J 2013

Nonlinear Regression Models for Determination of Nicotinamide Adenine Dinucleotide Content in Human Embryonic Stem Cells

SALYKIN, Anton, Petr KUZMIC, Olga KYRYLENKO, Jindra MUSILOVÁ, Zdeněk GLATZ et. al.

Basic information

Original name

Nonlinear Regression Models for Determination of Nicotinamide Adenine Dinucleotide Content in Human Embryonic Stem Cells

Authors

SALYKIN, Anton (643 Russian Federation, belonging to the institution), Petr KUZMIC (840 United States of America), Olga KYRYLENKO (246 Finland), Jindra MUSILOVÁ (203 Czech Republic, belonging to the institution), Zdeněk GLATZ (203 Czech Republic, belonging to the institution), Petr DVOŘÁK (203 Czech Republic, guarantor, belonging to the institution) and Sergiy KYRYLENKO (246 Finland, belonging to the institution)

Edition

Stem Cell Reviews and Reports, TOTOWA, HUMANA PRESS INC, 2013, 1550-8943

Other information

Language

English

Type of outcome

Článek v odborném periodiku

Field of Study

Genetics and molecular biology

Country of publisher

United States of America

Confidentiality degree

není předmětem státního či obchodního tajemství

Impact factor

Impact factor: 3.214

RIV identification code

RIV/00216224:14110/13:00066458

Organization unit

Faculty of Medicine

DOI

UT WoS

000327386500005

Keywords in English

Biochemical assay; Enzymatic cycling; hESC-derived fibroblasts; Human embryonic stem cells; NAD; Nonlinear regression

Tags

International impact, Reviewed
Změněno: 15/4/2014 15:46, Ing. Mgr. Věra Pospíšilíková

Abstract

V originále

Recent evidence suggests that energy metabolism contributes to molecular mechanisms controlling stem cell identity. For example, human embryonic stem cells (hESCs) receive their metabolic energy mostly via glycolysis rather than mitochondrial oxidative phosphorylation. This suggests a connection of metabolic homeostasis to stemness. Nicotinamide adenine dinucleotide (NAD) is an important cellular redox carrier and a cofactor for various metabolic pathways, including glycolysis. Therefore, accurate determination of NAD cellular levels and dynamics is of growing importance for understanding the physiology of stem cells. Conventional analytic methods for the determination of metabolite levels rely on linear calibration curves. However, in actual practice many two-enzyme cycling assays, such as the assay systems used in this work, display prominently nonlinear behavior. Here we present a diaphorase/lactate dehydrogenase NAD cycling assay optimized for hESCs, together with a mechanism-based, nonlinear regression models for the determination of NAD+, NADH, and total NAD. We also present experimental data on metabolic homeostasis of hESC under various physiological conditions. We show that NAD+/NADH ratio varies considerably with time in culture after routine change of medium, while the total NAD content undergoes relatively minor changes. In addition, we show that the NAD+/NADH ratio, as well as the total NAD levels, vary between stem cells and their differentiated counterparts. Importantly, the NAD+/NADH ratio was found to be substantially higher in hESC-derived fibroblasts versus hESCs. Overall, our nonlinear mathematical model is applicable to other enzymatic amplification systems.

Links

GAP206/11/0009, research and development project
Name: Kapilární electroforéza jako nástroj pro metabolomické studie (Acronym: CE metabolomics)
Investor: Czech Science Foundation
MSM0021622430, plan (intention)
Name: Funkční a molekulární charakteristiky nádorových a normálních kmenových buněk - identifikace cílů pro nová terapeutika a terapeutické strategie
Investor: Ministry of Education, Youth and Sports of the CR, Functional and molecular characteristics of cancer and normal stem cells - identification of targets for novel therapeutics and therapeutic strategies