Cytotoxic Effect of Advanced Glycation end Products

Abstract

ABSTRACTAdvanced glycation end products (AGE) are heterogeneous group of stable adducts, which include reactive derivatives from non-enzymatic glucose-protein condensation reactions, as well as lipids and nucleic acids exposed to reducing sugars. AGE may cause tissue injury both directly through trapping and cross-linking of proteins and indirectly by binding to specific receptors for AGE (RAGE) on the surface of various cells. AGE molecules (free, peptide-bound and protein bound) are found in blood plasma in high concentrations of diabetic patients. Diabetes mellitus is the most common maternal disease which leads to miscarriages and abnormalities in offspring. The mechanism of diabetic-associated embryopathy is still unclear and is considered as multifactorial. The objective of this study is to examine the effects of AOE on cell viability. In vitro model systems of cell cultures of amniotic and embryonal origin (WISH and MRC-5 cell lines) and cultivation of placental tissue explants were used. They were treated in normoglycaemic conditions with AGE modified bovine serum albumin (AGE-BSA) in variable concentrations and periods of time. Cell viability was assessed by Neutral red staining procedure. Apoptosis was evaluated by DNA laddering test, DAPI staining, cytokeratin 18 neoepitope formation and active caspase 3 expression. Results from this study showed that AGE-BSA has direct toxic effect on cell viability in WISH and MRC-5 cell cultures in time and dose dependent manner and causes DNA laddering. In placental villi explants AGE-BSA causes chromatin condensation and formation of apoptotic bodies as well as induction of cytokeratin 18 and active caspase 3 protein expression. These AGE-mediated changes may lead to impairment of embryo implantation and placentation as well as congenital malformations in fetus. This study clearly demonstrates that AGE modified proteins may have direct impact on the mechanisms of diabetes-associated pregnancy complications.