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1.
Int J STD AIDS ; 22(6): 310-4, 2011 Jun.
Article in English | MEDLINE | ID: mdl-21680665

ABSTRACT

We evaluated the accuracy of heat-denatured, amplification-boosted ultrasensitive p24 assay (Up24) compared with reverse transcriptase polymerase chain reaction (RT-PCR). We tested 394 samples from Ugandans infected with HIV-1 non-B subtypes. We compared Up24 levels (HIV-1 p24 Core Profile enzyme-linked immunosorbent assay (ELISA), NEN Life Science Products) to RNA viral loads (Amplicor HIV-1 Monitor 1.5, Roche) by linear regression, and calculated sensitivity, specificity, positive and negative predictive values. Median viral load was 4.9 log10 copies/mL (interquartile range [IQR], 2.6-5.5); 114 samples (29%) were undetectable (<400 copies/mL). Sensitivity of the Up24 assay to detect viral load ≥400 copies/mL was 69%, specificity was 67%, and positive and negative predictive values were 84% and 47%, respectively. Sensitivity of Up24 was 90%, 80%, 68%, 62% and 45% to detect viral loads of >500,000, 250,000-500,000, 100,000-250,000, 50,000-100,000 and 400-50,000 copies/mL, respectively. In conclusion, when compared with RT-PCR for patients infected with non-B subtypes, the Up24 demonstrated limited sensitivity especially at low viral loads. Moreover, the Up24 was positive in 33% of samples deemed undetectable by RT-PCR, which may limit the use of the Up24 to detect viral suppression.


Subject(s)
HIV Core Protein p24/analysis , HIV Infections/diagnosis , Adult , Developing Countries , Enzyme-Linked Immunosorbent Assay/methods , HIV Infections/blood , HIV Infections/immunology , HIV-1/isolation & purification , Humans , Linear Models , Protein Denaturation , Reverse Transcriptase Polymerase Chain Reaction/methods , Sensitivity and Specificity , Uganda , Viral Load/economics , Viral Load/methods
2.
Curationis ; 29(3): 15-23, 2006 Aug.
Article in English | MEDLINE | ID: mdl-17131605

ABSTRACT

African traditional health practitioners are an important source of health care for many South Africans. Thus, they are a health resource in this society. However, the integration of traditional health practitioners into the mainstream of health care is a complex process. Various factors contribute to this complexity, including the skepticism and reservation with which some western health practitioners view traditional health practitioners. This paper highlights the perceived strengths and weaknesses of the traditional healing system for people living with HIV/AIDS, as perceived by western health practitioners. The use of traditional practitioners as a choice of health care is attributed to both the strengths and weaknesses of this system of health care. The strength of the traditional healing system is in its sharing of the worldview and belief system of its users, it being an alternative to an inefficient western health care system (official system), privacy and absence of time limitations per consultation, treating patients psychologically, and scientifically unexplained physiological relief of the symptoms of specific illnesses. The perceived weaknesses of the traditional healing system include harmful treatment regimens, especially for people living with HIV/AIDS; prolonging the seeking of appropriate health care when traditional remedies fail to produce the desired effect; destroying interpersonal relationships of people living with HIV/AIDS through witchcraft accusations; psychological torment caused by the belief that HIV/AIDS can be cured by traditional remedies/intervention; and increasing the workload of western practitioners who are requested by patients to conduct multiple HIV tests after undergoing various traditional treatment regimens to cure HIV/AIDS. It is recommended that traditional practitioners be encouraged to adapt harmful traditional healing practices to the benefit of their patients in a non-judgemental and non-critical manner. In addition, joint workshops should be conducted with traditional and western practitioners to demystify traditional healing practices.


Subject(s)
Attitude of Health Personnel/ethnology , Community Health Workers/psychology , HIV Infections/ethnology , Medicine, African Traditional , Nursing Staff/psychology , Physicians, Family/psychology , AIDS Serodiagnosis/psychology , Female , HIV Infections/diagnosis , HIV Infections/therapy , Health Knowledge, Attitudes, Practice , Health Services Accessibility , Health Services Needs and Demand , Humans , Interpersonal Relations , Male , Nursing Methodology Research , Patient Acceptance of Health Care/ethnology , Qualitative Research , South Africa/epidemiology , Surveys and Questionnaires , Workload
3.
Nucleic Acids Res ; 29(19): 3965-74, 2001 Oct 01.
Article in English | MEDLINE | ID: mdl-11574678

ABSTRACT

The antisense activity of oligomers with 2'-O-methyl (2'-O-Me) phosphorothioate, 2'-O-methoxyethyl (2'-O-MOE) phosphorothioate, morpholino and peptide nucleic acid (PNA) backbones was investigated using a splicing assay in which the modified oligonucleotides blocked aberrant and restored correct splicing of modified enhanced green fluorescent protein (EGFP) precursor to mRNA (pre-mRNA), generating properly translated EGFP. In this approach, antisense activity of each oligomer was directly proportional to up-regulation of the EGFP reporter. This provided a positive, quantitative readout for sequence-specific antisense effects of the oligomers in the nuclei of individual cells. Nuclear localization of fluorescent labeled oligomers confirmed validity of the functional assay. The results showed that the free uptake and the antisense efficacy of neutral morpholino derivatives and cationic PNA were much higher than that of negatively charged 2'-O-Me and 2'-O-MOE congeners. The effects of the PNA oligomers were observed to be dependent on the number of L-lysine (Lys) residues at the C-terminus. The experiments suggest that the PNA containing Lys was taken up by a mechanism similar to that of cell-penetrating homeodomain proteins and that the Lys tail enhanced intracellular accumulation of PNA oligomer without affecting its ability to reach and hybridize to the target sequence.


Subject(s)
Cell Nucleus/drug effects , Oligonucleotides, Antisense/chemistry , Oligonucleotides, Antisense/pharmacology , Anions/chemistry , Biological Transport , Cations/chemistry , Cell Nucleus/metabolism , Genes, Reporter , Globins/genetics , Green Fluorescent Proteins , HeLa Cells , Humans , Indicators and Reagents/metabolism , Introns , Luminescent Proteins/biosynthesis , Luminescent Proteins/genetics , Lysine/physiology , Morpholines/metabolism , Morpholines/pharmacology , Oligonucleotides, Antisense/metabolism , Organothiophosphorus Compounds/metabolism , Organothiophosphorus Compounds/pharmacology , Peptides/chemistry , Point Mutation , RNA Precursors/genetics , RNA Splicing , Thionucleotides/pharmacology
4.
Proc Natl Acad Sci U S A ; 97(17): 9591-6, 2000 Aug 15.
Article in English | MEDLINE | ID: mdl-10944225

ABSTRACT

Mononuclear cells from peripheral blood of thalassemic patients were treated with morpholino oligonucleotides antisense to aberrant splice sites in mutant beta-globin precursor mRNAs (pre-mRNAs). The oligonucleotides restored correct splicing and translation of beta-globin mRNA, increasing the hemoglobin (Hb) A synthesis in erythroid cells from patients with IVS2-654/beta(E), IVS2-745/IVS2-745, and IVS2-745/IVS2-1 genotypes. The maximal Hb A level for repaired IVS2-745 mutation was approximately 30% of normal; Hb A was still detectable 9 days after a single treatment with oligonucleotide. Thus, expression of defective beta-globin genes was repaired and significant level of Hb A was restored in a cell population that would be targeted in clinical applications of this approach.


Subject(s)
Erythrocytes/metabolism , Genetic Therapy , Hemoglobin A/biosynthesis , Hemoglobin A/genetics , beta-Thalassemia/blood , beta-Thalassemia/therapy , Cell Nucleus/genetics , Cells, Cultured , Erythroid Precursor Cells/metabolism , Fluorescent Antibody Technique , Globins/genetics , Humans , Mutation/genetics , Oligonucleotides, Antisense/genetics , Oligonucleotides, Antisense/therapeutic use , RNA Precursors/genetics , RNA Precursors/metabolism , RNA Splicing/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , Spliceosomes/genetics , Time Factors , beta-Thalassemia/genetics
5.
Antisense Nucleic Acid Drug Dev ; 10(3): 163-76, 2000 Jun.
Article in English | MEDLINE | ID: mdl-10905553

ABSTRACT

The phosphorodiamidate Morpholino oligomers (PMO) are a new class of antisense agents that inhibit gene expression by binding to RNA and sterically blocking processing or translation. In a search for a Morpholino agent that would inhibit cell proliferation, it was found that oligomers directed against c-myc, a gene involved in control of the cell cycle, were effective. The sequence specificity and mechanism of action of one agent were determined. The 20-mer 126 lowers c-myc protein levels in treated cells and arrests cells in G0/G1 of the cell cycle. It also acts at the RNA level to inhibit normal pre-mRNA splicing and instead produces an aberrantly spliced mRNA. Irrelevant and mispair control oligomers indicated that the observed antiproliferative effect was sequence specific. This was confirmed in a reporter gene model system using a c-myc 5'-untranslated region (5'-UTR) fused to a cDNA copy of the insect luciferase gene. We conclude that 126 is acting through an antisense mechanism involving Watson-Crick hydrogen bonding to its target RNA. A specific antisense agent directed against a cell cycle-associated gene mRNA may be useful as a therapeutic in diseases characterized by excess cell proliferation, such as restenosis following balloon angioplasty or cancer.


Subject(s)
Genes, myc/drug effects , Growth Inhibitors/pharmacology , Morpholines/pharmacology , Oligonucleotides, Antisense/pharmacology , Animals , Cell Division/drug effects , Cell Line, Transformed/cytology , Cell Line, Transformed/drug effects , Fibroblasts/cytology , Fibroblasts/drug effects , Fibroblasts/metabolism , HeLa Cells , Humans , Protein Processing, Post-Translational/drug effects , Proto-Oncogene Proteins c-myc/genetics , Proto-Oncogene Proteins c-myc/metabolism , Rats , Sensitivity and Specificity , Stereoisomerism , Thionucleotides/pharmacology
6.
ASAIO J ; 45(1): 109-12, 1999.
Article in English | MEDLINE | ID: mdl-9952019

ABSTRACT

Physiologic clotting of a dialysis circuit was achieved using an in vitro method. The closed loop method used a circuit filled with defibrinated bovine blood. To achieve clotting, a slow infusion (5-15 ml/hr) of ACD bovine plasma was performed. The rate and amount of infusion of the plasma allowed for careful control of the amount of clotting in the circuit and dialyzer. By varying these, a range of clotting (10-40% dialyzer volume reduction) was achieved in a timeframe of approximately 100 minutes. The results of the technique closely mimicked clinical examples of dialysis circuits in appearance. Residual blood clots were present in the venous drip chamber filter, in nonstreamlined areas of the bloodline (e.g., bond joint gaps), and throughout the dialyzer fibers and header space. As an additional check of clotting distribution, the performance of the dialyzers compared favorably with previously reported effects of reuse. Solute clearances decreased with the fiber bundle volumes (FBV) of the devices. Correlations of small molecular weight (MW) (urea, creatinine) clearances with FBV were linear, whereas middle MW (vitamin B12) clearances and the ultrafiltration coefficient (kUF) had nonlinear correlations. The results indicate that this clotting method can be used as a valuable tool for a qualitative in vitro assessment of a hemodialysis extracorporeal circuit.


Subject(s)
Blood Coagulation , Renal Dialysis/methods , Animals , Cattle , Equipment Design , Extracorporeal Circulation , Linear Models , Models, Biological , Nonlinear Dynamics , Regression Analysis
7.
Biochim Biophys Acta ; 1489(1): 141-58, 1999 Dec 10.
Article in English | MEDLINE | ID: mdl-10807004

ABSTRACT

RNase H-competent phosphorothioates (S-DNAs) have dominated the antisense field in large part because they offer reasonable resistance to nucleases, they afford good efficacy in cell-free test systems, they can be targeted against sites throughout the RNA transcript of a gene, and they are widely available from commercial sources at modest prices. However, these merits are counterbalanced by significant limitations, including: degradation by nucleases, poor in-cell targeting predictability, low sequence specificity, and a variety of non-antisense activities. In cell-free and cultured-cell systems where one wishes to block the translation of a messenger RNA coding for a normal protein, RNase H-independent morpholino antisense oligos provide complete resistance to nucleases, generally good targeting predictability, generally high in-cell efficacy, excellent sequence specificity, and very preliminary results suggest they may exhibit little non-antisense activity.


Subject(s)
Morpholines/chemistry , Oligonucleotides, Antisense/chemistry , Ribonuclease H/metabolism , Alternative Splicing/drug effects , Cell-Free System/enzymology , Drug Carriers/chemistry , Drug Design , Gene Expression/drug effects , Gene Targeting/methods , HeLa Cells , Humans , Nucleic Acid Hybridization/genetics , Oligonucleotides, Antisense/genetics , Oligonucleotides, Antisense/metabolism , Oligonucleotides, Antisense/pharmacology , Protein Biosynthesis/drug effects , RNA Processing, Post-Transcriptional/drug effects , Substrate Specificity , Thionucleotides/chemistry , Thionucleotides/genetics , Thionucleotides/pharmacology
8.
Antisense Nucleic Acid Drug Dev ; 7(3): 151-7, 1997 Jun.
Article in English | MEDLINE | ID: mdl-9212905

ABSTRACT

Cell-free translation studies were carried out to compare the efficacy and specificity of four antisense structural types: DNA, phosphorothioate DNA (S-DNA), 2'-O-methyl RNA, and Morpholino oligos, a novel antisense structural type. In these studies, translational inhibition was assessed for two 20-mers of each structural type, where one 20-mer was complementary to its target sequence in rabbit alpha-globin mRNA and the other 20-mer contained three mispairs to that same target sequence. It is shown that at low concentration of antisense oligomer (50 nM) all four types provide high specificity, but the Morpholino oligos and 2'-O-methyl RNA afford better efficacy. At high oligomer concentration (3.5 microM), all four types provide high efficacy, but the Morpholino oligos and 2'-O-methyl RNA provide substantially better specificity than the DNA and S-DNA.


Subject(s)
DNA, Antisense/pharmacology , Globins/genetics , Protein Biosynthesis/drug effects , RNA, Antisense/pharmacology , RNA, Messenger/antagonists & inhibitors , Animals , Base Sequence , Cell-Free System , DNA, Antisense/chemistry , Globins/biosynthesis , Hot Temperature , Molecular Sequence Data , Molecular Structure , Morpholines/chemistry , Morpholines/pharmacology , Nucleic Acid Denaturation , RNA, Antisense/chemistry , Rabbits , Reticulocytes/chemistry , Ribonuclease H/metabolism , Structure-Activity Relationship , Thionucleotides/chemistry , Thionucleotides/pharmacology
9.
Antisense Nucleic Acid Drug Dev ; 7(3): 187-95, 1997 Jun.
Article in English | MEDLINE | ID: mdl-9212909

ABSTRACT

Antisense promised major advances in treating a broad range of intractable diseases, but in recent years progress has been stymied by technical problems, most notably inadequate specificity, ineffective delivery into the proper subcellular compartment, and unpredictable activity within cells. Herein is an overview of the design, preparation, and properties of Morpholino oligos, a novel antisense structural type that solves the sequence specificity problem and provides high and predictable activity in cells. Morpholino oligos also exhibit little or no nonantisense activity, afford good water solubility, are immune to nucleases, and are designed to have low production costs.


Subject(s)
Morpholines , Oligonucleotides, Antisense , Animals , Cell-Free System , Cells, Cultured , Endocytosis , Mice , Molecular Structure , Morpholines/chemical synthesis , Morpholines/isolation & purification , Morpholines/pharmacology , Nucleic Acid Denaturation , Oligonucleotides, Antisense/chemistry , Oligonucleotides, Antisense/isolation & purification , Oligonucleotides, Antisense/pharmacology , Rats , Solubility , Structure-Activity Relationship , Water
10.
Antisense Nucleic Acid Drug Dev ; 7(2): 63-70, 1997 Apr.
Article in English | MEDLINE | ID: mdl-9149841

ABSTRACT

Morpholino and phosphorothioate (S-DNA) antisense oligos were compared in both cell-free and in-cell translation systems. In the most stringent test of specificity in the cell-free system, a globin-targeted S-DNA oligo was found to inhibit its target sequence at concentrations of 10 nM and above, but the sequence-specific component of this inhibition dropped below 50% at concentrations of 100 nM and above. A corresponding Morpholino oligo achieved even higher inhibition at 10 nM, but in contrast to the S-DNA, with the Morpholino, the sequence-specific component of this inhibition remained above 93% at a concentration of 3000 nM. In this same cell-free test system, several S-DNA oligos exhibited substantial undesired nonantisense effects at concentrations of 300 nM and above, whereas corresponding Morpholino oligos exhibited little or no nonantisense activity through a concentration of 3000 nM. In scrape-loaded HeLa cells, both globin-targeted and HBV-targeted S-DNAs (both antisense and control oligos) generally failed to achieve significant translational inhibition at extracellular concentrations up to 3000 nM. In contrast, the Morpholino oligos achieved effective and specific translational inhibition at extracellular concentrations ranging from 30 nM to 3000 nM.


Subject(s)
Morpholines/metabolism , Oligonucleotides, Antisense/metabolism , Thionucleotides/metabolism , Animals , Cell-Free System , Globins/genetics , HeLa Cells , Hepatitis B virus/genetics , Humans , Luciferases/genetics , Plasmids , Protein Biosynthesis , RNA, Messenger/genetics , Rabbits
11.
Biotechniques ; 23(6): 1116-21, 1997 Dec.
Article in English | MEDLINE | ID: mdl-9421646

ABSTRACT

Nucleic acid isolation for amplification-based diagnostics requires techniques that do not co-purify inhibitors of DNA polymerases. Also, other requirements for an ideal sample preparation technology include ease of use, capability for automation, high recovery and the use of nontoxic reagents. Affinity purification techniques provide high purification factor with minimal sample processing. Hybridization is the affinity interaction specific to nucleic acids and thus provides a uniquely advantageous method for purifying DNA or RNA for subsequent manipulation. Nonionic (morpholino) probes (Neu-Probes, AntiVirals, Corvallis, OR, USA) have several unique hybridization properties, including resistance to nucleases and the ability to hybridize independently of salt concentration. Therefore, such probes provide advantages over DNA probes for sample preparation by hybridization capture. Three formats for hybridization-based purification of human immunodeficiency virus (HIV) RNA were evaluated using RNA transcripts spiked into crude lysates of normal human plasma. Indirect capture used streptavidin-coated microparticles to capture hybrids of biotinylated capture probes and HIV RNA. Direct capture used particles precoated with probes. In addition, a novel method for acceleration of sequence-specific hybridization was developed and shown to give consistently high recoveries.


Subject(s)
Morpholines , Oligonucleotide Probes , RNA, Viral/isolation & purification , DNA-Directed DNA Polymerase/genetics , HIV/chemistry , HIV/genetics , Humans , Hypotonic Solutions , Nucleic Acid Hybridization , Nucleic Acid Synthesis Inhibitors , Oligonucleotides, Antisense/genetics , Osmolar Concentration , Polymerase Chain Reaction , Salts , Sensitivity and Specificity
12.
Antisense Nucleic Acid Drug Dev ; 6(3): 169-75, 1996.
Article in English | MEDLINE | ID: mdl-8915501

ABSTRACT

We report a simple and effective means for delivering Morpholino antisense oligos into the cytosol of cultured anchorage-dependent animal cells. This method, referred to as scrape-loading, is carried out in a matter of seconds, uses a common inexpensive laboratory implement, and has minimal detrimental impact on the cells. Using this delivery method, a Morpholino oligo present at 0.1 microM and 1 microM in the extracellular medium inhibited its targeted genetic sequence within cultured Hela cells at levels of 56% and 85%, respectively. Lack of inhibition by two control Morpholino oligos at concentrations up to 3 microM indicates good sequence specificity by this structural type. Also described is a test system for simple, rapid, and sensitive quantitation of antisense activity in cultured cells.


Subject(s)
Cytological Techniques , Cytoplasm , Morpholines/chemistry , Oligonucleotides, Antisense/administration & dosage , Animals , Base Sequence , Cell Line , Enzyme Induction , HeLa Cells , Humans , Luciferases/biosynthesis , Molecular Sequence Data , Oligonucleotides, Antisense/chemistry , Plasmids/genetics , Rats , Sensitivity and Specificity , Time Factors , Transfection , Trypsin
13.
Biopolymers ; 32(10): 1351-63, 1992 Oct.
Article in English | MEDLINE | ID: mdl-1420963

ABSTRACT

CD spectra have been measured as a function of temperature for a number of ApA analogues with modified backbones. Oligonucleotides with these modified backbones are being used as antisense agents having potential as viral therapeutics. Results of these studies show that when a carbonyl is substituted for the phosphate to produce an uncharged backbone, the analogues that have either sugar or morpholino substitution do not stack. In contrast, when a morpholino group is substituted for the sugar and the phosphate is modified so as to be uncharged, there is strong base stacking. Stacking interactions in the phosphorus-linked morpholino analogues are at least as strong as those found in d(ApA). The stacking interactions in ApA are weak by comparison. Singular value decomposition demonstrates that the stacking is two state, and Taylor series decomposition yields a coefficient that measures base stacking interactions. The van't Hoff equation is applied to the base stacking coefficient from the Taylor series fitting to give thermodynamic parameters.


Subject(s)
Dinucleoside Phosphates/chemistry , Circular Dichroism , Molecular Structure , Nucleic Acid Conformation , Thermodynamics
14.
Antisense Res Dev ; 1(4): 319-27, 1991.
Article in English | MEDLINE | ID: mdl-1821653

ABSTRACT

Protein synthesis, which takes place within ribosomes, is essential for the survival of any living organism. Ribosomes are composed of both proteins and RNA. Specific interaction between the 3' end CCUCC sequence of prokaryotic 16S rRNA and a partially complementary sequence preceding the initiating codon of mRNA is believed to be a prerequisite for initiation of protein synthesis. Here we report the use of short (three to six nucleotides) synthetic DNA analogs complementary to this sequence to block protein synthesis in vitro and in vivo in Escherichia coli. In the DNA analogs the normal phosphodiester bond in the antisense DNA was replaced by methylcarbamate internucleoside linkages to enhance transport across plasma membranes. Of the analogs tested, those with the sequence AGG and GGA inhibit protein synthesis and colony formation by E. coli strains lacking an outer cell wall. Polyethylene glycol 1000 (PEG 1000) was attached to the 5' end of some of the test methylcarbamate DNAs to enhance solubility. Analogs of AGG and GGAG with PEG 1000 attached inhibited colony formation in normal E. coli. These analogs may be useful food additives to control bacterial spoilage and biomedically as antibiotics.


Subject(s)
Anti-Bacterial Agents/pharmacology , DNA, Antisense/pharmacology , Escherichia coli/drug effects , Oligonucleotides, Antisense/pharmacology , Protein Synthesis Inhibitors/pharmacology , RNA, Messenger/genetics , RNA, Ribosomal, 16S/genetics , 3T3 Cells , Animals , Base Sequence , Cell Division/drug effects , Escherichia coli/genetics , Escherichia coli/metabolism , Mice , Molecular Sequence Data , Protein Biosynthesis/drug effects
15.
Nucleic Acids Res ; 17(15): 6129-41, 1989 Aug 11.
Article in English | MEDLINE | ID: mdl-2771642

ABSTRACT

A novel oligonucleotide analog has been prepared from ribonucleoside derived morpholine subunits linked by carbamate groups. Oxidative cleavage of the 2',3' vicinal diol of cytidine followed by reductive amination of the resulting dialdehyde afforded the morpholine subunit. Coupling of the subunits are through carbamate moieties and the oligomers were characterized by 1H NMR and FAB MS. Evidence for interaction of the hexamer 19 with p(dG6) was found, but an atypical interaction of 19 with a RNA target was observed.


Subject(s)
Carbamates , Morpholines/chemical synthesis , Oligonucleotides/chemical synthesis , Chemical Phenomena , Chemistry , Chromatography , Hot Temperature , Hydrogen-Ion Concentration , Magnetic Resonance Spectroscopy , Mass Spectrometry , Molecular Structure , Nucleic Acid Denaturation , Solubility
16.
J Pers Soc Psychol ; 54(1): 142-8, 1988 Jan.
Article in English | MEDLINE | ID: mdl-3346803

ABSTRACT

We explored individual differences in health-seeking behavior and health status in a primary care population. Specifically, we compared high monitors (those who typically scan for threat-relevant information) with low monitors (those who typically ignore threat-relevant information), while controlling for depression. Overall, high monitors came to the physician with less severe medical problems than did low monitors. Nevertheless, high monitors reported equivalent levels of discomfort, dysfunction, and distress compared with low monitors. Furthermore, during the week following their visit, high monitors expressed less symptom improvement in both physical and psychological problems than did low monitors. Finally, high monitors demanded more tests, information, and counseling during their visit than did their low monitoring counterparts, yet desired a less active role in their own care. The theoretical and practical implications of these findings are discussed.


Subject(s)
Adaptation, Psychological , Arousal , Referral and Consultation , Sick Role , Adult , Female , Humans , Male , Patient Education as Topic , Psychological Tests
17.
Digestion ; 31(2-3): 77-81, 1985.
Article in English | MEDLINE | ID: mdl-4039693

ABSTRACT

It has previously been observed that 25% of human colorectal cancers contain specific receptors to deoxycholic acid (DCA). In the present study, the effect of intrarectal instillation of DCA on tumour number, distribution, size, and DCA receptor status was measured in rats receiving the colorectal carcinogen, azoxymethane. Rats treated with azoxymethane and intrarectal DCA developed significantly more colorectal cancers than rats receiving azoxymethane and intrarectal saline (median 11.5, range 8-17 vs. median 6.0, range 3-9 tumours/rat, respectively, p less than 0.01). This reflected a significantly higher number of tumours in the distal colon of the DCA-treated group (median 8.0, range 5-10 tumours/rat) compared to the saline-treated group (p less than 0.01). In those rats receiving DCA and azoxymethane, 5 of 12 tumours tested were found to be DCA receptor-positive, compared with only 1 of 11 in the saline and azoxymethane group. These results confirm the belief that DCA acts as a tumour promoter, and suggest a possible role for DCA receptors.


Subject(s)
Colonic Neoplasms/metabolism , Deoxycholic Acid/pharmacology , Disease Models, Animal , Rectal Neoplasms/metabolism , Animals , Azoxymethane/adverse effects , Colonic Neoplasms/analysis , Colonic Neoplasms/chemically induced , Male , Rats , Rats, Inbred Strains , Receptors, Steroid/analysis , Rectal Neoplasms/analysis , Rectal Neoplasms/chemically induced
18.
Med Oncol Tumor Pharmacother ; 1(4): 275-8, 1984.
Article in English | MEDLINE | ID: mdl-6544904

ABSTRACT

A more effective use of antibody in treating cancer appears to require derivatives with enhanced cytotoxic potential. Working with anti-idiotype antibodies directed against neoplastic lymphocytes, we have shown previously that univalent antibody derivatives with intact Fc-regions can avoid antigenic modulation while retaining the ability to recruit cytotoxic effectors such as complement. Chimeric univalent antibodies represent an extension of this approach. To prepare them Fab' gamma from antibody is linked by thioether bonds to half-cystine in normal Ig of the species to undergo immunotherapy. The derivatives FabIgG and FabFc utilize IgG and Fc gamma respectively as the effector partners of the antibody Fab' gamma. They appear superior to parent antibody in their ability to invoke complement and K-cell killing of target lymphocytes. They show promise of being minimally immunogenic and, because they present homologous Fc, should prove efficient recruiters of host effector functions.


Subject(s)
Immunization, Passive , Immunoglobulin Fab Fragments/immunology , Immunoglobulin Fc Fragments/immunology , Immunoglobulin Fragments/immunology , Immunoglobulin G/immunology , Leukemia/therapy , Lymphoma/therapy , Animals , Guinea Pigs , Rabbits
19.
Br J Surg ; 70(9): 549-51, 1983 Sep.
Article in English | MEDLINE | ID: mdl-6311314

ABSTRACT

Bile acids are thought to be involved in both the aetiology and development of colorectal cancer. In this study the existence of specific bile acid receptor proteins has been postulated. A receptor assay which involved labelling with 14C-deoxycholic acid was performed as well as autoradiography using 3H-deoxycholic acid. In an initial study resected colorectal cancer and adjacent histologically normal colorectal mucosa from 39 patients were studied, as were samples of normal gastric mucosa, cancers and benign colorectal tumours. Specific receptors to deoxycholic acid were detected in 12 (30.8 per cent) of the colorectal cancers, but in only 1 (2.6 per cent) of the samples from normal colorectal mucosa (X2 = 11.16, P less than 0.005). No deoxycholic acid receptors were detected in any other tissue studied. Autoradiographs of colorectal cancers showed binding of 3H-deoxycholic acid in receptor-positive tumour tissue. These findings might provide some explanation for the evidence linking bile acids with the disease.


Subject(s)
Colonic Neoplasms/analysis , Deoxycholic Acid/metabolism , Receptors, Cell Surface/analysis , Receptors, Steroid/analysis , Rectal Neoplasms/analysis , Autoradiography , Gastric Mucosa/analysis , Humans , Intestinal Polyps/analysis , Stomach Neoplasms/analysis
20.
Anal Biochem ; 133(1): 79-84, 1983 Aug.
Article in English | MEDLINE | ID: mdl-6356981

ABSTRACT

A method for isolating plasmids from Escherichia coli which requires less than 8 h from cell pellet to purified plasmid essentially free of protein, RNA, and chromosomal DNA is presented. By this procedure, amplified plasmid pBR322 was isolated from E. coli strain RR1. The final product had no detectable protein or RNA, and plasmid comprised approximately 99% of the total DNA. The procedure includes lysozyme treatment in hypertonic solution followed by lysis with a mild detergent in the presence of high salt and an RNase inhibitor--conditions which prevent unfolding of the bacterial nucleoid. After centrifuging out the nucleoid and cell debris, the nucleic acids are selectively precipitated with a neutral solution of sodium trichloroacetate and ethanol. RNA is degraded with RNase and the degradation products and RNase are eliminated through a second trichloroacetate/ethanol precipitation. Finally, the plasmid is resuspended and passed through a nitrocellulose filter to remove aggregates and any residual protein and single-stranded DNA--giving a plasmid preparation suitable for electrophoretic fractionation or cleavage with restriction nucleases.


Subject(s)
Bacteria/genetics , DNA, Bacterial/isolation & purification , Plasmids , Bacterial Proteins/analysis , Chromosomes, Bacterial/analysis , Escherichia coli/genetics
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