Pediatric Emergency Medicine Fellowship survey: 1991-1992.

This study was designed to determine the general characteristics, training expectations, and career goals of those individuals entering Pediatric Emergency Medicine Fellowships in 1992 through the NRMP Pediatric Emergency Medicine Match. A 20-item questionnaire was developed and sent to those individuals who successfully obtained fellowship positions through the 1991-1992 Match. All of the respondents will have completed a formal pediatric residency program, and 90% will have completed their residency since 1990. Eighty-eight percent of the respondents have not completed formal postgraduate training other than a pediatric residency, and none of the respondents were from training programs in emergency medicine. Ninety percent of the respondents are planning on two years of training, while 10% are either entering a three-year program or planning an optional third year. Ninety-four percent of the individuals who responded had not applied for Pediatric Emergency Medicine Fellowships in the past, but, while many of the individuals were concerned about obtaining a position, only 6% applied for a fellowship in an alternative field. If the respondents had not obtained positions this year, 79% felt that they would have reapplied next year. When asked why they are pursuing a Pediatric Emergency Medicine Fellowship, 85% listed opportunities in clinical medicine as their primary reason, while 10% claimed that research opportunity was the most important factor. When their fellowships are completed, 77% hope to practice at a university-based children's hospital, and 10% hope to practice at a private children's hospital.(ABSTRACT TRUNCATED AT 250 WORDS)


Plastic bronchitis: an unusual cause of respiratory distress in children.

This case illustrates an unusual cause of respiratory distress in the pediatric population. A high degree of suspicion is necessary to make the diagnosis of plastic bronchitis. Wheezing and cough will lead to the diagnosis of reactive airway disease and/or foreign body aspiration. Chest radiographs may yield additional information, but the diagnosis is made by bronchoscopy and removal of the casts. Any child with severe respiratory distress refractory to aggressive conventional medical therapy and with a history or radiograph suggestive of plastic bronchitis should be considered a candidate for bronchoscopy. As clinicians, we must always remember the dictum, "All that wheezes is not asthma."


Anomalous left coronary artery masquerading as infantile bronchiolitis.

Four infants less than six months of age with anomalous left coronary artery from the pulmonary artery who present with symptoms of wheezing are described. All had cardiomegaly on chest radiographs and because of wheezing received beta-agonist agents (albuterol alone or with epinephrine). One developed cardiopulmonary collapse secondary to supraventricular tachycardia after administration of these agents. The literature is reviewed for utility of chest radiographs in infants presenting with wheezing and for the efficacy of beta-adrenergic agents in infants less than six months of age. The authors suggest that physicians have a low threshold for obtaining a chest radiograph prior to treating a first-time wheezing infant less than six months of age with a beta-agonist agent.


Thermal epiglottitis after swallowing hot tea.

Acute infectious epiglottitis in children is a well-recognized clinical entity. We report the development of acute thermal epiglottitis after ingestion of hot tea by a three-year-old patient. Clinical and radiographic findings in our patient and others reported in the literature resemble acute infectious epiglottitis. In all cases of burns around the mouth, the possibility of intraoral and respiratory damage must be considered. Because of the high risk of upper airway obstruction, children in whom thermal epiglottitis is suspected should be observed in the intensive care unit and have appropriate airway management.


Testicular torsion versus epididymitis: a diagnostic challenge.

Unlike the patient who presents with a potentially acute abdomen, the child or adolescent with a potentially acute scrotum cannot simply be observed. If testicular torsion is present, the testicle must be detorted and orchiopexy performed as soon as possible for fertility to be maintained. Torsion of the appendix testis, however, can usually be managed without surgery. Since the presentations of epididymitis and testicular torsion overlap, it is sometimes difficult to rapidly make the correct diagnosis. Early genitourinary consultation is appropriate in this setting. Any patient in whom testicular torsion is strongly considered should undergo immediate exploratory surgery without diagnostic studies. If the findings overlap, immediate testicular radionuclide scanning should be arranged; alternatively, with experience, Doppler sonography can be carried out. If these radiographic studies cannot be arranged and interpreted within one to two hours, scrotal exploration should be performed. Any patient with an acute scrotal complaint and a negative scan should receive daily follow-up until the symptoms subside. Although our adolescent patient did well, his acute presentation and findings should have warranted immediate exploration. It is only through this aggressive approach that we can continue to increase testicular salvage rates.


Injuries associated with child safety seat misuse.

The charts of 370 children under the age of two years who presented to a pediatric emergency department between September 1988 and August 1989 were reviewed. Twenty-seven patients (7% of the total) had injuries associated with child safety seat (CSS) misuse. Thirteen were infants and toddlers injured as motor vehicle occupants when improperly restrained--CSS harness not properly connected (8), use of an improper device (3), and CSS not anchored to the car seat (2). Fourteen were infants under one year of age who were injured falling in their CSS. Injuries included minor head trauma (17), linear skull fracture (5), concussion (1), femoral fracture (1), depressed skull fracture with epidural hematoma (1), cervical vertebral fracture (1), and intraventricular hemorrhage (1). Nine patients were hospitalized. Injuries associated with CSS misuse may be more common than previously recognized and can result in significant injury. Educational efforts should focus on correct usage.


Grunting respirations: chest or abdominal pathology?

A large percentage of misdiagnosed appendicitis cases occur during childhood. Misdiagnosed patients have increased morbidity and mortality from the diagnostic delay. The patients excused from an emergency facility who are ultimately shown to suffer from appendicitis have higher rates of perforation with attendant abscess formation, peritonitis, sepsis, and potential death. The patients with misdiagnosed appendicitis are young and likely to have atypical signs and symptoms. Grunting respirations incorrectly attributed to respiratory infection may serve as a pathway for a misdiagnosed case of appendicitis.


[Lysozyme activity in undiluted and diluted blood of patients treated with cyclosporin A with prednisone or azathioprine with prednisone after kidney transplantation].

It is a general opinion that the renal transplant patients may show the decrease of immunity which manifests by their increased susceptibility to bacterial, viral and fungal infections. One of the important non-specific defensive factors of the organism participating in the protection against infections is lysozyme. It is the purpose of the present paper to estimate the lysozyme activity in the un- and diluted serum in the renal transplant patients and to determine the possible mechanism of the observed deviations. 32 renal transplant patients (24 men and 8 women) aged of 16 up to 50 were investigated, being in the period of 1 until 4 years after transplantation. In the investigation the kind of immunosuppressive treatment used was considered which served the purpose of dividing the total group of patients into two subgroups: one treated with cyclosporine (12 patients) and the other treated with azathioprine (20 patients). The control group consisted of 20 healthy persons, matching by age, sex and living place the group investigated. In both groups investigated and control, were determined: lysozyme activity in the serum of the undiluted and diluted blood, total leukocyte number, the absolute number of peripheral blood neutrophils as well as their adherence to fibres was evaluated. In the renal transplant patients compared with the control group and in the separated subgroups, no statistically significant differences were found concerning leukocyte count, absolute neutrophil count and adherence of those cells.(ABSTRACT TRUNCATED AT 250 WORDS)


[Erythropoietin level in the blood of patients with diabetes mellitus type 2].

52 patients with diabetes mellitus type II (23 of them were treated by diabetic diet, 12 were on oral antidiabetic drugs and 17 were treated by insulin), and in 31 healthy subjects the serum concentrations of erythropoietin (EPO), ferritin, iron, TIBC and hematocrit value, haemoglobin concentration and erythrocytes were measured. In diabetic patients higher but in insulin treated patients significantly higher plasma erythropoietin concentrations were found than in healthy subjects. In diabetic patients the physiological negative correlation between plasma erythropoietin concentration, haematocrit value, haemoglobin concentration and erythrocytes respectively was absent. In diabetic patients plasma ferritin concentrations were significantly higher than in normals. patients with diabetes mellitus type II are characterized by elevated plasma erythropoietin and ferritin concentrations and dissociation of the physiological feedback between plasma erythropoietin and intensity of erythropoiesis.


[Inhibition of growth hormone (GH) response to GHRH in diabetes mellitus type 1 after blockade of the cholinergic system with pirenzepine].

The effect of pirenzepine, a peripheric cholinergic antagonist, on GHRH-induced GH secretion was studied in 10 type I diabetic subjects and 8 healthy controls. GH response was significantly greater in diabetics than in the healthy controls (p < 0.02) with peak values of 59.6 and 15.9 ng/ml, respectively. Pirenzepine 20 mg in given 15 min before GHRH inhibited GH response in both groups. In diabetics GH response calculated as area under curve (AUC) was inhibited from 3666 to 260 ng/ml. min (p < 0.001) and in the controls from 831 to 255 ng/ml. min (p < 0.05). The great efficacy of pirenzepine in inhibiting GH hypersecretion in type I diabetes may find a clinical application in future.


[Comparison of the effectiveness of budesonide and prednisone in the maintenance treatment of pulmonary sarcoidosis].

The aim of this investigation was to compare inhaled budesonide vs oral prednisone in the maintenance phase treatment of pulmonary sarcoidosis. Double-blind controlled study was performed in 40 patients with stage II or III pulmonary sarcoidosis. After initial systemic 6 weeks treatment with prednisone (40 reduced to 20 mg daily) patients were allocated either to systematically (P) or topically (B) treated group. P patients continued with 10 mg prednisone daily, B patients were given inhaled budesonide 1.6 mg daily. The progress of treatment was assessed by serial radiography, spirometry, serum ACE activity and plasma cortisol levels. All patients completed the 12 months treatment. Using a numerical score to assess changes on the chest radiograms P patients improved by 1.7 +/- 0.66 points; B patients improved by 1.15 +/- 0.81 points. Spirometric changes were insignificant. Serum ACE fell from 107 +/- 51 U/L in the P group and 92 +/- 40 U/L in the B group to 46 +/- 11 U/L and 38 +/- 21 U/L respectively during the initial phase of treatment. In the maintenance phase ACE levels remained lower than initial ones in both groups. Morning cortisol plasma levels studied in 10 patients (5 in each group) decreased significantly during the initial phase. Thereafter cortisol levels remained low in the P patients returning to the lower limit of normal values in the B patients. We conclude that inhaled budesonide may be a safe and effective alternative to oral steroids in the maintenance treatment of pulmonary sarcoidosis especially in the early, stage II, disease.


Long-term prognosis after myocardial infarction. Who is at risk for sudden death?

Patients who have survived myocardial infarction may be at risk for recurrent infarction or sudden death within the first year. Clinical factors that identify high-risk patients include the following: Poor left ventricular function Presence of spontaneous ventricular ectopy Abnormal signal-averaged electrocardiogram Decreased heart rate variability Exercise-induced depression of the ST segment Inducible ventricular tachyarrhythmia When these high-risk patients are identified, appropriate strategies (eg, use of antiarrhythmic agents or an implantable cardioverter-defibrillator) may improve outcome.


Acute confusional state. Don't mistake it for dementia.

Acute confusional state is a condition that is commonly encountered in many types of medical practice. Certain patients, especially the elderly, are vulnerable to the disorder, and recognition of this fact can help prevent the development or minimize the severity of the condition. Thorough history taking and evaluation often point to the underlying cause, and attention to general principles of environmental and pharmacologic management can minimize morbidity.


Management of ascites in patients with cirrhosis. What to do when diuretics fail.

Resistant or refractory ascites is unusual in cirrhotic patients who comply with dietary sodium restriction and optimal diuretic therapy. Patients unresponsive to medical therapy often have end-stage liver disease and renal insufficiency, although reversible complicating factors must be excluded. For patients with truly refractory ascites, liver transplantation is the only option that improves chances of survival. When this is not feasible, therapeutic paracentesis is the procedure of choice for intractable ascites. Several surgical shunts have been used, but none have been found to be safer and more effective than large-volume paracentesis.


Listerial meningitis. Which patients are vulnerable?

The physician's index of suspicion for infection with Listeria monocytogenes should be elevated if a patient presents with symptoms of meningitis and has impaired cell-mediated immunity. Although diagnosis is aided by detection of an elevated white blood cell count and protein level in the cerebrospinal fluid, it requires isolation of the organism from the cerebrospinal fluid. Appropriate antibiotic treatment leads to recovery in most cases.


Community-acquired pneumonia. The changing picture.

Important changes in the initial management of community-acquired pneumonia have been prompted by the discovery of new respiratory pathogens, the changing susceptibility of traditional pathogens to antimicrobial agents, and the introduction of new antimicrobial agents. Although the clinical presentation may suggest a specific pathogen, findings overlap too much to reliably distinguish the specific cause of the pneumonia on a clinical basis. Useful laboratory studies include Gram's stain and culture of sputum, blood culture, serologic studies, and new tests such as the urinary antigen test for Legionella pneumophila. Empirical antimicrobial treatment must take into consideration that 20% to 30% of cases of community-acquired pneumonia are due to atypical pathogens that are not susceptible to beta-lactam agents.


Radiocontrast-induced nephropathy. Prevention is better than cure.

The principal predisposing factor in radiocontrast-induced nephropathy appears to be underlying renal insufficiency. Identifying patients at risk is of paramount importance when a diagnostic study is being chosen. Contrast-reliant studies should be avoided, if possible, in high-risk patients. If challenge with a contrast medium is essential, appropriate risk stratification and adequate patient preparation should be done beforehand. Ultimately, prevention is a better approach than cure.


Hypothermia. Saving patients from the big chill.

Although hypothermia is a serious and sometimes fatal condition, prompt recognition and institution of appropriate rewarming techniques may save even profoundly affected persons. The diagnosis of hypothermia should be considered when patients present with alterations of cerebral function without apparent explanation, especially in the presence of underlying predisposing illnesses and conditions. When hypothermia is suspected, an accurate core temperature must be obtained. Application of rewarming techniques appropriate to the degree of hypothermia may be lifesaving. Conservative use of pharmacotherapy is warranted.


Ovarian cancer screening. The search for cost-effective methods.

The search goes on for an effective screening test for ovarian cancer. This deadly disease has eluded attempts at early detection with the currently available methods. Dr Williams examines the research regarding various screening methods and suggests areas of promise for future study.


Cardiac imaging after acute myocardial infarction. Identification of patients at continued risk.

Diagnostic imaging performed early in the course of acute myocardial infarction provides anatomic and functional information that is useful in assessing patients at risk for future cardiac events and premature death. Early identification of left ventricular dysfunction or complications of myocardial infarction allows appropriate and timely management of high-risk patients and early transfer of stable patients from the intensive care environment. Noninvasive predischarge functional imaging to unmask patients with jeopardized myocardium identifies high-risk patients who may need invasive studies and surgical or interventional treatment. Postdischarge risk stratification with diagnostic imaging provides vital prognostic information in high- and low-risk patients, allowing for appropriate allocation of medical resources.


Selection for body weight at eight weeks of age. 20. Production traits and the B and C alloantigen systems.

The current study evaluated influence of genotypes of the B and C alloantigen systems on production traits in lines of White Plymouth Rock chickens. Lines had been selected previously for high (HWS) or low (LWS) 8-wk BW and after 27 generations of selection, a random sample of each selected line was used to initiate subpopulations in which selection was relaxed. For the present study, blood typing was used to identify allelic frequencies for the B and C systems for five consecutive generations of selection (Generations 30 through 34) and three consecutive generations of relaxation (Generations 5 through 7). Haplotypes for the B complex were assigned designations B32, B33, B34, and B35 and alleles of the C system were assigned C8, C9, C10, and C11. Production traits for males were BW at 4 and 8 wk of age, and for females were BW at 4, 8, and 38 wk of age, age and BW at production of first egg, percentage of normal eggs, and percentage of normal hen-day egg production. There were no differences in production traits among B genotypes or among C genotypes, although some time trends in gene frequencies suggested that more subtle effects may be present.


Effects of genetic strain and light management on the reproductive performance of turkeys.

The laying performance of six genetic strains of turkeys, which have been bred and maintained at the Ohio Agricultural Research and Development Center, Wooster, OH, was compared under three laying house lighting regimens over a period of 3 yr. Light Treatment 1 (L1) consisted of 14 h of continuous light (L) and 10 h of dark (D; 14L:10D) throughout the laying period. Treatment 2 (L2) consisted of 14 h of intermittent light (IL, 15 min L and 45 min D/h) followed by 10 h of continuous dark. The IL treatment was started following a period (6 wk) in which the hens were trained to use the trapnests. Thus, during the first 6 wk of their laying period, L2 hens were also provided 14L:10D. Treatment 3 (L3) hens were provided the same program as L1 for the first 14 wk of the laying period. They were then moved to a continuous period of 19L:5D for the remainder of the laying period. All eggs produced were recorded through 180 days after the first egg was laid. Traits studied included: the number of days to first egg after light stimulation; the number of eggs produced through 84, 120, and 180 days after the first egg was laid; the average clutch length; the maximum clutch length; the total days lost to broodiness; the rate of lay; and the effective length of the laying period. Highly significant differences (P less than or equal to .01) were observed among the strains used for all traits measured. Light treatments showed no significant effects on any trait measured. Thus, from the present studies, the delayed IL program provides an economically attractive management program for environmentally controlled turkey breeder houses.(ABSTRACT TRUNCATED AT 250 WORDS)


Growth, body composition, and plasma androgen concentration of male broiler chickens subjected to different regimens of photoperiod and light intensity.

Day-old male Hubbard broilers (960) were assigned to one of four treatments (two pens of 120 birds per treatment) to evaluate the effects of high (150 lx) versus low (5 lx) light intensity and constant 23 h light (L):1 h dark (D) versus increasing (6L:18D increasing 4 h/wk to 23L:1D) photoperiod in a 2 x 2 factorial arrangement of treatments. Birds were raised to 8 wk on a typical commercial four-diet program. Low-intensity birds were heavier than high-intensity birds from 2 to 8 wk (3.25% heavier at 8 wk). Birds raised under constant photoperiod were heavier than birds raised under increasing photoperiod from 2 to 5 wk and at 7 wk of age (1.71% heavier at 7 wk). High-intensity bird carcasses had lower percentage body fat, weight of fat, and higher percentage body protein at 8 wk compared with low-intensity bird carcasses (7.77, 10.76, and 1.77%, respectively). High-intensity birds had smaller abdominal fat pads (weight and percentage of body weight) at 8 wk compared with low-intensity birds (15.46 and 12.17%, respectively). Photoperiod did not affect body composition. Birds treated with increasing photoperiod had larger testes (weight and percentage of body weight) at 8 wk compared with birds under the constant photoperiod (29.36 and 30.51%, respectively). Birds treated under increasing photoperiod had higher plasma androgen concentrations at 7 wk compared with birds under constant photoperiod (testosterone, .270 versus .188 ng/mL; androstenedione, .632 versus .494 ng/mL).(ABSTRACT TRUNCATED AT 250 WORDS)


Beak trimming and sex effects on behavior and performance traits of large white turkeys.

The effects of beak trimming at day old on performance and behavioral activities of male and female Large White turkeys were evaluated. One hundred and twenty poults of each sex were evenly assigned to 20 treatment pens and evaluated for body weight, feed usage, and livability characteristics to 18 wk of age. Behavioral observations were conducted for feeding, drinking, sleeping, huddling, resting, and agonistic activities. Beak trimming affected body weight and feed usage levels for the sexes differently. From 6 wk, trimmed males were significantly heavier than untrimmed males, whereas untrimmed females were heavier than trimmed females from 12 wk. Similar to body weights, feed usage levels from 13 to 18 wk were higher for trimmed males compared with untrimmed males and lower for trimmed females compared with untrimmed females. Feed conversion ratios after 12 wk and survival to 18 wk were not affected by trimming treatment. Livability rates, however, were lowest for untrimmed males and highest for untrimmed females. Effects on behavioral activities were confined primarily to the brooding and early rearing phases. Beak trimming reduced feeding activity of females and drinking activity of both males and females during the first 2 wk. Sleeping, huddling, and resting activities were increased by beak trimming for both sexes during brooding. Agonistic acts were reduced by beak trimming main effect at 3 and 6 wk. Trimmed males committed fewer agonistic acts at 6 wk than untrimmed males.


Long segmented filamentous organisms in broiler chicks: possible relationship to reduced serum carotenoids.

Over the past several years, broiler-flocks at the author's laboratory have exhibited low serum carotenoid levels. This phenomenon has not been related to exceptionally low carotenoids in the feed or to known protozoan or viral infections. However, long segmented filamentous organisms (LSFO) have been identified in the pale chicks. An inverse correlation between LSFO counts and serum carotenoid levels was not observed, but prevention of LSFO infection consistently resulted in higher carotenoid values. Young chicks were very susceptible to naturally acquired LSFO infection, but the organisms decreased in number with time and became very difficult to find by the time the chicks were 7 wk old. Segments of LSFO containing endospores were detected in mucosal smears. This observation, along with the extreme ease of natural infection, suggests that LSFO are transmitted by dust-borne spores. Transmission can be prevented by fumigation of housing with formaldehyde vapors.


Anticoccidial efficacy and chicken toleration of potent new polyether ionophores. 1. The septamycin relative CP-82,009.

The anticoccidial activity of the ionophore CP-82,009 against laboratory isolates of four major species of poultry Eimeria was investigated. Parameters of anticoccidial efficacy that were evaluated were control of lesions and weight suppression. At 4 and 5 ppm, CP-82,009 demonstrated broad-spectrum anticoccidial efficacy in battery trials that was equivalent to reference commercial ionophores. When CP-82,009 was fed to uninfected broiler chickens at efficacious dose levels, growth rate and feed efficiency were found to be equivalent to commercial agents over a 21-day period in batteries and over a 49-day period in floor pens. From the present studies, it appears that CP-82,009 is an efficacious anticoccidial that is well tolerated by chickens, and that it ranks among the most potent anticoccidial ionophores described to date.


Anticoccidial efficacy and chicken toleration of potent new polyether ionophores. 2. The portmicin relative CP-84,657.

The current study investigated the anticoccidial activity of the ionophore CP-84,657 against laboratory strains of the five major pathogenic species of Eimeria that infect poultry. Based on lesion scores and weight gain, the ionophore CP-84,657 achieved broad-spectrum anticoccidial efficacy in battery trials at doses of 4 and 5 ppm that was equivalent to reference commercial ionophores. In uninfected chickens, 4 ppm of CP-84,657 was the highest dose that gave growth rate and feed efficiency equivalent to commercial agents over 21 days in batteries and 49 days in floor pens. Ionophore CP-84,657 is an efficacious, well-tolerated anticoccidial in chickens, with potency comparable to that of the most potent known ionophores.


Colostrum from cows immunized with Eimeria acervulina antigens reduces parasite development in vivo and in vitro.

Experiments were undertaken to determine whether passive immunization utilizing hyperimmune bovine colostrum (HBC) specific for Eimeria acervulina (EA) antigens conferred protection against coccidiosis in chickens. The HBC was produced by immunizing three pregnant, nonmilking Jersey cows with EA antigens administered via one intramuscular injection followed by three intramammary infusions at approximately 10, 8, 6, and 4 wk before parturition. One cow was immunized with sporozoites (SZ), the second with merozoites (MZ), and the third with recombinant merozoite antigen (rMZ). A fourth cow, unimmunized, provided normal colostrum (NC) for control purposes. Colostral whey from each cow was tested by ELISA for antibody against SZ, MZ, and rMZ antigens. In all immunized cows, antiparasite titers were elevated above those of the control. Antibodies from MZ- and rMZ-immunized cows recognized both MZ and rMZ antigen. Separate groups of 2-wk-old chickens received two oral doses of anti-SZ, -MZ, or -rMZ HBC or NC or PBS daily from 1 day before through 6 days after oral inoculation (DAI) with EA oocysts. Feces from each group were examined for oocysts. Intestines were examined for lesions 6 DAI. Histologic sections of duodenum were examined for asexual stages and gametocytes utilizing monoclonal antibody and fluorescence microscopy. In Experiments 1 and 2, oocyst production was reduced in all HBC-treated groups, except one treated with rMZ HBC, compared with PBS- or NC-treated groups. In Experiment 2, the severity of lesions was significantly reduced in all HBC-treated groups compared with those that received NC or PBS. Significantly fewer developmental stages were found in histological sections from all chickens treated with anti-SZ and anti-rMZ HBC than from controls. Anti-SZ HBC significantly reduced the number of intracellular SZ found 24 h after their inoculation into cultures of primary chicken kidney cells. These results suggest that HBC specific for certain EA antigens can inhibit parasite development and reduce severity of parasite-related gut lesions.


Preferential accumulation of n-3 fatty acids in the brain of chicks from eggs enriched with n-3 fatty acids.

Docosahexaenoic acid (DHA) is the major polyunsaturated fatty acid in the phospholipids of brain tissue. The extent to which maternal dietary n-3 fatty acid may influence the n-3 fatty acid composition of phosphatidylethanolamine (PE) and phosphatidylcholine (PC) fractions of brain and liver tissue of progeny was investigated. Hens were fed diets containing n-9, n-3, or n-6 fatty acids. The brain tissue of the chicks from eggs enriched with n-3 fatty acids contained higher DHA, resulting in an increased C22:6 n-3:C20:4 n-6 ratio compared with chicks from eggs enriched with n-9 or n-6 fatty acids. Incorporation of C22:5 n-3, and C22:6 n-3 was mainly in the PE fraction as compared with the PC fraction of the chick brain. For chicks hatched from eggs enriched with n-9, n-3, and n-6 fatty acids, the total n-3 fatty acids in the brain PE fraction were 29.3, 41.3, and 21.9% compared with 6.7, 8.8, and 4.0% in the brain PC fraction. In contrast, the n-3 fatty acids increased evenly in phospholipid PC and PE fractions. Total amounts of PE in the brain lipids of chicks from eggs enriched with n-3 and n-6 fatty acids were 40.8 and 41.9% compared with 33.1% for chicks from eggs enriched with n-9 fatty acids. Analyses of the remaining yolk sac revealed a preferential absorption of DHA from the egg PE fraction by the developing chick.


Growth, thyroid function, and serum macromineral levels in magnesium-deficient chicks.

Growth and thyroid function were studied in Mg-deficient chicks. Dietary levels of 80 to 315 ppm Mg were compared with control levels of 578 to 787 ppm Mg. Signs of Mg deficiency appeared rapidly and acutely within 2 to 5 days at dietary levels of 250 to 260 ppm or lower. Growth and feed intake decreased progressively as the deficiency became more severe. Control chicks pair-fed with the deficient chicks gained significantly more weight. Serum Mg decreased at all levels of Mg below control, but at 260 and 315 ppm it returned to control values after 21 days on treatment. Serum Ca diminished only when dietary Mg was 250 ppm or less. Serum K increased in severely deficient chicks but decreased over time in milder deficiencies. Thyroid gland weights were unchanged. However, very young chicks fed a Mg-deficient diet had lower serum 3,5,3'-triiodothyronine (T3) whereas serum thyroxine (T4) was generally unaffected. Beyond 1 wk of age chicks that had prior access to a Mg-sufficient diet had low serum T4 levels whereas serum T3 was unchanged. Therefore, peripheral thyroid hormone metabolism is altered in a Mg deficiency, but this effect is dependent on the age at which the deficiency occurs.


The effect of early protein restriction (zero to eight weeks) on skeletal development in turkey toms from two to eighteen weeks.

Turkey toms were fed protein- and lysine-deficient diets or protein- and lysine-adequate diets from 0 to 4 and 5 to 8 wk and similar diets from 9 to 18 wk. Beginning at 2 wk of age and at approximate 2-wk intervals thereafter, the length and width of the right tibia were measured in nine toms per treatment. The length and width of the femur were measured beginning at 8 wk. Three tibiae that were close to the average length for each age and treatment were selected for ashing. The top 25% (epiphyseal) and middle 10% (diaphyseal) segments from each of these bones were fat-extracted prior to ashing. Mean length of the tibia continued to increase throughout the experiment whereas tibia width reached a plateau at 16 wk. Femur length also continued to increase throughout the study whereas there were no significant changes in femur width after 16 wk. Compared with the control diet, the diet deficient in protein and lysine resulted in a significant decrease in tibia and femur width at all ages. There were no dietary effects on tibia length at 16 and 18 wk, however. There were no significant dietary effects on femur length after 14 wk. There were significant age effects but no significant dietary effects on epiphyseal or diaphyseal bone ash. Epiphyseal bone ash increased from 41 to 54% over the course of the study whereas diaphyseal bone ash increased only from 59 to 64%.


2-Hydroxy-4 (methylthio) butanoic acid as a drinking water supplement for broiler chicks.

Two experiments were conducted to study the effects on chick performance of a commercially available liquid methionine analog [2-hydroxy-4 (methylthio) butanoic acid; HMB] supplied through drinking water. Methionine activities of .025 and .05% were added to drinking water and compared with levels of .03, .06, or .09% included in a low-methionine basal diet from 0 to 21 days of age. Mortality was not significantly altered by any dietary treatment. Neither feed nor water intake was affected adversely by HMB inclusion in drinking water. Only male body weight in Experiment 1 did not respond to HMB feed supplementation. Equivalent levels of total sulfur amino acid intake among water treatments were associated with significantly heavier body weights than the control. In both experiments, body weight and feed efficiency of birds receiving .05% methionine activity as HMB in water were equal or superior to those of groups that received supplemented feed. The results indicate that, under the study conditions, HMB provided in drinking water can be effectively assimilated by broiler chicks, at least through 21 days of age.


Effect of heating on nutritional quality of conventional and Kunitz trypsin inhibitor-free soybeans.

Five 10-day chick growth experiments and an amino acid digestibility assay were conducted to assess the effect of steam heating on in vivo protein quality of raw, full-fat Kunitz trypsin inhibitor-free soybeans (KFSB) compared with raw, conventional full-fat soybeans (CSB). Protein solubility in .2% KOH was also evaluated as an in vitro test of in vivo protein quality for underprocessed CSB. The CSB and KFSB were autoclaved for 0, 3, 6, 9, 12, 15, 18, or 21 min at 121 C and 124 kPa. The soybeans were then fed to 8- or 9-day-old chicks as the sole source of protein in dextrose and soybean diets containing 23% protein. Growth performance of chicks fed raw KFSB was superior to that of chicks fed raw CSB. Growth performance of chicks fed autoclaved KFSB or CSB increased and pancreas weight (percentage of body weight) decreased as autoclaving time increased. Slightly less autoclaving time was consistently required to achieve maximum chick performance for KFSB compared with CSB. Less autoclaving time was also required to obtain maximum digestibility of amino acids in KFSB compared with CSB. Urease activity of the soybeans decreased as autoclaving time increased, whereas protein solubility in .2% KOH for CSB did not change consistently in response to heating time. The results of the present study indicate that raw KFSB must be heated to obtain maximum protein quality for chicks and that protein solubility in KOH is not a sensitive indicator of underprocessing of CSB soybeans.


The effect of enzyme supplementation on the apparent metabolizable energy and nutrient digestibilities of wheat, barley, oats, and rye for the young broiler chick.

The influence of enzyme supplementation on the bioavailable energy (AME(n)) and apparent digestibilities of lipid (ALD) and protein (APD) in young broiler chicks was examined for diets containing either wheat, hulled or hulless barley, naked oats, or spring rye. Dietary AME(n), APD, and ALD values were depressed (P less than or equal to .01) for all test grains (except hulled Bedford barley) as the inclusion rate of the grain replacing wheat increased. The antinutritives, beta-glucans (barley and oats) and pentosans (rye), had the most pronounced effect on ALD. The decreases in ALD were 43, 77, and 67% for chicks fed diets containing 70% Scout barley (hulless), Terra oats, and Gazelle rye, respectively, compared with those fed the control wheat diet. Enzyme supplementation increased (P less than or equal to .01) AME(n), APD, and ALD for all test cereals. The corresponding increases in the AME(n), of the enzyme-supplemented diets containing 70% HY320 wheat, Bedford barley, Scout barley, Terra oats, and Gazelle rye diets were 4, 7, 42, 33, and 14%, respectively, compared with their unsupplemented counterparts. Enzyme treatment also improved (P less than or equal to .01) weight gains and feed conversion efficiencies of chicks fed diets containing each of the cereals. Overall, the results demonstrate that the nutritive value of cereal grains such as wheat, barley, oats, and rye can be improved by the addition of crude fungal extracts to the diet of young chicks.


Ionic and endocrine characteristics of reproductive failure in calcium-deficient and vitamin D-deficient laying hens.

Whole blood ionized calcium and plasma total calcium, inorganic phosphorus, estradiol-17 beta, progesterone, and 1,25-di-hydroxycholecalciferol concentrations were measured in calcium- or vitamin D-deficient Single Comb White Leghorn hens. Control birds were serially sampled every 2 h for 26 h immediately following oviposition until the next oviposition. Deficient birds, which had ceased laying 10 to 14 days prior to sampling, were sampled at the same times. The control birds had significantly higher mean total and bound plasma calcium and inorganic phosphorus concentrations than the deficient hens. Control and vitamin D-deficient hens had similar mean ionized calcium concentrations. Control hens exhibited a cyclic pattern in ionized calcium and inorganic phosphorus concentrations over the sampling period that was related to shell calcification. Deficient hens showed no changes in ionized calcium concentration during this time. Plasma 1,25-dihydroxycholecalciferol concentrations were significantly higher in the calcium-deficient birds than the control or vitamin D-deficient hens. Mean plasma estradiol-17 beta and progesterone concentrations were consistently higher in the control hens than the deficient hens. Consistent with this observation were decreases in ovary and oviduct weights, which occurred in the nonlaying deficient hens.


Effect of dietary supplemental pyridoxine levels on the hatchability of turkey eggs.

Two identical experiments were conducted each with 120 Large White turkey hens in individual cages to determine the value of dietary pyridoxine supplementation for increasing hatchability. The hens were fed a basal corn and soybean meal diet. Weekly data responses were averaged across 5- to 6-wk timespans and recorded in three grouped time periods over the reproduction cycles. The hens were photostimulated at 31 wk of age and at 33 wk of age were assigned to dietary treatments containing 0, 6, 12, and 18 mg of supplemented pyridoxine/kg of diet. Thirty hens were fed each dietary treatment. In Experiment 1, eggs were collected for 4 days in the middle of each time period and egg yolk and albumen were assayed separately for vitamin B6. Although the vitamin B6 concentration in egg yolk was stable (1.9 micrograms/g dried basis), concentrations of vitamin B6 in egg albumen increased with incremental dietary pyridoxine levels; however, the average level of vitamin B6 in egg albumen was only 4% of the average level in egg yolk. About 37 micrograms of vitamin B6 per egg (82 g) was assayed in eggs from all treatments in the production periods. Incremental dietary levels of supplemented pyridoxine above the basal (unsupplemented pyridoxine) diet level did not result in increasing hatchability or egg vitamin B6 levels. Differences were not observed for 7-day or 28-day embryo deaths among treatments within the three 5- or 6-wk production periods of both experiments.


Effect of feed allowance during rearing and breeding on female broiler breeders. 1. Growth and carcass characteristics.

The influence of feed allowance on growth and carcass characteristics was investigated with female Indian River broiler breeders. Four feeding programs were imposed: FF, provided ad libitum access to feed throughout; FR, provided ad libitum access to feed from 0 to 18 wk of age and restricted thereafter; RF, provided ad libitum access to feed from 0 to 4 wk, restricted from 4 to 18 wk, and consumed feed ad libitum thereafter; RR, provided ad libitum access to feed from 0 to 4 wk of age and restricted thereafter. All birds received a starter diet (2,739 kcal ME/kg, 19.1% CP) from 0 to 3 wk of age, a grower diet (2,729 kcal ME/kg, 15.5% CP) from 3 to 22 wk of age, and a layer diet (2,889 kcal ME/kg, 14.6% CP) from 22 to 62 wk of age. Restricted feeding (limited amount, fed daily) was based on commercial BW guidelines. Feed restriction (37.2% of ad libitum intake) during rearing (4 to 18 wk) slowed growth. At 18 wk of age, restricted birds (RF, RR) had an average BW of 1.9 kg and a shank length of 9.2 cm compared with ad libitum-fed birds (FF, FR), which had an average BW of 4.2 kg and a shank length of 10.8 cm. Despite the large difference in BW, percentage carcass protein did not differ significantly between the two rearing treatments; however, percentage carcass fat was almost four times greater in ad libitum-fed than in feed-restricted birds. The growth deficit remained significant to 62 wk of age in RF birds compared with FF birds. During the prebreeding period (18 to 23 wk), feed intake of RF birds was 209% that of RR birds and 133% that of FF birds. At sexual maturity, the BW of RF birds was 3.6 kg, compared with 4.6, 4.5, and 2.6 kg for FF, FR, and RR birds, respectively. Stepwise regression indicated that BW was the most important (R2 = .363, P = .0001) among 11 variables in determining the number of large (greater than 10 mm) follicles in the ovaries of breeders at sexual maturity.


Effect of feed allowance during rearing and breeding on female broiler breeders. 2. Ovarian morphology and production.

The influence of feed allowance during rearing (4 to 18 wk) and breeding (18 to 62 wk) on ovarian morphology and egg production was examined. During rearing, beginning at 4 wk of age, 400 Indian River pullets that had been full-fed from 1 day of age either continued to be full-fed (F, n = 200) or were feed restricted (R, n = 200), following commercial practice. At 18 wk of age, 100 birds (within +15% of the mean BW) in each of F and R rearing groups were further assigned to be full-fed (n = 50) or feed restricted (n = 50) during breeding. Thus, based on feed allowance during rearing and breeding, four treatment groups were formed (FF, FR, RF, and RR). Feed restriction during rearing significantly delayed oviduct development and age at sexual maturity (age at first egg). Compared with feed restriction during both rearing and breeding (RR), ad libitum feeding beginning at 18 wk of age (RF) significantly increased the weight of the ovary and the number of large follicles at sexual maturity but did not significantly affect the age at sexual maturity. Feed restriction during rearing, breeding, or both significantly reduced the incidence of erratic ovipositions, defective eggs, and multiple ovulations. The incidence of erratic ovipositions was positively correlated (r = .692, P less than or equal to .001) with the laying of soft-shelled and shell-less eggs and negatively correlated (r = -.508, P less than or equal to .001) with the production of settable eggs to 62 wk of age. Compared with ad libitum feeding, controlled feeding during breeding significantly improved both settable and total egg production. Among the four feeding regimens, Treatment RR resulted in the highest fertility and hatchability.


Effect of feed allowance during rearing and breeding on female broiler breeders. 3. Ovarian steroidogenesis.

The influence of feed allowance during rearing (4 to 18 wk) and breeding (18 to 62 wk) on in vitro follicular steroidogenesis was examined. Four feeding programs were imposed: FF, consumed feed ad libitum throughout; FR, provided ad libitum access from 0 to 18 wk of age and restricted thereafter; RF, provided ad libitum access to feed from 0 to 4 wk, restricted from 4 to 18 wk, and consumed feed ad libitum thereafter; RR, consumed feed ad libitum from 0 to 4 wk of age and restricted thereafter. All birds received a starter diet (2,739 kcal ME/kg, 19.1% CP) from 0 to 3 wk of age, a grower diet (2,729 kcal ME/kg, 15.5% CP) from 3 to 22 wk of age, and a layer diet (2,889 kcal ME/kg, 14.6% CP) from 22 to 62 wk of age. Restricted feeding was based on commercial guidelines. At 33 wk of age, 10 to 12 (FF = 10, FR = 12, RF = 10, and RR = 11) birds from each feeding regimen were killed by cervical dislocation and the follicles removed for in vitro steroidogenesis. Feed allowance during breeding (18 to 62 wk) had a significant effect on in vitro follicular steroidogenesis at 33 wk of age. In broiler breeders fed ad libitum, some of the F2 (second largest) follicles had an endocrine profile characteristic of the F1 (largest) preovulatory follicles, secreting very little androstenedione and large amounts of progesterone. In feed-restricted birds, only the F1 follicles, which were destined to ovulate next, secreted any significant amount of progesterone into the culture media. Compared with restricted feeding, ad libitum feeding significantly increased the production of androstenedione in small white follicles (less than 1 mm).(ABSTRACT TRUNCATED AT 250 WORDS)


Post-mortem biochemistry of Pekin duckling and broiler chicken Pectoralis muscle.

Biochemical post-mortem changes between red and white avian muscle were determined using duckling and chicken Pectoralis muscles, respectively. Six live Pekin ducklings and six live broiler chickens, in each of two trials, were obtained from commercial plants and processed at a pilot facility. After evisceration, carcasses were held at 4 C, then Pectoralis muscles were removed at .25, 1, 4, and 24 h post-mortem, and sampled for pH, lactate, adenosine triphosphate (ATP) content, and R-value (inosine to adenine ratio). Duckling Pectoralis pH significantly (P less than .05) decreased from 6.25 to 5.66 from .25 to 24 h post-mortem, respectively, as compared with that of the chicken, which decreased from 6.41 to 5.62 for the same times. Duckling lactate values, from .25 to 24 h, increased from 15.86 to 28.86 mumol/g, respectively, and chicken lactate values increased from 27.62 to 53.51 mumol/g. The ATP content of the duckling Pectoralis muscle decreased from 1.59 to .14 mumol/g and chicken Pectoralis decreased from 3.42 to .21 mumol/g from .25 to 24h, respectively. The R-values of duckling and chicken Pectoralis significantly increased from .25 to 24 h (.98 to 1.37 and .80 to 1.51, respectively). Duckling and chicken biochemical measurements were significantly different at all post-mortem sampling times, except for 24-h values of pH (5.66 versus 5.62, respectively) and 24-h contents of ATP (.14 versus .21 mumol/g, respectively). The different post-mortem biochemical measurements between duckling and chicken Pectoralis muscle is evidence that different rates of post-mortem metabolism and rigor development exist between these red and white avian breast muscles.


Research note: in ovo administration of a competitive exclusion culture treatment to broiler embryos.

Exposure of chicks to salmonellae in the hatchery and hatchery environment limits the effectiveness of a competitive exclusion (CE) culture treatment. Therefore, in an attempt to apply treatment before chicks are exposed to salmonellae, the CE culture was introduced in ovo to unhatched embryos. An undefined, anaerobically grown CE culture, derived from cecal contents of healthy adult chickens, was diluted 1:1,000 or 1:1,000,000 and inoculated either into the air cell or beneath the inner air cell membrane of 17-day-old incubating hatching eggs. The treated chicks were more resistant than untreated chicks to varying challenge levels of Salmonella typhimurium, indicating that it may be possible to initiate protection of chicks to salmonellae challenge prior to hatching into a contaminated environment.


Could the pharmacological differences observed between angiotensin II antagonists and inhibitors of angiotensin converting enzyme be clinically beneficial?

Over the past several years, angiotensin I converting enzyme (ACE) inhibitors, compounds that block the formation of angiotensin II (ANG II), have become widely used in the treatment of cardiovascular disease. Recently, a new class of orally active, non-peptide inhibitors of the renin-angiotensin system, the ANG II receptor antagonists have also become available. Since both classes of compounds block the renin-angiotensin system, although at different sites, it remains to be determined whether blockade of ANG II receptors will have any specific advantage over inhibition of ACE. The following review assesses the actions of ANG II antagonists and suggests ways in which blockade of ANG II receptors may differ both pharmacologically and clinically from inhibition of ACE.


Effects of endosulfan and aldrin on muscle coordination and conditioned avoidance response in rats.

The deteriorative effects after chronic endosulfan exposure on muscle coordination, learning and memory of rats were compared with that produced by aldrin which has been reported to have similar effects in experimental animals. A rota-rod apparatus was used to study the muscle coordination and learning and memory were tested by recording the response to unconditioned and conditioned stimuli using a pole-climbing apparatus. Aldrin but not endosulfan inhibited motor coordination in both sexes. A greater motor deterioration occurred in male group. This finding, together with the previous data which shows inhibition by its metabolite of motor activity, suggests that its metabolic product is responsible for this action. Like aldrin, endosulfan inhibited both learning ability and conditioned avoidance response. A change in the activities of brain monoamines or inhibition of perception and reflexes or both were proposed for these behavioural effects, since the former was reported to be produced by both compounds and the latter was found to occur in aldrin treated rats.


Absence of CYP3A genetic polymorphism assessed by urinary excretion of 6 beta-hydroxycortisol in 102 healthy subjects on rifampicin.

A previous study has demonstrated that the urinary level of 6 beta-hydroxycortisol is a marker of liver CYP3A content after induction by rifampicin. To put in evidence an eventual genetic polymorphism for this cytochrome, the frequency distribution of 6 beta-hydroxycortisol excretion was investigated in 102 healthy Caucasians before and after 6 days of oral rifampicin administration (600 mg daily). After rifampicin treatment, a wide interindividual distribution was observed but no clear bimodality. Moreover the mean 6 beta-hydroxycortisol level was higher in women (n = 38) than in men (n = 64). These observations do not favour the existence of a CYP3A genetic polymorphism based on 6 beta-hydroxycortisol excretion but evoke a sexual dimorphism. However, CYP3A is composed of at least four enzymes and as the enzyme(s) responsible for cortisol 6 beta-hydroxylation is (are) not perfectly known, it can not be excluded that a genetic polymorphism does exist for one enzyme of this family.


Extrahepatic disposition of 3H-aflatoxin B1 in the rainbow trout (Oncorhynchus mykiss).

Whole-body autoradiography in rainbow trout (Oncorhynchus mykiss) after oral and intravenous administration of 3H-labelled aflatoxin B1 showed labelling of several extrahepatic tissues, such as the uveal melanin and the vitreous humour of the eyes, the trunk and head kidney, the olfactory rosettes and the pyloric caecae. Liquid chromatography of extracts of the vitreous humour showed that unmetabolized 3H-AFB1 was the main labelled material present at this site. Liquid chromatography of extracts of the uveal melanin showed presence of aflatoxicol and aflatoxin B1 in proportions of about 3:1. The binding to the pigment is probably due to a hydrophobic type of interaction with the melanin. Microautoradiography showed that melanin-containing cells in the trunk and head kidney and in the olfactory rosettes also accumulated high amounts of radioactivity. In the trunk kidney there was, in addition, a labelling of the second segment of the proximal tubules and of the distal tubules and the collecting ducts. Studies in vitro with microsomal and 12,000 x g supernatant preparations of the trunk kidney showed formation of DNA- and protein-bound metabolites from the aflatoxin B1. It is probable that the bioactivation of the aflatoxin B1 is confined to the cytoplasm of the cells, may be related to excretion and/or absorption processes. Microautoradiography of the olfactory rosettes, showed labelling of the sensory epithelium, but not the indifferent epithelium. A low formation of protein-bound aflatoxin B1-metabolites was found in incubations with microsomal preparations of this tissue. The same observation was made in incubations with microsomal preparations of the head kidney. In the pyloric caeca bound metabolites were observed in vivo at a level comparable to that found in the trunk kidney. Our results suggest that retention and metabolism in some extrahepatic tissues might be of importance as concerns the toxicologic potential of aflatoxin B1 in the rainbow trout.


Effect of antioxidants on hypoxia/reoxygenation-induced injury in isolated perfused rat liver.

Isolated perfused livers from rats fasted overnight were subjected to 30 min. of hypoxia followed by reoxygenation for 60 min., resulting in marked cytotoxicity as evidenced by an enhanced release of cytosolic enzymes (lactate dehydrogenase: 14-fold over controls, glutamate-pyruvate-transaminase: 12-fold over controls) and glutathione (twofold over controls) into the perfusate, by calcium accumulation (by a factor of 1.4) in the tissue and by an 80% inhibition of bile secretion. Virtually no mitochondrial injury became apparent and no evidence for lipid peroxidation could be found. In the presence of ascorbate, an augmentation of hepatic injury was observed. This might be due to the pro-oxidant activity of ascorbate in the presence of ionized iron, which is easily released from high molecular weight stores under reductive (e.g. hypoxic) conditions. The water soluble vitamin E analogue trolox C as well as propyl gallate clearly protected the liver against hypoxia/reoxygenation injury, yielding further evidence for a causative role of oxidative stress in this model. Due to their water solubility and their high efficacy as free radical scavengers, these antioxidants might be of therapeutic value.


Acute neurobehavioural effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in Han/Wistar rats.

The neurobehavioural effects of a single non-lethal dose (1000 micrograms/kg intraperitoneally) of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) were assessed in young male Han/Wistar rats, highly resistant to acute lethality of TCDD. TCDD decreased body weight significantly compared with ad libitum fed controls. TCDD did not change the behaviour or the motility of rats in the open field test 8 days after the treatment nor did it affect the spontaneous motor activity up to 27 days after the exposure. In the elevated plus-maze test for anxiety, TCDD-treated rats did not differ from either ad libitum fed controls or pair-fed controls. In the 24-hr passive avoidance test, the learning of TCDD-treated rats did not differ significantly from that of ad libitum fed controls or pair-fed controls from 8 hr to 16 days after the treatment. TCDD did not affect the motor coordination or the maintenance of balance on the rotating rod but it impaired them slightly in the elevated horizontal bridge test 16 hr after exposure. It did not affect nociception in the hot plate test 16 hr or 8 days after the injection. The results suggest that a single sublethal dose of TCDD does not alter markedly the general behaviour of Han/Wistar rats, in contrast to its striking effect on feeding behaviour which results in a marked decrease in body weight gain.


Lipid peroxidation of erythrocyte membrane induced by lipoamide dehydrogenase in the presence of ADP-Fe3+.

Lipid peroxidation of rat erythrocyte membranes was induced by lipoamide dehydrogenase (LADH) (EC 1.8.1.4) in the presence of ADP-Fe3+. Superoxide dismutase (SOD) (EC 1.15.1.1) strongly inhibited the peroxidation reaction but catalase did not. Hydroxyl radical scavengers, mannitol and dimethylsulfoxide did not inhibit the lipid peroxidation. These results indicated that the lipid peroxidation was a superoxide (O2-)-dependent reaction, but the hydroxyl radical was not involved. ADP-Fe3+, in the presence of LADH, was reduced more rapidly under aerobic than anaerobic conditions and SOD under aerobic conditions strongly inhibited the iron reduction, indicating that O2- plays a predominant role in iron reduction. Hydrogen peroxide enhanced O2- generation by LADH, but the peroxidation reaction was not affected. In the presence of lipoamide, lipid peroxidation was also induced but the reactions were not inhibited by SOD. Evidently, the lipid peroxidation induced in the presence of lipoamide was O2(-)-independent. Dihydrolipoamide may be involved in the peroxidation reaction.


Single-dose and steady-state pharmacokinetics of diltiazem administered in two different tablet formulations.

Single-dose and steady state pharmacokinetics of diltiazem administered in two different oral formulations were assessed with particular reference to rate and extent of absorption. Following single dose administration a significant difference in tmax was observed (2.9 +/- 1.9 and 6.8 +/- 2.6 hr respectively) whereas differences in AUC, t1/2 and Cmax were not significant. The AUC (mean +/- S.D.) values following single dose administration of Cardil and Cardizem were 678.4 +/- 321.5 and 948.6 +/- 580.6 ng.ml-1.hr respectively. The mean and the 95% confidence limits for the observed ratio AUCCardil/AUCCardizem are 0.89 and 0.44-1.34 respectively. At steady-state a significant difference between Cmax/Cmin and tmax was seen Cmax/Cmin being 4.9 and 3.2 respectively and Tmax being 2.7 +/- 2.0 and 6.0 +/- 2.8 hr respectively, whereas Cmax and AUC did not differ significantly. The AUC (mean +/- S.D.) values in steady state of Cardil and Cardizem were 880.1 +/- 399.8 and 1056.8 +/- 509.8 ng.ml-1.hr respectively. The mean and the 95% confidence limits for the observed ratio AUCCardil/AUCCardizem are 0.96 and 0.66-1.26 respectively. Although the observed ratios AUCCardil/AUCCardizem in both the single-dose and the steady-state study do not differ significantly from 1.0, the confidence limits exceed the acceptable values given by Poulsen & Juul (personal communication 1990) (a 20% decrease or increase of the ratio to 0.8 or 1.2).


[Risk of cancer following cholecystectomy, hysterectomy or appendectomy].

Patients with previous cholecystectomy, hysterectomy or appendectomy have an increased risk of developing cancer of other organs such as the bowel, breast kidney or ovary, particularly if they have undergone more than one operation. Previous nephrectomy, strumectomy, prostatectomy or gastric resection for gastroduodenal ulcer do not increase the risk of bowel cancer.


[A program for medical use of information systems. Validity of calculation of direct costs for the initial treatment of acute myeloblastic leukemia].

In order to validate the information systems medicalization program used since 1984 at the Hôtel-Dieu hospital, Paris, the direct costs given by this program were compared with those calculated from the medical records of 10 adult patients hospitalized for diagnosis and treatment of acute myeloblastic leukaemia. The mean direct cost of an 8 to 81 days hospital stay was estimated at 75.393 +/- 41.260 French francs by the program and 84.969 +/- 52.288 FF by calculations from the records. A fairly good correlation (r2 = 0.72; P = 0.002) was found between the 2 evaluations. There was no statistically significant difference between the figures obtained by the 2 methods, except for pharmaceutical expenditures (P = 0.005) which were grossly underevaluated by the program. A better correlation could be obtained if an attribution of drugs and transfusions per patient was entered in the program.


[Gastroesophageal reflux treated by posterior hemifundoplication. 251 cases].

Between September 1983 and March 1991, 251 consecutive patients with gastro-oesophageal reflux resistant to medical treatment underwent posterior hemifundoplication (modified Toupet procedure). One hundred and seventy-seven patients (71 percent) had peptic oesophagitis. pH monitoring showed a mean Kaye's score of 278 +/- 245 with a 29 percent part of total recording time at pH < 4. The mean low oesophageal sphincter pressure was 8.5 +/- 6.5 cm H2O. No patient died in the postoperative period. Morbidity consisted of 8 splenic injuries, as well as 8 pulmonary and 23 thromboembolic complications. Assessment of 199 patients (79 percent) with a mean follow-up of 32 +/- 21 months showed complete symptomatic relief in 96.5 percent, and complete endoscopic healing of oesophagitis was noted in 96 percent. Restoration of the pH profile to normal levels was obtained in 86 percent of the cases. The mean low oesophageal sphincter pressure had risen to 17 +/- 6 cm H2O. Early postoperative dysphagia was noted in 46 patients (18 percent); one of them required reoperation. Reflux symptoms persisted in 9 patients (4.5 percent). pH monitoring revealed abnormal levels in 3 patients. The results of this study demonstrate that effective gastro-oesophageal reflux control can be achieved with the modified Toupet procedure.


[Inflammatory pseudotumor of the urachus. A case].

The occurrence in a 9-year old boy of a painful, prominent and solid mid-hypogastric tumour is reported. This clinically isolated muscular tumour was suggestive of sarcoma. En bloc resection of the tumour including the attached vesical cupula was performed. Histology showed that this large tumour was purely inflammatory and had developed from remnants of the urachus. This diagnosis must be considered whenever a suprapubic tumour is discovered. Since malignant transformation of urachus cysts is possible, surgical resection is advised.


[Atrial natriuretic factor in heart insufficiency].

The atrial natriuretic factor is a peptide with vasodilator and natriuretic properties which is synthesized and released by atrial myocytes. In patients with chronic heart failure the ventricles contribute to the synthesis of this peptide, so that its plasma concentration rises in proportion to the heart filling pressure. The atrial natriuretic factor, therefore, seems to be a good biological marker of functional and haemodynamic severity of heart failure. However, the weight of this factor in the neuro-hormonal balance has not yet been fully evaluated, owing to the usual attenuation of its biological effects. This, and the necessity of continuous parenteral administration limits, for the moment, its therapeutic applications. Inhibitors of the enzyme involved in its metabolism are being tested, but only preliminary results are available in humans.


[Brucella vaccination in professionally exposed subjects. Prospective study].

This prospective phase IV study on cohort concerns a vaccine made of the phenol-insoluble fraction of Brucella abortus biotype 1 (B19 strain). Three hundred and three professionally exposed subjects entered the study; 161 out of 182 subjects (88.5 percent) with negative response to an intradermal test for detection of previous contamination accepted to be vaccinated. Booster injections were given 18 and 36 months after vaccination. Local pain was observed after 45.2 percent of injections and moderate systemic reactions after 5 percent of injections. Seropositivity after primary vaccination reached 80 percent. The booster injection, justified by a major decrease of this rate after 18 months, gave exactly the same response of the thymo-independent type. This vaccinal schedule did not result in detectable hypersensitivity. The clinical effectiveness of the vaccine could not be evaluated accurately because of the insufficient number of subjects. The possibility of subclinical infection in vaccinated subjects calls for wider comparative studies of vaccinated versus non-vaccinated subjects.


[Plasma beta chorionic gonadotropin between 14 and 20 weeks of amenorrhea: a sign of pregnancy-related hypertension].

A defect of placenta maturation has been described in hypertension of pregnancy. Plasma beta chorionic gonadotropins (beta HCG) of placental origin rise at the onset of pregnancy and reach a peak between 9 and 10 weeks of amenorrhoea. As we were making systematic assays between 14 and 20 weeks in a trisomy detection programme, we looked for differences in plasma beta HCG levels between women with pregnancy-induced arterial hypertension and pregnant women with normal blood pressure. We also studied the predictive value of such assays. Pregnancy-induced hypertension was found in 6 women in a population of 89 nulliparas and in 12 women in a population of 163 multiparas. beta HCG levels were significantly higher in women who later developed hypertension among both nulliparas (52,833 +/- 19,538 IU vs 24,499 +/- 16,485 IU) and multiparas (50,558 +/- 23,597 IU vs 20,911 +/- 11,677 IU). In nulliparas, taking 43,000 IU as threshold of pathology we found that the predictive value of beta HCG was higher than that of other tests which had gone through controlled studies (sensitivity 67 percent, specificity 91.6 percent, positive predictive value 36 percent, negative predictive value 97.4 percent, relative risk 5.4). In multiparas, taking 38,000 as threshold and combining this marker with obstetrical history it was possible to predict the occurrence of hypertension more precisely than with other markers which had gone through controlled studies (sensitivity 66.7 percent, specificity 98 percent, positive predictive value 61.4 percent, negative predictive value 97.3 percent, relative risk 8.4).


[Deforming arthropathy associated with the presence of anti Jo-1 antibody in polymyositis. A new case].

A deforming arthropathy confined to the hands developed in a woman with polymyositis of three year's duration. Roentgenograms showed distal subluxations of several fingers, especially the thumb, asymmetrical marginal erosions of the phalanges and periarticular calcifications. These roentgenographic findings are considered to be specific of polymyositis associated with anti-Jo-1 antibody.


Implicit knowledge: new perspectives on unconscious processes.

Recent evidence from cognitive science and neuroscience indicates that brain-damaged patients and normal subjects can exhibit nonconscious or implicit knowledge of stimuli that they fail to recollect consciously or perceive explicitly. Dissociations between implicit and explicit knowledge, which have been observed across a variety of domains, tasks, and materials, raise fundamental questions about the nature of perception, memory, and consciousness. This article provides a selective review of relevant evidence and considers such phenomena as priming and implicit memory in amnesic patients and normal subjects, perception without awareness and "blindsight" in patients with damage to visual cortex, and nonconscious recognition of familiar faces in patients with facial-recognition deficits (prosopagnosia). A variety of theoretical approaches to implicit/explicit dissociations are considered. One view is that all of the various dissociations can be attributed to disruption or disconnection of a common mechanism underlying conscious experience; an alternative possibility is that each dissociation requires a separate explanation in terms of domain-specific processes and systems. More generally, it is concluded that rather than reflecting the operation of affectively charged unconscious processes of the kind invoked by psychodynamic or Freudian theorists, dissociations between implicit and explicit knowledge are a natural consequence of the ordinary computations of the brain.


Application of DNA fingerprinting to the recovery program of the endangered Puerto Rican parrot.

The Puerto Rican parrot was reduced to approximately 13 animals in 1975 and as a conservation measure, a captive population was established from a few founders taken from the wild between 1973 and 1983. The number of successful breeding pairs in captivity has been low, and the captive breeding program has not been as productive as that of the closely related Hispaniolan parrot. Therefore, a genetic study was initiated to examine the relative levels of relatedness of the captive founders using levels of bandsharing in DNA fingerprints. Unrelated captive founder Puerto Rican parrots had the same average level of bandsharing (0.41) as second-degree relatives of the Hispaniolan parrot (0.38, P > 0.05), with an inbreeding coefficient of 0.04. High levels of bandsharing (> 40%) between pairs of males and females correlated with reproductive failure, suggesting that inbreeding depression is partly responsible for the low number of breeding pairs. Consequently, DNA profiling can be used to guide the captive breeding program for the Puerto Rican parrot, and other endangered species, by identifying pairs of males and females with low levels of bandsharing.


Requirement for coenzyme Q in plasma membrane electron transport.

Coenzyme Q is required in the electron transport system of rat hepatocyte and human erythrocyte plasma membranes. Extraction of coenzyme Q from the membrane decreases NADH dehydrogenase and NADH:oxygen oxidoreductase activity. Addition of coenzyme Q to the extracted membrane restores the activity. Partial restoration of activity is also found with alpha-tocopherylquinone, but not with vitamin K1. Analogs of coenzyme Q inhibit NADH dehydrogenase and oxidase activity and the inhibition is reversed by added coenzyme Q. Ferricyanide reduction by transmembrane electron transport from HeLa cells is inhibited by coenzyme Q analogs and restored with added coenzyme Q10. Reduction of external ferricyanide and diferric transferrin by HeLa cells is accompanied by proton release from the cells. Inhibition of the reduction by coenzyme Q analogs also inhibits the proton release, and coenzyme Q10 restores the proton release activity. Trans-plasma membrane electron transport stimulates growth of serum-deficient cells, and added coenzyme Q10 increases growth of HeLa (human adenocarcinoma) and BALB/3T3 (mouse fibroblast) cells. The evidence is consistent with a function for coenzyme Q in a trans-plasma membrane electron transport system which influences cell growth.


A putative ATP-dependent RNA helicase involved in Saccharomyces cerevisiae ribosome assembly.

We have isolated a cold-sensitive mutant of Saccharomyces cerevisiae in which there is a deficit of 60S ribosomal subunits. Cold sensitivity and the assembly defect are recessive and cosegregate, defining a single essential gene that we designated DRS1 (deficiency of ribosomal subunits). The wild-type DRS1 gene was cloned by complementation of the cold-sensitive phenotype of drs1. Sequence analysis reveals a high degree of similarity to a family of proteins that are thought to function as ATP-dependent RNA helicases. Pulse-chase analysis of ribosomal RNA synthesis and processing indicates that the drs1 mutant accumulates the 27S precursor of the mature 25S rRNA. These results suggest that, as in pre-mRNA splicing, RNA helicase activities are involved in ribosomal RNA processing.


Active complete in vitro replication of nodavirus RNA requires glycerophospholipid.

Flockhouse virus (FHV) is a member of the nodavirus group of positive-strand RNA viruses. In the absence of additional compounds, a template-dependent RNA-dependent RNA polymerase extracted from FHV-infected cells synthesizes complementary (-)-strand copies of added FHV RNA to yield a double-stranded RNA product. Upon addition of glycerophospholipid (GPL), this system reproducibly carries out complete highly active replication of added FHV RNA, producing newly synthesized (+)-strand RNA in predominantly single-stranded RNA form. This accounts for previously observed effects of Lipofectin (a mixture of GPL and cationic lipid) in the system. All tested neutral and negatively charged GPLs except phosphatidic acid support complete FHV RNA replication in this in vitro system, as do phospholipid extracts from uninfected and FHV-infected cells. Neither sphingomyelin, a membrane phospholipid that is not derived from glycerol, nor cholesterol supported FHV RNA replication. Testing of compounds derived from GPL shows that the ability of active GPL to support FHV (+)-strand RNA synthesis is dependent on the structures of both the head group and the acyl chains. Neither the phosphorylated head group nor the diacylglycerol lipid moiety alone supports RNA replication. The length and saturation of acyl chains strongly influence the ability of GPL to support RNA replication. Other characteristics of this in vitro RNA replication system and the possible role played by membranes and their components in FHV RNA replication are discussed.


Evidence that the packaging signal for nodaviral RNA2 is a bulged stem-loop.

Flock house virus is an insect virus belonging to the family Nodaviridae; members of this family are characterized by a small bipartite positive-stranded RNA genome. The larger genomic segment, RNA1, encodes viral replication proteins, whereas the smaller one, RNA2, encodes coat protein. Both RNAs are packaged in a single particle. A defective-interfering RNA (DI-634), isolated from a line of Drosophila cells persistently infected with Flock house virus, was used to show that a 32-base region of RNA2 (bases 186-217) is required for packaging into virions. RNA folding analysis predicted that this region forms a stem-loop structure with a 5-base loop and a 13-base-pair bulged stem.


At least three distinct proteins are necessary for the reconstitution of a specific multiprotein complex at a eukaryotic chromosomal origin of replication.

We have reconstituted in vitro a multistage assembly of a protein complex that specifically recognizes a yeast genomic origin of replication, the autonomously replicating sequence ARS121. The first step in the assembly was the interaction of the known origin-binding factor OBF1 and another factor, OBF2, with the ARS121 origin of replication to form the OBF1-OBF2-origin complex. This complex was the substrate for the ATP-dependent binding of a third DNA-binding activity, the core binding factor, CBF. Binding of CBF to the origin, identified by the retarded mobility of the origin DNA fragment in agarose gels, required, in addition to ATP and the OBF1-OBF2-origin complex, a functional essential core nucleotide sequence. ARS121 DNA containing mutations in the core, which inactivate the origin in vivo, did not sustain stable CBF binding, whereas ARS121 DNA mutated outside the boundaries of the essential core, which has normal origin function, bound CBF as wild type. This tight, direct correlation between the ability of the origin to bind CBF and its function as an origin of replication in vivo strongly suggest that the multiprotein complex reconstituted in vitro has a key role in the initiation of DNA replication.


Marked replicative advantage of human mtDNA carrying a point mutation that causes the MELAS encephalomyopathy.

The segregation of mutant and wild-type mtDNA was investigated in transformants constructed by transferring human mitochondria from individuals belonging to four pedigrees with the MELAS encephalomyopathy-associated mtDNA mutation (MELAS is mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes) into human mtDNA-less (rho 0) cells. Five of 13 clonal cell lines containing mixtures of wild-type and mutant mtDNAs were found to undergo a rapid shift of their genotype toward the pure mutant type. The other 8 cell lines, which included 6 exhibiting nearly homoplasmic mutant mtDNA, on the contrary, maintained a stable genotype. Subcloning experiments and growth rate measurements clearly indicated that an intracellular replicative advantage of mutant mtDNA was mainly responsible for the dramatic shift toward the mutant genotype observed in the unstable cell lines.


A yeast cyclophilin gene essential for lactate metabolism at high temperature.

The cyclophilins are a family of ubiquitous eukaryotic proteins first identified by high affinity for cyclosporin A (CsA). The immunosuppressant and cytotoxic effects of CsA are thought to result from formation of a toxic complex between cyclophilin and CsA rather than from inhibition of cyclophilin function. The physiological role(s) of the cyclophilins is unknown. Cyclophilins have in vitro peptidylprolyl cistrans isomerase (PPIase) activity, and thus may be involved in protein folding in vivo. We have isolated a yeast cyclophilin gene, CPR3, which encodes a presumptive mitochondrial isoform. While CPR3 disruption mutants lack any phenotype at 30 degrees C, they are unable to grow on L-lactate at 37 degrees C. Disruptions of two other cyclophilin genes (CPR1, CPR2) and of FPR1, the gene encoding an FK506 binding protein with PPIase activity, do not affect growth on L-lactate at 37 degrees C. L-Lactate metabolism requires transcriptional induction of CYB2, the gene encoding flavocytochrome b2; cpr3 mutants induce transcription of this gene normally. This result demonstrates a conditional lethal phenotype for a cyclophilin mutation and presents a system for genetic and biochemical analysis of cyclophilin function.


Isolation, characterization, and transcription of the gene encoding mouse mast cell protease 7.

A gene that encodes mouse mast cell protease (mMCP) 7 (also known as mouse mast cell tryptase 2) was isolated by genomic cloning with a cDNA that encodes mMCP-6, a tryptase in serosal mast cells. cDNAs encoding mMCP-7 were isolated from a bone-marrow-derived mast cell cDNA library. The mMCP-7 gene spans 2.3 kilobases and contains five exons rather than six, as found in the mMCP-6 and human mast cell tryptase I genes. Comparison of the 5' end of the transcript with the genomic sequence indicated that the region corresponding to the first intron in the mMCP-6 and human tryptase I genes is not spliced during transcription of mMCP-7 mRNA because of a point mutation at the intron 1 acceptor splice site; this results in a 5' untranslated region of 195 nucleotides, which is longer than that of any other known mast cell-specific transcript. mMCP-7 is 71-76% homologous with mMCP-6 and with dog and human mast cell tryptases, and it is the most acidic mast cell protease, with an overall net charge of -10. RNA blot analyses revealed that the mMCP-7 gene is transcribed in bone-marrow-derived mast cells but is not transcribed in mature serosal mast cells or in mucosal mast cell-enriched intestinal tissue of Trichinella spiralis-infected mice. Transcription of the mMCP-7 gene by differentiating bone-marrow-derived mast cells occurred within 1 week of bone-marrow culture but decreased dramatically after 3 weeks. Thus, the mMCP-7 gene displays a number of unusual structural characteristics and is distinctive in its transient and selective expression in immature mast cells maintained in interleukin 3-enriched medium.


Expression of a coriander desaturase results in petroselinic acid production in transgenic tobacco.

Little is known about the metabolic origin of petroselinic acid (18:1 delta 6cis), the principal fatty acid of the seed oil of most Umbelliferae, Araliaceae, and Garryaceae species. To examine the possibility that petroselinic acid is the product of an acyl-acyl carrier protein (ACP) desaturase, Western blots of coriander and other Umbelliferae seed extracts were probed with antibodies against the delta 9-stearoyl-ACP desaturase of avocado. In these extracts, proteins of 39 and 36 kDa were detected. Of these, only the 36-kDa peptide was specific to tissues which synthesize petroselinic acid. A cDNA encoding the 36-kDa peptide was isolated from a coriander endosperm cDNA library, placed under control of the cauliflower mosaic virus 35S promoter, and introduced into tobacco by Agrobacterium tumefaciens-mediated transformation. Expression of this cDNA in transgenic tobacco callus was accompanied by the accumulation of petroselinic acid and delta 4-hexadecenoic acid, both of which were absent from control callus. These results demonstrate the involvement of a 36-kDa putative acyl-ACP desaturase in the biosynthetic pathway of petroselinic acid and the ability to produce fatty acids of unusual structure in transgenic plants by the expression of the gene for this desaturase.


Fibrinogen blocks the autoactivation and thrombin-mediated activation of factor XI on dextran sulfate.

The intrinsic pathway of blood coagulation is activated when factor XIa, one of the three contact-system enzymes, is generated and then activates factor IX. Factor XI has been shown to be efficiently activated in vitro by surface-bound factor XIIa after factor XI is transported to the surface by its cofactor, high molecular weight kininogen (HK). However, individuals lacking any of the three contact-system proteins--namely, factor XII, prekallikrein, and HK--do not suffer from bleeding abnormalities. This mystery has led several investigators to search for an "alternate" activation pathway for factor XI. Recently, factor XI has been reported to be autoactivated on the soluble "surface" dextran sulfate, and thrombin was shown to accelerate the autoactivation. However, it was also reported that HK, the cofactor for factor XIIa-mediated activation of factor XI, actually diminishes the thrombin-catalyzed activation rate of factor XI. Nonetheless, it was suggested that thrombin was a more efficient activator than factor XIIa. In this report we investigated the effect of fibrinogen, the major coagulation protein in plasma, on the activation rate of factor XI. Fibrinogen, the preferred substrate for thrombin in plasma, virtually prevented autoactivation of factor XI as well as the thrombin-mediated activation of factor XI, while having no effect on factor XIIa-catalyzed activation. HK dramatically curtailed the autoactivation of factor XI in addition to the thrombin-mediated activation. These data indicate that factor XI would not be autoactivated in a plasma environment, and thrombin would, therefore, be unlikely to potentiate the activation. We believe that the "missing pathway" for factor XI activation remains an enigma that warrants further investigation.


Relationship between O6-alkylguanine-DNA alkyltransferase activity and N-methyl-N'-nitro-N-nitrosoguanidine-induced mutation, transformation, and cytotoxicity in C3H/10T1/2 cells expressing exogenous alkyltransferase genes.

While a great deal of evidence has directly implicated the importance of O6-alkylation of guanine in the mutagenicity of alkylating agents, evidence demonstrating the oncogenic potential of this lesion has been largely indirect. We have combined a well-studied in vitro neoplastic transformation system (using C3H/10T1/2 mouse cells) with a proven method of gene transfection for expressing the bacterial O6-alkylguanine-DNA alkyltransferase (AT; EC 2.1.1.63) repair genes ada and ogt to generate subclones which possess augmented repair capability toward specific DNA lesions. The products of these genes specifically and differentially repair O6-methylguanine (O6-MeGua), O4-methylthymine (O4-MeThy), and methylphosphotriesters. We show that the level of expression of either the ada or the ogt AT gene in C3H/10T1/2 cells directly correlates with protection against mutation to ouabain resistance by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Subclones expressing 70 fmol of AT per 10(6) cells exhibited a mutation frequency approximately 1/40th of that of clones expressing 15 fmol of AT per 10(6) cells when treated with MNNG at 0.4 micrograms/ml. Protection against mutagenesis by MNNG at 0.8 micrograms/ml, however, did not exceed 12-fold even in subclones expressing greater than 100 fmol of AT per 10(6) cells. As an MNNG dose of 0.6 micrograms/ml was sufficient to saturate more than 95% of the AT activity in any of the clones, the residual mutation frequency may have been caused by unrepaired O6MeGua lesions. In contrast to mutagenesis, protection against neoplastic transformation in vitro, in cells expressing high levels of AT, was most pronounced in cells treated with the highest dose of MNNG used (1.2 micrograms/ml). Low levels of transformation caused by MNNG at 0.4 and 0.8 micrograms/ml were not consistently inhibited in those clones. These data suggest that O6-MeGua formation is of major but not unique significance in the neoplastic transformation of C3H/10T1/2 cells by MNNG.


Specific inhibition of human immunodeficiency virus type 1 replication by antisense oligonucleotides: an in vitro model for treatment.

We have developed a culture system, simulating in vivo conditions of human immunodeficiency virus type 1 (HIV-1) infection, to evaluate the long-term efficacy of antisense oligonucleotide treatment. Five oligonucleotide phosphorothioates (28-mers), complementary to different regions of HIV-1 RNA, blocked replication of the virus in a sequence-specific manner at 1 microM concentration. Variations in antiviral activity were seen among the different oligonucleotides, revealing an effect of target selection. Mismatched or random oligonucleotide phosphorothioates delayed, but did not completely inhibit, HIV-1 replication. In the case of inhibition by a splice-acceptor-site antisense oligodeoxynucleotide, a break-through phenomenon occurred after 25 days of treatment, suggesting the development of an "escape mutant." This result did not occur when the inhibitory oligodeoxynucleotides were complementary to the primary-sequence areas of the rev-responsive element and rev-1 genes. Sequential treatment of HIV-1-infected cells with a combination of different antisense oligonucleotides, each administered once, also prevented the development of escape mutants. Our results suggest that chemotherapy based on specifically targeted antisense-oligonucleotide phosphorothioates may be an effective method for reducing the viral burden in HIV-1-infected individuals at clinically achievable oligonucleotide concentrations.


Transcription of the hypersensitive site HS2 enhancer in erythroid cells.

In the human genome, the erythroid-specific hypersensitive site HS2 enhancer regulates the transcription of the downstream beta-like globin genes 10-50 kilobases away. The mechanism of HS2 enhancer function is not known. The present study employs RNA protection assays to analyze the transcriptional status of the HS2 enhancer in transfected recombinant chloramphenicol acetyltransferase (CAT) plasmids. In erythroid K562 cells in which the HS2 enhancer is active, the HS2 sequence directs the synthesis of long enhancer transcripts that are initiated apparently from within the enhancer and elongated through the intervening DNA into the cis-linked CAT gene. In nonerythroid HL-60 cells in which the HS2 enhancer is inactive, long enhancer transcripts are not detectable. Splitting the HS2 enhancer between two tandem Ap1 sites abolishes the synthesis of a group of long enhancer transcripts and results in loss of enhancer function and transcriptional silencing of the cis-linked CAT gene. In directing the synthesis of RNA through the intervening DNA and the gene by a tracking and transcription mechanism, the HS2 enhancer may (i) open up the chromatin structure of a gene domain and (ii) deliver enhancer binding proteins to the promoter sequence where they may stimulate the transcription of the gene at the cap site.


General splicing factors SF2 and SC35 have equivalent activities in vitro, and both affect alternative 5' and 3' splice site selection.

The human pre-mRNA splicing factors SF2 and SC35 have similar electrophoretic mobilities, and both of them contain an N-terminal ribonucleoprotein (RNP)-type RNA-recognition motif and a C-terminal arginine/serine-rich domain. However, the two proteins are encoded by different genes and display only 31% amino acid sequence identity. Here we report a systematic comparison of the splicing activities of recombinant SF2 and SC35. We find that either protein can reconstitute the splicing activity of S100 extracts and of SC35-immunodepleted nuclear extracts. Previous studies revealed that SF2 influences alternative 5' splice site selection in vitro, by favoring proximal over distal 5' splice sites, and that the A1 protein of heterogeneous nuclear RNP counteracts this effect. We now show that SC35 has a similar effect on competing 5' splice sites and is also antagonized by A1 protein. In addition, we report that both SF2 and SC35 also favor the proximal site in a pre-mRNA containing duplicated 3' splice sites, but this effect is not modulated by A1. We conclude that SF2 and SC35 are distinct splicing factors, but they display indistinguishable splicing activities in vitro.


Multiple growth factors, cytokines, and neurotrophins rescue photoreceptors from the damaging effects of constant light.

Recent demonstrations of survival-promoting activity by neurotrophic agents in diverse neuronal systems have raised the possibility of pharmacological therapy for inherited and degenerative disorders of the central nervous system. We have shown previously that, in the retina, basic fibroblast growth factor delays photoreceptor degeneration in Royal College of Surgeons rats with inherited retinal dystrophy and that the growth factor reduces or prevents the rapid photoreceptor degeneration produced by constant light in the rat. This light-damage model now provides an efficient way to assess quantitatively the survival-promoting activity in vivo of a number of growth factors and other molecules. We report here that photoreceptors can be significantly protected from the damaging effects of light by intravitreal injection of eight different growth factors, cytokines, and neurotrophins that typically act through several distinct receptor families. In addition to basic fibroblast growth factor, those factors providing a high degree of photoreceptor rescue include brain-derived neurotrophic factor, ciliary neurotrophic factor, interleukin 1 beta, and acidic fibroblast growth factor; those with less activity include neurotrophin 3, insulin-like growth factor II, and tumor necrosis factor alpha; those showing little or no protective effect are nerve growth factor, epidermal growth factor, platelet-derived growth factor, insulin, insulin-like growth factor I, heparin, and laminin. Although we used at least one relatively high concentration of each agent (the highest available), it is still possible that other concentrations or factor combinations might be more protective. Injecting heparin along with acidic fibroblast growth factor or basic fibroblast growth factor further enhanced the degree of photoreceptor survival and also suppressed the increased incidence of macrophages produced by either factor, especially basic fibroblast growth factor. These results now provide the impetus for determining the normal function in the retina, mechanism(s) of rescue, and therapeutic potential in human eye diseases for each agent.


Binding and transport of gangliosides by prosaposin.

Prosaposin, the precursor of saposins A, B, C, and D, which activate lysosomal hydrolysis of sphingolipids, exists in various tissues and body fluids and is especially abundant in the nervous system. Prosaposin and saposins A,B, C, and D formed stable complexes with 13 different gangliosides as measured by an assay using column chromatography. Gangliosides of the gangliotetraose type (a series) were bound with high affinity, whereas b series gangliosides, O-acetylated gangliosides, and gangliosides with shorter carbohydrate chains, were bound with lower affinity. Prosaposin and saposins transferred gangliosides from donor liposomes to erythrocyte ghost membranes. Prosaposin also stimulated ganglioside GM1 beta-galactosidase more than mature saposins. Prosaposin exists as a secretory protein and as an integral membrane protein, and we propose that prosaposin is active as a ganglioside binding and transport protein in vivo.


Basal release of nitric oxide from aortic rings is greater in female rabbits than in male rabbits: implications for atherosclerosis.

Estradiol is known to exert a protective effect against the development of atherosclerosis, but the mechanism of this hormonal action is unknown. One of the early events in the development of atherosclerosis is the adhesion of macrophages to endothelial cells, and nitric oxide (NO) inhibits this process. We show that basal release of NO is greater with endothelium-intact aortic rings from female rabbits than those from males. Oophorectomy diminishes both circulating estradiol concentration and basal release of NO to levels seen in male rabbits. These data establish that basal NO release from endothelium-intact aortic rings depends on circulating estradiol concentration and offer an explanation for the protective effect of estradiol against the development of atherosclerosis.


Control of gal transcription through DNA looping: inhibition of the initial transcribing complex.

Involvement of DNA looping between two spatially separated gal operators, OE and OI, in repression of the gal operon has been demonstrated in vivo. An in vitro transcription assay using a minicircle DNA containing the gal promoter region with lac operators was employed to elucidate the molecular mechanism of repression. Wild-type lac repressors (LacI+ protein molecules), which are capable of associating into a tetramer and forming a DNA loop, repressed transcription from promoter sites P1 and P2, whereas a non-looping lac repressor mutant (LacI(adi)) failed to show normal repression of both of the gal promoters. Thus a DNA loop is also required for repression of transcription in vitro. Repression mediated by DNA looping resulted in the inhibition of the synthesis of complete as well as aborted transcripts, demonstrating that the repressive action was on the formation or activity of the initial transcribing complex. Under similar conditions, the gal repressor (GalR protein) did not repress the gal promoters effectively, apparently because it failed to loop DNA containing gal operators in the purified system. The component(s) or conditions that aid GalR in DNA looping remain to be identified.


NAD+ and NADH regulate an ATP-dependent kinase that phosphorylates enzyme I of the Escherichia coli phosphotransferase system.

Crude extracts of Escherichia coli contain a protein kinase, EI-K, that phosphorylates enzyme I (EI) of the phosphoenolpyruvate:glycose phosphotransferase system (PTS). Phosphorylation occurs at the active site histidine residue. The activity of EI-K was lost during purification. However, kinase activity was restored by adding NAD+ or NADP+.NADH reversed NAD+ activation of the kinase, and the level of EI-K activity was dependent on the NAD+/NADH ratio. Although crude preparations of EI-K showed no NAD+ requirement, they were completely inhibited by NADH, either in the assay mixture or when the enzyme was pretreated and the NADH was removed prior to the assay. NAD+ restored full activity to the NADH-pretreated inactive fractions. The results suggest that EI-K contains a bound cofactor that is lost during purification and that may be analogous to NAD+. EI-K activity may serve to link some of the diverse functions of the PTS, such as sugar transport, to the metabolic state of the cell.


Aerosol gene delivery in vivo.

The ability to express transgenes selectively within the lung will greatly facilitate the development of gene therapy for a variety of human diseases. We have demonstrated that aerosol administration of a chloramphenicol acetyltransferase (CAT) expression plasmid complexed to cationic liposomes produces high-level, lung-specific CAT gene expression in mice in vivo. Significant levels of CAT activity are seen in the lungs for at least 21 days following aerosolization. In situ immunostaining for intracellular CAT protein reveals that the majority of airway epithelial and alveolar lining cells are transfected in vivo. Histological analyses show no apparent treatment-related damage. These results have important implications for the development of human gene therapy.


Yeast RNA polymerase II initiation factor e: isolation and identification as the functional counterpart of human transcription factor IIB.

Yeast RNA polymerase II initiation factor e was purified to homogeneity and identified by biochemical criteria as the counterpart of human transcription factor IIB. Factor e was essential for initiation of transcription from yeast and mammalian promoters in a reconstituted yeast transcription system. Activity resided in a single polypeptide of approximately 41 kDa, identified by peptide sequence analysis as the product of the SUA7 gene. Factor e interacted specifically with RNA polymerase II, consistent with a proposed role in determining the start site of transcription.