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Biomedical subjects

L King

Publications and source records attributed to L King.

At least 19 recordsLinked to original sources

Behavioral response to diazepam in a residential laboratory.

Two groups of three healthy adult male volunteers without histories of sedative or other drug abuse participated in 15-day residential studies. Each day consisted of a private work period (10 AM to 4:30 PM), during which subjects participated in traditional laboratory performance tasks, and a social period (5 to 11:30 PM), during which subjects had access to recreational activities available under social or private conditions. Tobacco cigarettes and food were available throughout each day (9 AM to 12 PM). Diazepam (5 or 10 mg/70 kg) or placebo was administered orally twice daily in alternating three-consecutive-day intervals. Dosing order varied between groups. Diazepam had no effect on the total amount of time subjects spent in social conditions; however, the low dose increased verbal interaction, while the high dose decreased verbal interaction. Both doses disrupted performance on a second-order repeated-acquisition task but produced no effects on the other performance measures. Five of six subjects increased caloric intake following at least one dose, with the largest increases observed in subjects with the lowest baseline intake. Increases in subject reports of dose "Potency" and "Sedated" were also observed following the high dose. Diazepam doses routinely used in clinical settings influenced a variety of behaviors that are observed in the natural ecology, but not performance on accepted laboratory tasks.

Adult

Restricted use of fetal VH3 immunoglobulin genes by unselected B cells in the adult. Predominance of 56p1-like VH genes in common variable immunodeficiency.

The large VH3 family of human immunoglobulin genes is commonly used throughout B cell ontogeny. However, B cells of the fetus and certain autoantibody-producing clones are restricted to a recurrent subset of VH3 genes, and VH3 B cells are deficient in certain immunodeficiency diseases. In this study, we have sequenced a set of rearranged VH3 genes generated by genomic polymerase chain reaction (PCR) from normal adults and those with common variable immunodeficiency (CVI). In both groups, all cones were readily identifiable with the fetal VH3 subset, and were further distinguished by limited DH motifs and exclusive use of JH4. In CVI, the residual population of VH3 B cells were notable for predominant use of 56p1-like VH genes. All clones displayed sequence divergence (including somatic mutation) with evidence of strong selection against complementarity-determining region (CDR) coding change. A survey of other V gene families indicates that human V gene diversity may be restricted in general by germline mechanisms. These findings suggest that the expressed antibody repertoire in the human adult may be much smaller than anticipated, and selected by processes in part distinct from the paradigm of maximal antigen-binding diversity.

Antibody Diversity

Acute methanol poisoning: a case study.

Acute methanol poisoning produces severe anion gap metabolic acidosis caused by the toxic accumulation of metabolites, primarily formic acid. Formic acid produces serious neurologic sequelae. Therefore, an understanding of the mechanism of toxicity, treatment, and clinical course is essential in preventing permanent neurologic dysfunction. Prompt recognition and treatment are the keys to successful patient outcomes. This article presents a case report of a patient with severe methanol poisoning with clinical course, treatment, and outcome.

Adult

Glycerol reorientation during the conversion of phosphatidylglycerol to bis(monoacylglycerol)phosphate in macrophage-like RAW 264.7 cells.

Bis(monoacylglycero)phosphate (BMP) has the unique stereoconfiguration of 3-acyl-sn-glycero-1-phosphoryl-1'-sn-[3'-acylglycerol] (Brotherus, J., Renkonen, O., Herrmann, J., and Fischer, W. (1974) Chem. Phys. Lipids 13, 178-182) which differs from other known mammalian phospholipids that have the sn-glycero-3-phosphoryl configuration. This stereochemistry may contribute to its physiologic function. Here we describe studies using the macrophage-like cell line RAW 264.7 designed to determined how this unique stereoconfiguration occurs. These studies show that the stereoconfiguration of BMP produced from exogenous phosphatidylglycerol (PG) by RAW 264.7 cells has the expected stereoconfiguration of 3-acyl-sn-glycero-1-phosphoryl-1'-sn-[3'-acylglycerol]. Experiments using diacyl-sn-[2-3H]glycero-3-phosphoryl-sn-1'-[2-3H]glycerol demonstrate that this unique stereoconfiguration is not produced due to an oxidation/reduction mechanism involving the sn-2-glycerol carbon. When dioleoyl-sn-[1-14C]glycero-3-phosphoryl-rac-glycerol was converted to 14C-labeled BMP, the 14C label was found esterified to the phosphate moiety. These results suggest that a stereospecific enzyme is capable of reorienting the radiolabeled glycerol backbone of this PG substrate, effectively changing the stereochemistry of the lipid. We also show that this enzyme is stereoselective with regard to the base glycerol moiety of the substrate PG used. Finally, we propose a new pathway for the synthesis of BMP from PG.

Animals

Persistent B19 parvovirus infection in patients infected with human immunodeficiency virus type 1 (HIV-1): a treatable cause of anemia in AIDS.

OBJECTIVE: To determine the role of B19 parvovirus in red cell aplasia of patients infected with human immunodeficiency virus type 1 (HIV-1). DESIGN: Uncontrolled clinical trial, with assay of serum, peripheral blood cells, and bone marrow for virus using DNA hybridization and immunocytochemistry techniques; these assays were then correlated with clinical findings, results of immunoassays for antivirus antibodies, and with immunoglobulin (Ig) therapy. SETTING: Government medical referral center, and university and private hospitals. PATIENTS: Seven patients with pure red cell aplasia and serologic evidence of infection with HIV-1. MEASUREMENTS AND MAIN RESULTS: All patients had giant pronormoblasts in the bone marrow (present in transient aplastic crisis caused by acute B19 parvovirus infection). High concentrations of B19 parvovirus were demonstrated in sera, in several cases in samples separated by weeks or months. Viral DNA and capsid protein were present in the bone marrow of three patients studied, and active viral replication was detected by southern analysis. There was no antivirus IgG in capture immunoassay and no or very low levels of antivirus IgM. The patients did not have symptoms of fifth disease, the illness caused by this virus in immunologically normal persons. Six patients were treated with a regimen of intravenous commercial immunoglobulin. In all cases, this therapy resulted in rapid reduction in serum virus concentrations and full recovery of erythropoiesis. Relapses in two cases were predicted by DNA hybridization studies, and these cases were successfully retreated. CONCLUSIONS: The B19 parvovirus is a remediable cause of severe chronic anemia in HIV-infected patients. Recognition of and therapy for parvovirus in this population will avoid erythrocyte transfusion and should prevent transmission of the virus to other persons, including immunosuppressed persons and women of child-bearing age.

Acquired Immunodeficiency Syndrome

Metabolism of phosphatidylglycerol and bis(monoacylglycero)-phosphate in macrophage subcellular fractions.

Bis(monoacylglycero)phosphate (BMP) is synthesized from exogenous phosphatidylglycerol (PG) by macrophages (Cochran, F. R., Roddick, V. L., Connor, J. R., Thornburg, J. T., and Waite, M. (1987) J. Immunol. 138, 1877-1883). Previous work from our laboratory showed that arachidonic acid in BMP was released by the macrophages upon challenge of the cells with PMA (Cochran, F. R., Connor, J. R., Roddick, V. L., and Waite, M. (1985) Biochem. Biophys. Res. Commun. 130, 800-806). Here we extend those studies using a model cultured cell line of macrophages, RAW 264.7. When PG labeled with 32P- and [3H]glycerol in both moieties was added to the culture medium, 32P/[3H]BMP was synthesized in a time-dependent manner. Fractionation of cell homogenates on a discontinuous sucrose gradient in which the light membranes were floated from dense sucrose showed an enrichment of [3H]BMP in light membrane fractions. The precursor [3H]PG was also found in the light fractions but, relative to the [3H]BMP, was more abundant in the denser membrane fractions. The appearance of [3H]PG and [3H]BMP in the light membrane fraction was time-dependent which suggested that the initial uptake and metabolism of [3H]PG was into the denser membranes. Incubation of the light membranes under conditions that are optimal for the lysosomal phospholipase A1 led to significant metabolism of [3H]PG. Both degradation of [3H]PG to water-soluble compounds and its conversion to acylphosphatidylglycerol occurred while no lyso-PG was detected. On the other hand, little BMP was found to be degraded. From these studies we postulate that in lysosomes acylphosphatidylglycerol is a precursor of BMP and that the previously reported turnover of arachidonic acid by BMP may occur via transacylation rather than hydrolysis.

Animals

Antitumor activity of L6-ricin immunotoxin against the H2981-T3 lung adenocarcinoma cell line in vitro and in vivo.

The monoclonal antibody L6 recognizes a determinant that is expressed on lung, breast, colon, and ovarian carcinomas and is present only at trace levels in normal tissues. L6 was covalently linked to intact ricin by a thioether bond to produce an immunotoxin (IT). Gel analysis revealed that this IT was heterogeneous, but mostly one monoclonal antibody molecule linked to one ricin molecule. The L6-ricin IT selectively bound and was selectively toxic to L6-positive H2981-T3 adenocarcinoma cells in protein synthesis inhibition assays in which lactose was added to block the native ricin binding site. Clonogenic studies showed that 1 microgram/ml L6-ricin could inhibit about 99.99% of H2981-T3 growth in a limiting dilution assay, even in the presence of a 20-fold excess of human bone marrow cells. Treatment of bone marrow cells with the same dose of L6-ricin resulted in the growth of ample numbers of bone marrow progenitor cells (colony-forming units-mixed, colony-forming units granulocyte/macrophage, and blast-forming units erythroid) after 14 days. We also evaluated the antitumor effect of L6-ricin administered intratumorally with lactose against established H2981-T3 tumors in a nude mouse model. Thirty % of the tumor-bearing animals responded completely to single-dose treatment, while 60% gave partial responses. The in vivo effects were not absolutely specific, since irrelevant anti-CD5 IT also induced tumor regression in this model (10% responded completely, while 30% gave partial responses). However, irrelevant IT gave higher systemic toxicity (50% mortality) than L6-ricin (23% mortality). The nonspecific activity of IT was possibly due to Fc binding, which was demonstrated in vitro, or due to ricin B-chain binding. Ricin alone was too toxic for sustained tumor protection. Unconjugated L6 had no antitumor effect. The data suggest that L6-ricin may be useful for in vitro purging of autologous bone marrow from patients with solid tumors and marrow involvement and for in vivo regional therapy of L6-positive carcinomas.

Adenocarcinoma

15-Lipoxygenase products of arachidonate play a role in proliferation of transformed erythroid cells.

The role of specific products of the lipoxygenase pathway of arachidonic acid metabolism has been investigated in the Friend erythroleukemia cell line, a model system for erythroid cell differentiation. When triggered with agents such as hexamethylene-bis-acetamide, these cells mature as normal erythroid cells. 15-Hydroxyeicosatetraenoic acid (15-HETE) was identified by reverse-phase high-performance liquid chromatography and by radioimmunoassay as the principal lipoxygenase metabolite produced by Friend cells. Its production was significantly lower (903 +/- 73 pg/ml) in stationary-phase cells compared with logarithmic-phase cells (1,496 +/- 24 pg/ml). In addition, inhibitors of both the cyclooxygenase and lipoxygenase pathways (phenidone, BW 755C, caffeic acid, nordihydroguaiaretic acid and BW 4AC) significantly blocked DNA synthesis (P less than 0.05), whereas neither specific inhibitor of the cyclooxygenase pathway (aspirin or sodium meclofenate) blocked DNA synthesis. The addition of 15-hydroperoxyeicosatetraenoic acid as well as 15-HETE to Friend cells produced an increase in DNA synthesis as assessed by [3H]thymidine incorporation in differentiating cells but not in proliferating cells. These data support a role for 15-lipoxygenase products of arachidonic acid metabolism in maintaining DNA synthesis.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid

The association of blood pressure levels and change in renal function in hypertensive and nonhypertensive subjects.

We compared the changes in serum creatinine levels over time after a mean follow-up of 9.8 years in essential hypertensive (EH, n = 56) and control (n = 59) male veteran subjects. All subjects had normal serum creatinine levels (62 to 124 mumol/L) and "normal" urinalysis results on entry into the study. Subjects with comorbid renal diagnoses and diabetes mellitus were eliminated from the analysis. Although not statistically significant, the rate of change in the serum creatinine concentration over time was greater in the EH cohort compared with the control cohort (1.08 +/- 4.8 vs 0.027 +/- 3.5 mumol/L per year). The difference was especially marked in black EH subjects vs black control subjects (1.60 +/- 6.2 mumol/L per year vs -0.21 +/- 3.3 mumol/L per year). When age, race, body mass index, and a diagnosis of EH were entered into a logistic regression analysis, EH subjects had a statistically significantly greater rate of decline in renal function than did control subjects (1.5 +/- 8.3 mumol/L per year). When mean time-averaged systolic blood pressure for each subject was also included in the logistic regression analysis, only systolic time-averaged blood pressure was statistically significant (0.063 +/- 0.029 mumol/L per year). We conclude that in the absence of clinically detected parenchymal renal disease, EH subjects have a greater rate of decline in renal function than do nonhypertensive subjects. Time-averaged blood pressure is predictive of the change in serum creatinine concentration not only in EH subjects but also in nonhypertensive subjects. Thus, preservation of renal function may require a blood pressure lower than the currently accepted normotensive range.

Blood Pressure

Electroejaculatory stimulation of a quadriplegic man resulting in pregnancy.

Reproductive rehabilitation of men after spinal cord injury has been made possible by the application of assisted ejaculatory techniques such as electroejaculation. Although this technique may predictably overcome the barrier of semen recovery, the problem of seminal dysfunction still remains. This report describes one of the few pregnancies established by a quadriplegic man using electroejaculatory stimulation for semen acquisition and standard in vitro semen preparation methods for intrauterine insemination.

Adult

Thermal unfolding of myosin rod and light meromyosin: circular dichroism and tryptophan fluorescence studies.

Rabbit skeletal myosin rod, which is the coiled-coil alpha-helical portion of myosin, contains two tryptophan residues located in the light meromyosin (LMM) portion whose fluorescence contributes 27% to the fluorescence of the entire myosin molecule. The temperature dependence of several fluorescence parameters (quantum yield, spectral position, polarization) of the rod and its LMM portion was compared to the thermal unfolding of the helix measured with circular dichroism. Rod unfolds with three major helix unfolding transitions: at 43, 47, and 53 degrees C, with the 43 and 53 degrees C transitions mainly located in the LMM region and the 47 degrees C transition mainly located in the subfragment 2 region. The fluorescence study showed that the 43 degrees C transition does not involve the tryptophan-containing region and that the 47 degrees C transition produces an intermediate with different fluorescence properties from both the completely helical and fully unfolded states. That is, although the fluorescence of the 47 degrees C intermediate is markedly quenched, the tryptophyl residues do not become appreciably exposed to solvent until the 53 degrees C transition. It is suggested that although the intermediate that is formed in the 47 degrees C transition contains an extensive region which is devoid of alpha-helix, the unfolded region is not appreciably solvated or flexible. It appears to have the properties of a collapsed nonhelical state rather than a classical random coil.

Animals

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Fraud

Impact of percutaneous renal stone removal on renal function: assessment by urinary lysozyme activity.

Lysozyme in the urine in concentrations greater than 3 micrograms per milligram of creatinine reflects renal tubular disease or dysfunction in patients without bowel disease or leukemia. We therefore used urine lysozyme assays to assess renal response to percutaneous nephrostomy and stone removal in 42 patients. Eight patients had striking increases (4.2-21.1 [mean 7.58] micrograms/mg creatinine) immediately after nephrostomy puncture in urine obtained directly from the punctured kidney. Lysozyme declined sharply thereafter and was within normal limits in all cases by postoperative day 3. This increase appeared to result from bleeding into the urine from the tract. Five other patients had lysozymuria on admission, only 1 of whom had a sharp increase after nephrostomy puncture. In the remaining patients, the lysozyme levels remained within normal limits throughout the hospital course. These data are further evidence of the absence of significant deleterious effects of nephrostomy puncture on the kidney.

Acute Kidney Injury

Unique V gene usage by B-Ly1 cell lines, and a discordance between isotype switch commitment and variable region hypermutation.

We have previously described a series of Ly-1+ B cell lines obtained as spontaneous outgrowth cultures from mouse spleen cells (B-Ly1 cells). In this study, we report the variable region sequences utilized by independent cell lines derived from three mouse strains. Remarkably, V region utilization and junctional diversification were essentially identical in all three cell lines. These included: lambda 1 light chain; JH1; DSP2-7,8; and, a novel VH gene segement. The VH was highly homologous to CP4, a rare VH family previously found in a group of Ly-1 B cell-derived autoantibodies to bromelain-treated mouse red blood cells. The selective expression of this unusual lg species implies the action of distinctive molecular or immunophysiological processes in the outgrowth of B-Ly1 cell lines. Clarification of these factors may be relevant to understanding the peculiar biology of Ly-1 B cells in vivo. We have also evaluated switch-committed B-Ly1 clones to determine whether this phenotype is accompanied by variable region somatic mutation. However, no evidence for somatic mutation was observed in these induced clones. This may indicate that these two processes are independently regulated, despite their common concordance in vivo.

Amino Acid Sequence