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Biomedical subjects

P J McLaughlin

Publications and source records attributed to P J McLaughlin.

At least 19 recordsLinked to original sources
PubMed1992

Ontogeny of zeta (zeta), the opioid growth factor receptor, in the rat brain.

Opioid growth factor (OGF), [Met5]enkephalin, serves as an inhibitory influence on the developing nervous system and is especially targeted to cell proliferative events. OGF interacts with the zeta (zeta) opioid receptor to perform its function. Using [3H]-[Met5]enkephalin, the ontogeny of the zeta receptor in the whole brain and cerebellum of rats was explored. Specific and saturable binding was recorded at the earliest time sampled, prenatal day 15 (E15). In the whole brain, binding capacity (Bmax) was two-fold greater at E15 than at E18 and E20. The quantity of zeta receptor appeared to increase in the first postnatal week, reaching a maximum on postnatal day 8. Binding decreased the remainder of the 2nd week and between postnatal days 15 and 25 binding was no longer recorded. In the cerebellum, binding capacity increased from E20 to the 2nd postnatal week, reaching a maximum on postnatal days 8-10. The Bmax of the zeta receptor decreased precipitously on postnatal day 11, being 5.4-fold lower than on postnatal day 10. Between postnatal days 21 and 30, no binding was observed. The binding affinities of the whole brain and cerebellum were 2.3 and 2.7 nM, respectively, and no differences between ages could be detected. Continuous opioid receptor blockade from birth to postnatal day 6 increased body weight, the Bmax of the zeta receptor in the whole brain and cerebellum (but not the Kd), and increased the number of layers of germinal cells in the cerebellum.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Crystallization of the complex of actin with gelsolin segment 1.

Crystals of a 1:1 complex between human gelsolin segment 1 and actin have been grown from solutions containing polyethylene glycol 6000. The crystals are orthorhombic, space group P2(1)2(1)2(1); the axes are a = 57.4 A, b = 70.4 A, c = 184.5 A. They are moderately stable to X-rays and diffract to beyond 2.5 A. There is one molecule of complex in the asymmetric unit.

Actins↗
PubMed1992

Crystallization of human gelsolin.

Human gelsolin has been crystallized by microdialysis techniques to give single crystals that diffract to 3.5 A resolution. The crystals belong to space group P42(1)2 and have cell dimensions a = 175.0 A, c = 151.6 A. They contain two gelsolin molecules in the asymmetric unit.

Calcium-Binding Proteins↗
PubMed1992

An opioid growth factor regulates the replication of microorganisms.

An opioid growth factor (OGF), [Met5]-enkephalin, interacts with the zeta (zeta) opioid receptor to modulate development of eukaryotes. We have found that [Met5]-enkephalin, an endogenous opioid peptide serves to inhibit the growth of S. aureus. This effect on growth involves cell proliferative events and is under tonic control, since potent opioid antagonists accelerate cell replication. Both the OGF and zeta opioid receptor were associated with these microorganisms. Other opioid receptors (mu, delta and kappa) were not detected. OGF also controlled the growth of other bacteria: P. aeruginosa and S. marcesans. These results indicate that OGF and its receptor, known to be important in the regulation of mammalian development, also function in the growth of simple unicellular organisms. We suggest that the endogenous opioid system related to growth originated billions of years ago.

Enkephalin, Methionine↗
PubMed1992

Germ cell alkaline phosphatase in human seminal plasma following vasectomy.

Germ cell alkaline phosphatase (also termed placental-like alkaline phosphatase or Nagao isoenzyme) was assayed in seminal plasma using monoclonal antibody-based enzyme immunoassays in samples obtained both before and after vasectomy. In all cases, germ cell alkaline phosphatase fell to undetectable levels within 14 days of operation. A fall in seminal plasma germ cell alkaline phosphatase may thus be an early indication of successful vasectomy.

Alkaline Phosphatase↗
PubMed1991

Zeta (zeta), a growth-related opioid receptor in developing rat cerebellum: identification and characterization.

Endogenous opioids and opioid receptors (i.e. endogenous opioid systems) are expressed during neural ontogeny, and play a role in the development of the nervous system. Using [3H][Met5]-enkephalin, a potent ligand involved in neural growth, particularly cell proliferation, specific and saturable binding was detected in homogenates of 6-day-old rat cerebellum; the data were consistent with a single binding site. Scatchard analysis yielded a binding affinity (Kd) of 2.2 nM and a binding capacity (Bmax) of 22.3 fmol/mg protein. Binding was linear with protein concentration, dependent on time, temperature, and pH, and was sensitive to Na+, Mg2+, and guanyl nucleotides. Optimal binding required protease inhibitors, and pretreatment of the homogenates with trypsin markedly reduced binding, suggesting that the binding site was proteinaceous in character. The [Met5]-enkephalin binding site was an integral membrane protein located in the nuclear fraction. Competition experiments indicated that [Met5] enkephalin was the most potent displacer of [3H][Met5]-enkephalin, and that binding was stereospecific. In the adult rat cerebellum, non-opioid receptor binding of [3H][Met5]-enkephalin was recorded, mu and kappa receptors were also found in the developing rat cerebellum, while mu, delta, and kappa receptors were recorded in adult cerebellar tissue. The function, pharmacological and biochemical characteristics, subcellular distribution, and temporal expression of the [Met5]-enkephalin binding site suggest the presence of a unique opioid receptor, termed zeta (zeta), in the developing nervous system.

Animals↗
PubMed1991

Endogenous opioids regulate cell proliferation in the retina of developing rat.

The role of endogenous opioids and opioid receptors (endogenous opioid systems) in modulating cell proliferation in the developing mammalian retina was examined in 1-day-old rats. In contrast to a labeling index (LI) of 35.8% in control animals, administration of the opioid peptide [Met5]-enkephalin (100 micrograms/kg) significantly reduced (10.6%) the proportion of cells incorporating [3H]thymidine; concomitant injection of 1 mg/kg naloxone blocked the inhibitory effects of [Met5]-enkephalin on cell division. Naloxone (1 mg/kg) alone did not alter the LI. The interruption of endogenous opioid-opioid receptor interaction by naltrexone (50 mg/kg), a potent opioid antagonist, was accompanied by a significant increase (6.4%) in the LI relative to control levels. Immunocytochemical experiments revealed the presence of enkephalin-like immunoreactivity, with staining of the cortical cytoplasm of proliferating and differentiating retinal cells recorded; no immunoreactivity was noted in the adult retina. In vitro autoradiography using 125I-[Met5]-enkephalin indicated that [Met5]-enkephalin binding sites were localized to the developing retina; no binding of the radiolabeled ligand was recorded in the adult retina. These results demonstrate the presence of growth-related endogenous opioids and opioid receptors in the developing mammalian retina, but not in adult retina, and suggest that endogenous opioids serve as natural inhibitory trophic factors that tonically regulate cell proliferation.

Animals↗
PubMed1991

Identification of opioid peptides regulating proliferation of neurons and glia in the developing nervous system.

Endogenous opioid systems (i.e. opioids and opioid receptors) play a role in regulating neural development. Using the cerebellar cortex of 6-day-old rats, the most potent opioid peptides involved with cell proliferation were assessed. In both the external germinal (granule) layer (EGL), a germinative matrix giving rise to neurons, and the medullary layer (MED), a pool of cells that are the precursors of glia (astrocytes and oligodendrocytes), [Met5]enkephalin and peptide F were extremely potent in depressing the labeling index (LI) using [3H]thymidine and autoradiographic techniques; concentrations as low as 100 micrograms/kg reduced the LI of EGL cells by 24% and MED cells by 43%. This inhibition of DNA synthesis by opioid peptides was blocked by concomitant exposure to to naloxone, an opioid antagonist. Peptide action was apparent 2 h following drug administration, and concentrations of 80 micrograms/kg but not 1 or 10 micrograms/kg [Met5]enkephalin depressed the LI. These results identify a selective group of opioid peptides, derived from proenkephalin A, as the potent, natural, inhibitory factors targeted to cell proliferation of cells destined to be neurons and glia in the developing nervous system.

Animals↗
PubMed1991

Mental performance of air crew following layovers on transzonal flights.

Studies of transzonal travel indicate that desynchronization of performance and physiological rhythms occurs following long-haul flights. In an attempt to overcome the problems associated with such desynchronization, air crew are given layover periods en route. The extent to which such layovers are effective in removing performance deficit, however, has not been studied. The present study was designed to remedy this situation by investigating performance levels following layover periods on both eastward and westward long-haul flights. The results suggest that layovers may largely eliminate performance deficits but only if flight times are such that air crew may maintain a reasonably normal sleep-rest cycle.

Adult↗
PubMed1991

Evaluation of sequential plasma and urinary tumor necrosis factor alpha levels in renal allograft recipients.

The macrophage cytokine tumor necrosis factor-alpha is released early in immune activation and may be detected in the peripheral circulation. This study has investigated the occurrence of plasma and urinary TNF in 30 renal allograft recipients. Although circulating TNF may be detected in 20% of pretransplant or normal control samples, levels were significantly elevated during 65% of allograft rejection episodes. Plasma TNF levels did not rise in graft failure due to acute tubular necrosis, but were always highly raised in systemic infection. In contrast, urinary TNF was only detected in association with acute rejection (49%) or tubular necrosis (14%), and no controls had detectable urinary TNF. These findings indicate that evaluation of circulating and excreted TNF may give further insight into the immunobiology of graft rejection.

Communicable Diseases↗
PubMed1991

Immunolocalization of tumour necrosis factor alpha in lung tissue from patients dying with adult respiratory distress syndrome.

Using a mouse monoclonal antibody raised against recombinant human tumour necrosis factor alpha (TNF alpha), we have studied immunostained paraffin-embedded autopsy lung tissues from 13 patients dying with adult respiratory distress syndrome and 10 control patients dying of unrelated conditions. Material from 12 of the 13 cases showed positive staining, strong in six, less strong in the remaining six. TNF alpha was located principally within epithelial cells resembling type II pneumocytes in both early and late stages of adult respiratory distress syndrome. In the early stages the TNF alpha positive cells were scattered and appeared disorganized, but in later cases they formed an epithelium. The cells were confirmed as epithelial by their staining pattern with monoclonal antibodies reactive with epithelial, cytokeratin and macrophage antigens, and by double staining for epithelial membrane antigen and TNF alpha. In the control group, staining for TNF alpha was confined to very occasional plump type II pneumocytes, and a weak reaction in bronchial lining cells. Macrophages were only weakly stained in either group. It is suggested that TNF alpha is synthesized (or possibly absorbed) by type II pneumocytes and may be involved in the production of, or recovery from, adult respiratory distress syndrome.

Adult↗
PubMed1990

Crystal structure of ovalbumin as a model for the reactive centre of serpins.

The serpins are a widely distributed family of proteins with diverse functions; they include the key serine protease inhibitors of human plasma as well as noninhibitory homologues such as hormone-binding globulins, angiotensinogen and egg-white ovalbumin. Sequence alignment based on the crystal structure. On the cleaved form of the archetypal serpin, alpha 1-antitrypsin, indicates that the serpins share a common highly ordered structure. On cleavage of the reactive centre peptide bond, they characteristically undergo a remarkable conformational change, the newly generated C terminus moving some 70 A to the opposite pole of the molecule. The structure of this post-cleavage form is known, but the conformation of the intact serpins and in particular that of their reactive centre is not. Wright et al.'s structure of plakalbumin (ovalbumin cleaved by subtilisin) has provided evidence for the conformational change that results from cleavage. We have now determined the structure of native ovalbumin to 1.95 A resolution and have found that the intact peptide loop forming the analogue to the reactive centre of the inhibitory serpins takes the unexpected form of a protruding, isolated helix. This model of the intact structures of the serpins suggests how they may interact with their target proteases.

Amino Acid Sequence↗
PubMed1990

Adult and developing human cerebella exhibit different profiles of opioid binding sites.

The binding of [3H][D-Ala2,MePhe4,Gly-ol5]enkephalin (DAGO), [3H][D-Pen2,5]enkephalin (DPDPE), [3H]ethylketocyclazocine (EKC), and [3H][Met5]enkephalin (MET) was used to examine mu-, delta-, kappa-, and zeta-receptors, respectively, in the developing (birth to postnatal day 19) and adult human cerebellum. Specific and saturable binding of all ligands was recorded in developing brains, and of [3H]DAGO, [3H]DPDPE, and [3H]EKC in adult cerebellum; all data fit a single homogeneous binding site for each ligand. However, the ontogenic profile of opioid receptor subtypes differed. Delta- and kappa-receptor capacities were 7.8- and 3.6-fold, respectively, greater in infant cerebellum than in adults. The mu-receptor decreased over 7-fold in both binding affinity and capacity after day 2; by adulthood, the binding affinity was the same as in newborns but only one-half the binding capacity was recorded. The concentration of zeta-receptors was 20-fold greater in subjects 2-19 days of age than in newborns. These data demonstrate the presence, and distinct developmental profiles, of opioid receptors in human cerebellum. Although the function of mu-, delta-, and kappa-receptors in human cerebellum are unclear, the growth-related zeta-receptor is present at a time of cell replication and differentiation but is not detected in mature cerebellum.

Adult↗
PubMed1990

Demonstration and characterization of zeta (zeta), a growth-related opioid receptor, in a neuroblastoma cell line.

Endogenous opioids and opioid receptors (i.e. endogenous opioid systems) are involved in carcinogenesis. Using homogenates of S20Y neuroblastoma (NB) cells grown in culture, the binding of a growth-selective ligand, [Met5]enkephalin, was examined to ascertain the zeta (zeta) opioid receptor. Specific and saturable binding of [3H]-[Met5]enkephalin was detected in NB cells; the data were consistent with a single binding site. Scatchard analysis yielded a Kd of 1.6 nM and a binding capacity (Bmax) of 48.1 fmol/mg protein; 14,000 receptors per cell were estimated. Binding was dependent on protein concentration, time, temperature, and pH, and was sensitive to 100 nM, but not 5 nM, Na+, Ca2+, and Mg2+; GppNHp at concentrations of 100-500 mM had little effect on binding. Optimal binding required protease inhibitors, and pretreatment of the tumor cell homogenates with trypsin markedly reduced [3H]-[Met5]enkephalin binding, suggesting that the binding site was proteinaceous in character. Displacement experiments indicated that [Met5]enkephalin was the most potent displacer of [3H]-[Met5]enkephalin. Cell density (log, confluence, postconfluence) did not alter the Kd or Bmax. This study serves as the first demonstration and characterization of the zeta (zeta) opioid receptor in tissue culture cells. The homogeneous nature of NB cell cultures, along with the enrichment in receptor number, provides an excellent model system to isolate and purify the zeta receptor.

Animals↗
PubMed1990

Refined crystal structure of the phosphorylase-heptulose 2-phosphate-oligosaccharide-AMP complex.

The crystal structure of phosphorylase b-heptulose 2-phosphate complex with oligosaccharide and AMP bound has been refined by molecular dynamics and crystallographic least-squares with the program XPLOR. Shifts in atomic positions of up to 4 A from the native enzyme structure were correctly determined by the program without manual intervention. The final crystallographic R value for data between 8 and 2.86 A resolution is 0.201, and the overall root-mean-square difference between the native and complexed structure is 0.58 A for all protein atoms. The results confirm the previous observation that there is a direct hydrogen bond between the phosphate of heptulose 2-phosphate and the pyridoxal phosphate 5'-phosphate group. The close proximity of the two phosphates is stabilized by an arginine residue, Arg569, which shifts from a site buried in the protein to a position where it can make contact with the product phosphate. There is a mutual interchange in position between the arginine and an acidic group, Asp283. These movements represent the first stage of the allosteric response which converts the catalytic site from a low to a high-affinity binding site. Communication of these changes to other sites is prevented in the crystal by the lattice forces, which also form the subunit interface. The constellation of groups in the phosphorylase transition state analogue complex provides a structural basis for understanding the catalytic mechanism in which the cofactor pyridoxal phosphate 5'-phosphate group functions as a general acid to promote attack by the substrate phosphate on the glycosidic bond when the reaction proceeds in the direction of glycogen degradation. In the direction of glycogen synthesis, stereoelectronic effects contribute to the cleavage of the C-1-O-1 bond. In both reactions the substrate phosphate plays a key role in transition state stabilization. The details of the oligosaccharide, maltoheptaose, interactions with the enzyme at the glycogen storage site are also described.

Adenosine Monophosphate↗
PubMed1990

Ultrastructural localization of enkephalin-like immunoreactivity in developing rat cerebellum.

Methionine enkephalin, an endogenous opioid peptide, participates in the regulation of growth in the developing brain. In the present study, enkephalin-like immunoreactivity was localized in the cerebellum of developing and adult rats by immunoelectron microscopy. In 10-day-old animals, enkephalin-like immunoreactivity was found in the somata of proliferating, migrating and differentiating neural cells, and was associated with the plasma membrane, microtubules, filaments, mitochondria, endoplasmic reticulum and nuclear envelope. Both neurons and glia in the cerebellum of the preweaning rat displayed a similar profile of immunoreactivity. Reaction product was also detected in the dendrites and dendritic spines of Purkinje cells where it was concentrated in postsynaptic densities. The majority of internal granule neurons in 10-day-old animals were not immunoreactive, nor were axons, glial processes and postsynaptic elements (with the exception of mossy fiber terminals). At weaning (Day 21), enkephalin-like immunoreactivity was confined primarily to the somata of Purkinje, basket and stellate neurons, and to Purkinje cell dendrites and synaptic spines. Adult rats (day 75) exhibited no enkephalin-like immunoreactivity. These results establish that enkephalin or an enkephalin-like substance can be detected during the ontogeny of both neurons and glia in the cerebellar cortex, and appears to be associated with certain structural elements.

Aging↗