A simple ImageJ macro tool for analyzing mitochondrial network morphology in mammalian cell culture

Valente, Andrew; Maddalena, Lucas A.; Robb, Ellen L.; Moradi, Fereshteh; Stuart, Jeffrey A.

doi:10.1016/j.acthis.2017.03.001

Cited by 558 publications

(550 citation statements)

References 29 publications

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“…Acquired images were subsequently analyzed to allow the quantification of mitochondrial mass, morphological features and network integrity (Dagda et al, 2009; Valente et al, 2017; Figure 4A ). A decreased mass of polarized mitochondria was observed in fibroblasts from twin 2 and the mother compared to their respective age- and sex-matched NC.…”

Section: Resultsmentioning

confidence: 99%

“…More details are provided under Supplementary Information . For mitochondrial morphology quantification, images were further analyzed with two macro tools designed for ImageJ (Fiji) (Dagda et al, 2009; Valente et al, 2017) to allow the quantification of mitochondrial mass, morphological features and network integrity. Mitochondrial cellular distribution was evaluated using the Fiji surface plot feature, followed by plot profile analysis.…”

Section: Methodsmentioning

confidence: 99%

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Impact of FMR1 Premutation on Neurobehavior and Bioenergetics in Young Monozygotic Twins

et al. 2018

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Mitochondrial dysfunction (MD) has been identified in lymphocytes, fibroblasts and brain samples from adults carrying a 55–200 CGG expansion in the fragile X mental retardation 1 (FMR1) gene (premutation; PM); however, limited data are available on the bioenergetics of pediatric carriers. Here we discuss a case report of three PM carriers: two monozygotic twins (aged 8 years) harboring an FMR1 allele with 150–180 CGG repeats, with no cognitive or intellectual issues but diagnosed with depression, mood instability and ADHD, and their mother (asymptomatic carrier with 78 CGG repeats). Fibroblasts and lymphocytes from the twins presented a generalized OXPHOS deficit, altered mitochondrial network, accumulation of depolarized mitochondria, and increased mitochondrial ROS production, outcomes distinct and more severe than the mother’s ones, suggesting the involvement of modulatory effects mediated by CGG expansion, X-activation ratio, sex hormones and epigenetic factors (chronic inflammation, consequence of Lyme disease). The degree of the severity of MD appeared to segregate with the morbidity of the phenotype. The mitochondrial ROS-mediated HIF-1α stabilization was identified as a key player at contributing to the MD, pointing it as a novel target for future therapeutical intervention.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Impact of FMR1 Premutation on Neurobehavior and Bioenergetics in Young Monozygotic Twins

et al. 2018

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“…Images were collected by confocal microscopy (LSM710; Carl Zeiss, Jena, Germany) and analyzed with ImageJ 1.42 (NIH, Bethesda, MD, USA). Mitochondria networks were analyzed using a modified method, as described by Valente et al (32), using the FIJI distribution by ImageJ and an open-source macro tool, Mitochondrial Network Analysis (MiNA), designed by the authors. In brief, images were split into their redgreen-blue color channels, whereby only the mitochondria channel was analyzed.…”

Section: Confocal Microscopy and Image Analysismentioning

confidence: 99%

“…Mitochondrial networks are defined as any segment with at least 1 additional branch (blue arrows), whereas individual mitochondria are defined as unbranched (yellow arrows). E ) Images were analyzed using a publicly available ImageJ macro designed by Valente et al(32). A Student's t test (a = 0.05) was used to compare CBS knockdown siCBS (n = 16) with control siCTL (n = 20) CP20 cells.…”

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confidence: 99%

Cystathionine β‐synthase regulates mitochondrial morphogenesis in ovarian cancer

et al. 2018

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Deregulation of mitochondrial morphogenesis, a dynamic equilibrium between mitochondrial fusion and fission processes, is now evolving as a key metabolic event that fuels tumor growth and therapy resistance. However, fundamental knowledge underpinning how cancer cells reprogram mitochondrial morphogenesis remains incomplete. Here, we report that cystathionine β-synthase (CBS) reprograms mitochondrial morphogenesis in ovarian cancer (OvCa) cells by selectively regulating the stability of mitofusin 2 (MFN2). Clinically, high expression of both CBS and MFN2 implicates poor overall survival of OvCa patients, and a significant association between CBS and MFN2 expression exists in individual patients in the same data set. The silencing of CBS by small interfering RNA or inhibition of its catalytic activity by a small molecule inhibitor creates oxidative stress that activates JNK. Activated JNK phosphorylates MFN2 to recruit homologous to the E6-AP carboxyl terminus' domain-containing ubiquitin E3 ligase for its degradation via the ubiquitin-proteasome system. Supplementation with hydrogen sulfide or glutathione (the catalytic products of CBS enzymatic activity), anti-oxidants, or a JNK inhibitor restores MFN2 expression. In CBS-silenced orthotopic xenograft tumor tissues, MFN2 but not MFN1 is selectively downregulated. In summary, this report reveals a role for deregulated mitochondrial morphogenesis in OvCa, suggests one of the mechanisms for this deregulation, and provides a way to correct it through modulation of the metabolic enzyme CBS.-Chakraborty, P. K., Murphy, B., Mustafi, S. B., Dey, A., Xiong, X., Rao, G., Naz, S., Zhang, M., Yang, D., Dhanasekaran, D. N., Bhattacharya, R., Mukherjee, P. Cystathionine β-synthase regulates mitochondrial morphogenesis in ovarian cancer.

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“…MEFs ablated for either β cyto -actin, γ cyto -actin, or both actins displayed individual mitochondria which were significantly larger by surface area compared to mitochondrial size in control MEFs (Figure 5 A–G). Mitochondria in Actg1, Actb, and Actb/Actg1 KO MEFs compared to control (Actg1-CT or Actb-CT) MEFs have similar numbers of individual mitochondria, or mitochondria without branches (Figure 5 H) as measured using the Fiji plug-in MiDA [43]. However, MEFs lacking either β cyto -actin or γ cyto -actin have an increase in network or branched mitochondria (Figure 5 I).…”

Section: Resultsmentioning

confidence: 99%

Impaired muscle relaxation and mitochondrial fission associated with genetic ablation of cytoplasmic actin isoforms

et al. 2018

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While α-actin isoforms predominate in adult striated muscle, skeletal muscle-specific knockouts (KOs) of nonmuscle cytoplasmic β - or γ -actin each cause a mild, but progressive myopathy effected by an unknown mechanism. Using transmission electron microscopy, we identified morphological abnormalities in both the mitochondria and the sarcoplasmic reticulum (SR) in aged muscle-specific β - and γ -actin KO mice. We found β - and γ -actin proteins to be enriched in isolated mitochondrial-associated membrane preparations, which represent the interface between mitochondria and sarco-endoplasmic reticulum important in signaling and mitochondrial dynamics. We also measured significantly elongated and interconnected mitochondrial morphologies associated with a significant decrease in mitochondrial fission events in primary mouse embryonic fibroblasts lacking β - and/or γ -actin. Interestingly, mitochondrial respiration in muscle was not measurably affected as oxygen consumption was similar in skeletal muscle fibers from 12 month-old muscle-specific β - and γ -actin KO mice. Instead, we found that the maximal rate of relaxation after isometric contraction was significantly slowed in muscles of 12-month-old β - and γ -actin muscle-specific KO mice. Our data suggest that impaired Ca re-uptake may presage development of the observed SR morphological changes in aged mice while providing a potential pathological mechanism for the observed myopathy.

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A simple ImageJ macro tool for analyzing mitochondrial network morphology in mammalian cell culture

Cited by 558 publications

References 29 publications

Impact of FMR1 Premutation on Neurobehavior and Bioenergetics in Young Monozygotic Twins

Impact of FMR1 Premutation on Neurobehavior and Bioenergetics in Young Monozygotic Twins

Cystathionine β‐synthase regulates mitochondrial morphogenesis in ovarian cancer

Impaired muscle relaxation and mitochondrial fission associated with genetic ablation of cytoplasmic actin isoforms

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