Thioredoxin domain non-equivalence and anti-chaperone activity of protein disulfide isomerase mutants in vivo

Coexpression of the enzyme, protein disulfide isomerase (PDI), has been shown to increase soluble and secreted IgG levels from baculovirus-infected insect cells (Hsu, T.-A., Watson, S., Eiden, J. J., and Betenbaugh, M. J. (1996) Protein Expression Purif. 7, 281-288). PDI is known to include catalytic active sites in two separate thioredoxin-like domains, one near the amino terminus and another near the carboxyl terminus. To examine the role of these catalytic active sites in enhancing immunoglobulin solubility, baculovirus constructs were utilized with cysteine to serine mutations at the first cysteine of one or both of the CGHC active site sequences. Trichoplusia ni insect cells were coinfected with a baculovirus vector coding for IgG in concert with either the wild-type human PDI virus, amino-terminal mutant (PDI-N), carboxyl-terminal mutant (PDI-C), or mutant with both active sites altered (PDI-NC). Western blot analysis revealed that both immunoglobulins and PDI protein were expressed in the coinfected cells. To evaluate the effect of the PDI variants on immunoglobulin solubility and secretion, the infected cells were labeled with 35S-amino-acids for different periods, and the soluble immunoglobulins were immunoprecipitated from clarified cell lysates and culture medium using anti-IgG antibodies. Only coinfections with the wild-type PDI and PDI-N mutant led to increased immunoglobulin solubility and higher IgG secretion. In contrast, infection with the PDI-C and PDI-NC variants actually lowered immunoglobulin solubility levels below those achieved with a negative control virus. Immunoprecipitation with anti-PDI antibody revealed that heterologous PDI-C and PDI-NC were insoluble, even though PDI-N and wild-type PDI protein were detected in soluble form. The capacity for PDI-N to increase immunoglobulin solubility whereas the PDI-C mutant lowered solubility indicates that the amino- and carboxyl-terminal thioredoxin domains of PDI are functionally distinct in vivo following mutations to the active site. Furthermore, mutations at the active site of the carboxyl-terminal thioredoxin domain result in PDI variants that can act as anti-chaperones of immunoglobulin solubility in vivo as has been observed in vitro for lysozyme aggregation by wild-type PDI and PDI mutants (Puig, A., and Gilbert, H. F. (1994) J. Biol. Chem. 269, 7764-7771).     

A 21st century computerized injection system for local pain control

This article describes a new computerized local anesthetic injection system for pain control. The core technology of this system is the microprocessor-controlled delivery of anesthetic solution at a constant pressure and controlled volume, regardless of encountered variations in tissue resistance. This fine-tuned, high suffusion flow rate of anesthetic provides a rapid onset of anesthesia for most patients. Traditional block injections and infiltrations as well as palatal injections and periodontal ligament injections are administered with precision, ease, and patient comfort.     

Hierarchical polytomous regression models with applications to health services research

The analysis of variations is an important area of interest in health services and outcomes research and has two main goals: to identify and quantify variability across units, such as geographic regions or health care providers, in terms of procedure utilization and outcomes, and to explore the links between process, such as regional or hospital practice patterns, and outcomes, such as patient mortality and functional status. Hierarchical regression models are well suited for this type of analysis. In this paper we formulate a hierarchical polytomous regression model and apply it to the analysis of variations in the utilization of alternative cardiac procedures in a national cohort of elderly Medicare patients who had an acute myocardial infarction during 1987. The model is designed to accommodate clustered multinomial data with covariate vectors available on individual cases and on clusters. We present a Bayesian approach to fitting and checking the model using simulated values from the posterior distribution of the parameters. The simulation algorithms are based on Gibbs sampling in combination with Metropolis steps. Using the hierarchical polytomous regression model, we examine how the rates of cardiac procedures depend on patient-level characteristics, including age, gender and race, and whether there exist interstate differences and regional patterns in the use of these procedures.     

Limiting amino acids in meat and bone and poultry by-product meals

In situ, digestion, and growth studies were conducted to evaluate four meat and bone meals and six poultry by-product meals as sources of escape protein and to predict the first-limiting amino acid for growing calves. Escape protein values, determined by 12-h in situ incubation, ranged from 41.7 to 51.0% of CP for meat and bone meals; poultry by-product meals ranged from 32.0 to 39.8%. True protein digestion in the gastrointestinal tract of lambs differed among protein sources (P < .05), ranging from 79 to 95%. In each of three growth trials, 60 steers (258 +/- 24, 241 +/- 23, and 230 +/- 16 kg for Trials 1, 2, and 3, respectively) were supplemented with 4 of the 10 protein sources along with a urea supplement. Protein sources were fed at 30, 40, 50, and 60% of the supplemental CP, with urea supplying the remainder. Protein efficiency differed among treatments ( P < .10), ranging from .61 to 1.55. Amino acid composition was determined for each protein source, and the individual metabolizable amino acids were regressed on the protein efficiency values. Escape protein values were correlated (R2 = .75) with protein efficiency but had a negative slope. Metabolizable methionine was the only amino acid moderately correlated (R2 = .40, slope = 1.9) to protein efficiency, whereas other amino acids either correlated poorly or had negative slopes. These data indicate that the protein value of meat and bone meal and poultry by-product meal is limited by the amount of metabolizable methionine they contain.     

Effects of growth hormone, insulin-like growth factor-I, and cortisol on periparturient antibody response profiles of dairy cattle

The objectives of this study were to determine hormone and antibody response profiles from the prepartum period to peak lactation, and evaluate potential immunomodulatory effects of the classic endocrine hormones, growth hormone (GH), insulin-like growth factor-I (IGF-I) and cortisol. Specifically, 33 Holstein cows were immunized with ovalbumin (OVA) and Escherichia coli J5 at weeks -8 and -3 prior to parturition. At parturition (week 0), cows received an additional immunization of OVA. Blood was collected at weeks -8, -3, 0, 3 and 6 relative to parturition and various samples were used to determine plasma hormone concentration, serum immunoglobulin (Ig), and specific antibody response to OVA and E. coli. Colostrum and milk samples were also collected post-parturition to monitor local immunoglobulin and antibody responses. Results indicated that not all periparturient cows exhibited depressed immune response, and that antibody response to OVA could be used to partition cows into 3 groups recognizing animals with sustained measurable antibody response before and after parturition (Group 1), animals which responded poorly to immunization at parturition (Group 2), and animals which did not respond to immunizations at week -3 or parturition (Group 3). Cows with the highest antibody response to OVA (Group 1) also tended (P < or = 0.10) to have the highest response to E. coli J5 at parturition and had the lowest incidence of disease, particularly mastitis. Antibody response to OVA measured in milk tended to be higher in Group 1 cows, particularly at week 0 (P < or = 0.06) compared to cows of Group 3. IGF-I was higher (P < or = 0.05) in cows of Group 1 than Group 3 at peak lactation (week 6).     

Aging rat vestibular ganglion: II. Quantitative electron microscopic evaluation

This laboratory has shown that age-related vestibular ganglion cell loss does not occur in the Wistar rat as it does in humans. However, in that study, intracellular changes were evident. The purpose of the present study was to quantitate some of these changes. The volume densities of mitochondria, rough endoplasmic reticulum (RER), Golgi apparatus, and aging pigment, as well as the diameter of the vestibular ganglion cells, of young (3 to 5 months) and old (24 to 31 months) female Wistar rats were determined by electron microscopy and stereological techniques. The data show a significant decrease in the volume densities of mitochondria (11.4%), Golgi apparatus (8.1%), and RER (8.9%), a significant increase in aging pigment (327%), and no change in mean profile diameter. These results suggest a decreased capacity for oxidative metabolism and protein synthesis that may reflect a decrease in the number of hair cells innervated by each ganglion cell and/or in the number of central connections. In either case, these findings suggest impaired metabolic and functional capabilities.     

Augmented prenatal auditory stimulation alters postnatal perception, arousal, and survival in bobwhite quail chicks

Progress in biological psychiatry is contingent on progress in neurobiology and on research into proper characterisation and assessment of abnormal behavior. Advances in neurobiology are rapid and steady; diagnostic research does not keep pace. On the contrary, the diagnostic approach seems solidified, as today's basic premises are uncritically accepted. The diagnostic requirements for meaningful biological psychiatric research are discussed and contrasted with present-day diagnostic practices. Serious reflection on the state of psychiatric diagnosing is urgently needed.     

Manganese, nickel, selenium and cadmium in molluscs from the Magellan Strait, Chile.

The concentrations of manganese, selenium, nickel and cadmium were determined in 112 samples of molluscs belonging to mussels (Mytilus chilensis, n = 47) and limpets (Nacella deaurata, n = 65), which were collected from the coastline of the Magellan Strait, Chile. Four (6.2%) samples of limpets exceeded the maximum limits for cadmium established in Europe. Limpets showed higher mean manganese, nickel and cadmium concentrations than mussels, whilst the mean selenium concentration in mussels was higher. The consumption of one serving (100 g) of molluscs represents a considerable contribution to the dietary daily intake of selenium, and limpets make a significant contribution to the manganese and cadmium intakes. The sampling zone influenced the trace element concentrations, and different uptakes were observed between the mollusc species.