Fatal acute melioidosis in a tourist returning from Martinique Island, November 2010.

We report the fatal case of acute melioidosis in a patient returning from Martinique with fever in November 2010. Gram-negative rods were isolated from a blood culture and Burkholderia pseudomallei identified within 24 hours after first medical contact. The patient died two days after admission to hospital despite intravenous therapy with high doses of imipenem/cilastatin and intensive care. Clinicians seeing travellers returning from the subtropics or tropics with severe pneumonia or septicaemia should consider the possibility of acute melioidosis.

Significance of Periodontal Risk Assessment in the recurrence of periodontitis and tooth loss.

AIM: To investigate the association of the Periodontal Risk Assessment (PRA) model categories with periodontitis recurrence and tooth loss during supportive periodontal therapy (SPT) and to explore the role of patient compliance. MATERIAL AND METHODS: In a retrospective cohort, PRA was performed for 160 patients after active periodontal therapy (APT) and after 9.5 +/- 4.5 years of SPT. The recurrence of periodontitis and tooth loss were analysed according to the patient's risk profile (low, moderate or high) after APT and compliance with SPT. The association of risk factors with tooth loss and recurrence of periodontitis was investigated using logistic regression analysis. RESULTS: In 18.2% of patients with a low-risk profile, in 42.2% of patients with a moderate-risk profile and in 49.2% of patients with a high-risk profile after APT, periodontitis recurred. During SPT, 1.61 +/- 2.8 teeth/patient were lost. High-risk profile patients lost significantly more teeth (2.59 +/- 3.9) than patients with moderate- (1.02 +/- 1.8) or low-risk profiles (1.18 +/- 1.9) (Kruskal-Wallis test, p=0.0229). Patients with erratic compliance lost significantly (Kruskal-Wallis test, p=0.0067) more teeth (3.11 +/- 4.5) than patients compliant with SPT (1.07 +/- 1.6). CONCLUSIONS: In multivariate logistic regression analysis, a high-risk patient profile according to the PRA model at the end of APT was associated with recurrence of periodontitis. Another significant factor for recurrence of periodontitis was an SPT duration of more than 10 years.

p38 MAPK inhibition selectively mitigates inflammatory mediators and VEGF production in AF cells co-cultured with activated macrophage-like THP-1 cells.

OBJECTIVES: Recent data have suggested that macrophages are involved in the pathogenesis of discogenic back pain and enhance the secretion of inflammatory mediators in co-cultured annulus fibrosus (AF) cells. The purpose of these studies is to determine the role of p38 mitogen-activated protein kinase (p38 MAPK) signaling in the interactions between macrophage and AF cells. METHODS: Human AF cells were co-cultured with phorbol myristate acetate-stimulated macrophage-like THP-1 cells with and without p38 MAPK inhibition. Conditioned media from co-cultured cells were assayed for interleukin (IL)-6, IL-8, prostaglandin E2 (PGE2), PGF2alpha, and vascular endothelial growth factor (VEGF). Naïve and macrophage-exposed AF cell responses to 10ng/ml tumor necrosis factor-alpha (TNF-alpha) were compared using the same outcome measures. RESULTS: IL-6, IL-8, PGE2, PGF2alpha, and VEGF were secreted in greater quantities by cells maintained in co-culture compared to macrophages or AF cells cultured alone. SB202190 blunted IL-6, PGE2, and PGF2alpha production in a dose-dependent manner in co-culture. However, it did not suppress IL-8 and VEGF production. TNF-alpha-stimulated AF cell inflammatory mediators were up-regulated by macrophage exposure. SB202190 successfully suppressed IL-6, IL-8, PGE2, and PGF2alpha secretion in macrophage-exposed AF cells in response to TNF-alpha. CONCLUSIONS: Annular injury can result in macrophage infiltration, and this can cause enhanced inflammatory mediator and VEGF production by AF cells. The p38 MAPK pathway signals are responsible for much of IL-6 and PG secretion from AF cells with macrophage-like cells, suggesting that blockade of this signal may serve as a therapeutic approach to discogenic pain.

Noncytokinetic radiation injury: anticoagulants as radioprotective agents in experimental radiation hepatitis.

Rats that have been irradiated with a single dose of 1500 roentgens directed to the liver show the following abnormalities: increased clearance of colloidal gold by the liver at 30 days, a decreased albumin space index at 30 days, and increased alkaline phosphatase at 60 days. In each of these cases, animals receiving Depo-heparin in dose ranges reasonable for human application had essentially normal values. There was no radioprotective effect with salicylates at the doses used. Higher doses of salicylates resulted in death of the animals.

Detection of Pneumocystis jirovecii by two staining methods and two quantitative PCR assays.

BACKGROUND: Pneumocystis jirovecii is an opportunistic pathogen that causes pneumonia, particularly in immunodeficient hosts. MATERIALS AND METHODS: We retrospectively compared the results obtained by two staining methods (toluidine blue and calcofluor white) and two quantitative (q) real time PCR assays for the detection of P. jirovecii in bronchoalveolar lavage (BAL) specimens. For the qPCR assays, we used newly selected probes and primers targeting the Kex-1 gene, which codes for a serine endoprotease, and compared the results to those from the published assay targeting the beta-tubulin gene. RESULTS: A total of 1,843 BAL specimens were analyzed microscopically in parallel, and 74 (4.0%) were found to be positive with both stains, 23 (1.2%) were positive only with the toluidine blue stain, and six (0.3%) only with the calcofluor stain (p = 0.003). Of these, a selection of 186 consecutive BAL fluid samples were tested by qPCR using the respective different primer pairs. 21 of the 186 samples (11.3%) were microscopically positive with both stains as well as qPCR positive after 18-31 cycles (corresponding to 5.24 x 10(6) copies/ml to 640 copies/ml of native BAL) using the Kex-1 primer pair and between 21-33 cycles using the beta-tubulin assay. A good correlation between semi-quantitative microscopy and the number of PCR cycles needed for a positive signal was noted. Of the remaining 165 samples, 153 (82%) were both microscopically and PCR negative (PCR with the two sets of primers); the remaining 12 samples (7%) were Kex-1-based PCR positive (from cycles 33 to 41, corresponding to 160 copies/ml of BAL or less) but microscopically negative. Of these latter samples, ten (6%) were also positive (from cycles 34 to 38) with the primers targeting the beta-tubulin gene. Taking microscopy as a reference, the sensitivity of qPCR targeting the Kex-1 gene was 100%, and the specificity was 92.4%. CONCLUSION: The sensitive qPCR analysis proved to be a rapid and reliable method to detect P. jirovecii in BAL.

Lugol chromoendoscopy combined with brush cytology in patients at risk for esophageal squamous cell carcinoma.

BACKGROUND AND STUDY AIMS: Patients with achalasia or malignancies of the head and neck are at increased risk for esophageal squamous cell carcinoma. The discussion of a screening and surveillance program is controversial. The aim of the present study was to determine the diagnostic potential of Lugol chromoendoscopy combined with brush cytology to diagnose esophageal squamous cell carcinoma and high-grade dysplasia. Secondly, the benefit of additional biomarkers was investigated. PATIENTS AND METHODS: A total of 61 patients (21 patients with achalasia and 40 patients with malignancies of the head and neck) were included. Chromoendoscopy with 1.2% Lugol iodine solution with targeted biopsies and brush cytology processed by digital image cytometry (DICM) and fluorescence in situ hybridization (FISH) from unstained lesions (USLs) and stained mucosa were performed. RESULTS: Six of the 61 patients had USLs ≥2 cm. Four patients had high-grade dysplasia (HGD) or carcinoma in situ (CIS). One patient with HGD and one patient with CIS were detected only after Lugol chromoendoscopy. The sensitivity and specificity for detected HGD or CIS in USLs ≥2 cm were 100% and 96.5%. No dysplasia was found in USLs <2 cm. DNA ploidy by DNA cytometry and p53 loss of heterozygosity (LOH) by fluorescence in situ hybridization showed no additional impact on diagnostic accuracy. CONCLUSIONS: Lugol chromoendoscopy enhances the detection rate of high-risk lesions with dysplasia or carcinoma in situ in large unstained lesions. Biomarkers such as aneuploidy and p53 LOH from brush cytology were not of additional benefit in this setting.

Impaired nitric oxide-dependent cyclic guanosine monophosphate generation in glomeruli from diabetic rats. Evidence for protein kinase C-mediated suppression of the cholinergic response.

Nitric oxide (NO)-dependent cyclic guanosine monophosphate (cGMP) generation was examined in glomeruli isolated from 1-2-wk and 2-mo streptozotocin diabetic (D) and control (C) rats. After 1-2 wk of diabetes, ex vivo basal cGMP generation and cGMP responses to carbamylcholine (CCh) were significantly suppressed in glomeruli from D compared with those from C, whereas cGMP responses to the calcium ionophore A23187 and nitroprusside (NP) did not differ in glomeruli from D vs. those from C. After 2 mo, glomeruli from D did not respond to CCh, and responses to A23187 and NP were suppressed compared with those from C. Differences in basal, CCh, and A23187-responsive cGMP between D and C were abolished by the NO synthetase inhibitor NG-monomethyl-L-arginine. Soluble glomerular guanylate cyclase prepared from either D or C responded indistinguishably to NP, suggesting a role for NO quenching in the suppression of cGMP in intact glomeruli from D. Compared with those from C, glomeruli isolated from D demonstrated increased generation of thromboxane A2 (TXA2) and activation of protein kinase C (PKC). Both the TXA2/endoperoxide receptor antagonist Bay U3405 and inhibitors of PKC activity restored a cGMP response to CCh in glomeruli from D. Conversely, in glomeruli from C, the TXA2/endoperoxide analogue U46619 activated PKC and suppressed the cGMP response to CCh. Both of those actions were blocked by inhibitors of PKC. The results indicate a progressive impairment of NO-dependent cGMP generation in glomeruli from D which may be mediated in part by TXA2 and activation of PKC. This impairment may participate in glomerular injury in diabetes.

Renal inner medullary prostaglandin synthesis. A calcium-calmodulin-dependent process suppressed by urea.

Previous studies have demonstrated that hyperosmolar NaCl and mannitol stimulate immunoreactive prostaglandin E (iPGE) production by slices of inner medulla (IM), whereas urea inhibits this process. In the present study, the roles of Ca2+ and calmodulin in the control of PGE synthesis in IM and the basis for the differential actions of solutes were examined. A23187 increased [14C]arachidonate (AA) release and iPGE accumulation in the presence but not in the absence of media Ca2+ whereas stimulation by hypertonic NaCl or mannitol was well expressed with Ca2+ or in Ca2+-free buffer containing 2 mM EGTA. Hypertonic urea and trifluoperazine (TFP), an inhibitor of actions of the Ca2+-CaM complex, suppressed increases in [14C]AA release and iPGE induced by A23187, NaCl, or mannitol. By contrast, increases in iPGE in response to exogenous AA were not altered by urea or TFP. Ca2+ (25-100 microM) increased acyl hydrolase (AH) activity in EGTA washed (4 degrees C) 100,000 g particulate fractions of IM threefold, thereby restoring AH activity to the higher basal values of particulate fractions not washed with EGTA. This action of Ca2+ was blocked by hypertonic urea of TFP, whereas AH activity was not influenced by NaCl or mannitol in the presence or absence of Ca2+. In contrast to their effects on AH activity, hypertonic urea and TFP did not alter conversion of AA to PGE2, PGF2 alpha, or PGD2 by IM microsomal fractions. Ca2+-induced increases in particulate AH were blunted after partial depletion of endogenous CaM-like activity. Ca2+ action was restored by addition of purified exogenous CaM, but not by addition of other small acidic proteins, including troponin C. The findings support a role for CaM in the regulation of PGE synthesis in the IM at the level of Ca2+-responsive AH activity. They further imply that urea suppresses PGE synthesis in IM through inhibition of AH and a reduction in the availability of endogenous AA for conversion to PGE.

Increased angiotensin-I converting enzyme gene expression in the failing human heart. Quantification by competitive RNA polymerase chain reaction.

Local activation of the components of the renin angiotensin system in the heart is regarded as an important modulator of cardiac phenotype and function; however, little is known about their presence, regulation, and potential activation in the human heart. To investigate the gene expression of major angiotensin-II-forming enzymes in left ventricles of normal (n = 9) and failing human hearts (n = 20), we established a competitive RNA-polymerase chain reaction (PCR) for mRNA quantification of angiotensin-I converting enzyme (ACE) and human heart chymase. For each gene, competitor RNA targets with small internal deletions were used as internal standards to quantify the original number of transcripts and to control reverse transcription and PCR. In PCR, each target and the corresponding competitor were amplified by competing for the same primer oligonucleotides. The variability of ACE RNA-PCR was 11% indicating a high reproducibility of this method. In addition, ACE mRNA levels obtained by competitive RNA-PCR correlated favorably with traditional slot blot hybridization (r = 0.69, n = 10; P < 0.05). Compared with nonfailing hearts, the number of ACE transcripts referred to 100 ng of total RNA was increased threefold in patients with chronic heart failure (4.2 +/- 2.5 vs. 12.8 +/- 6 x 10(5); P < 0.0005). In contrast, no significant difference was found in chymase gene expression between normal and failing hearts. Thus, the expression of the cardiac ACE but not of human heart chymase is upregulated in failing human heart indicating an activation of the cardiac renin-angiotensin system in patients with advanced heart failure.

Na(+)-Ca2+ antiporter activity of rat hepatocytes. Effect of adrenalectomy on Ca2+ uptake and release from plasma membrane vesicles.

The presence and mode of Na(+)-Ca2+ antiporter activity were studied in hepatocytes isolated from sham-operated or adrenalectomized rats and in inside-out plasma membrane vesicles isolated from rat liver. Decreasing extracellular Na+ (Na+o) immediately increased cytosolic free calcium (Ca2+i). The rise in Ca2+i was proportional to the reduction in Na+o and was caused by an increased calcium influx, presumably on the Na(+)-Ca2+ antiporter operating in the reverse mode. Perfusing the cells with Ca(2+)-free media stimulated Ca2+ efflux and decreased Ca2+i, an effect dependent on Na+o. This suggests an activation of the forward mode of Na(+)-Ca2+ exchange. There was little difference in these parameters between sham and adx groups. In contrast, steady-state calcium uptake by inside-out plasma membrane vesicles was inhibited 40% after adrenalectomy. The decreased calcium uptake was not caused by a deficiency in the ATP-dependent Ca2+ pump, whose Km and Vmax were unaffected by adrenalectomy, but by an Na(+)-dependent leak from the vesicles. Ca2+ efflux was proportional to the extravesicular Na+ concentration, suggesting that the calcium leak may take place on a Na(+)-Ca2+ antiporter. This Na(+)-dependent calcium efflux was significantly increased in vesicles prepared from adx rat livers. These results suggest that hepatocytes have functional Na(+)-Ca2+ antiporters that can operate in both forward and reverse modes. Under normal conditions, the Na(+)-Ca2+ antiporter apparently operates in the reverse mode as a Ca2+ influx pathway. The increase in Na(+)-dependent Ca2+ efflux evoked by adrenalectomy in plasma membrane vesicles could explain the recent results we obtained in hepatocytes isolated from adx rats, showing increased calcium influx, increased Ca2+i, increased intracellular calcium sequestration, and increased plasmalemmal calcium cycling.

Adrenergic receptor properties of hepatocytes from male and female rats.

alpha 1- and beta-adrenergic receptor properties of intact hepatocytes from adult male and female rats were evaluated in ligand binding studies using [3H]prazosin and [3H]CGP-12177 (4-(t-butylamino-2-hydroxypropoxy)-[5,7-3H]benzimidazole-2-one-HCl), a hydrophilic beta antagonist. Prior work had suggested that the response of hepatocytes from males to alpha 1-adrenergic stimulation was greater than that of cells from females. However, little sexual difference in prazosin affinity, number of binding sites or kinetics of association/dissociation with the cells was found. Epinephrine, [3H]prazosin competition for binding sites on intact cells was performed at 2 degrees C and 80-90% of agonist sites remained in a high affinity state with an epinephrine Kd comparable to that previously found in glucose release and phosphorylase alpha activation studies. Agonist Kd inferred from these competition experiments also showed no sexual dimorphism. These data suggest that the greater rise in the concentration of cytosolic free calcium and release of 45Ca from cells of males in response to epinephrine stimulation is not due to male/female alpha 1-receptor differences but, rather, may be a function of the previously observed sexual difference in cell calcium metabolism. [3H]CGP binding to hepatocytes from females was stereospecific, saturable and identified a single, high affinity site. Comparable sites were not found on cells from males, however, [3H]CGP binding to crude membrane preparations from both sexes was identical. This suggests that the loss of hepatic beta-receptor function in the adult male is due to an inaccessibility of beta-receptors at the external surface of the plasma membrane of the intact cell. Further studies with other beta-receptor ligands are being carried out to confirm these initial findings.

Effect of adrenalectomy on cellular calcium metabolism and on the response to adrenergic stimulation of hepatocytes isolated from male and female rats.

The effects of adrenalectomy on cell calcium metabolism and on the effects of epinephrine on cAMP, phosphorylase a activity, and calcium efflux were studied in hepatocytes isolated from adult male and female rats. Adrenalectomy increased the total calcium of hepatocytes, all exchangeable calcium pools, and all calcium fluxes between the cellular pools in both sexes. After adrenalectomy, basal cAMP was elevated, phosphorylase a + b was decreased, but basal phosphorylase a activity was not changed. In adrenalectomized males and at all concentrations of epinephrine studied (1.10(-8)-1.10(-5)M) stimulation of calcium efflux was decreased and cAMP accumulation was enhanced, while the resulting phosphorylase a activation was depressed. In hepatocytes from adrenalectomized females there was a similar increase in cAMP accumulation induced by epinephrine, and a decrease in the stimulation of calcium efflux; however, the depression in phosphorylase a activation was much less and was significant only at 1.8(-8) and 1.10(-5)M epinephrine. In the male, while activation of phosphorylase a shifted from a pure alpha-adrenergic response mediated by calcium to one also involving a cAMP-mediated beta-adrenergic response, the contribution of the attenuated calcium signal was still significant. Hepatocytes from female rats did not show a comparable alpha- to beta-shift, since the relative contribution of calcium and cAMP to phosphorylase activation was similar in sham-operated and adrenalectomized animals.

Role for transforming growth factor beta in thromboxane-induced increases in mesangial cell fibronectin synthesis.

Thromboxane A2 (TXA2) has been implicated in the pathogenesis of progressive glomerulosclerosis and stimulates the synthesis of matrix protein by mesangial cells (MCs). This study examined the role of transforming growth factor beta (TGF-beta) in the mediation of the action of the stable TXA2/prostaglandin (PG) endoperoxide analog U-46619 to stimulate fibronectin (Fn) synthesis in cultured rat MC. Exogenous TGF-beta increased Fn synthesis by MC in a concentration- and time-dependent fashion, as reflected by incorporation of [35S]methionine into immunoprecipitable Fn. Submaximal concentrations of TGF-beta (1-2.5 pmol/l) increased Fn synthesis two- to threefold, a response comparable in magnitude to that observed with a maximal stimulatory concentration (1 mumol/l) of U-46619. Anti-TGF-beta antibody, but not isotypic IgG, blocked the increases in Fn synthesis induced by both U-46619 and exogenous TGF-beta. Endogenous TGF-beta bioactivity in MC culture media, assessed by the mink lung epithelial cell system, was significantly increased by 1 mumol/l U-46619 (1.7 +/- 0.3 pmol/l) compared with that of control media (0.6 +/- 0.1 pmol/l, P < 0.05). Total (active plus latent) TGF-beta bioactivity, assayed after heat activation of latent TGF-beta, was also significantly higher in media of MCs cultured with U-46619 (45 +/- 4 pmol/l) compared with control (24 +/- 4 pmol/l). Thus, U-46619 increased endogenous TGF-beta bioactivity to a level sufficient to account for the enhancement of Fn synthesis observed with U-46619, as reflected by the Fn synthetic response to exogenous TGF-beta.(ABSTRACT TRUNCATED AT 250 WORDS)

Role for protein kinase C in the mediation of increased fibronectin accumulation by mesangial cells grown in high-glucose medium.

The fibronectin content of RMC cultures grown for 8-14 days in medium containing 30 mM (540 mg/dl) D-glucose was increased 30-60% relative to that of control cells cultured in 10 mM (180 mg/dl) glucose. Addition of equiosmolar concentrations (20 mM, 360 mg/dl) of L-glucose, 3-O-methylglucose, or mannitol to 10 mM glucose media did not alter fibronectin accumulation compared with values observed in 10 mM glucose alone. The basal phosphorylation of the 80,000-M(r) MARKS protein, which is a substrate for PKC, and the phosphorylation induced by acute (15-min) exposure of cells to 15% FCS or to 0.1 microM (50 ng/ml) PDBu were all increased in cells grown in 30 mM compared with 10 mM glucose. By contrast, phosphorylation of the 80,000-M(r) protein in response to a maximal concentration of PDBu (1 microM, 500 ng/ml) was not different in cells grown in 30 mM compared with 10 mM glucose. The acute increases in phosphorylation of the 80,000-M(r) protein were blocked by the PKC inhibitor calphostin C. Chronic (7-day) exposure of mesangial cells grown in 10 mM glucose to 0.1 microM of the PKC agonist PDBu also resulted in a sustained 40% increase in 80,000-M(r) phosphorylation and a 20-30% increase in fibronectin accumulation. As assessed by [35S]methionine incorporation, mesangial cell fibronectin synthesis was increased by exposure to PDBu, a finding consistent with earlier observations with 30 mM glucose.(ABSTRACT TRUNCATED AT 250 WORDS)

Nitric oxide inhibition of transforming growth factor-beta and collagen synthesis in mesangial cells.

Culture of mesangial cells (MCs) in 5.6 vs. 30.0 mmol/l glucose for 3 weeks induced a sustained increase in protein kinase C (PKC) activity, transforming growth factor (TGF)-beta1 mRNA, bioactive TGF-beta, and collagen synthesis. Nitric oxide (NO), generated exogenously by the NO donor S-nitroso-N-acetyl, D,L-penicillamine (SNAP) or endogenously after the exposure of MC to interleukin-1beta (IL-1beta), suppressed bioactive TGF-beta in MCs cultured in 5.6 or 30.0 mmol/l glucose and suppressed or abolished increases in TGF-beta1 mRNA and collagen synthesis induced by high concentrations of glucose or phorbol 12,13-dibutyrate without altering values obtained with normal glucose concentrations. SNAP had a transient suppressive effect on PKC activity, which may explain at least in part some of the actions of SNAP. The selective inhibitor of PKC, bisindolylmaleimide (GFX), mimicked NO action. The ability of SNAP and IL-1beta to suppress TGF-beta and collagen synthesis was not mediated by cGMP, since the cGMP analog, 8-Br-PET-cGMP, did not mimic NO action and an antagonist of cGMP-dependent protein kinase, Rp-8-pCPT-cGMPs, did not prevent the inhibitory actions of SNAP. N-omega-L-arginine methyl ester (NMMA) increased TGF-beta in glomerular capillary endothelial cells (GCECs) and stimulated collagen synthesis by MC in a co-culture with GCECs. Captopril inhibited TGF-beta and collagen synthesis and increased cGMP in co-cultures of GCECs and MCs. These effects of captopril were abolished by NMMA, implying mediation by NO. Thus, endogenous NO produced by GCECs may modulate TGF-beta production by both GCECs and MCs and act to suppress matrix protein synthesis by MCs.

Expression, activity and functional significance of inducible nitric oxide synthase in the failing human heart.

OBJECTIVES: The study was designed to evaluate the functional impact of nitric oxide (NO) generation within the myocardium on cardiac contraction in the failing human heart. BACKGROUND: Heart failure is associated with activation of cytokines and expression of inducible nitric oxide synthase (NOS II), which generates NO from L-arginine. Nitric oxide has been shown to modulate myocardial performance, raising the possibility that cardiac generation of NO by NOS II modulates cardiac contraction in the failing human heart. METHODS: Left ventricular (LV) tissue of 24 patients with end-stage heart failure was obtained during cardiac transplantation. Gene expression of NOS II and endothelial NO-synthase (NOS III) was quantified by competitive reverse transcription-polymerase chain reaction and compared to tissues of five nonfailing donor hearts. Nitric oxide synthase II activity was determined by citrulline assay and related to changes in force of contraction induced by the beta-adrenergic agonist isoproterenol, NO-donors and/or N-mono-methyl-L-arginine (L-NMMA), an inhibitor of NOS. RESULTS: While NOS III mRNA was reduced in failing hearts, NOS II mRNA was increased in failing LV tissue and correlated with NOS II activity. High NOS II activity was associated with early relaxation and impaired responsiveness to beta-adrenergic stimulation, that is, the inotropic response to isoproterenol in failing hearts was inversely related to NOS II activity (r=0.61, p < 0.005). Nitric oxide donors or L-NMMA did not affect myocardial performance in failing hearts at baseline. However, L-NMMA enhanced the positive inotropic response to beta-adrenergic stimulation in failing hearts with high NOS II activity. Nitric oxide donors attenuated the isoproterenol-induced increase in force of contraction of failing hearts. CONCLUSIONS: Cardiac production of NO by NOS II attenuates the positive inotropic effects of beta-adrenergic stimulation and hastens relaxation in failing human hearts.

Cardiac Na+/Ca2+ exchange activity in patients with end-stage heart failure.

OBJECTIVE: The aim of the present study was to investigate the functional activity and expression of the sarcolemmal Na+/Ca2+-exchanger in the failing human heart. METHODS: Left ventricular samples were taken from eleven patients with end-stage heart failure and six organ donors (normal controls). The Na+/Ca2+-exchanger activity was assessed by measuring Na+ gradient-induced 45Ca2+ transport into sarcolemmal vesicles of quantitatively collected crude membrane preparations. The abundance of the Na+/Ca2+-exchanger protein was determined by Western blot analysis using a specific antiserum and the results were normalized to myocyte specific beta-myosin heavy chain protein content. RESULTS: In membrane preparations of failing human hearts, both the Na+ gradient-induced 45Ca2+ transport activity and the level of immunoreactive Na+/Ca2+-exchanger protein were increased (P < 0.01) by 87% and 160% compared to controls, respectively. CONCLUSIONS: In human end-stage heart failure the increased sarcolemmal Na+/Ca2+-exchanger activity appears to be due to an elevated expression of this protein. An increase in the expression and activity of the Na+/Ca2+-exchanger in the failing human heart may be of important functional significance: while a resulting increase in Ca2+ extrusion across the sarcolemma may limit diastolic Ca2+ overload, a corresponding influx of Na+ may be associated with membrane depolarization and enhanced arrhythmogenesis if the Na+/Ca2+-exchanger operates primarily in the forward mode.

Left ventricular wall stress and sarcoplasmic reticulum Ca(2+)-ATPase gene expression in renal hypertensive rats: dose-dependent effects of ACE inhibition and AT1-receptor blockade.

BACKGROUND: Cardiac hypertrophy is associated with altered Ca2+ handling and may predispose to the development of LV dysfunction and cardiac failure. At the cellular level, the re-expression of ANF represents a well-established marker of myocyte hypertrophy while the decreased expression of the sarcoplasmatic reticulum (SR) Ca(2+)-ATPase is thought o play a crucial role in the alterations of Ca2+ handling and LV function. We assessed the dose-dependent effect of chronic ACE inhibition or AT1 receptor blockade on cardiac function in relation to the cardiac expression of the SR Ca(2+)-ATPase and ANF. METHODS AND RESULTS: Renal hypertensive rats (2K-1C) were treated for 12 weeks with three different doses of the ACE inhibitor benazepril, the AT1-receptor antagonist valsartan (each drug 0.3, 3, and 10 mg/kg per day i.p.) or placebo. LV dimensions, hypertrophy and wall stress were determined in vivo by magnetic resonance imaging and the gene expressions of ANF and SR Ca(2+)-ATPase were quantified by Northern blot. Low doses of both drugs did not affect blood pressure, hypertrophy, systolic wall stress and the ANF and SR Ca(2+)-ATPase gene expression. High doses of each drug reduced systolic blood pressure, wall stress, and LV hypertrophy to a similar extent and to values comparable to normotensive, age-matched rats. In addition, high dose treatment reduced LV end-systolic and end-diastolic volume as compared to untreated 2K-1C animals and normalized the mRNA levels of both ANF and SR Ca(2+)-ATPase (as compared to normotensive animals). CONCLUSIONS: We conclude that in this model, high doses of ACE inhibition and AT1-receptor blockade are necessary to normalize systolic blood pressure, LV hypertrophy and systolic LV wall stress which, in turn, is associated with restoration of a normal cardiac phenotype with respect to SR Ca(2+)-ATPase and ANF and normalization of cardiac function.

Collembolan Transcriptomes Highlight Molecular Evolution of Hexapods and Provide Clues on the Adaptation to Terrestrial Life.

BACKGROUND: Collembola (springtails) represent a soil-living lineage of hexapods in between insects and crustaceans. Consequently, their genomes may hold key information on the early processes leading to evolution of Hexapoda from a crustacean ancestor. METHOD: We assembled and annotated transcriptomes of the Collembola Folsomia candida and Orchesella cincta, and performed comparative analysis with protein-coding gene sequences of three crustaceans and three insects to identify adaptive signatures associated with the evolution of hexapods within the pancrustacean clade. RESULTS: Assembly of the springtail transcriptomes resulted in 37,730 transcripts with predicted open reading frames for F. candida and 32,154 for O. cincta, of which 34.2% were functionally annotated for F. candida and 38.4% for O. cincta. Subsequently, we predicted orthologous clusters among eight species and applied the branch-site test to detect episodic positive selection in the Hexapoda and Collembola lineages. A subset of 250 genes showed significant positive selection along the Hexapoda branch and 57 in the Collembola lineage. Gene Ontology categories enriched in these genes include metabolism, stress response (i.e. DNA repair, immune response), ion transport, ATP metabolism, regulation and development-related processes (i.e. eye development, neurological development). CONCLUSIONS: We suggest that the identified gene families represent processes that have played a key role in the divergence of hexapods within the pancrustacean clade that eventually evolved into the most species-rich group of all animals, the hexapods. Furthermore, some adaptive signatures in collembolans may provide valuable clues to understand evolution of hexapods on land.

Toward knowledge-based liver surgery: holistic information processing for surgical decision support.

PURPOSE: Malignant neoplasms of the liver are among the most frequent cancers worldwide. Given the diversity of options for liver cancer therapy, the choice of treatment depends on various parameters including patient condition, tumor size and location, liver function, and previous interventions. To address this issue, we present the first approach to treatment strategy planning based on holistic processing of patient-individual data, practical knowledge (i.e., case knowledge), and factual knowledge (e.g., clinical guidelines and studies). METHODS: The contributions of this paper are as follows: (1) a formalized dynamic patient model that incorporates all the heterogeneous data acquired for a specific patient in the whole course of disease treatment; (2) a concept for formalizing factual knowledge; and (3) a technical infrastructure that enables storing, accessing, and processing of heterogeneous data to support clinical decision making. RESULTS: Our patient model, which currently covers 602 patient-individual parameters, was successfully instantiated for 184 patients. It was sufficiently comprehensive to serve as the basis for the formalization of a total of 72 rules extracted from studies on patients with colorectal liver metastases or hepatocellular carcinoma. For a subset of 70 patients with these diagnoses, the system derived an average of [Formula: see text] assertions per patient. CONCLUSION: The proposed concept paves the way for holistic treatment strategy planning by enabling joint storing and processing of heterogeneous data from various information sources.