Athanasia Mouzaki

Rifaximin Improves Systemic Hemodynamics and Renal Function in Patients With Alcohol-Related Cirrhosis and Ascites

Circulating levels of endotoxin, interleukin (IL)-6, and tumor necrosis factor (TNF)-α increase with intestinal bacterial overgrowth and translocation, and are believed to be involved in the pathogenesis of hyperdynamic circulatory syndrome and functional renal failure in patients with advanced cirrhosis. We investigated the effects of the antibiotic rifaximin on systemic hemodynamics and renal function in patients with alcohol-related cirrhosis and ascites. We measured mean arterial pressure, cardiac output (CO) by Doppler ultrasound, systemic vascular resistance (as the ratio of mean arterial pressure:CO), plasma renin activity, levels of plasma aldosterone, the glomerular filtration rate by plasma clearance of technetium-99m-DTPA, natriuresis, levels of plasma endotoxin, and serum levels of IL-6 and TNF-α in 13 patients at baseline and after 4 weeks of treatment with rifaximin. Rifaximin treatment significantly reduced CO and significantly increased systemic vascular resistance, in association with a significant decrease in plasma rennin activity. The therapy also significantly increased the glomerular filtration rate and natriuresis while reducing levels of endotoxin, IL-6, and TNF-α. Intestinal decontamination with rifaximin improved systemic hemodynamics and renal function in patients with advanced cirrhosis.

Molecular characterization of occult hepatitis B cases in Greek blood donors

The use of sensitive nucleic acid testing for hepatitis B virus in blood donors revealed a number of HBV DNA(+) cases among HBsAg(−) donors, a status known as occult HBV infection. The purpose of this study was the serological and molecular characterization of occult HBV infection in Greek blood donors. A prospective study was undertaken in order to identify occult HBV infection cases in blood donors. As part of the routine screening of blood donations in Greece, blood units were screened individually by a multiplex HIV‐1/HCV/HBV nucleic acid assay. Initially reactive samples were retested with discriminatory assays. HBV DNA(+)/HBsAg(−) samples were tested further for HBV serological markers and HBV DNA was quantified by real‐time PCR. Molecular characterization was performed by sequencing the envelope and polymerase genes of HBV. Preliminary screening revealed 21 occult cases with the following patterns: anti‐HBc only: 7 donors, anti‐HBc/anti‐HBs: 7 donors, anti‐HBc/anti‐HBe: 5 donors, anti‐HBc/anti‐HBs/anti‐HBe: 2 donors. In all cases, the HBV DNA load was <351 IU/ml. Sequencing was successful in 10 donors (classified within genotype D) revealing several amino acid substitutions related to diagnostic escape and antiviral resistance. HBsAg diagnostic failure and low viral replication in occult HBV infection carriers could possibly be attributed to multiple changes in envelope and polymerase regions, respectively. J. Med. Virol. 81:815–825, 2009.

Anti-TNF-α Antibody Therapies in Autoimmune Diseases

Autoimmune diseases affect about 3% of the world population, more frequently women than men, and their incidence is attributed to an immune response of a genetically predisposed individual to an environmental pathogen, under the influence of inadequate immuno-regulatory mechanisms. Advances in understanding the cellular activity pathways and cytokine expression profiles have led to new therapeutic regiments, like soluble receptors, monoclonal antibodies and molecular mimetics that have been employed to enhance or replace conventional immunosuppressive therapies. Among new biologicals that have been developed to target defined pathways of the adaptive immune response are TNF-alpha inhibitors. TNF-alpha is a proinflammatory cytokine elevated in many autoimmune lesions, and its deregulation characterizes many autoimmune diseases. TNF-alpha seems to exhibit an immunoregulatory role that can alter the balance of T regulatory cells and orchestrate acute immunological responses. More than half a million autoimmune patients have received therapy with anti-TNF-alpha antibodies, usually because they were refractory to conventional treatments. This review offers an update on TNF-alpha-targeted therapies used in patients suffering from various autoimmune diseases, based on the current knowledge of disease pathogenesis, with emphasis on the efficacy and safety that clinical trials have shown until now.

Citrullination of Linear and Cyclic Altered Peptide Ligands from Myelin Basic Protein (MBP87−99) Epitope Elicits a Th1 Polarized Response by T Cells Isolated from Multiple Sclerosis Patients: Implications in Triggering Disease

Derangement of cellular immunity is central in the pathophysiology of multiple sclerosis (MS) and is often manifested by abnormal cytokine production. We investigated cytokine secretion in peripheral blood mononuclear cells (PBMC) of 18 MS patients and 15 controls and correlated cytokine polarization with the nature of antigenic stimulus. We synthesized two novel citrullinated peptides, linear [Cit(91), Ala(96), Cit(97)]MBP(87-99) and cyclo(87-99)[Cit(91), Ala(96), Cit(97)]MBP(87-99) that resulted from citrullination of 91,97 Arg residues in antagonists, linear [Arg(91), Ala(96)]MBP(87-99) and cyclo(87-99)[Arg(91), Ala(96)]MBP(87-99) peptides. PBMC from MS patients and controls were cultured with citrullinated peptides, and both peptides caused a Th1 polarization in all MS patients studied. In contrast, culture with noncitrullinated MBP peptides resulted in heterogeneous cytokine secretion that differed between individual patients. Thus, citrullination of self-antigens may potentially trigger disease in susceptible individuals. This finding may open new avenues in drug design of new substances that inhibit citrullination and arrest epitope spreading and worsening of MS.

Treatment of experimental allergic encephalomyelitis (EAE) by a rationally designed cyclic analogue of myelin basic protein (MBP) epitope 72-85.

In this report the rational design, synthesis and pharmacological properties of an amide-linked cyclic antagonist analogue of the guinea pig myelin basic protein epitope MBP(72-85) are described. Design of the potent cyclic analogue was based on 2D NOESY nuclear magnetic resonance and molecular dynamics studies carried out in the linear antagonist Ala81MBP(72-85). The cyclic antagonist completely prevented the induction of experimental allergic/autoimmune encephalomyelitis when coinjected with linear and cyclic agonist analogues MBP(72-85) and cyclo(2-9)MBP(72-85).

Serum transforming growth factor-β1 is related to the degree of immunoparesis in patients with multiple myeloma

The expansion of myeloma celis is regulated by cytokines, among which !L-6 is a major growth factor, it has been recently suggested that serum transforming growth factorβi (TGFβi), a cytokine found in large amounts in α-granules of platelets, might play a role in multiple myeloma (MM). It was the purpose of this study to determine serum TGFβi levels in MM patients and to seek a correlation with disease parameters. Measurements were done by ELISA. We studied 35 MM patients (19 stage II, 16 stage III, 20lgG, 8lgA and 6BJ, 11gD) in different phases of the disease, 27 healthy individuals and 17 thrombocytopenic patients with other haematological diseases (three MDS, three congenital thrombocytopenia, 11ITP). Overall samples from MM patients were included: 10 at diagnosis, 18 in remission and 32 in relapse. In normal controls TGFβi serum levels ranged from 1 to 33 ng/ml (median 16.5 ng/ml). In both thrombocytopenic controls with other diseases and thrombocytopenic MM patients (seven samples), TGFβi serum levels were very low (median 3.2 and 4.5 ng/ml respectively). In MM patients with PLT > 100? 109/L (53 samples), TFGβi serum levels were in the normal range in patients without immunoparesis (1 to 27 ng/ml, median 16.6 ng/ml), whereas they were higher in patients with immunoparesis (polyclonal imrnunoglobulins (Igs) below lower normal reference values) ranging from 10.2 to 45 ng/ml (median 26.8 ng/ml) (P < 0.01). Serum TGFβi levels fluctuated in the same patient at different times but not according to relapse or remission. Correlation was found only between serum TGFβi levels and immunoparesis and not between serum TGFβi levels and disease stage or lg subtype nor with prognostic factors for MM (serum CRP, β2M or IL-6). This finding suggests that the remaining normal plasma cells are sensitive to the inhibitory action of TGFβi on Ig production. In conclusion TGFβi serum levels are very low in thrombocytopenic patients confirming that platelets are the major source of this cytokine. Furthermore, a strong correlation was found between TGFβi serum levels and immunoparesis in MM patients.

Different Immunosuppressive Combinations on T-Cell Regulation in Renal Transplant Recipients

Background/Aims: Recent studies indicate that regulatory T-cells (Tregs) promote transplant tolerance. We studied Treg levels in 39 stable renal transplant recipients to determine the sizes of the Treg populations and the effects of treatment regimens thereof. Methods: All patients (19 with good graft function and 20 with chronic allograft nephropathy) received induction therapy (basiliximab) and were on triple immunosuppressive regimens with calcineurin inhibitors (cyclosporine or tacrolimus), mycophenolate mofetil (MMF) or everolimus and steroids. Twenty healthy subjects served as controls. Whole blood samples were stained with anti-CD4, CD25, CD127, and FoxP3 antibodies and analyzed by flow cytometry to determine CD4+CD25highFoxP3± and CD4+ CD25highCD127–/low Treg levels. Results:All patients had significantly reduced CD4+CD25highFoxP3± but no CD4+ CD25highCD127–/low Treg levels compared to controls. Renal allograft function did not correlate with Treg levels. Statistically significant correlations between CD4+CD25highFoxp3+ Tregs and tacrolimus levels and CD4+CD25highFoxp3– Tregs and HLA-DR mismatching were detected. Patients receiving MMF had significantly higher CD4+CD25highFoxp3+ Tregs compared to patients on everolimus who were also receiving lower doses of calcineurin inhibitors. Conclusion:Overall, immunosuppression lowers CD4+CD25highFoxP3± Treg levels significantly in the periphery in renal transplant recipients. In addition, different immunosuppressive regimens have different impacts on CD4+CD25highFoxP3+ Tregs, a fact that may influence long-term allograft survival.

Rifaximin improves thrombocytopenia in patients with alcoholic cirrhosis in association with reduction of endotoxaemia

Thrombocytopenia is a major haematological disorder of cirrhosis with unclear pathogenesis. Endotoxaemia resulting from intestinal bacterial overgrowth could reduce platelet counts directly or through cytokine release.

Immunosuppressive Drugs in HIV Disease

Hyper activation of the immune system has emerged as an important clinical marker of HIV disease progression to AIDS. During the chronic phase of the disease, chronic immune activation is linked to systemic CD4 T-cell depletion and eventual immune failure. Additionally, the HIV virus per se seems to engage in a form of molecular parasitism for host T-cell signaling pathways and transcription factors (e.g. NFAT). Targeting host T-cell factors that mediate immune activation in conjunction with HAART (Highly Active Antiretroviral Therapy) could be the basis of novel immune-modulatory regimens that avoid the development of mutant viral strains. Hence the three-signal model of T-cell activation provides a framework for the rational selection of immunomodulatory therapies in HIV disease. Within this framework we examine the immunosuppressive, and antiretroviral properties of NFAT (calcineurin) inhibitors (cyclosporine and tacrolimus), the purine rescue pathway inhibitor mycophenolate mofetil and sirolimus (rapamycin). The results of small clinical studies to date are reviewed and they suggest that immunosuppressive medications might be a safe and effective adjunct to HAART in stable HIV disease, when such medications are used in full doses. Finally, we discuss the potential implications of such therapies for solid organ transplantation in HIV patients.

Silencing and trans-activation of the mouse IL-2 gene in Xenopus oocytes by proteins from resting and mitogen-induced primary T-lymphocytes.

The Xenopus oocyte system was used to test functionally, putative trans‐active elements involved in the transcriptional control of the mouse interleukin‐2 (IL‐2) gene in resting and mitogen‐induced primary T‐lymphocytes. The IL‐2 gene injected into the oocyte is active over a wide range of DNA concentrations. This basal activity is silenced by the addition of protein extracts from G0‐arrested spleen cells. Extracts from 8 h‐stimulated spleen cells do not silence but moderately increase transcription over basal level. When IL‐2 transcription is silenced first by an injection of extract from resting spleen cells, the addition of proteins from stimulated cells results in a strong increase in transcription (derepression). Use of proteins from purified splenic T‐lymphocytes shows that both silencer(s) and activator(s) are contributed by these cells. Extracts from control tissues have neither a silencing nor stimulatory effect. None of the proteins tested affects the activities of co‐injected control genes. Injections with IL‐2 promoter mutants indicate that the main target sequence of the silencing and activating factors is a purine region (Pu‐box) lying between positions −261 and −292 upstream of the IL‐2 gene. Bandshift assays show differential binding of the Pu‐box with proteins from resting or activated T‐cells.