170 results on '"Merril CR"'
Search Results
2. Stable transformation of CHO Cells and human NARP cybrids confers oligomycin resistance (oli(r)) following transfer of a mitochondrial DNA-encoded oli(r) ATPase6 gene to the nuclear genome: a model system for mtDNA gene therapy.
- Author
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Zullo SJ, Parks WT, Chloupkova M, Wei B, Weiner H, Fenton WA, Eisenstadt JM, Merril CR, Zullo, S J, Parks, W T, Chloupkova, M, Wei, B, Weiner, H, Fenton, W A, Eisenstadt, J M, and Merril, C R
- Published
- 2005
3. Preliminary study of an increase of a plasma apolipoprotein E variant associated with peripheral nerve damage. A finding in patients with chronic spinal pain.
- Author
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Cameron BM, VanderPutten DM, Merril CR, Cameron, B M, VanderPutten, D M, and Merril, C R
- Published
- 1995
4. Mitochondrial DNA Sequence Analysis of Human Skeletal Remains: Identification of Remains from the Vietnam War
- Author
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Holland, MM, Fisher, DL, Mitchell, LG, Rodriquez, WC, Canik, JJ, Merril, CR, and Weedn, VW
- Abstract
Deoxyribonucleic acid (DNA) sequence analysis of the control region of the mitochondrial DNA (mtDNA) genome was used to identify human skeletal remains returned to the United States government by the Vietnamese government in 1984. The postmortem interval was thought to be 24 years at the time of testing, and the remains presumed to be an American service member. DNA typing methods using nuclear genomic DNA, HLA-DQ alpha [1] and the variable number of tandem repeat (VNTR) locus D1S80 [2], were unsuccessful using the polymerase chain reaction (PCR) [3]. Amplification of a portion of the mtDNA control region was performed, and the resulting PCR product subjected to DNA sequence analysis. The DNA sequence generated from the skeletal remains was identical to the maternal reference sequence, as well as the sequence generated from two siblings (sisters). The sequence was unique when compared to more than 650 DNA sequences found both in the literature and provided by personal communications. The individual sequence polymorphisms were present in only 23 of the more than 1300 nucleotide positions analyzed. These results support the observation [4] that in cases where conventional DNA typing is unavailable, mtDNA sequencing can be used for human remains identification.
- Published
- 1993
- Full Text
- View/download PDF
5. Mapping and quantitation of proteins from discrete nuclei and other areas of the rat brain by two-dimensional gel electrophoresis
- Author
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Heydorn, WE, primary, Creed, GJ, additional, Goldman, D, additional, Kanter, D, additional, Merril, CR, additional, and Jacobowitz, DM, additional
- Published
- 1983
- Full Text
- View/download PDF
6. Addition of leucine precursors to the diet of leucine-starved mice
- Author
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Trigg, ME, primary, Geier, MR, additional, LaPolla, RJ, additional, Kamerow, HN, additional, and Merril, CR, additional
- Published
- 1975
- Full Text
- View/download PDF
7. Two hundred issues later: the dream comes true.
- Author
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Merril CR and Ghanbari HA
- Subjects
- History, 20th Century, History, 21st Century, Humans, Alzheimer Disease history, Dreams, Periodicals as Topic history
- Published
- 2015
- Full Text
- View/download PDF
8. Therapeutic and prophylactic applications of bacteriophage components in modern medicine.
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Adhya S, Merril CR, and Biswas B
- Subjects
- Animals, Bacterial Infections immunology, Bacterial Infections prevention & control, Disease Models, Animal, Humans, Muramidase therapeutic use, Vaccines, Anti-Bacterial Agents therapeutic use, Bacterial Infections therapy, Bacteriophages immunology
- Abstract
As the interactions of phage with mammalian innate and adaptive immune systems are better delineated and with our ability to recognize and eliminate toxins and other potentially harmful phage gene products, the potential of phage therapies is now being realized. Early efforts to use phage therapeutically were hampered by inadequate phage purification and limited knowledge of phage-bacterial and phage-human relations. However, although use of phage as an antibacterial therapy in countries that require controlled clinical studies has been hampered by the high costs of patient trials, their use as vaccines and the use of phage components such as lysolytic enzymes or lysozymes has progressed to the point of commercial applications. Recent studies concerning the intimate associations between mammalian hosts and bacterial and phage microbiomes should hasten this progress.
- Published
- 2014
- Full Text
- View/download PDF
9. Is sporadic Alzheimer's disease associated with diphtheria toxin?
- Author
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Merril CR
- Subjects
- Alzheimer Disease epidemiology, Animals, Diphtheria epidemiology, Diphtheria pathology, Diphtheria prevention & control, Humans, Alzheimer Disease pathology, Alzheimer Disease prevention & control, Diphtheria Toxin, Diphtheria-Tetanus-Pertussis Vaccine therapeutic use
- Abstract
The two major aspects of Alzheimer's disease (AD) that must be considered in a search for causative agents are its association with aging and its widespread epidemiology. While a number of agents have been identified, additional factors may play a role. An association with diphtheria toxin was suggested by observations that vaccinations may provide protective effects, and the observation that decreased proteins synthesis in cortical regions from AD patients is associated with modification of elongation factor 2, the target of diphtheria toxin. While protection against diphtheria toxin is provided by vaccination, the known decline in the immune system associated with aging would result in a renewed sensitivity to the toxin. An association with diphtheria toxin would be consistent with the observations that the bacteria associated with the toxin, Corynebacterium diphtheria, is often found in the nasopharynx and an early symptom of AD is the loss of smell with a disease progression from the entorhinal cortex to the hippocampus and the neocortical areas. If diphtheria toxin is involved in sporadic AD, booster vaccinations given to elderly individuals might result in a decreased incidence of this disease. As booster DPT vaccinations are already recommended for individuals over 65, cognitive testing at the time of the booster and 5 years later, along with similar cognitive testing in age-matched individuals who decline vaccination, might provide an inexpensive method to investigate whether diphtheria toxin plays a role in AD and the efficacy of DPT booster vaccines for AD.
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- 2013
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10. An amino acid substitution in a capsid protein enhances phage survival in mouse circulatory system more than a 1000-fold.
- Author
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Vitiello CL, Merril CR, and Adhya S
- Subjects
- Animals, Bacteriophage lambda genetics, Escherichia coli Infections microbiology, Female, Mice, Mice, Inbred BALB C, Amino Acid Substitution, Bacteremia microbiology, Bacteriophage lambda physiology, Blood virology, Capsid Proteins genetics, Escherichia coli K12 virology
- Abstract
In experiments with germ free mice, free from adaptive antibodies to the bacterial virus lambda phage, titers of the virus in the circulatory system have been reported to decrease by more than 10(9)pfu within 48 h of intraperitoneal intravenous or oral administration. Based on these observations, serial passage techniques have been used to select lambda phage mutants, with 13,000-16,000-fold greater capacity to remain in the mouse circulatory system 24h after intraperitoneal injection. In these prior studies the "long-circulating" phage, designated lambdaArgo phage, had at least three mutations including one in the major phage capsid (E) protein, which resulted in the change of glutamic acid to a lysine at residue 158. In the current study, we demonstrate that this single specific substitution in the E protein is sufficient to confer the "long-circulating" phenotype. The isogenic pair of phage developed in this study consisting of the long-circulating marker-rescued lambdaArgo phage, and the parental wild type phage can be used for studies of viral recognition mechanisms of the innate immune system and for the development of more effective antibacterial therapeutic phage strains.
- Published
- 2005
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11. The prospect for bacteriophage therapy in Western medicine.
- Author
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Merril CR, Scholl D, and Adhya SL
- Subjects
- Animals, Anti-Bacterial Agents, Bacteriophages genetics, Humans, Technology, Pharmaceutical methods, Anti-Infective Agents chemistry, Bacterial Infections drug therapy, Bacteriophages chemistry, Technology, Pharmaceutical trends
- Abstract
Bacteriophage (phage) have been used for clinical applications since their initial discovery at the beginning of the twentieth century. However, they have never been subjected to the scrutiny--in terms of the determination of efficacy and pharmacokinetics of therapeutic agents--that is required in countries that enforce certification for marketed pharmaceuticals. There are a number of historical reasons for this deficiency, including the overshadowing discovery of the antibiotics. Nevertheless, present efforts to develop phage into reliable antibacterial agents have been substantially enhanced by knowledge gained concerning the genetics and physiology of phage in molecular detail during the past 50 years. Such efforts will be of importance given the emergence of antibiotic-resistant bacteria.
- Published
- 2003
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12. Bacteriophage SP6 is closely related to phages K1-5, K5, and K1E but encodes a tail protein very similar to that of the distantly related P22.
- Author
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Scholl D, Adhya S, and Merril CR
- Subjects
- Amino Acid Sequence, Base Sequence, DNA Helicases genetics, DNA-Directed RNA Polymerases genetics, Molecular Sequence Data, Viral Tail Proteins chemistry, Virion genetics, Coliphages genetics, Salmonella Phages genetics, Viral Tail Proteins genetics
- Abstract
The lytic salmonella phage SP6 encodes a tail protein with a high degree of sequence similarity to the tail protein of the biologically unrelated lysogenic salmonella phage P22. The SP6 tail gene is flanked by an upstream region that contains a promoter and a downstream region that contains a putative Rho-independent transcription terminator, giving it a cassette or modular structure almost identical to the structure of the tail genes of coliphages K1E, K5, and K1-5. It now appears that SP6, K1-5, K5, and K1E are very closely related but have different tail fiber proteins, giving them different host specificities.
- Published
- 2002
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13. Bacteriophage therapy rescues mice bacteremic from a clinical isolate of vancomycin-resistant Enterococcus faecium.
- Author
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Biswas B, Adhya S, Washart P, Paul B, Trostel AN, Powell B, Carlton R, and Merril CR
- Subjects
- Animals, Antibodies, Viral blood, Antibodies, Viral immunology, Bacteriophages immunology, Bacteriophages ultrastructure, Disease Models, Animal, Enterococcus faecium growth & development, Female, Heating, Humans, Mice, Mice, Inbred BALB C, Microscopy, Electron, Time Factors, Bacteremia therapy, Bacteriophages physiology, Enterococcus faecium virology, Gram-Positive Bacterial Infections therapy, Vancomycin Resistance
- Abstract
Colonization of the gastrointestinal tract with vancomycin-resistant Enterococcus faecium (VRE) has become endemic in many hospitals and nursing homes in the United States. Such colonization predisposes the individual to VRE bacteremia and/or endocarditis, and immunocompromised patients are at particular risk for these conditions. The emergence of antibiotic-resistant bacterial strains requires the exploration of alternative antibacterial therapies, which led our group to study the ability of bacterial viruses (bacteriophages, or phages) to rescue mice with VRE bacteremia. The phage strain used in this study has lytic activity against a wide range of clinical isolates of VRE. One of these VRE strains was used to induce bacteremia in mice by intraperitoneal (i.p.) injection of 10(9) CFU. The resulting bacteremia was fatal within 48 h. A single i.p. injection of 3 x 10(8) PFU of the phage strain, administered 45 min after the bacterial challenge, was sufficient to rescue 100% of the animals. Even when treatment was delayed to the point where all animals were moribund, approximately 50% of them were rescued by a single injection of this phage preparation. The ability of this phage to rescue bacteremic mice was demonstrated to be due to the functional capabilities of the phage and not to a nonspecific immune effect. The rescue of bacteremic mice could be effected only by phage strains able to grow in vitro on the bacterial host used to infect the animals, and when such strains are heat inactivated they lose their ability to rescue the infected mice.
- Published
- 2002
- Full Text
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14. Bacteriophage K1-5 encodes two different tail fiber proteins, allowing it to infect and replicate on both K1 and K5 strains of Escherichia coli.
- Author
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Scholl D, Rogers S, Adhya S, and Merril CR
- Subjects
- Bacterial Capsules, Base Sequence, Coliphages genetics, Coliphages isolation & purification, Escherichia coli metabolism, Molecular Sequence Data, Mutation, Sewage virology, Viral Tail Proteins metabolism, Virion metabolism, Virus Replication, Antigens, Bacterial, Antigens, Surface metabolism, Coliphages physiology, Escherichia coli virology, Polysaccharides, Bacterial metabolism, Viral Tail Proteins genetics
- Abstract
A virulent double-stranded DNA bacteriophage, Phi K1-5, has been isolated and found to be capable of infecting Escherichia coli strains that possess either the K1 or the K5 polysaccharide capsule. Electron micrographs show that the virion consists of a small icosohedral head with short tail spikes, similar to members of the Podoviridae family. DNA sequence analysis of the region encoding the tail fiber protein showed two open reading frames encoding previously characterized hydrolytic phage tail fiber proteins. The first is the K5 lyase protein gene of Phi K5, which allows this phage to specifically infect K5 E. coli strains. A second open reading frame encodes a protein almost identical in amino acid sequence to the N-acetylneuraminidase (endosialidase) protein of Phi K1E, which allows this phage to specifically infect K1 strains of E. coli. We provide experimental evidence that mature phage particles contain both tail fiber proteins, and mutational analysis indicates that each protein can be independently inactivated. A comparison of the tail gene regions of Phi K5, Phi K1E, and Phi K1-5 shows that the genes are arranged in a modular or cassette configuration and suggests that this family of phages can broaden host range by horizontal gene transfer.
- Published
- 2001
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15. Dopamine induces cell death, lipid peroxidation and DNA base damage in a catecholaminergic cell line derived from the central nervous system.
- Author
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Masserano JM, Baker I, Venable D, Gong L, Zullo SJ, Merril CR, and Wyatt RJ
- Abstract
Dopamine can be autoxidized to superoxides and quinones. Superoxides can form hydroxyl radicals that are highly reactive with lipids, proteins and DNA leading to neuronal damage and cell death. We used a clonal catecholaminergic cell line (CATH.a) derived from the central nervous system to evaluate the effects of dopamine on cell death, lipid peroxidation and DNA base damage. Dopamine produces cell death in CATH.a cells and this is associated with an increase in annexin binding, which is an early indicator of apoptosis. Incubation of CATH.a cells with deferoximine, an iron chealator, partially antagonizes dopamine-induced cell death. In CATH.a cells, dopamine produces an increase in both lipid peroxidation, as measured by cis-parinaric acid fluorescence, and DNA oxidative base damage, as measured by 8-hydroxy-2'-deoxyguanosine formation. Cell death was inhibited 84-92% by the hydrophilic antioxidants, dithiothreitol, L-cysteine, and N-acetylcysteine. The lipophilic vitamins, retinol and vitamin E and the vitamin E analog, Trolox, inhibited dopamine-induced cell death by 18-33%. The lipophilic antioxidants probucol, propyl glycol and butylated hydroxyanisone had no inhibitory effect on dopamine-induced cell death. These data suggest that damage to DNA and lipids may be partially responsible for dopamine-induced cell death in CATH.a cells.
- Published
- 2000
- Full Text
- View/download PDF
16. Possible relationship between conditions associated with chronic hypoxia and brain mitochondrial DNA deletions; reduction of genomic 8-hydroxyguanine levels in human brain tissues containing elevated levels of the human mitochondrial DNA4977 deletion.
- Author
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Zullo SJ, Cerritos A, and Merril CR
- Subjects
- Brain pathology, Chronic Disease, DNA Adducts analysis, DNA Adducts genetics, DNA Damage genetics, Deoxyguanine Nucleotides analysis, Guanine analysis, Humans, Mitochondria genetics, Mitochondria physiology, Reactive Oxygen Species metabolism, Statistics as Topic, Brain metabolism, DNA, Mitochondrial genetics, Genome, Human, Guanine analogs & derivatives, Hypoxia metabolism, Sequence Deletion
- Published
- 1999
- Full Text
- View/download PDF
17. Localization by fluorescence in situ hybridization (FISH) of human mitochondrial polymerase gamma (POLG) to human chromosome band 15q24-->q26, and of mouse mitochondrial polymerase gamma (Polg) to mouse chromosome band 7E, with confirmation by direct sequence analysis of bacterial artificial chromosomes (BACs).
- Author
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Zullo SJ, Butler L, Zahorchak RJ, Macville M, Wilkes C, and Merril CR
- Subjects
- Animals, Base Sequence, Cloning, Molecular methods, DNA Polymerase gamma, Humans, In Situ Hybridization, Fluorescence, Mice, Mitochondria enzymology, Molecular Sequence Data, Sequence Analysis, DNA, Species Specificity, Chromosomes, Bacterial, Chromosomes, Human, Pair 15, DNA-Directed DNA Polymerase genetics, Gene Transfer Techniques, Mitochondria genetics
- Abstract
Cloned cDNAs for the human mitochondrial DNA polymerase gamma (POLG) were identified by homology with the yeast mitochondrial DNA polymerase catalytic subunit (MIP). Fluorescence in situ hybridization (FISH) of human and mouse bacterial artificial chromosomes (BACs), hybridized by radioactively labeled POLG cDNAs, mapped to human chromosome band 15q24-->q26, as well as to mouse chromosome band 7E. Direct sequencing of the BAC DNA without subcloning confirmed the presence of both human POLG and mouse mitochondrial DNA polymerase gamma (Polg) in the respective BACs.
- Published
- 1997
- Full Text
- View/download PDF
18. Plasma protein variations in monozygotic twins discordant for schizophrenia.
- Author
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Vander Putten DM, Torrey EF, Larive AB, and Merril CR
- Subjects
- Adult, Chromatography, Gel, Cognition, Haptoglobins metabolism, Humans, Twins, Monozygotic, Blood Proteins metabolism, Schizophrenia blood
- Abstract
Monozygotic twins discordant for schizophrenia were analyzed by two-dimensional (2-D) gel electrophoresis to identify extrahereditary factors important in the development of schizophrenia. Plasma protein patterns in 2-D gels of monozygotic twins discordant for schizophrenia were found to be significantly less alike than those of normal control monozygotic twins. Several polypeptide spots were found to be elevated in the plasma of the schizophrenic twin. One of these polypeptides, spot 782, was also found to be significantly (p < .001) elevated when schizophrenic patients were compared to unrelated normal control individuals. Spot 782 may be an isoform of haptoglobin. Quantitative variations in some plasma haptoglobin levels were seen between discordant twins, but not between unrelated schizophrenic and normal control individuals.
- Published
- 1996
- Full Text
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19. Long-circulating bacteriophage as antibacterial agents.
- Author
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Merril CR, Biswas B, Carlton R, Jensen NC, Creed GJ, Zullo S, and Adhya S
- Subjects
- Animals, Bacteremia therapy, Bacteriophage P22 genetics, Bacteriophage P22 physiology, Bacteriophage lambda genetics, Bacteriophage lambda physiology, Escherichia coli Infections therapy, Female, Mice, Mice, Inbred BALB C, Mutation, Salmonella Infections, Animal therapy, Salmonella typhimurium virology, Bacterial Infections therapy, Bacteriophages physiology
- Abstract
The increased prevalence of multidrug-resistant bacterial pathogens motivated us to attempt to enhance the therapeutic efficacy of bacteriophages. The therapeutic application of phages as antibacterial agents was impeded by several factors: (i) the failure to recognize the relatively narrow host range of phages; (ii) the presence of toxins in crude phage lysates; and (iii) a lack of appreciation for the capacity of mammalian host defense systems, particularly the organs of the reticuloendothelial system, to remove phage particles from the circulatory system. In our studies involving bacteremic mice, the problem of the narrow host range of phage was dealt with by using selected bacterial strains and virulent phage specific for them. Toxin levels were diminished by purifying phage preparations. To reduce phage elimination by the host defense system, we developed a serial-passage technique in mice to select for phage mutants able to remain in the circulatory system for longer periods of time. By this approach we isolated long-circulating mutants of Escherichia coli phage lambda and of Salmonella typhimurium phage P22. We demonstrated that the long-circulating lambda mutants also have greater capability as antibacterial agents than the corresponding parental strain in animals infected with lethal doses of bacteria. Comparison of the parental and mutant lambda capsid proteins revealed that the relevant mutation altered the major phage head protein E. The use of toxin-free, bacteria-specific phage strains, combined with the serial-passage technique, may provide insights for developing phage into therapeutically effective antibacterial agents.
- Published
- 1996
- Full Text
- View/download PDF
20. Possible relationship between conditions associated with chronic hypoxia and brain mitochondrial DNA deletions.
- Author
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Merril CR, Zullo S, Ghanbari H, Herman MM, Kleinman JE, Bigelow LB, Bartko JJ, and Sabourin DJ
- Subjects
- Base Sequence, Chronic Disease, Gene Deletion, Humans, Hypoxia pathology, Hypoxia psychology, Molecular Sequence Data, Brain pathology, DNA, Mitochondrial genetics, Hypoxia genetics
- Abstract
The brain relies heavily on aerobic metabolism which requires functional mitochondria. Mitochondria are subcellular organelles with their own genome which codes for 13 essential protein subunits. By employing PCR assays to examine brain tissue from 43 age-comparable individuals (between ages 34 and 73), we found a correlation between mitochondrial DNA deletion mutations, mtDNA4977 deletions, and conditions associated with chronic hypoxia. In prior studies, utilizing only 6 to 12 clinical samples, mtDNA4977 deletions were reported to increase in specific regions of the brain with aging. However, we found 12-fold and 5-fold higher levels of mtDNA4977 deletions in the putamen and the superior frontal gyrus of the cortex, respectively, from individuals who had conditions associated with chronic hypoxia when compared with individuals without evidence of such conditions. These findings suggest that chronic hypoxia should be more closely examined in the pathophysiology of central nervous system diseases.
- Published
- 1996
- Full Text
- View/download PDF
21. Use of silver staining to detect nucleic acids.
- Author
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Mitchell LG, Bodenteich A, and Merril CR
- Subjects
- DNA, Mitochondrial analysis, Gels, Nucleic Acids isolation & purification, Molecular Biology methods, Nucleic Acids analysis, Silver Staining methods
- Published
- 1996
- Full Text
- View/download PDF
22. The protein disease database.
- Author
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Lemkin PF, Orr GA, Goldstein MP, Creed J, Whitley E, Myrick JE, and Merril CR
- Subjects
- Electrophoresis, Gel, Two-Dimensional, Humans, Protein Folding, Databases, Factual, Disease, Proteins metabolism
- Published
- 1995
- Full Text
- View/download PDF
23. The protein disease database of human body fluids: I. Rationale for the development of this database.
- Author
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Merril CR, Goldstein MP, Myrick JE, Creed GJ, and Lemkin PF
- Subjects
- Humans, Body Fluids chemistry, Databases, Factual, Proteins analysis
- Abstract
We are developing a relational database to facilitate quantitative and qualitative comparisons of proteins in human body fluids in normal and disease states. For decades researchers and clinicians have been studying proteins in body fluids such as serum, plasma, cerebrospinal fluid and urine. Currently, most clinicians evaluate only a few specific proteins in a body fluid such as plasma when they suspect that a patient has a disease. Now, however, high resolution two-dimensional protein electrophoresis allows the simultaneous evaluation of 1,500 to 3,000 proteins in complex solutions, such as the body fluids. This and other high resolution methods have encouraged us to collect the clinical data for the body fluid proteins into an easily accessed database. For this reason, it has been constructed on the Internet World Wide Web (WWW) under the title Protein Disease Database (PDD). In addition, this database will provide a linkage between the disease-associated protein alterations and images of the appropriate proteins on high-resolution electrophoretic gels of the body fluids. This effort requires the normalization of data to account for variations in methods of measurement. Initial efforts in the establishment of the PDD have been concentrated on alterations in the acute-phase proteins in individuals with acute and chronic diseases. Even at this early stage in the development of our database, it has proven to be useful as we have found that there appear to be several common acute-phase protein alterations in the plasma and cerebrospinal fluid from patients with Alzheimer's disease, schizophrenia and major depression. Our goal is to provide access to the PDD so that systematic correlations and relationships between disease states can be examined and extended.
- Published
- 1995
24. The Protein Disease Database of human body fluids: II. Computer methods and data issues.
- Author
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Lemkin PF, Orr GA, Goldstein MP, Creed GJ, Myrick JE, and Merril CR
- Subjects
- Computer Communication Networks, Forecasting, Humans, Body Fluids chemistry, Database Management Systems, Databases, Factual, Proteins analysis
- Abstract
The Protein Disease Database (PDD) is a relational database of proteins and diseases. With this database it is possible to screen for quantitative protein abnormalities associated with disease states. These quantitative relationships use data drawn from the peer-reviewed biomedical literature. Assays may also include those observed in high-resolution electrophoretic gels that offer the potential to quantitate many proteins in a single test as well as data gathered by enzymatic or immunologic assays. We are using the Internet World Wide Web (WWW) and the Web browser paradigm as an access method for wide distribution and querying of the Protein Disease Database. The WWW hypertext transfer protocol and its Common Gateway Interface make it possible to build powerful graphical user interfaces that can support easy-to-use data retrieval using query specification forms or images. The details of these interactions are totally transparent to the users of these forms. Using a client-server SQL relational database, user query access, initial data entry and database maintenance are all performed over the Internet with a Web browser. We discuss the underlying design issues, mapping mechanisms and assumptions that we used in constructing the system, data entry, access to the database server, security, and synthesis of derived two-dimensional gel image maps and hypertext documents resulting from SQL database searches.
- Published
- 1995
25. Protein alterations in olfactory neuroblasts from Alzheimer donors.
- Author
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Johnson GS, Basaric-Keys J, Ghanbari HA, Lebovics RS, Lesch KP, Merril CR, Sunderland T, and Wolozin B
- Subjects
- Aged, Aged, 80 and over, Alzheimer Disease diagnosis, Biomarkers analysis, Electrophoresis, Gel, Two-Dimensional, Female, Humans, Male, Matched-Pair Analysis, Middle Aged, Neurons metabolism, Olfactory Nerve cytology, Alzheimer Disease metabolism, Nerve Tissue Proteins metabolism, Olfactory Nerve metabolism
- Abstract
Definitive diagnosis of Alzheimer's disease (AD) is made by pathologic examination of postmortem brain tissue in conjunction with a clinical history of dementia. To date, there are no good biological markers for a positive diagnosis of AD in the living patient. In an effort to identify biological markers useful both in the clinical and pathologic diagnosis of AD, we have investigated disease-specific protein alterations in cultured olfactory neurons. Olfactory neurons are readily accessible by biopsy, can be propagated in primary cell culture as olfactory neuroblasts (ONs), and exhibit several elements of AD brain pathophysiology making them powerful tools for the study of AD. Two-dimensional gel analysis of ON proteins from neuropsychologically evaluated AD donors revealed a set of five proteins (Mr 17-50 kD, pI 4.8-6.7) that were significantly altered in concentration when compared to cells from age-matched controls. Further characterization and microsequence analysis could lead to the identification of proteins that may have important diagnostic or therapeutic value in the treatment of AD.
- Published
- 1994
- Full Text
- View/download PDF
26. Use of silver staining to detect nucleic acids.
- Author
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Mitchell LG, Bodenteich A, and Merril CR
- Subjects
- Diamines, Electrophoresis, Polyacrylamide Gel, Nucleic Acids chemistry, Oxidation-Reduction, Photochemistry, Proteins analysis, Quaternary Ammonium Compounds, Silver Nitrate chemistry, Nucleic Acids analysis, Silver Staining methods
- Published
- 1994
- Full Text
- View/download PDF
27. Characterization of gene expression in the cerebral cortices of rat brains containing subcortical lesions.
- Author
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Wallace W, Brane D, Hsu N, Khowong N, Merril CR, and Haroutunian V
- Subjects
- Animals, Electrophoresis, Gel, Two-Dimensional, Heat-Shock Proteins metabolism, Isoelectric Point, Molecular Weight, N-Methylaspartate pharmacology, Nerve Tissue Proteins biosynthesis, Nerve Tissue Proteins metabolism, Polyribosomes metabolism, Rats, Rats, Sprague-Dawley, Silver Staining, Substantia Innominata drug effects, Substantia Innominata physiology, Cerebral Cortex metabolism, Gene Expression
- Abstract
Neurotoxic lesion of the nucleus basalis of Meynert in the rat brain, which results in the loss of subcortical cholinergic innervation to the cerebral cortex, is an animal model for the cortical cholinergic deficits that are characteristic of Alzheimer's disease. Previously, we have shown that amyloid precursor protein is induced in the cortex in response to this disrupted innervation. We have investigated the synthesis and accumulation of proteins in lesioned versus control cortices. Total proteins from cortices were separated by high resolution two-dimensional gel electrophoresis and visualized by silver stain. Of the greater than 1,000 polypeptides examined, only one exhibited a consistent alteration in the lesioned sample. This unidentified protein (Mr 34 kD, pI 5.5) was normally present in scant amounts but was virtually absent in the lesioned cortex (0.056% total integrated density (TID) and 0.008% TID, respectively; p < 0.04). To investigate gene expression more directly, polysomes purified from lesioned and control cortices were assayed in vitro. Examination of [35S] incorporation into translation products by two-dimensional gels and autoradiography revealed three newly synthesized polypeptide differences in the lesioned samples. One protein (M(r) 47 kD, pI 6.1) exhibited elevated levels with the lesion (0.05% to 0.16%; p = 0.02) while two other proteins (M(r) 34 kD, pI 5.5, and M(r) 33 kD, pI 5.7) exhibited reduced levels (0.20% to 0.04%, p < 0.02, and 0.34% to 0.12%, p = 0.04, respectively).
- Published
- 1994
28. Eliminating mitochondrial DNA competition for nuclear DNA primers.
- Author
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Zullo S, Kennedy JL, Gelernter J, Polymeropoulos MH, Tallini G, Pakstis AJ, Shapiro MB, Merril CR, and Kidd KK
- Subjects
- Base Sequence, DNA biosynthesis, DNA genetics, DNA Primers, Female, Humans, Male, Molecular Sequence Data, Pedigree, RNA biosynthesis, RNA genetics, RNA, Mitochondrial, Restriction Mapping, Sequence Homology, Nucleic Acid, Cell Nucleus metabolism, DNA analysis, Polymerase Chain Reaction methods, Polymorphism, Genetic, RNA analysis, Synaptophysin genetics
- Abstract
Mitochondrial DNA (mtDNA) sequences were synthesized with nuclear DNA (nucDNA) sequence-tagged site (STS) primers by mismatch priming in three independent studies of the human nuclear genome. Mismatch primer binding sites on the mtDNA were identified with from 6- to 10-bp identity at the 3' ends of the primers. In two of three cases, single-stranded mtDNA copies were gel-isolated with intended nucDNA PCR products. During routine screening of the STSs, the radiolabeled gel-isolated products hybridized to polymorphic mtDNA restriction fragments. Intense signals after overnight exposure of radiolabeled PCR probes on Southern blots suggest contaminating mtDNA PCR products. The theoretical annealing temperatures of the mismatches were well below the annealing temperatures of the PCR primers, demonstrating annealing reactions driven by the molar surplus of the primers, that is, mass action. The probability that two primers (either one of a pair or both), designed to amplify nucDNA, will bind to and amplify mtDNA may be as high as 1 in 64, assuming that an identical match with only the 3' hexanucleotide is sufficient for amplification. To circumvent this problem we have developed OLIGFIND, a program that has identified the 104 of 4096 possible hexamers that are not present in human mtDNA. Our results suggest that time could be saved by designing STS primers with one of these 104 hexamers at the 3' end. OLIGFIND can also evaluate primer 3' ends for potential PCR products from mtDNA.
- Published
- 1993
- Full Text
- View/download PDF
29. Chromosomal distribution of 320 genes from a brain cDNA library.
- Author
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Polymeropoulos MH, Xiao H, Sikela JM, Adams M, Venter JC, and Merril CR
- Subjects
- Animals, Cloning, Molecular, DNA, Complementary chemistry, Humans, Hybrid Cells, Information Systems, Lod Score, Polymerase Chain Reaction methods, Polymorphism, Genetic, Brain metabolism, Chromosome Mapping, Chromosomes, Human, DNA, Complementary genetics
- Abstract
We have determined the chromosomal assignment of 320 brain expressed genes by studying the segregation of polymerase chain reaction (PCR) products in human rodent somatic cell hybrids and by genetically mapping polymorphic cDNAs using the CEPH (Centre d'Etude du Polymophisme Humaine) reference pedigrees and database. These mapped genes can function as markers on the physical map of the human genome, as well as serve as candidate disease gene loci. Distribution of these genes to the human chromosomes correlates well with the GC content of the chromosomes. However, the distribution of these genes does not correlate well with the cytogenetic length of each chromosome.
- Published
- 1993
- Full Text
- View/download PDF
30. Search for a genetic event in monozygotic twins discordant for schizophrenia.
- Author
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Polymeropoulos MH, Xiao H, Torrey EF, DeLisi LE, Crow T, and Merril CR
- Subjects
- Adult, Chromosome Mapping, Female, Genetic Markers genetics, Humans, Male, Polymorphism, Genetic genetics, Schizophrenia diagnosis, Schizophrenia, Paranoid diagnosis, Schizophrenia, Paranoid genetics, Schizophrenia, Paranoid psychology, Twins, Monozygotic genetics, Diseases in Twins genetics, Schizophrenia genetics, Schizophrenic Psychology
- Abstract
When monozygotic twins are discordant for the diagnosis of schizophrenia, this discordance has been traditionally attributed to environmental factors acting upon a genome susceptible for the schizophrenia phenotype. The study presented here was designed to examine the occurrence of a genetic event, such as a postzygotic mitotic crossover, that could account for the discordance. Such a postzygotic event could affect cis-acting sequences and result in a phenotype of variable severity. We used molecular genetic methods to evaluate such an event with 94 microsatellite repeat polymorphic markers distributed on all autosomes and the X chromosome in five pairs of monozygotic twins discordant for schizophrenia. In this search, no genetic marker discordances were identified between the co-twins. The lack of a genetic difference may implicate nongenetic factors that are responsible in eliciting or suppressing the phenotype. However, the experiments performed in this study cannot eliminate the possibility that a tissue-specific mitotic crossover might have occurred in one of the discordant twins, which could not have been detected in our current study.
- Published
- 1993
- Full Text
- View/download PDF
31. Altered expression and phosphorylation of amyloid precursor protein in heat shocked neuronal PC12 cells.
- Author
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Johnson G, Refolo LM, Merril CR, and Wallace W
- Subjects
- Amyloid beta-Protein Precursor isolation & purification, Animals, Autoradiography, Electrophoresis, Gel, Two-Dimensional, Electrophoresis, Polyacrylamide Gel, Gene Expression, Heat-Shock Proteins isolation & purification, Hot Temperature, Methionine metabolism, PC12 Cells, Phosphates metabolism, Phosphorus Radioisotopes, Phosphorylation, Sulfur Radioisotopes, Amyloid beta-Protein Precursor biosynthesis, Amyloid beta-Protein Precursor metabolism, Gene Expression Regulation, Neoplastic, Heat-Shock Proteins biosynthesis, Neurons metabolism
- Abstract
The pathology of the Alzheimer's disease (AD) brain, including amyloid plaques, neurofibrillary tangles and neuronal degeneration, indicates that neurons affected by AD exist under conditions of stress. In fact, the brains of AD patients undergo many changes classically associated with the heat shock response, which is one form of a stress response. These changes include reduced protein synthesis, disrupted cytoskeleton, increased number of proteins associated with ubiquitin, and the induction of heat shock proteins. To investigate the response of neurons to stress, we examined neuronal PC12 cells incubated at either 37 degrees C (control cells) or 45 degrees C (heat-shocked cells). After a 30 min exposure at 45 degrees C, the heat-shocked cells exhibited several features characteristic of the classical heat shock response including a 45% reduction in total protein synthesis, the induction of heat shock protein 72, and an increased phosphorylation of the protein synthesis initiation factor eIF-2 alpha. We used this cellular model system to study the neuronal response to stress specifically focusing on protein synthesis elongation factor 2 (EF-2) and the Alzheimer's amyloid precursor protein (APP), the precursor form of beta-amyloid peptide. Hyperphosphorylation of EF-2 has been observed in the neocortex and hippocampus of AD brain. However, in our system, we find no hyperphosphorylation of EF-2 in response to heat shock. Heat-shocked neuronal PC12 cells exhibited two additional APP-like polypeptides not present in controls. We also found a significant decrease in the phosphorylation state of APP in response to heat shock.(ABSTRACT TRUNCATED AT 250 WORDS)
- Published
- 1993
- Full Text
- View/download PDF
32. Reversible phosphorylation of tau to form A68 in heat-shocked neuronal PC12 cells.
- Author
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Wallace W, Johnson G, Sugar J, Merril CR, and Refolo LM
- Subjects
- Animals, Antibodies, Autoradiography, Electrophoresis, Polyacrylamide Gel, Kinetics, Nerve Tissue Proteins biosynthesis, Nerve Tissue Proteins isolation & purification, PC12 Cells, Phosphorylation, Sulfur Radioisotopes, Time Factors, tau Proteins biosynthesis, tau Proteins isolation & purification, Hot Temperature, Nerve Tissue Proteins metabolism, Neurons metabolism, tau Proteins metabolism
- Abstract
A68, the primary protein constituent of Alzheimer's disease-associated neurofibrillary tangles, is an abnormally phosphorylated form of the microtubule-associated protein tau. We find that A68 is formed in neuronal PC12 cells when the cells are subjected to a heat shock (45 degrees C for 30 min). A68 was identified by immunoprecipitation with two different anti-tau antibodies (tau-2 and Alz50). Upon separation by SDS-polyacrylamide gel electrophoresis, the tau immunoprecipitates from heat-shocked cells exhibited an additional polypeptide of reduced electrophoretic mobility (approximately 68 kDa) when compared to control cells. A68 was formed with heat shock in the presence of cycloheximide, suggesting that its production occurred by post-translational modification of existing polypeptides. The tau/A68 polypeptides were identified as phosphoproteins by incorporation of 32P into the immunoprecipitates. The phosphorylation of tau to form A68 was reversed with recovery of the intact cells from the heat shock. Finally, immunoprecipitation of lysates from heat-shocked cells with antibodies to heat shock protein (hsp) 72/73 resulted in co-precipitation of tau with hsp 72, which indicates a stable complex formation between these two proteins. On the other hand, A68 remained unassociated with hsp during the heat shock. These results suggest that tau is reversibly phosphorylated to form A68 in neuronal PC12 cells under conditions of stress.
- Published
- 1993
- Full Text
- View/download PDF
33. Dinucleotide repeat polymorphism at the D11S982E locus.
- Author
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Xiao H, Ide SE, Merril CR, and Polymeropoulos MH
- Subjects
- Alleles, Base Sequence, DNA genetics, Gene Frequency, Genetic Markers, Humans, Molecular Sequence Data, Oligodeoxyribonucleotides genetics, Chromosomes, Human, Pair 11, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid
- Published
- 1993
- Full Text
- View/download PDF
34. Dinucleotide repeat polymorphism at the D14S99E locus.
- Author
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Polymeropoulos MH, Xiao H, Ide SE, and Merril CR
- Subjects
- Alleles, Base Sequence, Chromosomes, Human, Pair 14, DNA genetics, Gene Frequency, Genetic Markers, Humans, Molecular Sequence Data, Oligodeoxyribonucleotides genetics, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid
- Published
- 1993
- Full Text
- View/download PDF
35. Increased apolipoprotein-E concentrations in individuals suffering chronic low back syndrome identified by two-dimensional gel electrophoresis.
- Author
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VanderPutten DM, Cameron BM, and Merril CR
- Subjects
- Amino Acid Sequence, Biomechanical Phenomena, Electrophoresis, Gel, Two-Dimensional, Humans, Immunoblotting, Molecular Sequence Data, Pain blood, Apolipoproteins E metabolism, Low Back Pain blood
- Abstract
Non-biased screening of plasma proteins by two-dimensional gel electrophoresis from individuals suffering low back syndrome revealed a polypeptide spot that was increased 2-5-fold over the concentration found in normal control individuals. The apparent molecular weight (34-36 kDa) and pI (5.7) of this spot suggested that it might be apolipoprotein-E. Immunoblot analysis showed that the polypeptide was reactive with anti-apolipoprotein-E antibodies. N-terminal amino acid microsequence confirmed the identify of this polypeptide as apolipoprotein-E. We have determined that elevated plasma levels of apolipoprotein-E is associated with inflammation and nerve damage.
- Published
- 1993
36. Identification of a brain-specific human cerebrospinal fluid glycoprotein, beta-trace protein.
- Author
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Harrington MG, Aebersold R, Martin BM, Merril CR, and Hood L
- Subjects
- Adult, Amino Acid Sequence, Antibody Formation, Beta-Globulins immunology, Cerebrospinal Fluid Proteins immunology, Female, Glycosylation, Humans, Infant, Lipocalins, Male, Middle Aged, Molecular Sequence Data, Nervous System Diseases metabolism, Organ Specificity physiology, Beta-Globulins analysis, Brain Chemistry physiology, Cerebrospinal Fluid Proteins analysis, Intramolecular Oxidoreductases
- Abstract
A prominent human cerebrospinal fluid (CSF) protein, P5, identified at mass 19-24 kDa and charge 5.5, by two-dimensional electrophoresis (2DE) and silver staining, has been previously demonstrated to be reduced in quantity in the CSF of patients with multiple sclerosis and schizophrenia. We report the purification and partial amino acid sequences from five tryptic fragments of P5. These sequences are not those of any known sequence in the Protein Identification Resource (PIR release 31) database. Synthetic peptides from two of the sequences were used to raise rabbit polyclonal antibodies. These antibodies detected P5 on 2DE blots of normal CSF proteins and other proteins of the same mass with a charge distribution between 5.17-8.5. These proteins comprise 5-10% of the total CSF protein and their mass, charge, abundance and predominance in CSF over plasma are consistent with a protein that had been initially characterized with antibodies, beta-trace protein. Glycosidase studies confirm that most of these proteins are due to sialic acid modifications that are N-linked to an 18 kDa protein, but other charge and mass variations also exist. 2DE blots of 26 types of human tissue and body fluid were immunostained. Of these, anti-P5 serum detected proteins of the same mass and charge as beta-trace protein only in brain samples. Proteins of different mass and charge from beta-trace protein were clearly immunostained in samples of eight tissues.(ABSTRACT TRUNCATED AT 250 WORDS)
- Published
- 1993
37. Identification and use of constitutive proteins for the normalization of high resolution electrophoretograms.
- Author
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Merril CR, Creed GJ, Joy J, and Olson AD
- Subjects
- Animals, Genitalia, Male chemistry, Male, Moths, Algorithms, Electrophoresis, Gel, Two-Dimensional, Proteins analysis
- Abstract
Quantitative inter-gel comparisons of proteins separated by high resolution two-dimensional protein electrophoresis present a number of problems. These problems may arise from: variations in pipetting and other mechanical manipulations of samples, protein loss during transfer from the first to the second gel dimension, variations in staining, and/or variations in film development during autoradiography, in the case of radioactively labeled proteins. This study presents a discussion of these issues and a normalization algorithm to deal with variations, which relies on a class of proteins present in most biological samples which by their nature may be considered internal standards. This class consists of proteins which are controlled by constitutive genes. Constitutive genes are genes that are expressed constantly. We have developed an algorithm which is currently available as a subroutine, 'FINDCONS', in the computerized densitometry and protein comparison analysis program, developed by Olson & Miller (1988). This algorithm identifies potentially 'constitutive' proteins. A normalization method employing these potentially 'constitutive' proteins was compared to several others by examining 2D-electrophoretograms of proteins from developing gypsy moths (Lymantria dispar L.) insect tissue. Following normalization, inter-gel comparisons of spots, which were 'identified' as 'constitutive', were observed to vary less in density than when no normalization method was used, or when normalization based on total integrated spot density was used. In addition to its use as a normalization tool, this algorithm and the subroutine FINDCONS may be useful as an aid in biological studies to identify 'constitutive' proteins.
- Published
- 1993
38. Dinucleotide repeat polymorphism at the D18S74E locus.
- Author
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Polymeropoulos MH, Xiao H, and Merril CR
- Subjects
- Animals, Base Sequence, Genetic Markers, Humans, Hybrid Cells, Molecular Sequence Data, Oligodeoxyribonucleotides, Polymerase Chain Reaction, Chromosomes, Human, Pair 18, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid
- Published
- 1992
- Full Text
- View/download PDF
39. Tetranucleotide repeat polymorphism at the human myelin basic protein gene (MBP).
- Author
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Polymeropoulos MH, Xiao H, and Merril CR
- Subjects
- Base Sequence, Chromosomes, Human, Pair 18, Gene Frequency, Humans, Molecular Sequence Data, Myelin Basic Protein genetics, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid
- Published
- 1992
- Full Text
- View/download PDF
40. Immunochemical characterization of a monoclonal antibody specific for Alzheimer's disease associated protein.
- Author
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Bodenteich A, Mitchell LG, Merril CR, Miller BE, Ghanbari HA, and Heegaard NH
- Subjects
- Antibody Specificity, Antigen-Antibody Reactions, Brain metabolism, Humans, Immunoelectrophoresis, Immunohistochemistry, Regression Analysis, Alzheimer Disease immunology, Antibodies, Monoclonal immunology, Antigens immunology, Nerve Tissue Proteins immunology
- Abstract
In this study, the monoclonal antibody PHF-1 which recognizes epitopes unique to Alzheimer's disease associated proteins (ADAP) has been characterized. Crossed affinity immunoelectrophoresis was used to estimate the binding constant for the interaction of PHF-1 with ADAP and to estimate the fraction of PHF-1 reactive protein. The binding constant of PHF-1 was determined to be 1.3 x 10(-8) M. Furthermore, the effect of dephosphorylation on the electrophoretic pattern of the PHF-1 reactive protein and the ensuing changes in its immunoreactivity were demonstrated.
- Published
- 1992
- Full Text
- View/download PDF
41. Dinucleotide repeat polymorphisms at the D3S1246 and D3S1247 loci.
- Author
-
Xiao H, Merril CR, and Polymeropoulos MH
- Subjects
- Base Sequence, Female, Humans, Male, Molecular Sequence Data, Oligonucleotides, Pedigree, Chromosomes, Human, Pair 3, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid
- Published
- 1992
- Full Text
- View/download PDF
42. Dinucleotide repeat polymorphism at the D7S476 locus.
- Author
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Xiao H, Merril CR, and Polymeropoulos MH
- Subjects
- Animals, Base Sequence, Chromosomes, Human, Pair 7, DNA, Single-Stranded, Humans, Hybrid Cells, Molecular Sequence Data, Oligodeoxyribonucleotides, Polymerase Chain Reaction, Rodentia, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid
- Published
- 1992
- Full Text
- View/download PDF
43. Protein differences in tau mutant hamsters: candidate clock proteins.
- Author
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Joy JE, Johnson GS, Lazar T, Ralph MR, Hochstrasser AC, Menaker M, and Merril CR
- Subjects
- Animals, Circadian Rhythm, Cricetinae, Electrophoresis, Polyacrylamide Gel, Genotype, Photometry, Subcellular Fractions metabolism, Suprachiasmatic Nucleus chemistry, Suprachiasmatic Nucleus metabolism, Biological Clocks physiology, Nerve Tissue Proteins genetics
- Abstract
In the tau mutant hamster, the period of the circadian rhythm is shortened from about 24 h to about 22 h in heterozygotes and to about 20 h in homozygotes. Understanding the biochemical basis of the period changes in the tau mutant may elucidate the regulation of the vertebrate pacemaker. Using two-dimensional gel electrophoresis, we have found two sets of proteins that differ between the different genotypes. P33tau (about 33 kDa; pI 6.5) was found in all gels from wild type and heterozygous animals, but was absent in gels from all except one of the homozygous mutant animals. P32tau (about 32 kDa; pI 4.8) was a chain of spots, which showed a striking difference in pattern between gels from wild type animals and from mutant animals. P33tau was greatly enriched in soluble cellular fractions, whereas P32tau was found only in insoluble fractions. These differences between P33tau and P32tau were apparent in gels from both SCN and cortical tissue, suggesting that both proteins are distributed throughout the brain. These proteins should be useful as new tools to explore the biochemistry of circadian pacemakers.
- Published
- 1992
- Full Text
- View/download PDF
44. Dinucleotide repeat in the human mitochondrial D-loop.
- Author
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Bodenteich A, Mitchell LG, Polymeropoulos MH, and Merril CR
- Subjects
- Base Sequence, Humans, Molecular Sequence Data, Oligodeoxyribonucleotides, Polymerase Chain Reaction, DNA, Mitochondrial, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid
- Published
- 1992
- Full Text
- View/download PDF
45. Dinucleotide repeat polymorphism at the human c-myc oncogene locus (MYC).
- Author
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Polymeropoulos MH, Xiao H, and Merril CR
- Subjects
- Alleles, Base Sequence, Chromosome Mapping, Gene Frequency, Genes, Dominant, Heterozygote, Humans, Molecular Sequence Data, Oligodeoxyribonucleotides, Polymerase Chain Reaction methods, Chromosomes, Human, Pair 8, Genes, myc, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid
- Published
- 1992
- Full Text
- View/download PDF
46. Tetranucleotide repeat polymorphism at the human beta-actin related pseudogene H-beta-Ac-psi-2 (ACTBP2).
- Author
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Polymeropoulos MH, Rath DS, Xiao H, and Merril CR
- Subjects
- Base Sequence, Gene Frequency, Humans, Molecular Sequence Data, Oligodeoxyribonucleotides, Polymerase Chain Reaction, Actins genetics, Chromosomes, Human, Pair 6, Polymorphism, Genetic, Pseudogenes, Repetitive Sequences, Nucleic Acid
- Published
- 1992
- Full Text
- View/download PDF
47. Chromosomal assignment of 46 brain cDNAs.
- Author
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Polymeropoulos MH, Xiao H, Glodek A, Gorski M, Adams MD, Moreno RF, Fitzgerald MG, Venter JC, and Merril CR
- Subjects
- Animals, Base Sequence, Cricetinae, DNA isolation & purification, Human Genome Project, Humans, Hybrid Cells physiology, Mice, Molecular Sequence Data, Oligodeoxyribonucleotides, Polymerase Chain Reaction, Brain physiology, Chromosome Mapping, DNA genetics, Sequence Tagged Sites
- Abstract
Expressed sequence tags (ESTs) have been obtained from several hundred brain cDNAs as an initial effort to characterize expressed brain genes. These ESTs will become tools for human genome mapping and they will also provide candidate causative genes for inherited disorders affecting the central nervous system. We have developed a procedure for the rapid chromosomal assignment of these ESTs: cDNA sequences are first analyzed by a computer program to determine regions likely not to be interrupted by introns in the genomic DNA. A pair of oligonucleotide primers is then designed to amplify this region by the polymerase chain reaction using DNA template from human-rodent somatic cell hybrid chromosomal panels. The chromosomal assignment of the cDNA is determined by studying the segregation of the amplified products in these panels. In this paper we describe the mapping of 46 brain ESTs, as well as observations on the amplification of rodent sequences.
- Published
- 1992
- Full Text
- View/download PDF
48. Cerebrospinal fluid protein variations in common to Alzheimer's disease and schizophrenia.
- Author
-
Johnson G, Brane D, Block W, van Kammen DP, Gurklis J, Peters JL, Wyatt RJ, Kirch DG, Ghanbari HA, and Merril CR
- Subjects
- Acute-Phase Proteins physiology, Adult, Alzheimer Disease etiology, Humans, Immunoenzyme Techniques, Iron metabolism, Male, Middle Aged, Schizophrenia etiology, Alzheimer Disease cerebrospinal fluid, Cerebrospinal Fluid Proteins analysis, Electrophoresis, Gel, Two-Dimensional, Haptoglobins cerebrospinal fluid, Schizophrenia cerebrospinal fluid, Silver Staining
- Abstract
Analysis of silver stained two-dimensional (2D) gels of cerebrospinal fluid (CSF) from 27 patients with schizophrenia (SCZ) and 10 patients with Alzheimer's disease (AD) revealed an increase in the relative amount of a polypeptide of 18,000M(r) and isoelectric point of 6.5 when compared to the appropriate controls. This protein was identified by its electrophoretic characteristics and by immune analysis of Western blots as an isoform of alpha-2 haptoglobin, provisionally identified as alpha-2FS haptoglobin. Alzheimer's disease versus control CSF samples showed a 6.8-fold increase in the percent mean density value of this haptoglobin isoform (n = 10 AD vs 11 control; P > 0.025) while a 4.4-fold increase was observed in the schizophrenic patients (n = 17 SCZ vs 10 control; P > 0.001). Two additional polypeptides (proteins '127' and '128') of 40,000 M(r) and isoelectric points 5.7 and 5.9, respectively, described previously by this laboratory, were found in the CSF of 27% of schizophrenics, 23% of the Alzheimer's disease patients, and 4% of the controls in the current study. The presence of proteins 127 and 128, as well as the increased concentrations of alpha-2 haptoglobin in the CSF of Alzheimer's disease and schizophrenic patients, may be useful as diagnostic biological markers. They may also indicate a common pathophysiology between these diseases.
- Published
- 1992
49. Haloperidol induced CSF protein variations in schizophrenic patients: as studied by two-dimensional electrophoresis.
- Author
-
Johnson G, Brane D, van Kammen DP, Gurklis J, Peters JL, Perel JM, Ghanbari HA, and Merril CR
- Subjects
- Adult, Cerebrospinal Fluid Proteins isolation & purification, Electrophoresis, Gel, Two-Dimensional, Haloperidol administration & dosage, Humans, Male, Middle Aged, Cerebrospinal Fluid Proteins drug effects, Haloperidol pharmacology, Schizophrenia cerebrospinal fluid, Schizophrenia drug therapy
- Abstract
High resolution two-dimensional electrophoresis of cerebrospinal fluid (CSF) from 10 schizophrenic patients demonstrated a 21% average difference in the number of proteins which could be detected in patients undergoing haloperidol therapy when compared with CSF from the same patients after withdrawal from neuroleptic treatment. Proteins affected were trace proteins, as we found no significant variation in either the total CSF protein content or the integrated protein density on each electrophoretic gel. Three mechanisms which might account for these observations are: (1) a small change in liver protein synthesis or degradation would have little if any visible effect on the concentration of major blood or CSF proteins, such as albumin, but it could significantly alter trace proteins, such as alpha 2-haptoglobin, since their concentrations are orders of magnitude less than that of the major proteins, (2) haloperidol might alter the blood-CSF protein filtration system which could affect the visibility of the trace proteins, and (3) proteins synthesized in the Central Nervous System (CNS) or enhanced in the CSF could be differentially affected by haloperidol. While additional research will be required to determine the basis for the effects of haloperidol on CSF proteins, the current studies provide information which may be helpful in delineating disease specific protein alterations from those induced by drug therapy.
- Published
- 1992
50. A lifetime of retinal light exposure does not appear to increase mitochondrial mutations.
- Author
-
Bodenteich A, Mitchell LG, and Merril CR
- Subjects
- Aged, Aging radiation effects, Base Sequence, Blotting, Southern, Cloning, Molecular, DNA Repair genetics, Electron Transport Complex IV genetics, Female, Humans, Molecular Sequence Data, NADH Dehydrogenase genetics, RNA, Transfer, Gly genetics, Ultraviolet Rays adverse effects, Aging genetics, DNA, Mitochondrial genetics, Mitochondria radiation effects, Mutation, Retina radiation effects
- Abstract
Recently, there have been a number of reports of an accumulation of mutations in the mitochondrial (mt) genome with age. Such mutations may be due in part to the mt oxidative metabolic pathways which provide most of the cell's energy, but also generate free radicals. In addition, the mt genome in some tissues, such as the retina, may also accumulate mutations from the effects of ultraviolet light. To obtain information concerning the possible accumulation of retinal mt mutations with age, we cloned retinal mt DNA from a 71-year-old person. Thirty-two kilobases of sequence from 83 independently isolated clones representing two regions, a coding and a noncoding region, of the mt genome were obtained. Three polymorphisms between these sequences and the standard 'Anderson sequence' were discovered. Only one heteroplasmic mutation was found. These results confirm the low somatic mutation rate found in prior studies utilizing different types of human tissues. In addition, these results suggest that there is little if any accumulated damage to the mt DNA of the retina during normal aging.
- Published
- 1991
- Full Text
- View/download PDF
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