Determination of free aspartic acid enantiomers in rat brain by capillary electrophoresis with laser-induced fluorescence detection.

Quantification of aspartic acid enantiomers in rat brain by using a chiral capillary electrophoresis procedure is described. Amino acids were pre-column derivatized with naphthalene-2,3-dialdehyde. Enantiomeric separation was achieved by micellar electrokinetic chromatography in the presence of methanol and beta-cyclodextrin as chiral selector. The chiral separation was coupled with laser-induced fluorescence detection. Contents of D- and L-aspartic acids in rats at different stages of growth (from 1 day before birth to 90 days after birth) were determined. D-Aspartic acid was detected in all the brain tissue samples tested, but at different levels. In the cerebrum of rats 1 day before birth, D-aspartic acid was found to be at the highest concentration of 81 nmol/g wet tissue. The level of D-aspartic acid in rat brain falls rapidly after birth, while the L-aspartic acid level increases with age.

Case study: rapid emergence of a cytomegalovirus UL97 mutant in a heart-transplant recipient on pre-emptive ganciclovir therapy.

A cytomegalovirus (CMV) strain containing a mutation in the UL97 gene (Leu595Ser), which is known to confer ganciclovir resistance, emerged rapidly in a heart-transplant recipient after 54 days of antigenaemia-guided ganciclovir therapy. The emergence of this viral mutant, while the patient was receiving oral ganciclovir 1000 mg three times daily, was associated with increasing CMV pp65 antigenaemia levels despite the re-instatement of intravenous ganciclovir 5 mg/kg twice daily. The antiviral regimen was then switched to intravenous foscarnet 90 mg/kg, administered either twice daily, once daily or every other day, which resulted in a rapid decrease of CMV antigenaemia levels and prevented the development of CMV-associated disease. Frequent monitoring of the CMV viral load and/or mutational studies are warranted in highly immunocompromised transplant recipients to prevent the development of CMV disease caused by ganciclovir-resistant strains.

Desmethyl tirilazad improves neurologic function after hypoxic ischemic brain injury in piglets.

OBJECTIVE: Desmethyl tirilazad is a lipid-soluble free radical quencher. Deferoxamine reduces free radicals by chelating iron and reducing hydroxyl formation. Free radical inhibitors have shown promise in several hypoxic ischemic brain injury models, and we wished to see if this work could be extended to our newborn piglet model. DESIGN: Randomized controlled trial. SUBJECTS: Piglets (0 to 3 days old). INTERVENTION: Carotid snares and arterial and venous catheters were placed under 1.5% isoflurane anesthesia. In Experiment 1, piglets were randomly assigned to receive either 3 mg/kg desmethyl tirilazad or vehicle at -15 and 90 mins. In Experiment 2, piglets were randomly assigned to receive either 20 mg/kg desmethyl tirilazad at -15 mins followed by 8 mg/kg/hr for 90 mins or 100 mg/kg deferoxamine at -15 mins or vehicle. At time 0, both carotid arteries were clamped and blood was withdrawn to reduce the blood pressure to two-thirds normal. At 15 mins, inspired oxygen was reduced to 6%. At 30 mins, the carotid snares were released, the withdrawn blood was reinfused, and the oxygen was switched to 100%. On the third day after the hypoxic ischemic injury, the animals were killed by perfusing their brains with 10% formalin. We tested the timing of lipid peroxidation and inhibition of lipid peroxidation by these agents by freezing the brains of a subset of pigs in liquid nitrogen. MEASUREMENTS: Neurologic examination and brain pathology were scored by blinded observers. Thiobarbituric acid-reactive substance and oxidized and reduced glutathione were measured on frozen brains. MAIN RESULTS: Desmethyl tirilazad (20 mg/kg) and 100 mg/kg deferoxamine inhibit lipid peroxidation. Desmethyl tirilazad (20 mg/kg) improves neurologic exam, but 3 mg/kg Desmethyl tirilazad or 100 mg/kg deferoxamine does not. Neither desmethyl tirilazad nor deferoxamine improves pathologic results. CONCLUSIONS: High-dose desmethyl tirilazad improves neurologic function after hypoxic ischemic brain injury in the newborn piglet.

N-tosyl-L-phenylalanyl-chloromethylketone reduces hypoxic-ischemic brain injury in rat pups.

N-tosyl-L-phenylalanyl-chloromethylketone (TPCK) in vitro blocks apoptotic pathways leading to cell death. We wished to see if TPCK would reduce brain injury in vivo. Seven-day-old rat pups had the right carotid artery ligated and then received either vehicle or TPCK (5 to 100 mg/kg i.p.). They were then given 8% oxygen for 2.25 h. Twenty-two days later, the cerebral hemispheres were weighed to determine the reduction in size in the right hemisphere. TPCK decreased the reduction in right hemisphere weight from 15+/-3% (vehicle, n=20), to 4+/-2% (10 mg/kg, n = 19, P<0.01). TPCK reduced the number of cells staining for DNA breaks 3 days after injury from 1729+/-275 mm(-2) (vehicle, n = 8) to 550+/-236 mm(-2) (10 mg/kg TPCK, n = 9, P<0.01), decreased the amount of DNA fragmentation 3 days after injury by gel electrophoreses (20 mg/kg, n = 16, P<0.01) and eliminated the increase in nitric oxide metabolites 6 h after injury (vehicle 1.5+/-0.4, n = 10; and 20 mg/kg TPCK 0.0+/-0.1 nM/mg protein, n = 10, P<0.001). TPCK pretreatment in the newborn rat model of hypoxic-ischemic brain injury reduces DNA fragmentation, nitric oxide production and brain injury.

Changes in left ventricular mass and function after aortic valve replacement: a comparison between stentless and stented bioprosthetic valves.

The objective of this study was to compare stentless bioprostheses with stented bioprostheses with regard to the postoperative changes in left ventricular (LV) mass and function. Forty patients with aortic stenosis undergoing valve replacement with a stentless (20 patients) or a stented (20 patients) bioprosthesis were evaluated early (baseline), 1 year, and 2 years after operation. Left ventricular mass index was calculated with the corrected American Society of Echocardiography formula. The relative changes between end-diastole and end-systole in LV mid-wall radius, length, and volume (ejection fraction) were determined with a previously validated model for dynamic geometry of the left ventricle. Overall, a significant decrease was found in LV mass index (from 155 +/- 30 to 112 +/- 23 g/m(2); P <.001) and a significant increase in longitudinal shortening (from 0.12 +/- 0.11 to 0.22 +/- 0.08; P <. 001), and ejection fractions (from 0.67 +/- 0.11 to 0.71 +/- 0.10; P =.017). No significant change was found in the mid-wall radius shortening fraction. Two years after surgery, the extent of LV mass regression was greater in stentless bioprostheses (-51 +/- 18 vs -35 +/- 17 g/m(2); P =.01), though the average mass index was similar in both groups (114 +/- 26 vs 110 +/- 20 g/m(2)). Also at 2 years, the longitudinal shortening fraction was greater in patients with a stentless bioprosthesis (0.25 +/- 0.07 vs 0.18 +/- 0.08; P =.03). In conclusion, this study suggests that the superior hemodynamic performance of stentless bioprostheses may have some benefits with regard to LV mass regression and function after aortic valve replacement. The significance of these benefits in terms of prognosis remains to be determined.

Role of postnatal penicillin prophylaxis in prevention of neonatal group B streptococcus infection.

This prospective study was designed to identify the role of postnatal penicillin prophylaxis in the prevention of neonatal group B streptococcus (GBS) infection. We studied 10 998 infants. Of these, 5389 were in the penicillin prophylaxis group (PP) and 5609 infants did not receive penicillin prophylaxis (NPP). Infants were allocated to treatment by month of birth, alternating 3-mo blocks or 2-mo blocks to the two groups after the first block was randomly assigned. The use of PP reduced the incidence of clinical sepsis (1.7% PP versus 2.5% NPP, p < 0.01), GBS infection (0.4% PP versus 0.9% NPP, p < 0.001) and deaths from sepsis (0.1% PP versus 0.3% NPP, p < 0.05). We conclude that the routine use of postnatal penicillin prophylaxis appears to be effective in reducing the incidence of clinical sepsis and death from sepsis in neonates.

Long-term outcome of infants at the margin of viability.

The outcome of babies at extremely short gestational age (22 to 26 weeks) effects our clinical decisions regarding their care. We looked at survival and presence of disability at 25 +/- 11 months of age in 246 of these infants born at our hospital between 1992 and 1996 who were average weight for gestational age. Babies were evaluated in our follow up clinic by a pediatrician, and a physical therapist for cerebral palsy, blindness and deafness, and by a psychologist with the Bayley II. Chances for survival without disability exceeded 50% of live born infants at 25 weeks gestation or a birth weight of 700 to 800 grams. Chances for survival exceeded 50% of live born infants at 24 weeks gestation or a birth weight of 600 to 700 grams. Chances for intact survival reached 50% of survivors at 23 weeks gestation or a birth weight of 400 to 500 grams.

Hemodynamic features of the freestyle aortic bioprosthesis compared with stented bioprosthesis.

BACKGROUND: The Freestyle prosthesis is a new stentless aortic bioprosthesis. Anticipated benefits are improved hemodynamics and increased longevity. METHODS: Doppler echocardiograms were performed early and at 3 to 6 months, 1 year, and 2 years after operation in 157 patients (69 men, 88 women, aged 48 to 85 years) with this prosthesis, and results were compared with hemodynamic data in patients with Intact and Mosaic stented bioprostheses. RESULTS: Distinctive features of the prosthesis compared with stented prostheses are (1) an increase in effective orifice area (+0.15+/-0.26 cm2; p < 0.05) and a decrease in mean gradient (-3.5+/-4.0 mm Hg; p < 0.001) during the first 3 to 6 months postoperatively and stabilization thereafter; (2) a markedly lower mean gradient at 1 year after operation (average, 6+/-4 mm Hg) than in stented prostheses (Intact, 22+/-8 mm Hg; Mosaic, 12+/-6 mm Hg); (3) in contrast to stented prostheses, in vivo effective orifice areas much lower (-0.91+/-0.35 cm2) than those calculated in vitro; (4) as in stented prostheses, the indexed effective orifice area (cm2/m2) is the best predictor (r = 0.77 at 1 year) of the mean gradient after operation; and (5) similar incidence of aortic regurgitation (trivial or mild, 34% versus 29% in Intact). CONCLUSIONS: The hemodynamics of the Freestyle are very satisfactory and represent a marked improvement in comparison to stented prosthesis.

The effect of glucose during ischemia on brain ATP, lactate, and glutamate in piglets.

Glucose worsens hypoxic-ischemic brain injury in 0- to 3-day-old piglets. Piglets were randomly assigned to have blood glucose increased with glucose infusion (n = 12), or decreased with insulin (n = 13), or a sham group (n = 10). In the insulin and glucose groups at time 0, both carotid arteries were clamped, and blood was withdrawn to reduce the blood pressure one third. At time 15 min FiO2 was reduced to 6%. At time 30 min cerebrospinal fluid (CSF) was taken for glutamate, and the brains were frozen. The shams had CSF removed and brains frozen without hypoxia or ischemia. Brain ATP was 1.7 +/- 0.09 mumol/g wet weight in the shams, 0.98 +/- 0.09 in the glucose group (p < 0.01 vs. sham), and 0.52 +/- 0.10 in the insulin group (p < 0.01 vs. glucose). Brain lactate levels were 4.3 +/- 1.0 mumol/g wet weight in the shams, 18.3 +/- 1.9 in the insulin group (p < 0.01 vs. sham), and 29.4 +/- 2.6 in the glucose group (p < 0.01 vs. insulin). CSF glutamate was 9.3 +/- 3.6 microM in the glucose group, 9.6 +/- 3.5 in the insulin group, and 2.2 +/- 0.9 in the shams (p < 0.05, glucose and insulin > sham). The Bmax for MK-801 binding was 2.3 +/- 0.2 pmol/mg protein in the glucose group, 2.6 +/- 0.1 in the insulin group (p < 0.05 vs. sham), and 2.0 +/- 0.2 in the shams, and the Kd was the same in the three groups (p = nonsignificant). Brain Na+,K(+)-ATPase was the same in the three groups (p = nonsignificant). Providing additional glucose preserves ATP during hypoxic-ischemic brain injury in the piglet but does not change CSF glutamate or brain-801 binding and probably worsens outcome by elevating brain lactate levels above the threshold for cellular injury.

Effect of birth order on incidence of periventricular-intraventricular hemorrhage in very low birth weight twins.

The purpose of the study was to determine whether the incidence of periventricular-intraventricular hemorrhage (PV-IVH) is significantly different between the firstborn and secondborn of very low birth weight twins. The firstborn and secondborn of 102 sets of twins were compared by incidence of PV-IVH, mode of delivery, birth weight, 5-minute Apgar score, pH, duration of oxygen therapy, duration of ventilator support, and occurrence of patent ductus arteriosus, air leak, bronchopulmonary dysplasia, or hyaline membrane disease. PV-IVH was graded by the Papile classification. Statistical data were analyzed by either the paired t test or McNemar's test. Major PV-IVH (grades III and IV) occurred in 11 firstborn and 14 secondborn infants. Minor PV-IVH (grades I and II) occurred in 11 firstborn and 8 secondborn twins. Neither these findings nor those from other measured variables showed a statistically significant difference.

AMPA antagonist LY293558 does not affect the severity of hypoxic-ischemic injury in newborn pigs.

BACKGROUND AND PURPOSE: LY293558 is a systemically active alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) excitatory amino acid antagonist. AMPA antagonists have shown promise in several adult hypoxic-ischemic brain injury models, and we wanted to see if this work could be extended to a newborn animal. METHODS: Seventy-six (beta error < .10) 0- to 3-day-old piglets under 1.5% isoflurane anesthesia underwent placement of carotid snares and arterial and venous catheters. While paralyzed with succinylcholine under 0.5% isoflurane, 50% nitrous oxide, piglets were randomly assigned to receive either 5 mg/kg or 15 mg/kg of LY293558 or saline at time--10 minutes and again 10 hours later. At time 0, both carotid arteries were clamped, and blood was withdrawn to reduce the blood pressure to two thirds of normal. At time 15 minutes, inspired oxygen was reduced to 6%. At time 30 minutes, the carotid snares were released, the withdrawn blood was reinfused, and the oxygen was switched to 100%. On the third day after the hypoxic-ischemic injury, the animals were killed by perfusion of the brain with 10% formalin. Brain pathology was scored by a blinded observer. RESULTS: There were no significant differences between the drug-treated and control groups. CONCLUSIONS: The systemically active AMPA antagonist LY293558, when given at a dose of 5 mg/kg or 15 mg/kg before injury and 10 hours later, does not affect the severity of hypoxic-ischemic brain injury in newborn piglets. Neither AMPA receptor activity nor NMDA receptor activity are important in brain injury in this model.

Overall hemodynamic studies after the chronic inhibition of endothelial-derived nitric oxide in rats.

Previous studies have demonstrated that an acute intravenous administration of nitro-L-arginine methyl ester (L-NAME) causes a sustained hypertension and widespread vasoconstriction. However, little information is available regarding the chronic effect of L-NAME on circulatory hemodynamics. Therefore, the purpose of the present study was to characterize both the systemic and regional hemodynamics after the chronic inhibition of endothelium-derived nitric oxide in male Sprague Dawley rats. The rats were divided into two groups: control (n = 8) and L-NAME (n = 8). The rats in the control group received only tap water and the rats in the L-NAME group received oral L-NAME solution at a dose of 0.1 mg/mL in the drinking water ad libitum. Four weeks after L-NAME or tap water treatment the rats were anesthetized with inactin, and mean arterial blood pressure, cardiac output, and individual organ flows were measured. Cardiac output and individual organ flows were measured using radioactive microspheres. Chronic administration of L-NAME resulted in a significant increase in mean arterial blood pressure from a control value of 118 +/- 4 mm Hg to 174 +/- 8 mm Hg (P < .01). Cardiac output decreased from a control value of 29 +/- 2 mL/min/100 g to 20 +/- 2 mL/min/100 g (P < .01) and total peripheral resistance increased from a control value of 4.3 +/- 0.3 mm Hg/mL/min/100 g to 9.7 +/- 1.4 mm Hg/mL/min/100 g (P < .01). In addition, chronic L-NAME treatment resulted in a widespread vasoconstriction and decrease in regional blood flows.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation of the needle technique for producing an arteriovenous fistula.

The purpose of the present study were to evaluate the needle technique of creating an arteriovenous (a-v) fistula and to quantitatively determine the hemodynamic responses in rats with three different fistula sizes. The fistula was made in male Sprague-Dawley rats between the aorta and vena cava below the renal arteries by using 20-, 18-, and 16-gauge angiocath needles. Five weeks after a sham operation or creation of an a-v fistula, mean arterial blood pressure (MAP), right atrial pressure, shunt flow, cardiac index, systemic flow, individual organ flows, and heart weight were quantitatively determined. All flow measurements were made using radioactive microspheres. The flow to the lungs was used as a measure of shunt flow. The shunt flow in the 20-, 18-, and 16-gauge fistula animals was significantly increased from a value of 2 +/- 1% (SE) to 50.0 +/- 0.1, 78 +/- 3, and 76 +/- 3% of total cardiac output, respectively. Average cardiac index in the 20-, 18-, and 16-gauge fistula animals increased by 105, 270, and 250%, respectively, compared with control. Right atrial pressure and heart weight were increased in proportion to the size of the fistula. MAP in the control and 20-, 18-, and 16-gauge fistula animals was 122 +/- 5, 126 +/- 3, 118 +/- 3, and 111 +/- 4 mmHg, respectively. There were no significant differences in MAP or systemic flow among any of these groups. The calculated total peripheral resistance in all fistula groups was significantly decreased compared with control.(ABSTRACT TRUNCATED AT 250 WORDS)

Postnatal penicillin prophylaxis and the incidence of group B streptococcal sepsis in neonates.

We conducted a retrospective chart review of infants, born over a 3-year period, who had positive urine latex agglutination and/or positive blood culture for group B streptococci (GBS). Infants routinely received intramuscular aqueous penicillin for the first half of the study period, and no penicillin was given for the subsequent 18 months. Overall, infants who received penicillin prophylaxis had a decreased incidence of clinical sepsis and positive blood culture for GBS (4.8/1,000 versus 8/1,000 and 1.3/1,000 versus 5.4/1,000, respectively). The incidence of GBS sepsis during the time of penicillin prophylaxis was not different from that in previously reported studies. When analyzed by weight groups, no difference in clinical sepsis or positive blood cultures for GBS was seen in the subset of infants weighing < or = 2,500 g at birth. There were fewer positive blood cultures in the infants who received penicillin and met the criteria for clinical sepsis. Mortality from GBS sepsis was unchanged during these two study periods in all weight groups.