Elevated levels of homocysteine predict cardiovascular death, nonfatal myocardial infarction, and symptomatic bypass graft disease at 2-year follow-up following coronary artery bypass surgery.

Severe graft disease occurs in patients at a rate of approximately 15% within the first year of coronary artery bypass grafting (CABG). In this study, the authors examined predictors of the combined end point of death, nonfatal myocardial infarction (MI), and bypass graft disease at 2-year follow-up after CABG. One hundred twenty-one consecutive patients were included in this study after informed consent was obtained. In univariate analysis, there was a significantly (P<.05) higher homocysteine level (11.0 ng/mol vs 9.7 ng/mol, P=.04) in patients who met the combined end point vs those who did not. There were no statistically significant differences in the following: low-density lipoprotein cholesterol, high-sensitivity C-reactive protein, and lipoprotein(a) values; age; body mass index; smoking and diabetes status; statin or aspirin use; creatinine level; hematologic markers; left ventricular ejection fraction; number of bypass grafts; and distribution of coronary artery disease. Logistic regression analysis modeling for low-density lipoprotein cholesterol, lipoprotein(a), fibrinogen, and homocysteine showed that homocysteine value (P=.016) was an independent predictor of the primary combined end point.

Lymphocytic choriomeningitis virus infection of the developing brain: critical role of host age.

OBJECTIVE: Lymphocytic choriomeningitis virus (LCMV) is a common human pathogen that causes substantial injury to the developing brain when the infection occurs during pregnancy. However, among children with congenital LCMV infection, there is considerable variability in the site, nature, and severity of neuropathology and in the clinical outcome. We hypothesize that the variability in neuropathology and outcome is due to differences in the gestational timing of LCMV infection. METHODS: We utilized an animal model of human congenital LCMV infection, in which developing rat pups were inoculated with LCMV at a series of postnatal ages, including postnatal days 1, 4, 6, 10, 21, 30, and 60. Cellular targets of infection were determined immunohistochemically, viral titers were determined by plaque assay, and pathology was determined by histological analysis, neuronal quantification, and immunostaining for lymphocytic subclasses. RESULTS: Host age at the time of infection profoundly affected the cellular targets of infection, maximal viral titers, immune response to the viral infection, and the severity, nature, and location of the neuropathology. All of the pathological changes observed in children with congenital LCMV infection were reproduced in the rat model by infecting the rat pups at different ages. INTERPRETATION: The effect of LCMV infection on the developing brain strongly depends on host age at the time of infection. Much of the variability in neuropathology and outcome among children with congenital LCMV infection probably depends on the gestational age at which the infection occurs.

Severe alcohol-induced neuronal deficits in the hippocampus and neocortex of neonatal mice genetically deficient for neuronal nitric oxide synthase (nNOS).

Alcohol can severely damage the developing brain, and neuronal loss is a critical component of this injury. Thus, identification of molecular factors that ameliorate alcohol-induced neuronal loss is of great importance. Previous in vitro work has demonstrated that nitric oxide (NO) protects neurons against alcohol toxicity. We tested the hypothesis that neonatal mice carrying a null mutation for neuronal nitric oxide synthase (nNOS), the enzyme that synthesizes NO in neurons, have an increased vulnerability to alcohol-induced neuronal loss in the neocortex and hippocampus. Wildtype mice and nNOS-/- mice received ethanol (0.0, 2.2, 3.3, or 4.4 g/kg) daily over postnatal days (P) 4-9 and were sacrificed on P10. The number of hippocampal CA1 and CA3 pyramidal cells, dentate gyrus granule cells, and neocortical neurons were determined using stereological methods. Alcohol pharmacokinetics did not differ between wildtype and nNOS-/- strains. Alcohol induced dose-dependent reductions in all four neuronal populations, and the losses were substantially more severe in the nNOS-/- mice than in wildtype. Furthermore, the threshold dose of alcohol to induce cell death was lower in the nNOS-/- mice than in the wildtype mice for all neuronal populations. While nNOS deficiency worsened alcohol-induced neuronal losses, the magnitude of this exacerbation varied among brain regions and depended on alcohol dose. These results demonstrate that nNOS deficiency decreases the ability of developing neurons in vivo to survive the toxic effects of alcohol and strengthen the hypothesis that NO exerts a neuroprotective effect against alcohol toxicity in the developing brain.

Use of frozen sections to determine neuronal number in the murine hippocampus and neocortex using the optical disector and optical fractionator.

Stereology is an important technique for the quantification of neurons in subregions of the central nervous system. A commonly used method of stereology relies upon embedment of tissue in glycol methacrylates to allow production of sections that are resistant to shrinkage in thickness. However, the use of glycol methacrylates for stereology has several disadvantages, including severe constraints on the size of tissue that can be processed and the long duration of time often required for infiltration. We describe a novel method of stereology utilizing tissue sections cut in the frozen state. This new methodology relies upon the staining of sections as free-floating sections and upon the mounting of these sections onto slides with a water-based mounting media. Sections cut in the frozen state and processed by these methods undergo little or no shrinkage in thickness and are ideal for stereological cell counts utilizing either the optical disector or optical fractionator methods of stereology. We demonstrate that frozen sections can be utilized to estimate neuronal number with high degrees of precision and with low coefficients of error. Because large tissue blocks can be cut as frozen sections, this method expands the range of tissues that can be processed efficiently for stereology and readily allows quantification of neurons from multiple brain regions from the same tissue sections. We applied this new methodology to estimate neuronal numbers in the neocortex and hippocampus of 10-day-old mice. The method was useful for estimation of both large, sparsely packed cell populations, such as the neocortex, and small, densely packed cells, such as the dentate gyrus granule cells. Thus, frozen section methodology offers many potential advantages over the use of glycol methacrylate embedment for stereology. These advantages include expansion of the size of tissue blocks that can be processed, reduction in expended time and costs, and ability to quantify multiple brain regions from a single set of sections.