubMed1987

Modification of the internal pH sensitivity of the Na+/H+ antiporter by parathyroid hormone in a cultured renal cell line.

Sodium-proton antiporter activity can be modulated through changes Vmax and/or intracellular proton sensitivity of the antiporter. To characterize a parathyroid hormone (PTH)-induced decrease in antiporter activity in a continuous renal cell line (opossum kidney cells), the extracellular sodium and intracellular proton dependence of amiloride-inhibitable 22Na uptake was studied. The Km for extracellular sodium at intracellular pH 6.32 was 28 mM and was unaltered by PTH, whereas the Vmax was decreased by 26%. When intracellular pH was set over the range 5.87-7.57 by the potassium-nigericin method, antiporter activity increased as intracellular pH decreased. Hill analysis revealed Hill coefficients of 1.25 and 1.01 and half-maximal antiporter activity at intracellular pH values of 6.90 and 6.35 for control and PTH-treated cells, respectively. PTH decreased the apparent Vmax at low pH by 15% and the intracellular pH at which Na+/H+ exchange is half-maximal by 0.55 pH units.

Animals↗
PubMed1987

The effects of cycloheximide on Na+/H+ antiporter activity in cultured opossum kidney cells.

These studies examined the effects of cycloheximide on the Na+/H+ antiporter in cultured opossum kidney cells. The effects of cycloheximide on antiporter activity depended on the basal level of activity. These data suggest that the Na+/H+ antiporter may be regulated by several processes which are sensitive to protein synthesis inhibition.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1986

Melanotic schwannoma of the acoustic nerve.

Melanotic schwannoma is a rare neoplasm, with only 23 previously reported cases. We report an additional case of a melanotic schwannoma arising from the acoustic nerve in a 74-year-old man who had a probable forme fruste of neurofibromatosis. We briefly review the literature of this neoplasm and discuss theories regarding the histogenesis of these tumors.

Aged↗
PubMed1984

Pathophysiology and management of severe asthma.

Severe asthma is characterized by marked airway obstruction caused by mucus plugging, airway edema and inflammation, and contractions of airway smooth muscle. This leads to a complex series of pathophysiologic events that can be reversed with vigorous treatment given for a sufficient period of time. Standard treatment includes not only bronchodilating agents and corticosteroids but in some instances mechanical ventilation.

Adrenal Cortex Hormones↗