Biomedical subjects

A Banerjee

Publications and source records attributed to A Banerjee.

At least 19 recordsLinked to original sources

PubMed1999

SNARE proteins regulate H(+)-ATPase redistribution to the apical membrane in rat renal inner medullary collecting duct cells.

The interaction of soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins provides the necessary steps for vesicle docking fusion. In inner medullary collecting duct (IMCD) cells, acid secretion is regulated in part by exocytotic insertion and endocytotic retrieval of an H(+)-ATPase to and from the apical membrane. We previously suggested a role for SNARE proteins in exocytotic insertion of proton pumps in IMCD cells. The purpose of the present study was to determine whether SNARE proteins are associated with the 31-kDa subunit of H(+)-ATPase in IMCD cells during exocytosis and to determine the effects of clostridial toxins on SNARE-mediated trafficking of H(+)-ATPase. Cell acidification induced a marked increment of H(+)-ATPase in the apical membrane. However, pretreating cells with clostridial toxins blocked the cellular translocation of the 31-kDa subunit. Immunoprecipitation of IMCD cell homogenate, using antibodies against either the 31-kDa subunit of H(+)-ATPase or vesicle-associated membrane protein-2, co-immunoprecipitated N-ethylmaleimide-sensitive factor, alpha-soluble NSF attachment protein (alpha-SNAP), synaptosome-associated protein-23, syntaxin, and vesicle-associated membrane protein-2. Pretreatment with clostridial toxin resulted in reduced co-immunoprecipitation of H(+)-ATPase and syntaxin. These experiments document, for the first time, a putative docking fusion complex in IMCD cells and a physical association of the H(+)-ATPase with the complex. The sensitivity to the action of clostridial toxin indicates the docking-fusion complex is a part of the exocytotic mechanism of the proton pump.

Animals↗

PubMed1999

A confocal microscopic study relating the autofluorescence of carious dentine to its microhardness.

OBJECTIVE: An in-vitro study to examine the correlation between the distribution of the autofluorescent signal emitted from carious dentine (detected using confocal laser scanning microscopy) and its microhardness, within the depths of human dentine lesions. MATERIALS AND METHODS: Twelve carious teeth were sectioned longitudinally, the cut faces marked with a grid reference line system and colour photomicrographs taken. The same samples were imaged using confocal laser-scanning microscopy for autofluorescence and then subjected to microhardness testing using a Knoop microhardness indenter. Adjacent sound dentine was used as a control reference. Digital image superimposition allowed direct comparisons to be made between the colour, autofluorescence and microhardness of each lesion. RESULTS: Sound enamel and dentine did not autofluorescence. Autofluorescence distribution from carious dentine correlated with the highly softened tissue (detected using the Knoop indenter) and terminated at a level superficial to the translucent zone. This zone was still pigmented. Normal, sound dentine hardness levels were found deep to the translucent zone. CONCLUSIONS: A correlation existed between the zone of autofluorescence and carious dentine that was markedly softened by the carious process. These findings highlighted a possibility that the autofluorescence might be used as an in-vitro, objective histological marker for the softened, carious dentine requiring clinical excavation.

Color↗

PubMed1999

Disordered N-terminal residues affect the folding thermodynamics and kinetics of maltose binding protein.

Maltose binding protein (MBP) exhibits a slow phase of folding at pH 7.4, 298 K. The kinetics of this phase has been characterized as a function of denaturant concentration and temperature. Denaturant double-jump experiments and the activation energy for folding indicate that the slow phase involves processes other than proline isomerization. Although the first five N-terminal residues are disordered in the MBP crystal structure, mutations in this region slow down folding and destabilize the native structure. This is the first report showing that disordered N-terminal residues can affect folding kinetics and stability.

ATP-Binding Cassette Transporters↗

PubMed1999

Inhibition of myocardial TNF-alpha production by heat shock. A potential mechanism of stress-induced cardioprotection against postischemic dysfunction.

Overproduction of tumor necrosis factor-alpha (TNF-alpha) contributes to cardiac dysfunction associated with systemic or myocardial stress, such as endotoxemia and myocardial ischemia/reperfusion (I/R). Heat shock has been demonstrated to enhance cardiac functional resistance to I/R. However, the protective mechanisms remain unclear. The purpose of this study was to determine: (1) whether cardiac macrophages express heat shock protein 72 (HSP72) after heat shock, (2) whether induced cardiac HSP72 suppresses myocardial TNF-alpha production during I/R, and (3) whether preservation of postischemic myocardial function by heat shock is correlated with attenuated TNF-alpha production during I/R. Rats were subjected to heat shock (42 degrees C for 15 min) and 24 h recovery. Immunoblotting confirmed the expression of cardiac HSP72. Immunofluorescent staining detected HSP72 in cardiac interstitial cells including resident macrophages rather than myocytes. Global I/R caused a significant increase in myocardial TNF-alpha. The increase in myocardial TNF-alpha was blunted by prior heat shock and the reduced myocardial TNF-alpha level was correlated with improved cardiac functional recovery. This study demonstrates for the first time that heat shock induces HSP72 in cardiac resident macrophages and inhibits myocardial TNF-alpha production during I/R. These observations suggest that inhibition of myocardial TNF-alpha production may be a mechanism by which HSP72 protects the heart against postischemic dysfunction.

Animals↗

PubMed1999

p38 MAPK inhibition decreases TNF-alpha production and enhances postischemic human myocardial function.

INTRODUCTION: TNF-alpha is a proinflammatory cytokine implicated in myocardial dysfunction following ischemia/reperfusion (I/R). I/R results in myocardial production of TNF-alpha and TNF-alpha suppresses myocardial contractility. p38 mitogen-activated protein kinase (MAPK) is a redox-sensitive protein kinase involved in intracellular signaling leading to TNF-alpha production. It remains unknown if the human heart produces TNF-alpha after I/R and, if so, whether p38 MAPK is involved. HYPOTHESIS: p38 MAPK inhibition enhances human myocardial post-I/R contractile function by inhibition of myocardial TNF-alpha production. METHODS: Human atrial trabeculae were suspended in organ baths, field simulated at 1 Hz, and force development was recorded. Following a 90-min equilibration, trabeculae were exposed to a p38 MAPK inhibitor (SB 203580, 1 microM) or vehicle (each n = 6) prior to simulated ischemia (45 min hypoxia, substrate-free, rapid pacing at 3 Hz) followed by 120 min reoxygenation. Myocardial TNF-alpha levels were measured by ELISA at end reoxygenation. RESULTS: I/R increased human myocardial TNF-alpha levels from 26.9 +/- 9.3 to 83.9 +/- 19.2 pg/g wet tissue (P < 0.05 perfusion vs I/R; ANOVA Bonferroni/Dunn), while p38 MAPK inhibition decreased post-I/R myocardial TNF-alpha levels to 32.3 +/- 8.0 pg/g wet tissue (P > 0.05 p38 MAPK inhibition vs I/R). p38 MAPK inhibition improved postischemic force development from 18.5 +/- 2.1 to 37.0 +/- 2.0% baseline developed force (%BDF; P < 0.05 I/R vs p38 MAPK inhibition). CONCLUSIONS: (1) The human heart produces TNF-alpha after I/R, (2) p38 MAPK mediates myocardial I/R-induced TNF-alpha production, (3) p38 MAPK inhibition limits functional impairment after I/R, and (4) inhibition of ischemia-induced TNF-alpha production may represent a potent therapeutic strategy for improving myocardial function after angioplasty, coronary bypass, or heart transplantation.

Calcium-Calmodulin-Dependent Protein Kinases↗

PubMed1999

LPS-Induced NF-kappaB activation and TNF-alpha release in human monocytes are protein tyrosine kinase dependent and protein kinase C independent.

BACKGROUND: Tumor necrosis factor alpha (TNF-alpha) is an important mediator of septic shock. Endotoxin (LPS) signal transduction in human monocytes leads to activation of nuclear factor-kappa B (NF-kappaB) and TNF-alpha release. Previous studies have implicated activation of both protein kinase C (PKC) and protein tyrosine kinases (PTK) in LPS-induced NF-kappaB activation and TNF-alpha production. We hypothesized that inhibition of either PKC or PTK would decrease LPS-induced NF-kappaB DNA binding and TNF-alpha release in human monocytes. MATERIALS AND METHODS: Human monocytes were stimulated with PMA (50 ng/ml) alone or LPS (100 ng/ml) with and without a nonspecific serine/threonine protein kinase inhibitor staurosporine (Stauro), a specific pan-PKC inhibitor bisindolylmaleimide (Bis), or an inhibitor of PTK genistein (Gen). TNF-alpha release in culture supernatants was measured by an ELISA. NF-kappaB DNA binding was evaluated by electrophoretic mobility shift assay. RESULTS: LPS increased NF-kappaB DNA binding and TNF-alpha release in human monocytes. Nonspecific protein kinase inhibition inhibited NF-kappaB activation and TNF-alpha release, while specific PKC inhibition with Bis had no effect on LPS-induced NF-kappaB DNA binding or TNF-alpha release. PTK inhibition with Gen attenuated both LPS-induced NF-kappaB DNA binding and TNF-alpha production in human monocytes. Direct activation of PKC with PMA induced both NF-kappaB activation and TNF-alpha production by human monocytes. CONCLUSIONS: These results suggest that LPS-induced NF-kappaB activation and TNF-alpha release in human monocytes are independent of PKC activity. Furthermore, our results provide evidence that PTK plays a role in LPS-induced NF-kappaB activation and TNF-alpha release in human monocytes and thus could be a potential therapeutic target in inflammatory states.

Cells, Cultured↗

PubMed1999

Liposomal delivery of purified inhibitory-kappaBalpha inhibits tumor necrosis factor-alpha-induced human vascular smooth muscle proliferation.

Vessel injury results in the elaboration of various cytokines, including tumor necrosis factor-alpha (TNF-alpha), which may influence vascular smooth muscle cell (VSMC) function and contribute to atherogenesis. We tested the hypothesis that TNF-alpha-induced VSMC proliferation requires activation of the transcription factor nuclear factor-kappaB (NF-kappaB), which could be prevented by delivery of the NF-kappaB inhibitory peptide, IkappaBalpha. TNF-alpha induced concentration-dependent human VSMC proliferation, and neutralizing antibody to interleukin-6 reduced TNF-alpha-induced VSMC proliferation by 65%. In TNF-alpha-stimulated VSMCs, there was a 3-fold increase in NF-kappaB-dependent luciferase reporter activity that was associated with degradation of IkappaBalpha. To determine an essential role for NF-kappaB in TNF-alpha-induced VSMC proliferation, recombinant IkappaBalpha was introduced into VSMCs via liposomal delivery. Under these conditions, TNF-alpha-induced NF-kappaB nuclear translocation and DNA binding were inhibited, NF-kappaB-dependent luciferase activity was reduced by 50%, there was no degradation of native IkappaBalpha detected, interleukin-6 production was reduced by 54%, and VSMC proliferation was decreased by 60%. In conclusion, the mitogenic effect of TNF-alpha on human arterial VSMCs is dependent on NF-kappaB activation and may be prevented by exogenously delivered IkappaBalpha. Furthermore, liposomal delivery of endogenous inhibitory proteins may represent a novel, therapeutically accessible method for selective transcriptional suppression in the response to vascular injury.

Cell Division↗

PubMed1999

A monoclonal antibody to alpha-tubulin: purification of functionally active alpha-tubulin isoforms.

Both alpha- and beta-tubulin exist as numerous isotypic forms that originate from different primary sequences as well as a variety of posttranslational modifications. Recent studies show that tubulin dimers differing in the beta-subunit differ significantly in their subcellular distribution as well as in their functional properties such as assembly, dynamics, conformation, and interaction with antimitotic drugs; however, very little is known about the functional significance of the different alpha-tubulin isoforms and their posttranslational modifications. In an effort to get a better understanding about the alpha-tubulin isoforms, a monoclonal antibody, AYN.6D10, was prepared against the mammalian alpha-tubulin C-terminal sequence Glu-Glu-Gly-Glu-Glu-Tyr. Using an immunoaffinity column, bovine brain tubulin was fractionated into three functionally active alphabeta heterodimers which were identified by immunoblotting with alpha-tubulin-specific antibodies and sequence analysis. Assembly studies in the presence of glycerol and Mg2+ show that one of the fractions, that contains mainly the tyrosinated form of alpha1/2, assembled poorly, while the nontyrosinated form assembled normally. The results indicate that tubulin dimers differing in their alpha-tubulin may differ in their functional properties. Future studies with the isoforms may yield valuable information regarding the role of alpha-tubulin and its posttranslational modifications in regulating microtubule assembly and function in vivo.

Amino Acid Sequence↗

PubMed1999

Echocardiographic characteristics of successful deployment of the Das AngelWings atrial septal defect closure device: initial multicenter experience in the United States.

The AngelWings device is a newer transcatheter device used for closure of secundum atrial septal defects (ASD) and patent foramen ovale (PFO), which consists of a self-centering, 2-disk system. Transesophageal echocardiography (TEE) plays a pivotal role in the deployment of the 2 disks of this device, on the appropriate sides of the atrial septum. The objective of this study is to describe the echocardiographic findings associated with successful deployment of the AngelWings device for closure of ASD and PFO. We evaluated the TEE studies of 70 patients enrolled in 4 United States centers, for closure of ASD and PFO with the AngelWings device. The TEE characteristics of successful and unsuccessful deployments were analyzed. Residual shunts across the atrial septum were assessed by TEE at the end of the procedure, 24 hours later by transthoracic echocardiography, and at 6 months by TEE. The deployment of the device was successful in 65 patients (93%). In the unsuccessful group, ASD size by TEE was larger (13.4 +/- 3.1 vs 8.9 +/- 4.7 mm, p <0.05). TEE was successful in identifying snagging of the device by intracardiac structures and prolapse of corners of the left or right atrial disk through the ASD, features that were difficult to identify by fluoroscopy. The echocardiographic characteristics outlined here are important guidelines for successful deployment of the AngelWings device.

Adolescent↗

PubMed1999

Interaction of novel thiocolchicine analogs with the tubulin isoforms from bovine brain.

The antimitotic alkaloid colchicine binds to tubulin and inhibits microtubule assembly. Recently a new series of colchicine derivatives has been synthesized in which the seven-membered B-ring was shortened to a six-membered ring. In an effort to study the role of the B-ring substituents in this new series, we have studied the interaction of two compounds of this series, THC 5 and THC 18, with tubulin isoforms from bovine brain. We find that THC 18, which has a side chain with a pi-bonded SP2 conformation, binds differently to the tubulin isoforms, while THC 5 with a slightly different side chain does not. The results indicate that the conformation of the B-ring domain plays a major role in the differential interaction of a colchicine derivative with different tubulin isoforms. The results will be very important in designing potent antitumor derivatives of colchicine.

Animals↗

PubMed1999

PKC-dependent preconditioning with norepinephrine protects sarcoplasmic reticulum function in rat trabeculae following metabolic inhibition.

The authors have previously shown that norepinephrine (NE) pretreatment attenuates Ca2+ overloading in cardiac rat trabeculae during metabolic inhibition, and improves contractile function during a subsequent recovery period. The present study investigated: (i) whether protection of sarcoplasmic reticulum (SR) function during metabolic inhibition (MI) is involved in the preconditioning-like effect of NE-pretreatment, and (ii) whether or not this process is PKC-dependent. A 15 min preincubation period was used with 1 micromol/l exogenous NE to precondition isolated, superfused rat trabeculae against contractile dysfunctioning following 40 min of MI in 2 mmol/l NaCN containing Tyrode (gassed with 95% O2/5% CO2; pH 7.4, 24 degrees C) without glucose at 1-Hz stimulation frequency. Contractile recovery was studied during a subsequent 60 min recovery period (RP) in glucose containing Tyrode at 0.2 Hz. Force and intracellular free calcium ([Ca2+]ii) were monitored throughout the experimental protocol. Pretreatment of trabeculae with NE (group NE) substantially diminished the Ca2+ rise from the onset of rigor development during MI, compared to preparations which were pretreated with NE, in the presence of specific PKC blocker chelerythrine (2 micromol/l; group NE+CHEL). After 40 min of MI, resting [Ca2+]i in group NE and NE+CHEL was increased to 0.50+/-0.03 and 2.08+/-0.20 micromol/l, respectively (P<0.05), whereas total intracellular ATP levels were similar in both groups (approximately 0.20 micromol/g dry wt). This corresponded with an increase in active force development (119%) and a decrease in twitch force relaxation time (77%) during subsequent RP in group NE, compared to pre-MI values of the same group. In contrast, a significant decrease in force recovery (54%) and an increase in twitch force relaxation time (123%) was observed in group NE+CHEL. Values for [Ca2+]i, contractile recovery, and twitch force relaxation time in untreated controls as well as CHEL preparations corresponded to those measured in the NE+CHEL group. Rapid cooling contractures (RCCs), which provide information on both SR-Ca2+ loading and Ca2+ re-uptake activity, revealed a 2-fold higher SR Ca2+ content during RP in group NE compared to controls and group NE+CHEL. In addition, kinetic analysis of the RCC rewarming spike (RWS) showed that this was accompanied by greater than a 28% increase in the maximum rate of RWS relaxation (-dF/dt/rws) in group NE compared to group NE+CHEL. The change of -dF/dt/rws in the NE group during RP following MI persisted after SR Ca2+-release channel blockade by ryanodine treatment (100 micromol/l), which suggests involvement of NE-induced, PKC-dependent protection of SR Ca2+-ATPase activity. The results of the present study point to an inverse relationship between the Ca2+ rise during MI and SR functioning, in which PKC appears to play a key role. It is concluded that the preconditioning-like effect of NE-pretreatment on contractile recovery is at least partly mediated by protection of SR function.

Adenosine Triphosphate↗

PubMed1999

Calcium preconditioning, but not ischemic preconditioning, bypasses the adenosine triphosphate-dependent potassium (KATP) channel.

BACKGROUND: Recent evidence has implicated the KATP channel as an important mediator of ischemic preconditioning (IPC). Indeed, patients taking oral sulfonylurea hypoglycemic agents (i.e., KATP channel inhibitors) for treatment of diabetes mellitus are resistant to the otherwise profoundly protective effects of IPC. Unfortunately, many cardiopulmonary bypass patients, who may benefit from IPC, are chronically exposed to these agents. Calcium preconditioning (CPC) is a potent form of similar myocardial protection which may or may not utilize the KATP channel in its mechanism of protection. The purpose of this study was to determine whether CPC may bypass the KATP channel in its mechanism of action. If so, CPC may offer an alternative to IPC in patients chronically exposed to these agents. METHODS: Isolated rat hearts (n = 6-8/group) were perfused (Langendorff) and received KATP channel inhibition (glibenclamide) or saline vehicle 10 min prior to either a CPC or IPC preconditioning stimulus or neither (ischemia and reperfusion, I/R). Hearts were subjected to global warm I/R (20 min/40 min). Postischemic myocardial functional recovery was determined by measuring developed pressure (DP), coronary flow (CF), and compliance (end diastolic pressure, EDP) with a MacLab pressure digitizer. RESULTS: Both CPC and IPC stimuli protected myocardium against postischemic dysfunction (P < 0.05 vs I/R; ANOVA with Bonferroni/Dunn): DP increased from 52 +/- 4 (I/R) to 79 +/- 2 and 83 +/- 4 mmHg; CF increased from 11 +/- 0.7 to 17 +/- 2 and 16 +/- 1 ml/min; and EDP decreased (compliance improved) from 50 +/- 7 to 27 +/- 5 and 31 +/- 7 mmHg. However, KATP channel inhibition abolished protection in hearts preconditioned with IPC (P < 0.05 vs IPC alone), but not in those preconditioned with CPC (P > 0.05 vs CPC alone). CONCLUSIONS: (1) Both IPC and CPC provide similar myocardial protection; (2) IPC and CPC operate via different mechanisms; i.e., IPC utilizes the KATP channel whereas CPC does not; and (3) CPC may offer a means of bypassing the deleterious effects of KATP channel inhibition in diabetic patients chronically exposed to oral sulfonylurea hypoglycemic agents.

Adenosine Triphosphate↗

PubMed1999

Peritoneal cytology in colorectal cancer.

Explore the source record for details and available documents.

Antineoplastic Agents↗

PubMed1999

The NFkappaB inhibitory peptide, IkappaBalpha, prevents human vascular smooth muscle proliferation.

BACKGROUND: Vessel injury results in an inflammatory response characterized by the elaboration of cytokines and growth factors, which ultimately influence vascular smooth muscle cell (VSMC) growth and contribute to atherogenesis. Nuclear factor-kappa B (NFkappaB) is a central transcription factor important in mediating stress and inflammatory-induced signals. We hypothesized that strategies aimed at inhibiting NFkappaB would abrogate mitogen-induced human VSMC proliferation. METHODS: Human aortic VSMC were stimulated with basic fibroblast growth factor (FGF) and tumor necrosis factor-alpha (TNF), and proliferation was quantified by a colormetric assay. The influence of NFkappaB on VSMC proliferation was examined by both nonspecific NFkappaB blockade with calpain inhibitor-1 (CI-1) and dexamethasone (Dex) and specific NFkappaB blockade with liposomal delivery of the NFkappaB inhibitory peptide, IkappaBalpha. RESULTS: FGF and TNF induced concentration-dependent VSMC proliferation (p < 0.002). Neither CI-1, Dex, nor liposomal IkappaBalpha influenced proliferation of unstimulated VSMC. However, both FGF- and TNF-stimulated VSMC proliferation was inhibited to the level of control with CI-1, Dex, and liposomal IkappaBalpha (p < 0.001). CONCLUSION: The mitogenic effect of FGF and TNF on human arterial VSMC may be prevented by inhibiting NFkappaB. Furthermore, liposomal delivery of endogenous inhibitory proteins such as IkappaBalpha may represent a novel, therapeutically accessible method for selective transcriptional suppression in the response to vascular injury.

Aorta↗

PubMed1999

Clonal heterogeneity of dendritic cells derived from patients with chronic myeloid leukemia and enhancement of their T-cells stimulatory activity by IFN-alpha.

Adoptive immunotherapy in form of donor leukocyte infusions is effective in a significant number of patients with chronic myeloid leukemia (CML) that have relapsed after allogeneic bone marrow transplantation (BMT). However, the therapy is associated with clinically significant side effects such as graft-versus-host disease (GVHD) and bone marrow (BM) hypoplasia that may be avoided through the administration of T cells with specific antileukemic activity. Dendritic cells (DC) functioning as potent antigen presenting cells (APC) may play an important role in the generation of T cells with specificity against CML. We examined a subpopulation of CD1a+/CD14- DC generated in vitro from BM of normal subjects and patients with CML using granulocyte-macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor-alpha (TNF-alpha) and interleukin-4 (IL-4). These DC derived from both the BM of normal subjects and of patients with CML, differentiated and matured in culture in a similar way. However, DC derived from patients with CML, displayed decreased activity when tested with allogeneic T cells in a mixed lymphocyte reaction (MLR). Addition of interferon-alpha (IFN-alpha) to DC cultures significantly upregulated the expression of major histocompatibility complex (MHC) molecules (class I and class II) and costimulatory molecules (B7.1 and B7.2) on DC from normal donors and CML patients. However, DC grown from CML patients required a higher concentration of IFN-alpha. IFN-alpha also significantly improved the capacity of CML DC to stimulate T-lymphocyte responses. Fluorescence in situ hybridization (FISH) showed that only some CD1a+/CD14- DC derived from BM of patients with CML expressed the bcr/abl fusion gene. Incubation with INF-alpha decreased the proportion of bcr/abl positive DC.

Antigens, CD↗

PubMed1999

Tumor necrosis factor-alpha and interleukin-1beta synergistically depress human myocardial function.

OBJECTIVE: Proinflammatory cytokines such as tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta have been implicated in the pathogenesis of myocardial dysfunction in ischemia-reperfusion injury, sepsis, chronic heart failure, viral myocarditis, and cardiac allograft rejection. Although circulating TNF-alpha and IL-1beta are both often elevated in septic shock, it remains unknown whether TNF-alpha or IL-1beta are the factors induced during sepsis that directly depress human myocardial function, and if so, whether the combination synergistically depresses myocardial function. Furthermore, the mechanism(s) by which these cytokines induce human myocardial depression remain unknown. We hypothesized the following: a) TNF-alpha and IL-1beta directly depress human myocardial function; b) together, TNF-alpha and IL-1beta act synergistically to depress human myocardial function; and c) inhibition of ceramidase or nitric oxide synthase attenuates myocardial depression induced by TNF-alpha or IL-1beta by limiting proximal cytokine signaling or production of myocardial nitric oxide (NO). DESIGN: Prospective, randomized, controlled study. SETTING: Experimental laboratory in a university hospital. SUBJECTS: Freshly obtained human myocardial trabeculae. INTERVENTIONS: Human atrial trabeculae were obtained at the time of cardiac surgery, suspended in organ baths, and field simulated at 1 Hz, and the developed force was recorded. After a 90-min equilibration, TNF-alpha (1.25, 12.5, 125, or 250 pg/mL for 20 mins), IL-1beta (6.25, 12.5, 50, or 200 pg/mL for 20 mins), or TNF-alpha (1.25 pg/mL) plus IL-1beta (6.25 pg/mL) were added to the bath, and function was measured for the subsequent 100 mins after the 20-min exposure. To assess the roles of the sphingomyelin and NO pathways in TNF-alpha and IL-1beta cross-signaling, the ceramidase inhibitor N-oleoyl ethanolamine (1 microM) or the NO synthase inhibitor N(G)-monomethyl-L-arginine (10 microM) was added before TNF-alpha (125 pg/mL) or IL-1beta (50 pg/mL). MEASUREMENTS AND MAIN RESULTS: TNF-alpha and IL-1beta each depressed human myocardial function in a dose-dependent fashion (maximally depressing to 16.2 + 1.9% baseline developed force for TNF-alpha and 25.7 + 6.3% baseline developed force for IL-1beta), affecting systolic relatively more than diastolic performance (each p < .05). However, when combined, TNF-alpha and IL-1beta at concentrations that did not individually result in depression (p > .05 vs. control) resulted in contractile depression (p < .05 vs. control). Inhibition of myocardial sphingosine or NO release abolished the myocardial depressive effects of either TNF-alpha or IL-1beta. CONCLUSIONS: TNF-alpha and IL-1beta separately and synergistically depress human myocardial function. Sphingosine likely participates in the TNF-alpha and IL-1beta signal leading to human myocardial functional depression. Therapeutic strategies to reduce production or signaling of either TNF-alpha or IL-1beta may limit myocardial dysfunction in sepsis.

Amidohydrolases↗

PubMed1999

Reduction of infarct size in the rat heart by LPS preconditioning is associated with expression of angiogenic growth factors and increased capillary density.

Inflammation induces the expression of angiogenic growth factors in tissues, which leads to microvascular growth. Bacterial lipopolysaccharide (LPS) provokes a transient inflammatory response in the heart and induces delayed cardiac resistance to post-ischemic contractile dysfunction. In this study, we examined: 1) the effects of LPS on myocardial expression of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF), 2) whether an increase in the density of myocardial microvessels follows the expression of angiogenic growth factors, and 3) the effect of LPS on myocardial resistance to infarction and its relationship with microvascular growth. Rats were treated with LPS (from Salmonella typhimurium, 0.5 mg/kg i.p.). The expression of bFGF and VEGF in the myocardium was examined at 6 and 12 h after LPS treatment by immunofluorescent staining. Myocardial capillary and arteriole densities were determined 3 days after LPS treatment by morphometry, using immunofluorescent staining of von Willebrand factor (a marker protein of endothelial cells) and alpha-smooth muscle actin (a marker protein of smooth muscle cells). To examine cardiac resistance to infarction, hearts were subjected to 40 min of regional ischemia and 2 h of reperfusion by reversible occlusion of left coronary artery at 3 days after LPS treatment. LPS induced cardiac bFGF and VEGF at 6 and 12 h after treatment. The expression of these growth factors was followed by an increase in myocardial capillary density (2032 +/- 78/mm2 vs. 1617 +/- 47/mm2 in saline control, P < 0.05), but not arteriole density, at 3 days. Meanwhile, infarct size was significantly reduced by LPS preconditioning (infarct/left ventricle 12.3 +/- 1.04% vs. 21.7 +/- 1.65% in saline control, 43% reduction, P < 0.05). These results suggest that LPS preconditioning induces cardiac bFGF and VEGF, and an increase in myocardial capillary density. This increased myocardial capillary density is associated with a reduced infarct size after in vivo regional ischemia-reperfusion.

Animals↗

PubMed1999

Inhibition of PARS attenuates endotoxin-induced dysfunction of pulmonary vasorelaxation.

Endotoxin (Etx) causes excessive activation of the nuclear repair enzyme poly(ADP-ribose) synthase (PARS), which depletes cellular energy stores and leads to vascular dysfunction. We hypothesized that PARS inhibition would attenuate injury to mechanisms of pulmonary vasorelaxation in acute lung injury. The purpose of this study was to determine the effect of in vivo PARS inhibition on Etx-induced dysfunction of pulmonary vasorelaxation. Rats received intraperitoneal saline or Etx (Salmonella typhimurium; 20 mg/kg) and one of the PARS inhibitors, 3-aminobenzamide (3-AB; 10 mg/kg) or nicotinamide (Nic; 200 mg/kg), 90 min later. After 6 h, concentration-response curves were determined in isolated pulmonary arterial rings. Etx impaired endothelium-dependent (response to ACh and calcium ionophore) and -independent (sodium nitroprusside) cGMP-mediated vasorelaxation. 3-AB and Nic attenuated Etx-induced impairment of endothelium-dependent and -independent pulmonary vasorelaxation. 3-AB and Nic had no effect on Etx-induced increases in lung myeloperoxidase activity and edema. Lung ATP decreased after Etx but was maintained by 3-AB and Nic. Pulmonary arterial PARS activity increased fivefold after Etx, which 3-AB and Nic prevented. The beneficial effects were not observed with benzoic acid, a structural analog of 3-AB that does not inhibit PARS. Our results suggest that PARS inhibition with 3-AB or Nic improves pulmonary vasorelaxation and preserves lung ATP levels in acute lung injury.

Adenosine Triphosphate↗