2-Fluoro-4-pyridinylmethyl analogues of linopirdine as orally active acetylcholine release-enhancing agents with good efficacy and duration of action

In an effort to improve the pharmacokinetic and pharmacodynamic properties of the cognition-enhancer linopirdine (DuP 996), a number of core structure analogues were prepared in which the 4-pyridyl pendant group was systematically replaced with 2-fluoro-4-pyridyl. This strategy resulted in the discovery of several compounds with improved activity in acetylcholine (ACh) release-enhancing assays, in vitro and in vivo. The most effective compound resulting from these studies, 10, 10-bis[(2-fluoro-4-pyridinyl)methyl]-9(10H)-anthracenone (9), is between 10 and 20 times more potent than linopirdine in increasing extracellular hippocampal ACh levels in the rat with a minimum effective dose of 1 mg/kg. In addition to superior potency, 9 possesses an improved pharmacokinetic profile compared to that of linopirdine. The half-life of 9 (2 h) in rats is 4-fold greater than that of linopirdine (0.5 h), and it showed a 6-fold improvement in brain-plasma distribution over linopirdine. On the basis of its pharmacologic, pharmacokinetic, absorption, and distribution properties, 9 (DMP543) has been advanced for clinical evaluation as a potential palliative therapeutic for treatment of Alzheimer's disease.     

Mutations in the carboxyl terminus of the agouti protein decrease agouti inhibition of ligand binding to the melanocortin receptors

Several mutations that cause ectopic expression of the agouti gene result in obesity, hyperinsulinemia, and yellow coat color. A candidate pathway for agouti induced obesity and hyperinsulinemia is through altered signaling by melanocortin receptors, as agouti normally regulates coat coloration through antagonism of melanocortin receptor 1. Furthermore, melanocortin peptides mediate functions including steroidogenesis, lipolysis, and thermoregulation. We report apparent inhibition dissociation constants for mouse and human agouti protein inhibition of ligand binding to the melanocortin receptors, to determine which of these receptors might be involved in agouti induced diabetes. The similarity in the apparent K(I) values for agouti inhibition of ligand binding to the brain melanocortin receptors 3 and 4 (mouse: K(I) app = 190 +/- 74 and 54 +/- 18 nM; human: K(I) app = 140 +/- 56 and 70 +/- 18 nM, respectively) suggests that the MC3-R is a potential candidate for a receptor mediating the effects of agouti protein overexpression. Agouti residues important for melanocortin receptor inhibition were identified through the analysis of deletion constructs and site-specific variants. Val83 is important for inhibition of binding to MC1-R (K(I) app for Val83Ala agouti increased 13-fold relative to wild-type protein). Arg85, Pro86, and Pro89 are important for selective inhibition of binding between MC1-R and MC3-R and MC4-R as their apparent K(I) values are essentially unchanged at MC1-R, while they have increased 6-10-fold relative to wild-type protein at MC3-R and MC4-R.     

Force and myosin content variation in isolated intact single muscle fibres from Rana temporaria

We studied the relation between force normalized by dry mass per unit length and the myosin fraction of muscle dry mass. The two tibialis anterior muscles were dissected from 12 frogs (Rana temporaria). Then, from one muscle, two single fast-twitch fibres were isolated. Each fibre was mounted isometrically in Ringer's solution, and electrically stimulated using a standardized protocol. Peak force production, normalized by the fibre's dry mass per unit length, varied by a factor of 1.4. Little variation in normalized force was measured between fibres from the same animal, whereas between animals a significant difference was found (P<0.05). The contralateral muscle was used to determine the myosin fraction of the dry mass. The relationship between the fraction myosin of the dry mass and force normalized by dry mass per unit length showed a high correlation (r = 0.81; n = 12). From this we conclude that variation in normalized tetanic force is determined greatly (65%) by variations in myosin content.     

Rhabdomyolysis, acute renal failure and hepatopathy induced by lovastatin monotherapy

The immunoglobulin VH gene rearrangement in a primary cutaneous, large-cell (centroblastic and immunoblastic) B-cell lymphoma was analyzed using a micromanipulation/single-cell polymerase chain reaction technique. In all single B cells obtained from CD20-stained skin sections that gave a polymerase chain reaction product (eight of 27 in biopsy I), the same VHDJH rearrangement, consisting of DP-54-DIR1-JH3a genes, was detected, with no intraclonal nucleotide diversity. Comparison with the most closely related germline counterpart showed significantly altered complementarity determining gene regions as a result of somatic mutations, suggesting an antigen-driven selection and expansion ofthis particular B-cell clone. Interestingly, in a biopsy obtained from the patient 9 mo later, during disease progression (deep muscle infiltration), the lymphoma cells again contained the same VHDJH gene rearrangement (six of 18 in biopsy II) without any further somatic mutations. Therefore, it is suggested that the cutaneous lymphoma characterized throughout this study descended from postgerminal center B-cells.     

Accuracy of the implant torque wrench following time in clinical service

Dermatofibrosarcoma protuberans (DFSP) with fibrosarcoma (FS)-like areas (DFSP-FS) is a peculiar neoplasm that combines microscopic findings of DFSP and FS. Because of the scarce number of cases published, tumor prognosis remains controversial. The clinical histories and the histologic material of 27 cases of DFSP were reviewed. Four of them showed fibrosarcomatous areas. Follow-up data, ranging from 12 to 125 months, were obtained in all four cases. Two patients had repeated local recurrences. One patient developed pulmonary metastases and died of disease 49 months after diagnosis. In the other two patients, no recurrences or metastases were detected at 12 and 70 months after local excision, respectively. Progressive increase of FS areas, cellular density, cellular atypia, and mitotic activity were observed during the recurrences. All cases showed diffuse positive immunostaining for CD34 in DFSP areas. Three cases were also CD34-positive in FS areas. Based on a careful review of the literature and our personal experience, we conclude that DFSP-FS is a rare variant of DFSP with a higher rate of local recurrences and more distant metastases than typical DFSP.     

Nutrient flows in agriculture in The Netherlands with special emphasis on pig production

Annual nitrogen (N), phosphorus (P), and potassium (K) flows in agriculture in The Netherlands were identified and quantified in 1990, with special emphasis on pig production. Also, the effects that various management strategies in pig production have on NPK emission in 1990 were compared using a static deterministic simulation model. Ammonia emission from pig production in 1990 (60.9 Gg N) exceeded the defined target for the year 2000 (12.7 Gg N). Measures that affect volatilization of ammonia directly (i.e., introduction of low-emission stables, manure storage facilities, or manure application techniques) reduced ammonia emission most effectively. These measures, however, should be combined with a reduction in application of artificial N fertilizer to avoid an increase in N losses through leaching, run-off, or denitrification. Targets for ammonia emission in the year 2010 require a reduction in the pig population of 24 to 62%, in addition to implications of measures described in this article. National NPK losses in 1990 through leaching, run-off, or denitrification, predicted at 223.5 kg/ha for N, 32.7 kg/ha for P, and 67 kg/ha for K, exceeded government targets for the year 2010 (185 kg N/ha; 8.7 kg P/ha; norm not set for K). Reducing application of artificial NPK fertilizer reduced national NPK losses most effectively. For P, use of phytase and feeding pigs in accordance with their P requirements is required, in addition to limited use of artificial P fertilizer to meet targets for the year 2010. Hence, from an environmental point of view, pig production in The Netherlands is limited primarily by ammonia emission targets for the year 2010.     

Peroxisome proliferator-activated receptor gene expression in human tissues. Effects of obesity, weight loss, and regulation by insulin and glucocorticoids

The peroxisome proliferator activated receptor (PPAR gamma) plays a key role in adipogenesis and adipocyte gene expression and is the receptor for the thiazolidinedione class of insulin-sensitizing drugs. The tissue expression and potential for regulation of human PPAR gamma gene expression in vivo are unknown. We have cloned a partial human PPAR gamma cDNA, and established an RNase protection assay that permits simultaneous measurements of both PPAR gamma1 and PPAR gamma2 splice variants. Both gamma1 and gamma2 mRNAs were abundantly expressed in adipose tissue. PPAR gamma1 was detected at lower levels in liver and heart, whereas both gamma1 and gamma2 mRNAs were expressed at low levels in skeletal muscle. To examine the hypothesis that obesity is associated with abnormal adipose tissue expression of PPAR gamma, we quantitated PPARgamma mRNA splice variants in subcutaneous adipose tissue of 14 lean and 24 obese subjects. Adipose expression of PPARgamma 2 mRNA was increased in human obesity (14.25 attomol PPAR gamma2/18S in obese females vs 9.9 in lean, P = 0.003). This increase was observed in both male and females. In contrast, no differences were observed in PPAR gamma1/18S mRNA expression. There was a strong positive correlation (r = 0.70, P < 0.001) between the ratio of PPAR gamma2/gamma1 and the body mass index of these patients. We also observed sexually dimorphic expression with increased expression of both PPAR gamma1 and PPAR gamma2 mRNAs in the subcutaneous adipose tissue of women compared with men. To determine the effect of weight loss on PPAR gamma mRNA expression, seven additional obese subjects were fed a low calorie diet (800 Kcal) until 10% weight loss was achieved. Mean expression of adipose PPAR gamma2 mRNA fell 25% (P = 0.0250 after a 10% reduction in body weight), but then increased to pretreatment levels after 4 wk of weight maintenance. Nutritional regulation of PPAR gamma1 was not seen. In vitro experiments revealed a synergistic effect of insulin and corticosteroids to induce PPAR gamma expression in isolated human adipocytes in culture. We conclude that: (a) human PPAR gamma mRNA expression is most abundant in adipose tissue, but lower level expression of both splice variants is seen in skeletal muscle; to an extent that is unlikely to be due to adipose contamination. (b) RNA derived from adipose tissue of obese humans has increased expression of PPAR gamma 2 mRNA, as well as an increased ratio of PPAR gamma2/gamma1 splice variants that is proportional to the BMI; (c) a low calorie diet specifically down-regulates the expression of PPAR gamma2 mRNA in adipose tissue of obese humans; (d) insulin and corticosteroids synergistically induce PPAR gamma mRNA after in vitro exposure to isolated human adipocytes; and (e) the in vivo modulation of PPAR gamma2 mRNA levels is an additional level of regulation for the control of adipocyte development and function, and could provide a molecular mechanism for alterations in adipocyte number and function in obesity.     

Effect of noninvasive positive pressure ventilation on mortality in patients admitted with acute respiratory failure: a meta-analysis

Changes in angiogenesis and expression of extracellular matrix-degrading enzymes have been substantiated during progression of solid tumours, whereas information on haematological tumours remains circumstantial. In this study, 57 biopsies of mycosis fungoides (MF), a haematological tumour of T-cell lineage, were investigated immunohistochemically for the extent of angiogenesis, and by in situ hybridisation for the expression of matrix metalloproteinases 2 (MMP-2, collagenase A) and 9 (MMP-9, collagenase B). The biopsies we grouped according to the stage of progression: patch-->plaque-->nodular (most advanced). The extent of angiogenesis, as microvessel area, of MF lesions as a whole was significantly higher than that of normal uninjured skin, used as a control. When the stages of MF progression were compared, the values of MF patch stage overlapped that of control skin, while values were significantly higher in the plaque stage and even higher in the nodular stage. In these stages, microvessels were widely scattered in the tumour tissue, in close association with tumour cells, and they frequently displayed arborisation and microaneurysmatic dilation. In contrast, in the patch stage microvessels were irregularly distributed around the tumour aggregates, and arborisation or dilated structures were only rarely seen. The expression of MMP-2 and MMP-9 mRNAs underwent significant upregulation in relation to advancing stage. Indeed, the upstaging was significantly associated with higher proportions of lesions positive for each mRNA or for both, and with lesions with the greatest intensity of expression for each mRNA. Besides tumour cells, the MMP-2 mRNA was expressed by microvascular endothelial cells of intratumour and peri-tumour vessels, and by fibroblasts which were especially abundant in the stroma adjacent to the tumour nodules. The MMP-9 mRNA was found to be present in a subset of tissue macrophages which were more frequently located in close vicinity to the tumour nodules. In contrast, in control skin, a weak positivity for the MMP-2 mRNA in very few microvascular endothelial cells and no signal for the MMP-9 mRNA were observed. These in situ data suggest that angiogenesis and degradation of the extracellular matrix occur simultaneously during MF progression. They imply that interaction between tumour cells and their microvasculature are all the more likely to occur during progression, occasionally with the contribution of tumour-associated stromal cells.     

Thrombosis of the deep cerebral veins: CT and MRI findings with pathologic correlation

Deep cerebral vein thrombosis can present with acute, severe neurological symptoms and may be rapidly fatal as in the 20-year-old woman reported here. Although MRI is superior for establishing the diagnosis, CT is usually the first examination performed in the clinical setting. It is therefore important to recognise certain indicators such as extensive bithalamic low density. These and certain other less specific signs are correlated with the MRI and autopsy findings.     

Phase II study of combined levamisole with recombinant interleukin-2 in patients with advanced malignant melanoma

Adoptive immunotherapy (AI) with interleukin-2 (IL-2) and lymphokine-activated killer cells (LAK) is an antineoplastic modality in which immune-activated cells are administered to a host with advanced cancer in an attempt to mediate tumor regression. Levamisole (LEV), an immune stimulant, has been suggested to have therapeutic effectiveness in a variety of cancers. After a phase I trial of recombinant IL-2 plus LEV, a phase II trial of this combination was conducted in patients with advanced malignant melanoma. Nineteen patients were entered in the trial. They received IL-2 at 3 x 10(6) U/m2 subcutaneously daily x 5 plus LEV 50 mg/ m2 orally three times daily (p.o. t.i.d.) x 5. Patients were reevaluated at four-week intervals. None of the patients achieved a partial or complete regression (PR, CR). The median time to treatment failure (refusal, progression, or off study due to toxicity) was 56 days. Grade IV toxicities included vomiting (3 patients), lethargy (1 patient), and musculoskellar pain (1 patient). This regimen is not recommended for further testing in patients with advanced malignant melanoma.