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Mitoapocynin, a mitochondria targeted derivative of apocynin induces mitochondrial ROS generation and apoptosis in multiple cell types including cardiac myoblasts: a potential constraint to its therapeutic use

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Abstract

Mitoapocynin is a triphenylphosphonium conjugated derivative of apocynin that specifically locates to the mitochondria. It has been developed as a mitochondrially targeted therapeutic antioxidant. We attempted to attenuate the mitochondrial ROS induced in H9c2 cardiac myoblast cells treated with norepinephrine. Mitoapocynin was a poor quencher of total ROS as detected by the fluoroprobe DCFH-DA. Using mitochondrial superoxide specific probe MitoSoxRed, we found that 5–10 µM mitoapocynin itself induces superoxide over and above that is generated by the norepinephrine treatment. A supposedly control molecule to mitoapocynin, the synthetic compound PhC11TPP, having the triphenylphosphonium group and a benzene moiety with C11 aliphatic chain spacer was also found to be a robust inducer of mitochondrial ROS. Subsequent assays with several cell lines viz., NIH3T3, HEK293, Neuro2A, MCF-7 and H9c2, showed that prolonged exposure to mitoapocynin induces cell death by apoptosis that can be partially prevented by the general antioxidant N-acetyl cysteine. Analyses of mitochondrial electron transport complexes by Blue Native Polyacrylamide gel electrophoresis showed that both mitoapocynin and PhC11TPP disrupt the mitochondrial Complex I and V, and in addition, PhC11TPP also damages the Complex IV. Our data thus highlights the limitations of the therapeutic use of mitoapocynin as an antioxidant.

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Abbreviations

ROS:

Reactive oxygen species

RET:

Reverse electron transport

SIRT1:

Sirtuin 1

DCFH-DA:

Dichloro-dihydro-fluoresceindiacetate

NAC:

N-acetyl cysteine

Nox:

NADPH oxidase

YFP:

Yellow fluorescent protein

PI:

Propidium Iodide

Nrf2:

The nuclear factor erythroid 2-related factor 2

TPP:

Triphenylphosphonium

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Acknowledgements

Authors thankfully acknowledge funding support from the Science & Engineering Research Board (SERB), Government of India, under number EMR/2016/001832 to SKG. AM is a recipient of DS-Kothari Fellowship from the University Grants Commission, India. PB is a recipient of Senior Research Fellowship from the Council of Scientific and Industrial Research, India. MY acknowledges postdoctoral fellowship support from SERB-DST (DST File No: PDF/2017/000439), New Delhi, India. Sumona Ghosh, Technical Assistant performed some of the experiments shown in Figs. 3 and 4. We thank Director, CSIR-IICT (Ms. No. IICT/Pubs./2020/196) for providing necessary facilities to synthesize mitoapocynin.

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Author notes

  1. Md Yousuf

    Present address: Department of Chemistry, Ramnagar College, Purba Medinipur, Shikarbar, West Bengal, 721453, India

  2. Amena Mahmood and Padmini Bisoyi have contributed equally to this work.

Authors and Affiliations

  1. School of Life Sciences, Jawaharlal Nehru University, New Delhi, 110067, India

    Amena Mahmood, Padmini Bisoyi & Shyamal K. Goswami

  2. Applied Biology Division, CSIR-Indian Institute of Chemical Technology, Hyderabad, 500 007, India

    Rajkumar Banerjee & Md Yousuf

  3. DS-Kothari Fellow, UGC, New Delhi, India

    Amena Mahmood

  4. DST, SERB-National Postdoctoral Fellow, New Delhi, India

    Md Yousuf

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  1. Amena Mahmood
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Mahmood, A., Bisoyi, P., Banerjee, R. et al. Mitoapocynin, a mitochondria targeted derivative of apocynin induces mitochondrial ROS generation and apoptosis in multiple cell types including cardiac myoblasts: a potential constraint to its therapeutic use. Mol Cell Biochem 476, 2047–2059 (2021). https://doi.org/10.1007/s11010-020-04039-4

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