Hippocampal scrapie infection impairs operant DRL performance in mice.

In differential reinforcement of low rates of responding (DRL) tasks, animals are trained to respond for rewards that become available only after some set time has elapsed since the animal's previous response. DRL performance is impaired by hippocampal lesions regardless of their precise location, and can be measured using automated operant equipment, whereas spatial tasks are selectively impaired by dorsal, but not ventral hippocampal lesions, and are typically conducted by hand. Earlier studies of prion infection following dorsal hippocampal micro-injections of scrapie have shown clear impairments of spatial alternation, but these occurred significantly later than dysfunction in hippocampus-dependent 'domestic' tasks such as nesting or burrowing. In the present experiment, mice were trained to respond on an automated DRL schedule prior to dorsal hippocampal ME7 scrapie injection. Post-operative DRL performance was monitored, along with performance on 'domestic' and other tests, which provided additional measures of disease progression. Animals with scrapie developed a clear DRL deficit at approximately the same time as their deficits on the other tests became apparent, and long before clinical signs were detectable. DRL deficits thus appeared earlier in the sequence of disease progression than previously reported for spatial alternation, suggesting that early signs of scrapie infection are caused in part by neuronal dysfunction extending beyond the dorsal hippocampal region of initial infection.

Measurement of von Willebrand factor activity: relative effects of ABO blood type and race.

Tests based on three different principles are reported to measure the activity of von Willebrand factor (VWF): ristocetin cofactor (VWF:RCo), collagen binding (VWF:CB), and the so-called "activity ELISA" (VWF:MoAb). We measured these and other diagnostic parameters in a population of 123 randomly selected female study controls, age 18-45 years. Type O subjects had significantly lower levels than non-O subjects in each test. Race differences were seen in all tests except VWF:RCo, with Caucasians having significantly lower levels than African-Americans. ABO differences accounted for 19% of the total variance in VWF:Ag (P < 0.0001) and race for 7% (P < 0.0001), for a total of 26%. Both effects were mediated through VWF:Ag and were independent. VWF:Ag level was the primary determinant of VWF function, accounting for approximately 60% of the variance in VWF:RCo and VWF:CB and 54% of the variance in factor VIII. The ratio VWF:RCo/VWF:Ag differed significantly by race within blood group. The median ratios were 0.97 for type O Caucasians vs. 0.79 for type O African-Americans and 0.94 for non-O Caucasians vs. 0.76 for non-O African-Americans. The ratio VWF:CB/VWF:Ag did not vary. This suggests racial differences in the interaction of VWF with GP1b but not with subendothelium. Alternatively, VWF:RCo may be regulated to maintain a relatively constant plasma level in the presence of excessive VWF:Ag. This heterogeneity within the normal population is partially responsible for the difficulty in defining diagnostic limits for von Willebrand disease.

Novel reduced benzo[j]fluranthen-3-ones from Cladosporium cf. cladosporioides with cytokine production and tyrosine kinase inhibitory properties.

A series of reduced benzo[j]fluoranthen-3-ones (1-4) was isolated from fermentations of a fungal strain CBUK20700 (CBS 100220), classified as Cladosporium cf. cladosporioides, during a microbial extract screening programme to identify inhibitors of anti-CD28-induced interleukin-2 (IL-2) production by Jurkat E6-1 cells as potential immunosuppressive agents. These compounds were also found to be tyrosine kinase inhibitors. The structures of compounds 1-4 were elucidated by spectroscopic methods including the HMQC, HMBC and NOESY NMR experiments. The most potent compound in the series, (6bS,7R,8S)-7-methoxy-4,8,9-trihydroxy-1,6b,7,8-tetrahydro-2H-benzo[j]fluoranthen-3-one (1) inhibited anti-CD28-induced IL-2 production and Abl tyrosine kinase with IC50 values of 400 and 60 nM respectively. The 6b-stereoisomeric 2 was a moderate inhibitor of both IL-2 production and Abl tyrosine kinase while the 8-oxo derivative 3 was inactive in both assays. The 8-O-methyl ether 4 was a moderate inhibitor of IL-2 production but exhibited potent inhibition of Abl tyrosine kinase with an IC50 of 45 nM.

Differential regulation of CD3- and CD28-induced IL-2 and IFN-gamma production by a novel tyrosine kinase inhibitor XR774 from Cladosporium cf. cladosporioides.

Inhibition of CD28 signalling after an immune response impedes T cell activation and can lead to immunosuppression. To identify inhibitors of anti-CD28 induced IL-2 production, a library of fungal metabolites was screened in a cell-based, high throughput assay. A reduced novel benzofluoranthene, tentatively named as (6bS, 7R, 8S)-7-methoxy-4, 8, 9-trihydroxy-1, 6b, 7, 8-tetrahydro-2H-benzo[j] fluoranthen-3-one (XR774), from Cladosporium cf. cladosporioides, was isolated. XR774 inhibited IL-2 mRNA and protein expression induced by anti-CD28 and anti-CD3 but had no effect on IL-2 induction by PMA and ionomycin. Moreover, XR774 inhibited the activity of the tyrosine kinases, Fyn, Lck, Abl and epidermal growth factor receptor (EGFR) with nanomolar activity, whereas micromolar concentrations of XR774 were ineffective on the serine-threonine kinase, PKA. Kinetic analysis of Fyn kinase inhibition was consistent with XR774 as a competitive inhibitor with respect to ATP. In peripheral blood, mononuclear cells (PBMC), XR774 inhibited anti-CD3 and anti-CD28 induced IL-2 and IL-2R alpha chain (CD25) expression but was consistently less active for inhibition of IFN-gamma production. On stimulation with PMA and anti-CD28, XR774 inhibited IL-2 production but had no effect on CD25 expression and enhanced IFN-gamma production. In contrast, the ansamycin, geldanamycin, inhibited both IL-2 and IFN-gamma production induced by anti-CD3 and anti-CD28 or PMA and anti-CD28. No significant associated cytotoxicity or inhibition of protein synthesis was observed at concentrations up to 14 microM. Thus, XR774 represents a novel class of pharmacological agent with selective biological activities that distinguish it from other natural product inhibitors, such as the ansamycins.

Venous thrombosis in relation to fibrinogen and factor VII genes among African-Americans.

We evaluated the relation between venous thrombosis and plasma fibrinogen levels, the HaeIII and BcI polymorphisms of the beta fibrinogen gene, and the MspI polymorphisms of the factor VII gene in a case-control study of African-Americans. The study included 91 venous thrombosis cases and 185 control subjects obtained from a hospital in Atlanta, Georgia. High plasma fibrinogen was associated with increased risk of venous thrombosis, but the finding was not statistically significant. There was little association between the HaeIII polymorphisms and the BclI polymorphisms and the risk of venous thrombosis. The prevalence of the M2/M2 genotype of the factor VII gene was higher among cases than controls, but the difference was not statistically significant. The prevalence of the HaeIII H2 allele and the BclI B2 allele of the beta fibrinogen gene, both of which have been associated with slightly higher levels of plasma fibrinogen in most studies, is considerably lower among African-Americans in this study than it is among Whites in the United States and among Northern Europeans. The study is limited by its small size. However, despite this limitation, it supports the belief that increased plasma fibrinogen levels are associated with increased venous thrombosis risk. The study also indicated that the HaeIII and the BclI polymorphisms of the beta fibrinogen gene and the MspI polymorphisms of the factor VII gene are not strong determinants of venous thrombosis.

The role of the t-PA I/D and PAI-1 4G/5G polymorphisms in African-American adults with a diagnosis of myocardial infarction or venous thromboembolism.

To determine whether or not the PAI-1 4G/5G and t-PA I/D polymorphisms in African-Americans were linked to cardiovascular disease, the association of these polymorphisms to disease expression was analyzed in a recently completed case-control study of myocardial infarction or venous thromboembolism among African-Americans. All African-Americans patients with a history of venous thromboembolism attending an anticoagulant clinic, and patients with a history of a MI attending a cardiology clinic at a large local urban public hospital were eligible for inclusion as cases in the study. In this study it was observed that there was a statistically significant association between the D allele of the t-PA I/D polymorphism and venous thromboembolism and a nonsignificant association between the D allele and myocardial infarction among African-Americans. t-PA antigen levels were statistically significantly higher among both myocardial infarction and venous thromboembolism cases compared with control subjects. The genotypes were unrelated to t-PA plasma levels. There was no association between either myocardial infarction or venous thromboembolism and the 4G/5G PAI-1 genotype. It was also found that genotype frequencies for both PAI-1 4G/5G and t-PA I/D polymorphisms in African-American adults were different from those reported for both U.S. Causcians and Europeans.

Inhibition of endotoxin-induced TNF-alpha production in macrophages by 5Z-7-oxo-zeaenol and other fungal resorcylic acid lactones.

Resorcylic acid lactones are fungal metabolites that exhibit a wide range of biological properties which includes oestrogenic, antifungal, phytotoxic and anti-inflammatory activity. The capacity of 5Z-7-oxo-zeaenol, a resorcylic lactone of fungal origin and six naturally occurring analogues to inhibit lipopolysaccharide (LPS)-induced cytokine production in phorbol 12-myristate-13-acetate (PMA)-treated cultured myelomonocytic cells (U937) was compared. The activity of the natural analogues in the U937 assay varied over 10(4)-fold, with 5Z-7-oxo-zeaenol the most potent of those tested inhibiting tumour necrosis factor-alpha (TNF alpha) production in these cells with IC50 of 6 nM. The isomeric 7-oxo-zeaenol and structurally more distant monorden (radicicol) were the next most active compounds with IC50 approximately 500 nM, and zearalenone, the least active with IC50 > 400 microM. 5Z-7-oxo-zeaenol retained activity in LPS-stimulated peripheral blood mononuclear cells with an IC50 of 10-25 nM. This compound also inhibited LPS-induced TNF alpha production in whole blood experiments (IC50 100-1000 nM) and lowered serum levels of TNF alpha in mice when administered prior to LPS. 5Z-7-oxo-zeaenol was shown to inhibit the phosphorylation and activation of mitogen-activated protein kinase (MAPK) induced by LPS. These data are consistent with a mechanism of action at or upstream of MAPK with resultant downstream effects. This series of naturally occurring analogues represents an interesting group of compounds with diverse biological properties. Of this series, 5Z-7-oxo-zeanenol has exceptionally potent anti-inflammatory properties exhibited by its strong inhibition of cytokine production.

The effects of hippocampal and fimbria-fornix lesions on prepulse inhibition.

The present experiments tested the effects of conventional (dorsal aspiration and electrolytic) and excitotoxic (N-methyl-D-aspartate [NMDA]) hippocampal lesions and fimbria-fornix (FF) transection on prepulse inhibition (PPI) of startle response and on open-field activity. Activity was increased by FF transection and by conventional but not excitotoxic hippocampal lesions; complete NMDA lesion increased amphetamine-induced activity. Whereas dorsal hippocampal aspiration lesion disrupted PPI, the phenomenon was not affected by dorsal hippocampal electrolytic lesion, partial or complete excitotoxic (NMDA) hippocampal lesions, or complete FF transection, which interrupted the cholinergic input to the hippocampus as well as the hippocampal-subicular input to the nucleus accumbens. Systemic apomorphine disrupted PPI in both FF-transected rats and their controls. It is suggested that the hippocampus is essential for PPI disruption rather than for PPI expression.

The relationship between polymorphisms in the endothelial cell nitric oxide synthase gene and the platelet GPIIIa gene with myocardial infarction and venous thromboembolism in African Americans.

STUDY OBJECTIVES: To determine whether the polymorphic dinucleotide repeats found in intron 4 of the endothelial cell nitric oxide synthase (ecNOS) gene and the platelet GPIIIa PLA(1)/A(2) polymorphism are associated with myocardial infarction (MI) and venous thromboembolism (VTE) in African Americans. Because these two genes may interact physiologically, the third objective was to determine if there was a relationship between the polymorphisms with respect to MI and VTE. DESIGN: A hospital-based case-control study. After informed consent was obtained, blood used for DNA extraction was drawn from the subjects. SETTING: The study was conducted in the Anticoagulant Clinic and the Cardiology Clinic at Grady Memorial Hospital in Atlanta Georgia. PATIENTS: Subjects were recruited from African-American patients with a reported history of MI (n = 110) or VTE (n = 91). Control subjects (n = 185) without a history of cardiovascular or venous disease were recruited from an outpatient clinic. MEASUREMENTS AND RESULTS: The 393 ecNOS allele was more common among MI cases (36%; p = 0.01) and VTE cases (35%; p = 0.04) than among control subjects (26%). There was no association between the GPIIIa genotypes and either MI or VTE. However, among the MI subjects, there was a strong association between the ecNOS 393/393 genotype and the Pl(A2) allele. It was also found that the frequency of the 393 allele was higher in African-American persons (0.26) compared with what has been reported for Australian Caucasians (0. 14) and Japanese (0.10). CONCLUSIONS: The 393 allele but not the Pl(A2) allele was significantly associated with both MI and VTE in African Americans. Homozygosity for the 393 allele was significantly associated to the diagnosis of MI prior to the age of 45. The combination of the 393 allele and a Pl(A2) allele was also highly associated with MI. The frequency of the 393 allele was significantly higher in African Americans than what has been reported for other populations. This study furthers not only extends the association of the 393 allele to VTE but has demonstrated an interaction with the Pl(A2) allele with respect to MI.

Patient management of cerebral origin spasticity with intrathecal baclofen.

Controlled, continuous intrathecal infusion of baclofen injection relieves severe spasticity for a wide range of patients. This therapy has become a standard treatment option in spasticity management programs. Multidisciplinary teams, coordinated by an experienced neuroscience practitioner, provide treatment in five phases. Experience from clinical trials and commercial use of this treatment provides a guide for others who are initiating this therapy at their facility. Further prospective research is needed to accurately determine best clinical practice guidelines for cost effective use of this therapy.

Relation of three genetic traits to venous thrombosis in an African-American population.

A mutation in the Factor V gene (Factor V Leiden), a variant in the 5,10-methylenetetrahydrofolate reductase gene (MTHFR), and an insertion/deletion polymorphism of the angiotensin I-converting enzyme gene (ACE) may be related to abnormal blood clotting. The authors examined the associations between these genetic traits and venous thrombosis among African Americans. This study comprised 93 patients with venous thrombosis and 185 control subjects attending clinics at an urban, public hospital in Atlanta, Georgia, in 1995-1996. Subjects' DNA was extracted from blood and assayed for these genetic traits. Odds ratios were obtained from logistic regression and used as a measure of association between each genetic trait and venous thrombosis. Factor V Leiden was unrelated to venous thrombosis, but the mutation ws too rare among our African-American subjects to evaluate adequately its relation to venous thrombosis. The homozygous and heterozygous genotypes for the V allele of the MTHFR gene were unrelated to venous thrombosis (odds ratio = 0.9, 95% confidence interval 0.5-1.8). Subjects with the deletion/deletion ACE polymorphism experienced a moderate increase in venous thrombosis risk compared with persons with the other genotypes (odds ratio = 1.5, 95% confidence interval 0.9-2.6). However, women with this ACE genotype experienced no increased risk (odds ratio = 0.9, 95% confidence interval 0.5-1.9), whereas men with this genotype had nearly three times the risk (odds ratio = 2.8, 95% confidence interval 1.2-6.2; p value for interaction = 0.06). These data indicate that the prevalence of Factor V Leiden and the V allele of the MTHFR gene is low among African Americans. The D allele of the ACE gene is equally prevalent among African Americans and whites and may be related to venous thrombosis among African-American men.

Differential effects on TNF alpha production by pharmacological agents with varying molecular sites of action.

This study describes the activation conditions for tumor necrosis factor-alpha (TNF alpha) production in myelomonocytic U937 cells and human primary peripheral blood monocytes in response to lipopolysaccharide (LPS) and/or phorbol 12-myristate 13-acetate (PMA). PMA itself induced only low levels of TNF alpha production with delayed kinetics (e.g. 0.758 +/- 0.128 ng/ml from U937 cells after 48 h) while LPS induced greater levels of TNF alpha production in less time (e.g. 2.083 +/- 0.96 ng/ml from monocytes in 24 h). Pharmacological agents with various molecular sites of action were used to validate the two systems, with the protein serine-threonine kinase inhibitors staurosporine and Ro-31-8220, the protein tyrosine kinase inhibitor herbimycin A (HBA) and dexamethasone exhibiting the greatest potency (IC50S 5-350 nM). In contrast to the effect on TNF alpha production, PMA induced strong phosphorylation/activation of p42/p44mapk in monocytes by 10 min determined in a mobility shift assay, while LPS was a weaker inducer. Additionally, staurosporine (to LPS and PMA) and HBA (to LPS only) inhibited the activation of these mitogen-activated protein kinase (MAPK) isoforms at doses 10-100 fold higher than those required to inhibit maximal TNF alpha production. These data indicate the involvement of the p42/p44mapk signalling pathway in LPS-induced pro-inflammatory cytokine production but suggest that other signalling pathways are also implicated in this phenomenon.

A racial difference in the prevalence of the Arg506-->Gln mutation.

Several recent studies have reported that the factor V Arg506-->Gln mutation is present in 3-10% of adults of European descent. To determine if the prevalence is comparable among Blacks, we have initiated a case-control study in a large urban hospital in Atlanta which serves a substantial black population. We have evaluated 131 black subjects with confirmed venous or arterial thrombosis and 61 black subjects without a history of thrombosis. Only one case and one control were positive for the Arg506-->Gln mutation. We conclude that the mutation is more common among Whites than Blacks.

Intrathecal baclofen for spasticity of cerebral palsy: project coordination and nursing care.

Spasticity caused by cerebral palsy is painful and disabling. Infusion of an intrathecal antispasmodic for relief is investigated in a multicenter, interdisciplinary clinical trial. Clinical nurse specialists coordinate local team endeavors. The nursing process serves as a functional framework for project development, protocol implementation and long-term patient follow-up.