Efficient Assay and Marker Significance of NAD+ in Human Blood

Natalia V Balashova; Lev G Zavileyskiy; Artem V Artiukhov; Leonid A Shaposhnikov; Olga P Sidorova; Vladimir I Tishkov; Angela Tramonti; Anastasia A Pometun; Victoria I Bunik

doi:10.3389/fmed.2022.886485

Efficient Assay and Marker Significance of NAD⁺ in Human Blood

Front Med (Lausanne). 2022 May 19:9:886485. doi: 10.3389/fmed.2022.886485. eCollection 2022.

Authors

Natalia V Balashova^{1

2}, Lev G Zavileyskiy³, Artem V Artiukhov^{4

5}, Leonid A Shaposhnikov⁶, Olga P Sidorova⁷, Vladimir I Tishkov^{6

8}, Angela Tramonti⁹, Anastasia A Pometun^{6

8}, Victoria I Bunik^{3

4

5}

Affiliations

¹ Department of Clinical Laboratory Diagnostics, Faculty of Advanced Medicine, M.F. Vladimirsky Moscow Regional Research and Clinical Institute (MONIKI), Moscow, Russia.
² Department of Dietetics and Clinical Nutritionology, Faculty of Continuing Medical Education, RUDN Medical Institute, Moscow, Russia.
³ Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, Moscow, Russia.
⁴ Department of Biokinetics, A. N. Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow, Russia.
⁵ Department of Biochemistry, Sechenov University, Moscow, Russia.
⁶ Department of Chemical Enzymology, Faculty of Chemistry, Lomonosov Moscow State University, Moscow, Russia.
⁷ Department of Neurology, Faculty of Advanced Medicine, M.F. Vladimirsky Moscow Regional Research and Clinical Institute (MONIKI), Moscow, Russia.
⁸ Bach Institute of Biochemistry, Federal Research Centre "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Moscow, Russia.
⁹ Institute of Molecular Biology and Pathology, Italian National Research Council, Department of Biochemical Sciences "A. Rossi Fanelli," Sapienza University of Rome, Rome, Italy.

Abstract

Oxidized nicotinamide adenine dinucleotide (NAD⁺) is a biological molecule of systemic importance. Essential role of NAD⁺ in cellular metabolism relies on the substrate action in various redox reactions and cellular signaling. This work introduces an efficient enzymatic assay of NAD⁺ content in human blood using recombinant formate dehydrogenase (FDH, EC 1.2.1.2), and demonstrates its diagnostic potential, comparing NAD⁺ content in the whole blood of control subjects and patients with cardiac or neurological pathologies. In the control group (n = 22, 25-70 years old), our quantification of the blood concentration of NAD⁺ (18 μM, minimum 15, max 23) corresponds well to NAD⁺ quantifications reported in literature. In patients with demyelinating neurological diseases (n = 10, 18-55 years old), the NAD⁺ levels significantly (p < 0.0001) decrease (to 14 μM, min 13, max 16), compared to the control group. In cardiac patients with the heart failure of stage II and III according to the New York Heart Association (NYHA) functional classification (n = 24, 42-83 years old), the blood levels of NAD⁺ (13 μM, min 9, max 18) are lower than those in the control subjects (p < 0.0001) or neurological patients (p = 0.1). A better discrimination of the cardiac and neurological patients is achieved when the ratios of NAD⁺ to the blood creatinine levels, mean corpuscular volume or potassium ions are compared. The proposed NAD⁺ assay provides an easy and robust tool for clinical analyses of an important metabolic indicator in the human blood.

Keywords: Charcot-Marie-Tooth disease; NAD+ in human blood; cardiac patient; formate dehydrogenase; metabolic markers; neurological patient.