Effects of speaker variability and phoneme location on speech perception.

The effects of variation in a speaker's voice and temporal phoneme location were assessed through a series of speeded classification experiments. Listeners monitored speech syllables for target consonants or vowels. The results showed that speaker variability and phoneme-location variability had detrimental effects on classification latencies for target sounds. In addition, an interaction between variables showed that the speaker variability effect was obtained only when temporal phoneme location was fixed across trials. A subadditive decrement in latencies produced by the interaction of the two variables was also obtained, suggesting that perceptual loads may not affect perceptual adjustments to a speaker's voice in the same way that memory loads do.

Octreotide enhances positive calcium balance in Duchenne muscular dystrophy.

Although receptors for somatostatin are found in bone cells, the effect of somatostatin analogs on calcium metabolism is unknown. The authors studied, in a metabolic ward, the effect of octreotide (a long-acting somatostatin analog) and a placebo in two 6-day calcium balance periods in 8 children with Duchenne muscular dystrophy. As expected, octreotide (2 micrograms/kg, subcutaneously, every 8 hours) reduced serum growth hormone and somatomedin (IGF-1) to levels found in growth hormone deficiency. Octreotide enhanced calcium retention by 30% (96 mg daily [P < 0.04]) in 7 boys for whom complete data (diet, urine, and fecal calcium) were available. In 6 children with urinary calcium excretion (Uca) greater than 50 mg daily, octreotide markedly lowered Uca, from 114 +/- 23 mg daily to 61 +/- 9 mg daily (P < 0.03). Calcium retention occurred in patients with or without initial hypercalciuria, but the higher the basal Uca, the greater was the inhibition by octreotide (r = 0.79; P < 0.03). Inactive, nonambulatory patients had a more pronounced response of Uca to octreotide (P < 0.02). Octreotide caused a mild, nonsignificant reduction in fecal calcium, with no major changes in serum calcium, phosphorus, parathyroid hormone, urinary excretion of sodium and potassium, or in creatinine clearance. Based on the current observations and the presence of receptors for somatostatin in bone cells, this hormone may have, at least on a short-term basis, an anabolic effect on calcium, perhaps favoring its deposition in bone.

Postprandial lipoprotein responses in hypertriglyceridemic subjects with and without cardiovascular disease.

Three groups of age- and weight-matched men (aged 40 to 70 years) without diabetes were studied: controls (n = 10), plasma triglycerides (TG) less than 180 mg/dL and no cardiovascular disease (CVD); HTG-CVD (n = 11), hypertriglyceridemic (HTG) (TG > 240 mg/dL) without CVD; and HTG+CVD (n = 10), HTG (TG > 240 mg/dL) with documented CVD. HTG+CVD subjects had higher fasting and post-oral glucose tolerance test insulin levels than the other two groups, respectively. Very-low-density lipoprotein (VLDL)+chylomicrons (CMs), intermediate-density lipoprotein (IDL), low-density lipoprotein (LDL), and three high-density lipoprotein (HDL) subfractions (HDL-L, HDL-M, and HDL-D, from least to most dense) were isolated by gradient ultracentrifugation. Fasting lipoproteins were similar in HTG groups, except for higher VLDL lipid to apolipoprotein (apo) B ratios (P < .04) in the HTG+CVD group. Subjects were fed a high-fat mixed meal, and lipoprotein composition was determined at 3, 6, 9, and 12 hours postprandially. Postprandial responses of the core lipids (TG and cholesterol esters [CE]) in all of the lipoprotein subfractions were similar in the two HTG groups at each time point. However, both controls and HTG-CVD subjects had increases in HDL-M phospholipid (PL) at 9 and 12 hours with no change in HDL-D PL. The HTG+CVD group, on the other hand, had no increase in HDL-M PL and had a substantial reduction in HDL-D PL. These changes resulted in significant increases in HDL-M and HDL-D PL to apo A-I ratios in both controls and HTG-CVD subjects between 6 and 12 hours, whereas there was no increase seen in the HTG+CVD group. The HTG-CVD group also had a significantly greater increase in the VLDL+CM PL to apo B ratio (P = .038) at 3 hours than the HTG+CVD group. This diminished amount of surface lipid per VLDL particle may account for the late decrease in the HDL-D PL to apo A-I ratio seen in HTG+CVD patients. There were no other postprandial lipid or apolipoprotein differences between the two HTG groups. We conclude therefore that the major postprandial lipoprotein abnormality in these HTG+CVD patients was a failure to increase the PL content per particle in VLDL+CM, HDL-M, and HDL-D. This abnormality could prevent the usual increase in reverse cholesterol transport seen in postprandial plasma and therefore contribute to their increased incidence of CVD. The greater insulin resistance seen in these patients also appears to contribute significantly to their CVD.

Identification of Mengo virus T helper cell epitopes.

To identify Mengo virus-specific T cell epitopes in mice (the natural host for the virus), lymph node cells were obtained from BALB/c (H-2d) mice, previously immunized with u.v.-inactivated virus, and stimulated in vitro with each of 116 overlapping peptides (10 to 18 residues long) covering the entire capsid coding region (834 amino acids). T cell epitopes were defined on the basis of specific peptide-induced lymphocyte proliferation. Where proliferation occurred, immunological characterization showed that it was the CD4+ T helper (Th) cell subpopulation that was responsible for the Mengo virus-specific response. Surprisingly, no Mengo virus Th cell epitopes were found in capsid protein VP1 or VP4. Six peptides in VP2 (residues 1 to 15, 99 to 108, 118 to 132, 133 to 147, 227 to 236 and 247 to 256) identified the positions of separate Th cell epitopes, and two overlapping peptides (residues 173 to 182 and 178 to 192) defined an additional Th cell immunogenic sequence. Three individual peptides in VP3 (residues 46 to 58, 136 to 150 and 198 to 212) and two overlapping peptides (residues 1 to 15 and 11 to 20) also represent Th cell epitopes. Similar assays with C57BL/6 (H-2b) and SJL/J (H-2s) mice showed that the pattern of recognition of these peptides was H-2 restricted. Each of the previously identified sites of B cell antigenicity in VP2 and VP3 are associated with one Th epitope. Comparison of the experimentally determined Th epitopes with potential T cell epitopes identified by several predictive strategies revealed only a low correlation between authentic and predicted epitopes.

Neutralizing antibody to Mengo virus, induced by synthetic peptides.

A peptide from the carboxyl-terminal region of the Mengo virus capsid protein VP1, representing residues 259 to 277, can induce serum neutralizing (SN) antibodies in both the mouse and guinea-pig. This peptide, termed F164, also induces high levels of protective neutralizing antibodies in mice subsequent to immunization; 87 to 100% of mice are refractory to the effects of an intraperitoneal challenge of 100 LD50 of Mengo virus. The mouse model discussed herein will prove useful for studying the immune response to Mengo virus and evaluating the immunogenicity of individual viral components.

Maintenance of normal circulating levels of delta 4-androstenedione and dehydroepiandrosterone in simple obesity despite increased metabolic clearance rates: evidence for a servo-control mechanism.

To study the effect of obesity on the metabolism of adrenal androgens not bound to testosterone-estradiol-binding globulin, the MCRs of delta 4-androstenedione (A) and dehydroepiandrosterone (DHEA) were determined using constant infusion of unlabeled steroids to steady state in 8 normal weight and 19 obese nonhirsute eumenorrheic women. The blood production rates (PR) were calculated as the product of the MCR and the 24-h integrated serum concentrations (IC). The mean MCR and PR of A and DHEA were significantly higher in the obese women than in the normal weight women. There was, however, no difference in the mean IC of each androgen in the 2 groups. The MCR and PR of A and DHEA were each correlated with the body mass index (BMI; kilograms per m2). The MCR and PR of A and the MCR of DHEA were also correlated with the ratio of waist circumference to hip circumference (WHR). However, the PR of DHEA was not correlated with WHR. There was no correlation between the IC of either androgen and BMI or WHR. However, partial correlation analysis revealed that correction of the BMI for WHR resulted in a significant negative correlation between BMI and IC of A. We conclude that the MCR and PR of A and DHEA were increased in obese nonhirsute eumenorrheic women; there was a strong correlation between BMI and the MCR and PR of A and DHEA; upper segment obesity, as measured by WHR, was correlated with the MCR and PR of A and the MCR of DHEA, but not with the PR of DHEA; and circulating DHEA and A were maintained at normal levels in the obese eumenorrheic women despite an increase in the MCR, which suggests that a servo-mechanism is operative which registers the body size and adjusts the PR according to the MCR.

Immune response to uncoupled peptides of foot-and-mouth disease virus.

Uncoupled synthetic peptide representing the sequence of amino acids 141-160 of foot-and-mouth disease virus (FMDV) protein VP1 induced a virus-neutralizing antibody response in guinea-pigs. This response required incomplete Freund's adjuvant (IFA) for the primary inoculation and was dependent on the presence of an added cysteine residue with an unblocked sulphydryl group at the carboxy-terminus. Secondary immunization could be carried out in the absence of adjuvant. A study of the relative activities of nested sets of uncoupled peptides from 150-160 to 135-160 and 141-160 to 141-155 indicated that amino acids 146-156 were critical for the induction of virus-neutralizing antibodies and that extension to 137-160 further improved this response. Results of in vitro proliferation studies demonstrated that the carboxy-terminal residues on this peptide may form a T-cell epitope. The significance of these observations in the broader context of synthetic peptide vaccines is discussed.

Reduction of hyperinsulinemia and insulin resistance by opiate receptor blockade in the polycystic ovary syndrome with acanthosis nigricans.

We previously reported that circulating beta-endorphin levels are increased in obese hirsute women and that plasma immunoreactive insulin (IRI) levels are increased in proportion to the degree of hyperandrogenism in women with the polycystic ovary (PCO) syndrome. We, therefore, tested the hypothesis that endogenous opiates are at least partially responsible for the hyperinsulinemia and insulin resistance in this syndrome. In the first study, acute naloxone administration significantly reduced the plasma IRI response and IRI/glucose ratio in three euglycemic obese women with PCO and acanthosis nigricans (AN) and marked insulin resistance, but did not alter the glucose response. Naloxone had no effect on these parameters in the normal weight control subjects. In the second study, nalmefene, a new, orally active opiate antagonist, reduced IRI and the IRI/glucose ratio in four women with PCO-AN and marked hyperinsulinemia in a randomized, double blind, crossover protocol. We conclude that endogenous opiates are at least partially responsible for the hyperinsulinemia and insulin resistance in PCO-AN.

Increased serum concentrations of 1,25(OH)2 vitamin D in children with fasting hypercalciuria.

Inappropriately elevated concentrations of 1,25(OH)2 vitamin D in serum appear to be responsible for excessive gastrointestinal absorption of dietary calcium in patients with absorptive hypercalciuria. We have examined serum 1,25(OH)2 vitamin D concentrations in another group of children with hypercalciuria in whom urinary calcium excretion was excessive after an overnight fast. Eleven children with idiopathic fasting hypercalciuria (IH) (urinary calcium excretion greater than 4 mg/kg/24 hr and fasting urinary calcium/urinary creatinine ratio greater than 0.21) and seven healthy children were observed while they were eating a diet containing 1 gm calcium per day. Fasting serum 1,25(OH)2 vitamin D concentrations were elevated in children with IH compared with control values (35.3 +/- 3.2 vs 21 +/- 2 pg/ml, P = 0.003), whereas fasting serum parathyroid hormone, 25-OH vitamin D, phosphorus, and ionized calcium concentrations were similar in the two groups. These data suggest that disordered 1,25(OH)2 vitamin D metabolism occurs in children with fasting IH. Absorptive and fasting IH may represent a spectrum of a single disorder characterized by excessive urinary calcium excretion and inappropriately elevated serum concentrations of 1,25(OH)2 vitamin D.

Localization of neutralizing regions of the envelope gene of feline leukemia virus by using anti-synthetic peptide antibodies.

We synthesized 27 synthetic peptides corresponding to approximately 80% of the sequences encoding gp70 and p15E of Gardner-Arnstein feline leukemia virus (FeLV) subtype B. The peptides were conjugated to keyhole limpet hemocyanin and injected into rabbits for preparation of antipeptide antisera. These sera were then tested for their ability to neutralize a broad range of FeLV isolates in vitro. Eight peptides elicited neutralizing responses against subtype B isolates. Five of these peptides corresponded to sequences of gp70 and three to p15E. The ability of these antipeptide antisera to neutralize FeLV subtypes A and C varied. In certain circumstances, failure to neutralize a particular isolate corresponded to sequence changes within the corresponding peptide region. However, four antibodies which preferentially neutralized the subtype B viruses were directed to epitopes in common with Sarma subtype C virus. These results suggest that distal changes in certain subtypes (possibly glycosylation differences) alter the availability of certain epitopes in one virus isolate relative to another. We prepared a "nest" of overlapping peptides corresponding to one of the neutralizing regions of gp70 and performed slot blot analyses with both antipeptide antibodies and a monoclonal antibody which recognized this epitope. We were able to define a five-amino-acid sequence required for reactivity. Comparisons were made between an anti-synthetic peptide antibody and a monoclonal antibody reactive to this epitope for the ability to bind both peptide and virus, as well as to neutralize virus in vitro. Both the anti-synthetic peptide and the monoclonal antibodies bound peptide and virus to high titers. However, the monoclonal antibody had a 4-fold-higher titer against virus and a 10-fold-higher neutralizing titer than did the anti-synthetic peptide antibody. Competition assays were performed with these two antibodies adjusted to equivalent antivirus titers against intact virions affixed to tissue culture plates. The monoclonal antibody had a greater ability to compete for virus binding, which suggested that differences in neutralizing titers may relate to the relative affinities of these antisera for the peptide conformation in the native structure.

A new generation of vaccines.

New technology in recombinant DNA, gene sequencing, and peptide synthesis will make possible a new generation of vaccines. These new vaccines will be safer, more stable, and can be produced in a more predictable manner than the present vaccines. This should lead to a wider use of vaccines and a greater control of infectious diseases.

The genome-linked proteins of aphthoviruses: specific immunoprecipitation of the three species detected on virus RNA and identification of possible precursors.

Synthetic peptides have been made corresponding to the C-terminal portion of each of the three presumptive genome-linked proteins (VPgs) of foot-and-mouth disease virus type A10. Antisera against each of these peptides efficiently precipitated only the homologous VPg, and the reactions were inhibited by prior absorption with homologous, but not heterologous synthetic peptide. The peptide antisera precipitated a number of proteins from infected cell extracts with mol. wt. of 100, 84, 56, 36, 27, 25 and 20, all X 10(3); all these reactions were inhibited by absorption with homologous peptide, indicating that they were probable precursors of VPg. The relationship between these proteins is at present unclear.

Relative sensitivity and responsivity of serum cortisol and two adrenal androgens to alpha-adrenocorticotropin-(1-24) in normal and obese, nonhirsute, eumenorrheic women.

The alpha ACTH-(1-24) threshold dose and the response slope were determined for cortisol (F), delta 4-androstenedione (A), and dehydroepiandrosterone (DHEA) in 10 normal and 16 obese eumenorrheic nonhirsute women matched for age. Each woman received 1 mg dexamethasone at 2300 h and again at 0700 h the next morning. At 0700 h, a continuous alpha ACTH-(1-24) infusion was begun at an initial dose of 30 ng/1.5 m2 body surface area X hr. The ACTH infusion rate was doubled every hour for 5 consecutive h to a maximum dose of 480 ng/1.5 m2 X h. Blood samples were collected for steroid assays before the infusion and at the end of each hour. The ACTH threshold dose was defined as the dose that produced a steroid response significantly above the basal level. The ACTH threshold dose for serum F and DHEA stimulation was not different between the groups, but the threshold dose for A was significantly lower in the obese women. Basal and stimulated serum DHEA to F ratios were significantly higher in the obese women. In both groups, the mean F response slope was significantly higher than that for DHEA, which in turn, was significantly higher than that for A. The mean DHEA response slope was significantly greater in the obese women. The F and A response slopes were not different between the groups. We conclude that the relative responsivity of the steroids to ACTH was the same in both groups: F greater than DHEA greater than A; in the obese women, the ACTH threshold dose for F stimulation was lower (greater sensitivity) than for DHEA or A stimulation; and in the obese women, the ACTH threshold dose for A was significantly lower (increased sensitivity) and the slope of the DHEA response to ACTH was steeper (greater responsivity) than in normal women.

Immunological priming with synthetic peptides of foot-and-mouth disease virus.

A sub-immunizing dose of a synthetic peptide corresponding to the amino acids 141 to 160 region of protein VP1 from foot-and-mouth disease virus (FMDV), serotype O1, coupled to keyhole limpet haemocyanin (141-160KLH) has been shown to prime the immune system of guinea-pigs for an FMDV serotype-specific neutralizing antibody response to a second sub-immunizing dose of the same peptide. Optimal priming required an interval of 42 days between the priming dose and the booster dose. No priming was observed in the absence of adjuvant. The secondary response was not restricted by the carrier since animals primed with 141-160KLH could be boosted with uncoupled 141-160 or 141-160 coupled to tetanus toxoid. It has also been shown that uncoupled peptide 141-160 will prime for a neutralizing antibody response when it is incorporated into a relatively non-immunogenic carrier such as small unilamellar liposomes. These results indicate that the 141-160 peptide of FMDV, as well as containing an important neutralizing antibody site, can initiate its own T-helper cell response.

Synthetic peptides from four separate regions of the poliovirus type 1 capsid protein VP1 induce neutralizing antibodies.

Peptides from different regions of the poliovirus type 1 capsid protein VP1 were synthesized. Antibodies raised against these peptides in rabbits and rats recognized the cognate peptides and denatured VP1. Peptides from four regions of VP1 generated antisera with neutralizing titers specifically against poliovirus type 1. Antisera against all other regions of VP1 failed to neutralize virus infectivity, although some of the antisera clearly bound to native virions. Thus, the neutralizing determinants on VP1 reside in specific noncontiguous regions of the protein and can be defined by specific peptides from these regions.

Recent trends in veterinary vaccines.

Recent developments in the fields of molecular biology and immunology have made possible the synthesis of proteins and peptides that closely mimic antigenic sites. The potential for developing vaccines from this technology is reviewed.

Chemical basis of antigenic variation in foot-and-mouth disease virus.

One of the difficulties in controlling foot and mouth disease by vaccination is the occurrence of the virus as seven distinct serotypes because immunity conferred by vaccination against one serotype leaves the animals susceptible to infection by the other six. Moreover, the antigenic variation, even within a serotype, can be so great that immunity against the homologous strain of virus need not necessarily ensure protection against infection by other viruses within that serotype. Here we report the separation of three natural antigenic variants, distinguishable in cross-neutralization tests from an isolate of foot-and-mouth disease virus (FMDV). The serological differences could also be demonstrated by antisera elicited by synthetic peptides corresponding to residues 141-160 of the capsid polypeptide VP1, showing that this region contains a major immunogenic site of the virus. The results have practical implications for the choice of viruses for vaccine production.