Distribution of cytosolic and mitochondrial aspartate aminotransferase in normal, ischemic, and necrotic myocardium. An immunohistochemical study

We utilized immunoperoxidase methods to study the distribution of both cytosolic or soluble(s) and mitochondrial (m) aspartate aminotransferase (AspAT) in normal, ischemic, and necrotic myocardium. Human myocardium was obtained from autopsy (n = 9) and surgery (n = 6). Cardiac tissue from 26 dogs with experimental myocardial infarction induced by closed-chest balloon occlusion and four dogs with myocardial ischemia without necrosis induced by a 50% reduction in left main coronary artery flow for 3 hours were studied. Duration of occlusion was 45 minutes (n = 1), 3 hours (n = 11), 5 to 6 hours (n = 10), or 15 to 24 hours (n = 4). Highly purified m- and s-AspAT and specific antibodies were prepared in our laboratory. In all cases, control experiments were performed. Microscopically normal human and dog myocardium uniformly stained for m- and s-AspAT. Necrotic myocardium from patients with infarcts showed markedly reduced immunostaining. In those dogs with myocardial necrosis, all dogs with coronary occlusion of 5 to 24 hours, and eight of 11 dogs with 3-hour occlusions, immunostaining was significantly reduced for both s- and m-AspAT in regions confirmed to be necrotic by triphenyl tetrazolium chloride and hematoxylin and eosin staining. Myocardial necrosis was confirmed in the 3-hour infarcts by electron microscopy. In the four dogs with a 50% reduction in left main flow for 3 hours, and one dog with a 45-minute coronary occlusion, ischemia was demonstrated by glycogen loss in period acid-Schiff-stained sections but there was no evidence of necrosis by electron microscopy or triphenyl tetrazolium chloride staining and there was no loss of immunostaining evident for s- or m-AspAT. Thus, s- and m-AspAT were visualized in normal and ischemic myocardium with decreased staining in necrotic tissue using immunoperoxidase techniques. Loss of both s- and m-AspAT can be demonstrated in human myocardium and in experimental canine myocardium as early as 3 hours after coronary occlusion and appears to be specific for irreversible myocardial injury. No depletion of isoenzyme can be detected by immunohistochemical techniques in tissue that is ischemic but not necrotic. Furthermore, by these immunoperoxidase techniques, loss of s- and m-AspAT from necrotic myocardium appears to be simultaneous.     

Intracerebroventricular passive immunization in transgenic mouse models of Alzheimer's disease

Since uncontrolled production of beta-amyloid (Abeta) is considered a key seeding event underlying progression of Alzheimer's disease (AD), elimination of excessive Abeta and preventing its reaccumulation constitute the primary therapeutic goal in preventing and treating AD. To date, immunoneutralization has been the most effective strategy in removing pre-existing cerebral Abeta. Both active and systemic passive immunizations are known to reduce cerebral Abeta and improve memory in transgenic murine models of AD. However, active immunization is associated with adverse effects such as encephalitis with perivascular inflammation and hemorrhage, while passive immunization has the potential to disrupt cerebral vasculature that is laden with amyloid and exposed to high levels of antibody in the blood. Intriguingly, intracerebroventricular passive immunization established in the authors' laboratory circumvented these problems. The authors demonstrated that a single intracerebroventricular injection of anti-Abeta antibody reduced the cerebral Abeta burden and Abeta-related astrocytosis, retarded reaccumulation of Abeta and restored Abeta-induced depletion of presynaptic SNAP-25, for at least 1 month and reduced inflammatory reactions for 1 week in AD murine models without producing inflammation, microhemorrhage or systemic histotoxicity. These facts suggest that intracerebroventricular anti-Abeta may be a safe method for the rapid clearance of pre-existing Abeta and retarding reaccumulation of Abeta in AD. Intracerebroventricular administration via a catheter and reservoir, may be combined with the development of humanized monoclonal antibody against Abeta. Intraventricular shunts and ventriculostomy are frequently employed with acceptable risk-to-benefit ratios in the treatment of various brain disorders, while humanized antibodies are currently used in clinical trials of brain diseases such as multiple sclerosis and lymphoma.     

Function and phenotype of immature CD4+ lymphocytes in healthy infants and early lymphocyte activation in uninfected infants of human immunodeficiency virus-infected mothers.

The function and phenotypes of CD4+ lymphocytes in infants are different than in adults and are modulated by maturational changes and exposure to environmental antigens. Infants of non-human immunodeficiency virus (HIV)-infected mothers and uninfected infants of HIV-infected mothers, 0 to 6 months of age, were examined for CD4+ lymphocyte function by in vitro interleukin-2 (IL-2) production and for CD4+ phenotypes by three-color flow cytometry. A minority of these uninfected infants (28%) had functional responses similar to those of healthy adult women (IL-2 production in response to anti-CD3, alloantigen, and mitogen), while the remainder were capable of responding to alloantigen and mitogen but not to anti-CD3. We did demonstrate reduced phytohemagglutinin-stimulated IL-2 production in uninfected infants born to HIV-seropositive mothers compared to that in infants from seronegative mothers. The proportions of CD3+ CD4+, CD4+ HLA-DR- CD38+, and CD4+ CD45RA+ RO- (naive) lymphocytes were much higher in infants than in adults, and the proportions of CD4+ CD45RA- RO+ (memory) and CD4+ CD25+ (IL-2 receptor-bearing) lymphocytes were lower in infants than in adults. The proportions of activated (CD4+ HLA-DR+ CD38+) and memory (CD4+ CD45RA- RO+) lymphocytes were increased in uninfected infants of HIV-infected mothers compared to infants of uninfected mothers. Therefore, T-helper-cell function is immature in many infants, but the CD4+ lymphocytes of some HIV-exposed, uninfected infants have been stimulated by antigen at an early age.     

Levels of quality management of blood transfusion services in Europe

The European blood legislation has defined several key quality elements to achieve Good Manufacturing Practice (GMP) in the field of blood transfusion. During the recent years, the blood legislation is in the process of implementation throughout its member states. Following the Directive 2002/98/EC, Directive 2005/62/EC has given further requirements for quality-management systems to be fulfilled by blood establishments. In addition, GMP/Good Laboratory Practice (GLP) and ISO standards are used inter alia by blood establishments. In order to support the implementation of the blood legislation, the European Public Health Work Plan (2005/2007) has cofunded two projects, led by the German Red Cross and supported by the European Blood Alliance, delivering a common European Standard Operating Procedure (SOP) methodology (EU-Q-Blood-SOP) and criteria and standards for the inspection of blood establishments (EUBIS). The EU-SOP manual will assist blood establishments in preparing for the inspection of their services related to the implementation of quality relevant elements required by the EU Directive 2002/98/EC and its technical annexes. The standards and criteria for inspection of blood establishments will cross-reference existing quality standards to the directive requirements and define requirements for the structure of quality-management systems based on the directive 2002/98/EC and its technical annexes. Based on these requirements, inspection standards and criteria are developed to assist in the independent assessment of quality systems established by individual blood establishments. These assessments are done in relation to the requirements defined by the European Union legislation on blood, in order to safeguard the quality of blood and to achieve continuous improvement of its quality throughout Europe.     

Sensation Seeking and Cortical Augmenting-Reducing

The experiment was designed to establish the relationship between the Sensation Seeking Scales (SSS) and cortical augmenting-reducing. Forty-nine male undergraduate Ss were used. Ss were presented with five intensities of light flashes in randomly presented blocks of trials at each intensity. Averaged evoked response (AER) amplitudes were measured at each intensity of light. Augmenting-reducing was measured for each S as the slope of the relationship between stimulus intensity and amplitude of response. This slope measure correlated very significantly (r= .59) with the Disinhibition subscale of the SSS and positively, but not significantly, with other subscales. Comparing the low and high scorers on the Disinhibition scale, a significant interaction between groups and stimulus intensities was found but no main effects of stimulus intensity or groups were found. The high Disinhibitors did not differ from the lows at the low stimulus intensities but did differ significantly at the highest intensity where the lows showed a marked reducing tendency. The results show an interesting convergence between the Disinhibition type of sensation seeking, manic tendencies, and the AER.     

Development of Coombs positive haemolytic anaemia in NZB mice injected with Freund's complete adjuvant

Intraperitoneal injection of young adult NZB mice with Freund's complete adjuvant emulsion, alone or incorporating BALB/c mouse erythrocytes, resulted in accelerated direct Coombs conversion and development of splenomegaly. Incorporation of adjuvant with NZB erythrocytes caused an accelerated reactivity in some animals, but in others resulted in a delayed Coombs conversion. In contrast, positive Coombs tests failed to develop at all in NZB mice injected with a Freund's complete adjuvant preparation containing ten times the usual amount of Mycobacterium. These observations might be explained in part by the relative influences of adjuvancy and of antigenic competition on the availability of precursor cells capable of responding to autoantigenic stimulation.     

Permeation of human ovarian tissue with cryoprotective agents in preparation for cryopreservation

The recent improvements in the treatment of cancer by chemo- and radiotherapy have led to a significant increase in the survival rates of patients with malignant disease, but at the expense of distressing side effects. One major problem, especially for younger patients, is that aggressive therapy destroys a significant proportion of the follicular population, which can result in either temporary or permanent infertility. Freeze-banking pieces of ovarian cortex prior to treatment is one strategy for preserving fecundity. When the patient is in remission, fertility could, theoretically, be restored by autografting the thawed tissue at the orthotopic site or by growing isolated follicles to maturity in vitro. Recent studies have found good follicular survival in frozen-thawed human ovarian tissue but to optimize the process an effective cryopreservation method needs to be developed. An essential part of such a technique is to permeate the tissue with a cryoprotectant to minimize ice formation and the extent of this equilibration is an important determinant of post-thaw cellular survival. In the current study, we have investigated the diffusion of four cryoprotective agents into human tissue at both 4 degrees C and 37 degrees C. We have also studied the effect of adding different concentrations of the non penetrating cryoprotective agent, sucrose, to the freezing media using the release of lactate dehydrogenase as a measure of its protective effect. At 4 degrees C propylene glycol and glycerol penetrated the tissue significantly slower than either ethylene glycol or dimethyl sulphoxide. At the higher temperature of 37 degrees C all four cryoprotectants penetrated at a faster rate, however concern about enhanced toxicity prevents the use of these conditions in practice. Thus, the results suggest that the best method of preparing tissue for freezing is exposure for 30 min to 1.5 M solutions of ethylene glycol or dimethyl sulphoxide at 4 degrees C; this achieved a mean tissue concentration that was almost 80% that of the bathing solution. We also report that the addition of low concentrations of sucrose to the freezing medium does not have a significant protective effect against freezing injury.