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Biomedical subjects

W R Fisher

Publications and source records attributed to W R Fisher.

At least 37 records · Page 2Linked to original sources
PubMed1985

Immune complex hyperlipidemia induced by an apolipoprotein-reactive immunoglobulin A paraprotein from a patient with multiple myeloma. Characterization of this immunoglobulin.

An antibodylike paraprotein has been isolated from a patient with multiple myeloma and autoimmune hyperlipoproteinemia. The paraprotein bound to apolipoprotein B (apo B)-containing lipoproteins that formed macromolecular aggregates, and globules thought to be aggregated complexes of lipoproteins and reactive immunoglobulins were observed circulating within the retinal blood vessels of this patient. This binding specificity permitted purification of the paraprotein from both the agglutinated immune complexes and from the plasma. The protein is an IgA, kappa-immunoglobulin which exists primarily in a polymeric state. Capillary immunoprecipitation demonstrated reactivity with very low density lipoproteins (VLDL) and low density proteins (LDL), but not with high density lipoproteins (HDL). Delipidated apo B and apo E, but not apo A or apo C, formed precipitates with this immunoglobulin. In using a radioimmunoassay format, the affinity of the immunoglobulin was greatest for VLDL and decreases sequentially for intermediate density lipoproteins and LDL. No binding occurred with a dispersion of LDL lipids or with HDL. Deglycosylation did not change the binding to LDL. The apolipoproteins B and E bound with similar affinity, but no binding occurred with apo A-I or apo A-II. Weak binding appeared to occur with apo C. This paraprotein immunoprecipitated apo B-containing lipoproteins from all classes of vertebrates tested. Displacement of the lipids of LDL by Triton X-100 resulted in the formation of an apo B-Triton complex which, however, did not bind to the immunoglobulin; apparently the binding site on apo B was lost. Upon enzymatic digestion with the IgA-specific protease from Streptococcus sanguis the immunoglobulin was cleaved into Fc and Fab fragments, and the binding of LDL occurred only with the latter, consistent with the behavior of an immunoglobulin. The immunoreactivity of this paraprotein with apo B and apo E raises the interesting possibility that it may be binding to a site on these apolipoproteins which is reactive with the apo B, E receptor of the plasma membrane, a site which is conserved throughout the vertebrate phylum.

Aged↗
PubMed1984

A method for the measurement of the sedimentation coefficient and molecular weight of microgram quantities of proteins in 6 M guanidine hydrochloride.

A technique has been perfected for measuring the sedimentation coefficient of microgram quantities of a reduced protein in 6 M guanidine hydrochloride. The protein is sedimented through a gradient of 5-8 M guanidine-HCl in the presence of dithiothreitol in a SW 50.1 swinging-bucket rotor. Run conditions are calibrated by a simultaneous measurement using a single reference protein. Thus, the need for running a calibration curve involving several standard proteins simultaneously with a sample is eliminated. Because of the trace quantity of protein used, the technique yields an estimate of the sedimentation coefficient at zero concentration (s0) directly without extrapolation. Since s0 is a function of the molecular weight of a reduced protein in this solvent, the method also allows an estimate of the subunit molecular weight of the protein. The results of the application of the method to known proteins are reported.

Centrifugation, Density Gradient↗
PubMed1984

Structural studies on apolipoprotein B: controllable heterogeneity of the complex formed with the surfactant, Triton X-100.

Apolipoprotein B complexed with Triton X-100 (T-ApoB) has been isolated from human low density lipoprotein (LDL). Preparations are heterogeneous when analyzed by sedimentation velocity, with a major 12 S species and minor 17 S species present. The 12 S T-ApoB complex possesses a molecular weight of 880,000 containing 400,000 daltons of protein. Hydrodynamic measurements on this complex are consistent with a prolate ellipsoid model having an axial ratio of 13:1 and 0.22 g/g of bound water. Heterogeneity results from the irreversible aggregation of 12 S complexes into discrete 17 S and faster sedimenting components. A significant finding is that three determinants of this T-apoB heterogeneity could be elucidated and controlled. First, the initial state of aggregation is mainly influenced by the technique by which Triton and LDL are mixed. Second, once isolated, T-ApoB complexes slowly but spontaneously undergo further aggregation at 4 degrees C; the rate and extent of aggregation is enhanced remarkably with increasing temperature. Finally, reagents that unfold and expose protein structure (perchlorate, thiocyanate, and reducing reagents) lead to increased aggregation. The ability to control heterogeneity carries important implications for other studies concerning interactions of apoB with surfactants and lipids.

Apolipoproteins B↗
PubMed1983

A comparative study of enzymatic digestion profiles of apolipoprotein B from four human subjects.

A methodological approach for comparative structural study of apolipoprotein B has been developed. Low-density lipoproteins from four human subjects were digested in three separate enzyme systems, utilizing trypsin, chymotrypsin and Staphylococcus aureus protease V8, each in the presence of 1% sodium dodecyl sulfate. The peptides were separated by electrophoresis on polyacrylamide gels in SDS; the stained gels were scanned spectrophotometrically to produce characteristic profiles. Comparison of the profiles revealed good reproducibility and a high degree of similarity among the different subjects. Of the four subjects studied, one subject had one apparent difference in the tryptic digest profile and also in the S. aureus protease V8 digest profile. The structural significance of these variations can be evaluated only after a larger number of subjects, including those presented here, have been examined; this study is now in preparation.

Apolipoproteins↗
PubMed1983

Primary empty sella, hyperprolactinemia, and isolated ACTH deficiency after postpartum hemorrhage.

In a 20-year-old woman, a complicated full-term delivery was followed by a 14-month history of galactorrhea, amenorrhea, and symptoms of hypocortisolism. Evaluation revealed the presence of an empty sella, hyperprolactinemia, and an isolated pituitary deficiency of ACTH, resulting in secondary adrenal insufficiency. The defect in ACTH secretion was apparently due to intrinsic pituitary rather than hypothalamic disease, because administration of lysine vasopressin did not stimulate ACTH release. An empty sella with hyperprolactinemia has been described before. However, to the authors' knowledge, isolated ACTH deficiency as a complication of postpartum hypopituitarism (atypical Sheehan's syndrome) in association with an empty sella and hyperprolactinemia has not previously been reported.

Adrenocorticotropic Hormone↗
PubMed1981

Medical student research: a program of self-education.

Student participation in research has traditionally been part of the academic program in American medical schools. A survey of student research in 115 U.S. medical schools showed that 74 percent had M.D./Ph.D. programs but that only about 0.6 percent of the students participated in them. Eighty percent had research programs not directed toward a Ph.D.; and although these involved only 6 percent of the students, the enthusiasm of the students for their research experience was very high. The University of Florida has long fostered medical student research. A survey of 1969--1978 participants in the Florida programs yielded a 74 percent endorsement of research as an important, positive educational experience, and nearly all agreed that a research option should be part of the medical school curriculum. Since research provides the students with an intellectually challenging, self-learning experience that they endorse enthusiastically and since there is small student participation, it seems timely to reassess the role of student research in the medical school curriculum.

Education, Medical↗
PubMed1980

The metabolism of apolipoprotein B in subjects with hypertriglyceridemia and polydisperse LDL.

This research concerns the metabolism of apolipoprotein B (apoB) in hypertriglyceridemic subjects with polydisperse or heterogeneous LDL. Five subjects maintained under constant dietary control had blood samples fractionated into very low density lipoprotein (VLDL), Sf 20, Sf 10, and Sf 4 LDL, and plasma free leucine, and in three subjects VLDL was further fractionated by size. Apo B was isolated, and the masses of the plasma apo B pools were measured for these lipoproteins. Following injection of [3H]leucine as a metabolic tracer the specific activity of apo B in these lipoproteins and of plasma leucine were measured over 7 or 14 days. The kinetic data were examined using multicompartmental analysis and interpreted in terms of our previous model of apo B metabolism (1975. Federation Proc. 24: 2263.). Newly synthesized apo B, secreted as large VLDL, is metabolized by a delipidation chain yielding intermediate density lipoprotein (IDL), consisting of small VLDL and Sf 20 LDL, and eventually forms small, Sf4 LDL. LDL is metabolized in a steplike process from Sf 20 to Sf 10 and Sf 4 LDL. A second major biosynthetic input in apo B enters directly into IDL and 1/4 to 2/3 of newly synthesized apo B enters plasma by this route. Total apo B synthesis in these subjects is 5- to 10-fold greater than reported for normals. The rate of transport of VLDL apo B and IDL is slower than normal with a residence time which is increased about twofold; however, the VLDL apo B pool is enlarged 5- to 10-fold, and thus the quantity of apo B entering and leaving this pool per hour is much greater than in the normal. Two major pathways for apo B catabolism occur. Between 1/3 and 2/3 of apo B is metabolized through LDL, disappearing from plasma as Sf 4 LDL. The remainder of apo B disappears from plasma IDL directly. The four major findings in this kinetic study of apo B metabolism in hypertriglyceridemic subjects with polydisperse LDL are: 1) The marking increase in apo B synthesis; 2) the biosynthetic input of much of this apo B directly into IDL; 3) the large catabolic pathway of apo B which leaves IDL, and 4) the stepwise metabolism of LDL by which Sf 20, Sf 10 and Sf 4 LDL are generated.

Adult↗
PubMed1979

The absence of a role for the carbohydrate moiety in the binding of apolipoprotein B to the low density lipoprotein receptor.

The binding of low density lipoprotein (LDL) to fibroblasts occurs through apolipoprotein B, a glycoprotein. The role of the carbohydrate in binding was assessed in two ways: (1) LDL, freed of sialic acid and most of the glucosamine and hexoses by digestion with a mixture of glycosidases, bound to fibroblasts as does native LDL. (2) The glycopeptides liberated from apoprotein B by trypsin and pronase failed to inhibit LDL binding to fibroblasts. Apparently the carbohydrate moiety of LDL does not interact with the plasma membrane receptor.

Apolipoproteins↗
PubMed1979

Apolipoprotein B: its role in the control of fibroblast cholesterol biosynthesis and in the regulation of its own binding to cellular receptors.

Apolipoprotein B transports cholesterol in plasma as low density lipoprotein (LDL) and targets its delivery to cells by binding to a specific plasma membrane receptor. The cellular consequences of apoB binding to its receptor were investigated to determine whether it suppresses cholesterol biosynthesis and reduces the number of cellular receptors for the apoprotein. Upon preincubation of fibroblasts with lipoprotein-deficient medium alone or supplemented with either LDL or apoB complexed to BSA (apoB-BSA), LDL suppressed cholesterol biosynthesis, but apoB enhanced it. Similarly, fibroblasts preincubated in medium supplemented with LDL bound decreased amounts of either (125)I-labeled LDL or (125)I-labeled apoB-BSA to their receptors, while preincubation with apoB-BSA increased the binding relative to the controls. These latter results occurred in association with a decrease in cellular cholesterol content, indicating that apoB in the medium bound cholesterol and removed it from the cells, thus stimulating both cholesterol synthesis and cellular binding of apoB. Accordingly, fibroblast cholesterol synthesis and the number of functional LDL receptors are not suppressed by the binding of the apoprotein to the receptor, and the known role of apoB remains that of transporting cholesterol in plasma and delivering it to the cell. A possible physiologic role for apoB in depleting cells of cholesterol is presently unknown since apoB is not known to exist free in plasma; however, these findings demonstrate such a functional capability for this apoprotein.-Shireman, R. B., and W. R. Fisher. Apolipoprotein B: its role in the control of fibroblast cholesterol biosynthesis and in the regulation of its own binding to cellular receptors.

Apolipoproteins↗
PubMed1978

Proteolytic digestion in the elucidation of the structure of low density lipoprotein.

The apoprotein (apoB) of low density lipoprotein (LDL) is reported to be a large polypeptide, and it is proposed that there are two similar-sized subunit proteins in LDL (Smith, Dawson, and Tanford. 1972. J. Biol. Chem. 247: 3376-3381.). When apoB is isolated under conditions that minimize artifactual proteolysis, only a single, large molecular weight protein appears on polyacrylamide gel electrophoresis in SDS. To investigate the organization of apoB as it exists within native LDL, limited proteolysis with trypsin has been used as a structural probe. Tryptic digestion for 1 hr at pH 7.6 with enzyme-to-protein ratios of 1:100 and 1:5 results in the liberation of approximately 10% and 30% of apoB as smaller, water-soluble peptides. These peptides may be separated from the partially digested but still intact tryptic core (T-core) of the lipoprotein by chromatography on Sephadex G-75. Repeatedly, the 1:5 T-core of native LDL is found to contain a family of polypeptides of 14,000-100,000 molecular weight. Although they have lost significant quantities of apoprotein, these T-cores sustain an appearance of homogeneity, as studied by analytical ultracentrifugation. Their measured molecular weights do not differ appreciably from those of the native LDL, and the carbohydrate content of the 1:5 tryptic T-core of LDL is similar to that of the native LDL. In normolipemic individuals, LDL generally exists in a monodisperse state, but, in different individuals, monodisperse LDL may range in molecular weight from 2.4 to 3.9 x 10(6). Limited tryptic digestions were used to probe the organization of apoB in these different molecular weight LDL. As assayed by SDS-acrylamide gel electrophoresis of the larger polypeptides and fingerprinting of the smaller released peptides, those regions of LDL exposed to trypsin digestion are identical in monodisperse LDL of 2.5 and 3.4 x 10(6) molecular weight. Thus, the different quantities of lipid bound in these various LDL must interact with apoB so that the same regions of the apoprotein are exposed to the action of trypsin in these different molecular weight lipoproteins.

Amino Acids↗
PubMed1977

Solubilization of apolipoprotein B and its specific binding by the cellular receptor for low density lipoprotein.

Low density lipoprotein (LDL) and very low density lipoprotein (VLDL) bind specifically to a receptor on fibroblasts, and it has been postulated that the apoprotein of LDL (apo B) confers the specificity of cellular binding. This hypothesis has been tested in the present study with a watersoluble apo B-bovine serum albumin complex. The binding of (125)I-labeled apo B to cultured fibroblasts was temperature-dependent. Specific binding ranged between 183 and 859 ng/mg of cell protein at a concentration of 5 mug/ml; at 37 degrees , 750-2199 ng/mg was bound and internalized. The binding of apo B greatly exceeded the amount of (125)I-labeled LDL bound at 4 degrees and 37 degrees in the same experiment. Fibroblasts from a subject homozygous for hyper-beta-lipoproteinemia showed minimal binding of (125)I-labeled LDL, consistent with the absence of the cellular LDL receptor. Such cells also had depressed binding of (125)I-labeled apo B. Lymphocytes grown in lipoprotein-deficient medium demonstrated specific binding of LDL; however, freshly isolated lymphocytes did not show such binding. The binding of (125)I-labeled apo B to lymphocytes paralleled the binding of (125)I-labeled LDL. Unlabeled LDL and apo B-albumin complex both competitively inhibited the binding of (125)I-labeled apo B and (125)I-labeled LDL to fibroblasts. When labeled LDL was incubated with fibroblasts for 6 hr at 37 degrees , it underwent cellular internalization and degradation, as measured by the release of (125)I-labeled fragments into the medium. This degradation was inhibited by unlabeled apo B. Conversely, (125)I-labeled apo B also was internalized and degraded by fibroblasts, and this process was inhibited by LDL. These findings demonstrate that apo B binds specifically to the LDL receptor and that the cellular binding of LDL is determined by this apoprotein.

Apolipoproteins↗
PubMed1976

Components of school anxiety: developmental trends and sex differences.

Dunn's School Anxiety Questionnaire was administered to 104 third through eighth graders (Study I) and 122 eighth graders (Study II). Significant decrements in anxiety with increasing age were found for worry (but not emotionality), report card anxiety, and failure anxiety. In Study I girls scored significantly higher than boys on emotionality (but not worry) and test anxiety. Sex differences were greater in Study II, girls scoring consistently higher. In Study II a worry-emotionality questionnaire administered immediately preceding a final examination correlated highly with School Anxiety Questionnaire scores. Results are taken as generally supportive of the multidimensional approach to the study of school anxiety.

Achievement↗
PubMed1976

The common hyperlipoproteinemias: an understanding of disease mechanisms and their control.

Our review focuses on low density lipoprotein (LD lipoprotein) and very low density lipoprotein (VLD lipoprotein) in their roles as transporters of cholesterol and triglyceride and as factors contributing to premature arteriovascular disease. We describe the clinical manifestations of the common, primary hyperlipoproteinemias--that is, hyper-beta-lipoproteinemia, combined hyperlipoproteinemia, hyper-pre-beta-lipoproteinemia, and sporadic hyperlipoproteinemia--and discuss the variations in lipoprotein structure and metabolism that occur in these diseases. Based on an understanding of the physiologic control of lipoprotein metabolism, it is possible for the physician to alter the concentrations of LD lipoprotein and VLD lipoprotein by selecting a course of therapy appropriate to the specific disease. We describe the effects of obesity, diet, insulin, ethanol, estrogens, and the drugs clofibrate, nicotinic acid, and cholestyramine.

Cholestyramine Resin↗
PubMed1975

A genetic determinant of the phenotypic variance of the molecular weight of low density lipoprotein.

The molecular weight of monodisperse human plasma low densitylipoprotein has been measured in 69 individuals and found to vary over the range of 2.4 to 3.9 X 10-6. By contrast, the molecular weight of low density lipoprotein measured on two separate occasions for specific individuals shows a mean difference of 0.07 X 10-6 and a standard deviation of 0.08 X 10-6; hence low density lipoprotein differing in molecular weight by greater than 0.2 X 10-6 may be considered different macomolecules. The distribution of the molecular weight of low density lipoprotein does not differ as a function of age or sex. Hyperlipemic subjects having monodisperse low density lipoprotein show similar molecular weight distribution to normal subjects, as do subjects with premature coronary artery disease. Family studies reveal a correlation coefficient of 0.82 between average molecular weights of parents and offspring, with significance at 0.01. In order to assess the influence of environment on molecular weight of low density lipoprotein, the correlation coefficient between the fathers' and mothers' low density lipoprotein was measured and no statistically significant correlation was found. These data are interpreted as strong evidence for a genetic determination of molecular weight of low density lipoprotein. A study of individuals in five families yields molecular weight data consistent with a single gene locus genetic mode of inheritance without dominance. The regression coefficient of the mean low denisty lipoprotein parental molecular weight on the offspring molecular weight is 0.30. If the variability of molecular weight is considered as an expression of phenotypic variance, then the regression analysis identified 30% of this phenotypic variance as arising from additive gene action presumably at a single locus. Segregation in the family data is consistent. Since the differences in molecular weight of low density lipoprotein arise from differences in the amount of lipid bound to the apoprotein, it is likely that an additional portion of the phenotypic variance of the molecular weight results from individual variations in the metabolism of low density lipoprotein, which yield differences in lipid content. The individual variation in molecular weight is only approximately 5%; hence those metabolic sequences that influence molecular weight of low density lipoproteins must be precisely controlled.

Adolescent↗
PubMed1975

Preliminary model for human lipoprotein metabolism in hyperlipoproteinemia.

A model is proposed for the metabolism of plasma lipoprotein apoproteins based on studies of a hyperlipoproteinemic subject who received 2.5 mCi[3H]leucine intravenously. Measurements included apoprotein specific activities (apo-B and apo-C) of very low density lipoprotein (VLDL) and of three low density lipoprotein (LDL) subspecies, Sf 17 LDL, Sf 10 LDL, and Sf 4 LDL. Activities of plasma albumin were also determined. The data were analyzed using a compartmental model and the SAAM computer program. A chain-like series of compartments were necessary to simulate plasma VLDL kinetics, suggesting a multistep delipidation process. The data are consistent with the notion that VLDL is the dominant LDL precursor. Two modes of conversion from VLDL to LDL are required. After partial delipidation some VLDL is converted to the Sf 17 LDL, while the remainder undergoes further delipidation before being converted to Sf 4 LDL, the major plasma LDL component. Some direct release of LDL into plasma had to be introduced to fit the data, about 24% of total LDL production. The three LDL subspecies follow a precursor-product relationship (Sf 17 leads to Sf 10 leads to Sf 4). The analysis also indicates that in using labeled leucine as a tracer, the slow exchange of leucine with the total body protein pool must be considered in trying to resolve the LDL subsystem and in the estimation of steady-state apoprotein levels. In view of the fact that the proposed model is based predominantly on the data from a single patient, no generalizations can be made about parameter values. The study is most valuable, however, in pointing out metabolic pathways not considered before and in calling attention to variables that must be considered in the design of experiments to study lipoprotein kinetics.

Apoproteins↗