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Biomedical subjects

L S Jefferson

Publications and source records attributed to L S Jefferson.

At least 19 recordsLinked to original sources
PubMed1992

Age-dependent decrease in the amount of eukaryotic initiation factor 2 in various rat tissues.

Recent studies have suggested that the decline in protein synthesis that occurs in rat liver and brain during development and aging is associated with a decrease in the activity of eukaryotic initiation factor 2 (eIF-2). One way in which eIF-2 activity could be decreased in tissue extracts would be through a decrease in the activity of the GDP exchange factor, eIF-2B. In the present study, the activity of eIF-2B was measured in tissue extracts and was found to be less in older than in younger rats. Thus a decrease in eIF-2B activity could account for part of the decrease in protein synthesis that occurs during aging. Another way in which eIF-2 activity could be decreased would be through a decrease in amount of the protein. Therefore the amount of eIF-2 in various tissues was quantified by protein immunoblot analysis. We found that the amount of eIF-2 relative to total protein tended to fall with increasing age. Furthermore, eIF-2 content was directly proportional to the rate of protein synthesis in the tissues examined. Finally, slot-blot analysis of polyadenylated RNA revealed no significant change in the relative abundance of eIF-2 alpha mRNA with age. The last-mentioned experiments suggest that the synthesis of eIF-2 may be regulated through changes in the deficiency of translation of eIF-2 alpha mRNA rather than through changes in gene transcription.

Aging↗
PubMed1992

Severe laryngotracheobronchitis complicating measles.

OBJECTIVE: To determine the incidence of severe measles-related laryngotracheobronchitis in patients hospitalized during a recent measles epidemic and to evaluate factors associated with severity of airway injury and its management. DESIGN: Clinical description of patient series. SETTING: Children's hospital and county general hospital, Houston, Tex. PATIENTS: One hundred twenty-four children (aged 1 month to 19 years) admitted with a diagnosis of measles. INTERVENTIONS: None. MEASUREMENTS/RESULTS: Twenty-seven patients had significant laryngotracheobronchitis, including 10 who had not received appropriate immunization. Six patients required endotracheal intubation for relief of upper airway obstruction. The median age of patients requiring intubation was 12 months (range, 4 to 24 months). Two patients died of complications of superinfection. Two patients survived but required prolonged intubation. Two patients underwent early diagnostic laryngoscopy and bronchoscopy and required shorter artificial airway maintenance. CONCLUSIONS: Severe laryngotracheobronchitis frequently occurs in patients younger than 2 years hospitalized with measles and may be related to bacterial or viral super-infection. Early diagnostic laryngoscopy and bronchoscopy for injury assessment and possible endotracheal tube exchange are recommended and, in some severe cases, tracheostomy should be considered to shorten artificial airway maintenance and decrease the incidence of airway complications.

Adolescent↗
PubMed1992

Regulation of protein synthesis by modulation of intracellular calcium in rat liver.

The rate of protein synthesis can be modulated in intact cells by varying the concentration and subcellular distribution of intracellular calcium. Because the biochemical reactions required for the pathway of protein synthesis occur in the cytosol of the cell, it might be expected that protein synthesis would be controlled by free cytosolic calcium rather than the sequestered cation. However, a recent report proposed that maintenance of optimal rates of protein synthesis depends on the amount of calcium sequestered in the endoplasmic reticulum rather than free cytosolic calcium (C.O. Brostrom and M. A. Brostrom, Annu. Rev. Physiol. 52: 577-590, 1990). In the present study, rat livers were perfused with buffer containing various compounds previously shown to alter intracellular calcium concentration and distribution in isolated cells. It was found that conditions designed to cause a rise in free cytosolic calcium had no effect on protein synthesis. In contrast, conditions designed to cause depletion of sequestered calcium resulted in an inhibition of protein synthesis characterized by a reduction in peptide-chain initiation relative to elongation. The inhibition of protein synthesis was further localized to a decrease in the activity of eukaryotic initiation factor (eIF) 2B as measured in extracts from perfused livers. The inhibition of eIF-2B activity was associated with a 2.4-fold increase in the proportion of the alpha-subunit of eIF-2 in the phosphorylated form. In summary, the results of the present study support a model whereby mobilization of calcium sequestered in the endoplasmic reticulum results in an inhibition of protein synthesis in rat liver.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Effects of insulin on total RNA, poly(A)+ RNA, and mRNA in primary cultures of rat hepatocytes.

The purpose of this study was to examine mechanisms involved in the regulation of protein synthesis in primary cultures of rat hepatocytes. Hepatocytes were maintained in a chemically defined serum-free medium in the presence or absence of insulin. The rate of protein synthesis in hepatocytes deprived of insulin between days 2 and 5 of culture was reduced to 67% of the rate observed in insulin-maintained controls. The decrease in protein synthetic rate was accompanied by a proportional fall in the content of both total RNA and poly(A)+RNA, suggesting that the capacity for protein synthesis was reduced in the absence of insulin. Both total RNA and poly(A)+ RNA contents and the protein synthetic rate were returned to control values after 3 days of insulin resupplementation. In addition, the effect of insulin on the expression of specific mRNAs was assessed by in vitro translation of total RNA followed by two-dimensional gel analysis of radiolabeled translation products. Only 13 of the greater than 150 spots discernible on the two-dimensional gels were altered in response to insulin. The mRNAs that were altered include examples of repression and stimulation of expression in response to insulin deprivation. Thus, in isolated rat hepatocytes, insulin regulates the capacity of both overall protein synthesis as well as the capacity for the synthesis of specific proteins.

Animals↗
PubMed1991

Inhibition of microsomal calcium sequestration causes an impairment of initiation of protein synthesis in perfused rat liver.

The present study examined the effect of 2,5-di-(tert-butyl)-hydroquinone (tBuHQ), an inhibitor of liver microsomal calcium sequestration, on initiation of protein synthesis in perfused rat liver. Perfusion of livers with a concentration of tBuHQ previously shown to completely inhibit microsomal calcium sequestration in isolated hepatocytes caused a 50% inhibition of protein synthesis. The inhibition was characterized by an increase in liver content of free ribosomal particles and a decrease in polysomes indicating that peptide-chain initiation was slowed relative to elongation. Furthermore, the inhibition was associated with a 7.5-fold increase in the proportion of the alpha-subunit of eukaryotic initiation factor 2 (eIF-2) present in the phosphorylated form and a reduction in the activity of eukaryotic initiation factor 2B (eIF-2B) to 37% of the control value. The results suggest that protein synthesis in rat liver is regulated directly by changes in intracellular calcium concentration through a mechanism involving modulation of the phosphorylation state of eIF-2 alpha.

Animals↗
PubMed1991

Mechanism of inhibition of peptide chain initiation by amino acid deprivation in perfused rat liver. Regulation involving inhibition of eukaryotic initiation factor 2 alpha phosphatase activity.

In previous studies, initiation of protein synthesis was shown to be inhibited in perfused rat livers deprived of single essential amino acids. In the present study, histidinol, a competitive inhibitor of histidinyl-tRNA synthetase, was used to amplify the effects of histidine deprivation on protein synthesis in perfused liver to facilitate investigation of mechanisms involved in the inhibition of peptide chain initiation. Protein synthesis was reduced to 77% of the control rate in livers deprived of histidine and to 13% of the control rate in livers deprived of histidine and exposed to 2.0 mM histidinol. The inhibition of protein synthesis caused by histidine deprivation alone was accompanied by a 2-fold increase in the number of free ribosomal particles, a 29% decrease in Met-tRNA(i) binding to 43 S preinitiation complexes, and a 31% reduction in activity of eukaryotic initiation factor 2B (eIF-2B). By comparison, histidine deprivation combined with histidinol addition resulted in a 3-fold increase in free ribosomal particles, a 66% decrease in Met-tRNAi binding, and a 78% reduction in eIF-2B activity. The proportion of the alpha-subunit of eukaryotic initiation factor two (eIF-2) in the phosphorylated form increased from 8.9 +/- 0.8% in control livers to 52.4 +/- 5.5% in response to histidinol. The increase in the amount of eIF-2 alpha in the phosphorylated form apparently was not due to an increase in kinase activity, because there was no change in eIF-2 alpha kinase activity in extracts of liver perfused with medium containing histidinol compared to controls. Instead, the increased phosphorylation of eIF-2 alpha was associated with an inhibition of eIF-2 alpha phosphatase activity. Thus, in contrast to other systems that have been examined, the mechanism involved in the increase in the phosphorylation state of eIF-2 alpha appears to involve an inhibition of eIF-2 alpha phosphatase activity rather than activation of an eIF-2 alpha kinase.

Amino Acids↗
PubMed1991

Regulation of initiation of protein synthesis by insulin in skeletal muscle.

Protein synthesis is impaired in skeletal muscle and heart from diabetic rats. In muscles composed primarily of slow-twitch fibres (e.g. heart or soleus), the inhibition of protein synthesis can be accounted for entirely by a decrease in the amount of RNA. In contrast, in muscles of mixed fibre composition (e.g. gastrocnemius or psoas), the inhibition of protein synthesis is associated with an impairment in peptide-chain initiation. We have found that the inhibition of peptide-chain initiation that occurs in muscles composed of mixed fast-twitch fibres involves eukaryotic initiation factor 2B (eIF-2B). Thus, eIF-2B activity is inhibited in gastrocnemius and psoas but not heart or soleus from diabetic rats. In other systems eIF-2B activity is regulated by phosphorylation of the alpha-subunit of a second initiation factor, eIF-2. However, we have found no change in the phosphorylation state of eIF-2 alpha in either fast- or slow-twitch muscles from diabetic compared to control animals. Instead, the available evidence suggests that eIF-2B activity may be modulated by an alternate mechanism such as a change in the extent of phosphorylation of the 82,000 Mr subunit of the factor or a change in the NADPH/NADP+ ratio.

Animals↗
PubMed1991

Use of the Natural Death Act in pediatric patients.

OBJECTIVE: To review the use of Natural Death Act declarations (living will procedures) in pediatric patients. The implementation of such declarations for children is now possible in six states, including Texas, by specific statutory provisions. DESIGN: Retrospective study. SETTING: Pediatric ICU in a university hospital. PATIENTS: Records of patients who had a Texas Natural Death Act declaration, either discussed and signed or discussed only, were studied. Patients who had another vehicle of limiting care (e.g., a do-not-resuscitate order) were excluded from the study. MEASUREMENTS AND MAIN RESULTS: Reviewed characteristics included age, primary diagnosis, and concurrent complications. Also examined were who raised the issue of limiting care (parent or physician), the initial reaction of the other party, what support was withdrawn, what support was added, the final outcome (including the time from implementing limited care to death), and the description of witnesses. Discussions were held with parents of 17 patients, and 13 Natural Death Act declarations were actually implemented. In all but three instances, the patient died within 4 hrs from the time support was withdrawn. The main supports that were withdrawn were ventilators and catecholamines. In half of the cases, morphine sulfate was added for anticipated pain relief and sedation. All decisions were reached by close consultation between the family and the physicians, with the physicians raising the issue in 11 of the 17 cases and the family raising the issue in six cases. In 15 of the 17 patients, consultation with the Bioethics Committee was not necessary. The majority of difficulties involved resolving issues that beset patients with HIV infections, and finding appropriate witnesses as prescribed by the statute. CONCLUSIONS: We conclude that the Natural Death Act works well in situations involving dying children and their parents.

Adolescent↗
PubMed1991

Effect of diabetes and insulin treatment of diabetic rats on total RNA, poly(A)+ RNA, and mRNA in skeletal muscle.

We have assessed the time course of alterations in several biochemical parameters and expression of specific mRNAs in gastrocnemius muscle following both the induction of diabetes and the administration of insulin to diabetic rats. Muscle mass, total RNA, and total protein were reduced, whereas poly(A)+ RNA relative to total RNA was increased following the induction of diabetes. All the above parameters, with the exception of poly(A)+ RNA, were reciprocally and rapidly altered following administration of insulin to 3-day diabetic animals. These changes suggest that during the induction of diabetes 1) total cellular protein is reduced at a rate that is less than the reduction in gastrocnemius mass, whereas RNA is reduced at a rate 1.5 times the reduction in tissue mass, and 2) poly(A)+ RNA is elevated relative to total RNA. After insulin administration, there appears to be coordinate synthesis of both poly(A)+ RNA and ribosomal RNA, assuming 85% of total RNA is ribosomal. Therefore, we conclude that poly(A)+ RNA is more stable than ribosomal RNA during diabetes, whereas the amounts of poly(A)+ RNA and ribosomal RNA are increased at the same rates following insulin administration to diabetic animals. Analysis of expression of specific gene products over the same time course, as assessed by in vitro translation of total RNA followed by two-dimensional gel analysis, suggests that there are a few mRNAs that are very rapidly altered in response to insulin administration. The mRNAs that are altered demonstrate variable temporal patterns of either repression or full or transient expression. These rapid, but limited, alterations in gene expression may prove important in the development of the defects that occur in skeletal muscle in response to diabetes.

Animals↗
PubMed1991

Management of malignant air leak in a child with a neonatal high-frequency oscillatory ventilator.

A 10-year-old, 36-kg child with a malignant air leak who failed conventional mechanical ventilation and high-frequency jet ventilation was successfully treated with a neonatal high-frequency oscillatory ventilator for 31 days. Since the air leak resolved with minimal hemodynamic compromise, this technique may have application in the management of respiratory failure and air leak in the older and larger child for prolonged periods of time.

Bronchial Fistula↗
PubMed1990

Mechanism of the inhibition of protein synthesis by vasopressin in rat liver.

A recent study reported that protein synthesis was inhibited in rat livers perfused with medium containing vasopressin (Chin, K. -V., Cade, C., Brostrom, M. A., and Brostrom, C. O. (1988) Int. J. Biochem. 20, 1313-1319). The inhibition of protein synthesis caused by vasopressin was associated with a disaggregation of polysomes, suggesting that peptide chain initiation was slowed relative to elongation. In contrast, Redpath and Proud (Redpath, N. T., and Proud, C. G. (1989) Biochem. J. 262, 69-75) recently reported an inhibition of peptide chain elongation by a calcium/calmodulin-dependent mechanism. Therefore, the question remained whether only peptide chain initiation was inhibited or both initiation and elongation were affected by vasopressin. In the present study, vasopressin was found to inhibit protein synthesis in both perfused rat livers and isolated rat hepatocytes. Ribosomal half-transit times in isolated hepatocytes averaged 1.9 +/- 0.1 min with or without vasopressin present in the media, demonstrating that the rate of peptide chain elongation was unaffected by vasopressin. Instead, the inhibition of protein synthesis induced by vasopressin was manifested at the level of peptide chain initiation. Vasopressin treatment resulted in both a 2-fold increase in the number of free ribosomal particles and a greater than 50% decrease in the amount of [35S]methionine bound to 43 S preinitiation complexes. In addition, the activity of eukaryotic initiation factor (eIF) 2B in crude extracts from perfused livers was reduced to 53% of the control value in response to vasopressin. The inhibition of eIF-2B activity was associated with an increase in the proportion of the alpha-subunit of eIF-2 in the phosphorylated form from 9.6% in control livers to 30.7% in livers perfused with medium containing vasopressin. The results demonstrate the novel finding that the inhibition of protein synthesis in vasopressin-treated livers is caused by a reduction in eIF-2B activity due to an increase in phosphorylation of eIF-2 alpha.

Animals↗
PubMed1990

High-dose, short-duration ribavirin aerosol therapy in children with suspected respiratory syncytial virus infection.

Nine children (aged 6 weeks to 7 years) with suspected respiratory syncytial virus infection received aerosal treatment with ribavirin, 60 mg/ml for 2-hour periods three times daily for up to 5 days. Five children received treatment via an endotracheal tube and four via an oxygen hood. Blood samples (3 to 17 per patient) and respiratory secretions (4 to 23 per patient) were assayed for ribavirin with reverse-phase high-performance liquid chromatography. Ribavirin triphosphate in erythrocytes was determined by ion-exchange high-performance liquid chromatography. The mean (+/- SD) peak ribavirin level after the first dose was 1725 +/- 2179 mumol/L in secretions and 3.8 +/- 2.6 mumol/L in plasma. Ribavirin in the secretions was rapidly cleared, with a mean (+/- SD), half-life of 1.9 +/- 0.8 hours. Plasma ribavirin increased with treatments to reach a steady state of 5 to 10 mumol/L. Mean peak ribavirin triphosphate levels were 15- to 300-fold higher than plasma ribavirin levels by the end of therapy. More than 98% reduction of viral load without the emergence of resistant virus was noted on day 3 of therapy. High-dose treatment was compatible with the aerosol equipment routinely used (small-particle aerosol generator, model 2-6000) for ribavirin administration and with ventilators. High-dose, short-duration ribavirin therapy was well tolerated by all patients, permitted easier accessibility for patient care, and may result in less environmental exposure of health care workers.

Administration, Inhalation↗
PubMed1990

Back pain with cardiovascular collapse in a pediatric emergency department patient.

Severe back pain in the pediatric patient is an infrequent complaint. The following case report illustrates the disastrous outcome for this patient with back pain secondary to aortic dissection. The most common predisposing disease process is hypertension, which exposes the aortic wall to high pressures and flows. Just as in the adult patient, the possibility of aortic dissection should be included in the differential diagnosis of acute onset of severe back pain with preexisting hypertension in the pediatric patient. A high index of suspicion is warranted.

Adolescent↗
PubMed1989

Effect of amino acid deprivation on initiation of protein synthesis in rat hepatocytes.

Conditions were defined for maintaining optimal protein synthetic activity in suspensions of freshly isolated rat hepatocytes. Under these conditions, isolated hepatocytes exhibited rates of protein synthesis and levels of polysomal aggregation equivalent to those observed in vivo and in perfused liver. Deprivation of total amino acids or single, essential amino acids resulted in a rapid decrease in the rate of protein synthesis, which was readily reversed by readdition of the deficient amino acid(s). The decrease was accompanied by a disaggregation of polysomes and an inhibition of 43S initiation complex formation, which was indicative of a limitation in the rate of initiation of protein synthesis. Extracts prepared from perfused liver deprived of amino acids were inhibitory to initiation of protein synthesis in reticulocyte lysate. The inhibition in reticulocyte lysate was accompanied by an increase in phosphorylation of the alpha-subunit of eukaryotic initiation factor 2 (eIF-2), suggesting activation of an eIF-2 alpha kinase or inhibition of a phosphatase in amino acid-deprived hepatocytes. This suggestion was confirmed by prelabeling hepatocytes with 32Pi before amino acid deprivation. Incorporation of 32Pi into eIF-2 alpha was two- to threefold higher in lysine-deprived cells than in hepatocytes incubated in fully supplemented medium. Overall, the results indicated that an increase in eIF-2 alpha phosphorylation was responsible for the defect in initiation of protein synthesis caused by amino acid deprivation.

Adenosine Triphosphate↗
PubMed1989

Initiation of protein synthesis in a cell-free system prepared from rat hepatocytes.

A cell-free system, which maintained a linear rate of protein synthesis for up to 20 min of incubation, was prepared from isolated rat hepatocytes. The rate of protein synthesis in the cell-free system was approximately 20% of the rate obtained in isolated hepatocytes or perfused liver. More than 70% of total protein synthesis in the cell-free system was due to reinitiation, as indicated by addition of inhibitors of initiation, i.e., edeine or polyvinyl sulfate. The rate of protein synthesis and formation of 43S initiation complexes in the cell-free system were reduced to 60 and 30% of the control values, respectively, after incubation of hepatocytes in medium deprived of an essential amino acid. Therefore, the cell-free system maintained the defect in initiation induced in the intact cells by amino acid deprivation. The defect in initiation was corrected by addition of either rat liver eukaryotic initiation factor 2 or the guanine nucleotide exchange factor (GEF) to the cell-free system. A role for GEF in the defect in initiation was further implicated by experiments that showed that the activity of the factor was decreased in extracts from livers perfused with medium deficient in amino acids. The cell-free system should provide a valuable tool for investigation of mechanisms involved in the regulation of initiation of protein synthesis.

Adenosine Triphosphate↗
PubMed1988

Effect of diabetes on guanine nucleotide exchange factor activity in skeletal muscle and heart.

The present study examined the effects of diabetes and insulin treatment of diabetic rats on the activity of the protein synthesis initiation factor, the guanine nucleotide exchange factor. In extracts from gastrocnemius and psoas muscles from two-day diabetic rats, guanine nucleotide exchange factor activity was reduced to 80% and 67% of control values, respectively. Insulin treatment (2 h) restored guanine nucleotide exchange factor activity to control values in both muscles. In contrast, guanine nucleotide exchange factor activity was unchanged in extracts from either soleus muscle or heart from diabetic rats compared to controls. Also, insulin treatment did not increase guanine nucleotide exchange factor activity in extracts from soleus and heart. The results suggest that the diabetes-induced impairment in peptide-chain initiation in fast-twitch skeletal muscle (i.e. gastrocnemius and psoas) is related to an inhibition of guanine nucleotide exchange factor activity and that slow-twitch muscle is spared from the effect on initiation due to the preservation of guanine nucleotide exchange factor activity.

Animals↗
PubMed1988

Quantitation of proteins by dot-immunobinding assay. A comparison of visualization methods using eukaryotic initiation factor 2 and a monospecific antibody.

Various visualization methods were compared for quantitation of proteins by the dot-immunobinding assay. Comparisons were carried out using a multi-subunit protein, eukaryotic initiation factor 2, and monospecific antibodies directed against two of the factor's subunits. The protein was spotted onto nitrocellulose and the membranes were incubated with primary antibody. The antigen-antibody complex was visualized by one of six methods using either alkaline phosphatase-, horseradish peroxidase-, or glucose oxidase-conjugated IgG, or colloidal gold-labelled IgG, colloidal gold-labelled IgG with silver enhancement, or 125I-labelled protein A. The amount of secondary antibody bound was quantitated by densitometric scanning of the nitrocellulose membrane after staining or autoradiography. The sensitivity of each of the methods was similar; each of the visualization methods could detect less than 1 ng of protein by the dot-immunobinding assay. Curves of protein concentration vs. densitometric absorbance were found to fit a parabolic relationship (r2 = 0.99) over a wide range of concentrations.

Alkaline Phosphatase↗