Innate production of interleukin-10 and tumor necrosis factor affects the risk of multiple sclerosis

Multiple sclerosis (MS) typically presents with a relapsing-remitting onset. This can be distinguished from primary progressive MS. Typical MS is characterized by a profound inflammatory reaction in which anti-inflammatory cytokine interleukin-10 (IL-10) and pro-inflammatory cytokine tumor necrosis factor (TNF) may play a pivotal role. We tested the hypothesis that patients with MS have a distinct innate cytokine production that contributes to the susceptibility for and outcome of MS. The innate cytokine production of patients was estimated as the average production of cytokines in lipopolysaccharide -stimulated whole-blood cultures of 2 to 5 first-degree healthy family members. A total of 126 family members of 50 patients with typical MS, 61 family members of 25 patients with primary progressive MS, and 129 control subjects of 54 families were enrolled in this study. We found that members of families with low IL-10 and high TNF production had a fourfold increased risk of developing typical MS compared with members of families with high IL-10 and low TNF production. Patients with MS were eightfold more likely to develop typical MS than primary progressive MS when they belonged to families with low IL-10 and high TNF production. The presence of human leukocyte antigen-DR2 was associated with MS but not with TNF production. This study shows that typical MS is associated with an innate pro-inflammatory cytokine profile in contrast to primary progressive MS.     

From "our world" to the "real world": Exploring the views and behaviour of policy-influential Australian public health researchers

Research and researchers influence the genesis and development of public health policy in limited but essential ways. Surveys and interviews with 36 peer-nominated "highly influential" Australian public health researchers found they engaged in a breadth of strategies that included rigorous but targeted research design, multilateral collaboration, multiple methods of research dissemination and promotion (including tactical use of the media), and purposeful development of bridging relationships. Researchers' ability to understand the worlds of research, policy and the media and to speak their languages (or to work with others who fulfilled this role) was a key factor. Advocacy was seen as fundamental by some but was disparaged by others. Influential behaviours were guided by values and beliefs about the principles underlying traditional science and the contrasting ethos of contemporary research. This study may help researchers consider their own policy-related roles, strategies and relationships in the context of increasing calls for research that serves economic and/or social goals.     

Distribution and quantitation of the anti-trypanosomal diamidine 2,5-bis(4-amidinophenyl)furan (DB75) and its N-methoxy prodrug DB289 in murine brain tissue

The current epidemic of sleeping sickness, also known as human African trypanosomiasis in sub-Saharan Africa places nearly 60 million people at risk for developing this life threatening infection. Although effective treatments for early-stage sleeping sickness exist, these drugs usually require extended dosing schedules and intravenous administration. New treatments are also needed for cerebral (late) stage trypanosomiasis. 2,5-Bis(4-amidinophenyl)furan (DB75), a pentamidine analog, has potent in vitro and in vivo anti-trypanosomal activity. However, DB75 does not exhibit significant oral bioavailability and has proved to be ineffective against mouse models of late-stage sleeping sickness regardless of administration route. To circumvent the limited oral bioavailability of DB75, an N-methoxy prodrug 2,5-bis(4-amidinophenyl)furan-bis-O-methylamidoxime (DB289) was designed and developed initially as a compound to treat AIDS-related Pneumocystis carinii pneumonia (PCP). Despite excellent oral activity against early-stage sleeping sickness, oral administration of DB289 exhibited limited efficacy in mouse models of late-stage disease. DB289 has recently entered Phase II(b) clinical trials to treat primary-stage sleeping sickness in Central Africa. The current study takes advantage of the innate fluorescence of DB75 and DB289 along with specific and sensitive quantitative analyses to examine plasma and brain distribution of these compounds. Animals were dosed with intravenous DB75, oral DB289, and intravenous DB289. Following intravenous administration, DB75 was readily detectable in whole brain extracts and persisted for long periods. Fluorescence microscopy revealed that DB75 did not penetrate into brain parenchyma, however, but was sequestered within cells lining the blood-brain and blood-cerebrospinal fluid barriers. In contrast, brain tissue of mice treated with oral DB289 exhibited diffuse fluorescence within the brain parenchyma, suggesting that the prodrug was not trapped within blood-brain barrier cells (BBB). However, maximal brain concentrations of the active compound DB75 were very low (13 nmol/mg of tissue at 24 h). Intravenous administration of DB289 resulted in a qualitatively similar fluorescence pattern to oral DB289, indicating again that DB289 and DB75 were present within brain parenchyma, not only in barrier regions. Furthermore, peak DB75 tissue levels were higher (61 nmol/mg of tissue at 24 h) than with oral prodrug. The near five-fold increase in brain levels of DB289 combined with parenchymal localization of compound fluorescence after intravenous administration suggest that the unaltered prodrug penetrates the blood-brain barrier, and may be subject to in situ biotransformation. Intravenous administration of DB289 should be evaluated in mouse models of late-stage sleeping sickness.     

Tissue plasminogen activator activity and inhibition in acute myocardial infarction and angiographically normal coronary arteries

Parameters of blood coagulation, blood platelet reactivity and fibrinolysis were analyzed in 18 patients with acute myocardial infarction (AMI) and angiographically normal coronary arteries. This study group was compared with a patient control group of 18 AMI patients with 1-vessel obstructive coronary artery disease. Patients were matched for sex, age and AMI date. A healthy control group consisted of 18 sex- and age-matched volunteers. Blood coagulation measurements and platelet reactivity were similar in the 3 groups, except for fibrinogen, which was significantly higher in the patient control group. Plasma activity of tissue plasminogen activator (tPA) was detectable in only 2 patients in the study group, 9 in the patient control group (p less than 0.02) and 12 in the healthy control group (p less than 0.0001). Median plasma tPA inhibitory activity was higher in the study group (10.3 IU/ml) and the patient control group (8.1 IU/ml) than in the healthy control group (2.7 IU/ml, p less than 0.0001 and p less than 0.03). Thus, reduced activity and enhanced inhibition of plasma tPA may be important factors in the origin of coronary thrombosis, especially in the absence of coronary artery disease.     

Activated complement components and complement activator molecules on the surface of cell-derived microparticles in patients with rheumatoid arthritis and healthy individuals

OBJECTIVES: In vitro, microparticles can activate complement via the classical pathway. If demonstrable ex vivo, this mechanism may contribute to the pathogenesis of rheumatoid arthritis (RA). We therefore investigated the presence of activated complement components and complement activator molecules on the surface of cell-derived microparticles of RA patients and healthy individuals. METHODS: Microparticles from synovial fluid (n = 8) and plasma (n = 9) of 10 RA patients and plasma of sex- and age-matched healthy individuals (n = 10) were analysed by flow cytometry for bound complement components (C1q, C4, C3) and complement activator molecules (C-reactive protein (CRP), serum amyloid P component (SAP), immunoglobulin (Ig) M, IgG). RESULTS: Microparticles with bound C1q, C4, and/or C3 were abundant in RA synovial fluid, while in RA and control plasma much lower levels were present. Microparticles with bound C1q correlated with those with bound C3 in synovial fluid (r = 0.961, p = 0.0001), and with those with bound C4 in plasma (RA: r = 0.908, p = 0.0007; control: r = 0.632, p = 0.0498), indicating classical pathway activation. In synovial fluid, microparticles with IgM and IgG correlated with those with C1q (r = 0.728, p = 0.0408; r = 0.952, p = 0.0003, respectively), and in plasma, microparticles with CRP correlated with those with C1q (RA: r = 0.903, p = 0.0021; control: r = 0.683, p = 0.0296), implicating IgG and IgM in the classical pathway activation in RA synovial fluid, and CRP in the low level classical pathway activation in plasma. CONCLUSIONS: This study demonstrates the presence of bound complement components and activator molecules on microparticles ex vivo, and supports their role in low grade complement activation in plasma and increased complement activation in RA synovial fluid.     

Two novel and one recurrent missense mutation in the factor XIII A gene in two Dutch patients with factor XIII deficiency

Congenital factor XIII (FXIII) deficiency is a rare autosomal recessive disorder, usually attributed to a defect in the FXIII A subunit, whose genetic basis has been studied in a number of cases. We describe here the genetic variations found in two unrelated patients with FXIII deficiency. Both patients, under prophylactic substitution with FXIII concentrate, showed low plasma FXIII A subunit antigen levels with undetectable A subunit antigen in the platelets and normal plasma B antigen levels, which indicate that the defects are present in the A subunit of the molecule. Both probands were heterozygous for a previously reported G-->A transversion in exon 8 of the FXIII A subunit gene (Arg326Gln substitution). Proband 1 was also heterozygous for a novel G-->T transversion in exon 7, which predicts a Val316Phe substitution. Two of her sons were heterozygous for this mutation and showed low FXIII activity and FXIII A subunit antigen levels. Val316 is a well-conserved amino acid among the transglutaminase family, located within the core domain, close to the Cys314 member of the catalytic triad. Proband 2 had a unique 2-bp (TT) insertion in one of the alleles within or adjacent to the -7 to -20 T tail of intron A. This insertion was not found in 50 healthy individuals, which supports this being the second mutation in this patient.     

Cellular origin and procoagulant properties of microparticles in meningococcal sepsis

Patients with meningococcal sepsis generally suffer from disseminated intravascular coagulation (DIC). The aim of this study was to address whether these patients have elevated numbers of circulating microparticles that contribute to the development of DIC. Plasma samples from 5 survivors, 2 nonsurvivors, and 5 healthy volunteers were analyzed for the presence of microparticles by flow cytometry. Ongoing coagulation activation in vivo was quantified by enzyme-linked immunosorbent assay of plasma prothrombin fragment F(1 + 2), and procoagulant properties of microparticles in vitro were estimated by thrombin-generation assay. On admission, all patients had increased numbers of microparticles originating from platelets or granulocytes when compared with controls (P =.004 and P =.008, respectively). Patients had elevated levels of F(1 + 2) (P =.004), and their microparticles supported thrombin generation more strongly in vitro (P =.003) than those of controls. Plasma from the patient with the most fulminant disease course and severe DIC contained microparticles that expressed both CD14 and tissue factor, and these microparticles demonstrated extreme thrombin generation in vitro. We conclude that patients with meningococcal sepsis have elevated numbers of circulating microparticles that are procoagulant. These findings may suggest a novel therapeutic approach to combat clinical conditions with excessive coagulation activation.     

Microparticle-associated P-selectin reflects platelet activation in preeclampsia

Platelet activation in preeclampsia is reflected by elevated levels of platelets exposing P-selectin. In plasma, a non-cell bound (soluble) form of P-selectin is present. Elevated levels of this soluble form have been reported in preeclampsia. Plasma P-selectin may consist of two fractions: microparticle (MP)--associated P-selectin and non-MP--associated P-selectin. In the present cross-sectional study, we investigated to which extent plasma P-selectin is MP--associated and whether such MP are elevated in preeclamptic patients. Preeclamptic patients (n?=?10) were matched with normotensive pregnant women (n?=?10) and non-pregnant controls (n?=?10). Plasma P-selectin was measured by ELISA. MP were isolated, double labelled with anti-CD61 (GPIIIa) and anti-CD62P (P-selectin) and subsequently analyzed with flowcytometry. Plasma P-selectin concentration was elevated in preeclamptic patients compared to non-pregnant controls (p?=?0.007), but not compared to normotensive pregnant women (p?=?0.210). Plasma P-selectin is partially MP--associated (3–5%). In pregnancy, the fraction of P-selectin exposing platelet-derived MP (PMP) (10.9%) was increased compared to non-pregnant controls (8%). This fraction further increased in preeclamptic patients (15.4%), and significantly differed from normotensive pregnant women (p?=?0.02). A minor fraction of plasma P-selectin is associated with PMP. The fraction of PMP exposing P-selectin is increased in preeclamptic patients and to a lesser extent in normotensive pregnancy. Because MP associated P-selectin exclusively originates from platelets, this fraction indicates platelet activation. Platelet activation is prominent in preeclampsia and this study proves that at least a part of the plasma P-selectin originates from platelets.