BK Virus Infection and BK-Virus-Associated Nephropathy in Renal Transplant Recipients

Poliomavirus BK virus (BKV) is highly infective, causing asymptomatic infections during childhood. After the initial infection, a stable state of latent infection is recognized in kidney tubular cells and the uroepithelium with negligible clinical consequences. BKV is an important risk factor for BKV-associated diseases, and, in particular, for BKV-associated nephropathy (BKVN) in renal transplanted recipients (RTRs). BKVN affects up to 10% of renal transplanted recipients, and results in graft loss in up to 50% of those affected. Unfortunately, treatments for BK virus infection are restricted, and there is no efficient prophylaxis. In addition, consequent immunosuppressive therapy reduction contributes to immune rejection. Increasing surveillance and early diagnosis based upon easy and rapid analyses are resulting in more beneficial outcomes. In this report, the current status and perspectives in the diagnosis and treatment of BKV in RTRs are reviewed

Hydrogen Sulfide, a Toxic Gas with Cardiovascular Properties in Uremia: How Harmful Is It?

Hydrogen sulfide (H2S) is a poisonous gas which can be lethal. However, it is also produced endogenously, thus belonging to the family of gasotransmitters along with nitric oxide and carbon monoxide. H2S is in fact involved in mediating several signaling and cytoprotective functions, for example in the nervous, cardiovascular, and gastrointestinal systems, such as neuronal transmission, blood pressure regulation and insulin release, among others. When increased, it can mediate inflammation and apoptosis, with a role in shock. When decreased, it can be involved in atherosclerosis, hypertension, myocardial infarction, diabetes, sexual dysfunction, and gastric ulcer; it notably interacts with the other gaseous mediators. Cystathionine γ-lyase, cystathionine β-synthase, and 3-mercaptopyruvate sulfurtransferase are the principal enzymes involved in H2S production. We have recently studied H2S metabolism in the plasma of chronic hemodialysis patients and reported that its levels are significantly decreased. The plausible mechanism lies in the transcription inhibition of the cystathionine γ-lyase gene. The finding could be of importance considering that hypertension and high cardiovascular mortality are characteristic in these patients

Plasma proteins containing damaged L-isoaspartyl residues are increased in uremia: Implications for mechanism

Plasma proteins containing damaged L-isoaspartyl residues are increased in uremia: Implications for mechanism.BackgroundSeveral alterations of protein structure and function have been reported in uremia. Impairment of a transmethylation-dependent protein repair mechanism possibly related to a derangement in homocysteine metabolism is also present in this condition, causing erythrocyte membrane protein damage. Homocysteine may affect proteins via the accumulation of its parent compound S-adenosylhomocysteine (AdoHcy), a powerful in vivo methyltransferase inhibitor. However, since plasma homocysteine is mostly protein bound, a direct influence on protein structures cannot be ruled out. We measured the levels of L-isoaspartyl residues in plasma proteins of uremic patients on hemodialysis. These damaged residues are markers of molecular age, which accumulate when transmethylation-dependent protein repair is inhibited and/or protein instability is increased.MethodsL-isoaspartyl residues in plasma proteins were quantitated using human recombinant protein carboxyl methyl transferase (PCMT). Plasma concentrations of homocysteine metabolites were also measured under different experimental conditions in hemodialysis patients.ResultsThe concentration of damaged plasma proteins was increased almost twofold compared to control (controls 147.83 ± 17.75, uremics 282.80 ± 26.40 pmol of incorporated methyl groups/mg protein, P < 0.003). The major protein involved comigrated with serum albumin. Although hyperhomocysteinemia caused a redistribution of thiols bound to plasma proteins, this mechanism did not significantly contribute to the increase in isoaspartyl residues. The S-adenosylmethionine (AdoMet)/AdoHcy concentration ratio, an indicator of the flux of methyl group transfer, was altered. This ratio was partially corrected by folate treatment (0.385 ± 0.046 vs. 0.682 ± 0.115, P < 0.01), but protein L-isoaspartate content was not.ConclusionsPlasma protein damage, as determined by protein L-isoaspartyl content, is increased in uremia. This alteration is to be ascribed to an increased protein structural instability, rather than the effect of hyperhomocysteinemia

Uremic Toxin Lanthionine Interferes with the Transsulfuration Pathway, Angiogenetic Signaling and Increases Intracellular Calcium

(1) The beneficial effects of hydrogen sulfide (H2S) on the cardiovascular and nervous system have recently been re-evaluated. It has been shown that lanthionine, a side product of H2S biosynthesis, previously used as a marker for H2S production, is dramatically increased in circulation in uremia, while H2S release is impaired. Thus, lanthionine could be classified as a novel uremic toxin. Our research was aimed at defining the mechanism(s) for lanthionine toxicity. (2) The effect of lanthionine on H2S release was tested by a novel lead acetate strip test (LAST) in EA.hy926 cell cultures. Effects of glutathione, as a redox agent, were assayed. Levels of sulfane sulfur were evaluated using the SSP4 probe and flow cytometry. Protein content and glutathionylation were analyzed by Western Blotting and immunoprecipitation, respectively. Gene expression and miRNA levels were assessed by qPCR. (3) We demonstrated that, in endothelial cells, lanthionine hampers H2S release; reduces protein content and glutathionylation of transsulfuration enzyme cystathionine--synthase; modifies the expression of miR-200c and miR-423; lowers expression of vascular endothelial growth factor VEGF; increases Ca2+ levels. (4) Lanthionine-induced alterations in cell cultures, which involve both sulfur amino acid metabolism and calcium homeostasis, are consistent with uremic dysfunctional characteristics and further support the uremic toxin role of this amino acid

Nanostructured Photoelectrochemical Biosensing Platform for Cancer Biomarker Detection

The innovative photoelectrochemical properties of multifunctional nanomaterials are here investigated for the development of biosensing platforms for rapid and sensitive detection of a class of cancer biomarker candidates, known as microRNAs. Many different transducers have been proposed, so far, for microRNA detection. Recently, with the emergence of novel photoelectrochemically active species and new detection schemes, photoelectrochemistry has received increasing attention. Gold nanostructures have been, here, used to modify TiO2 electrodes. The surface of the nanostructured platform has been modified by nucleic acid capture probes (CPs). Biotinylated target miRNAs have been recognized by the specific CPs. The biosensing platform has been incubated with streptavidin alkaline phosphatase and exposed to a proper substrate. The product of the enzymatic reaction has been photoelectrochemically monitored. A compact and hand-held analytical device has been developed in order to have a final prototype in line with the concept of point of care testing. (C) 2017 The Authors. Published by Elsevier Ltd

Protein Isoaspartate Methyltransferase Prevents Apoptosis Induced by Oxidative Stress in Endothelial Cells: Role of Bcl-Xl Deamidation and Methylation

BACKGROUND:Natural proteins undergo in vivo spontaneous post-biosynthetic deamidation of specific asparagine residues with isoaspartyl formation. Deamidated-isomerized molecules are both structurally and functionally altered. The enzyme isoaspartyl protein carboxyl-O-methyltransferase (PCMT; EC 2.1.1.77) has peculiar substrate specificity towards these deamidated proteins. It catalyzes methyl esterification of the free alpha-carboxyl group at the isoaspartyl site, thus initiating the repair of these abnormal proteins through the conversion of the isopeptide bond into a normal alpha-peptide bond. Deamidation occurs slowly during cellular and molecular aging, being accelerated by physical-chemical stresses brought to the living cells. Previous evidence supports a role of protein deamidation in the acquisition of susceptibility to apoptosis. Aim of this work was to shed a light on the role of PCMT in apoptosis clarifying the relevant mechanism(s). METHODOLOGY/PRINCIPAL FINDINGS:Endothelial cells transiently transfected with various constructs of PCMT, i.e. overexpressing wild type PCMT or negative dominants, were used to investigate the role of protein methylation during apoptosis induced by oxidative stress (H(2)O(2); 0.1-0.5 mM range). Results show that A) Cells overexpressing "wild type" human PCMT were resistant to apoptosis, whereas overexpression of antisense PCMT induces high sensitivity to apoptosis even at low H(2)O(2) concentrations. B) PCMT protective effect is specifically due to its methyltransferase activity rather than to any other non-enzymatic interactions. In fact negative dominants, overexpressing PCMT mutants devoid of catalytic activity do not prevent apoptosis. C) Cells transfected with antisense PCMT, or overexpressing a PCMT mutant, accumulate isoaspartyl-containing damaged proteins upon H(2)O(2) treatment. Proteomics allowed the identification of proteins, which are both PCMT substrates and apoptosis effectors, whose deamidation occurs under oxidative stress conditions leading to programmed cell death. These proteins, including Hsp70, Hsp90, actin, and Bcl-xL, are recognized and methylated by PCMT, according to the general repair mechanism of this methyltransferase. CONCLUSION/SIGNIFICANCE:Apoptosis can be modulated by "on/off" switch partitioning the amount of specific protein effectors, which are either in their active (native) or inactive (deamidated) molecular forms. Deamidated proteins can also be functionally restored through methylation. Bcl-xL provides a case for the role of PCMT in the maintenance of functional stability of this antiapoptotic protein

Homocysteinylated Albumin Promotes Increased Monocyte-Endothelial Cell Adhesion and Up-Regulation of MCP1, Hsp60 and ADAM17

RATIONALE:The cardiovascular risk factor homocysteine is mainly bound to proteins in human plasma, and it has been hypothesized that homocysteinylated proteins are important mediators of the toxic effects of hyperhomocysteinemia. It has been recently demonstrated that homocysteinylated proteins are elevated in hemodialysis patients, a high cardiovascular risk population, and that homocysteinylated albumin shows altered properties. OBJECTIVE:Aim of this work was to investigate the effects of homocysteinylated albumin - the circulating form of this amino acid, utilized at the concentration present in uremia - on monocyte adhesion to a human endothelial cell culture monolayer and the relevant molecular changes induced at both cell levels. METHODS AND RESULTS:Treated endothelial cells showed a significant increase in monocyte adhesion. Endothelial cells showed after treatment a significant, specific and time-dependent increase in ICAM1 and VCAM1. Expression profiling and real time PCR, as well as protein analysis, showed an increase in the expression of genes encoding for chemokines/cytokines regulating the adhesion process and mediators of vascular remodeling (ADAM17, MCP1, and Hsp60). The mature form of ADAM17 was also increased as well as Tnf-α released in the cell medium. At monocyte level, treatment induced up-regulation of ICAM1, MCP1 and its receptor CCR2. CONCLUSIONS:Treatment with homocysteinylated albumin specifically increases monocyte adhesion to endothelial cells through up-regulation of effectors involved in vascular remodeling

DNA Methylation Dysfunction in Chronic Kidney Disease

Renal disease is the common denominator of a number of underlying disease conditions, whose prevalence has been dramatically increasing over the last two decades. Two aspects are particularly relevant to the subject of this review: (I) most cases are gathered under the umbrella of chronic kidney disease since they require&mdash;predictably for several lustrums&mdash;continuous clinical monitoring and treatment to slow down disease progression and prevent complications; (II) cardiovascular disease is a terrible burden in this population of patients, in that it claims many lives yearly, while only a scant minority reach the renal disease end stage. Why indeed a review on DNA methylation and renal disease? As we hope to convince you, the present evidence supports the role of the existence of various derangements of the epigenetic control of gene expression in renal disease, which hold the potential to improve our ability, in the future, to more effectively act toward disease progression, predict outcomes and offer novel therapeutic approaches