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Biomedical subjects

V J Marder

Publications and source records attributed to V J Marder.

At least 37 records · Page 2Linked to original sources
PubMed1999

Thrombogenic factors and recurrent coronary events.

BACKGROUND: Thrombosis is a pivotal event in the pathogenesis of coronary disease. We hypothesized that the presence of blood factors that reflect enhanced thrombogenic activity would be associated with an increased risk of recurrent coronary events during long-term follow-up of patients who have recovered from myocardial infarction. METHODS AND RESULTS: We prospectively enrolled 1045 patients 2 months after an index myocardial infarction. Baseline thrombogenic blood tests included 6 hemostatic variables (D-dimer, fibrinogen, factor VII, factor VIIa, von Willebrand factor, and plasminogen activator inhibitor-1), 7 lipid factors [cholesterol, triglycerides, HDL cholesterol, LDL cholesterol, lipoprotein(a), apolipoprotein (apo)A-I, and apoB], and insulin. Patients were followed up for an average of 26 months, with the primary end point being coronary death or nonfatal myocardial infarction, whichever occurred first. The hemostatic, lipid, and insulin parameters were dichotomized into their top and the lower 3 risk quartiles and evaluated for entry into a Cox survivorship model. High levels of D-dimer (hazard ratio, 2.43; 95% CI, 1.49, 3.97) and apoB (hazard ratio, 1.82; 95% CI, 1.10, 3.00) and low levels of apoA-I (hazard ratio, 1.84; 95% CI, 1.10, 3.08) were independently associated with recurrent coronary events in the Cox model after adjustment for 6 relevant clinical covariates. CONCLUSIONS: Our findings indicate that a procoagulant state, as reflected in elevated levels of D-dimer, and disordered lipid transport, as indicated by low apoA-1 and high apoB levels, contribute independently to recurrent coronary events in postinfarction patients.

Adult↗
PubMed1999

Hematologic neoplasia and the central nervous system.

Central nervous system (CNS) involvement with malignant cells is a well recognized complication of hematologic neoplasms. A number of disorders such as acute lymphoblastic leukemia and high grade lymphoma frequently involve the CNS and prophylactic therapy is advised. Disorders such as acute myeloid leukemia (AML) and multiple myeloma are less likely to be associated with CNS involvement. This series describes three cases of CNS involvement by malignant hematologic disease: myelomatous meningitis, CNS chloromas complicating AML, and primary lymphomatous meningitis.

Adult↗
PubMed1999

A soluble fibrin standard: comparable dose-response with immunologic and functional assays.

A soluble fibrin (SF) preparation has been developed as a potential standard by the Scientific and Standardization Committee for use in assays evaluating in vitro preparations and patient plasma samples. The SF standard was prepared by reaction of factor XIII-free fibrinogen with thrombin, followed by neutralization with hirudin and solubilization of the fibrin in acetic acid. As characterized by SDS-PAGE, the polypeptide chain structure shows the anticipated loss of fibrinopeptides and lack of gamma or alpha chain crosslinking. The standard was added to pooled normal plasma at concentrations from 12.5 microg/ml to 340 microg/ml and tested with four commercially available assays based on immunologic reactions using ELISA or latex agglutination or on t-PA cofactor activity for plasminogen to plasmin conversion. Absolute "soluble fibrin" concentrations were calculated using the manufacturers' calibrators and showed distinct dose-response relationships for each assay. Expression of the results following log-transformation produced a series of parallel lines, indicating that this SF preparation can serve as a standard, effectively normalizing the disparate proprietary internal calibrators currently used for each assay.

Biological Assay↗
PubMed1999

Automated latex agglutination and ELISA testing yield equivalent D-dimer results in patients with recent myocardial infarction. THROMBO Research Investigators.

Our previous prospective study of post-infarction patients described a strong and significant association of increased plasma D-dimer concentrations in those who experienced a subsequent coronary death or non-fatal myocardial infarction. In the present study, we compare results on stored plasma obtained two months after the index myocardial infarction from 1,038 patients of this trial, using a simple automated latex agglutination (LA) assay in parallel with the standard ELISA test. Results show a somewhat higher mean value for the LA assay (702+/-1092 vs. 638+/-986 ng/ml, p = 0.0002), a strong linear correlation of the two assays (r = 0.86) and 88% agreement for values below 500 ng/ml by the ELISA test. D-dimer concentrations determined by each assay were highly correlated in patients with subsequent coronary artery events (p = 0.93) and quartile values for both the LA and ELISA were equally predictive of such events (p = 0.003 and p = 0.001, respectively). This is the first demonstration that a latex agglutination assay for D-dimer can be used to assess the prognostic risk of recurrent coronary thrombotic disease after myocardial infarction

Adult↗
PubMed1999

Phospholipid-bound tissue factor modulates both thrombin generation and APC-mediated factor Va inactivation.

Hemostasis is initiated by tissue factor (TF) exposed on cellular phospholipid (PL) membranes, leading to thrombin generation. The binding of thrombin to thrombomodulin (TM), activates the protein C pathway, resulting in the inactivation of factors Va and VIIIa by activated protein C (APC) and a negative feedback effect on thrombin generation. A new assay system was developed for simultaneous measurement of thrombin and APC generation in defibrinated plasma induced by large unilamellar PL vesicles complexed with full-length recombinant TF (TF:PL). TF:PL preparations with a low TF concentration induced an initial rate of thrombin generation below 100 nM/min, and resulted in less thrombin formation in the presence of TM than in its absence. In contrast, TF:PL preparations with a high concentration of TF induced a higher rate of thrombin generation, and APC-mediated feedback inhibition did not occur, despite maximal APC generation. We used the same TF:PL surfaces to study factor Va inactivation by APC in a non-plasma reaction system, and found an inverse correlation between TF surface density and the rate of factor Va inactivation. This observation suggests a previously unrecognized hemostatic effect of TF, namely a non-enzymatic surface density-based inhibition of the anticoagulant effect of APC. In this model, high concentrations and surface density of TF exert complementary effects by promoting the regular procoagulant cascade and by inhibiting the protein C pathway, thereby maximizing hemostasis after vascular injury.

Animals↗
PubMed1999

Reactivity of soluble fibrin assays with plasmic degradation products of fibrin and in patients receiving fibrinolytic therapy.

The ability to identify the products of thrombin and plasmin action on fibrinogen is important in patients with thrombotic and fibrinolytic disorders. New assays have been developed for "soluble fibrin" which represents soluble derivatives other than fibrinopeptides formed from fibrinogen by thrombin. These assays are either immunological, using antibodies for fibrin-specific neoepitopes, or functional and based on the cofactor activity of soluble fibrin in the tissue plasminogen activator (t-PA)-mediated conversion of plasminogen to plasmin. As plasmic derivations of fibrin share structural features with soluble fibrin, they may be reactive with assays for soluble fibrin. Therefore, we prepared plasmic digests of fibrin and determined the degree of reactivity with four soluble fibrin assays. Three assays used Mabs directed toward the fibrin-specific neoepitopes at alpha17-23 (A), gamma312-324 (B) and alpha17-78 (D). A fourth (C) was based on t-PA co-factor activity. Tests A and C demonstrated marked crossreactivity with fibrin degradation products, and digests containing the largest derivatives showed greatest reactivity. Plasmic derivatives of crosslinked fibrin had greater reactivity than those of non-crosslinked fibrin. Tests B and D demonstrated minimal reactivity with plasmic derivatives of crosslinked or of non-crosslinked fibrin. Samples from patients with lower limb peripheral arterial occlusion were assayed for soluble fibrin, D-dimer and fibrinogen at presentation and eight hours after thrombolytic therapy. Variable results were seen at presentation with elevations in 13, 1, 0 and 4 of 19 patients using Tests A, B, C and D, respectively. After fibrinolytic therapy, marked increases in soluble fibrin levels were observed up to 600-fold above normal. A strong correlation between baseline levels was observed with Test B and Test D, which showed the least cross-reactivity with plasmic derivations. After thrombolytic therapy there were either weak or no correlations among the different assays. The results demonstrate that assays for soluble fibrin may react with plasmic derivatives of fibrin and this must be considered in interpreting clinical results.

Anticoagulants↗
PubMed1998

NF-kappa B-dependent inhibition of apoptosis is essential for host cellsurvival during Rickettsia rickettsii infection.

The possibility that bacteria may have evolved strategies to overcome host cell apoptosis was explored by using Rickettsia rickettsii, an obligate intracellular Gram-negative bacteria that is the etiologic agent of Rocky Mountain spotted fever. The vascular endothelial cell, the primary target cell during in vivo infection, exhibits no evidence of apoptosis during natural infection and is maintained for a sufficient time to allow replication and cell-to-cell spread prior to eventual death due to necrotic damage. Prior work in our laboratory demonstrated that R. rickettsii infection activates the transcription factor NF-kappa B and alters expression of several genes under its control. However, when R. rickettsii-induced activation of NF-kappa B was inhibited, apoptosis of infected but not uninfected endothelial cells rapidly ensued. In addition, human embryonic fibroblasts stably transfected with a superrepressor mutant inhibitory subunit Ikappa B that rendered NF-kappa B inactivatable also underwent apoptosis when infected, whereas infected wild-type human embryonic fibroblasts survived. R. rickettsii, therefore, appeared to inhibit host cell apoptosis via a mechanism dependent on NF-kappa B activation. Apoptotic nuclear changes correlated with presence of intracellular organisms and thus this previously unrecognized proapoptotic signal, masked by concomitant NF-kappa B activation, likely required intracellular infection. Our studies demonstrate that a bacterial organism can exert an antiapoptotic effect, thus modulating the host cell's apoptotic response to its own advantage by potentially allowing the host cell to remain as a site of infection.

Apoptosis↗
PubMed1998

Transcriptional regulation of endothelial cell tissue factor expression during Rickettsia rickettsii infection: involvement of the transcription factor NF-kappaB.

The vascular endothelial cell (EC) is a primary target of infection with Rickettsia rickettsii, the etiologic agent of Rocky Mountain spotted fever. Changes in gene transcription elicited by intracellular infection, including EC expression of the coagulation pathway initiator known as tissue factor (TF), may contribute to the vascular pathology observed during disease. Nuclear run-on analysis of uninfected and infected, cultured human endothelial cells revealed that the rate of TF mRNA transcription is enhanced more than twofold at 3 h following infection, thus coinciding with increased steady-state levels of TF mRNA. TF mRNA remained relatively unstable during infection, with a half-life of 1.6 h. The eukaryotic protein synthesis inhibitor cycloheximide did not block R. rickettsii-induced increase in TF mRNA levels and actually resulted in its superinduction, thus revealing that de novo synthesis of host cell protein was not prerequisite to this transcriptional response. Involvement of the transcription factor NF-kappaB in R. rickettsii-induced TF expression was demonstrated by using two unrelated inhibitors of NF-kappaB activation. The antioxidant pyrrolidinedithiocarbamate and the proteasome inhibitor N-tosyl-L-phenylalanine chloromethyl ketone blocked expression of TF mRNA and activity during infection. This study demonstrates that R. rickettsii infection results in transcriptional activation of the TF gene and that this response involves activation of the transcription factor NF-kappaB.

Cells, Cultured↗
PubMed1998

Proteasome-independent activation of nuclear factor kappaB in cytoplasmic extracts from human endothelial cells by Rickettsia rickettsii.

Interaction of many infectious agents with eukaryotic host cells is known to cause activation of the ubiquitous transcription factor nuclear factor kappaB (NF-kappaB) (U. Siebenlist, G. Franzoso, and K. Brown, Annu. Rev. Cell Biol. 10:405-455, 1994). Recently, we reported a biphasic pattern of NF-kappaB activation in cultured human umbilical vein endothelial cells consequent to infection with Rickettsia rickettsii, an obligate intracellular gram-negative bacterium and the etiologic agent of Rocky Mountain spotted fever (L. A. Sporn, S. K. Sahni, N. B. Lerner, V. J. Marder, D. J. Silverman, L. C. Turpin, and A. L. Schwab, Infect. Immun. 65:2786-2791, 1997). In the present study, we describe activation of NF-kappaB in a cell-free system, accomplished by addition of partially purified R. rickettsii to endothelial cell cytoplasmic extracts. This activation was rapid, reaching maximal levels at 60 min, and was dependent on the number of R. rickettsii organisms added. Antibody supershift assays using monospecific antisera against NF-kappaB subunits (p50 and p65) confirmed the authenticity of the gel-shifted complexes and identified both p50-p50 homodimers and p50-p65 heterodimers as constituents of the activated NF-kappaB pool. Activation occurred independently of the presence of endothelial cell membranes and was not inhibited by removal of the endothelial cell proteasome. Lack of involvement of the proteasome was further confirmed in assays using the peptide-aldehyde proteasome inhibitor MG 132. Activation was not ATP dependent since no change in activation resulted from addition of an excess of the unhydrolyzable ATP analog ATPgammaS, supplementation with exogenous ATP, or hydrolysis of endogenous ATP with ATPase. Furthermore, Western blot analysis before and after in vitro activation failed to demonstrate phosphorylation of serine 32 or degradation of the cytoplasmic pool of IkappaB alpha. This lack of IkappaB alpha involvement was supported by the finding that R. rickettsii can induce NF-kappaB activation in cytoplasmic extracts prepared from T24 bladder carcinoma cells and human embryo fibroblasts stably transfected with a superrepressor phosphorylation mutant of IkappaB alpha, rendering NF-kappaB inactivatable by many known signals. Thus, evidence is provided for a potentially novel NF-kappaB activation pathway wherein R. rickettsii may interact with and activate host cell transcriptional machinery independently of the involvement of the proteasome or known signal transduction pathways.

Cells, Cultured↗
PubMed1997

Unusual anemias.

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Adult↗
PubMed1997

Primary fibrinolysis during supraceliac aortic clamping.

PURPOSE: An increased incidence of bleeding complications has been observed after supraceliac aortic clamping (SCC). This study was performed to identify possible hemostatic abnormalities that contribute to this problem. METHODS: A prospective cohort study over a 3-month period was performed by comparing hemostatic parameters in 10 consecutive patients who required elective SCC with those of eight concurrent randomly selected control subjects who required infrarenal clamping (IRC) for abdominal aortic reconstruction. Measures of coagulation, fibrinolysis, platelet function, temperature, hemodilution, and hepatic function were performed at selected times before, during, and after operation. RESULTS: Aneurysm size, fibrinogen, D-dimers, prothrombin, partial thromboplastin time, platelet counts, bleeding times, hemodilution, and temperature were comparable in both groups. Patients in the SCC group, however, consistently developed a primary fibrinolytic state within 20 minutes after supraceliac clamping, reflected by significantly decreased euglobulin clot lysis times (ECLT; p < 0.0001), elevated tissue plasminogen activator (t-PA) levels (p < 0.0006), elevated t-PA-to-plasminogen activator inhibitor-1 ratios (p < 0.0001), and reduced alpha 2-antiplasmin levels (p < 0.002). SCC produced hepatocellular injury documented by elevations in both aspartate transaminase (p < 0.0001) and lactate dehydrogenase (p < 0.009). CONCLUSIONS: SCC rapidly induces a primary fibrinolytic state manifested by increased circulating t-PA, reduced alpha 2-antiplasmin, and increased fibrinolytic activator-to-inhibitor ratios. These effects may be a result of hepatic hypoperfusion caused by SCC leading to insufficient clearance of t-PA. Antifibrinolytic agents may be of benefit if bleeding develops after aortic procedures that require supraceliac clamping.

Aged↗
PubMed1997

Rickettsia rickettsii infection of cultured human endothelial cells induces NF-kappaB activation.

Rickettsia rickettsii, the etiologic agent of Rocky Mountain spotted fever, is an obligate intracellular bacterial organism that infects primarily the vascular endothelial cells (EC). A component of the EC response to infection is transcriptional activation, which may contribute to the thrombotic and inflammatory consequences of disease. In this study, we explore R. rickettsii-induced activation of the nuclear factor-kappaB/Rel (NF-kappaB) family of transcription factors involved in early transcriptional responses to injurious stimuli. Two NF-kappaB species were activated by infection and reacted with a double-stranded oligonucleotide probe corresponding to the kappaB binding domain of the murine kappa light-chain gene enhancer. Gel supershift analysis demonstrated the reactivity of these complexes with antibodies against p65 and p50, and the induced species were tentatively identified as p50-p50 homodimers and p50-p65 heterodimers. Semiquantitative reverse transcription-PCR analysis revealed dramatic increases in the steady-state levels of mRNA coding for the inhibitory subunit of NF-kappaB (IkappaB alpha), transcription of which is enhanced by the binding of NF-kappaB within the IkappaB alpha promoter region. NF-kappaB activation was first detected 1.5 h following infection and was biphasic, with an early peak of activation at approximately 3 h, a return to baseline levels at 14 h, and even higher levels of activation at 24 h. It is likely that NF-kappaB activation requires cellular uptake of R. rickettsii, since treatment of EC with cytochalasin B during infection to block entry inhibited activation by only 70% at 3 h. R. rickettsii-induced activation of NF-kappaB may be an important controlling factor in the transcriptional responses of EC to infection with this obligate intracellular organism.

Cells, Cultured↗
PubMed1997

Polarized secretion of fibrinogen by lung epithelial cells.

The lung epithelium has recently been identified as a novel site of fibrinogen (FBG) biosynthesis. A coordinated upregulation of A alpha, B beta, and gamma chain FBG gene transcription occurs upon stimulation of A549 lung epithelial cells with dexamethasone (DEX) and the proinflammatory mediator interleukin-6 (IL-6). Subsequently, the cells synthesize and secrete fully assembled FBG. This study addresses the polarity of such FBG secretion by A549 cells cultured on polycarbonate membrane filters. After induction with IL-6 and DEX, cells were metabolically labeled, and FBG was immunopurified from the apical and basolateral chambers. Analysis by gel electrophoresis revealed that A549 cells secreted newly synthesized FBG in a polarized manner, with the majority (80%) of FBG secreted basolaterally. Consistent with this observation, immunoelectron microscopy using Protein A-gold labeling showed FBG within secretory vesicles in close proximity to the basolateral aspect of the A549 cell membrane. Polarized secretion was microtubule-dependent since depolymerization using colchicine significantly reduced the basolateral component of secretion, causing intracellular retention of FBG. These data provide evidence that FBG is secreted by lung alveolar epithelial cells vectorially toward the basement membrane, which may reflect in vivo processes associated with local injury, inflammation, and repair mechanisms.

Cell Line↗
PubMed1997

Prevention of deep-vein thrombosis after total hip arthroplasty. Comparison of warfarin and dalteparin.

The effectiveness and safety of warfarin were compared with those of a low-molecular-weight heparin (dalteparin) for the prevention of deep-vein thrombosis after total hip arthroplasty in a prospective, randomized, multi-institutional trial. Patients who were older than eighteen years of age and were scheduled to have an elective primary or revision total hip arthroplasty were eligible; 580 patients were randomized, 550 had the operation and received prophylaxis, and 382 had evaluable venograms. Prophylaxis was provided either with warfarin beginning the night before the operation or with dalteparin beginning two hours before the operation and was continued until venography was performed. Bleeding was assessed on the basis of intraoperative blood loss, transfusion requirements, a decrease in hematocrit, and clinically identified bleeding complications. The prevalence of deep-vein thrombosis was found to be significantly lower in the patients who had received dalteparin than in those who had received warfarin (twenty-eight [15 per cent] of 192 patients compared with forty-nine [26 per cent] of 190 patients; p = 0.006). Deep-vein thrombosis occurred in the calf veins of twenty-one patients (11 per cent) who had received dalteparin and of forty-three patients (23 per cent) who had received warfarin; this difference was significant (p = 0.003). Proximal deep-vein thrombosis occurred in ten patients (5 per cent) who had received dalteparin and in sixteen patients (8 per cent) who had received warfarin; however, with the numbers available, no significant difference could be detected (p = 0.185). We also could not detect a significant difference with regard to the intraoperative and postoperative blood loss, the decrease in hematocrit, and the prevalence of major bleeding complications between the two groups; however, the patients who had received dalteparin had a significantly higher prevalence of bleeding complications involving the operative site (p = 0.03), and a significantly greater percentage required postoperative transfusions (p = 0.001). We concluded that preoperative prophylaxis with dalteparin is significantly more effective than that with warfarin in preventing deep-vein thrombosis after total hip arthroplasty. The greater effectiveness of dalteparin must be considered, however, in light of an increased need for postoperative transfusions and an increase in the prevalence of wound-related bleeding complications.

Anticoagulants↗