Circulating thyroid hormone autoantibodies.
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Animals↗
Biomedical subjects
J Robbins
Publications and source records attributed to J Robbins.
Evidence for dopaminergic innervation on kitten retinal ganglion cells.
Effects of iontophoretically applied dopamine and its receptor antagonists on physiologically identified retinal ganglion cells were studied in the optically intact eye of pentobarbitone anaesthetised kittens (7-9 weeks of age) and the results were compared with the effects obtained from adult cats (18-22 weeks of age). In both the adult and the kitten, dopamine had an inhibitory effect on visually evoked and spontaneous activity of the retinal ganglion cells, irrespective of cell type. However, unlike in the adult, the effects of dopamine in kittens were variable and not dependent on retinal eccentricity. In adult cells, only L-sulpiride (a potent D2 receptor antagonist) reduced the inhibitory effect of exogenous dopamine, whereas in kitten cells, both alpha-flupenthixol (a potent D1 receptor antagonist) and L-sulpiride did so. When applied alone, neither alpha-flupenthixol nor affected the activity of ganglion cells in adults, but in kittens both antagonists produced an excitatory effect in some cells. Physiologically active dopaminergic innervation seemed, therefore, to be present on the immature ganglion cells, but was subsequently 'eliminated' during the course of postnatal development. Furthermore, in immature cells, both D1 and D2 type receptors are present but only D2 receptors remain in adult. Therefore, there is a mismatch between dopamine receptors and dopamine in the adult retina and this mismatch may be the consequence of a developmental event.
Action Potentials↗
Chromosomal localization of the met proto-oncogene in the mouse and cat genome.
The met proto-oncogene was mapped in the mouse and cat genomes with the use of mouse X hamster and cat X rodent somatic cell hybrid DNA panels. Based on these analyses we assigned the met gene to mouse chromosome 6 and to cat chromosome A2. We also assigned the cat raf-1 proto-oncogene to the A2 chromosome; met and raf-1 are the first cloned DNAs mapped to this linkage group. Using an interspecies backcross we further localized met on mouse chromosome 6 to a position proximal to the beta chain of the T-cell receptor. This places met near the obese locus in a region of mouse chromosome 6 that appears to be homologous with the long arm of human chromosome 7. The close linkage of met to the gene responsible for cystic fibrosis in humans suggests that further genetic analysis of mouse chromosome 6 may be useful in developing a mouse model for the disease.
Animals↗
Clinical assessment of oropharyngeal motor development in young children.
A clinical protocol was developed for the purpose of assessing the oral and speech motor abilities of children. An 86-item test was administered to 90 normally developing children aged 2:6-6:11. Evaluations of the structural integrity of the vocal tract did not show developmental change, although evaluations of oral and speech motor functioning changed significantly with age. The functional portion of the protocol was most sensitive to developmental change up to age 3:6, with an asymptote in performance thereafter. Clinical application of the protocol is discussed.
Age Factors↗
Role of sodium in thyroid hormone uptake by rat skeletal muscle.
Whether Na+ movement through the plasma membrane plays a role in thyroid hormone uptake was investigated in intact rat soleus muscles. After preincubation for 120 min at 37 degrees C in modified Krebs-Ringer bicarbonate containing 140 or 5 mM Na+ plus choline or lithium to maintain osmolarity, muscles were incubated with 50 pM [125I]triiodo-L-thyronine (T3) or [125I]L-thyroxine (T4) for 60 min. T3 uptake was decreased when extracellular Na+ was replaced by either choline or lithium, the amount of decrease corresponding to the specific (or saturable) uptake component. Monensin, an ionophore that stimulates Na+ entry, increased T3 uptake at 140 mM Na+ but not at 5 mM Na+. Amiloride, a Na+/H+ exchange inhibitor, had no effect on T3 uptake under basal conditions or when Na+ was replaced by choline, but reversed the action of lithium. Ouabain, an inhibitor of Na+/K+ ATPase, reduced specific T3 uptake. T4 uptake was unaffected by low extracellular Na+. These results are consistent with a major role of Na+ movement in T3 uptake by skeletal muscle, but not in T4 uptake, and suggest an involvement of membrane pumps in this process.
Amiloride↗
Further studies on the role of glycosylation in thyroxine-binding globulin secretion by human hepatoma (HEP G2) cells.
To investigate the role of glycosylation of T4-binding globulin (TBG) in the secretion of the protein, human hepatoma (Hep G2) cells were continuously labeled with [35S] methionine or [3H]mannose or pulse-chase labeled with [35S] methionine in the absence or presence of 1 microgram/ml swainsonine, an inhibitor of Golgi alpha-mannosidase II and lysosomal alpha-mannosidase. In the presence of this alkaloid, TBG was released into the medium at a faster rate than in control cells (50% being secreted after 35 min and 47 min, respectively), owing to accelerated intracellular transport of the newly synthesized protein. TBG secreted from swainsonine-treated cultures moved faster in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, probably because of the reduced sialylation of TBG consequent to the perturbed processing of the oligosaccharide units. Furthermore, secreted TBG was sensitive to endo-beta-N-acetylglucosaminidase H digestion as shown by the shift in the apparent molecular size in sodium dodecyl sulfate-polyacrylamide gel electro-phoresis from 50,000 to 45,000 daltons. Sensitivity to endo H indicated the presence of hybrid-type oligosaccharide chains with high mannose structures. This was also suggested by the higher incorporation of [3H]mannose in swainsonine-treated cultures. In conclusion, the results of the present study demonstrate that swainsonine accelerates the release of TBG from Hep G2 cells and that complete processing of oligosaccharide moieties is not required for TBG secretion.
Alkaloids↗
Intracellular transport of 3,5,3'-triiodo-L-thyronine in rat skeletal myoblasts.
Since in skeletal muscle circulating L-T3 is the only source for the hormone bound to nuclei, we investigated the intracellular and intranuclear transport of L-T3 in L6E9 rat skeletal muscle cells. The characteristics of this process were assessed by analyzing the nuclear bound L-T3 as a marker of the internalized hormone and by determining the initial rate of L-T3 uptake. [125I]L-T3 cellular uptake at 37 C reached a plateau at 2 h when the nuclear uptake, after an initial lag phase, was still increasing and represented 4.7% of the cellular uptake. Incubation at 4 C caused [125I]L-T3 cellular uptake to decrease by 77% and nuclear uptake to be abolished. A similar effect on [125I]L-T3 nuclear uptake was obtained after myoblasts were incubated at 37 C with a 1000-fold excess of unlabeled L-T3. The addition of various inhibitors of ATP production, cytoskeleton integrity, endocytosis, and Na+, K+-ATPase that did not interfere with [125I]L-T3 binding to the cell surface or to isolated nuclei caused a dose-dependent reduction of both extranuclear and nuclear uptake, ranging from 34-85%. Scatchard analysis revealed the presence on the myoblast surface of L-T3 high affinity (Ka = 1.6 X 10(9) M-1) and low affinity (Ka = 5.4 X 10(6) M-1) binding sites; other iodothyronines exhibited lower affinity for both sites. Kinetic analysis of L-T3 transport after 60-sec incubation at 23 C defined a process with a Km of 17 +/- 5.6 nM and a maximum velocity of 83 +/- 35 pmol/mg DNA. These results indicate the existence in rat myoblasts of a temperature-dependent, energy-requiring, saturable, and stereospecific L-T3 uptake mechanism, probably mediated through an endocytotic pathway, located on the myoblast plasma membrane, that may regulate L-T3 action in skeletal muscle.
Animals↗
Swallowing in ALS and motor neuron disorders.
The dysphagia of ALS is characterized by an impaired oral stage of swallowing, which has a direct impact on the seemingly more robust pharyngeal stage. Increased duration of the volitional stage (PPRT) of swallowing, including DT and lingual motility deficits, appear to be the major contributors to the dysphagia. The abnormal temporospatial events during the oral stage of swallowing have a direct and significant effect on the more automatic, pharyngeal stage. Specifically, abnormal lingual movement may result in pharyngeal residue that is aspirated after the swallow is completed and respiration is resumed. ALS patients with bulbar involvement demonstrate more severe swallowing problems (such as aspiration); however, the current work indicates that predominantly nonbulbar ALS patients may also demonstrate dysphagia. The semisolid material was more effective than liquids in eliciting temporospatial abnormalities of swallowing in the nonbulbar group. Thus, more viscous materials may be most sensitive for eliciting behaviors indicative of the onset of bulbar dysfunction in patients who otherwise appear nonbulbar, or different neurophysiologic mechanisms are involved in swallowing the distinct types of substances.
Adult↗
Nutrition and immune function in the geriatric population.
Aging in experimental animals and in man is associated with a significant defect of helper/inducer T-lymphocytes. This quantity deficiency is paralleled by a qualitative deficiency of T-lymphocyte number and function. Similar defects have been observed in protein calorie malnutrition and zinc deficiency, and there is evidence that both may exist in the aged ill and perhaps even in aged healthy individuals. Nutritional support improves the immune defect in aged ill patients and it may be of value in improving this deficiency, even when administered to aged healthy subjects.
Age Factors↗
Effect of free fatty acids and nonlipid inhibitors of thyroid hormone binding in the immunoradiometric assay of thyroxin-binding globulin.
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Binding, Competitive↗
Pharmacist-operated drug information centers in the United States--1986.
Pharmacist-operated drug information centers (DICs) were surveyed early in 1986 to obtain current information concerning the status, availability, and scope of their services. Questionnaires were sent to 191 institutions believed to be active in disseminating drug information; 127 responses were usable. Descriptive information on the respondents is tabulated, and a comprehensive directory of pharmacist-operated DICs is provided. Significant changes have occurred in the affiliations of DICs since previous surveys, and sources of DIC funding seem to be related to these affiliations. Many DICs appear to have the potential to answer more questions than they actually answer. Since few of them are self-sustaining, DICs should attempt to document how their activities affect prescribing practices and, hence, overall hospital finances.
Computers↗
The chicken myosin heavy chain family.
We have used a myosin heavy chain gene fragment to probe two chicken genomic libraries. The fragment, derived from a gene expressed in the fast-white fibers of adult chickens, contains 1400 bases upstream from the translational start site and 1600 bases downstream from the initiation codon. Thirty-one unique nonoverlapping clones were isolated. All of the genes showed homologies in the nucleotide sequences which code for the globular head portion of the myosin protein while no extensive homologies were detected in the 5'-flanking sequences. The relationships between the genes were studied using oligomeric sequences as probes. The hybridization patterns showed that seven of the genes fall within a well defined subgroup. The exon containing a domain of the nucleotide (ATP) binding site was sequenced for all seven of these genes and shown to be, except for 2 nucleotides in one of the genes, completely conserved. The lack of sequence conservation in the 5'-nontranslated portions of the genes was exploited in the preparation of transcript-specific probes. We have used these probes to show that two of the isoforms are expressed in a tissue-specific and developmental stage-specific manner in the chicken.
Adenosine Triphosphate↗
A canonical sequence organization at the 5'-end of the myosin heavy chain genes.
The sequences encoding the 5'-ends of three chicken fast-white myosin heavy chain (MHC) genes have been determined. When compared with the sequences of two other MHC genes it is apparent that both the exon and intron positions are conserved. All exon sequences are highly conserved; there is absolute amino acid conservation in the second and third exons. In addition, while the first and third introns diverge among the genes, the second intron is highly conserved between the five. This intron contains a 24-bp sequence that is repeated twice in one of the introns and once in the other four. Analyses indicate that this sequence, which is partially homologous to 7SL RNA, appears to be largely restricted to the MHC gene family. Analysis of the 5'-flanking sequences show that while small homologies are present between some of the genes, they have extensively diverged in this region.
Amino Acid Sequence↗
Blood use in urgent operations for patients with fractures of the femoral neck.
Unnecessary preoperative cross-matching of blood wastes time and money and may increase morbidity and mortality by delaying a necessary operation. One hundred and ninety-three consecutive patients who underwent semi-urgent operative treatment for fractures of the neck of the femur at a large district general hospital were evaluated. Of the patients with haemoglobin values of 11 g/dl or more, only 12 per cent required transfusion for the operation. None of the patients with fractures treated by 'pinning' required a transfusion. It is our opinion that patients with normal preoperative haemoglobin levels can undergo operative treatment for fractures of the neck of femur after typing but without the necessity of cross-matching blood.
Blood Grouping and Crossmatching↗
Characteristics of speech production after tracheoesophageal puncture: voice onset time and vowel duration.
Voice onset time (VOT) and vowel duration characteristics of speakers following the Singer-Blom technique of tracheoesophageal puncture (1980) were compared to those of traditional esophageal and laryngeal speakers. Fifteen subjects in each of the three speaker groups produced the words /pik/, /kap/, /kup/ in a carrier phrase while audio recordings were obtained. Broadband spectrograms were made of the consonant-vowel-consonant (CVC) utterances and vowel duration and VOT were measured. Analysis of variance (ANOVA) procedures revealed that the tracheoesophageal speakers produced significantly shorter VOTs and longer vowel durations than the laryngeal speakers. However, the longer vowel durations for the tracheoesophageal speakers were not completely accounted for by the shorter VOTs found for that group. Spectrographic examination suggests that delayed voice offset time for the tracheoesophageal speakers also contributes to their longer vowel durations. Overall findings indicate that the physical characteristics and motor control properties of the neoglottis, even when driven by pulmonary air as in tracheoesophageal speakers, exert a major influence on alaryngeal voice production.
Aged↗
Purification and partial characterization of a novel thyroxine-binding protein (27K protein) from human plasma.
T4-binding globulin (TBG) prepared from human plasma by the standard three-step procedure (T4-agarose affinity chromatography, anion exchange chromatography, and gel filtration) often shows in sodium dodecyl sulfate-polyacrylamide gel electrophoresis in addition to the expected 54K band, another with a mol wt of 27,000 (27K protein). The two proteins can be separated after the three-step procedure by chromatofocusing (because of different isoelectric points, 4.2-4.8 for TBG and 5.0-5.2 for 27K protein) or by T4-aragose chromatography eluting with a linear gradient of T4 (TBG is eluted between 10(-10) and 10(-9) M T4, 27K protein between 10(-8) and 10(-7) M T4). The 27K protein does not appear to be a fragment of TBG since 1) it does not displace [125I]TBG bound to anti-TBG monoclonal antibodies; and 2) absorption of polyclonal antibody reacting with both TBG and 27K protein with sera from TBG-deficient patients completely prevents [125I]27K protein binding, while only slightly affecting [125I]TBG binding. On the other hand, 27K protein is not simply a contaminant devoid of biological activity, but is a T4-binding protein, as supported by the following findings: 1) it covalently binds [125I]T4 by photoaffinity labeling, and this binding can be almost completely prevented by excess T4; 2) equilibrium dialysis shows two equivalent T4-binding sites per 66K, with an association constant of 0.85 X 10(7) M-1, intermediate between albumin and prealbumin; and 3) tryptophanyl fluorescence analysis shows quenching of 37% of the fluorescence when the protein is titrated with T4. The 27K protein appears as a single 27K band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, pH 8.8, but under nondenaturing nonreducing conditions mostly remains at the origin of the gel; a fraction enters the gel and migrates slightly ahead of albumin. This electrophoretic pattern is distinct from those of albumin, prealbumin, and TBG. In immunoelectrophoresis in agar at pH 8.6, 27K protein moves slightly faster than TBG. The results of equilibrium sedimentation indicate a mol wt of 66,000, suggesting that the 27K protein might exist as a dimer. These data indicate that the 27K protein is a previously unrecognized T4-binding protein with a low affinity for the hormone. Further studies are required to clarify its physiological role in the transport of circulating thyroid hormones.
Antibodies, Monoclonal↗
Biosynthesis of a novel thyroxine-binding protein (27K protein) in human hepatoma (Hep G2) cells.
Human hepatoma (Hep G2) cells were shown to synthesize and secrete a novel T4-binding protein, called 27K protein for its apparent mol wt on sodium dodecyl sulfatepolyacrylamide gel electrophoresis. The mRNA coding for this protein was characterized by immunoprecipitation of [125I]T4 bound to 27K protein secreted into the medium of oocytes injected with total Hep G2 RNA. Sucrose gradient fractionation of RNA from Hep G2 cells showed that TBG mRNA and 27K mRNA had different sizes, indicating that TBG and 27K protein are two distinct proteins. In vitro translation of RNA in a rabbit reticulocyte lysate demonstrated that the translation product immunoprecipitated by anti-27K serum had the same mol wt as the immunoprecipitated protein from whole cells labeled with [35S]methionine, thus suggesting that 27K protein is neither derived from TBG nor synthesized through a larger mol wt precursor, and also that it does not contain carbohydrates. The absence of carbohydrates was further supported by the observation that [3H]mannose was not covalently bound to the 27K protein when Hep G2 cells were labeled with [3H]mannose, nor was there a shift in apparent mol wt when the cells were treated with the glycosylation inhibitor tunicamycin. The kinetics of secretion of 27K protein were similar to those of albumin and faster than those of TBG, which is also in keeping with the nonglycoprotein nature of 27K protein.
Animals↗
Effect of the antileukemic agent L-asparaginase on thyroxine-binding globulin and albumin synthesis in cultured human hepatoma (HEP G2) cells.
L-Asparaginase (ASNase), a drug widely used in the treatment of acute lymphoblastic leukemia, has been reported to decrease serum T4-binding globulin (TBG) levels, while results of serum albumin determinations were conflicting. This effect in vivo has been attributed to depressed liver protein synthesis, but this hypothesis has not been proved. To investigate this problem, human hepatoma (Hep G2) cells were continuously labeled for 4 h with 100 microCi/ml [35S]methionine in the absence or presence of graded amounts of ASNase (from 0.1 nM to 0.1 mM). Media and cell lysates were collected, immunoprecipitated with antialbumin or anti-TBG serum and protein A, and submitted to sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Gels were sliced, and the radioactivity was counted in a beta-counter. A dose-dependent inhibition of TBG and albumin biosynthesis (as well as of total protein synthesis) was demonstrable, but TBG appeared to be more sensitive to the action of the drug. In fact, TBG biosynthesis was reduced by 8% with 0.1 nM ASNase, while an effect on albumin was observed only at 1 nM ASNase; 50% inhibition was obtained with 30 nM ASNase in the case of TBG and with 800 nM in the case of albumin. At the highest concentration (0.1 mM), TBG biosynthesis was reduced by 94%, and albumin biosynthesis by 75%. ASNase also proved to have a time-dependent effect, as assessed by the measurement of radioimmunoassayable TBG in the media from Hep G2 cells grown in the presence of 10 nM ASNase for 1-4 days. The TBG concentration was progressively reduced, by 40% after 1 day to 85% after 4 days. In pulse-chase experiments, a reduction of total (intracellular plus secreted) immunoprecipitable TBG and, to a lesser extent, albumin was observed, suggesting that the drug also affected the catabolism of newly synthesized proteins. These results provide the first in vitro evidence that ASNase actually inhibits TBG biosynthesis. This effect is not specific for TBG, but this protein appears to be more susceptible than albumin to ASNase action. This can explain why in patients treated with ASNase for leukemia, a decrease in serum TBG concentrations has not always been associated with a reduction in serum albumin levels.
Asparaginase↗