Biomedical subjects

M J Glimcher

Publications and source records attributed to M J Glimcher.

At least 109 records · Page 6Linked to original sources

PubMed1986

Resolution and identification of O-phosphoserine, O-phosphothreonine, O-phosphotyrosine, and gamma-carboxyglutamic acid as their fluorescent o-phthalaldehyde derivatives by high performance liquid chromatography.

High performance liquid chromatography was used to resolve O-phosphoserine, O-phosphothreonine, and O-phosphotyrosine as their fluorescent o-phthalaldehyde derivatives. By adjusting the buffer system, very small amounts of O-phosphothreonine could be detected and quantitated in the presence of very large amounts of O-phosphoserine. In addition, gamma-carboxyglutamic acid and glutamic acid were also separated and quantitated. Depending on the buffer used, various combinations of these amino acids could be resolved in a single run.

1-Carboxyglutamic Acid↗

PubMed1986

Proline hydroxylation of collagens synthesized at different temperatures in vivo by two poikilothermic species.

Extent of prolyl hydroxylation in newly synthesized viper collagen is decreased at 10 degrees C to approximately 23% of normal on skin and to approximately 57% of normal in bone collagen. At 20 degrees C, prolyl hydroxylation is approximately 50% of normal in skin and normal in bone. At 10 degrees C and 20 degrees C, prolyl hydroxylation is decreased approximately 32% in the skin collagen of carp. In contrast, prolyl hydroxylation is unchanged at 10 and 20 degrees C in bone, scale and lepidotrichia. Prolyl hydroxylation of cartilaginous endoskeleton showed an approximately 25% decrease at 20 degrees C.

Acclimatization↗

PubMed1985

Reaction products formed after strong acid treatment of alpha-amino-delta-hydroxyvaleric acid.

High-voltage paper electrophoresis and analytical and preparative amino acid ion-exchange chromatography were used to investigate the products resulting from strong acid treatment and strong acid treatment and subsequent alkalinization of alpha-amino-delta-hydroxyvaleric acid. After strong acid treatment, alpha-amino-delta-hydroxyvaleric acid and proline, as well as several unidentified components, presumably lactones, were recovered. No definitive identification of the putative unstable lactones was accomplished either by physical-chemical means or by ion-exchange chromatography or high-voltage paper electrophoresis. Nevertheless, after acid treatment and alkalinization, only alpha-amino-delta-hydroxyvaleric acid and proline were recovered, in approximately equal amounts. The formation of such large amounts of proline from the acid treatment and alkalinization of alpha-amino-delta-hydroxyvaleric acid is therefore an important consideration in quantitating the number of gamma-glutamyl phosphate residues present in various proteins.

Acids↗

PubMed1985

Structural studies of phosvitin in solution and in the solid state.

Phosvitin, a highly phosphorylated glycoprotein, represents the major fraction of hen egg yolk phosphoproteins. Circular dichroism, Fourier transform infrared spectroscopy, and Fourier transform infrared photoacoustic and fluorescence spectroscopic methods were employed to determine the secondary structure of the protein in both the solid and solution phases. This was supplemented by a Chou-Fasman type of predictive algorithm for the first 25 residues at the N terminus of the dephosphorylated protein. A three-compartment model consisting of alpha-helical, beta-sheet, and beta-turn components with beta-turns occurring at the interface between alpha-helical and beta-sheet regions in the proximity of O-phosphoserine residues is suggested from the combined analyses. Beta-sheets appear to be the dominant secondary structural component in phosvitin in the solid and solution phases. The suggested model bears many similarities to other phosphoproteins reported in the literature. The secondary structure of phosvitin is observed to be sensitive to environmental factors as previously reported although the present studies differ in some respects from earlier results. Preliminary results suggest that Ca2+ ions trigger a decrease in beta-sheet structure at pH 2.

Amino Acid Sequence↗

PubMed1985

Synthesis and degradation in vivo of a phosphoprotein from rat dental enamel. Identification of a phosphorylated precursor protein in the extracellular organic matrix.

The cellular enamel organ and the cell-free organic matrix of developing enamel of female rats injected intravascularly with [3H]serine and [3H]proline were extracted in a number of solvents and examined by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and h.p.l.c. in 6M-guanidinium chloride at intervals varying from 5 min to 1 week after injection. Three major species soluble in NH4HCO3 with Mr values of approx. 100 000, 25 000 and 11 000 were identified in the cellular enamel organ. The Mr 100 000 and 11 000 components were not secreted but remained intracellular for periods of up to 1 week after injection of the radioactively labelled amino acids. In contrast, the Mr 25 000 species was secreted from the cells and was first detected in the extracellular organic matrix approx. 15-30 min after injection. With time, labelled components, first of Mr approx. 11 000 and subsequently approx. 6500, were detected in the organic matrix concomitant with a relative decrease in the Mr 25 000 component, demonstrating that the lower Mr species were derived from degradation of the putative extracellular precursor protein (Mr 25 000). All of the extracellular components were found to contain O-phosphoserine. No radioactively labelled component with an Mr greater than approx. 25 000, either an amelogenin or an enamelin, was observed in the extracellular organic matrix or in an intracellular component which subsequently was lost from the intracellular pool. The Mr of the highest Mr protein or class of proteins is calculated to be approx. 22 000-26 000 when standard proteins are used as markers, but only 15 000-18 000 when using the CNBr peptides of alpha 1 chains of rat tail tendon collagen as markers.

Amino Acids↗

PubMed1985

Identification and quantitation of O-phosphoserine in human plasma fibronectin.

O-Phosphoserine was positively identified as the phosphorylated moiety in human plasma fibronectin by 31P-NMR of intact peptides. These data correlated completely with chemical analyses which demonstrated the presence of O-phosphoserine at a concentration of 2 residues/molecule. Neither O-phosphothreonine nor O-phosphotyrosine was detected in partial acid and partial alkaline hydrolysates, respectively.

Fibronectins↗

PubMed1985

Relative molecular mass determination of a major, highest relative molecular mass extracellular amelogenin of developing bovine enamel.

Proteins of developing bovine enamel were fractionated by molecular sieving and ion-exchange chromatography. The major fraction corresponding to the highest Mr amelogenin of Mr approximately 26 000-30 000 was isolated and its Mr determined by SDS-PAGE, molecular sieving on G-100 resin and high performance liquid chromatography and by sedimentation-equilibrium ultracentrifugation, the latter three procedures in guanidine hydrochloride. SDS-PAGE and HPLC molecular sieving, employing commonly used Mr standards, gave Mr values of approximately 22 000-26 000. SDS-PAGE and HPLC molecular sieving, using proline-rich CNBr peptides of collagen as standards, and sedimentation-equilibrium ultracentrifugation, gave Mr values of approximately 15 000-18 000 and approximately 17 385, respectively. These latter values correspond well with those reported earlier and with the Mr of the major amelogenin computed from recent amino acid sequence data (approximately 19 000). It is concluded that the recently described, highest Mr amelogenin of Mr = 26 000-30 000 is not a new component but is identical to the proline-rich components having relative molecular masses ranging from 15 000 to 18 000 described much earlier by several groups of workers.

Amelogenin↗

PubMed1985

Accumulative cell stress: the multifactorial etiology of idiopathic osteonecrosis.

Most cases of idiopathic osteonecrosis are caused by many factors and rarely by an instantaneous, sudden, or solitary phenomenon. The cells in bony tissues become progressively sick or ill and therefore stressed. Depending on the condition, the cell sickness may intensify as the underlying disease progresses or as other factors are added that further decompensate the cells to the point at which they can no longer maintain cellular equilibrium. In many diseases, the final stress that overwhelms the cell is a course of corticosteroid therapy that could cause necrosis by several mechanisms, including increased stores of marrow fat, fat embolization, intraosseous hypertension, and a direct cellular toxic effect (see Fig. 3). The reasons why bone cells within the femoral head and condyles, humeral head, and certain other anatomic locations are especially susceptible remain obscure. However, in each of these sites, the blood supply enters from one direction only, and this relative ischemia could limit the reparative abilities of the cells.

Adrenal Cortex Hormones↗

PubMed1985

Pathogenesis of idiopathic osteonecrosis: the ubiquitous crescent sign.

The repair of a large subchondral osteonecrotic lesion follows a precise sequence of biologic events. The repair of osteonecrosis becomes clinically significant with the development of a subchondral fracture manifested radiographically as the crescent sign. Once the subchondral fracture occurs, collapse and subsequent degenerative arthritis are generally inevitable. The repair process, radiologic appearance, and clinical features (pain and loss of function) are identical regardless of the site of the disease.

Femur Head Necrosis↗

PubMed1984

Solid-state phosphorus-31 nuclear magnetic resonance studies of synthetic solid phases of calcium phosphate: potential models of bone mineral.

Phosphorus-31 NMR spectra have been obtained from a variety of synthetic, solid calcium phosphate mineral phases by magic angle sample spinning. The samples include crystalline hydroxyapatite, two type B carbonatoapatites containing 3.2 and 14.5% CO3(2-), respectively, a hydroxyapatite in which approximately 12% of the phosphate groups are present as HPO4(2-), an amorphous calcium phosphate, monetite, brushite, and octacalcium phosphate. Spectra were observed by the standard Bloch decay and cross-polarization techniques, as well as by a dipolar suppression sequence, in order to distinguish between protonated and unprotonated phosphate moieties. The spectra of the synthetic calcium phosphates provide basic information that is essential for interpreting similar spectra obtained from bone and other calcified tissues.

Bone and Bones↗

PubMed1984

Investigation of the mineral phases of bone by solid-state phosphorus-31 magic angle sample spinning nuclear magnetic resonance.

Phosphorus-31 magic angle sample spinning NMR spectra have been employed to investigate the structure and composition of the mineral deposits in chicken bone. Three different pulse sequences, Bloch decay, cross-polarization, and dipolar suppression, were employed to obtain spectra from bone specimens of varying age. These were compared to the spectra obtained from a variety of crystalline and noncrystalline synthetic calcium phosphate solids used as reference standards. The results suggest that the most suitable model for the major solid calcium phosphate mineral phase in bone is a hydroxyapatite containing approximately 5-10% CO32- and approximately 5-10% HPO42- groups, the latter in a brushite-like configuration. From the NMR line shapes it was deduced that the fraction of HPO42- groups was highest in the youngest bone and decreased progressively with increasing age of the specimen.

Animals↗

PubMed1984

Recent studies of the mineral phase in bone and its possible linkage to the organic matrix by protein-bound phosphate bonds.

The most widely accepted hypothesis to account for maturational changes in the X-ray diffraction characteristics of bone mineral has been the 'amorphous calcium phosphate theory', which postulates that an initial amorphous calcium phosphate solid phase is deposited that gradually converts to poorly crystalline hydroxyapatite. Our studies of bone mineral of different ages by X-ray radial distribution function analysis and 31P n.m.r. have conclusively demonstrated that a solid phase of amorphous calcium phosphate does not exist in bone in any significant amount. 31P n.m.r. studies have detected the presence of acid phosphate groups in a brushite-like configuration. Phosphoproteins containing O-phosphoserine and O-phosphothreonine have been isolated from bone matrix and characterized. Tissue and cell culture have established that they are synthesized in bone, most likely by the osteoblasts. Physiochemical and pathophysiological studies support the thesis that the mineral and organic phases of bone and other vertebrate mineralized tissues are linked by the phosphomonester bonds of O-phosphoserine and O-phosphothreonine, which are constituents of both the structural organic matrix and the inorganic calcium phosphate crystals.

Aging↗

PubMed1984

Phosphoproteins of chicken bone matrix. Proof of synthesis in bone tissue.

Twelve-day-old embryonic chick mandibles were cultured in vitro for 6 days. Measurements of the weights of the explants, their mineral and protein components, and the EDTA-extractable proteins established that bone tissue synthesizes O-phosphoserine- and O-phosphothreonine-containing phosphoproteins which are similar to those present in embryonic and postnatal chicken bone matrix. The synthesis of the phosphoproteins was further confirmed by the demonstration that radioactively labeled O-phosphoserine and O-phosphothreonine were identified in bone and in the EDTA-extractable phosphoproteins after pulse-labeling chick mandibles in vitro with radioactively labeled serine and threonine, respectively.

Animals↗

PubMed1984

Failure to detect an amorphous calcium-phosphate solid phase in bone mineral: a radial distribution function study.

X-ray diffraction radial distribution function analysis was used to determine if a significant amount of an amorphous solid phase of calcium phosphate exists in bone, and if so, whether the amount varies as a function of age and maturation. Unfractionated cortical bone from embryonic and posthatch chicks of various ages and a low-density fraction of embryonic bone were studied. No evidence was found for the presence of an amorphous solid phase of calcium phosphate in any of the samples studied, including the recently deposited bone mineral of the low density fraction of embryonic bone. As little as 12.5% of synthetic amorphous calcium phosphate (ACP) added to bone was readily detected by the radial distribution function technique used. The results clearly indicate that the concept that ACP is the initial solid mineral phase deposited in bone, and the major mineral constituent of young bone is no longer tenable. The concept does not provide an accurate description of the nature of the initial bone mineral deposited, or the changes that occur with maturation, nor can it account for the compositional and X-ray diffraction changes that the mineral component undergoes during maturation and aging.

Age Factors↗

PubMed1984

Failure to detect crystalline brushite in embryonic chick and bovine bone by X-ray diffraction.

We have reexamined the question of whether brushite (CaHPO4 X 2H2O) occurs in embryonic bone. On the basis of the experiments reported here, and a reexamination of data previously obtained in this laboratory, we have concluded that crystalline brushite as a separate phase identifiable by X-ray diffraction does not occur in embryonic chick or bovine bone. Crystalline brushite previously identified in this laboratory in the less-mineralized fractions of embryonic chick bone was apparently formed artifactually during the fractionation process, possibly from preexisting domains of HPO4(-2) groups in a noncrystalline brushite-like configuration.

Animals↗

PubMed1984

Radioautographic visualization and biochemical identification of O-phosphoserine- and O-phosphothreonine-containing phosphoproteins in mineralizing embryonic chick bone.

We injected NaH2(33)PO4 into normal 14-d-old embryonic chicks and examined the long bones by both radioautography and biochemical analyses from 10 to 240 min after the injection was completed. At 30 min, determination of the radiographic grain density revealed that 33P was concentrated principally in fibroblasts, preosteoblasts, and osteoblasts. With time, there was a progressive increase in the density of silver grains located over both the osteogenic cells and the regions of uncalcified (osteoid) and calcified extracellular organic matrices. Biochemical analyses identified 33P-O-phosphoserine as the major 33P component in glutaraldehyde-treated whole demineralized bone tissue and in EDTA-soluble, nondiffusible proteins extracted from the bones, both at the same time periods that 33P-induced silver grains were visualized by radioautography. 33P-O-phosphothreonine was also identified in experiments using a dosage of 10 mCi per embryo. The results provide the first combined direct biochemical and radioautographic identification that phosphoproteins are synthesized in bone and are located morphologically at the sites of mineralization. The data provide further evidence that phosphoproteins play a critical role in the biological calcification of vertebrate tissues.

Animals↗

PubMed1984

Is the chemistry of collagen in intervertebral discs an expression of Wolff's Law? A study of the human lumbar spine.

The collagen content, proportion of Types I and II collagen, and the relative concentrations of the reducible crosslinks of human lumbar intervertebral discs have been found to vary with age and location and to be highly dependent on the topography of the tissue. From adolescence to mature adulthood, the most significant change is an increase in the content of Type I collagen at the expense of genetically distinct Type II collagen in the outer lamella of the posterior quadrant, while just the reverse is true of the anterior quadrant. These changes are accompanied by similar but smaller alterations in the total collagen content and in the crosslink hydroxylysinohydroxynorleucine . The same differences in the distribution of Types I and II collagens occur in the annuli on the concave and convex sides of scoliotic curves. Together, these data establish that active cellular activity and tissue remodelling occur in the annuli fibrosi and suggest that these specific changes are initiated in response to overall increases in compressive loading on the concave side and tensile loading on the convex side of the spine and the subsequent changes they induce in the magnitude and distribution of internal stresses within the annuli. In its most general formulation, the biological behavior of annuli fibrosi to mechanical forces appears to follow Wolff's Law.

Adolescent↗

PubMed1983

Phosphoproteins of bovine dentin: evidence for polydispersity during tooth maturation.

The ethylenediaminetetraacetic acid extractable phosphoproteins of dentin from very young fetal calves consisted principally of a monodisperse species with an apparent molecular weight of at least 94 000 and a phosphorus content of 8.0% (w/w). In contrast, the soluble phosphoproteins extracted from mature adult animals under identical conditions ranged in molecular weight from 18000 to 94000 and contained less phosphorus. The data support the conclusion that the phosphoproteins of bovine dentin are degraded in vivo during mineralization and maturation of the tissue.

Adult↗