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Biomedical subjects

R A Smith

Publications and source records attributed to R A Smith.

At least 19 recordsLinked to original sources
PubMed1992

Comparative behaviour of calpain and cathepsin B toward peptidyl acyloxymethyl ketones, sulphonium methyl ketones and other potential inhibitors of cysteine proteinases.

Peptidyl acyloxymethyl ketones, previously established as potent inactivators of the lysosomal cysteine proteinase cathepsin B, were evaluated against smooth-muscle calpain, a member of the family of Ca(2+)-dependent cysteine proteinases. Only modest rates of time-dependent inhibition could be achieved, even with peptidyl affinity groups optimized for calpain and linked to a carboxylate leaving group of very low pKa [2,6-(CF3)2PhCOO-, pKa 0.58]. Selective inactivation of cathespin B versus calpain was consistently observed with this type of inhibitor. Examination of other potential inhibitors revealed a rank order of potency against calpain to be: peptidyl sulphonium methyl ketones > fluoromethyl ketones, diazomethyl ketones >> acyloxymethyl ketones, an order which differs sharply from that found for cathespin B.

Amino Acid Sequence↗
PubMed1992

Loop mobility in a four-helix-bundle protein: 15N NMR relaxation measurements on human interleukin-4.

15N NOE, T1, and T2 measurements have been carried out on uniformly 15N-labeled human interleukin-4. Analysis of the results in terms of order parameters (S2) shows that although the helical core of this four-helix-bundle protein exists as a well-defined structure with limited conformational flexibility (S2 congruent to 0.9), other regions of the molecule experience substantial fluctuations in the conformation of the main chain (S2 = 0.3-0.8). These regions include both the N- and C-termini and two of the loops joining the helices. The majority of these internal motions are fast compared with the overall rotational correlation time (tau R = 7.6 ns at 35 degrees C) and are localized in regions that are relatively ill-defined in the NMR structures previously determined for this protein [Smith, L. J., Redfield, C., Boyd, J., Lawrence, G. M. P., Edwards, R. G., Smith, R. A. G., & Dobson, C. M. (1992) J. Mol. Biol. 224, 899-904]. Other motions are on a slower time scale and appear to be associated with two of the three disulfide bonds and the beta-sheet region in the protein. The dynamic properties of interleukin-4 in solution have been compared with features of the X-ray structures of other four-helix-bundle proteins. The results suggest that the dynamic properties observed here may be general for this class of proteins and may be significant for the interpretation of both their structural and functional properties.

Humans↗
PubMed1992

Sequential 1H NMR assignments and secondary structure of the kringle domain from urokinase.

The sequence-specific 1H NMR assignments of the 89-residue recombinant kringle domain from human urokinase are presented. These were achieved primarily by utilizing TOCSY and NOESY spectra in conjunction with COSY spectra recorded at 500 MHz and 600 MHz. Regular secondary structure elements have been derived from a qualitative interpretation of nuclear Overhauser enhancement, JNH alpha coupling constant, and amide proton exchange data. Two helices have been identified. One helix, involving Ser40-Gly46, corresponds to that reported for t-PA kringle 2 (Byeon et al., 1991), but does not exist in other kringles with known structures. The second helix, in the region Asn26-Gln33, is thus far unique to the urokinase kringle. Three antiparallel beta-sheets and three tight turns have also been identified, which correspond exactly to those identified in t-PA kringle 2 both in solution and in the crystalline state (de Vos et al., 1992). Despite the very different ligand binding properties of the urokinase kringle, NOE data indicate that the tertiary fold of the molecule conforms closely to that found for other kringles.

Amino Acid Sequence↗
PubMed1992

Acrolein initiates rat urinary bladder carcinogenesis.

Acrolein, a reactive, alpha,beta-unsaturated aldehyde which is ubiquitous in the environment, forms DNA adducts, is mutagenic, and is teratogenic. However, studies have not indicated a carcinogenic effect in rodent bioassays. Since it is present in cigarette smoke and is the toxic metabolite of cyclophosphamide with respect to the urinary tract, we investigated the possibility that acrolein might have carcinogenic activity toward the rat urinary bladder. We also evaluated whether it possessed initiating and/or promoting activity. To evaluate initiating activity, acrolein was administered at a dose of 2 mg/kg i.p. twice a week for 6 weeks followed by uracil as 3% of the diet for 20 weeks and then control diet for 6 weeks. N-[4-(5-Nitro-2-furyl)-2-thiazolyl]formamide (FANFT) as 0.2% of the diet followed by uracil was used as a positive control, and a negative control group was administered solvent control (water) i.p. during the 6-week initiation period followed by uracil. Acrolein followed by uracil produced an incidence of 18 of 30 rats (60%) with papilloma compared to 8 of 30 rats (27%) treated with solvent control followed by uracil. FANFT followed by uracil produced an incidence of 70% carcinomas and 30% papillomas, clearly indicating that it is a much more potent initiating agent than acrolein. Acrolein for 6 weeks followed by control diet produced no tumors. To evaluate promoting activity, groups of rats were fed FANFT for 6 weeks followed by acrolein. Acrolein administered during the initial 6 weeks and continued for the second phase of the experiment (to evaluate complete carcinogenic activity) resulted in severe toxicity. Administration of acrolein had to be terminated after 21 weeks of the experiment. The animals were maintained for 53 weeks of the experiment without further chemical treatment, and there was no evidence of papilloma or carcinoma development. This study clearly indicates that acrolein has initiating activity for the urinary bladder when administered by i.p. injection to the male F344 rat, but toxicity precluded evaluation of its promoting or complete carcinogenic activity.

Acrolein↗
PubMed1992

Breast and cervical cancer surveillance, United States, 1973-1987.

Breast and cervical cancer incidence and mortality rates were reviewed for the period 1973-1987. For breast cancer, mortality has been relatively stable, increasing from 26.9/100,000 women in 1973 to 27.1 in 1987. Alternatively, data from the National Cancer Institute's Surveillance, Epidemiology, End Results Program (SEER) showed a 36% increase in the incidence of this malignancy over the same period. In 1987, overall incidence of invasive breast cancer was 111.9/100,000 women. White women experienced lower overall mortality rates and higher overall incidence than black women; however, these differences varied by age. Examination of breast cancer incidence by stage of disease at diagnosis revealed that rates for distant and regional disease have remained relatively stable since 1973. In contrast, rates of localized and in situ cancers exhibited an increase in the 1980s that may correspond to increased use of mammography in this country. The rate of decline in cervical cancer incidence and mortality has slowed in recent years. In 1987, 3.0 cervical cancer deaths/100,000 women occurred. SEER incidence for invasive disease for that year was 8.2/100,000. Rates varied by race, age, state, and stage of disease. In general, black women experienced much higher incidence and mortality from invasive cervical cancer than white women. For both races, rates of in situ disease were highest among young women and decreased rapidly with age. Rates of in situ cervical cancer were consistently higher than rates of invasive cancer for the time period studied.

Adult↗
PubMed1992

Human interleukin 4. The solution structure of a four-helix bundle protein.

Heteronuclear 13C and 15N three-dimensional nuclear magnetic resonance (n.m.r.) techniques have been used to determine the solution structure of human interleukin 4, a four-helix bundle protein. A dynamical simulated annealing protocol was used to calculate an ensemble of structures from an n.m.r. data set of 1735 distance restraints, 101 phi angle restraints and 27 pairs of hydrogen bond restraints. The protein structure has a left-handed up-up-down-down topology for the four helices with the two long overhand loops in the structure being connected by a short section of irregular antiparallel beta-sheet. Analysis of the side-chains in the protein shows a clustering of hydrophobic residues, particularly leucines, in the core of the bundle with the side-chains of charged residues being located on the protein surface. The solution structure has been compared with a recent structure prediction for human interleukin 4 and with crystal structures of other helix bundle proteins.

Computer Graphics↗
PubMed1992

Barriers to screening for breast cancer.

Despite strong epidemiologic evidence that screening for breast cancer with mammography and clinical breast examination results in mortality reductions, and the considerable effort to communicate this message to women and health-care providers, most US women are not screened according to recommended guidelines. Recent investigations have focused on trends in use, and factors associated with physicians' and women's knowledge, attitudes, and practices associated with mammography. Even as use of mammography has increased, the literature suggests that a number of significant impediments to participation in routine screening will need to be addressed to achieve high rates of screening among US women according to recommended guidelines.

Breast Neoplasms↗
PubMed1992

The use of fresh frozen allogeneic bone for maxillary and mandibular reconstruction.

The use of fresh frozen bone (FFB) alone, or in combination with autogenous bone (AB), for bony augmentation of the maxilla and mandible in preparation for dental reconstruction with endosseous implants has been studied. Ten patients received FFB +/- AB for augmentation of a severely atrophic mandible (n = 6) or for reconstruction of a jaw defect secondary to trauma or tumor resection (n = 4). Average follow-up was 26.3 +/- 5.4 months. At the time of implant placement, the bone grafts were found to be firm in consistency, well incorporated, and well vascularized in all 10 patients. Twenty-nine endosseous implants were placed an average of 8.3 +/- 3.1 months following bone grafting. One implant failed and was replaced, and one implant remains buried as a nonfunctional unit. All patients have been restored prosthetically by means of 28 of the 29 implants. This preliminary study indicates that FFB may be used alone or in combination with autogenous bone for augmentation or reconstruction of the atrophic maxilla and mandible. The resultant ridge is adequate to support loaded endosseous implants. A potential disadvantage is the minimal risk of disease transmission.

Bone Transplantation↗
PubMed1992

The use of bovine fibrin-streptokinase films for the determination of recombinant human plasminogen.

Plasminogen is a key component of the haemostatic system in man and the plasma-derived protein molecule has been actively investigated. Within the last few years cDNA and the gene encoding plasminogen have been cloned and the protein has been expressed in a number of eukaryotic or prokaryotic systems. Yields of expressed plasminogen are frequently low. Currently available assays for plasminogen generally rely on the determination of antigen or utilize tripeptide substrates for measuring functional activity, and they have certain limitations. Assays employing relevant protein substrates offer an alternative way to measure function and overcome the drawbacks associated with the other tests. The use of fibrin films for the assay of low levels of recombinant plasminogen has not been described fully before. The two fibrin film-based assays described in this paper are significant additions to the array of assays available for plasminogen molecules.

Animals↗
PubMed1992

The development of a microwave fluid-bed processor. I. Construction and qualification of a prototype laboratory unit.

The static bed- and planetary-type microwave dryers currently available to process pharmaceutical materials are not designed to use hot-air fluidization for the purpose of maximizing microwave energy inputs and particle drying. To take advantage of the benefits offered by fluidization, a 1-kg Uni-Gatt laboratory fluid bed processor was modified to support microwave-assisted fluid bed drying of several representative pharmaceutical granulations. The construction, design features, and validation of this new microwave fluid bed processor are presented.

Calibration↗
PubMed1992

Dextromethorphan: enhancing its systemic availability by way of low-dose quinidine-mediated inhibition of cytochrome P4502D6.

There has been a substantial amount of interest in the anticonvulsant and neuroprotective actions of dextromethorphan. Its therapeutic efficacy, however, is limited by its extensive first-pass elimination by way of the cytochrome P4502D6 enzyme in humans. The purpose of this research was to determine whether quinidine (a selective inhibitor of cytochrome P4502D6) could improve dextromethorphan systemic delivery in patients with amyotrophic lateral sclerosis (a neurodegenerative disease). In the absence of quinidine, 60 mg dextromethorphan every 12 hours resulted in plasma concentrations of only 12 +/- 13 ng/ml (range, less than 5 to 40 ng/ml; n = 7). The same dose of dextromethorphan in the presence of 75 mg quinidine every 12 hours resulted in dextromethorphan plasma concentrations of 241 +/- 94 ng/ml (range, 157 to 402 ng/ml; n = 5). The achievement of higher dextromethorphan plasma concentrations was also associated with an increased occurrence of adverse effects in some patients. Based on the brain/plasma ratio for dextromethorphan in rats, it is estimated that brain dextromethorphan concentrations of 1.0 to 10 micrograms/gm may be attainable in humans by inhibition of cytochrome P4502D6 activity with quinidine.

Adult↗
PubMed1992

Absence of ras oncogene activation in rat urinary bladder carcinomas induced by N-methyl-N-nitrosourea or N-butyl-N-(4-hydroxybutyl)nitrosamine.

Previously, we demonstrated point mutations of the H-ras gene in N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide (FANFT)-induced rat urinary bladder carcinomas. In this study, ras oncogene activation was examined in urinary bladder carcinomas induced by N-(4-hydroxybutyl)nitrosamine (BBN) or N-methyl-N-nitrosourea (MNU) administration followed by uracil treatment. In the first experiment, MNU (20 mg/kg body wt) was i.p. injected into 11 male F344 rats twice a week for 4 weeks, followed by feeding 3% uracil for 20 weeks (MNU/uracil group). Ten rats were given only 3% uracil without MNU pretreatment. In the second experiment, 20 male F344 rats were given 0.05% BBN in the drinking water for 4 weeks, then fed 3% uracil for 20 weeks (BBN/uracil group). Another 20 rats were fed 3% uracil without the BBN pretreatment. Transitional cell carcinomas were induced in the urinary bladder of all rats in the MNU/uracil and BBN/uracil groups. Papillomas and hyperplasias were present in the rats given uracil without prior BBN or MNU. DNA and protein were extracted from the tumors (MNU/uracil or BBN/uracil groups) or from the scraped bladder epithelium (uracil alone groups). Sequences around codons 12, 13 and 61 of H-, K- and N-ras genes were examined by direct sequencing after polymerase chain reaction, and p21 was examined by Western blotting. No mutation was found within the examined sequences and p21 showed no changes in mobility. There was no difference in the level of p21 expression between rats treated with MNU/uracil or BBN/uracil compared to corresponding uracil alone groups. These results indicate that the ras oncogene was not activated in urinary bladder carcinomas induced by BBN or MNU in combination with uracil treatment, in contrast to previous findings with FANFT.

Animals↗
PubMed1992

Pressure support ventilation decreases inspiratory work of breathing during general anesthesia and spontaneous ventilation.

Spontaneous ventilation may offer advantages over controlled mechanical ventilation (CMV), but increase in work of breathing may diminish its usefulness. During general anesthesia, respiratory depression and increased work of breathing often preclude spontaneous ventilation, and patients then receive CMV. We compared the inspiratory work of breathing of anesthetized patients who breathed with pressure support ventilation (PSV) with that associated with a demand gas flow and a standard anesthesia circle system. We studied nine consenting patients who underwent general inhaled anesthesia with or without regional supplementation. An anesthesia/ventilator system (Siemens 900D, Solna, Sweden) provided PSV (5 cm H2O) or demand gas flow during spontaneous inspiration. Gas flow during demand breathing and PSV was initiated when inspiration produced a 2-cm H2O reduction in airway pressure. An anesthesia machine (Dräger Narkomed 3, Telford, Pa.) provided a gas flow rate of 6 L/min through a standard semiclosed circle system. Airway pressure, airway gas flow rate, and esophageal pressure were continuously transduced, and data or signals were conveyed to a computer. Tidal volume and respiratory rate were computed from the flow curve. The inspiratory work of breathing was calculated as the integral of the area subserved by a plot of esophageal pressure and tidal volume during inspiration. Heart rate and mean arterial blood pressure were recorded, and arterial blood was sampled for gas tension and pH analysis. No differences were found in pHa, Paco2, Pao2, tidal volume, respiratory rate, heart rate, or mean arterial blood pressure among the three modes of ventilation.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1992

Tissue distribution of 125I-labeled fetal thyroid hormone in nude mice xenografted with Ewing sarcoma.

Biodistribution of fetal thyroid hormone (RT3) was studied in nude mice with Ewing's sarcoma xenografts. At 30 min and 1, 2, 4, 8, and 12 hr after injection, blood, tumor and normal organs were measured to determine the amount of radioactivity per gram of tissue. The amount of radioactivity in the blood of tumor-bearing mice decreased sharply from 7.64% of injected dose per gram of tissue (% ID/g) at 30 min after inoculation to 0.45%/ID/g at 12 hr. The % of injected dose per gram for 125I-labeled RT3 in the tumor reached 1.92 at 1 hr after injection and decreased to 0.22 at 12 hr. The radiolocalization indices for tumor to other organs at 12 hr ranged from 1.35 to 4.43. This study suggests increased localization of RT3 in the Ewing's sarcoma as compared to other tissues in the nude mouse.

Animals↗
PubMed1992

The size of the mature membrane receptor for follicle-stimulating hormone is larger than that predicted from its cDNA.

A 240 kDa protein isolated from bovine calf testis has been shown to have properties characteristic of an FSH receptor. However, rat testis FSH receptor has, on the basis of cloning experiments, been found to have a much lower molecular mass of 75 kDa (peptide only). To examine this point, the size of the FSH receptor in membranes obtained from cultured Sertoli cells of immature rats was determined after polyacrylamide gel electrophoresis under non-reducing conditions, followed by transfer to polyvinylidene difluoride membranes and direct identification of the FSH receptor by ligand blot analysis utilizing radioiodinated human FSH. In this system, the rat Sertoli cell membrane FSH receptor also showed a molecular mass of 240 kDa. Bovine testis contains LH and FSH receptors. We compared the sizes of FSH and LH receptors present in the same bovine testis membrane preparation by ligand blot analysis. The FSH receptor again showed a molecular mass of 240 kDa, whereas the LH receptor showed a molecular mass of 90 kDa. The latter value is similar to that deduced by cloning techniques (75 kDa, peptide only). The evidence seems to suggest that, whereas the molecular mass deduced for the LH receptor on the basis of its cDNA is similar to that of the mature membrane receptor, the size of the FSH membrane receptor is considerably different from that deduced on the basis of its cDNA, presumably as a result of post-translational processing. The marked difference in size between mature FSH (240 kDa) and LH (90 kDa) receptors may reflect significant structural differences of importance with regard to mechanisms of signal transduction.

Animals↗
PubMed1992

Risk factors associated with dental implants in healthy and medically compromised patients.

A total of 104 consecutive patients treated with 313 Nobelpharma implants was studied to determine the medical risks associated with dental implants. There did not appear to be an increased implant failure rate or an increase in perioperative morbidity in patients with a compromised medical status. Age; sex; and concurrent use of hypoglycemic agents, supplemental female hormones, or steroids also did not correlate with increased implant failure or perioperative morbidity. Implant procedures using a variety of pain-/anxiety-control agents failed to reveal any increase in anesthetic-related complications. However, the number of implants placed per patient did correlate with implant failure. It appears that implant surgery and the required anesthetic appear to be safe procedures even in the medically compromised patient.

Adolescent↗