A multifaceted hospitalist quality improvement intervention: Decreased frequency of common labs.

PURPOSE: Common labs such as a daily complete blood count or a daily basic metabolic panel represent possible waste and have been targeted by professional societies and the Choosing Wisely campaign for critical evaluation. We undertook a multifaceted quality-improvement (QI) intervention in a large community hospitalist group to decrease unnecessary common labs. METHODS: The QI intervention was composed of academic detailing, audit and feedback, and transparent reporting of the frequency with which common labs were ordered as daily within the hospitalist group. We performed a pre-post analysis, comparing a cohort of patients during the 10-month baseline period before the QI intervention and the 7-month intervention period. Demographic and clinical data were collected from the electronic medical record. The primary endpoint was number of common labs ordered per patient-day as estimated by a clustered multivariable linear regression model clustering by ordering hospitalist. Secondary endpoints included length of stay, hospital mortality, 30-day readmission, blood transfusion, amount of blood transfused, and laboratory cost per patient. RESULTS: The baseline (n = 7824) and intervention (n = 5759) cohorts were similar in their demographics, though the distribution of primary discharge diagnosis-related groups differed. At baseline, a mean of 2.06 (standard deviation 1.40) common labs were ordered per patient-day. Adjusting for age, sex, and principle discharge diagnosis, the number of common labs ordered per patient-day decreased by 0.22 (10.7%) during the intervention period compared to baseline (95% confidence interval [CI], 0.34 to 0.11; P < 0.01). There were nonsignificant reductions in hospital mortality in the intervention period compared to baseline (2.2% vs 1.8%, P = 0.1) as well as volume of blood transfused in patients who received a transfusion (127.2 mL decrease; 95% CI, -257.9 to 3.6; P = 0.06). No effect was seen on length of stay or readmission rate. The intervention decreased hospital direct costs by an estimated $16.19 per admission or $151,682 annualized (95% CI, $119,746 to $187,618). CONCLUSION: Implementation of a multifaceted QI intervention within a community-based hospitalist group was associated with a significant, but modest, decrease in the number of ordered lab tests and hospital costs. No effect was seen on hospital length of stay, mortality, or readmission rate. This intervention suggests that a community-based hospitalist QI intervention focused on daily labs can be effective in safely reducing healthcare waste without compromising quality of care.

Pharmacological evaluation of adipose dysfunction via 11ß-hydroxysteroid dehydrogenase type 1 in the development of diabetes in diet-induced obese mice with cortisone pellet implantation.

Signals from intracellular glucocorticoids (GCs) via 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) in adipose tissues have been reported to serve as amplifiers leading to deterioration of glucose metabolism associated with obesity. To elucidate adipose dysfunction via 11ß-HSD1 activation in the development of obesity-related diabetes, we established novel diabetic mice by implanting a cortisone pellet (CP) in diet-induced obesity (DIO) mice. Cortisone pellet-implanted DIO mice (DIO/CP mice) showed hyperglycemia, insulin resistance, hyperlipidemia, and ectopic fat accumulation, whereas cortisone pellet implantation in lean mice did not induce hyperglycemia. In DIO/CP mice, indexes of lipolysis such as plasma glycerol and nonesterified fatty acids (NEFAs) increased before hyperglycemia appeared. Furthermore, the adipose mRNA level of 11ß-HSD1 was up-regulated in DIO/CP mice compared with sham-operated DIO mice. RU486 (mifepristone, 11ß-[p-(dimethylamino)phenyl]-17ß-hydroxy-17-(1-propynyl)estra-4,9-dien-3-one), a glucocorticoid receptor antagonist, decreased adipose mRNA levels of 11ß-HSD1 as well as adipose triglyceride lipase. RU486 also improved plasma NEFA, glycerol, and glucose levels in DIO/CP mice. These results demonstrate that lipolysis in adipose tissues caused by GC activation via 11ß-HSD1 serves as a trigger for diabetes with ectopic fat accumulation. Our findings also indicate the possibility of a vicious circle of GC signals via 11ß-HSD1 up-regulation in adipose tissues, contributing to deterioration of glucose metabolism to result in diabetes. Our DIO/CP mouse could be a suitable model of type 2 diabetes to evaluate adipose dysfunction via 11ß-HSD1.

Is your clinical documentation improvement program compliant?

Hospitals and health systems should consider four strategies for improving documentation: Develop a short list of the most commonly underdocumented or incorrectly documented clinical conditions at the facility. Develop definitions for each of the conditions on this list. Ask the medical director of each specialty area to educate the clinicians in their group on these definitions. Measure and manage documentation performance.

Adrenergic regulation of AMP-activated protein kinase in brown adipose tissue in vivo.

AMP-activated protein kinase (AMPK), an evolutionarily conserved serine-threonine kinase that senses cellular energy status, is activated by stress and neurohumoral stimuli. We investigated the mechanisms by which adrenergic signaling alters AMPK activation in vivo. Brown adipose tissue (BAT) is highly enriched in sympathetic innervation, which is critical for regulation of energy homeostasis. We performed unilateral denervation of BAT in wild type (WT) mice to abolish neural input. Six days post-denervation, UCP-1 protein levels and AMPK α2 protein and activity were reduced by 45%. In ß(1,2,3)-adrenergic receptor knock-out mice, unilateral denervation led to a 25-45% decrease in AMPK activity, protein expression, and Thr(172) phosphorylation. In contrast, acute α- or ß-adrenergic blockade in WT mice resulted in increased AMPK α Thr(172) phosphorylation and AMPK α1 and α2 activity in BAT. But short term blockade of α-adrenergic signaling in ß(1,2,3)-adrenergic receptor knock-out mice resulted in decreased AMPK activity in BAT, which strongly correlated with enhanced phosphorylation of AMPK on Ser(485/491), a site associated with inhibition of AMPK activity. Both PKA and AKT inhibitors attenuated AMPK Ser(485/491) phosphorylation resulting from α-adrenergic blockade and prevented decreases in AMPK activity. In vitro mechanistic studies in BAT explants showed that the effects of α-adrenergic blockade appeared to be secondary to inhibition of oxygen consumption. In conclusion, adrenergic pathways regulate AMPK activity in vivo acutely via alterations in Thr(172) phosphorylation and chronically through changes in the α catalytic subunit protein levels. Furthermore, AMPK α Ser(485/491) phosphorylation may be a novel mechanism to inhibit AMPK activity in vivo and alter its biological effects.

Irbesartan enhances GLUT4 translocation and glucose transport in skeletal muscle cells.

Irbesartan, an angiotensin II type 1 receptor blocker has been reported to alleviate metabolic disorder in animal studies and human clinical trials. Although this effect may be related to the ability of irbesartan to serve as a partial agonist for the peroxisome proliferator-activated receptor (PPAR)-γ, the target tissues on which irbesartan acts remain poorly defined. As muscle glucose transport plays a major role in maintaining systemic glucose homeostasis, we investigated the effect of irbesartan on glucose uptake in skeletal muscle cells. In C2C12 myotubes, 24-h treatment with irbesartan significantly promoted both basal and insulin-stimulated glucose transport. In L6-GLUT4myc myoblasts, irbesartan caused a significant increase in glucose transport and GLUT4 translocation to the cell surface in a concentration-dependent manner. Valsartan, another angiotensin II type 1 receptor blocker had no effect on either glucose uptake or GLUT4 translocation, implying that these actions on glucose transport are independent of angiotensin II receptor blockade. Moreover, irbesartan exerted these effects in an additive manner with insulin, but not with acute treatment for 3 h, suggesting that they may require the synthesis of new proteins. Finally, in insulin-resistant Zucker fatty rat, irbesartan (50 mg/kg/day for 3 weeks) significantly ameliorated insulin resistance without increasing weight gain. We conclude that irbesartan has a direct action, which can be additive to insulin, of promoting glucose transport in skeletal muscle. This may be beneficial for ameliorating obesity-related glucose homeostasis derangement.

Identification of glycosylated exendin-4 analogue with prolonged blood glucose-lowering activity through glycosylation scanning substitution.

Exendin-4, a glucagon-like peptide 1 receptor agonist, is a potent therapeutic xenopeptide hormone for the treatment of type 2 diabetes. In order to further improve in vivo activity, we examined the introduction of sialyl N-acetyllactosamine (sialyl LacNAc) to exendin-4. The glycosylated analogue having sialyl LacNAc at position 28 was found to have improved in vivo activity with prolonged glucose-lowering activity.

Neuronal protein tyrosine phosphatase 1B deficiency results in inhibition of hypothalamic AMPK and isoform-specific activation of AMPK in peripheral tissues.

PTP1B(-/-) mice are resistant to diet-induced obesity due to leptin hypersensitivity and consequent increased energy expenditure. We aimed to determine the cellular mechanisms underlying this metabolic state. AMPK is an important mediator of leptin's metabolic effects. We find that alpha1 and alpha2 AMPK activity are elevated and acetyl-coenzyme A carboxylase activity is decreased in the muscle and brown adipose tissue (BAT) of PTP1B(-/-) mice. The effects of PTP1B deficiency on alpha2, but not alpha1, AMPK activity in BAT and muscle are neuronally mediated, as they are present in neuron- but not muscle-specific PTP1B(-/-) mice. In addition, AMPK activity is decreased in the hypothalamic nuclei of neuronal and whole-body PTP1B(-/-) mice, accompanied by alterations in neuropeptide expression that are indicative of enhanced leptin sensitivity. Furthermore, AMPK target genes regulating mitochondrial biogenesis, fatty acid oxidation, and energy expenditure are induced with PTP1B inhibition, resulting in increased mitochondrial content in BAT and conversion to a more oxidative muscle fiber type. Thus, neuronal PTP1B inhibition results in decreased hypothalamic AMPK activity, isoform-specific AMPK activation in peripheral tissues, and downstream gene expression changes that promote leanness and increased energy expenditure. Therefore, the mechanism by which PTP1B regulates adiposity and leptin sensitivity likely involves the coordinated regulation of AMPK in hypothalamus and peripheral tissues.

Chemoenzymatic synthesis of glycosylated glucagon-like peptide 1: effect of glycosylation on proteolytic resistance and in vivo blood glucose-lowering activity.

Glucagon-like peptide 1 (7-36) amide (GLP-1) has been attracting considerable attention as a therapeutic agent for the treatment of type 2 diabetes. In this study, we applied a glycoengineering strategy to GLP-1 to improve its proteolytic stability and in vivo blood glucose-lowering activity. Glycosylated analogues with N-acetylglucosamine (GlcNAc), N-acetyllactosamine (LacNAc), and alpha2,6-sialyl N-acetyllactosamine (sialyl LacNAc) were prepared by chemoenzymatic approaches. We assessed the receptor binding affinity and cAMP production activity in vitro, the proteolytic resistance against dipeptidyl peptidase-IV (DPP-IV) and neutral endopeptidase (NEP) 24.11, and the blood glucose-lowering activity in diabetic db/db mice. Addition of sialyl LacNAc to GLP-1 greatly improved stability against DPP-IV and NEP 24.11 as compared to the native type. Also, the sialyl LacNAc moiety extended the blood glucose-lowering activity in vivo. Kinetic analysis of the degradation reactions suggested that the sialic acid component played an important role in decreasing the affinity of peptide to DPP-IV. In addition, the stability of GLP-1 against both DPP-IV and NEP24.11 incrementally improved with an increase in the content of sialyl LacNAc in the peptide. The di- and triglycosylated analogues with sialyl LacNAc showed greatly prolonged blood glucose-lowering activity of up to 5 h after administration (100 nmol/kg), although native GLP-1 showed only a brief duration. This study is the first attempt to thoroughly examine the effect of glycosylation on proteolytic resistance by using synthetic glycopeptides having homogeneous glycoforms. This information should be useful for the design of glycosylated analogues of other bioactive peptides as desirable pharmaceuticals.

A high-fat, ketogenic diet induces a unique metabolic state in mice.

Ketogenic diets have been used as an approach to weight loss on the basis of the theoretical advantage of a low-carbohydrate, high-fat diet. To evaluate the physiological and metabolic effects of such diets on weight we studied mice consuming a very-low-carbohydrate, ketogenic diet (KD). This diet had profound effects on energy balance and gene expression. C57BL/6 mice animals were fed one of four diets: KD; a commonly used obesogenic high-fat, high-sucrose diet (HF); 66% caloric restriction (CR); and control chow (C). Mice on KD ate the same calories as mice on C and HF, but weight dropped and stabilized at 85% initial weight, similar to CR. This was consistent with increased energy expenditure seen in animals fed KD vs. those on C and CR. Microarray analysis of liver showed a unique pattern of gene expression in KD, with increased expression of genes in fatty acid oxidation pathways and reduction in lipid synthesis pathways. Animals made obese on HF and transitioned to KD lost all excess body weight, improved glucose tolerance, and increased energy expenditure. Analysis of key genes showed similar changes as those seen in lean animals placed directly on KD. Additionally, AMP kinase activity was increased, with a corresponding decrease in ACC activity. These data indicate that KD induces a unique metabolic state congruous with weight loss.

Diet-induced obesity alters AMP kinase activity in hypothalamus and skeletal muscle.

AMP-activated protein kinase (AMPK) is a key regulator of cellular energy balance and of the effects of leptin on food intake and fatty acid oxidation. Obesity is usually associated with resistance to the effects of leptin on food intake and body weight. To determine whether diet-induced obesity (DIO) impairs the AMPK response to leptin in muscle and/or hypothalamus, we fed FVB mice a high fat (55%) diet for 10-12 weeks. Leptin acutely decreased food intake by approximately 30% in chow-fed mice. DIO mice tended to eat less, and leptin had no effect on food intake. Leptin decreased respiratory exchange ratio in chow-fed mice indicating increased fatty acid oxidation. Respiratory exchange ratio was low basally in high fat-fed mice, and leptin had no further effect. Leptin (3 mg/kg intraperitoneally) increased alpha2-AMPK activity 2-fold in muscle in chow-fed mice but not in DIO mice. Leptin decreased acetyl-CoA carboxylase activity 40% in muscle from chow-fed mice. In muscle from DIO mice, acetyl-CoA carboxylase activity was basally low, and leptin had no further effect. In paraventricular, arcuate, and medial hypothalamus of chow-fed mice, leptin inhibited alpha2-AMPK activity but not in DIO mice. In addition, leptin increased STAT3 phosphorylation 2-fold in arcuate of chow-fed mice, but this effect was attenuated because of elevated basal STAT3 phosphorylation in DIO mice. Thus, DIO in FVB mice alters alpha2-AMPK in muscle and hypothalamus and STAT3 in hypothalamus and impairs further effects of leptin on these signaling pathways. Defective responses of AMPK to leptin may contribute to resistance to leptin action on food intake and energy expenditure in obese states.

Exposure of blood from patients with sickle cell disease to air changes the morphological, oxygen-binding, and sickling properties of sickled erythrocytes.

We collected venous blood samples from 7 steady-state patients with homozygous sickle cell disease under venous oxygen pressure without exposure to air (UnExp-blood) and compared the morphological, oxygen-binding, and sickling properties with those of SS cells in aliquots of the same venous blood samples that were oxygenated in room air or at a PO2 near 180 mmHg (Exp-blood). Results showed that (1) upon deoxygenation under nitrogen, UnExp-blood generated a significantly higher percentage of elongated reversibly sickled cells (RSCs) than did Exp-blood; (2) upon gradual oxygenation of completely deoxygenated sickled cells, RSCs in UnExp-blood converted to discocytes at a higher oxygen pressure than did those in Exp-blood; (3) the degree of hysteresis between the sickling/desickling curves of UnExp-blood was greater than that of Exp-blood; and (4) deoxy-Hb S in hemolysate prepared from SS cells in UnExp-blood polymerized without a delay time, while those from Exp-blood polymerized with a distinct delay time. The in vivo properties of RSCs significantly changed upon oxygenation. We also found that the various properties of blood samples collected from patients with SCD by the ordinary method were similar to those of Exp-blood, probably because such blood samples are exposed to oxygen through air in the needle, syringe, and Vacutainer. Once SS cells were oxygenated, the in vivo properties of RSCs could not be recovered by partial deoxygenation to venous oxygen pressure.

Blood samples collected under venous oxygen pressure from patients with sickle cell disease contain a significant number of a new type of reversibly sickled cells: constancy of the percentage of sickled cells in individual patients during steady state.

We found various levels of a new type of reversibly sickled cell (RSC) with blunt edges in 44 blood samples obtained from 32 steady-state patients with sickle cell disease (SCD) without exposure to air (UnExp-blood). Because these RSCs could be generated in vitro by partial oxygenation of once-deoxygenated SS cells to venous oxygen pressure, we named them "partially oxygenated sickled cells" (POSCs). These RSCs were classified into elongated and non-elongated RSCs, depending on the ratio of the short axis to long axis. The presence of these cells was previously unknown because the standard blood collection method oxygenates most of the POSCs to discocytes due to oxygen in the air space in the needle, syringe, and blood collection tube (Exp-blood). Although the shape of elongated POSCs is similar to that of irreversibly sickled cells (ISCs), POSCs revert to discocytes upon exposure to air. We found the following: (1) the percentage of total sickled cells (total POSCs + ISCs) in UnExp-blood (29.0 +/- 14.5%) was significantly higher than the percentage of sickled cells (mainly ISCs) in Exp-blood (7.3 +/- 5.7%); (2) the percentage of sickled cells in UnExp-blood was specific to individual patients during steady state, while it decreased at the onset of a vaso-occlusive event; and (3) the percentage of sickled cells in UnExp-blood varied widely among steady-state patients (4-56%). This new type of RSC may be used as an internal biomarker to evaluate the disease state of individual patients.

Novel binding sites of 15-deoxy-Delta12,14-prostaglandin J2 in plasma membranes from primary rat cortical neurons.

15-Deoxy-Delta12,14-prostaglandin J2 (15d-Delta12,14-PGJ2) is an endogenous ligand for a nuclear peroxysome proliferator activated receptor-gamma (PPAR). We found novel binding sites of 15d-Delta12,14-PGJ2 in the neuronal plasma membranes of the cerebral cortex. The binding sites of [3H]15d-Delta12,14-PGJ2 were displaced by 15d-Delta12,14-PGJ2 with a half-maximal concentration of 1.6 microM. PGD2 and its metabolites also inhibited the binding of [3H]15d-Delta12,14-PGJ2. Affinities for the novel binding sites were 15d-Delta12,14-PGJ2 > Delta12-PGJ2 > PGJ2 > PGD2. Other eicosanoids and PPAR agonists did not alter the binding of [3H]15d-Delta12,14-PGJ2. In primary cultures of rat cortical neurons, we examined the pathophysiologic roles of the novel binding sites. 15d-Delta12,14-PGJ2 triggered neuronal cell death in a concentration-dependent manner, with a half-maximal concentration of 1.1 microM. The neurotoxic potency of PGD2 and its metabolites was also 15d-Delta12,14-PGJ2 > Delta12-PGJ2 > PGJ2 > PGD2. The morphologic and ultrastructural characteristics of 15d-Delta12,14-PGJ2-induced neuronal cell death were apoptotic, as evidenced by condensed chromatin and fragmented DNA. On the other hand, we detected little neurotoxicity of other eicosanoids and PPAR agonists. In conclusion, we demonstrated that novel binding sites of 15d-Delta12,14-PGJ2 exist in the plasma membrane. The present study suggests that the novel binding sites might be involved in 15d-Delta12,14-PGJ2-induced neuronal apoptosis.

Effect of Gas6 on secretory phospholipase A(2)-IIA-induced apoptosis in cortical neurons.

Gas6, a product of the growth-arrest-specific gene 6, protects cortical neurons from amyloid beta protein (Abeta)-induced apoptosis. Neuronal apoptosis is also caused by human group IIA secretory phospholipase A(2) (sPLA(2)-IIA), which is expressed in the cerebral cortex after brain ischemia. sPLA(2)-IIA induces Ca(2+) influx via L-type voltage-sensitive calcium channels (L-VSCCs), leading to its neurotoxicity. In the present study, we investigated effects of Gas6 on sPLA(2)-IIA-induced cell death in primary cultures of rat cortical neurons. sPLA(2)-IIA caused neuronal cell death in a concentration- and time-dependent manner. Gas6 significantly prevented neurons from sPLA(2)-IIA-induced cell death. Gas6 suppressed sPLA(2)-IIA-induced apoptotic features such as the condensation of chromatin and the fragmentation of DNA. Prior to cell death, sPLA(2)-IIA increased the influx of Ca(2+) into neurons through L-VSCCs. Gas6 significantly inhibited the sPLA(2)-IIA-induced Ca(2+) influx. The blocker of L-VSCCs also suppressed sPLA(2)-IIA-induced neuronal cell death. The cortical cultures contained few non-neuronal cells, indicating that Gas6 affected the survival of neurons directly, but not indirectly via non-neuronal cells. In conclusion, we demonstrate that Gas6 rescues cortical neurons from sPLA(2)-IIA-induced apoptosis. Furthermore, the present study indicates that inhibition of L-VSCC contributes to the neuroprotective effect of Gas6.

Human group IIA secretory phospholipase A2 potentiates Ca2+ influx through L-type voltage-sensitive Ca2+ channels in cultured rat cortical neurons.

Mammalian group IIA secretory phospholipase A2 (sPLA2-IIA) generates prostaglandin D2 (PGD2) and triggers apoptosis in cortical neurons. However, mechanisms of PGD2 generation and apoptosis have not yet been established. Therefore, we examined how second messengers are involved in the sPLA2-IIA-induced neuronal apoptosis in primary cultures of rat cortical neurons. sPLA2-IIA potentiated a marked influx of Ca2+ into neurons before apoptosis. A calcium chelator and a blocker of the L-type voltage-sensitive Ca2+ channel (L-VSCC) prevented neurons from sPLA2-IIA-induced neuronal cell death in a concentration-dependent manner. Furthermore, the L-VSCC blocker ameliorated sPLA2-IIA-induced morphologic alterations and apoptotic features such as condensed chromatin and fragmented DNA. Other blockers of VSCCs such as N type and P/Q types did not affect the neurotoxicity of sPLA2-IIA. Blockers of L-VSCC significantly suppressed sPLA2-IIA-enhanced Ca2+ influx into neurons. Moreover, reactive oxygen species (ROS) were generated prior to apoptosis. Radical scavengers reduced not only ROS generation, but also the sPLA2-IIA-induced Ca2+ influx and apoptosis. In conclusion, we demonstrated that sPLA2-IIA potentiates the influx of Ca2+ into neurons via L-VSCC. Furthermore, the present study suggested that eicosanoids and ROS generated during arachidonic acid oxidative metabolism are involved in sPLA2-IIA-induced apoptosis in cooperation with Ca2+.

Porcine pancreatic group IB secretory phospholipase A2 potentiates Ca2+ influx through L-type voltage-sensitive Ca2+ channels.

Secretory phospholipase A(2) (sPLA(2)) exhibits neurotoxicity in the central nervous system. There are high-affinity binding sites of the porcine pancreatic group IB sPLA(2) (sPLA(2)-IB) in the brain. sPLA(2)-IB causes neuronal cell death via apoptosis in the rat cerebral cortex. Although apoptosis is triggered by an influx of Ca(2+) into neurons, it has not yet been ascertained whether the Ca(2+) influx is associated with the neurotoxicity of sPLA(2)-IB. We thus examined the possible involvement of Ca(2+) in the neurotoxicity of sPLA(2)-IB in the primary culture of rat cortical neurons. sPLA(2)-IB induced neuronal cell death in a concentration- and time-dependent manner. This death was accompanied by condensed chromatin and fragmented DNA, exhibiting apoptotic features. Before apoptosis, sPLA(2)-IB markedly enhanced the influx of Ca(2+) into neurons. A calcium chelator suppressed neurons from sPLA(2)-IB-induced neuronal cell death in a concentration-dependent manner. An L-type voltage-sensitive Ca(2+) channel (L-VSCC) blocker significantly protected the sPLA(2)-IB-potentiated influx of Ca(2+). On the other hand, blockers of N-VSCC and P/Q-VSCC did not. An L-VSCC blocker protected neurons from sPLA(2)-IB-induced neuronal cell death. In addition, the L-VSCC blocker ameliorated the apoptotic features of sPLA(2)-IB-treated neurons. Neither an N-VSCC blocker nor P/Q-VSCC blockers affected the neurotoxicity of the enzyme. In conclusion, these findings demonstrate that the influx of Ca(2+) into neurons play an important role in the neurotoxicity of sPLA(2)-IB. Furthermore, the present study suggests that L-VSCC contribute to the sPLA(2)-IB-potentiated influx of Ca(2+) into neurons.

Prostaglandin E2 rescues cortical neurons from amyloid beta protein-induced apoptosis.

Cerebrospinal fluid prostaglandin E(2) (PGE(2)) levels are elevated in patients with Alzheimer's disease (AD), suggesting an involvement of PGE(2) in the neurodegeneration. AD is characterized by deposits of amyloid beta protein (Abeta) in various regions of the brain, e.g. the cerebral cortex. In the present study, we investigated the effects of PGE(2) on neuronal survival in primary cultures of rat cortical neurons. PGE(2) had no effect on neuronal cell viability or its morphology. Therefore, we examined the synergistic effects of PGE(2) with Abeta, a neurotoxin. Abeta caused neuronal cell death via apoptosis. PGE(2) significantly suppressed Abeta neurotoxicity, but did not promote the neurotoxicity. Furthermore, PGE(2) ameliorated Abeta-induced apoptotic features such as the condensation of chromatin and the fragmentation of DNA. Abeta increased the influx of Ca(2+) into neurons before cell death. Nimodipine, an inhibitor of the L-type voltage-sensitive calcium channel (L-VSCC), significantly reduced Abeta-potentiated Ca(2+) uptake. On the other hand, there was no effect on the Abeta-induced Ca(2+) influx by an N-VSCC blocker or P/Q-VSCC blockers. Moreover, the inhibitor of L-VSCC suppressed Abeta-induced neuronal cell death, whereas neither an N-VSCC blocker nor P/Q-VSCC blockers affected the neurotoxicity of Abeta. PGE(2) also suppressed the Abeta-induced Ca(2+) influx in a concentration-dependent manner. This study demonstrated that PGE(2) rescues cortical neurons from Abeta-induced apoptosis by reducing Ca(2+) influx in the primary culture. Furthermore, the present study suggested that the inhibition of L-VSCC contributes to the neuroprotective effect of PGE(2).

Effects of endothelin B receptor agonists on amyloid beta protein (25-35)-induced neuronal cell death.

Endothelin (ET), a vasoconstrictive peptide, acts as an anti-apoptotic factor, and endothelin receptor B (ET(B) receptor) is associated with neuronal survival in the brain. In the Alzheimer's disease (AD) brain, accumulation of amyloid beta protein (Abeta) is thought to cause neuronal cell death via apoptosis. In the present study, we investigated effects of ET(B) receptor agonists on Abeta-induced neuronal cell death. In primary cultures of rat cortical neurons, Abeta(25-35) caused neuronal cell death in a concentration- and time-dependent manner. Abeta(25-35)-induced neuronal cell death was accompanied by chromatin condensation and DNA fragmentation, exhibiting apoptotic features. ET-3 and IRL-1620, ET(B) receptor agonists, significantly prevented neurons from undergoing Abeta(25-35)-induced cell death. Prior to cell death, Abeta increased concentration of intracellular Ca(2+) ([Ca(2+)](i)). Nimodipine, an L-type voltage-sensitive Ca(2+) channel (L-VSCC) blocker, suppressed the Abeta-induced Ca(2) influx, and attenuated Abeta-induced neuronal apoptosis. On the other hand, omega-conotoxin GIVA, an N-type VSCC blocker and omega-conotoxin MVIIC and omega-agatoxin IVA, P/Q-type VSCC blockers, had no effect. ET-3 and IRL-1620 significantly blocked Abeta(25-35)-induced Ca(2) influx. Furthermore, BQ788, an ET(B) receptor antagonist, inhibited both an anti-apoptotic effect and an L-VSCC-inactivating effect of ET(B) receptor agonists. In conclusion, ET(B) receptor agonists exhibit a protective effect against neurotoxicity of Abeta. Furthermore, these agonists appear to act as anti-apoptotic factors by blocking of L-VSCCs.

Group IB secretory phospholipase A(2)induces cell death in the cultured cortical neurons: a possible involvement of its binding sites.

In primary cultures of rat cortical neurons, group IB secretory phospholipase A(2) (sPLA(2)-IB) induced cell death. In rat cortical membranes, there were high affinity binding sites of [125I]sPLA(2)-IB. The high-affinity binding sites were decreased by sPLA(2)-IB and anti-sPLA(2) receptor immunoglobulin G (anti-sPLA(2)R IgG). Furthermore, anti-sPLA(2)R IgG caused neuronal cell death in a concentration-dependent manner. The present study suggests that sPLA(2)-IB induces neuronal cell death via its high-affinity binding sites.

Group IB secretory phospholipase A2 induces neuronal cell death via apoptosis.

Group IB secretory phospholipase A2 (sPLA2-IB) mediates cell proliferation, cell migration, hormone release and eicosanoid production via its receptor in peripheral tissues. In the CNS, high-affinity binding sites of sPLA2-IB have been documented. However, it remains obscure whether sPLA2-IB causes biologic or pathologic response in the CNS. To this end, we examined effects of sPLA2-IB on neuronal survival in primary cultures of rat cortical neurons. sPLA2-IB induced neuronal cell death in a concentration-dependent manner. This death was a delayed response requiring a latent time for 6 h; sPLA2-IB-induced neuronal cell death was accompanied with apoptotic blebbing, condensed chromatin, and fragmented DNA, exhibiting apoptotic features. Before cell death, sPLA2-IB liberated arachidonic acid (AA) and generated prostaglandin D2 (PGD2) from neurons. PGD2 and its metabolite, Delta12-PGJ2, exhibited neurotoxicity. Inhibitors of sPLA2 and cyclooxygenase-2 (COX-2) significantly suppressed not only AA release, but also PGD2 generation. These inhibitors significantly prevented neurons from sPLA2-IB-induced neuronal cell death. In conclusion, we demonstrate a novel biological response, apoptosis, of sPLA2-IB in the CNS. Furthermore, the present study suggests that PGD2 metabolites, especially Delta12-PGJ2, might mediate sPLA2-IB-induced apoptosis.