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Biomedical subjects

R L Miller

Publications and source records attributed to R L Miller.

At least 163 records · Page 9Linked to original sources
PubMed1986

Phospholipid methylation in starfish spermatozoa is linked to sperm chemoattraction.

The mechanism whereby ovarian peptides cause sperm attraction was studied in the starfish. Phospholipid methylation and protein-O-carboxyl methylation, reactions linked to chemotactic responses in a variety of systems, were studied in starfish sperm. When sperm were preincubated with [methyl-3H]methionine and then exposed to the attractant, a rapid drop in radioactivity occurred in the phospholipid fraction. Methylated phospholipids decreased by 90% in the first 2 sec; however, no change was observed in endogenous methylation of protein carboxyl groups. The effect on phospholipid methylation was dose dependent, with a 40% reduction in radioactive phospholipids in sperm occurring with the minimal amount of attractant necessary to obtain a positive response in a sperm attraction bioassay. Attractants from species of starfish with little or no cross-reactivity in the bioassay had a limited effect on phospholipid methylation. The transmethylase inhibitor, homocysteine, caused a marked decrease in the accumulation of methylated phospholipids under basal conditions, which was correlated with as much as a 50-fold increase in sperm sensitivity to the attractant. The addition of chemoattractant resulted in a reduction in the amount of all individual methylated phospholipids, but the amount of phosphatidylmono[3H]methylethanolamine relative to the other methylated phospholipid decreased by a factor of 4 after stimulation. Homocysteine had the same effect. The reduction in methylated phospholipids by attractants suggests that phospholipid methylation is linked to the mechanism of action of these peptides. Methylation of phospholipids may play a role in the rapid desensitization of sperm cells to the attractant, which would be required for the orientation of the spermatozoa in the gradient of ovarian peptide.

Animals↗
PubMed1985

Lectin histochemistry of secretory and cell-surface glycoconjugates in the ovine submandibular gland.

Information about the chemical structure of ovine submandibular glycoconjugates was obtained in situ by means of a battery of peroxidase-conjugated lectins with affinity for specific terminal or internal sugars or sugar sequences in conjunction with neuraminidase digestion and periodate oxidation. Stored secretions in all mucous acinar cells contained disaccharide side chains consisting of N-acetylneuraminic acid linked to penultimate alpha-N-acetylgalactosamine localizing the predominant disaccharide demonstrated biochemically. A previously unrecognized disaccharide consisting of terminal N-acetylneuraminic acid and penultimate beta-galactose was found in 20-30% of mucous acinar cells. Occasional clusters of acini composed purely of serous cells contained an additional unrecognized glyco-conjugate with oligosaccharides terminated by sialic acid with O-acetylated polyhydroxyl side chains and penultimate beta-galactose. Serous demilunes, however, lacked detectable complex carbohydrate other than glycogen. Terminal sialic acid-beta-galactose dimers were present on the apical surface of all ducts except for intercalated ducts coated only with neutral glycoconjugate. Fucose assayed biochemically as a minor component occurred in abundance in glycoconjugates at the apical surface of all intercalated and most striated duct cells and within some striated duct cells. Terminal alpha-galactose not previously detected biochemically was localized at the apex of all duct cells. These results provide new knowledge concerning the structure of ovine submandibular glycoconjugates. They also illustrate the value of histochemical methods for elucidating the diversity of complex carbohydrates in an organ, locating different glycoconjugates in different types or subtypes of epithelial cells and demonstrating intracellular sites that contain complex carbohydrate.

Animals↗
PubMed1985

Desmoplastic fibroma of the maxilla.

A case of desmoplastic fibroma of the right maxilla is reported. The lesion presented as a painless mass in the right posterior alveolar ridge of a 22-year-old white man. Histologically, the lesion was composed of interlacing fascicles of benign-appearing fibroblasts in a varying ground substance of collagenous and myxoid tissue. The lesion was not encapsulated histologically and was invading between bone trabeculae resulting in resorption of the bone. A review of the literature reveals that a total of 30 cases of desmoplastic fibroma of the jaws have been reported. All of the mandibular lesions except two were reported to have occurred in the left side, with the molar-ramus region favored. The case reported here is, to the best of our knowledge, the first reported case of desmoplastic fibroma occurring in the maxillary alveolar process.

Adult↗
PubMed1985

Normal pulmonary function measurements and airway reactivity in childhood after mild bronchiolitis.

Concern about the long-term sequelae of bronchiolitis has been raised through studies of children hospitalized for bronchiolitis, but the long-term sequelae of mild bronchiolitis have not been studied. We assessed the hypothesis that 25 children with mild bronchiolitis (index subjects) were at greater risk for abnormalities of pulmonary function or airway reactivity to cold air between the ages of 8 and 12 years than were randomly selected, matched controls. There were no consistent differences in pulmonary function or airway reactivity between index and control groups. Airway hyperreactivity was found in five control subjects and three index subjects, and all children with symptomatic asthma were identified by cold air challenge. Our data suggest that children with a history of mild bronchiolitis are not at increased risk between ages 8 and 12 years for airway hyperreactivity or for abnormalities in pulmonary function.

Adult↗
PubMed1984

Nucleoside hydrolases from Trypanosoma cruzi.

Four distinct nucleoside cleaving enzymes were detected in extracts of Trypanosoma cruzi epimastigotes. Two of these were nucleoside phosphorylases: one was specific for pyrimidines and the other for purines. The other two enzymes were nucleoside hydrolases. These hydrolytic enzymes were partially purified and their substrate and inhibitor specificities were studied. One of the hydrolases was designated inosine/guanosine hydrolase. It required purine substrates containing a 9-beta-D-ribofuranosyl substituent. Neither 2'-deoxyinosine, 2'-deoxyguanosine, nor hypoxanthine arabinoside served as substrates. The kinetic patterns obtained from combined product analysis and the mutual competitive inhibition of this enzyme by inosine and guanosine suggested that these substrates were cleaved at a common catalytic site. Inhibitor studies with several deoxyinosines have demonstrated the importance of the 2'-, 3'-, and 5'-hydroxyl groups for efficient binding to the enzyme. Adenosine, which did not serve as a substrate, was a potent competitive inhibitor (Ki = 8 microM) with respect to both inosine and guanosine. This enzyme had a particle weight of 106,000 +/- 10,000 as determined by Sephadex chromatography. The other hydrolase was specific for 2'-deoxyinosine. It only accepted purine substrates with a 6-oxo- and a 9-beta-D-2'-deoxyribofuranosyl substituent. Inosine, hypoxanthine arabinoside and a variety of deoxyinosines were not substrates nor did they strongly inhibit this enzyme. This enzyme had a particle weight of 19,000 +/- 2,000 as determined by Sephadex chromatography.

Animals↗
PubMed1984

Somatic cell hybrid mapping panels.

The recent advances in human gene mapping have been largely due to the development of interspecies cell hybrids containing human chromosomes and their fragments. The importance of characterized panels of these hybrid lines has grown exponentially with the application of recombinant DNA technologies to human genetics. In this article, we discuss current strategies employed in the construction of somatic cell hybrid mapping panels.

Animals↗
PubMed1984

Regulation of epithelial electrolyte transport by metabolites of arachidonic acid.

Intestine (both small and large) secretes chloride in response to prostaglandins and agents, such as bradykinin, that increase prostaglandin synthesis. The colon, but not the small intestine, can also be stimulated to secrete chloride by certain lipoxygenase products, such as 5-hydroperoxy-eicosatetraenoic acid and 5-hydroxy-eicosatetraenoic acid, but not by leukotrienes. Intestine generates these compounds and leukotriene B4 (but not C, D, and E) and their synthesis is increased by melittin, calcium ionophore A23187, and bradykinin.

Animals↗
PubMed1984

Human gingival fibroblast collagenase: purification and properties of precursor and active forms.

Human gingival fibroblast procollagenase has been purified to apparent homogeneity from serum-free and serum-supplemented fibroblast culture medium by a combination of ammonium sulfate precipitation, CM-cellulose chromatography, and gel-filtration on Bio-Gel P-150. Sodium dodecylsulfate polyacrylamide gel electrophoretic studies suggests that the purified fibroblast proenzyme is comprised of two closely related zymogens with the estimated Mr of 57,000 and 52,000. Upon densitometric scanning of the gels, the ratio of the two proenzyme forms was about 1 : 4 (57 : 52 kdal). Limited proteolysis of the fibroblast procollagenase with trypsin resulted in the conversion of both proenzyme forms into active enzyme forms of Mr 48,000 and 44,000, respectively. Amino acid analysis of the active enzymes and proenzyme forms revealed that the active enzymes contained fewer basic amino acids than do the proenzyme forms. The purified trypsin-activated fibroblast collagenase hydrolyzed type I collagen fibrils, cleaved tropocollagen in solution at 24 degrees C into TCA and TCB fragments, and cleaved the synthetic peptide substrate, DNP-peptide III, at the Gly-Ile bond. The gingival fibroblast collagenase exhibited a pH optimum of 7.5, was completely inhibited by EDTA or dithioerythritol but was not inhibited by N-ethylmaleimide or phenylmethylsulfonyl fluoride, and appeared to cleave human type III collagen approximately 10-fold faster than homologous type I collagen. In addition, comparison of the biochemical properties of the precursor and active forms of human gingival fibroblast collagenase with the precursor and active forms of human skin fibroblast collagenase, previously characterized by Stricklin and co-workers (Biochemistry 17: 2331-2337, 1978), revealed that they were similar in Mr, amino acid composition, and substrate specificity. Furthermore, the human gingival and skin fibroblast procollagenases were immunologically identical.

Amino Acids↗
PubMed1984

Intercompartmental transport in the Golgi complex is a dissociative process: facile transfer of membrane protein between two Golgi populations.

The transfer of the vesicular stomatitis virus-encoded glycoprotein (G protein) between Golgi populations in fused cells (Rothman, J. E., L. J. Urbani, and R. Brands. 1984. J. Cell Biol. 99:248-259) is exploited here to study and to help define the compartmental organization of the Golgi stack and to characterize the mechanism of intercompartmental transport. We find that G protein that has just received its peripheral N-acetylglucosamine in the Golgi complex of one cell is efficiently transferred to the Golgi complex of another cell to receive galactose (Gal). Remarkably, this transport occurs at the same rate between these two compartments whether they are present in the same or different Golgi populations. Therefore, a dissociative (presumably vesicular) transport step moves G protein from one part of the Golgi in which N-acetylglucosamine is added to another in which Gal is added. Minutes later, upon receiving Gal, the same G protein molecules are very poorly transferred to an exogenous Golgi population after cell fusion. Therefore, once this intercompartmental transfer has already taken place (before fusion), it cannot take place again (after fusion); i.e., transport across the compartment boundary in the Golgi complex that separates the sites of N-acetylglucosamine and Gal incorporation is a vectorial process. We conclude that transfers between Golgi cisternae occur by a stochastic process in which transport vesicles budding from cisternae dissociate, can diffuse away, and then attach to and fuse with the appropriate target cisterna residing in the same or in a different stack, based on a biochemical pairing after a random encounter. Under these circumstances, a transported protein would almost always randomize among stacks with each intercisternal transfer; it would not progress systematically through a single stack. Altogether, our studies define three sequential compartments in the Golgi stack.

Acetylglucosamine↗
PubMed1984

Inhibition of cellular alpha DNA polymerase and herpes simplex virus-induced DNA polymerases by the triphosphate of BW759U.

The triphosphate form of the acyclovir analog BW759U (9-[[2-hydroxy-1-(hydroxymethyl)ethoxy]methyl]guanine) inhibited the DNA polymerases (EC 2.7.7.7) from several strains of herpes simplex virus type 1. Two acyclovir triphosphate-resistant DNA polymerases were as sensitive to BW759U-triphosphate as were the DNA polymerases induced by wild-type viruses (Ki = 0.05 to 0.1 microM). The Ki value for cellular alpha DNA polymerase was 35- to 50-fold greater than those for the DNA polymerases induced by the various herpes simplex virus strains investigated. Incubation of Vero cells infected by the KOS strain of herpes simplex virus type 1 with the acyclovir analog resulted in the formation of substantial quantities of (9-[[2-hydroxy-1-(hydroxymethyl)ethoxy]methyl]guanine) triphosphate.

Acyclovir↗
PubMed1984

Standardized automated assay for functional alpha 1-antitrypsin.

We describe an enzymic assay for the functional activity of alpha 1-antitrypsin, in which tosyl-Gly-Pro-Lys-p-nitroanilide acetate (Chromozym PL) is the trypsin substrate. For the indicating reaction, the concentration of substrate is about double the Km of its reaction with trypsin. This assay is an improvement over reactions involving N-alpha-benzoyl-DL-arginine p-nitroanilide as substrate, the solubility of which in water is less than the Km of its reaction with trypsin. The assay is standardized in terms of active sites of trypsin inhibited per liter of serum, with p-nitrophenyl-p'-guanidinobenzoate as the active site titrant. The results agreed well with those by an immunonephelometric assay for alpha 1-antitrypsin (r = 0.953). The within-run and run-to-run precision was approximately 3%. The results of the assay varied linearly with alpha 1-antitrypsin concentration from 0 to 60 mumol/L of serum. The minimum sample size was 50 microL, and analysis of 24 samples took less than 20 min.

Aniline Compounds↗