Mechanistic and Kinetic Insights into Cellular Uptake of Biomimetic Dinitrosyl Iron Complexes and Intracellular Delivery of NO for Activation of Cytoprotective HO-1

Han Chiu; Anyelina Chau Fang; Yi-Hong Chen; Ru Xin Koi; Kai-Ching Yu; Li-Hung Hsieh; Yueh-Ming Shyu; Tarik Abdelkareem Mostafa Amer; Yi-Jen Hsueh; Yu-Ting Tsao; Yang-Jin Shen; Yun-Ming Wang; Hung-Chi Chen; Yu-Jen Lu; Chieh-Cheng Huang; Tsai-Te Lu

doi:10.1021/jacsau.4c00064

Mechanistic and Kinetic Insights into Cellular Uptake of Biomimetic Dinitrosyl Iron Complexes and Intracellular Delivery of NO for Activation of Cytoprotective HO-1

JACS Au. 2024 Mar 29;4(4):1550-1569. doi: 10.1021/jacsau.4c00064. eCollection 2024 Apr 22.

Authors

Han Chiu¹, Anyelina Chau Fang¹, Yi-Hong Chen¹, Ru Xin Koi¹, Kai-Ching Yu¹, Li-Hung Hsieh¹, Yueh-Ming Shyu¹, Tarik Abdelkareem Mostafa Amer², Yi-Jen Hsueh³, Yu-Ting Tsao³, Yang-Jin Shen^{4

5}, Yun-Ming Wang², Hung-Chi Chen^{3

4}, Yu-Jen Lu^{4

5}, Chieh-Cheng Huang¹, Tsai-Te Lu^{1

6

7}

Affiliations

¹ Institute of Biomedical Engineering, National Tsing Hua University, Hsinchu 30013 Taiwan.
² Department of Biological Science and Technology, Institute of Molecular Medicine and Bioengineering, College of Biological Science and Technology, National Yang Ming Chiao Tung University, Hsinchu 300, Taiwan.
³ Department of Ophthalmology and Center for Tissue Engineering, Chang Gung Memorial Hospital, Taoyuan 33305, Taiwan.
⁴ College of Medicine, Chang Gung University, Kwei-San, Taoyuan 33302, Taiwan.
⁵ Department of Neurosurgery, Chang Gung Memorial Hospital, Taoyuan 33305, Taiwan.
⁶ Department of Chemistry, National Tsing Hua University, Hsinchu 30013 Taiwan.
⁷ Department of Chemistry, Chung Yuan Christian University, Taoyuan 32023, Taiwan.

Abstract

Dinitrosyl iron unit (DNIU), [Fe(NO)₂], is a natural metallocofactor for biological storage, delivery, and metabolism of nitric oxide (NO). In the attempt to gain a biomimetic insight into the natural DNIU under biological system, in this study, synthetic dinitrosyl iron complexes (DNICs) [(NO)₂Fe(μ-SCH₂CH₂COOH)₂Fe(NO)₂] (DNIC-COOH) and [(NO)₂Fe(μ-SCH₂CH₂COOCH₃)₂Fe(NO)₂] (DNIC-COOMe) were employed to investigate the structure-reactivity relationship of mechanism and kinetics for cellular uptake of DNICs, intracellular delivery of NO, and activation of cytoprotective heme oxygenase (HO)-1. After rapid cellular uptake of dinuclear DNIC-COOMe through a thiol-mediated pathway (t_max = 0.5 h), intracellular assembly of mononuclear DNIC [(NO)₂Fe(SR)(S_Cys)]^n-/[(NO)₂Fe(SR)(S_Cys-protein)]^n- occurred, followed by O₂-induced release of free NO (t_max = 1-2 h) or direct transfer of NO to soluble guanylate cyclase, which triggered the downstream HO-1. In contrast, steady kinetics for cellular uptake of DNIC-COOH via endocytosis (t_max = 2-8 h) and for intracellular release of NO (t_max = 4-6 h) reflected on the elevated activation of cytoprotective HO-1 (∼50-150-fold change at t = 3-10 h) and on the improved survival of DNIC-COOH-primed mesenchymal stem cell (MSC)/human corneal endothelial cell (HCEC) under stressed conditions. Consequently, this study unravels the bridging thiolate ligands in dinuclear DNIC-COOH/DNIC-COOMe as a switch to control the mechanism, kinetics, and efficacy for cellular uptake of DNICs, intracellular delivery of NO, and activation of cytoprotective HO-1, which poses an implication on enhanced survival of postengrafted MSC for advancing the MSC-based regenerative medicine.