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Abstract

Toxicity with advanced glycation end products (AGEs) is a major problem in uremic patients. Treatment with peritoneal dialysis (PD) exacerbates AGE formation as a result of bioincompatibility of the conventional peritoneal dialysis fluid (PDF). The presence of glucose degradation products (GDPs) in PDF is the main cause of its bioincompatibility. Carnosine is an endogenous dipeptide with a powerful antiglycation/antioxidant activity. In an attempt to improve PDF biocompatibility, we evaluated the effect of carnosine in human peritoneal mesothelial cells (HPMC) incubated with PDF or GDPs in vitro. HPMC were incubated for short or prolonged time with PDF in the presence or absence of carnosine. Similarly, HPMC were incubated in the same condition but with a combination of GDPs. Following the incubation, cells were tested for their viability, protein carbonyl content and reactive oxygen species (ROS) production. Results demonstrated a significant protective effect of carnosine to HPMC in both acute and chronic conditions with PDF or GDPs as judged by the enhancement of cell viability, preserved protein from modification and decreased ROS production. Carnosine enhanced HPMC viability against the toxic effect of GDPs probably through protection of cellular protein from modification and from ROS-mediated oxidative damage. The salutary effect of carnosine may render it a desirable candidate for improving PDF biocompatibility and reducing AGE complications in PD patients. Copyright 2007 S. Karger AG, Basel.

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A tetrazolium-based colorimetric MTT assay to quantitate human monocyte mediated cytotoxicity against leukemic cells from cell lines and patients with acute myeloid leukemia.

A.A. van de Loosdrecht, R.H.J. Beelen, G.J. Ossenkoppele … (1994)

The MTT-colorimetric monocyte mediated cytotoxicity assay, based upon the ability of living cells to reduce 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide (MTT) into formazan, was evaluated using leukemic cells from five representative human leukemic cell lines and from 28 patients with acute myeloid leukemia (AML). An excellent linearity between absorbance and leukemic cell number was observed up to 5 x 10(4) cells/well and 50 x 10(4) cells/well for all cell lines and patients samples tested, respectively, in a 96-wells microtiter culture system. A huge variability in the susceptibility of leukemic cells to purified and IFN-gamma-activated human monocytes could be observed at effector-to-target cell (E:T) ratios of 1. The mean signal-to-noise ratio of the MTT assay for monocyte-leukemic cell mixtures from patients was 2.69 +/- 0.39 at E:T 1. In conclusion, the MTT based monocyte mediated cytotoxicity assay should be useful for studying the susceptibility of a variety of leukemic cells from cell lines and from patients with AML to monocytes in a rapid, sensitive and semi-automated manner.

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Alterations in nonenzymatic biochemistry in uremia: origin and significance of "carbonyl stress" in long-term uremic complications.

Yosuke Kurokawa, Sarah Miyata, A J Baynes … (1999)

Advanced glycation end products (AGEs), formed during Maillard or browning reactions by nonenzymatic glycation and oxidation (glycoxidation) of proteins, have been implicated in the pathogenesis of several diseases, including diabetes and uremia. AGEs, such as pentosidine and carboxymethyllysine, are markedly elevated in both plasma proteins and skin collagen of uremic patients, irrespective of the presence of diabetes. The increased chemical modification of proteins is not limited to AGEs, because increased levels of advanced lipoxidation end products (ALEs), such as malondialdehydelysine, are also detected in plasma proteins in uremia. The accumulation of AGEs and ALEs in uremic plasma proteins is not correlated with increased blood glucose or triglycerides, nor is it determined by a decreased removal of chemically modified proteins by glomerular filtration. It more likely results from increased plasma concentrations of small, reactive carbonyl precursors of AGEs and ALEs, such as glyoxal, methylglyoxal, 3-deoxyglucosone, dehydroascorbate, and malondialdehyde. Thus, uremia may be described as a state of carbonyl overload or "carbonyl stress" resulting from either increased oxidation of carbohydrates and lipids (oxidative stress) or inadequate detoxification or inactivation of reactive carbonyl compounds derived from both carbohydrates and lipids by oxidative and nonoxidative chemistry. Carbonyl stress in uremia may contribute to the long-term complications associated with chronic renal failure and dialysis, such as dialysis-related amyloidosis and accelerated atherosclerosis. The increased levels of AGEs and ALEs in uremic blood and tissue proteins suggest a broad derangement in the nonenzymatic biochemistry of both carbohydrates and lipids.

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Isolation, culture and characterization of human peritoneal mesothelial cells.

Eleni Stylianou, Lucy Jenner, Malcolm Davies … (1990)

This study establishes a reproducible technique for the culture of human peritoneal mesothelial cells. Direct explants, as well as enzymatically degraded specimens, of human omentum have been used as the source of cells. Cells were grown on collagen and gelatin coated matrices and were maintained in supplemented Ham's F-12 medium containing 10% (vol/vol) Fetal calf serum. Morphologically and ultrastructurally, the cells formed a homogeneous population. They were polygonal when confluent and devoid of contaminating fibroblasts, endothelial cells and macrophages. Cultured mesothelial cells co-expressed cytokeratin and vimentin and synthesized laminin, fibronectin, mesosecrin, non-specific esterase and collagen Types I and III but not Type IV. Ultrastructural features included numerous surface microvilli, cytoplasmic vesicles and an abundant endoplasmic reticulum. The stimulation of mesothelial cells by the calcium ionophore A23187 demonstrated that the two major products of arachidonic acid metabolism were prostacyclin and prostaglandin E2. The peritoneal mesothelial cell may be pivotal in the initiation of the inflammatory response during peritonitis and its establishment in culture will provide the basis for an in vitro model of peritoneal inflammation.

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PubMed ID:: 17664891

DOI:: 10.1159/000106509

ScienceOpen disciplines: Chemistry

Keywords: Antioxidants,pharmacology,Carnosine,Cell Survival,drug effects,physiology,Cells, Cultured,Epithelium,metabolism,Glucose,Glycosylation End Products, Advanced,antagonists & inhibitors,Humans,Peritoneal Cavity,cytology

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Antiglycation and antioxidant effect of carnosine against glucose degradation products in peritoneal mesothelial cells.

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Karger: Nephrology

Most cited references 25

A tetrazolium-based colorimetric MTT assay to quantitate human monocyte mediated cytotoxicity against leukemic cells from cell lines and patients with acute myeloid leukemia.

Alterations in nonenzymatic biochemistry in uremia: origin and significance of "carbonyl stress" in long-term uremic complications.

Isolation, culture and characterization of human peritoneal mesothelial cells.

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