12 research outputs found
Rosiglitazone Induces Mitochondrial Biogenesis in Differentiated Murine 3T3-L1 and C3H/10T1/2 Adipocytes
Growing evidence indicates that PPARγ agonists, including rosiglitazone (RSG), induce adipose mitochondrial biogenesis. By systematically analyzing mitochondrial gene expression in two common murine adipocyte models, the current study aimed to further establish the direct role of RSG and capture temporal changes in gene transcription. Microarray profiling revealed that in fully differentiated 3T3-L1 and C3H/10T1/2 adipocytes treated with RSG or DMSO vehicle for 1, 2, 4, 7, 24, and 48 hrs, RSG overwhelmingly increased mitochondrial gene transcripts time dependently. The timing of the increases was consistent with the cascade of organelle biogenesis, that is, initiated by induction of transcription factor(s), followed by increases in the biosynthesis machinery, and then by increases in functional components. The transcriptional increases were further validated by increased mitochondrial staining, citrate synthase activity, and O2 consumption, and were found to be associated with increased adiponectin secretion. The work provided further insight on the mechanism of PPARγ-induced mitochondrial biogenesis in differentiated adipocytes
Mitochondria-localized AMPK responds to local energetics and contributes to exercise and energetic stress-induced mitophagy
Mitochondria form a complex, interconnected reticulum that is maintained through coordination among biogenesis, dynamic fission, and fusion and mitophagy, which are initiated in response to various cues to maintain energetic homeostasis. These cellular events, which make up mitochondrial quality control, act with remarkable spatial precision, but what governs such spatial specificity is poorly understood. Herein, we demonstrate that specific isoforms of the cellular bioenergetic sensor, 5′ AMP-activated protein kinase (AMPKα1/α2/β2/γ1), are localized on the outer mitochondrial membrane, referred to as mitoAMPK, in various tissues in mice and humans. Activation of mitoAMPK varies across the reticulum in response to energetic stress, and inhibition of mitoAMPK activity attenuates exercise-induced mitophagy in skeletal muscle in vivo. Discovery of a mitochondrial pool of AMPK and its local importance for mitochondrial quality control underscores the complexity of sensing cellular energetics in vivo that has implications for targeting mitochondrial energetics for disease treatment
miR‐206 family is important for mitochondrial and muscle function, but not essential for myogenesis in vitro
miR-206, miR-1a-1, and miR-1a-2 are induced during differentiation of skeletal myoblasts and promote myogenesis in vitro. miR-206 is required for skeletal muscle regeneration in vivo. Although this miRNA family is hypothesized to play an essential role in differentiation, a triple knock-out (tKO) of the three genes has not been done to test this hypothesis. We report that tKO C2C12 myoblasts generated using CRISPR/Cas9 method differentiate despite the expected derepression of the miRNA targets. Surprisingly, their mitochondrial function is diminished. tKO mice demonstrate partial embryonic lethality, most likely due to the role of miR-1a in cardiac muscle differentiation. Two tKO mice survive and grow normally to adulthood with smaller myofiber diameter, diminished physical performance, and an increase in PAX7 positive satellite cells. Thus, unlike other miRNAs important in other differentiation pathways, the miR-206 family is not absolutely essential for myogenesis and is instead a modulator of optimal differentiation of skeletal myoblasts
Label-Free Quantification of Intracellular Mitochondrial Dynamics Using Dielectrophoresis
Mitochondrial dynamics
play an important role within several pathological
conditions, including cancer and neurological diseases. For the purpose
of identifying therapies that target aberrant regulation of the mitochondrial
dynamics machinery and characterizing the regulating signaling pathways,
there is a need for label-free means to detect the dynamic alterations
in mitochondrial morphology. We present the use of dielectrophoresis
for label-free quantification of intracellular mitochondrial modifications
that alter cytoplasmic conductivity, and these changes are benchmarked
against label-based image analysis of the mitochondrial network. This
is validated by quantifying the mitochondrial alterations that are
carried out by entirely independent means on two different cell lines:
human embryonic kidney cells and mouse embryonic fibroblasts. In both
cell lines, the inhibition of mitochondrial fission that leads to
a mitochondrial structure of higher connectivity is shown to substantially
enhance conductivity of the cell interior, as apparent from the significantly
higher positive dielectrophoresis levels in the 0.5–15 MHz
range. Using single-cell velocity tracking, we show ∼10-fold
higher positive dielectrophoresis levels at 0.5 MHz for cells with
a highly connected versus those with a highly fragmented mitochondrial
structure, suggesting the feasibility for frequency-selective dielectrophoretic
isolation of cells to aid the discovery process for development of
therapeutics targeting the mitochondrial machinery
Optimized chemical probes for REV-ERB alpha
REV-ERB alpha has emerged as an important target for regulation of circadian rhythm and its associated physiology. Herein, we report on the optimization of a series of REV-ERB alpha agonists based on G5K4112 (1) for potency, selectivity, and bioavailability.(1) Potent REV-ERB alpha agonists 4, 10, 16, and 23 are detailed for their ability to suppress BMAL and IL-6 expression from human cells while also demonstrating excellent selectivity over LAR alpha. Amine 4 demonstrated in vivo bioavailability after either iv or oral dosing
Optimized Chemical Probes for REV-ERBα
REV-ERBα has emerged as an
important target for regulation of circadian rhythm and its associated
physiology. Herein, we report on the optimization of a series of REV-ERBα
agonists based on GSK4112 (<b>1</b>) for potency, selectivity,
and bioavailability. Potent REV-ERBα
agonists <b>4</b>, <b>10</b>, <b>16</b>, and <b>23</b> are detailed for their ability to suppress BMAL and IL-6
expression from human cells while also demonstrating excellent selectivity
over LXRα. Amine <b>4</b> demonstrated in vivo bioavailability
after either iv or oral dosing