Biomedical subjects
Publications and source records attributed to S Jespersen.
We introduce a model of random layered media, extending the Matheron-de Marsily model: Here we allow for the flows to change in time. For such layered structures, we solve exactly the equations of motion for single particles and also for polymers modelled as Rouse chains. The results show a rich variety of dynamical patterns.
Recently, Watts and Strogatz introduced the so-called small-world networks in order to describe systems that combine simultaneously properties of regular and random lattices. In this work we study diffusion processes defined on such structures by considering explicitly the probability for a random walker to be present at the origin. The results are intermediate between the corresponding ones for fractals and Cayley trees.
Small-world networks (SWN), obtained by randomly adding to a regular structure additional links (AL), are of current interest. In this paper we explore (based on physical models) a new variant of SWN, in which the probability of realizing an AL depends on the chemical distance between the connected sites. We assume a power-law probability distribution and study random walkers on the network, focusing especially on their probability of being at the origin. We connect the results to Levy flights, which follow from a mean-field variant of our model.
Dissected tissue pieces of the pituitary pars intermedia from the amphibian Xenopus laevis was directly subjected to matrix-assisted laser desorption/ionization (MALDI) mass analysis. The obtained MALDI peptide profile revealed both previously known and unexpected processing products of the proopiomelanocortin gene. Mass spectrometric peptide sequencing of a few of these neuropeptides was performed by employing MALDI combined with postsource decay (PSD) fragment ion mass analysis. The potential of MALDI-PSD for sequence analysis of peptides directly from unfractionated tissue samples was examined for the first time for the known desacetyl-alpha-MSH-NH2 and the presumed vasotocin neuropeptide. In addition, the sequence of an unknown peptide which was present in the pars intermedia tissue sample at mass 1392.7 u was determined. The MALDI-PSD mass spectrum of precursor ion 1392.7 u contained sufficient structural information to uniquely identify the sequence by searching protein sequence databases. The determined amino acid sequence corresponds to the vasotocin peptide with a C-terminal extension of Gly-Lys-Arg ("vasotocinyl-GKR"), indicating incomplete processing of the vasotocin precursor protein in the pituitary pars intermediate of X. laevis. Both vasotocin and vasotocinyl-GKR are nonlinear peptides containing a disulfide (S-S) bridge between two cysteine residues. Interpretation of the spectra of these two peptides reveals three different forms of characteristic fragment ions of the cysteine side chain: peptide-CH2-SH (regular mass of Cys-containing fragment ions), peptide-CH2-S-SH (regular mass + 32 u) and peptide = CH2 (regular mass -34 u) due to cleavage on either side of the sulfur atoms.
We have previously shown that the melanotrope population of the pituitary intermediate lobe of Rana ridibunda is composed of two subpopulations, of low (LD) and high density (HD), that show distinct ultrastructural features and display different synthetic and secretory rates. To investigate whether LD and HD melanotrope cells also differ in proopiomelanocortin (POMC) processing, we have analyzed the POMC-end products in single cells from both subpopulations by means of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The mass spectra revealed the presence of 8 POMC-derived peptides in HD and LD melanotrope cells, indicating a similar processing of the precursor in both subpopulations. However, the relative abundance of three POMC-end products (i.e. lys-gamma1-MSH, acetyl-alpha-MSH, and CLIP fragment) was higher in the HD subset. Moreover, two peptides with molecular weights of 1030 and 1818 Da, respectively, were detected that could not be assigned to any product deduced from the frog POMC sequence. The relative amount of the 1030 Da peptide was higher in LD melanotrope cells. Taken together, our results suggest that POMC processing is differentially regulated in the two melanotrope cell subsets.
BACKGROUND: Although epidemiological studies suggest that people with low birthweight are at higher risk of coronary heart disease and stroke in adult life than those of higher birthweight, the mechanisms underlying this relation are unclear. We investigated whether impaired fetal growth leads to an increased risk of atherosclerosis by assessment of the carotid and lower-limb arteries in a group of people aged around 70 years for whom birthweight data were available. METHODS: 2232 singleton babies were born to married mothers at a maternity hospital in Sheffield, UK, between 1922 and 1926, and had complete birth records. We traced 829 survivors and invited 395 who still lived in Sheffield to take part in this study. 322 agreed; they were interviewed at home and 181 underwent duplex ultrasonographic examination of the extracranial parts of the carotid arteries. Atherosclerosis in the arteries of the lower limb was assessed in 186 people by the ankle-brachial-pressure index (ABPI). FINDINGS: The prevalence and severity of carotid atherosclerosis was greatest in those with the lowest recorded birthweight. The risk of carotid stenosis was greater for people who had weighed 6.5 lb or less at birth than for those who weighed over 7.5 lb (odds ratio 5.3 [95% CI 2.0-14.0]), after adjustment for cardiovascular risk factors and gestational age at birth. The odds ratio for atherosclerotic disease in the lower limbs (low ABPI) was highest in people with the lowest recorded birthweight, but this relation was not significant (odds ratio 2.3 [1.0-5.6]). INTERPRETATION: Increased atherogenesis may be one mechanism mediating the observed epidemiological link between impaired fetal growth and cardiovascular disease. Adaptations made by the fetus in response to influences that retard its growth may have long-term consequences for the structure and metabolism of the vascular system.
To explore the relation between reduced fetal growth and impaired glucose tolerance in adult life, an oral glucose tolerance test (75 g glucose) was carried out on 218 men and women, now aged around 50 years, who had been measured in detail at birth. Measurements of plasma concentrations of glucose and insulin were made at 0, 30, and 120 min. Fasting plasma concentrations of proinsulin and 32-33 split proinsulin were also measured. People in the highest category of birthweight tended to have the lowest plasma concentrations of insulin as adults at both 0 and 120 min, though both these relations were weak. Plasma insulin concentrations in adult life were more strongly related to abdominal circumference at birth than to birthweight. After adjusting for sex and body mass index, mean insulin concentrations at 0 min fell from 50 pmol l-1 to 46 pmol l-1 (p = 0.04) and at 120 min from 235 pmol l-1 to 144 pmol l-1 (p = 0.003) between people whose abdominal circumference at birth had been less than 11.5 in and those who abdominal circumference had been greater than 13 in. Plasma glucose concentrations at 120 min also fell with increasing abdominal circumference at birth. Because abdominal circumference at birth is an indicator of the growth of the liver in fetal life, one interpretation of these findings is that the sensitivity of the liver to insulin is permanently reduced if the intrauterine development of this organ is impaired.
A number of potential matrix candidates were investigated with regard to the importance of the pH in the matrix-assisted laser desorption/ionization mass spectrometry (MALDI/MS) analysis of non-covalently bound protein complexes. The matrices examined were 2,5-dihydroxybenzoic acid (DHB), 4-hydroxy-alpha-cyanocinnamic acid (HCCA), 2-aminonicotinic acid (ANA), 4-nitroaniline (NA), 2-amino-4-methyl-5-nitropyridine (AMNP) and 3-hydroxypicolinic acid (HPA). In solution these matrix compounds permitted the preparation of MALDI samples at pH in the range 2-7. Among the matrices tested, complex formation, by specific non-covalent interactions, could only be observed when HPA (pH 3.8) was used as the matrix for the MALDI analysis. Under these conditions, specific non-covalent complex formation of recombinant streptavidin and glutathione-S-transferases were observed but not for human hemoglobin. The MALDI spectra obtained with the neutral compounds ANA (pH 4.4), NA (pH 6.4) and AMNP (pH 7.1) as matrices contain only peaks of the subunit with no signal of the non-covalent bound complexes present. Considering the results obtained in this study with basic and acidified matrix materials, there does not appear to be a strong correlation between the pH of the matrix solution and the utility of a matrix for the analysis of non-covalently bound complexes.
The use of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) in identifying proopiomelanocortin (POMC) processing products in melanotrope cells of the pituitary intermediate lobe of Xenopus laevis was explored. Mass spectra were obtained with such a high sensitivity of detection that the peptides could be identified in a single melanotrope cell. In addition to known POMC processing products of the Xenopus melanotrope cell, the presence of previously unidentified POMC-derived peptides was demonstrated. Together these POMC processing products accounted for the entire length of the POMC precursor. Furthermore, Xenopus possesses two genes for POMC and the sensitivity and accuracy of the MALDI-MS technique allowed identification of processing products of both the POMCA and POMCB gene. In addition, differences were obtained between the mass spectra of melanotrope cells from Xenopus laevis adapted to different conditions of background illumination. These results show that MALDI-MS is a valuable tool in the study of the expression of peptides in single (neuroendocrine) cells.
The irreversible and reversible inhibition of glutathione S-transferases (GSTs) by eugenol was studied in rat, mouse and man. Using liver cytosol of human, rat and mouse, species differences were found in the rate of irreversible inhibition of GSTs by eugenol in the presence of the enzyme tyrosinase. Tyrosinase was used to oxidize eugenol. No inhibition was observed in the absence of tyrosinase. The rate of irreversible inhibition of GSTs was highest in mouse cytosol, and lowest in rat cytosol. In addition, the irreversible inhibition of human and rat GSTs by eugenol was studied using purified isoenzymes of man and rat. The human GST isoenzymes A1-1, M1a-1a and P1-1 and the rat GST isoenzymes 1-1, 2-2, 3-3, 4-4 and 7-7 were irreversibly inhibited by eugenol in the presence of tyrosinase. In this respect human GST P1-1 and rat GST 7-7 were by far the most sensitive enzymes; human GST A2-2 was not inhibited. Indications were found that human GST P1-1 may be inhibited via three mechanisms: in addition to the well documentated nucleophilic addition of quinones and oxidation of cysteine residues, a covalent subunit cross-linking was also observed. The reversible inhibition of human and rat GST by eugenol, eugenol methyl ether, isoeugenol methyl ether, 2-allylphenol and 4-propylphenol was also studied using purified isoenzymes. The reversible inhibition of human and rat GSTs, using 1-chloro-2,4-dinitrobenzene as substrate, was expressed as I25. All compounds caused moderate reversible inhibition (I25 ranged from 0.2 to 5.4 mM for human GSTs and from 0.4 to 4.9 mM for rat GSTs). In rat, eugenol methyl ether was the strongest inhibitor. In human, the overall inhibiting capacities of eugenol, eugenol methyl ether, isoeugenol methyl ether and 4-propyl phenol were more or less similar; 2-allylphenol was the poorest inhibitor.
A mass spectrometric method providing qualitative site-specific information regarding covalent modification of proteins is described. The method involves comparison of unmodified and modified proteins by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) peptide mapping in combination with site-specific mutagenesis of possible target amino acids. The approach is demonstrated through the mapping of glutathione-S-transferases (GSH transferases) before and after inhibition with the glutathione conjugate 2-(S-glutathionyl)-3,5,6-trichloro-1,4-benzoquinone (GSTCBQ). The results demonstrate the utility of site-specific mutagenesis in combination with MALDI MS peptide mapping. Evidence is presented that three residues in or near the active site, including the hydroxyl groups of Tyr6 and Tyr115 and the sulphydryl group of Cys114, are target sites for GSTCBQ. Although only one GSTCBQ molecule per active site was detected, it appears to be distributed among all three target sites. In addition, MALDI MS peptide mapping covered 81% of the cDNA deduced amino acid sequence for GSH transferase and site-directed mutagenesis corresponding to a single amino acid substitution were verified.
High molecular weight precursors of insulin-like growth factor II (IGF-II) were isolated from Cohn fraction IV of human plasma by ultrafiltration, affinity chromatography and reversed-phase high-performance liquid chromatography. Molecular weight determination by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of two high molecular weight IGF-II preparations revealed heterogeneous glycosylation. A combination of enzymatic degradation and MALDI-MS were applied for further structural characterization of the glycosylated precursors of IGF-II. The first step was molecular weight determination of intact high molecular weight IGF-IIs prior to and after treatment with neuraminidase and O-glycosidase. This, together with a comparison of molecular weight information available from the cDNA, revealed that both high molecular weight IGF-II species contain an identical C-terminal extension of 20 residues but different degrees of glycosylation. Second, comparative Endo Glu-C digestion of the preparations prior to and after enzymatic release of carbohydrates and subsequent remeasurement of the molecular weight by MALDI-MS confirmed the primary structure of precursor IGF-II1-87. The O-linked carbohydrates were found to be associated with the C-terminal extension and the heterogeneity was identified as varied sialylated forms of one and two HexNAc-Hex groups.
An innovative method, direct peptide profiling of small samples of nervous tissue by matrix-assisted laser desorption ionization mass spectrometry, in combination with peptide characterization, immunocytochemistry in conjunction with specific neuronal labeling by backfilling of the penis nerve, and bioassay of peptides was used to study the intrinsic neuronal expression patterns of distinct sets of related FMRFa peptides and their significance for the organization of male copulation behavior in the mollusk, Lymnaea stagnalis. Previous studies indicate that the sets of FMRFa-related and GDPFLRFa-related peptides are encoded by two alternatively spliced transcripts of the single FMRFa gene. Direct mass spectrometry revealed that both FMRFa-related and GDPFLRFa-related peptides are present in the penis nerve, the sole nerve that innervates the penis complex. Accordingly, authentic FMRFa, GDPFLRFa, and related peptides were purified from the penis complex. The loci of synthesis of FMRFa and related peptides could be traced to the right cerebral ventral lobe, those of GDPFLRFa and related peptides to the B group neurons in the right parietal ganglion and to a few unidentified neurons in the right pleural ganglion. Notwithstanding their related structures, the two sets of peptides have distinctly different actions on the penis retractor muscle.
OBJECTIVES: To examine the relation between disproportionate fetal growth and adult blood pressure and to investigate whether arterial compliance in adult life is related to early development. DESIGN: A follow up study of a group of men and women whose birth weights and other measurements of body size had been recorded at birth. SETTING: Home and outpatient study. SUBJECTS: 337 men and women born in the Jessop Hospital, Sheffield, between 1939 and 1940. MAIN OUTCOME: Adult systolic and diastolic blood pressures and arterial compliance as measured by pulse wave velocity in two arterial segments. RESULTS: Both systolic and diastolic blood pressures were higher in people whose birth weight was low, who were short or who had small abdominal or head circumferences at birth. Systolic blood pressure decreased by 2.7 mm Hg (95% CI 0.8 to 4.6) for each pound (454 g) gain in birth weight and by 3.4 mm Hg (95% CI 1.4 to 5.4) for each inch (2.54 cm) increase in crown-heel length. Diastolic pressure fell by 1.9 mm Hg (95% CI 0.9 to 2.9) for each pound (454 g) gain in birth weight and by 2.4 mm Hg (95% CI 1.4 to 3.5) for each inch (2.54 cm) increase in length. Systolic blood pressure was also higher in individuals whose mother's intercristal pelvic diameter was small or whose mother's blood pressure had been raised during pregnancy but these effects were statistically independent of the effects of low birth weight and other measurements that indicate fetal growth retardation. Arterial compliance was lower in those who had been small at birth. CONCLUSION: Impairment of fetal growth is associated with raised blood pressure in adult life and decreased compliance in the conduit arteries of the trunk and legs.
The mode of inactivation of glutathione S-transferase isoenzyme 3-3 from rat by the active site-directed inhibitor 2-(S-glutathionyl)-3,5,6-trichloro-1,4-benzoquinone (GSTCBQ) has been investigated by a combination of site-specific mutagenesis and mass spectrometric analysis of the sites of reaction of the reagent with the enzyme. This very reactive reagent is shown to target 3 residues in or near the active site, including the hydroxyl groups of Tyr-6 and Tyr-115 and the sulfhydryl group of Cys-114. Although the covalent attachment of one 2-(S-glutathionyl)dichloro-1,4-benzoquinonyl group/active site is sufficient to inactivate the enzyme ( < 5% residual activity), the 1 mol of reagent appears to be distributed among all three target sites. Mutant enzymes in which the reactive functional groups of these 3 residues have been individually removed remain susceptible to GSTCBQ. Evidence from amino acid sequencing and peptide maps visualized by matrix-assisted laser desorption/ionization mass spectrometry suggests that both Tyr-6 and Tyr-115 are primary targets of the reagent in the native enzyme. Docking of a model of GSTCBQ in a model of the active site derived from the crystal structure of the enzyme indicates that the trichlorobenzoquinonyl group can be positioned so that both tyrosine hydroxyl groups can act as nucleophiles to add to the reagent or alternatively act as electrophiles to assist in the nucleophilic addition of the other. The reaction of GSTCBQ with Cys-114 appears to require a conformation different from that in the crystal structure.
The complete primary structures of two variants of a protein, Abd-5, isolated from the endocuticles of the migratory locust Locusta migratoria and the desert locust Schistocerca gregaria, have been determined. The proteins from the two species are N-terminally blocked with pyroglutamic acid. Their sequences differed only in two positions. Comparison of the sequences to those of other cuticular proteins shows that moderate homologies exist to 11 other cuticular proteins from insects representing four different orders. Amino acid residues in certain positions appear to be strictly conserved.
The parameters for recovery of sample from the nitrocellulose support used in plasma desorption are optimized. The losses in washing, in situ reactions, and extraction procedures are quantitatively evaluated. At least 80% of the sample can be effectively extracted or transferred to a polyvinyl difluoride membrane with 2-propanol-water mixture in the range 1:1-2:3 v/v. Quantitative losses of insulin during washing procedures vary from 0 to 50% depending on the washing procedure used. The losses in in situ reactions are negligible. Optimization of the procedures allows several successive procedures to be carried out after adsorption of 1 nmol of insulin on the nitrocellulose support. These include in situ reaction, extraction of A and B chains followed by S-alkylation, chain separation by high-performance liquid chromatography, mass spectrometric analysis of the separated chains, and finally automatic sequence after transfer to the sequenator.
Explore the source record for details and available documents.