The role of nitric oxide in the pathogenesis of spontaneous murine autoimmune disease: increased nitric oxide production and nitric oxide synthase expression in MRL-lpr/lpr mice, and reduction of spontaneous glomerulonephritis and arthritis by orally administered NG-monomethyl-L-arginine.

MRL-lpr/lpr mice spontaneously develop various manifestations of autoimmunity including an inflammatory arthropathy and immune complex glomerulonephritis. This study examines the role of nitric oxide, a molecule with proinflammatory actions, in the pathogenesis of MRL-lpr/lpr autoimmune disease. MRL-lpr/lpr mice excreted more urinary nitrite/nitrate (an in vivo marker of nitric oxide production) than did mice of normal strains and MRL-(+/+) and B6-lpr/lpr congenic strains. In addition, MRL-lpr/lpr peritoneal macrophages had an enhanced capacity to produce nitric oxide in vitro as well as increased nitric oxide synthase activity, and certain tissues from MRL-lpr/lpr mice had increased expression of inducible nitric oxide synthase (NOS) mRNA and increased amounts of material immunoreactive for inducible NOS. Oral administration of NG-monomethyl-L-arginine, a nitric oxide synthase inhibitor, prevented the development of glomerulonephritis and reduced the intensity of inflammatory arthritis in MRL-lpr/lpr mice. By using interspecific backcross mice, the gene for inducible NOS (Nosi) was mapped to mouse chromosome 11. This chromosomal localization was different from those loci that we have previously demonstrated to be linked to enhanced susceptibility to renal disease in an MRL-lpr/lpr cross. However, the chromosomal location of the NOS gene was consistent with an insulin-dependent diabetes locus identified in an analysis of nonobese diabetic (NOD) mice. These results suggest that elevated nitric oxide production could be important in the pathogenesis of autoimmunity, and that treatments to block the production of nitric oxide or block its effects might be valuable therapeutically.

Mitotic chromatin condensation in vitro using somatic cell extracts and nuclei with variable levels of endogenous topoisomerase II.

We report the development of a new method for producing mitotic extracts from tissue culture cells. These extracts reproducibly promote the condensation of chromatin in vitro when incubated with purified interphase nuclei. This condensation reaction is not species specific, since nuclei from chicken, human, and hamster cell lines all undergo chromatin condensation upon incubation with the extract. We have used this extract to investigate the role of DNA topoisomerase II (topo II) in the chromosome condensation process. Chromatin condensation does not require the presence of soluble topo II in the mitotic extract. However, the extent of formation of discrete chromosome-like structures correlates with the level of endogenous topo II present in the interphase nuclei. Our results further suggest that chromatin condensation in this extract may involve two processes: chromatin compaction and resolution into discrete chromosomes.

ScII: an abundant chromosome scaffold protein is a member of a family of putative ATPases with an unusual predicted tertiary structure.

Here, we describe the cloning and characterization of ScII, the second most abundant protein after topoisomerase II, of the chromosome scaffold fraction to be identified. ScII is structurally related to a protein, Smc1p, previously found to be required for accurate chromosome segregation in Saccharomyces cerevisiae. ScII and the other members of the emerging family of SMC1-like proteins are likely to be novel ATPases, with NTP-binding A and B sites separated by two lengthy regions predicted to form an alpha-helical coiled-coil. Analysis of the ScII B site predicted that ScII might use ATP by a mechanism similar to the bacterial recN DNA repair and recombination enzyme. ScII is a mitosis-specific scaffold protein that colocalizes with topoisomerase II in mitotic chromosomes. However, ScII appears not to be associated with the interphase nuclear matrix. ScII might thus play a role in mitotic processes such as chromosome condensation or sister chromatid disjunction, both of which have been previously shown to involve topoisomerase II.

Hippocampal neurons encode information about different types of memory episodes occurring in the same location.

Firing patterns of hippocampal complex-spike neurons were examined for the capacity to encode information important to the memory demands of a task even when the overt behavior and location of the animal are held constant. Neuronal activity was recorded as rats continuously alternated left and right turns from the central stem of a modified T maze. Two-thirds of the cells fired differentially as the rat traversed the common stem on left-turn and right-turn trials, even when potentially confounding variations in running speed, heading, and position on the stem were taken into account. Other cells fired differentially on the two trial types in combination with behavioral and spatial factors or appeared to fire similarly on both trial types. This pattern of results suggests that hippocampal representations encode some of the information necessary for representing specific memory episodes.

Does assimilation into schemas involve systems or cellular consolidation? It's not just time.

A comment by Rudy and Sutherland [Rudy, J. R., & Sutherland, R. J. (2008). Is it systems or cellular consolidation? Time will tell. An alternative interpretation of the Morris Group's recent Science Paper. Neurobiology of Learning and Memory] has suggested an alternative account of recent findings concerning very rapid systems consolidation as described in a recent paper by Tse et al [Tse, D., Langston, R. F., Kakeyama, M., Bethus, I., Spooner, P. A., & Wood, E. R., et al. (2007). Schemas and memory consolidation. Science, 316, 76-82]. This is to suppose that excitotoxic lesions of the hippocampus cause transient disruptive neural activity outside the target structure that interferes with cellular consolidation in the cortex. We disagree with this alternative interpretation of our findings and cite relevant data in our original paper indicating why this proposal is unlikely. Various predictions of the two accounts are nonetheless outlined, together with the types of experiments needed to resolve the issue of whether systems consolidation can occur very rapidly when guided by activated neural schemas.

The characterization of novel, dual ErbB-2/EGFR, tyrosine kinase inhibitors: potential therapy for cancer.

The type I receptor tyrosine kinases constitute a family of transmembrane proteins involved in various aspects of cell growth and survival and have been implicated in the initiation and progression of several types of human malignancies. The best characterized of these proteins are the epidermal growth factor receptor (EGFR) and ErbB-2 (HER-2/neu). We have developed potent quinazoline and pyrido-[3,4-d]-pyrimidine small molecules that are dual inhibitors of ErbB-2 and EGFR. The compounds demonstrate potent in vitro inhibition of the ErbB-2 and EGFR kinase domains with IC(50)s <80 nM. Growth of ErbB-2- and EGFR-expressing tumor cell lines is inhibited at concentrations <0.5 microM. Selectivity for tumor cell growth inhibition versus normal human fibroblast growth inhibition ranges from 10- to >75-fold. Tumor growth in mouse s.c. xenograft models of the BT474 and HN5 cell lines is inhibited in a dose-responsive manner using oral doses of 10 and 30 mg/kg twice per day. In addition, the tested compounds caused a reduction of ErbB-2 and EGFR autophosphorylation in tumor fragments from these xenograft models. These data indicate that these compounds have potential use as therapy in the broad population of cancer patients overexpressing ErbB-2 and/or EGFR.

Fibronectin induction abrogates the BRAF inhibitor response of BRAF V600E/PTEN-null melanoma cells.

The mechanisms by which some melanoma cells adapt to Serine/threonine-protein kinase B-Raf (BRAF) inhibitor therapy are incompletely understood. In the present study, we used mass spectrometry-based phosphoproteomics to determine how BRAF inhibition remodeled the signaling network of melanoma cell lines that were BRAF mutant and PTEN null. Short-term BRAF inhibition was associated with marked changes in fibronectin-based adhesion signaling that were PTEN dependent. These effects were recapitulated through BRAF siRNA knockdown and following treatment with chemotherapeutic drugs. Increased fibronectin expression was also observed in mouse xenograft models as well as specimens from melanoma patients undergoing BRAF inhibitor treatment. Analysis of a melanoma tissue microarray showed loss of PTEN expression to predict for a lower overall survival, with a trend for even lower survival being seen when loss of fibronectin was included in the analysis. Mechanistically, the induction of fibronectin limited the responses of these PTEN-null melanoma cell lines to vemurafenib, with enhanced cytotoxicity observed following the knockdown of either fibronectin or its receptor α5ß1 integrin. This in turn abrogated the cytotoxic response to BRAF inhibition via increased AKT signaling, which prevented the induction of cell death by maintaining the expression of the pro-survival protein Mcl-1. The protection conveyed by the induction of FN expression could be overcome through combined treatment with a BRAF and PI3K inhibitor.

Neurotoxic hippocampal lesions have no effect on odor span and little effect on odor recognition memory but produce significant impairments on spatial span, recognition, and alternation.

Recent work has shown that lesions of the hippocampus in monkeys cause deficits in the capacity to remember increasing numbers of objects, colors, and spatial locations (). However, others have observed that hippocampectomized monkeys can show intact memory for a list of objects or locations (). We wished to explore the effects of hippocampal damage on the capacity of memory in the rodent and, to do so, developed novel "span" tasks in which a variable number of odors or locations had to be remembered. In the odor span task (experiment 1), rats were trained on a nonmatching to sample task in which increasing numbers of odors had to be remembered. Half of the trained rats received ibotenic acid lesions of the hippocampus. Postoperatively, hippocampectomized animals did not differ from control animals even when required to remember up to 24 odors. However, when tested on delayed retention of a list of 12 odors, rats with hippocampal lesions were impaired at a long delay. Also, these rats were impaired on a subsequent test of delayed spatial alternation. In a spatial span task (experiment 2), naive rats were trained on a nonmatching to sample task in which a variable number of locations had to be remembered. After this, half of the animals received ibotenic acid lesions. Postoperatively, hippocampectomized animals performed above chance levels when required to remember a single cup location, but were unable to remember more. Subsequent testing on another spatial delayed alternation task suggested that hippocampectomized rats could recognize, but could not inhibit their approach to previously visited locations.

Acute care costs of the oldest old: they cost less, their care intensity is less, and they go to nonteaching hospitals.

BACKGROUND: The cost of acute hospital care is often believed to increase with age among older persons. Our clinical experience in the acute hospital setting suggested that people aged 90 years and older may be a distinct cohort who have different health care needs and who use health resources differently from younger aged groups. METHODS: To determine the cost of care for the oldest old and younger old patients in Massachusetts acute care hospitals, 1992 and 1993 discharge data from all nonfederal Massachusetts hospitals were examined (678 954 discharges) according to five age groups: 60 to 69 years (n=210 270), 70 to 79 years (n=256 781), 80 to 89 years (n=171 725), 90 to 99 years (n=39 170), and 100 or more years (n=1008). Average estimated total and ancillary costs per discharge and per diagnosis related group were calculated. Differences by gender and survivorship were also examined. RESULTS: Hospitalization costs peaked in the 70- to 79-year age group and declined with age thereafter. Case mix was an important determinant of this trend. Despite lower cost per stay, average length of stay was longer for the oldest age groups. Ancillary costs accounted for 53% of the total costs per stay among the 60- to 69-year-olds and only 32% among the 100 or more-year- olds. For hospitalizations during which the patient died, the average cost per discharge decreased 61%, from $16886 for 60- to 69-year-olds to $6523 for centenarians. Costs were greater for decedents than for survivors, although these differences decreased dramatically with increasing age. Those aged 80 years and older tended to be hospitalized in nonteaching hospitals. CONCLUSIONS: In the acute hospital setting, the oldest old cost less per admission than younger elderly patients. This finding must be considered when future health care costs are predicted for this fastest growing segment of our population.

The effects of the novel, reversible epidermal growth factor receptor/ErbB-2 tyrosine kinase inhibitor, GW2016, on the growth of human normal and tumor-derived cell lines in vitro and in vivo.

The epidermal growth factor receptor (EGFR) and ErbB-2 transmembrane tyrosine kinases are currently being targeted by various mechanisms in the treatment of cancer. GW2016 is a potent inhibitor of the ErbB-2 and EGFR tyrosine kinase domains with IC50 values against purified EGFR and ErbB-2 of 10.2 and 9.8 nM, respectively. This report describes the efficacy in cell growth assays of GW2016 on human tumor cell lines overexpressing either EGFR or ErbB-2: HN5 (head and neck), A-431 (vulva), BT474 (breast), CaLu-3 (lung), and N87 (gastric). Normal human foreskin fibroblasts, nontumorigenic epithelial cells (HB4a), and nonoverexpressing tumor cells (MCF-7 and T47D) were tested as negative controls. After 3 days of compound exposure, average IC50 values for growth inhibition in the EGFR- and ErbB-2-overexpressing tumor cell lines were < 0.16 microM. The average selectivity for the tumor cells versus the human foreskin fibroblast cell line was 100-fold. Inhibition of EGFR and ErbB-2 receptor autophosphorylation and phosphorylation of the downstream modulator, AKT, was verified by Western blot analysis in the BT474 and HN5 cell lines. As a measure of cytotoxicity versus growth arrest, the HN5 and BT474 cells were assessed in an outgrowth assay after a transient exposure to GW2016. The cells were treated for 3 days in five concentrations of GW2016, and cell growth was monitored for an additional 12 days after removal of the compound. In each of these tumor cell lines, concentrations of GW2016 were reached where outgrowth did not occur. Furthermore, growth arrest and cell death were observed in parallel experiments, as determined by bromodeoxyuridine incorporation and propidium iodide staining. GW2016 treatment inhibited tumor xenograft growth of the HN5 and BT474 cells in a dose-responsive manner at 30 and 100 mg/kg orally, twice daily, with complete inhibition of tumor growth at the higher dose. Together, these results indicate that GW2016 achieves excellent potency on tumor cells with selectivity for tumor versus normal cells and suggest that GW2016 has value as a therapy for patients with tumors overexpressing either EGFR or ErbB-2.

Impaired object recognition memory in rats following ischemia-induced damage to the hippocampus.

Transient cerebral ischemia can produce irreversible neuronal damage and permanent learning and memory impairments in humans. This study examined whether ischemia-induced brain damage in rats results in impairments on the delayed nonmatching-to-sample (DNMS) task, a nonspatial recognition task analogous to tests on which amnesic patients display impairments. Male Wistar rats received either sham surgery or 20-min forebrain ischemia induced by bilateral carotid occlusion and hypotension. Four weeks after surgery, ischemic rats were significantly impaired in both learning and performing the DNMS task at retention intervals up to 5 min. Extensive presurgical training did not reduce this impairment. Observable cell loss in ischemic rats was limited to CA1 pyramidal neurons and a subset of cells in the dentate gyrus. The results indicate that ischemic damage to the hippocampus in rats results in recognition memory deficits similar to those produced by ischemic damage in humans.

Ischemia-induced object-recognition deficits in rats are attenuated by hippocampal ablation before or soon after ischemia.

The literature on the role of the hippocampus in object-recognition contains a paradox: Transient forebrain ischemia (ISC) produces hippocampal damage and severe deficits on the delayed nonmatching-to-sample (DNMS) task, yet hippocampal ablation (ABL) produces milder deficits. Experiment 1 confirmed that pretrained rats display severe DNMS deficits following ISC, but not ABL. Ischemia produced loss of CA1 neurons, but no obvious extrahippocampal damage. In Experiments 2 and 3, ISC rats from Experiment 1 received ABL, and ABL rats received ISC; neither treatment affected DNMS performance. In Experiment 4, rats that received ISC followed 1 hr later by ABL displayed only mild deficits. It is hypothesized that ISC-induced DNMS deficits are due to extrahippocampal damage produced by pathogenic processes that involve the hippocampus.

A glycine antagonist 7-chlorokynurenic acid attenuates ischemia-induced learning deficits.

Transient global ischemia can result in permanent neuronal damage and impairments in learning and memory. We investigated the therapeutic potential of 7-Chlorokynurenic acid, a potent antagonist at the glycine-modulatory site on the NMDA receptor, in terms of both neuroprotection and behavioral outcome in rats following transient forebrain ischemia. Intraventricular administration of the drug immediately before ischemia significantly attenuated ischemia-induced CA1 pyramidal cell loss. Moreover, ischemic rats treated with 7-Chlorokynurenic acid showed unimpaired acquisition of a delayed nonmatching to sample task 8 weeks following surgery, whereas saline-treated ischemic rats were significantly impaired. These data provide preliminary evidence that the glycine site may be an appropriate target for therapeutic agents in ischemia.

Extracellular dopamine in the rat striatum during ischemia and reperfusion as measured by in vivo electrochemistry and in vivo microdialysis.

The effects of transient global forebrain ischemia and reperfusion on striatal extracellular dopamine levels were analyzed using both in vivo electrochemistry and in vivo microdialysis in urethane-anesthetized rats. Electrochemical records showed that extracellular dopamine levels increased once during the period of ischemia, and a second time during reperfusion. This biphasic pattern was not detected by microdialysis, probably because of the relatively low time resolution of this technique. Microdialysis provided evidence that the voltammetric signal was a measure of dopamine, and also allowed measurement of the metabolites dihydroxyphenylacetic acid and homovanillic acid, both of which decreased during ischemia. The biphasic dopamine pattern seen in rats is similar to that reported previously in gerbils, suggesting that it is a phenomenon common to transient ischemia and reperfusion across different species and models of transient global ischemia. This phenomenon may have important implications for therapeutic intervention in cerebral ischemia.

A glycine antagonist reduces ischemia-induced CA1 cell loss in vivo.

Excessive activation of the N-methyl-D-aspartate (NMDA) receptor-channel complex has been implicated as one of the mechanisms by which ischemia-induced neuronal damage is mediated. Elevated glycine levels during ischemia may contribute to damage mediated by the NMDA receptor as glycine binding potentiates NMDA responses, and may be necessary for channel opening. We investigated the protective effects of 7-chlorokynurenic acid--a competitive antagonist at the glycine binding site associated with the NMDA receptor--against hippocampal CA1 cell loss induced by transient forebrain ischemia in rats. Intraventricular administration of the drug immediately before the onset of ischemia significantly attenuated neuronal loss compared to vehicle-treated animals.

Weight loss maintenance 1 year after individual counseling.

Many individual and group weight loss programs can boast short-term success. There has been little documentation on the long-term success of any particular program. A study was conducted to examine total weight loss for individually counseled patients seen in a hospital outpatient nutrition clinic February to October 1986. All patients who lost weight were contacted via telephone 1 year later to ascertain weight loss maintenance. During this period, 157 patients were seen. In an average of five visits, 119 (75%) lost an average of 9 lb. One year later, 73 (61%) of these patients were contacted, 54 of whom (74%) reported that they had either maintained or lost additional weight during the year. Sixteen (30%) of these 54 individuals lost additional weight during the year--an average of 13 additional lb. Results were similar for men and women. Although based on a small sample size, the results suggest that weight loss counseling can be successful in maintaining as well as losing weight. Such results should encourage dietetic practitioners to track their own patients for long-term results and to share the results with physicians, administrators, and insurers who are concerned about whether their money has been well spent.

Evaluation of a hospital-based education program for patients with diabetes.

Ninety-three hospitalized patients with diabetes were followed for a 4-month period after discharge. Fifty-three of them attended an inpatient diabetes education program. The primary objectives of this research were to (a) determine the effectiveness of the program in improving diabetes-related behaviors and outcomes of the patients attending the program and (b) study the effects of time on those behaviors and outcomes. Examples of self-care behaviors included eating three meals per day, daily urine testing, and maintaining an exercise program. Outcome measures included reduced fasting blood glucose levels, reduced insulin requirements, fewer hospitalizations, and fewer emergency room visits. At 1 and 4 months after discharge, the group that attended the program reported better compliance for all self-care behaviors than did the group that did not attend the program. Outcomes were also better for the group that attended the education program.

A recurring question--how many beds for dementia?

Experience at Crichton Royal in dealing with dementia over a thirty year period has been used to develop two models of bed requirements for dementia sufferers. The first based on results from the periods 1957 to 1959 and 1974 to 1976 and the second on experience in the years 1974-75 and 1984-85. The variables considered were demographic change, admission of a constant proportion of the at risk group and changes in individual patient survival between groups of patients admitted. Projections based on the earlier model suggested that the SHAPE provision of 10 beds per 1000 of the over 64 population would be insufficient to maintain established standards of care. The later model, however, finds SHAPE numbers appropriate. The reasons for the change of view--inaccuracy in estimate of increasing male survival and significant under-estimates of population growth--nullify each other. The conclusion is that SHAPE is a useful model provided the demographic changes and patient survival change are carefully monitored.

Psychogeriatrics 1974 to 1984. Expanding problems and fixed resources.

The impact of a 16% increase between censuses in the population aged 65 and over, on a psychogeriatric department whose bed complement remained static between 1974 and 1984, has been studied. Demand for service, as measured by new referrals, rose by 150%, while admissions fell by 14%. The increase in new referrals was uniform across the diagnostic spectrum, but the fall in admissions was not. Functional admissions fell in all age-groups except that of women of 85 and over, while organic-case admissions other than for cases of dementia virtually collapsed. Overall admissions for dementia rose to the predicted level, but the distribution of the increase was irregular and unexpected. Admissions of males and females aged 65-74 and females of 85 and over fell relatively and absolutely, while those of women aged 75-84 and men of 85 and over were little changed. Only admissions of men aged 75-84 alone increased in real terms.