Subcellular structure and behaviour in fungal hyphae.

This work briefly surveys the diversity of selected subcellular characteristics in hyphal tip cells of the fungal kingdom (Mycota). Hyphae are filamentous cells that grow by tip extension. It is a highly polarised mechanism that requires a robust secretory system for the delivery of materials (e.g. membrane, proteins, cell wall materials) to sites of cell growth. These events result it the self-assembly of a Spitzenkörper (Spk), found most often in the Basidiomycota, Ascomycota, and Blastocladiomycota, or an apical vesicle crescent (AVC), present in the most Mucoromycota and Zoopagomycota. The Spk is a complex apical body composed of secretory vesicles, cytoskeletal elements, and signaling proteins. The AVC appears less complex, though little is known of its composition other than secretory vesicles. Both bodies influence hyphal growth and morphogenesis. Other factors such as cytoskeletal functions, endocytosis, cytoplasmic flow, and turgor pressure are also important in sustaining hyphal growth. Clarifying subcellular structures, functions, and behaviours through bioimagining analysis are providing a better understanding of the cell biology and phylogenetic relationships of fungi. LAY DESCRIPTION: Fungi are most familiar to the public as yeast, molds, and mushrooms. They are eukaryotic organisms that inhabit diverse ecological niches around the world and are critical to the health of ecosystems performing roles in decomposition of organic matter and nutrient recycling (Heath, 1990). Fungi are heterotrophs, unlike plants, and comprise the most successful and diverse phyla of eukaryotic microbes, interacting with all other forms of life in associations that range from beneficial (e.g., mycorrhizae) to antagonistic (e.g., pathogens). Some fungi can be parasitic or pathogenic on plants (e.g., Cryphonectria parasitica, Magnaporthe grisea), insects (e.g., Beauveria bassiana, Cordyceps sp.), invertebrates (e.g., Drechslerella anchonia), vertebrates (e.g., Coccidioides immitis, Candia albicans) and other fungi (e.g., Trichoderma viride, Ampelomyces quisqualis). The majority of fungi, however, are saprophytes, obtaining nutrition through the brake down of non-living organic matter.

Expression of coxsackie and adenovirus receptor distinguishes transitional cancer states in therapy-induced cellular senescence.

Therapy-induced cellular senescence describes the phenomenon of cell cycle arrest that can be invoked in cancer cells in response to chemotherapy. Sustained proliferative arrest is often overcome as a contingent of senescent tumor cells can bypass this cell cycle restriction. The mechanism regulating cell cycle re-entry of senescent cancer cells remains poorly understood. This is the first report of the isolation and characterization of two distinct transitional states in chemotherapy-induced senescent cells that share indistinguishable morphological senescence phenotypes and are functionally classified by their ability to escape cell cycle arrest. It has been observed that cell surface expression of coxsackie and adenovirus receptor (CAR) is downregulated in cancer cells treated with chemotherapy. We show the novel use of surface CAR expression and adenoviral transduction to differentiate senescent states and also show in vivo evidence of CAR downregulation in colorectal cancer patients treated with neoadjuvant chemoradiation. This study suggests that CAR is a candidate biomarker for senescence response to antitumor therapy, and CAR expression can be used to distinguish transitional states in early senescence to study fundamental regulatory events in therapy-induced senescence.

Prevention of alcohol-induced developmental delays and learning abnormalities in a model of fetal alcohol syndrome.

OBJECTIVE: Prenatal alcohol exposure results in fetal death and neurobehavioral complications including learning impairment. Previously synthetic peptides derived from activity-dependent neurotrophic factor have been shown to prevent aspects of alcohol-induced damage in pregnancy. The objective of this work was to evaluate whether activity-dependent neurotrophic factor-12 could prevent alcohol-induced damage in a model of fetal alcohol syndrome. STUDY DESIGN: Using a well-characterized model, C57Bl6/J mice on gestational day 8 were treated with placebo, alcohol (30% volume/volume alcohol 0.03 mL/kg), alcohol plus activity-dependent neurotrophic factor-12 30 minutes prior to alcohol, or activity-dependent neurotrophic factor-12 alone. Fetal death was assessed on gestational day 18 (25 litters were evaluated: alcohol, n = 5; placebo, n = 9; alcohol plus activity-dependent neurotrophic factor-12, n = 11). Neonatal behavior tests were performed on postnatal days 1 through 21 with the offspring of 12 dams (alcohol, n = 16; placebo, n = 46; alcohol plus activity-dependent neurotrophic factor-12, n = 23; and activity-dependent neurotrophic factor-12, n = 35). Adult males were tested in the Morris water maze for learning assessment and with the hole punch activity test for exploratory activity. Statistical analysis included Kruskal-Wallis and analysis of variance. RESULTS: Fetal death was greater in alcohol (67% +/- 13%) vs placebo (8.4% +/- 3%, P < .001). Pretreatment with activity-dependent neurotrophic factor-12 prevented the alcohol-induced fetal death (2.2% +/- 8.1%) with levels similar to control (P = .12). Alcohol exposure caused a delay in achieving developmental milestones, with alcohol achieving milestones later than all other groups (all P < .001). Pretreatment with activity-dependent neurotrophic factor-12 prevented the alcohol-induced milestone delays. In the Morris water maze, the placebo learned, decreasing their latency to find the hidden platform over 70% (P < .01). Alcohol plus activity-dependent neurotrophic factor-12 also significantly learned, with a learning curve not different from placebo (all P > .5) and significantly better than alcohol on days 4, 6, and 7 (all P < .05). Alcohol exposure resulted in significantly less time in hole punch activity (P < .02) than control. Activity-dependent neurotrophic factor-12 pretreatment prevented the alcohol-induced decline, with levels the same as control (P = .1). CONCLUSION: The novel peptide activity-dependent neurotrophic factor-12 prevents alcohol-induced fetal death and developmental and learning abnormalities in a model of fetal alcohol syndrome. This demonstrates that a single treatment with a peptide is efficacious and may be of value in the prevention of alcohol-induced damage.

Antifungal and cancer cell growth inhibitory activities of 1-(3',4',5'-trimethoxyphenyl)-2-nitro-ethylene.

The antifungal and cancer cell growth inhibitory activities of 1-(3',4',5'-trimethoxyphenyl)-2-nitro-ethylene (TMPN) were examined. TMPN was fungicidal for the majority of 132 reference strains and clinical isolates tested, including those resistant to fluconazole, ketoconazole, amphotericin B or flucytosine. Minimum fungicidal concentration/minimum inhibitory concentration (MFC/MIC) ratios were < or = 2 for 96% of Cryptococcus neoformans clinical isolates and 71% of Candida albicans clinical isolates. TMPN was fungicidal for a variety of other basidiomycetes, endomycetes and hyphomycetes, and its activity was unaffected by alterations in media pH. The frequency of occurrence of fungal spontaneous mutations to resistance was <10(-6). Kill-curve analyses confirmed the fungicidal action of TMPN, and demonstrated that killing was concentration- and time-dependent. At sub-MIC exposure to TMPN, C. albicans did not exhibit yeast/hyphae switching. TMPN was slightly cytotoxic for murine and human cancer cell lines (GI50=1-4 microg ml(-1)), and weakly inhibited mammalian tubulin polymerization (IC50=0.60 microg ml(-1)).

Metoclopramide versus ondansetron in prophylaxis of nausea and vomiting for laparoscopic cholecystectomy.

BACKGROUND: Postoperative nausea and vomiting are significant problems in laparoscopic surgery. This double-blind, randomized, prospective trial compares the prophylactic use of metoclopramide, ondansetron, and placebo for the treatment of postoperative nausea and vomiting in patients undergoing outpatient laparoscopic cholecystectomy. METHODS: Two hundred thirty-two patients aged 18 to 73 years were randomized into three groups. Patients received intravenously 10 mg of metoclopramide, 4 mg of ondansetron, or placebo in a double-blinded manner prior to surgery. RESULTS: The incidence of nausea was 32% for metoclopramide, 45% for ondansetron, and 44% for placebo in the postanesthesia care unit or day surgery, which was not statistically significant. The incidence of vomiting was 8% for metoclopramide, 4% for ondansetron, and 22% for placebo in the postanesthesia care unit or day surgery. These differences were statistically significant when comparing both drugs to placebo but not when comparing both drugs to each other. CONCLUSION: Prophylactic administration of metoclopramide or ondansetron significantly reduces the incidence of postoperative vomiting for laparoscopic cholecystectomy, but neither drug was found to be significantly more effective than the other. Metoclopramide is a more cost-effective treatment.

Long-range repression by multiple polycomb group (PcG) proteins targeted by fusion to a defined DNA-binding domain in Drosophila.

A tethering assay was developed to study the effects of Polycomb group (PcG) proteins on gene expression in vivo. This system employed the Su(Hw) DNA-binding domain (ZnF) to direct PcG proteins to transposons that carried the white and yellow reporter genes. These reporters constituted naive sensors of PcG effects, as bona fide PcG response elements (PREs) were absent from the constructs. To assess the effects of different genomic environments, reporter transposons integrated at nearly 40 chromosomal sites were analyzed. Three PcG fusion proteins, ZnF-PC, ZnF-SCM, and ZnF-ESC, were studied, since biochemical analyses place these PcG proteins in distinct complexes. Tethered ZnF-PcG proteins repressed white and yellow expression at the majority of sites tested, with each fusion protein displaying a characteristic degree of silencing. Repression by ZnF-PC was stronger than ZnF-SCM, which was stronger than ZnF-ESC, as judged by the percentage of insertion lines affected and the magnitude of the conferred repression. ZnF-PcG repression was more effective at centric and telomeric reporter insertion sites, as compared to euchromatic sites. ZnF-PcG proteins tethered as far as 3.0 kb away from the target promoter produced silencing, indicating that these effects were long range. Repression by ZnF-SCM required a protein interaction domain, the SPM domain, which suggests that this domain is not primarily used to direct SCM to chromosomal loci. This targeting system is useful for studying protein domains and mechanisms involved in PcG repression in vivo.

Biochemical characterization of an active pyrophosphate-dependent phosphofructokinase from Treponema pallidum.

An active pyrophosphate-dependent phosphofructokinase containing a six residue polyhistidine tag has been cloned from Treponema pallidum, and characterized biochemically. The phosphofructokinase has pH optima for activity of 8.0 for both the forward and reverse reactions. The apparent K(m) for pyrophosphate was 0.042 mM (V(max) of 141 U mg(-1) protein) and for fructose-6-phosphate, 0.529 mM. The apparent K(m) for the reverse reaction for fructose-1,6-diphosphate was 0.267 mM (V(max) of 42.4 U mg(-1) protein). The enzyme appears to be both a dimer and non-allosteric.

Safety and immunogenicity of improved Shigella O-specific polysaccharide-protein conjugate vaccines in adults in Israel.

Data suggest that the O-specific polysaccharide (O-SP) domain of the lipopolysaccharide (LPS) of Shigella species is both an essential virulence factor and a protective antigen and that a critical level of serum immunoglobulin G (IgG) to this antigen will confer immunity to shigellosis. Because covalent attachment of polysaccharides to proteins increases their immunogenicity, especially in infants and in young children, the O-SP of Shigella species were bound to medically useful proteins, and the safety and immunogenicity of the resultant conjugates were confirmed in adults and 4- to 7-year-old children. Succinylation of the carrier protein improved the immunogenicity of Shigella conjugates in mice and increased their yield. Based on these results, a clinical trial of O-SP conjugates of Shigella sonnei and Shigella flexneri 2a bound to succinylated mutant Pseudomonas aeruginosa exotoxin A (rEPAsucc) or native or succinylated Corynebacterium diphtheriae toxin mutant (CRM9 or CRM9succ) was conducted in healthy adults. The conjugates were safe and immunogenic. S. sonnei-CRM9, S. sonnei-CRM9succ, and S. sonnei-rEPAsucc elicited significant rises of geometric mean (GM) IgG anti-LPS within 1 week of injection (P < 0.001). At 26 weeks, the GM anti-LPS levels elicited by these three conjugates were similar and higher than their prevaccination levels (P < 0.0001). GM IgG anti-LPS levels elicited by S. flexneri 2a-rEPAsucc were significantly higher than those elicited by S. flexneri 2a-rCRM9succ at all intervals after injection. At 26 weeks, the levels of IgG anti-LPS in vaccinees were higher than their prevaccination levels (P < 0.0001). The serum antibody responses were specific, as there was no significant rise of anti-LPS to the heterologous O-SP in any vaccinee. Both conjugates elicited statistically significant rises of serum antibodies to the injected carrier protein. At 6 months, these five Shigella conjugates elicited higher fold rises than similar conjugates (D. N. Taylor et al., Infect. Immun. 61:3678-3687, 1993). Based on these data, we chose S. sonnei-CRM9 and S. flexneri 2a-rEPAsucc for evaluation in children.

Cytoplasmic cleavage in living zoosporangia of Allomyces macrogynus.

We have used the vital fluorescent dye, FM4-64, as a marker of membrane development during zoospore formation in living zoosporangia of Allomyces macrogynus. Membrane development was visualized and documented using standard epifluorescence and laser scanning confocal microscopy. Video-enhanced light microscopy and transmission electron microscopy, using cryopreparation methods, were also employed in this study. In the first 10-12 min after the induction of zoospore formation, only the plasma membrane labelled with FM4-64. During this time, nuclei were strictly located in the cortical cytoplasm with their associated centrosomes positioned immediately adjacent to the plasma membrane (Lowry & Roberson, 1997). Between 12 and 20 min post-induction, increased fluorescence appeared along regions of the plasma membrane adjacent to the nuclei. From these sites, membranes (i.e. cleavage elements) extended laterally within the cortex and then, in conjunction with nuclear migration, rapidly elongated into the sporangial cytoplasm. By 25-35 min post-induction, cleavage elements had ramified throughout the cytoplasm forming a complex, interconnected membranous network. Transmission electron microscopy revealed that cleavage elements were paired membrane sheets with a lumen consisting of an electron opaque, granular matrix. Cleavage elements developed into a highly ordered network by 35-40 min post-induction, which fully delimited zoospore initials into polyhedral-shaped cells. Zoospore discharge occurred between 40 and 50 min post-induction. Our results have shown that cleavage elements undergo four stages of development during zoospore formation in A. macrogynus: (i) development of membrane initials, (ii) cortical extension, (iii) cytoplasmic elongation and ramification and (iv) zoospore initial delimitation.

Microtubules Are required for motility and positioning of vesicles and mitochondria in hyphal tip cells of Allomyces macrogynus.

We have used video-enhanced light microscopy and digital image processing to characterize the intracellular motility and positioning of vesicles ( approximately 1-microm diameter) and mitochondria in growing hyphal tip cells of Allomyces macrogynus. These observations were coupled with cytoskeletal inhibitory experiments to define the roles of the microtubule and actin cytoskeletons in organelle translocation and positioning. Vesicles and mitochondria were abundant in apical and subapical hypha regions. Vesicles traveled along paths that were parallel to the longitudinal axis of the cell. Anterograde (i.e., toward the hyphal apex) and retrograde (i.e., away from the hyphal apex) movements of vesicles occurred at average rates of 4.0 and 2.2 microm/s, respectively. Bidirectional travel of vesicles along common paths was noted in the cortical cytoplasm. Mitochondria were aligned mostly parallel to the long axis of the hypha, except those extending into the hyphal apex, which were oriented toward the Spitzenkörper. In regions of the subapical hypha mitochondria were often restricted to the cortical cytoplasm and nuclei occupied the central cytoplasmic region. Mitochondria displayed rapid anterograde movements reaching speeds of 3.0 microm/s, but primarily maintained a constant position relative to either the advancing cytoplasm or the lateral cell wall. Cytoskeletal disruption experiments showed that the positioning of mitochondria and motility of vesicles and mitochondria were microtubule-based and suggested that the actin cytoskeleton played uncertain roles.

A neomorphic syntaxin mutation blocks volatile-anesthetic action in Caenorhabditis elegans.

The molecular mechanisms underlying general anesthesia are unknown. For volatile general anesthetics (VAs), indirect evidence for both lipid and protein targets has been found. However, no in vivo data have implicated clearly any particular lipid or protein in the control of sensitivity to clinical concentrations of VAs. Genetics provides one approach toward identifying these mechanisms, but genes strongly regulating sensitivity to clinical concentrations of VAs have not been identified. By screening existing mutants of the nematode Caenorhabditis elegans, we found that a mutation in the neuronal syntaxin gene dominantly conferred resistance to the VAs isoflurane and halothane. By contrast, other mutations in syntaxin and in the syntaxin-binding proteins synaptobrevin and SNAP-25 produced VA hypersensitivity. The syntaxin allelic variation was striking, particularly for isoflurane, where a 33-fold range of sensitivities was seen. Both the resistant and hypersensitive mutations decrease synaptic transmission; thus, the indirect effect of reducing neurotransmission does not explain the VA resistance. As assessed by pharmacological criteria, halothane and isoflurane themselves reduced cholinergic transmission, and the presynaptic anesthetic effect was blocked by the resistant syntaxin mutation. A single gene mutation conferring high-level resistance to VAs is inconsistent with nonspecific membrane-perturbation theories of anesthesia. The genetic and pharmacological data suggest that the resistant syntaxin mutant directly blocks VA binding to or efficacy against presynaptic targets that mediate anesthetic behavioral effects. Syntaxin and syntaxin-binding proteins are candidate anesthetic targets.

Regulation of apolipoprotein secretion by biliary lipids in newborn swine intestinal epithelial cells.

Biliary lipids, composed of bile acids, cholesterol, and phosphatidylcholine, are a major source of luminal lipid in the small intestine. In the present study in a newborn swine intestinal epithelial cell line (IPEC-1), taurocholate and phosphatidylcholine were found to have no effect on apolipoprotein B (apo B) secretion but did significantly increase the basolateral secretion of apo A-I. This regulation of apo A-I secretion occurred at the pretranslational level for taurocholate and at the posttranslational level for phosphatidylcholine. The regulation of apo A-I secretion by phosphatidylcholine did not involve changes in apo A-I degradation and may involve mobilization of a preformed pool of apo A-I. Cholesterol, whether solubilized with taurocholate or phosphatidylcholine, had no effect on the secretion of either apo B or apo A-I. However, when taurocholate, phosphatidylcholine, and cholesterol were combined, apo B secretion was decreased, and the increase in apo A-I secretion noted with taurocholate and phosphatidylcholine alone was ablated. Another primary bile acid, taurochenodeoxycholate, was found to decrease apo B secretion but had no effect on apo A-I secretion. However, the significance of this effect is uncertain, since this bile acid caused significant cellular membrane injury, as evidenced by increased apical medium lactate dehydrogenase activity. Phosphatidylcholine, but not taurocholate, dramatically increased the basolateral secretion of radiolabeled phospholipid with a modest increase in cellular triglyceride radiolabeling. Furthermore, this effect of phosphatidylcholine on lipid synthesis did not require significant hydrolysis or uptake of the phosphatidylcholine molecule. Studies using radiolabeled taurocholate did not demonstrate active transport of taurocholate by these cells.

Oral arginine-lysine does not increase growth hormone or insulin-like growth factor-I in old men.

BACKGROUND: Older adults tend to have reduced growth hormone (GH) secretion and insulin-like growth factor I (IGF-I) levels as well as changes in body composition which are partially reversed by GH injections. Arginine stimulates GH release, and lysine may amplify this response. We investigated whether oral arginine/lysine could be used to increase basal IGF-I and GH levels in non-obese old men (age 69 +/- 5 years; mean +/- SD) to values similar to those of untreated young men (age 26 +/- 4 years). METHODS: Two groups of 8 healthy old men were treated with 3 g of arginine plus 3 g of lysine or with placebo capsules twice daily for 14 days. Before and on day 14 of each treatment GH levels were determined in blood samples taken at 20-minute intervals from 2000-0800 h, IGF-I was measured at 0800 h, and a 1 microgram/kg GHRH stimulation test was done. RESULTS: At baseline, mean GH peak amplitude (p < .02) and serum IGF-I (p < .0001) were lower, whereas GHRH responses were similar, in old vs young men. Arginine/lysine did not significantly alter spontaneous or GHRH-stimulated GH levels, or serum IGF-I. Arginine absorption was age-independent. The correlation (p < .005) between measured increments in serum arginine and increases in serum GH after a single dose of arginine/lysine was similar in old and young groups. CONCLUSIONS: Our data suggest that oral arginine/lysine is not a practical means of chronically enhancing GH secretion in old men.

Continuous subcutaneous infusions of growth hormone (GH) releasing hormone 1-44 for 14 days increase GH and insulin-like growth factor-I levels in old men.

Twice daily sc injections of GHRH increase serum GH and IGF-I levels in healthy old men to values like those of untreated young men by producing high amplitude GH peaks after the injections. In the present study, we measured baseline insulin-like growth factor-I (IGF-I) 24-h profiles of GH release, and responses to GHRH stimulation tests in healthy young and old men. Old men were then given, in random order, 1 and 2 mg continuous sc GHRH 1-44 infusions daily for 14 days with an intervening 14-day treatment-free period. The study protocol was repeated on day 14 of each treatment. At baseline, mean duration of GH peaks (P < 0.005) and IGF-I levels (P < 0.0001) were lower in old men. Both doses increased (vs. old basal) mean 24-h GH, integrated area under the GH curve, and GH peak number (P < 0.05), as well as serum IGF-I (P < 0.001). Interpeak GH levels also increased during low (P < 0.01) and high (P < 0.05) dose treatment. Significant increases in mean GH and integrated area under the GH curve occurred only during the day (0800-2000 h) during both low (P < 0.01) and high (P < 0.05) dose treatment. Treatment also decreased nocturnal peak GH amplitude and duration. GH responses to GHRH stimulation tests did not differ with age at baseline, or in old men after treatment. Thus, continuous, short-term sc administration of GHRH to healthy old men restores subnormal GH secretion and IGF-I levels by increasing GH peak frequency and interpeak secretion, particularly during the day.

Growth hormone (GH)-releasing hormone-(1-29) twice daily reverses the decreased GH and insulin-like growth factor-I levels in old men.

Aging is associated with decreased GH and insulin-like growth factor-I (IGF-I) levels and lean body mass, and increased body fat. Recombinant human GH treatment of old men partially reverses body composition changes. Administration of GH-releasing hormone (GHRH) to GH-deficient children and young adults increases GH and IGF-I levels while preserving physiological GH release. We investigated whether GHRH injections restore GH and IGF-I levels in old men to the levels in young men. Healthy young (n = 9; 26.2 +/- 4.1 yr; mean +/- SD) and old (n = 10; 68.0 +/- 6.2 yr) nonobese men underwent baseline blood sampling for measurements of IGF-I and 24-h profiles of GH release, followed by iv bolus GHRH stimulation tests. Old men then took, randomly, both low (0.5 mg) and high (1 mg) dose GHRH-(1-29) sc injections twice daily for 14 days, with an intervening 14-day nontreatment period. The study protocol was repeated on day 14 of each treatment. At baseline, the mean peak duration of spontaneous GH release (P less than 0.005) and IGF-I levels (P less than 0.0001) were lower in the old men. GHRH treatment evoked dose-related increases in all parameters, with significant differences (vs. old basal values) in mean 24-h GH (P less than 0.001), area under peaks (P less than 0.001), peak amplitude (P less than 0.05), and IGF-I (P less than 0.005) only at the high dose. After high dose treatment, there were no significant differences in these parameters between age groups. Peak and integrated responses to iv GHRH stimulation tests did not differ between young and old men either before or during GHRH treatment. Baseline serum levels of both testosterone (P less than 0.01) and phosphate (P less than 0.05) were lower in the older men. Phosphate levels increased (P less than 0.05) during GHRH treatment. GHRH treatment did not affect fasting glucose, urinary C-peptide, blood pressure, or chemistry and hematology profiles. Thus, short term sc administration of GHRH to healthy old men reverses age-related decreases in GH and IGF-I, suggesting that prolonged treatment could improve age-related alterations in body composition.

Toxin-neutralizing antibodies in patients with pertussis, as determined by an assay using Chinese hamster ovary cells.

Antibodies to pertussis toxin were measured by a neutralization test (Chinese hamster ovary cell assay) in paired serum samples from 122 patients with whooping cough. The results were compared with previously published titers of IgG, IgM, and IgA measured by ELISA. Significant (fourfold or more) increases in neutralizing antibodies developed in 69 patients. Sixty-eight of them also had significant increases in at least one antibody class as measured by ELISA. Seventeen patients had significant increases in at least one antibody class without increases in neutralizing antibodies. All 163 serum samples with neutralizing antibodies also had detectable IgG, whereas 14 samples with IgG and 18 with IgM were negative by neutralization test. Thus, toxin-neutralizing serum antibodies develop in most patients with pertussis. Our ELISA, however, was more sensitive in detecting low levels of antibodies and in demonstrating significant increases between acute- and convalescent-phase serum samples.

Clinical, metabolic, and antibody responses of adult volunteers to an investigational vaccine composed of pertussis toxin inactivated by hydrogen peroxide.

A toxoid vaccine, composed of purified pertussis toxin inactivated with H2O2 (NICHD-Ptxd), was developed on the basis of evidence that serum neutralizing antibodies (antitoxin) would confer immunity to pertussis. In vivo and in vitro assays of NICHD-Ptxd showed only trace or nondetectable levels of pyrogenic, adenosine diphosphate-ribosyltransferase, binding and pharmacologic activities. Nevertheless, about 40% of the antigenicity of pertussis toxin was retained. Adult volunteers were injected, two times 6 weeks apart, with either 10 (n = 21), 50 (n = 25), or 75 (n = 30) micrograms/dose of one lot, Ptx-06, adsorbed onto AI(OH)3. Neither fever nor changes in the levels of leukocytes, lymphocytes, fasting blood glucose, or insulin were observed in the volunteers. The optimal immunizing dose, 50 micrograms, induced levels of antitoxin (geometric mean (GM) 302 U) comparable to those found in eight adults convalescent from pertussis (GM 269 U) and greater than those found in 18-month-old children after their fourth dose of diphtheria and tetanus toxoids and pertussis vaccine (GM 20.0 U, p less than 0.001). These data indicate that NICHD-Ptxd is safe and immunogenic in adults, and they justify its evaluation in infants and children.

Direct and maternal genetic effects on preweaning characters of Brahman, Hereford and Brahman-Hereford crossbred cattle.

Birth weight, preweaning gain and weaning weight (adjusted 180-d weight) data, collected at McGregor, Texas, were analyzed for genetic differences. Breedtypes represented in the data were Brahman, Hereford and various Brahman-Hereford crosses. Preweaning gain was calculated as adjusted 180-d weight less birth weight. All statistical models included effects of dam age, year, season and sex. Analyses were performed using a breedtype model and a regression model that redefined breedtype as direct additive, direct heterotic, maternal additive and maternal heterotic effects. Brahman dams produced calves with lightest birth weights. Brahman-sired calves were heaviest at birth compared with those by other sire breedtypes. The estimated Brahman direct additive effect on birth weight was 4.6 kg greater than Hereford. The Brahman maternal additive effect was 7.5 kg less than Hereford. Direct and maternal heterotic effects on birth weight were 2.2 and .6 kg, respectively. Calves from F1 dams had larger preweaning gains than those of the other breedtypes. The Brahman direct additive effect on preweaning gain was 17.7 kg less than Hereford and the Brahman maternal additive effect was 20 kg greater than Hereford. Direct and maternal heterotic effects on preweaning gain were 19.6 and 19.5 kg, respectively. Results of weaning weight analyses were similar to preweaning gain analyses. The largest effects on weaning weight were direct and maternal heterosis, which were 21.6 and 19.8 kg, respectively.

Pharmacological modification of corneal endothelial intracellular pH, intracellular electrical potential difference, and corneal swelling and deswelling rates.

Rabbit corneal endothelial intracellular pH and electrical potential difference were measured using radioactive tracer techniques. A variety of drugs and chemicals were used to modify specific aspects of cell membranes in order to determine, in this steady state system, whether a close relationship existed between intracellular pH and potential. Corneal swelling and deswelling rates were also determined in the presence and absence of the drugs. Longer times of immersion of tissue with drugs (3 hours) tended to provide more support for a link between intracellular pH and the inability of the cornea to maintain a constant thickness. Drugs that altered corneal thickness without altering either intracellular pH or potential difference presumably acted on extracellular pathways. No conclusive evidence was obtained for a direct link between intracellular pH and the intracellular electrical potential difference of the endothelial cell.