Evaluation of biochemical changes during in vivo erythrocyte senescence in the dog

One hypothesis to explain the age-dependent clearance of red blood cells (RBCs) from circulation proposes that denatured/oxidized hemoglobin (hemichromes) arising late during an RBC's life span induces clustering of the integral membrane protein, band 3. In turn, band 3 clustering generates an epitope on the senescent cell surface leading to autologous IgG binding and consequent phagocytosis. Because dog RBCs have survival characteristics that closely resemble those of human RBCs (ie, low random RBC loss, approximately 115-day life span), we decided to test several aspects of the above hypothesis in the canine model, where in vivo aged cells of defined age could be evaluated for biochemical changes. For this purpose, dog RBCs were biotinylated in vivo and retrieved for biochemical analysis at various later dates using avidin-coated magnetic beads. Consistent with the above hypothesis, senescent dog RBCs were found to contain measurably elevated membrane-bound (denatured) globin and a sevenfold enhancement of surface-associated autologous IgG. Interestingly, dog RBCs that were allowed to senesce for 115 days in vivo also suffered from compromised intracellular reducing power, containing only 30% of the reduced glutathione found in unfractionated cells. Although the small quantity of cells of age >/=110 days did not allow direct quantitation of band 3 clustering, it was nevertheless possible to exploit single-cell microdeformation methods to evaluate the fraction of band 3 molecules that had lost their normal skeletal linkages and were free to cluster in response to hemichrome binding. Importantly, band 3 in RBCs >/=112 days old was found to be 25% less restrained by skeletal interactions than band 3 in control cells, indicating that the normal linkages between band 3 and the membrane skeleton had been substantially disrupted. Interestingly, the protein 4.1a/protein 4.1b ratio, commonly assumed to reflect RBC age, was found to be maximal in RBCs isolated only 58 days after labeling, implying that while this marker is useful for identifying very young populations of RBCs, it is not a very sensitive marker for canine senescent RBCs. Taken together, these data argue that several of the readily testable elements of the above hypothesis implicating band 3 in human RBC senescence can be validated in an appropriate canine model.     

Differential reactivity of the rat S100A4(p9Ka) gene to sodium bisulfite is associated with differential levels of the S100A4 (p9Ka) mRNA in rat mammary epithelial cells

Elevated intracellular levels of S100A4, an S100-related calcium-binding protein, induce metastatic capability in benign mammary tumor-derived epithelial cells and in transgenic mice bearing oncogene-induced benign mammary tumors. The S100A4(p9Ka) gene in rat mammary epithelial cells expressing low levels of S100A4 yields a reduced number of fragments upon digestion with the methylation-sensitive restriction enzyme, HpaII, compared with the gene from high S100A4-expressing cells. Genomic sequencing of two potential regulatory elements in the S100A4 gene, an intronic enhancer and TATA box region, revealed that in low S100A4-expressing cells, most cytosine bases exhibited high levels of resistance to conversion to thymine by sodium bisulfite. In derivative cell lines, which express high levels of S100A4, only a small number of cytosine bases were resistant to treatment with sodium bisulfite. In contrast, cytosine bases in the DNA surrounding an upstream regulatory region, which binds inhibitory GC factor in the low-expressing cell lines, are sensitive to conversion to thymine by sodium bisulfite in both low- and high-expressing cell lines. The results suggest that the rat S100A4 gene is maintained in a different state in the low-expressing cell lines and that this state might be a consequence of the pattern of methylation in this regulated gene that does not contain a CpG island.     

Computer viruses

Computer viruses have been around since the mid 1980s. Over 40,000 different viruses have been cataloged so far and the number of viruses is increasing dramatically. The damage they cause is estimated to be several billions of U.S. dollars per year. Most often, the origin of the virus is difficult to trace. Various kinds of anti-virus software have been developed which detect viruses and take corrective actions. The anti-virus software needs to be continually updated to cope with newer types of viruses. The proliferation of the Internet and Web, have enabled viruses to spread quickly on a massive scale, by taking advantage of several security loopholes. The continual challenge is to have quick and effective responses to these virus attacks     

Intestinal tuberculosis

Since the symptoms and clinical presentation of intestinal tuberculosis is nonspecific, the diagnosis is frequently delayed and may be achieved at autopsy only. Intestinal tuberculosis is very rare in Denmark, but may now be seen more often because of increasing numbers of immigrants from countries of the third world with a high prevalence of tuberculosis. A case of intestinal tuberculosis in a 28 year old Somalian female is reported. Methods of diagnosing intestinal tuberculosis are commented, and the frequent necessity of starting medical treatment before a bacteriological diagnosis is reached is emphasized.     

Stromal estrogen receptors mediate mitogenic effects of estradiol on uterine epithelium

Estradiol-17beta (E2) acts through the estrogen receptor (ER) to regulate uterine growth and functional differentiation. To determine whether E2 elicits epithelial mitogenesis through epithelial ER versus indirectly via ER-positive stromal cells, uteri from adult ER-deficient ER knockout (ko) mice and neonatal ER-positive wild-type (wt) BALB/c mice were used to produce the following tissue recombinants containing ER in epithelium (E) and/or stroma (S), or lacking ER altogether: wt-S + wt-E, wt-S + ko-E, ko-S + ko-E, and ko-S + wt-E. Tissue recombinants were grown for 4 weeks as subrenal capsule grafts in intact female nude mice, then the hosts were treated with either E2 or oil a week after ovariectomy. Epithelial labeling index and ER expression were determined by [3H]thymidine autoradiography and immunohistochemistry, respectively. In tissue recombinants containing wt-S (wt-S + wt-E, wt-S + ko-E), E2 induced a similar large increase in epithelial labeling index compared with oil-treated controls in both types of tissue recombinants despite the absence of epithelial ER in wt-S + ko-E tissue recombinants. This proliferative effect was blocked by an ER antagonist, indicating it was mediated through ER. In contrast, in tissue recombinants prepared with ko-S (ko-S + ko-E and ko-S + wt-E), epithelial labeling index was low and not stimulated by E2 despite epithelial ER expression in ko-S + wt-E grafts. In conclusion, these data demonstrate that epithelial ER is neither necessary nor sufficient for E2-induced uterine epithelial proliferation. Instead, E2 induction of epithelial proliferation appears to be a paracrine event mediated by ER-positive stroma. These data in the uterus and similar studies in the prostate suggest that epithelial mitogenesis in both estrogen and androgen target organs are stromally mediated events.     

Contemporary percutaneous treatment of unprotected left main coronary stenoses: initial results from a multicenter registry analysis 1994-1996

BACKGROUND: Coronary artery bypass surgery (CABG) has been considered the therapy of choice for patients with unprotected left main (ULMT) coronary stenoses. Selected single-center reports suggest that the results of percutaneous intervention may now approach those of CABG. METHODS AND RESULTS: To assess the results of percutaneous ULMT treatment from a wide variety of experienced interventional centers, we requested data on consecutive patients treated after January 1, 1994, from 25 centers. One hundred seven patients were identified who were treated either electively (n=91) or for acute myocardial infarction (n=16). Of patients treated electively, 25% were considered inoperable, and 27% were considered high risk for bypass surgery. Primary treatment included stents (50%), directional atherectomy (24%), and balloon angioplasty (20%). Follow-up was 98.8% complete at 15+/-8 months. Results varied considerably, depending on presentation and treatment. For patients with acute myocardial infarction, technical success was achieved in 75%, and survival to hospital discharge was 31%. For elective patients, technical success was achieved in 98.9%, and in-hospital survival was strongly correlated with left ventricular ejection fraction (P=.003). Longer-term event (death, infarction, or bypass surgery) -free survival was correlated with ejection fraction (P<.001) and was inversely related to presentation with progressive or rest angina (P<.001). Surgical candidates with ejection fractions > or = 40% had an in-hospital survival of 98% and a 9-month event-free survival of 86+/-5%, whereas patients with ejection fractions < 40% had 67% and 22+/-12% in-hospital and 9-month event-free survivals, respectively. Nine hospital survivors (10.6%) experienced cardiac death within 6 months of hospital discharge. CONCLUSIONS: While results for selected patients appear promising, until early post-hospital discharge cardiac death can be better understood and minimized, percutaneous revascularization of ULMT stenosis should not be considered an alternative to bypass surgery for most patients. When percutaneous revascularization of ULMT is required, directional atherectomy and stenting appear to be the preferred techniques, and follow-up angiography 6 to 8 weeks after treatment is probably advisable.     

Report from the Maryland Epidemiology Schizophrenia Linkage Study: no evidence for linkage between schizophrenia and a number of candidate and other genomic regions using a complex dominant model

Our collaborative group has undertaken a linkage study of schizophrenia, using a systematic sample of patients admitted to Maryland hospitals. An initial sample of 39 families, each having two or more affecteds, was available for genotyping candidate genes, candidate regions, and highly polymorphic markers randomly distributed throughout the genome. We used a single complex dominant model (with a disease gene frequency of 0.005 and age-dependent penetrance for affected phenotype: for under 35, penetrance = .45; for 35 and older, penetrance = .85). We report here 130 markers, which met the exclusion criteria of LOD score < -2.00 at theta > 0.01 in at least 10 informative families, and no evidence for heterogeneity. We also report here markers that were tested as candidates for linkage to the schizophrenic phenotype. They were selected based on the following criteria: a) proximity to reported chromosomal rearrangements (both 5q and 11q), b) suggestions of linkage from other families (5q), or c) presence of a candidate gene (5q, 11q, 3q: Dopamine receptors 1, 2, and 3, respectively). We also tested for mutations of codon 717 in exon 17 of the amyloid precursor protein (APP) gene and were unable to detect the C to T substitution in our schizophrenic group.     

Ethylene glycol poisoning with a normal anion gap caused by concurrent ethanol ingestion: importance of the osmolal gap

Ethylene glycol poisoning classically presents as a metabolic acidosis with an increased anion gap. Metabolism of ethylene glycol to organic acids, and increased production of lactate, are responsible for the increased gap. We report the case of an alcohol user who consumed ethanol and ethylene glycol concurrently, and presented without acidosis, with a normal anion gap. Several hours later, when his serum ethanol level had declined, he developed severe acidosis with an elevated anion gap. An increased osmolal gap, not accounted for by the serum ethanol level, proved to be an important clue to the diagnosis. In this patient, ingestion of ethanol inhibited the hepatic metabolism of ethylene glycol to organic acids, obscuring the diagnosis. In intoxicated alcohol users, even in the absence of metabolic acidosis, serum osmolality measurements and calculation of the osmolal gap may facilitate the rapid diagnosis of ethylene glycol poisoning.     

Multi-component T1 relaxation and magnetisation transfer in peripheral nerve

Corneas of diabetic patients have abnormal healing and epithelial adhesion, which may be due to alterations of the corneal extracellular matrix (ECM) and basement membrane (BM). To identify such alterations, various ECM and BM components and integrin receptors were studied by immunofluorescence on sections of normal and diabetic human corneas. Age-matched corneas from 15 normal subjects, 10 diabetics without diabetic retinopathy (DR), and 12 diabetics with DR were used. In DR corneas, the composition of the central epithelial BM was markedly altered, compared to normal or non-DR diabetic corneas. In most cases the staining for entactin/nidogen and for chains of laminin-1 (alpha1beta1gamma1) and laminin-10 (alpha5beta1gamma1 was very weak, discontinuous, or absent over large areas. Other BM components displayed less frequent changes. The staining for alpha3beta1 (VLA-3) laminin binding integrin was also weak and discontinuous in DR corneal epithelium. Components of stromal ECM remained unchanged even in DR corneas. Therefore, distinct changes were identified in the composition of the epithelial BM in DR corneas. They may be due to increased degradation or decreased synthesis of BM components and related integrins. These alterations may directly contribute to the epithelial adhesion and wound healing abnormalities found in diabetic corneas.