Comment on: Neurotoxicity of prion peptide 106-126 not confirmed by Beat Kunz, Erika Sandmeier, Philipp Christen, FEBS letters 458 (1999) 65-68.
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Cells, Cultured↗
Biomedical subjects
D R Brown
Publications and source records attributed to D R Brown.
Potentiation of anaphylaxis in guinea pig ileal mucosa by a selective delta-opioid receptor agonist.
Immediate hypersensitivity reactions in the intestinal mucosa evoke active chloride secretion which enhances the elimination of luminal antigens. The prosecretory actions of histamine and other soluble mediators of anaphylaxis are mediated by submucosal neurons, as are the antisecretory actions of opioid antidiarrheal medications. We tested the hypothesis that the selective delta-opioid receptor agonist [D-Pen2, D-Pen5]enkephalin (DPDPE) alters anaphylaxis-associated ileal anion secretion in vitro. Sheets of ileal mucosa with attached submucosa from guinea pigs sensitized to cow's milk were mounted in Ussing chambers under short-circuit conditions. Mucosal sheets responded to the serosal application of the milk protein, beta-lactoglobulin, with a rapid rise in transepithelial short-circuit current (Isc); in contrast, the egg protein, ovalbumin, was without effect. Pretreatment of tissues with the neuronal conduction blocker, saxitoxin, or the H1 histamine receptor antagonist, diphenhydramine, but not the opioid receptor antagonist, naloxone, significantly reduced mucosal responses to antigen. [D-Pen-2, D-Pen5]enkephalin (0.1 microM, serosal addition) decreased baseline Isc, but potentiated mucosal responses to antigen; its effects were abolished in tissues pretreated with naloxone. These results suggest that immediate hypersensitivity reactions in the guinea pig ileal mucosa are mediated by submucosal neural circuits that are phasically modulated by both mast cell products and opioids.
Analgesics↗
Association of herpesvirus with fibropapillomatosis of the green turtle Chelonia mydas and the loggerhead turtle Caretta caretta in Florida.
Sea turtle fibropapillomatosis (FP) is a disease marked by proliferation of benign but debilitating cutaneous fibropapillomas and occasional visceral fibromas. Transmission experiments have implicated a chloroform-sensitive transforming agent present in filtered cell-free tumor homogenates in the etiology of FP. In this study, consensus primer PCR methodology was used to test the association of a chelonian herpesvirus with fibropapillomatosis. Fibropapilloma and skin samples were obtained from 17 green and 2 loggerhead turtles affected with FP stranded along the Florida coastline. Ninety-three cutaneous and visceral tumors from the 19 turtles, and 33 skin samples from 16 of the turtles, were tested. All turtles affected with FP had herpesvirus associated with their tumors as detected by PCR. Ninety-six percent (89/93) of the tumors, but only 9% (3/33) of the skin samples, from affected turtles contained detectable herpesvirus. The skin samples that contained herpesvirus were all within 2 cm of a fibropapilloma. Also, 1 of 11 scar tissue samples from sites where fibropapillomas had been removed 2 to 51 wk earlier from 5 green turtles contained detectable herpesvirus. None of 18 normal skin samples from 2 green and 2 loggerhead turtles stranded without FP contained herpesvirus. The data indicated that herpesvirus was detectable only within or close to tumors. To determine if the same virus infected both turtle species, partial nucleotide sequences of the herpesvirus DNA polymerase gene were determined from 6 loggerhead and 2 green turtle samples. The sequences predicted that herpesvirus of loggerhead turtles differed from those of green turtles by only 1 of 60 amino acids in the sequence examined, indicating that a chelonian herpesvirus exhibiting minor intratypic variation was the only herpesvirus present in tumors of both green and loggerhead turtles. The FP-associated herpesvirus resisted cultivation on chelonian cell lines which support the replication of other chelonian herpesviruses. These results lead to the conclusion that a chelonian herpesvirus is regularly associated with fibropapillomatosis and is not merely an incidental finding in affected turtles.
Amino Acid Sequence↗
Nucleotide sequence and characterization of human papillomavirus type 83, a novel genital papillomavirus.
Studies of human papillomaviruses (HPV) are hampered by the lack of a conventional culture system, because HPV completes its life cycle only in fully differentiated human tissue. To overcome this obstacle, the athymic mouse xenograft system has been used to study the pathogenesis of a limited number of HPV types. We recently reported the propagation of a novel HPV type in the mouse xenograft system and the cloning of its genome. Consensus primer PCR had previously identified this virus as MM7, LVX82, or PAP291. Here we report the nucleotide sequence of the 8104-bp genome of this virus, now called HPV 83. HPV 83 is most closely related to HPV 61 and HPV 72, placing it in the papillomavirus genome homology group A3. Based on limited epidemiological data, the histological appearance of infected human foreskin implants, and the structure of the predicted HPV 83 E7 protein, this virus is probably of at least intermediate cancer risk. Like other papillomaviruses, HPV 83 produces an E1 E4, E5 transcript, but the position of the splice acceptor differs from that of other HPVs. The presence of an E5 open reading frame in the HPV 83 genome is uncertain; the most likely candidate to be the HPV 83 E5 protein has some structural similarity to the bovine papillomavirus 1 E5 oncoprotein, and is unlike most other HPV E5 proteins. HPV 83 is a relatively prevalent genital papillomavirus that has the largest genome of any characterized HPV and several other novel structural features that merit further study.
Amino Acid Sequence↗
Selective oxidation of methionine residues in prion proteins.
Prion proteins are central to the pathogenesis of several neurodegenerative diseases through the postulated conversion of the endogenous cellular isoform (PrPc) into a pathogenic isoform (PrPSc). Although the cellular function of normal prion protein remains unresolved a number of studies have shown that prion proteins may be involved in the cellular response to oxidative stress. Here, using purified recombinant sources of mouse and chicken PrP refolded in the presence of copper (II) we show that the methionine residues of the protein are uniquely susceptible to oxidation. We suggest that Met residues may form an essential part of the mechanism of the antioxidant activity exhibited by normal prion protein.
Amino Acid Sequence↗
Reliability of information on physical activity and other chronic disease risk factors among US women aged 40 years or older.
Data on chronic disease risk behaviors and related variables, including barriers to and attitudes toward physical activity, are lacking for women of some racial/ethnic groups. A test-retest study was conducted from July 1996 through June 1997 among US women (n = 199) aged 40 years or more who were white, black, American Indian/Alaska Native, or Hispanic. The sample was selected and interviews were conducted using a modified version of the methods of the Behavioral Risk Factor Surveillance System. For behavioral risk factors such as physical inactivity, smoking, and low fruit and vegetable consumption, group prevalences were generally similar between interviews 1 and 2. However, kappa values for selected physical activity variables ranged from 0.26 to 0.51 and tended to be lower for black women. Discordance was low for variables on cigarette smoking and exposure to environmental tobacco smoke (kappa = 0.64-0.92). Discordance was high (kappa = 0.33) for low consumption of fruits and vegetables. Additional variables for barriers to and access to exercise ranged widely across racial/ethnic groups and in terms of measures of agreement. These methods illustrate an efficient way to sample and assess the reliability of data collected from women of racial/ethnic minority groups.
Adult↗
Astrocytic glutamate uptake and prion protein expression.
Factors influencing glutamate uptake by astrocytes may indirectly influence neuronal survival. Elevated extracellular glutamate may be excitotoxic or may exacerbate neurodegeneration in various neurological diseases. By using a cell culture model, we have investigated the influence of astrocytic prion protein (PrPc) expression on glutamate uptake. Type 1 astrocytes expressing PrPc have a higher rate of Na+-dependent glutamate uptake than PrPc-deficient type 1 astrocytes. This difference is exacerbated when serum free media is used to culture the astrocytes. Further analysis suggested that a decrease in substrate affinity is responsible for the sensitivity of PrP-deficient astrocytic glutamate uptake to culture conditions. PrPc has been shown to bind copper. Greater sensitivity of cells to copper concentrations may be responsible for the decreased substrate affinity observed. PrPc-deficient cerebellar cells are more sensitive to glutamate toxicity in the presence of copper. These results show that glutamate uptake from astrocytes is dependent on PrPc expression which in turn may be related to copper metabolism.
Animals↗
Cloning, expression and functional role of a nociceptin/orphanin FQ receptor in the porcine gastrointestinal tract.
The heptadecapeptide nociceptin/orphanin FQ is the cognate ligand for the opioid receptor-like orphanin FQ (OFQ) receptor, a member of the G protein-coupled receptor superfamily. The gastrointestinal tract is a major site of opioid action, and preliminary evidence suggests that an OFQ receptor may be expressed in rat small intestine. We addressed the hypothesis that this receptor is expressed in the gastrointestinal tract of the pig, a model for the human digestive system. A 1205-bp cDNA was isolated from porcine forebrain which contained the 370 amino acid open reading frame encoding the OFQ receptor. The receptor mRNA is likely to arise from a single gene, as determined by Southern blotting of porcine genomic DNA restriction digests using a porcine OFQ receptor cDNA probe. A semi-nested reverse transcriptase-polymerase chain reaction survey of receptor mRNA indicates that it is expressed in the porcine cerebral cortex and kidney, and along the length of the gastrointestinal tract. OFQ decreased initial contractile responses of porcine ileal smooth muscle strips to trains of electrical field stimulation with an IC50 value of 1.3 nM; its effects were resistant to the opioid antagonist, naloxone. The peptide, at concentrations > or =3 nM, also attenuated Isc elevations evoked by electrical transmural stimulation of mucosa-submucosa sheets from porcine ileum. The actions of OFQ appeared to differ from those previously reported for opioid receptor agonists in these tissue preparations. These results indicate that an OFQ receptor is expressed in the porcine intestine which modulates the neural control of intestinal smooth muscle contractility and mucosal transport.
Amino Acid Sequence↗
Opportunities for depression disease management: a pharmacy benefit manager's experience.
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Clinical Pharmacy Information Systems↗
Neurons depend on astrocytes in a coculture system for protection from glutamate toxicity.
Glutamate can be toxic to neurons although it is a neurotransmitter. Regulation of extracellular glutamate levels is essential for prevention of glutamate neurotoxicity. Astrocytes play a major role in clearance of glutamate released by neurons. A coculture system combining cerebellar cells and astrocytes was employed to investigate the astrocytic control of glutamate toxicity. Coculture of astrocytes with cerebellar neurons enhanced uptake of glutamate by astrocytes. Inhibition of glutamate uptake in a coculture system led to death of cerebellar cells. This toxicity could be inhibited by MK801. However, in the presence of the glutamate uptake inhibitor, there was no increase in glutamate in the cultures compared to when the neurons were not cocultured. This indicated that neurons become more susceptible to glutamate toxicity in the presence of astrocytes and thus become dependent on astrocytes for prevention of glutamate toxicity. Astrocytes treated with conditioned medium from cerebellar cells did not show an increase in glutamate uptake but medium from astrocytes exposed to neuron conditioned medium was toxic to cerebellar cells. This toxicity was due to glutamate present in the medium. This suggests that a soluble factor released by neurons signals to astrocytes that neurons are present and stimulates a signal back to neurons which causes an increased sensitivity to glutamate toxicity.
Animals↗
Catecholaminergic, cholinergic and peptidergic innervation of gut-associated lymphoid tissue in porcine jejunum and ileum.
With its abundance of neurons and immunocytes, the gut is a potentially important site for the study of the interaction between the nervous and immune systems. Using immunohistochemical techniques, we tested the hypothesis that gut-associated lymphoid tissue in the porcine small intestine might receive catecholaminergic, cholinergic and peptidergic innervation. Antibodies against protein gene product (PGP) 9.5 were employed to detect neuronal membranes; antibodies against tyrosine hydroxylase (TH), type 2 vesicular monoamine transporter (VMAT-2) and choline acetyltransferase (ChAT) were used to detect catecholaminergic and cholinergic neurons; and antibodies to neuromedin U-8 (NMU-8), substance P (SP) and vasoactive intestinal peptide (VIP) were also used. PGP9.5-immunoreactive nerve fibers were observed between jejunal Peyer's patch (PP) follicles and in submucosal ganglia localized at the base of continuous ileal PP. Many ChAT-positive and a few TH-/VMAT-2-immunoreactive neurons or axons adjacent to jejunal and ileal PP were observed. Neurons and fibers from ganglia situated between or at the base of PP follicles manifested robust immunoreactivities to VIP and NMU-8; relatively less SP immunoreactivity was observed at these locations. All neuromedin-U 8-positive neurons observed exhibited immunoreactivity to ChAT as did some VIP-positive neurons. The specific chemical coding of enteric neurons in close apposition to jejunal and ileal PP and the differential localization of neuropeptides within the jejunal and ileal PP are indicative of neuroimmunomodulation at these sites.
Animals↗
Cervical human papillomavirus deoxyribonucleic acid persists throughout pregnancy and decreases in the postpartum period.
OBJECTIVE: Our goal was to determine the persistence of human papillomavirus infection of the cervix in a prospectively evaluated cohort of pregnant women observed from the first trimester until after delivery. STUDY DESIGN: A group of 232 women were enrolled in the first trimester of pregnancy and had cervico-vaginal lavage specimens collected for detection of the deoxyribonucleic acid of human papillomavirus. They underwent sampling again in the third trimester (146 patients available) and at 4 to 12 weeks after delivery (83 patients available). Human papillomavirus deoxyribonucleic acid was detected by means of the Hybrid Capture assay. RESULTS: In the first trimester of pregnancy, 31% of the patients had positive test results for human papillomavirus deoxyribonucleic acid, whereas 35.6% had positive results in the third trimester (P = 1.0). A comparison of first-trimester test results with postpartum results (paired data available from 83 patients) showed a decline from 39.8% positivity to 26.5% (P =.04). Comparing third-trimester results with postpartum results (paired data available from 74 patients) showed a decline from 35.1% to 25. 7% positivity (P =.12). When specimens positive for human papillomavirus were divided between those containing "high cancer risk" types (9 virus types often associated with dysplasia or malignancy) and "low cancer risk" types (5 types usually found in benign lesions), similar trends were seen, although not all comparisons were statistically significant. CONCLUSION: The increased prevalence, during pregnancy, of detectable human papillomavirus deoxyribonucleic acid, which was previously reported (Fife et al, Am J Obstet Gynecol 1996;174:1487-93), persists at a similar level throughout pregnancy but declines in the postpartum period. This observation is most consistent with activation of the virus by the physiologic changes of pregnancy.
Adult↗
Physical activity patterns of Chippewa and Menominee Indians: the Inter-Tribal Heart Project.
INTRODUCTION: Leisure-time (LTPA), occupational, transportation, and household physical activity were evaluated among participants in the Inter-Tribal Heart Project (ITHP). METHODS: Age-stratified random samples of persons aged > or =25 years were drawn from three communities of Chippewa and Menominee Indians in Minnesota and Wisconsin. Participants (843 women and 501 men) completed an interviewer-administered questionnaire. Logistic regression analyses were performed to assess age-adjusted associations between no reported LTPA and potential correlates. RESULTS: During leisure time, 12% of women and 17% of men were active 7-12 months in the past year; 33% of women and 21% of men reported no LTPA. During a typical workday for employed persons, approximately 90% of both genders walked > or =20 minutes, for carrying/lifting moderate to heavy objects the median value for men was 1 hour and women 0 hours. The median value of weekly household activity for men was 3 hours compared to 10 hours for women. Little transportation activity was reported by either gender. Age, household income, smoking, and poor/fair self-perceived health were associated with leisure-time inactivity in women (p < 0.05). Age, poor/fair self-perceived health, and smoking were associated with leisure-time inactivity in men (p < 0.05). Walking was the most prevalent activity in the population. CONCLUSIONS: Prevalence of leisure-time inactivity is higher than the national health objective of 15%, however, ITHP participants obtain a substantial amount of occupational and household activity that may lower risks of chronic diseases. Health promotion efforts to increase LTPA may lead not only to reduced chronic disease risk, but to additional benefits of enjoyment and improved psychological well-being.
Adult↗
Poly(ADP-ribose) immunostaining to detect apoptosis induced by a neurotoxic fragment of prion protein.
PrP106-126 is a synthetic peptide representing codons 106-126 of the prion protein, which spontaneously forms amyloid fibrils and exerts neurotoxic effects on primary mouse brain cell cultures. Neurotoxicity by this peptide is commonly used as a model for the neurotoxicity observed in prion diseases and involves the formation of reactive oxygen species which, in turn, can cause DNA damage, including DNA strand breaks. Strand breaks in nuclear DNA can activate poly(ADP-ribose) polymerase to covalently modify nuclear proteins with poly(ADP-ribose). We, therefore, examined by immunofluorescence whether or not PrP106-126 triggers poly(ADP-ribose) formation. We observed strong poly(ADP-ribose) immunofluorescence signals in a fraction of cells, typically arranged in a clustered pattern, by 30-48 h after peptide addition. A few positive cells were also present in untreated cultures. Cell morphology was suggestive of apoptosis, and this was confirmed by positivity in the terminal deoxynucleotidyltransferase-mediated dUTP nick-end labelling (TUNEL) assay. On the other hand, our immunofluorescence assay did not detect any 'early' activation of poly(ADP-ribose) polymerase in morphologically normal cells that could have resulted from peptide-induced formation of reactive oxygen species. We conclude that poly(ADP-ribose) immunostaining is a convenient and reliable method for visualizing cells undergoing apoptosis induced by PrP106-126.
Animals↗
Cellular uptake of the prion protein fragment PrP106-126 in vitro.
The aetiological agent of prion disease is proposed to be an aberrant isoform of the cell surface glycoprotein known as the prion protein (PrP(c)). This pathological isoform (PrP(Sc)) is abnormally deposited in the extracellular space of diseased CNS. Neurodegeneration in these disease has been shown to be associated with accumulation of PrP(Sc) in affected tissue. To investigate the possible uptake mechanisms that may be required for PrP(Sc)-induced neurodegeneration we studied the cellular trafficking of the neurotoxic fragment, PrP106-126. We were able to detect, by fluorescence microscopy, PrP106-126 inclusions in murine neurones, astrocytes and microglia in vitro. These inclusions were abundant after 24 hour exposure and still present 48h post-exposure. Shorter exposure times yielded only occasional cells with inclusions. Large extracellular aggregates of PrP106-126 could also be detected, which appeared in a time dependent manner. The appearance of inclusions or aggregates was not dependent on PrP(c) expression as determined by exposure of peptides from PrP-null mice. Using transmission electron microscopy and gold particle detection, positively labelled osmiophilic inclusions of peptide could be detected in the cytoplasm of exposed cells. These results demonstrate that cultured cells are capable of sequestering PrP106-126 and may indicate uptake pathways for PrP(Sc) in various cell types. Toxicity of PrP106-126 may thus be mediated via a sequestration pathway that is not effective for this peptide in PrP-null cells.
Animals↗
Dependence of neurones on astrocytes in a coculture system renders neurones sensitive to transforming growth factor beta1-induced glutamate toxicity.
Transforming growth factor beta1 (TGF-beta1) has been implicated in formation of astrocyte scars, which prevents axonal regeneration. A coculture system of astrocytes and cerebellar cells was used to investigate possible neurotoxic effects of TGF-beta1. Although not directly neurotoxic, TGF-beta1 was toxic to cerebellar cells in the presence of astrocytes. This toxicity is based on an effect of the cytokine on astrocytes, as conditioned medium from astrocyte cultures treated with TGF-beta1 was more toxic by a similar mechanism. This neurotoxicity was mediated by glutamate present in the culture medium as demonstrated by inhibition by MK-801. Astrocytic ability to metabolise glutamate was compromised by TGF-beta1, as this cytokine increased glutamate concentration. The astrocytes in the coculture system responded to the presence of neurones by secreting neuroprotective interleukin-6, which was partly protective against the TGF-beta1-induced toxicity. In the coculture system, neurones responded to the presence of astrocytes by a reduction in resistance to glutamate toxicity. On addition of TGF-beta1, which compromised astrocytic clearance of glutamate, this reduction in resistance to glutamate toxicity led to a reduction in neuronal survival. These results suggest that when neurones are cocultured with astrocytes they become dependent on astrocytes for survival. This dependence makes neurones susceptible to damage when astrocytes are activated by substances such as TGF-beta1.
Animals↗
Prion protein peptide neurotoxicity can be mediated by astrocytes.
A peptide based on amino acids 106-126 of the sequence of human prion protein (PrP106-126) is neurotoxic in culture. A role for astrocytes mediating PrP106-126 toxicity was investigated. The toxicity of PrP106-126 to cerebellar cell cultures was reduced by aminoadipate, a gliotoxin. Normally, PrP106-126 is not toxic to cultures containing neurones deficient in the cellular isoform of prion protein (PrPc). However, PrP106-126 was toxic to cerebellar cells derived from Prnp(0/0) mice (deficient in PrPc expression) when those cerebellar cells were cocultured with astrocytes. This toxicity was found to occur only in the presence of PrPc-positive astrocytes and to be mediated by glutamate. Furthermore, PrPc-positive astrocytes were shown to protect Prnp(0/0) cerebellar cells from glutamate toxicity. This effect could be inhibited by PrP106-126. PrP106-126 did not enhance the toxicity of glutamate to neurones directly. When cerebellar cells were cocultured with astrocytes, the neurones became dependent on astrocytes for protection from glutamate toxicity and expressed an increased sensitivity to glutamate. In such a system, the protective effects of astrocytes against glutamate toxicity to neurones were inhibited by PrP106-126, resulting in a greater reduction in neuronal survival than would have been caused by PrP106-126 when astrocytes were not present. This new model provides a possible mechanism by which the gliosis in prion disease may accelerate the neurodegeneration seen in the later stages of the disease.
Amino Acid Sequence↗
Racial differences in the multiple social roles of older women: implications for depressive symptoms.
The relationship between multiple role participation and depressive symptoms experienced by African American (n = 547) and White (n = 2,152) women aged 55-61 was explored. Data were obtained from the Health and Retirement Study (HRS). Racial differences in the social roles of marriage, employment, grandmother, care provider, and volunteer and their influence on level of depressive symptoms were examined. African Americans reported higher levels of depressive symptoms than Whites. Additionally, marriage, employment, and total number of social roles were the most powerful predictors of depressive symptoms for both African American and White women. However, employment was more important in diminishing depressive symptoms among African American than White women occupying multiple social roles.
Black or African American↗