Showing papers by "University of Texas Southwestern Medical Center published in 1985"
TL;DR: The LDL receptor appears to be a mosaic protein built up of exons shared with different proteins, and it therefore belongs to several supergene families.
Abstract: The multifunctional nature of coated pit receptors predicts that these proteins will contain multiple domains. To establish the genetic basis for these domains (LDL) receptor. This gene is more than 45 kilobases in length and contains 18 exons, most of which correlate with functional domains previously defined at the protein level. Thirteen of the 18 exons encode protein sequences that are homologous to sequences in other proteins: five of these exons encode a sequence similar to one in the C9 component of complement; three exons encode a sequence similar to a repeat sequence in the precursor for epidermal growth factor (EGF) and in three proteins of the blood clotting system (factor IX, factor X, and protein C); and five other exons encode nonrepeated sequences that are shared only with the EGF precursor. The LDL receptor appears to be a mosaic protein built up of exons shared with different proteins, and it therefore belongs to several supergene families.
800 citations
TL;DR: The molecular size of the plasma LDL receptor synthesized by cultured fibroblasts from a patient with the internalization-defective form of familial hypercholesterolemia (FH 274) was smaller by 10,000 daltons, and a 5-kilobase deletion appeared to be caused by a novel intrastrand recombination between two repetitive sequences of the Alu family that were oriented in opposite directions.
Abstract: The molecular size of the plasma LDL (low density lipoprotein) receptor synthesized by cultured fibroblasts from a patient with the internalization-defective form of familial hypercholesterolemia (FH 274) was smaller by 10,000 daltons than the size of the normal LDL receptor. The segment of the gene encoding the truncated portion of the FH 274 receptor was cloned into bacteriophage lambda. Comparison of the nucleotide sequences of the normal and FH 274 genes revealed a 5-kilobase deletion, which eliminated the exons encoding the membrane-spanning region and the carboxyl terminal cytoplasmic domain of the receptor. The deletion appeared to be caused by a novel intrastrand recombination between two repetitive sequences of the Alu family that were oriented in opposite directions. The truncated receptors lack membrane-spanning regions and cytoplasmic domains; they are largely secreted into the culture medium, but a small fraction remains adherent to the cell surface. The surface-adherent receptors bind LDL, but they are unable to cluster in coated pits, thus explaining the internalization-defective phenotype.
429 citations
TL;DR: In this article, the authors used objective functional capacity measurement techniques to guide a treatment program for a group of 66 chronic back pain patients and found that the functional capacity measures collected for the treatment group improved in approximately 80% of the patients.
Abstract: Objective functional capacity measurement techniques were used to guide a treatment program for a group of 66 chronic back pain patients. These patients were compared with a group of 38 chronic patients who were not administered the treatment program. Outcome data were collected by telephone survey at an average 1 year follow-up. In addition, functional capacity measures were collected for treatment group patients on admission and follow-up evaluations. Results demonstrated that the functional capacity measures collected for the treatment group improved in approximately 80% of the patients. These changes were also accompanied by positive changes in psychologic measures. In addition, at 1 year follow-up, the treatment group had approximately twice the rate of patients who returned to work, relative to the comparison group. Additional surgery rates were comparable for both groups (6% in the treatment and 7% in the comparison group), but the frequency of additional health-care professional visits was substantially higher in the comparison group. The findings suggest that quantitative functional capacity measures can give objective evidence of patient physical abilities and degree of effort and can significantly guide the clinician in administering an effective treatment program.
404 citations
TL;DR: It is demonstrated that B cell stimulatory factor (BSF-1) and B cell differentiation factor (BCDF-gamma) are the same lymphokine and that BSF-1 acts on both resting and activated B cells to induce different effects.
Abstract: By three criteria, we have demonstrated that B cell stimulatory factor (BSF-1) and B cell differentiation factor (BCDF-gamma) are the same lymphokine. Highly purified preparations of high performance liquid chromatography-purified or affinity-purified BSF-1 had BCDF-gamma activity but not BCDF-mu activity. A monoclonal anti-BSF-1 antibody coupled to Sepharose depleted both BSF-1 and BCDF-gamma activity but not BCDF-mu activity from two different T cell supernatants. Soluble monoclonal anti-BSF-1 blocked the BSF-1 and BCDF-gamma but not the BCDF-mu responses. These results suggest that BSF-1 acts on both resting and activated B cells to induce different effects.
388 citations
TL;DR: It is concluded that the membrane-bound domain of reductase plays a crucial role in the rapid and regulated degradation of this ER protein.
374 citations
TL;DR: It could be concluded that gel reorganization required physical rearrangement of pre-existing collagen fibrils rather than degradation of the original collagen and resynthesis of a new matrix.
Abstract: During reorganization of collagen gels by human skin fibroblasts the total protein content of the gels remained approximately constant. Only 5% of the collagen was degraded, although the volume of the gels decreased by 85% or more. It could be concluded, therefore, that gel reorganization required physical rearrangement of pre-existing collagen fibrils rather than degradation of the original collagen and resynthesis of a new matrix. Collagen molecules in the gels were not covalently crosslinked or otherwise modified enzymically during gel reorganization, as determined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis and collagen repolymerization studies. Serum was required for gel reorganization and, in the absence of serum, cell spreading was predominantly filipodial, i.e. there was little cytoplasmic reorganization. At the electron-microscopic level it was found that many more collagen fibrils became associated with the cells in the presence of serum than in its absence. Serum was also found to promote the synthesis and secretion of proteins by the cells, and conditioned medium could take the place of serum in promoting gel reorganization. The involvement of cell-secreted factors was also demonstrated by the ability of cycloheximide to inhibit gel reorganization. Finally, when gel reorganization was stopped by adding cytochalasin D to the incubations or removing cells by detergent treatment, a small but significant re-expansion of the collagen fibrils was observed. Consequently, a portion of the collagen that had been physically reorganized by the gels was unstable and could not hold its position without continued force exerted by the cells.
298 citations
TL;DR: Findings indicated that receptor-mediated LDL transport was suppressed approximately equal to 30% by cholesterol feeding alone and this was unaffected by the addition of polyunsaturated triacylglycerols to the diet.
Abstract: The liver plays a key role in the regulation of circulating levels of low density lipoproteins (LDL) because it is both the site for the production of and the major organ for the degradation of this class of lipoproteins In this study, the effects of feeding polyunsaturated or saturated triacylglycerols on receptor-dependent and receptor-independent hepatic LDL uptake were measured in vivo in the hamster In control animals, receptor-dependent LDL transport manifested an apparent Km value of 85 mg/dl (plasma LDL-cholesterol concentration) and reached a maximum transport velocity of 131 micrograms of LDL-cholesterol/hr per g, whereas receptor-independent uptake increased as a linear function of plasma LDL levels Thus, at normal plasma LDL-cholesterol concentrations, the hepatic clearance rate of LDL equaled 120 and 9 microliter/hr per g by receptor-dependent and receptor-independent mechanisms, respectively As the plasma LDL-cholesterol was increased, the receptor-dependent (but not the receptor-independent) component declined When cholesterol (012%) alone or in combination with polyunsaturated triacylglycerols was fed for 30 days, receptor-dependent clearance was reduced to 36-42 microliter/hr per g, whereas feeding of cholesterol plus saturated triacylglycerols essentially abolished receptor-dependent LDL uptake (5 microliter/hr per g) When compared to the appropriate kinetic curves, these findings indicated that receptor-mediated LDL transport was suppressed approximately equal to 30% by cholesterol feeding alone and this was unaffected by the addition of polyunsaturated triacylglycerols to the diet In contrast, receptor-dependent uptake was suppressed approximately equal to 90% by the intake of saturated triacylglycerols As compared to polyunsaturated triacylglycerols, the intake of saturated lipids was also associated with significantly higher plasma LDL-cholesterol concentrations and lower levels of cholesteryl esters in the liver
292 citations
TL;DR: The association of agonists with muscarinic receptors in membranes from bovine brain was affected only slightly by guanine nucleotides, but solubilization of these membranes with deoxycholate and subsequent removal of detergent resulted in a preparation of receptors with increased affinity for agonists and a large increase in response to guanines.
281 citations
TL;DR: A model for the secondary structure and membrane orientation of 3-hydroxy-3-methylglutaryl coenzyme A reductase, the glycoprotein of the endoplasmic reticulum that controls the rate of cholesterol biosynthesis, is presented and evaluated.
278 citations
TL;DR: Genomic DNA libraries from cells of two individuals with internalization-defective familial hypercholesterolemia suggest that the signal for targeting the LDL receptor to coated pits resides in the cytoplasmic domain of the molecule.
269 citations
TL;DR: The results suggest that in vertebrate skeletal muscle, P-light chain phosphorylation increases the force level at submaximal Ca2+ concentrations, probably by affecting the interaction between the myosin cross-bridge and the thin filament.
TL;DR: Using this procedure, human placental ChAT was purified to homogeneity with high recovery of enzyme activity (50–60%) and was used to raise a monospecific anti‐human ChAT polyclonal antibody in rabbits.
Abstract: A rapid and efficient immunoaffinity purification procedure has been developed for human placental choline acetyltransferase (ChAT). Using this procedure, human placental ChAT was purified to homogeneity with high recovery of enzyme activity (50–60%). Purified ChAT was used to raise a monospecific anti-human ChAT polyclonal antibody in rabbits. A comparison of the physical properties of ChAT was made between the enzymes purified from human brain and human placenta. Only one form of the enzyme exists in either tissue, having identical molecular weights of 68,000 and a single apparent pI of 8.1. A more detailed comparison of the two enzymes using peptide mapping and epitope mapping indicates identity between the brain and placental enzymes.
TL;DR: While additional proteins may modulate adenylate cyclase activity in native membranes, these results show that these three proteins are sufficient for the expression of hormone-stimulatedadenylates cyclase.
Journal Article•
TL;DR: The results suggest that the increasing cardiovascular stresses of late pregnancy, especially when intensified by parturition, seriously compromise women with ischemic heart disease and efforts should be made to limit myocardial oxygen demand/consumption throughout pregnancy, and particularly during parturitions.
TL;DR: It is concluded that increased estrone production associated with aging may result from an increase in the specific activity of the aromatase enzyme in adipose stromal cells and is not affected by changes in gonadotropin concentrations associated with menopause.
Abstract: Adipose tissue is the principal site of estrogen formation in postmenopausal women; with advancing age as well as with increased body weight, there is an increase in the fractional conversion of circulating androstenedione to estrone. We have studied the effects of aging as well as body weight on aromatase activity of adipose tissue specimens taken from 50 women of various ages and weights. Since aromatase activity of adipose tissue is detectable primarily in stromal cells, these cells were incubated with [1-3H]androstenedione (150 nM), and estrogen formation was assayed by measuring the incorporation of tritium into [3H]water. The aromatization rate, when normalized on the basis of equal numbers of cells, increased with increasing age (P ¼ 0.03; r = 0.43). In contrast, when expressed as a function of body weight, no change in aromatase activity of adipose stromal cells were found. Aromatization of androstenedione by cells from young women who had undergone oophorectomy was not increased compared with tha...
TL;DR: A complementary DNA clone encoding the alpha subunit of the adenylate cyclase stimulatory G protein (Gs) was isolated and identified and RNA that hybridizes with probes made from the clone is detected in wild-type S49 cells; however, cyc- S 49 cells, which are deficient in Gs alpha activity, are devoid of this messenger RNA.
Abstract: A complementary DNA clone encoding the alpha subunit of the adenylate cyclase stimulatory G protein (Gs) was isolated and identified. A bovine brain complementary DNA library was screened with an oligonucleotide probe derived from amino acid sequence common to known G proteins. The only clone that was obtained with this probe has a complementary DNA insert of approximately 1670 base pairs. An antibody to a peptide synthesized according to deduced amino acid sequence reacts specifically with the alpha subunit of Gs. In addition, RNA that hybridizes with probes made from the clone is detected in wild-type S49 cells; however, cyc- S49 cells, which are deficient in Gs alpha activity, are devoid of this messenger RNA.
TL;DR: These studies suggest that rates of hepatic cholesterol synthesis and receptor-dependent LDL uptake are regulated independently and the primary response of the liver to changes in cholesterol availability is regulation of sterol synthesis and only when the capacity of this compensatory mechanism is exceeded is the rate of LDL transport altered.
TL;DR: Kinetic studies with pure human renal NEP showed that the chemotactic peptide fMet-Leu-Phe was one of the best biologically active substrates and indicates that NEP in neutrophils may have important functions in inflammation and chemotaxis.
Abstract: Membrane metallo-endopeptidase (NEP; neutral endopeptidase, kidney-brush-border neutral proteinase, enkephalinase, EC 3.4.24.11) cleaves peptides at the amino side of hydrophobic amino acids. While the enzyme is known to be in organs such as kidney and brain, we found it in human neutrophils. These cells cleaved the NEP substrate glutaryl (Glut)-Ala-Ala-Phe-(4-methoxynaphthylamine) (Glut-Ala-Ala-Phe-MNA) at a rate of 9.5 nmol X hr-1 per 10(6) cells, and phosphoramidon (1 microM) inhibited the hydrolysis by 90%. Intact neutrophils from donors who smoked had NEP activities about twice that of nonsmokers. Subcellular fractionation and sucrose density gradient centrifugation of lysed neutrophils showed that most of the NEP activity was membrane bound. A washed membrane fraction from human neutrophils rapidly cleaved 0.5 mM Glut-Ala-Ala-Phe-MNA (96 nmol X min-1 X mg-1) and the hydrolysis was inhibited by phosphoramidon and by specific antiserum to human renal NEP. The washed membrane fraction also rapidly cleaved 0.1 mM bradykinin (34 nmol X min-1 mg-1) and 0.1 mM fMet-Leu-Phe (49 nmol X min-1 X mg-1). The membrane-bound enzyme cleaved the peptide substrates at the same site as the homogeneous human renal NEP, and phosphoramidon and thiorphan inhibited the hydrolysis. Kinetic studies with pure human renal NEP showed that the chemotactic peptide fMet-Leu-Phe was one of the best biologically active substrates (Km, 59 X 10(-6) M; kcat, 3654 min-1). Immunocytochemistry at the light microscopic level revealed a high concentration of NEP on the cell membrane of neutrophils. This was confirmed with electron microscopy using the immunogold technique on ultrathin cryosections. These studies indicate that NEP in neutrophils may have important functions in inflammation and chemotaxis.
TL;DR: It is concluded that short-term potassium supplementation that ameliorates diuretic-induced hypokalemia may induce a significant fall in blood pressure.
Abstract: Changes in potassium balance have been found to have variable effects on the blood pressure of animals, and the administration of potassium supplements has been reported to lower the blood pressure of normokalemic hypertensive patients. To assess the effect of potassium repletion in hypokalemic hypertension, we administered either potassium chloride, 60 mmol per day, or placebo tablets, each for six weeks, in a randomized, double-blind, crossover trial to 16 hypertensive patients who had diuretic-induced hypokalemia and who continued to take a constant amount of diuretic. We selected patients whose control serum potassium levels were below 3.5 mmol per liter. In association with an average rise in the serum potassium concentration of 0.56 mmol per liter, the mean blood pressure fell by an average of 5.5 mm Hg (P = 0.004), with at least a 4 mm Hg fall observed in 9 of the 16 patients. The fall in blood pressure correlated with a fall in plasma renin activity (r = 0.568, P = 0.043) but not with cha...
TL;DR: The goals of this study were to measure isometric and isokinetic strength of trunk flexors, extensors and rotators, and explore relationships among these muscle groups; and compare a torque to body weight adjustment measure versus lean body weight.
Abstract: The goals of this study were to: (1) evaluate the repeatability of a method of measuring trunk strength; (2) measure isometric and isokinetic strength of trunk flexors, extensors and rotators; (3) explore relationships among these muscle groups; and (4) compare a torque to body weight adjustment mea
TL;DR: It is concluded that ototoxic screening should be reserved for patients defined as “at risk” and those patients receiving more than 400 mg of DDP under the conditions stated in this report.
Abstract: Twenty-four patients with head and neck neoplasms were prospectively evaluated for cisplatin (DDP)-induced ototoxicity. Patients were selected from a larger population based on the uniformity of their chemotherapy regimen, renal status, lack of prior or concurrent exposure to ototoxic agents, and availability for repetitive audiometric testing in the same setting. Scanning electron microscopy of the inner ear was performed on four temporal bones. Hearing impairment was found to be dose-related, irreversible within the confines of the study period, and primarily in the higher frequencies. Vestibular toxicity was rare and well-documented by our testing methods in only one patient. Based on the results of this study, and a review of animal and human data on DDP ototoxicity, the authors concluded that ototoxic screening should be reserved for patients defined as “at risk” and those patients receiving more than 400 mg of DDP under the conditions stated in this report. Cancer 56: 1934-1939, 1985.
TL;DR: Every reductase-CAT plasmid that showed transcriptional activity also showed inhibition by sterols, indicating that the sites for promotion and inhibition of transcription are closely associated.
TL;DR: This work has shown that the amino acid sequences of the human low-density lipoprotein (LDL) receptor and the human precursor for epidermal growth factor (EGF) show 33 percent identity over a stretch of 400 residues, suggesting that the homologous region may have resulted from a duplication of an ancestral gene.
Abstract: The amino acid sequences of the human low-density lipoprotein (LDL) receptor and the human precursor for epidermal growth factor (EGF) show 33 percent identity over a stretch of 400 residues. This region of homologous is encoded by eight contiguous exons in each respective gene. Of the nine introns that separate these exons, five are located in identical positions in the two protein sequences. This finding suggests that the homologous region may have resulted from a duplication of an ancestral gene and that the two genes evolved further by recruitment of exons from other genes, which provided the specific functional domains of the LDL receptor and the EGF precursor.
TL;DR: It was found that the IgG from the serum of the rabbit immunized with the 55-kDa protein bound specifically to a protein of 55 kDa in human placental microsomes, indicative that both the monoclonal and polyclonal IgGs are specific for human cytochrome P-450AROM.
TL;DR: In this paper, the GTPase activity of the stimulatory guanine nucleotide-binding regulatory protein (Gs) of hormone-sensitive adenylate cyclase was investigated using purified rabbit hepatic Gs and either [alpha-32P]- or [gamma- 32P] GTP as substrate.
TL;DR: Although there is excellent agreement between Fick and both thermodilution and dye measurements of CO, ther modilution is preferable to dye in patients with a low CO and those with aortic or mitral regurgitation.
Abstract: This study was performed to assess the relation between Fick and indicator dilution measurements of cardiac output (CO) in a large number of subjects and to evaluate this relation in patients with a low CO, a high CO, and left-sided cardiac regurgitation. In 808 patients (428 men, 380 women, mean age 50 +/- 11), CO was measured by Fick and either thermodilution (right atrium to pulmonary artery)(n = 252) or indocyanine green dye ("dye")(pulmonary artery to systemic artery)(n = 556) within 10 minutes of each other. There was excellent agreement between Fick and both thermodilution and dye. The difference between Fick and indicator dilution measurements was 9 +/- 9%; it was 10% or less in 67% and 20% or less in 91% of patients. The disparity between Fick and indicator dilution measurements was increased in patients with a low CO (less than 2 liters/min/m2)(n = 152) (difference 14 +/- 11%, p less than 0.001) and those with aortic or mitral regurgitation (n = 83) (difference 13 +/- 11%, p less than 0.001). In these groups, the disparity between Fick and thermodilution measurements was not exaggerated, but the disparity between Fick and dye measurements was greater. Thus, although there is excellent agreement between Fick and both thermodilution and dye measurements of CO, thermodilution is preferable to dye in patients with a low CO and those with aortic or mitral regurgitation.
TL;DR: Evidence is provided that the open reading frame encodes a protein required for intron transposition and support the role of the double-strand break in the process.
TL;DR: The study showed that a significant fraction of VLDL-B is removed directly from the circulation and never reaches LDL regardless of the type of patients, and that in patients with obesity and in those with CHD, synthesis rates of V LDL particles may be abnormally high.
Abstract: This study was designed to examine the integrated metabolism of apolipoprotein B (apo B) in very low density lipoproteins (VLDL), intermediate density lipoproteins (IDL), and low density lipoproteins (LDL) in normal subjects, obese patients, and a group of patients with coronary heart disease (CHD) Turnover rates of 131I-VLDL-B, 131I-IDL-B, 125I-LDL-B, and [3H]VLDL-triglycerides (TG) were determined by the multicompartmental analysis that used the model described in the preceding article (Beltz, WF, et al 1985 J Clin Invest 76: 575-585) Compared with five normal subjects, four obese subjects had increased synthesis rates of both VLDL-B and VLDL-TG Production of LDL-B was inconsistently raised in these same patients Five patients with CHD had enhanced production of both VLDL-B and LDL-B, but secretion rates of VLDL-TG were not increased Thus, in patients with obesity and in those with CHD, synthesis rates of VLDL particles may be abnormally high In the obese patients, the VLDL appeared to be of normal composition, but in patients with CHD, the VLDL were relatively poor in TG The study also showed that a significant fraction of VLDL-B is removed directly from the circulation and never reaches LDL regardless of the type of patients The fraction that does reach LDL is one factor that determines LDL concentrations
TL;DR: Results strongly support direct interaction of miconazole with microsomal cytochrome P-450 in human placental microsomes with high affinity resulting in the inhibition of aromatase activity.
TL;DR: With indirect immunofluorescence using a monoclonal antibody, it was shown that the testis' interstitial tissue is well-endowed with cells bearing class II histocompatibility antigens, a prerequisite for antigen-processing capability.
Abstract: The significance of several local factors in prolonging allograft survival in the rat testis has been investigated. The lower temperature of this organ was shown to have no effect, because parathyroid gland allografts implanted s.c. in the ear, which is similarly hypothermic, were rejected as promptly as in conventional sites. Conversely, testes secured in the abdominal cavity afforded privilege to grafts just as well as normal testes. With indirect immunofluorescence using a monoclonal antibody, it was shown that the testis' interstitial tissue is well-endowed with cells bearing class II histocompatibility antigens, a prerequisite for antigen-processing capability. When intratesticular allograft recipients were pretreated with estrogen to suppress Leydig cell synthesis of testosterone, most grafts were rejected promptly, suggesting that local steroid secretion is important in testicular immune privilege. Both testes and s.c. sites supported inflammatory reactions to cotton pledgets, indicating that there is no interference with the nonspecific inflammatory process necessary for graft rejection, but more likely with antigen processing itself.