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Biomedical subjects

D L Walker

Publications and source records attributed to D L Walker.

At least 55 records · Page 3Linked to original sources
PubMed1990

Improved methods for preparation of culture media for in-vitro fertilization and gamete intra-fallopian transfer.

A new method for preparation of culture media for IVF-ET and GIFT was developed which eliminated the requirement for volumetrics and glassware. Water weight was used instead of volumetrics for preparation of media. Media prepared by the volumetric and water weight methods were compared for (i) preparation time, (ii) pH, (iii) osmolarity and (iv) the percentage of two-cell murine embryos developing to blastocysts. The time required for preparation of media was significantly less for the water weight method. Following equilibration with 5% CO2, no differences were observed between the two methods for pH, osmolarity or development of embryos to the blastocyst stage. Time for media preparation and osmolarity was less variable among preparation days for the water weight method. These results suggest that media can be prepared more efficiently and precisely with the water weight method than with the standard volumetric method used by most IVF laboratories. The former method eliminates considerable technician time which must be devoted to proper cleaning/sterilization of volumetrics and the possibility of media contamination by residual substances remaining on volumetrics following improper cleaning.

Animals↗
PubMed1990

Comparison of concurrent pregnancy rates for in-vitro fertilization--embryo transfer, pronuclear stage embryo transfer and gamete intra-fallopian transfer.

Concurrent pregnancy and implantation (sacs/embryos transferred) rates were compared for 84, 77 and 49 cases of in-vitro fertilization-embryo transfer (IVF-ET), pronuclear stage embryo transfer (PROST) and gamete intra-Fallopian transfer (GIFT), respectively. All cases reported occurred during an 18-month interval since the initiation of PROST by our programme. Leuprolide acetate was used with follicle stimulating hormone and human menopausal gonadotrophin for follicular stimulation of all but donor oocyte cases (n = 9). Clinical pregnancy (per transfer) and implantation rates were significantly higher (P less than 0.03) for PROST (52.4%, 20.2%) in comparison with IVF-ET (26.9%, 11.4%). Rates for GIFT (48.9%, 18.4%) were not significantly higher (P = 0.10, 0.14) than for IVF-ET. This was probably due to the lower number of GIFT than PROST procedures performed. The total pregnancy rate for GIFT (biochemical, ectopic and clinical combined) was significantly greater (P less than 0.05) than for IVF-ET. Pregnancy and implantation rates for PROST and GIFT were similar. These results support the use of PROST rather than IVF-ET for all cases in which the woman has one functional Fallopian tube. Furthermore, to maintain equivalent rates of pregnancy with PROST and GIFT, it is suggested that GIFT should not be used for cases of male-factor infertility without first documenting normal rates of in-vitro fertilization with PROST.

Adult↗
PubMed1990

Immunolabeling of JC virus large T antigen in neonatal hamster brain before tumor formation.

Using immunolabeling methods, the JC virus (JCV) early or regulatory protein, large T antigen, was demonstrated in frozen sections of neonatal hamster brains before tumor formation. Three days after intracerebral inoculation of 2500 hemagglutinating units of JCV, T antigen was expressed predominantly in nuclei of cells in the external granular layer and newly forming internal granular layer of the cerebellum and also in cell nuclei located in the hippocampus, periventricular areas, and the olfactory bulb. At 7 days postinoculation (p.i.), most cerebellar T antigen-containing cells had migrated to the internal granular layer, but by 15 days p.i., cells that expressed T antigen was greatly reduced in number or absent. However, by 30 days p.i., the internal granular layer of the cerebellum again contained T antigen-positive cells. In contrast to the scattered cells seen at 3 or 7 days p.i., these cells appeared in dense clusters thought to represent pretumor foci. Since JCV capsid proteins were not detected at any time, JCV may establish a latent or abortive infection in cells during their mitotic phase and these cells initially express T-antigen during migration or become immunoreactive later before tumor formation.

Animals↗
PubMed1989

Nucleotide sequence of the human polyomavirus AS virus, an antigenic variant of BK virus.

The complete DNA sequence of the human polyomavirus AS virus (ASV) is presented. Although ASV can be differentiated antigenically from the other human polyomaviruses (BK and JC viruses), it shares 94.9% homology at the nucleotide level with the Dunlop strain of BK virus. Differences found in ASV relative to BK virus include the absence of tandem repeats in its regulatory region, the deletion of 32 nucleotides in the late mRNA leader region (altering the initiation codon for the agnoprotein), the presence of a cluster of base pair substitutions within the coding region of the major capsid protein, VP1, and the absence of 4 amino acids in the carboxy-terminal region of the early protein, T antigen. The 43 nucleotides deleted in the Dunlop strain of BK virus relative to the Gardner prototype strain of BK virus are present in ASV. Possible reasons for the distinct antigenicity of the ASV capsid, given the high degree of nucleotide homology with BK virus, are discussed. To reflect the high degree of sequence homology between ASV and BK virus, we suggest ASV be renamed BKV(AS).

Antigens, Viral↗
PubMed1989

JC virus-simian virus 40 genomes containing heterologous regulatory signals and chimeric early regions: identification of regions restricting transformation by JC virus.

The papovavirus JC virus (JCV) is highly oncogenic in experimental animals but, unlike simian virus 40 (SV40), is severely restricted in its ability to transform cells in culture. We exploited the close genetic relatedness of these two viruses to delimit region(s) of the T protein which can restrict transforming activity. Novel chimeric genomes were produced by exchanging various segments of the JCV and SV40 T-protein-coding regions. These DNA constructs specified early proteins with in-frame substitutions of analogous amino acid sequences. A second set of genomes was prepared which, in addition to chimeric early proteins, contained substituted regulatory regions. The transformation efficiencies of these chimeric genomes were intermediate between those of SV40 and JCV, with the source of T protein exerting a greater effect than that of the regulatory region. The ability of certain constructs to induce efficient transformation required the presence of an SV40 regulatory region or specific sequences within the SV40 early coding region. Cloned cell lines prepared from representative transformants were characterized; the ability to form colonies in soft agarose was investigated, and the presence of viral T and cellular p53 proteins was determined. The various T proteins differed in amount, stability, and the ability to form stable complexes with p53.

Animals↗
PubMed1988

A double-label method detects both early (T-antigen) and late (capsid) proteins of JC virus in progressive multifocal leukoencephalopathy brain tissue from AIDS and non-AIDS patients.

A new double-label immunocytochemical method detects JC virus (JCV) early (T-antigen) and late (capsid) proteins simultaneously in cryostat sections of progressive multifocal leukoencephalopathy (PML) brain tissue from both acquired immunodeficiency syndrome (AIDS) and non-AIDS patients. T-antigen is detected with a monoclonal antibody (PAb 416) followed by goat anti-mouse IgG and mouse Clono-PAP, while capsid proteins are detected by a rabbit polyclonal antiserum to capsid proteins followed by biotinylated goat anti-rabbit IgG and streptavidin-alkaline phosphatase conjugate. The substrates are 3,3'-diaminobenzidine and Vector Red I, respectively. With this method some infected glial cells stain for late (capsid) antigens in the nucleus, while others show early protein (large T-antigen) immunoreactivity. The latter are likely to be astrocytes infected abortively or oligodendrocytes in the early stages of a productive JCV infection.

Acquired Immunodeficiency Syndrome↗
PubMed1988

A monoclonal antibody to SV40 large T-antigen labels a nuclear antigen in JC virus-transformed cells and in progressive multifocal leukoencephalopathy (PML) brain infected with JC virus.

Thirty monoclonal antibodies to SV40 large T-antigen were tested for reactivity on the JC virus-transformed hamster glial cell line known as HJC-15. Two of them (PAb 416 and PAb 108) detected a nuclear antigen in both SV40-transformed CCL 75.1 cells and in HJC-15 cells, but not in control cells lacking T-antigen. These same antibodies also labeled a nuclear antigen in hamster tumor tissue derived from HJC-15 cells. In addition, the monoclonal antibody PAb 416 detected a nuclear antigen in progressive multifocal leukoencephalopathy (PML) tissue infected with JC virus, but not in normal brain tissue or tissue from other neurological diseases. Staining by PAb 416 was reduced by prior incubation with hamster anti-JCV tumor serum, suggesting that the polyclonal antiserum to JCV T-antigen may compete for an epitope at or near the PAb 416 binding site.

Animals↗
PubMed1988

Concentration of glycerol required for optimal survival and in vitro fertilizing capacity of frozen sperm is dependent on cryopreservation medium.

Sperm survival and in vitro fertilizing capacity were examined following cryopreservation with three concentrations of glycerol (0, 2, and 7.5%) and four cryopreservation media. Post-thaw motility and motility index increased with increasing concentrations of glycerol. Post-thaw velocity, linearity, percentage of zona-free hamster oocytes penetrated, and penetrations per oocyte were greater following freezing with 2 or 7.5% glycerol than with 0% glycerol. A significant interaction between media and glycerol was observed for motility, velocity, linearity, motility index, and percentage of oocytes penetrated. These results suggest that the concentration of glycerol required for optimal survival and in vitro fertilizing capacity of human sperm following cryopreservation is dependent on the type of medium used for freezing.

Cell Survival↗
PubMed1988

Blood glucose and brain function: interactions with CNS cholinergic systems.

We recently found that glucose injections attenuate amnesia and hyperactivity produced by scopolamine, a muscarinic antagonist. The present study examined whether glucose would augment behavioral effects produced by a muscarinic agonist, physostigmine. In experiment I, doses were first determined for which neither glucose (10 mg/kg) nor physostigmine (0.05 mg/kg) altered scopolamine-induced hyperactivity. However, combined glucose-physostigmine injections significantly reduced scopolamine hyperactivity. Experiment II evaluated the effects of glucose on physostigmine-induced tremors. Glucose (10, 100, and 250 mg/kg) or saline injections were given 20 min before physostigmine injections (0.4 or 0.05 mg/kg). Observations of glucose effects on the severity of physostigmine-induced tremors were then obtained at 5-min intervals for 25 min after physostigmine injections. Glucose (100 mg/kg) significantly facilitated the onset of tremors when injected before either dose of physostigmine, and augmented (at 100 and 250 mg/kg) tremor severity when injected before the lower dose of physostigmine. These findings indicate that glucose can facilitate the actions of a cholinergic agonist on two behaviors, locomotor activity and tremors, adding support to the view that circulating glucose levels can modulate central cholinergic function. More generally, the results provide additional evidence that circulating glucose levels can influence brain function.

Animals↗
PubMed1988

Age distribution of progressive multifocal leukoencephalopathy.

The age distribution is given of 79 cases of progressive multifocal leukoencephalopathy confirmed as JC virus infection. The data are compared with a published age distribution of multiple sclerosis onset in Vestfold County, Norway, and with published data from Rochester, Minnesota of the age distribution of viral encephalitis. In contrast to viral encephalitis, 61% of which occurs in children under the age of 10, PML has been identified only once in this age group. PML, like MS, is a disease of adult onset, peaking in the sixth decade of life. The reason for this late onset of a CNS disease caused by a ubiquitous childhood infection is still uncertain, but it may be related to maturation of susceptible cells in the brain, as well as to declining immunity associated with chronic disease and age.

Adolescent↗
PubMed1988

Progressive multifocal leukoencephalopathy and retroviral encephalitis in acquired immunodeficiency syndrome.

Antigens of human polyomaviruses, the etiologic agents of progressive multifocal leukoencephalopathy (PML), and of human immunodeficiency virus were localized in paraffin sections from brains of six patients who died with the acquired immunodeficiency syndrome. Immunostaining revealed polyomaviral antigens in oligodendrocytes and in some astrocytes. Human immunodeficiency (retro) virus antigens were immunostained in mononuclear macrophages, glial cells, and vascular endothelial cells. Both viral types were found ultrastructurally. The lesions of PML were more destructive than is usually seen in cases without the acquired immunodeficiency syndrome. The retroviral encephalitis could have occurred before the onset of PML. However, a secondary retroviral encephalitis could have resulted if the monocytes responding to an initial polyomaviral lesion were already infected with human immunodeficiency virus before they differentiated into macrophages.

Acquired Immunodeficiency Syndrome↗
PubMed1987

Upper endoscopy for occult bleeding in renal failure.

We studied 41 patients with renal failure prospectively to assess the value of upper gastrointestinal endoscopy in detecting occult bleeding after an abrupt fall in hemoglobin (Hb) level. Thirty-four patients were on maintenance dialysis, six on plasma exchange, and one on dialysis and plasma exchange. At gastroscopy two patients (4.9%) had bleeding lesions, five patients (12%) peptic ulcers without bleeding, and 18 patients (44%) mild esophagitis, gastritis, or duodenitis. Although mucosal abnormalities in the upper gastrointestinal tract were common, clinically inapparent gastrointestinal bleeding was only rarely responsible for sudden changes of Hb in these patients.

Adolescent↗
PubMed1987

Derivation and characterization of POJ cells, transformed human fetal glial cells that retain their permissivity for JC virus.

The study of the medically important polyomavirus JC virus is limited to only a few laboratories, primarily because the permissive cell system most often used, primary human fetal glial cells, is difficult to obtain and propagate. We have introduced mutations at the origin of DNA replication of JC virus and transformed glial cells with the replication-defective genomes. Although normal glial cell cultures rapidly lose their permissivity for the virus after subculture, the transformed cells (designated POJ) had a greatly expanded life span and remained permissive for JC virus even after 30 passages in vitro. POJ cells constitutively express a functional T protein that complements the replication defect of lethal early-region mutations in JC virus. We expect that these cells will greatly facilitate the study of this human virus.

Antigens, Polyomavirus Transforming↗
PubMed1986

Immunocytochemical search for JC papovavirus large T-antigen in multiple sclerosis brain tissue.

The large T-antigens of papovaviruses JC (JCV) and BK share a C-terminal subsequence with myelin basic protein (MBP). Since this sequence functions as a phosphate acceptor site in MBP, expression of a competing T-antigen sequence in oligodendroglia might adversely affect their ability to post-translationally process MBP and thus to maintain myelin. We have used techniques which demonstrate JCV T-antigen in small oligodendroglial cells from progressive multifocal leukoencephalopathy tissue to search for a possible latent JCV infection expressing T-antigen in nine cases of multiple sclerosis (MS) and three normal brains. No cells expressing T-antigen were detected in plaque or periplaque regions of the MS brains or in control CNS tissue.

Adult↗
PubMed1986

JC papovavirus large tumor (T)-antigen expression in brain tissue of acquired immune deficiency syndrome (AIDS) and non-AIDS patients with progressive multifocal leukoencephalopathy.

Progressive multifocal leukoencephalopathy (PML) is a JC papovavirus infection of the central nervous system in immunocompromised patients. It is well established that demyelination in PML is caused by JC virus infection of oligodendroglia, but whether the nonstructural regulatory protein, large tumor (T) antigen, is detectable in infected human tissue was not known. Using a modification of the peroxidase-antiperoxidase technique, we found T antigen expressed in the nuclei of cells in virus-infected sites in five cases of PML studied, including two with acquired immune deficiency syndrome (AIDS). PML occurs in AIDS at a much higher frequency than in other immunosuppressive disorders, and PML in AIDS may represent a more severe form of JC virus infection of the central nervous system.

Acquired Immunodeficiency Syndrome↗