In vivo single-cell detection of metabolic oscillations in stem cells.

Through the use of bulk measurements in metabolic organs, the circadian clock was shown to play roles in organismal energy homeostasis. However, the relationship between metabolic and circadian oscillations has not been studied in vivo at a single-cell level. Also, it is unknown whether the circadian clock controls metabolism in stem cells. We used a sensitive, noninvasive method to detect metabolic oscillations and circadian phase within epidermal stem cells in live mice at the single-cell level. We observe a higher NADH/NAD+ ratio, reflecting an increased glycolysis/oxidative phosphorylation ratio during the night compared to the day. Furthermore, we demonstrate that single-cell metabolic heterogeneity within the basal cell layer correlates with the circadian clock and that diurnal fluctuations in NADH/NAD+ ratio are Bmal1 dependent. Our data show that, in proliferating stem cells, the circadian clock coordinates activities of oxidative phosphorylation and glycolysis with DNA synthesis, perhaps as a protective mechanism against genotoxicity.

[1]  K. Obrietan,et al.  Segregation of expression of mPeriod gene homologs in neurons and glia: possible divergent roles of mPeriod1 and mPeriod2 in the brain. , 2009, Human molecular genetics.

[2]  N. Ramanujam,et al.  Metabolic mapping of MCF10A human breast cells via multiphoton fluorescence lifetime imaging of the coenzyme NADH. , 2005, Cancer research.

[3]  Joseph S. Takahashi,et al.  Circadian Integration of Metabolism and Energetics , 2010, Science.

[4]  W R Brown,et al.  A review and mathematical analysis of circadian rhythms in cell proliferation in mouse, rat, and human epidermis. , 1991, The Journal of investigative dermatology.

[5]  Allon M. Klein,et al.  Interfollicular Epidermal Stem Cells Self-Renew via Autocrine Wnt Signaling , 2013, Science.

[6]  A. Kudlicki,et al.  Logic of the Yeast Metabolic Cycle: Temporal Compartmentalization of Cellular Processes , 2005, Science.

[7]  J. Denu,et al.  Circadian Clock NAD+ Cycle Drives Mitochondrial Oxidative Metabolism in Mice , 2013, Science.

[8]  B. Schoener,et al.  Intracellular Oxidation-Reduction States in Vivo , 1962, Science.

[9]  A. Heikal,et al.  Two-photon autofluorescence dynamics imaging reveals sensitivity of intracellular NADH concentration and conformation to cell physiology at the single-cell level. , 2009, Journal of photochemistry and photobiology. B, Biology.

[10]  Enrico Gratton,et al.  Phasor Fluorescence Lifetime Microscopy of Free and Protein-Bound NADH Reveals Neural Stem Cell Differentiation Potential , 2012, PloS one.

[11]  Padhraic Smyth,et al.  Brain and muscle Arnt-like protein-1 (BMAL1) controls circadian cell proliferation and susceptibility to UVB-induced DNA damage in the epidermis , 2012, Proceedings of the National Academy of Sciences.

[12]  S Banerjee,et al.  Histochemical studies on the distribution of certain dehydrogenases in squamous cell carcinoma of cheek. , 1989, Indian journal of cancer.

[13]  Sylvain Brohée,et al.  Distinct contribution of stem and progenitor cells to epidermal maintenance , 2012, Nature.

[14]  R. Hamanaka,et al.  Warburg Effect and Redox Balance , 2011, Science.

[15]  R. Jordan,et al.  Rhythms in Human Gastrointestinal Mucosa and Skin , 2002, Chronobiology international.

[16]  O. Warburg.  [Origin of cancer cells]. , 1956, Oncologia.

[17]  John B. Hogenesch,et al.  Mop3 Is an Essential Component of the Master Circadian Pacemaker in Mammals , 2000, Cell.

[18]  E. Gratton,et al.  Wnt signaling directs a metabolic program of glycolysis and angiogenesis in colon cancer , 2014, The EMBO journal.

[19]  A. Heikal.  Intracellular coenzymes as natural biomarkers for metabolic activities and mitochondrial anomalies. , 2010, Biomarkers in medicine.

[20]  P. Sassone-Corsi,et al.  Metabolism and the circadian clock converge. , 2013, Physiological reviews.

[21]  Iris Riemann,et al.  High-resolution multiphoton tomography of human skin with subcellular spatial resolution and picosecond time resolution. , 2003, Journal of biomedical optics.

[22]  C. Potten,et al.  CIRCADIAN RHYTHMS OF PRESUMPTIVE STEM CELLS IN THREE DIFFERENT EPITHELIA OF THE MOUSE , 1977, Cell and tissue kinetics.

[23]  N. Ramanujam,et al.  In vivo multiphoton microscopy of NADH and FAD redox states, fluorescence lifetimes, and cellular morphology in precancerous epithelia , 2007, Proceedings of the National Academy of Sciences.

[24]  Eduard Batlle,et al.  The circadian molecular clock creates epidermal stem cell heterogeneity , 2011, Nature.

[25]  W. Webb,et al.  Neural Activity Triggers Neuronal Oxidative Metabolism Followed by Astrocytic Glycolysis , 2004, Science.

[26]  R. Smart,et al.  Control of skin cancer by the circadian rhythm , 2011, Proceedings of the National Academy of Sciences.

[27]  Michael P. Murphy,et al.  How mitochondria produce reactive oxygen species , 2008, The Biochemical journal.

[28]  Enrico Gratton,et al.  Metabolic trajectory of cellular differentiation in small intestine by Phasor Fluorescence Lifetime Microscopy of NADH , 2012, Scientific Reports.

[29]  Paolo Sassone-Corsi,et al.  Metabolism and cancer: the circadian clock connection , 2009, Nature Reviews Cancer.

[30]  C. Chuong,et al.  Local circadian clock gates cell cycle progression of transient amplifying cells during regenerative hair cycling , 2013, Proceedings of the National Academy of Sciences.

[31]  Ben Lehner,et al.  Human epidermal stem cell function is regulated by circadian oscillations. , 2013, Cell stem cell.

[32]  Enrico Gratton,et al.  A novel fluorescence lifetime imaging system that optimizes photon efficiency , 2008, Microscopy research and technique.

[33]  E. Gratton,et al.  The phasor approach to fluorescence lifetime imaging analysis. , 2008, Biophysical journal.

[34]  J. Lakowicz,et al.  Fluorescence lifetime imaging of free and protein-bound NADH. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[35]  M. Toledano,et al.  ROS as signalling molecules: mechanisms that generate specificity in ROS homeostasis , 2007, Nature Reviews Molecular Cell Biology.

[36]  E. Gratton,et al.  Phasor approach to fluorescence lifetime microscopy distinguishes different metabolic states of germ cells in a live tissue , 2011, Proceedings of the National Academy of Sciences.

[37]  Benjamin D. Simons,et al.  A single type of progenitor cell maintains normal epidermis , 2007, Nature.

[38]  M. Antoch,et al.  Early aging and age-related pathologies in mice deficient in BMAL1, the core componentof the circadian clock. , 2006, Genes & development.