Rapid quantification of mitochondrial fractal dimension in individual cells

Vargas, Isaac; Alhallak, Kinan; Kolenc, Olivia I.; Jenkins, Samir V.; Griffin, Robert J.; Dings, Ruud P.M.; Rajaram, Narasimhan; Quinn, Kyle P.

doi:10.1364/boe.9.005269

Cited by 10 publications

(7 citation statements)

References 29 publications

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“…NADH fluorescence intensity maps were utilized to assess the spatial arrangement of mitochondria using a custom MATLAB code. Specifically, a modified blanket method was utilized to obtain the mitochondrial fractal dimension, a metric defining the closeness of mitochondrial arrangement, on a pixel-by-pixel basis 20 . At least two to four fields of view were taken from five to six separate samples per time point, and per treatment group 18,19 .…”

Section: Two-photon Excited Fluorescence (Tpef) Imaging and Analysismentioning

confidence: 99%

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Label-free metabolic biomarkers for assessing valve interstitial cell calcific progression

Tandon

Kolenc

Cross

et al. 2020

Sci Rep

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Calcific aortic valve disease (CAVD) is the most common form of valve disease where the only available treatment strategy is surgical valve replacement. Technologies for the early detection of CAVD would benefit the development of prevention, mitigation and alternate therapeutic strategies. Two-photon excited fluorescence (TPEF) microscopy is a label-free, non-destructive imaging technique that has been shown to correlate with multiple markers for cellular differentiation and phenotypic changes in cancer and wound healing. Here we show how specific TPEF markers, namely, the optical redox ratio and mitochondrial fractal dimension, correlate with structural, functional and phenotypic changes occurring in the aortic valve interstitial cells (VICs) during osteogenic differentiation. The optical redox ratio, and fractal dimension of mitochondria were assessed and correlated with gene expression and nuclear morphology of VICs. The optical redox ratio decreased for VICs during early osteogenic differentiation and correlated with biological markers for CAVD progression. Fractal dimension correlated with structural and osteogenic markers as well as measures of nuclear morphology. Our study suggests that TPEF imaging markers, specifically the optical redox ratio and mitochondrial fractal dimension, can be potentially used as a tool for assessing early CAVD progression in vitro. Calcific aortic stenosis or calcific aortic valve disease (CAVD) is a progressive disease involving multiple signaling pathways, endothelial dysfunction, cytokine infiltration, collagen remodeling, as well as lipid and calcium deposition 1-4. The symptoms and markers for CAVD manifest via both degenerative (apoptotic) and active (osteogenic) mechanisms 5,6. Aortic valve endothelial and interstitial cells differentiate into an osteoblast-like phenotype, the extracellular matrix becomes thicker and stiffer and calcium mineralization occurs throughout the tissue 7,8. Aortic stenosis and sclerosis have an increased prevalence in the elderly and contribute to a 50% elevated risk of infarction and other potentially fatal cardiovascular pathologies 2,9,10. Currently, valve replacement is the preferred treatment method, as other strategies, such as the retardation of calcific progression, prevention and early diagnosis, are non-existent 11. Diagnostic techniques like echocardiography, cardiac MRI and cardiac CT are widely used, but are only sensitive during later stages of the disease once there is tissue mineralization, and hemodynamic and geometric impairment 12,13. Aortic valve interstitial cells (VICs) are the primary cells in the heart valves and are involved in tissue maintenance, repair and remodeling 14. VICs exist in a quiescent state under healthy conditions and are activated due to injury or disease 15 , potentially differentiating into an osteogenic-like phenotype to potentiate calcification 2. Current in vitro biochemical techniques to assess CAVD are typically destructive as they involve cell lysis or fixation and do not facilitate the longitu...

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Section: Two-photon Excited Fluorescence (Tpef) Imaging and Analysismentioning

confidence: 99%

“…to assess the mitochondrial organization by determining mitochondrial fractal dimension (FD) 18,20 . Previously, TPEF was used to monitor the progression of osteogenic differentiation in human mesenchymal stem cells (MSCs) 18,21,22 and ORR and mitochondrial FD was shown to correlate with the osteogenic differentiation of human MSCs 18 .…”

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confidence: 99%

Label-free metabolic biomarkers for assessing valve interstitial cell calcific progression

Tandon

Kolenc

Cross

et al. 2020

Sci Rep

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“…The same method was used to investigate mitochondrial structural dynamics in human skin in vivo (53). A recent study used an improved image processing method in the spatial domain to rapidly quantify the local fractal dimension (FD) within individual cells in response to radiation therapy (64). This analysis found a significant decrease in FD (or an increase in mitochondrial clustering) of radiation-resistant lung cancer cells between 12- and 24-h post-radiation compared with pre-radiation measurements.…”

Section: Optical Microscopymentioning

confidence: 99%

“…The images were obtained at time periods of 1- and 24-h post radiation. (B) Summary data demonstrates significant temporal changes in the mitochondrial organization of radiation resistant cell line at 24 h [Reproduced with permission from Vargas et al (64)]. * p < 0.0001.…”

Section: Optical Microscopymentioning

confidence: 99%

Optical Imaging Approaches to Investigating Radiation Resistance

Dadgar

Rajaram

2019

Front. Oncol.

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Radiation therapy is frequently the first line of treatment for over 50% of cancer patients. While great advances have been made in improving treatment response rates and reducing damage to normal tissue, radiation resistance remains a persistent clinical problem. While hypoxia or a lack of tumor oxygenation has long been considered a key factor in causing treatment failure, recent evidence points to metabolic reprogramming under well-oxygenated conditions as a potential route to promoting radiation resistance. In this review, we present recent studies from our lab and others that use high-resolution optical imaging as well as clinical translational optical spectroscopy to shine light on the biological basis of radiation resistance. Two-photon microscopy of endogenous cellular metabolism has identified key changes in both mitochondrial structure and function that are specific to radiation-resistant cells and help promote cell survival in response to radiation. Optical spectroscopic approaches, such as diffuse reflectance and Raman spectroscopy have demonstrated functional and molecular differences between radiation-resistant and sensitive tumors in response to radiation. These studies have uncovered key changes in metabolic pathways and present a viable route to clinical translation of optical technologies to determine radiation resistance at a very early stage in the clinic.

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“…The ratio of the fluorescence intensity of these factors [e.g., FAD/(FAD+NAD(P)H)], called optical redox ratio (ORR), can reveal insights into the interplay between glucose catabolism and oxidative phosphorylation (11,12,(19)(20)(21). NAD(P)H autofluorescence can also be used to assess the mitochondrial organization via the mitochondrial fractal dimension (FD) parameter (20,22,23).…”

Section: Introductionmentioning

confidence: 99%

Label-Free Multiphoton Microscopy for the Detection and Monitoring of Calcific Aortic Valve Disease

Tandon

Quinn

Balachandran

2021

Front. Cardiovasc. Med.

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Calcific aortic valve disease (CAVD) is the most common valvular heart disease. CAVD results in a considerable socio-economic burden, especially considering the aging population in Europe and North America. The only treatment standard is surgical valve replacement as early diagnostic, mitigation, and drug strategies remain underdeveloped. Novel diagnostic techniques and biomarkers for early detection and monitoring of CAVD progression are thus a pressing need. Additionally, non-destructive tools are required for longitudinal in vitro and in vivo assessment of CAVD initiation and progression that can be translated into clinical practice in the future. Multiphoton microscopy (MPM) facilitates label-free and non-destructive imaging to obtain quantitative, optical biomarkers that have been shown to correlate with key events during CAVD progression. MPM can also be used to obtain spatiotemporal readouts of metabolic changes that occur in the cells. While cellular metabolism has been extensively explored for various cardiovascular disorders like atherosclerosis, hypertension, and heart failure, and has shown potential in elucidating key pathophysiological processes in heart valve diseases, it has yet to gain traction in the study of CAVD. Furthermore, MPM also provides structural, functional, and metabolic readouts that have the potential to correlate with key pathophysiological events in CAVD progression. This review outlines the applicability of MPM and its derived quantitative metrics for the detection and monitoring of early CAVD progression. The review will further focus on the MPM-detectable metabolic biomarkers that correlate with key biological events during valve pathogenesis and their potential role in assessing CAVD pathophysiology.

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Rapid quantification of mitochondrial fractal dimension in individual cells

Cited by 10 publications

References 29 publications

Label-free metabolic biomarkers for assessing valve interstitial cell calcific progression

Label-free metabolic biomarkers for assessing valve interstitial cell calcific progression

Optical Imaging Approaches to Investigating Radiation Resistance

Label-Free Multiphoton Microscopy for the Detection and Monitoring of Calcific Aortic Valve Disease

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