Product Citations: 185

Fig.1.B showing Western Blotting from the publication: Glutaminolysis is associated with mitochondrial pathway activation and can be therapeutically targeted in glioblastoma.
Fig.3.A showing Western Blotting from the publication: Glutaminolysis is associated with mitochondrial pathway activation and can be therapeutically targeted in glioblastoma.
Fig.7.B showing Western Blotting in a Homo sapiens (Human) sample from the publication: Dose-Dependent Effects of Radiation on Mitochondrial Morphology and Clonogenic Cell Survival in Human Microvascular Endothelial Cells.
Fig.5.I showing Western Blotting in a Mus musculus (House mouse) sample from the publication: Tissue Transglutaminase Knock-Out Preadipocytes and Beige Cells of Epididymal Fat Origin Possess Decreased Mitochondrial Functions Required for Thermogenesis.
Fig.6.C showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
Fig.1.B showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
Fig.2.F showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
Fig.2.E showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
Fig.3.L showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
Fig.3.D showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
Fig.4.C showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
Fig.4.A showing Western Blotting from the publication: Caveolin-3 deficiency associated with the dystrophy P104L mutation impairs skeletal muscle mitochondrial form and function.
Fig.7.C showing Western Blotting from the publication: Caveolin-3 deficiency associated with the dystrophy P104L mutation impairs skeletal muscle mitochondrial form and function.
Fig.8.A showing Western Blotting from the publication: Proinflammatory NFkB signalling promotes mitochondrial dysfunction in skeletal muscle in response to cellular fuel overloading.
Fig.2.A showing Western Blotting in a Homo sapiens (Human) sample from the publication: GSKIP-Mediated Anchoring Increases Phosphorylation of Tau by PKA but Not by GSK3beta via cAMP/PKA/GSKIP/GSK3/Tau Axis Signaling in Cerebrospinal Fluid and iPS Cells in Alzheimer Disease.
Fig.2.B showing Western Blotting in a Homo sapiens (Human) sample from the publication: GSKIP-Mediated Anchoring Increases Phosphorylation of Tau by PKA but Not by GSK3beta via cAMP/PKA/GSKIP/GSK3/Tau Axis Signaling in Cerebrospinal Fluid and iPS Cells in Alzheimer Disease.
Fig.4.C showing Western Blotting from the publication: Amyotrophic Lateral Sclerosis associated FUS mutation shortens mitochondria and induces neurotoxicity.
Fig.7.A showing Western Blotting from the publication: Amyotrophic Lateral Sclerosis associated FUS mutation shortens mitochondria and induces neurotoxicity.
Fig.5.E showing Western Blotting from the publication: Empagliflozin normalizes the size and number of mitochondria and prevents reduction in mitochondrial size after myocardial infarction in diabetic hearts.
Fig.5.C showing Western Blotting from the publication: Empagliflozin normalizes the size and number of mitochondria and prevents reduction in mitochondrial size after myocardial infarction in diabetic hearts.
Fig.2.A showing Western Blotting in a Mus musculus (House mouse) sample from the publication: Slower Dynamics and Aged Mitochondria in Sporadic Alzheimer's Disease.
Fig.13.C showing Immunocytochemistry-immunofluorescence from the publication: Deep transcriptome annotation enables the discovery and functional characterization of cryptic small proteins.
Fig.13.B showing Western Blotting from the publication: Deep transcriptome annotation enables the discovery and functional characterization of cryptic small proteins.
Fig.1.C showing Western Blotting from the publication: c-Abl-mediated Drp1 phosphorylation promotes oxidative stress-induced mitochondrial fragmentation and neuronal cell death.
Fig.1.D showing Western Blotting from the publication: c-Abl-mediated Drp1 phosphorylation promotes oxidative stress-induced mitochondrial fragmentation and neuronal cell death.
48 images found

Electrochemical Metabolic Profiling Reveals Mitochondrial Hyperactivation and Enhanced Neural Progenitor Proliferation in MLC1-Mutant Human Cortical Organoids

Preprint on BioRxiv : the Preprint Server for Biology on 20 April 2025 by Koo, K., Choi, J., et al.

SUMMARY Mitochondrial function is critical for neural progenitor regulation, yet its dysregulation during early human brain development remains poorly defined. Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a neurodevelopmental disorder caused by MLC1 mutations, previously attributed to postnatal astrocyte dysfunction. Using patient-derived human cortical organoids, we show that MLC1 is expressed in early neuroepithelial cells. To assess mitochondrial state in live organoids, we developed the MAGO (Matrigel-coated gold nanostructure) platform for real-time, label-free detection of redox activity. MLC1 mutant organoids showed mitochondrial hyperactivation, increased ATP and ROS, reduced membrane potential, and altered fusion protein expression. These changes were accompanied by enhanced BrdU incorporation and expansion of PAX6⁺/SOX2⁺ progenitors. To assess the causal role of MLC1 mutation, we generated isogenic organoids using CRISPR prime editing, which recapitulated redox hyperactivation and increased proliferation. Our findings redefine MLC as a disorder of early mitochondrial and progenitor dysregulation and establish a tractable platform to study metabolic mechanisms in neurodevelopmental disease.

  • Biochemistry and Molecular biology
  • Cell Biology
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Mfn2 regulates calcium homeostasis and suppresses PASMCs proliferation via interaction with IP3R3 to mitigate pulmonary arterial hypertension.

In Journal of Translational Medicine on 24 March 2025 by Wang, R., Wang, J., et al.

Pulmonary arterial hypertension (PAH) is a chronic disorder characterized by the excessive proliferation of pulmonary arterial smooth muscle cells (PASMCs). Recent studies indicate that Mitochondrial fusion protein 2 (Mfn2) maintains intracellular calcium (Ca2+) homeostasis via the mitochondria-associated endoplasmic reticulum membranes (MAMs) pathway, thereby inhibiting PASMCs proliferation and reducing pulmonary artery pressure. However, the precise mechanisms remain unclear.
This study explored the roles of Mfn2 and IP3R3 in PAH progression by assessing their expression in lung tissues of a monocrotaline (MCT)-induced PAH rat model. Immunoprecipitation assays were performed to confirm the interaction between Mfn2 and IP3R3. PASMCs were treated with either silenced or overexpressed Mfn2 and exposed to TNF-ɑ to observe effects on ER stress, IP3R3 expression, mitochondrial Ca2+ transport, and mitochondrial integrity. We also evaluated the effects of 4-phenylbutyric acid (4-PBA) and cistanche phenylethanol glycosides (CPGs) on the Mfn2-IP3R3 interaction in a TNF-α-induced PAH cell model, focusing on Ca2+ transport and mitochondrial structure.
Mfn2 expression was significantly down-regulated in the MCT-induced PAH rat model. Inhibition of ER stress upregulated Mfn2 expression, downregulated IP3R3 expression, increased mitochondrial Ca2+ concentration, and reduced autophagy, improving pulmonary hemodynamics and vascular remodeling. Overexpression of Mfn2 reduced ER stress, decreased IP3R3 expression, decreased mitochondrial Ca2+ transport, and restored mitochondrial integrity. Immunoprecipitation assays confirmed the interaction between Mfn2 and IP3R3. Inhibition of IP3R3 elevated Mfn2 levels, yielding similar beneficial effects as Mfn2 overexpression. 4-PBA and CPGs modulated the Mfn2-IP3R3 signaling axis, effectively inhibiting PAH progression.
Mfn2 mediates mitochondrial Ca2+ transport via IP3R3, suppressing PASMCs proliferation and pulmonary vascular remodeling, underscoring Mfn2's potential in regulating metabolic processes and vascular remodeling in PAH. These findings provide new insights for developing PAH-targeted therapeutics and establish a theoretical basis for traditional Chinese medicine in PAH prevention and treatment.
© 2025. The Author(s).

  • Cardiovascular biology
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Glutaminolysis is associated with mitochondrial pathway activation and can be therapeutically targeted in glioblastoma.

In Cancer Metabolism on 19 November 2024 by Miki, K., Yagi, M., et al.

Glioblastoma is an aggressive cancer that originates from abnormal cell growth in the brain and requires metabolic reprogramming to support tumor growth. Metabolic reprogramming involves the upregulation of various metabolic pathways. Although the activation of specific metabolic pathways in glioblastoma cell lines has been documented, the comprehensive profile of metabolic reprogramming and the role of each pathway in glioblastoma tissues in patients remain elusive.
We analyzed 38 glioblastoma tissues. As a test set, we examined 20 tissues from Kyushu University Hospital, focusing on proteins related to several metabolic pathways, including glycolysis, the one-carbon cycle, glutaminolysis, and the mitochondrial tricarboxylic acid cycle. Subsequently, we analyzed an additional 18 glioblastoma tissues from Kagoshima University Hospital as a validation set. We also validated our findings using six cell lines, including U87, LN229, U373, T98G, and two patient-derived cells.
The levels of mitochondria-related proteins (COX1, COX2, and DRP1) were correlated with each other and with glutaminolysis-related proteins (GLDH and GLS1). Conversely, their expression was inversely correlated with that of glycolytic proteins. Notably, inhibiting the glutaminolysis pathway in cell lines with high GLDH and GLS1 expression proved effective in suppressing tumor growth.
Our findings confirm that glioblastoma tissues can be categorized into glycolytic-dominant and mitochondrial-dominant types, as previously reported. The mitochondrial-dominant type is also glutaminolysis-dominant. Therefore, inhibiting the glutaminolysis pathway may be an effective treatment for mitochondrial-dominant glioblastoma.
© 2024. The Author(s).

  • WB
  • Cancer Research
  • Cell Biology
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Lineage-specific changes in mitochondrial properties during neural stem cell differentiation.

In Life Science Alliance on 1 July 2024 by Soares, R., Lourenço, D. M., et al.

Neural stem cells (NSCs) reside in discrete regions of the adult mammalian brain where they can differentiate into neurons, astrocytes, and oligodendrocytes. Several studies suggest that mitochondria have a major role in regulating NSC fate. Here, we evaluated mitochondrial properties throughout NSC differentiation and in lineage-specific cells. For this, we used the neurosphere assay model to isolate, expand, and differentiate mouse subventricular zone postnatal NSCs. We found that the levels of proteins involved in mitochondrial fusion (Mitofusin [Mfn] 1 and Mfn 2) increased, whereas proteins involved in fission (dynamin-related protein 1 [DRP1]) decreased along differentiation. Importantly, changes in mitochondrial dynamics correlated with distinct patterns of mitochondrial morphology in each lineage. Particularly, we found that the number of branched and unbranched mitochondria increased during astroglial and neuronal differentiation, whereas the area occupied by mitochondrial structures significantly reduced with oligodendrocyte maturation. In addition, comparing the three lineages, neurons revealed to be the most energetically flexible, whereas astrocytes presented the highest ATP content. Our work identified putative mitochondrial targets to enhance lineage-directed differentiation of mouse subventricular zone-derived NSCs.
© 2024 Soares et al.

  • Cell Biology
  • Stem Cells and Developmental Biology
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Regulation of TSC2 lysosome translocation and mitochondrial turnover by TSC2 acetylation status.

In Scientific Reports on 31 May 2024 by Marques, P., Burillo, J., et al.

Sirtuin1 (SIRT1) activity decreases the tuberous sclerosis complex 2 (TSC2) lysine acetylation status, inhibiting the mechanistic target of rapamycin complex 1 (mTORC1) signalling and concomitantly, activating autophagy. This study analyzes the role of TSC2 acetylation levels in its translocation to the lysosome and the mitochondrial turnover in both mouse embryonic fibroblast (MEF) and in mouse insulinoma cells (MIN6) as a model of pancreatic β cells. Resveratrol (RESV), an activator of SIRT1 activity, promotes TSC2 deacetylation and its translocation to the lysosome, inhibiting mTORC1 activity. An improvement in mitochondrial turnover was also observed in cells treated with RESV, associated with an increase in the fissioned mitochondria, positive autophagic and mitophagic fluxes and an enhancement of mitochondrial biogenesis. This study proves that TSC2 in its deacetylated form is essential for regulating mTORC1 signalling and the maintenance of the mitochondrial quality control, which is involved in the homeostasis of pancreatic beta cells and prevents from several metabolic disorders such as Type 2 Diabetes Mellitus.
© 2024. The Author(s).

  • Cell Biology
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Glutaminolysis is associated with mitochondrial pathway activation and can be therapeutically targeted in glioblastoma.

In Cancer Metab on 19 November 2024 by Miki, K., Yagi, M., et al.

Fig.1.B
Fig.1.B showing Western Blotting from the publication: Glutaminolysis is associated with mitochondrial pathway activation and can be therapeutically targeted in glioblastoma.
  • WB
Protein expression of the metabolic pathway of tumor tissue. a Schematic representation of the study showing the relationship between each enzyme and mitochondria. b Expression of proteins of the metabolic pathway in tumor tissue, including COX1, COX2, DRP1, HK2, SHMT2, GLS1, GLDH, and β-actin more
Protein expression of the metabolic pathway of tumor tissue. a Schematic representation of the study showing the relationship between each enzyme and mitochondria. b Expression of proteins of the metabolic pathway in tumor tissue, including COX1, COX2, DRP1, HK2, SHMT2, GLS1, GLDH, and β-actin less

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Glutaminolysis is associated with mitochondrial pathway activation and can be therapeutically targeted in glioblastoma.

In Cancer Metab on 19 November 2024 by Miki, K., Yagi, M., et al.

Fig.3.A
Fig.3.A showing Western Blotting from the publication: Glutaminolysis is associated with mitochondrial pathway activation and can be therapeutically targeted in glioblastoma.
  • WB
Protein expression of the metabolic pathway of tumor tissues (used as a validation set). Expression of mitochondria-related proteins is correlated with each other and with that of glutaminolysis proteins. a Proteins associated with metabolic pathways in tumor tissues include COX1, COX2, DRP1, HK2... more
Protein expression of the metabolic pathway of tumor tissues (used as a validation set). Expression of mitochondria-related proteins is correlated with each other and with that of glutaminolysis proteins. a Proteins associated with metabolic pathways in tumor tissues include COX1, COX2, DRP1, HK2, SHMT2, GLS1, GLDH, and actin. b Correlation coefficient between each protein. The correlation values (r) are calculated using the Spearman method using GraphPad Prism version 9. c COX1 expression correlates with COX2 expression (r = 0.88). d COX1 expression correlates with DRP1 expression (r = 0.89). e COX1 expression correlates with GLS1 (r = 0.72). f COX1 expression tends to correlate with GLDH (r = 0.71) less

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Dose-Dependent Effects of Radiation on Mitochondrial Morphology and Clonogenic Cell Survival in Human Microvascular Endothelial Cells.

In Cells on 23 December 2023 by Wang, L., Rivas, R., et al.

Fig.7.B
Fig.7.B showing Western Blotting in a Homo sapiens (Human) sample from the publication: Dose-Dependent Effects of Radiation on Mitochondrial Morphology and Clonogenic Cell Survival in Human Microvascular Endothelial Cells.
  • WB
  • Homo sapiens (Human)
The impact of doses on mitochondrial fusion and fission machinery in irradiated HMEC-1. The examination was conducted on day 4 post-IR. (A,C) 1 Gy and 0.5 Gy upregulated Mfn1 expression, as determined by Western blot analysis. However, no differences in Mfn2 expression were observed across all do... more
The impact of doses on mitochondrial fusion and fission machinery in irradiated HMEC-1. The examination was conducted on day 4 post-IR. (A,C) 1 Gy and 0.5 Gy upregulated Mfn1 expression, as determined by Western blot analysis. However, no differences in Mfn2 expression were observed across all doses tested, as shown in panels (A,D). (A,E) High doses decreased Opa1 expression. (B,F) There were no differences in Drp1 expression observed across all doses tested. There were three samples per radiation dose, and the statistical analysis conducted using one-way ANOVA and Dunnett’s multiple comparisons test revealed significant differences between irradiated cells and control cells, denoted by * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001. The mitochondrial loading control in Western blot analyses was the voltage-dependent anion channel (VDAC), whereas the cytosol loading control was glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The abbreviations used in the figures are as follows: Mfn1 represents mitofusin 1; Mfn2 represents mitofusin 2; Opa1 represents optic atrophy 1; and Drp1 represents dynamin-related protein 1. Data are expressed as mean + SEM. less

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Tissue Transglutaminase Knock-Out Preadipocytes and Beige Cells of Epididymal Fat Origin Possess Decreased Mitochondrial Functions Required for Thermogenesis.

In Int J Mol Sci on 5 May 2022 by Lénárt, K., Banko, C., et al.

Fig.5.I
Fig.5.I showing Western Blotting in a Mus musculus (House mouse) sample from the publication: Tissue Transglutaminase Knock-Out Preadipocytes and Beige Cells of Epididymal Fat Origin Possess Decreased Mitochondrial Functions Required for Thermogenesis.
  • WB
  • Mus musculus (House mouse)
High content screening of the preadipocytes and beige cells, and detection of mitochondrial fission- and fusion-related proteins: (A) Representative high content screening images showing mitochondrial morphology in TG2+/+ and TG2−/− preadipocytes, scale bars represent 50 μm; (B) fractions of frag... more
High content screening of the preadipocytes and beige cells, and detection of mitochondrial fission- and fusion-related proteins: (A) Representative high content screening images showing mitochondrial morphology in TG2+/+ and TG2−/− preadipocytes, scale bars represent 50 μm; (B) fractions of fragmented and tubular mitochondrial morphology in preadipocytes (%); (C) quantitative analysis of mitochondrial morphology in preadipocytes; (D) representative high content screening images showing mitochondrial morphology in TG2+/+ and TG2−/− beige cells; (E) fractions of fragmented and tubular mitochondrial morphology in beige cells (%); (F) quantitative analysis of mitochondrial morphology in beige cells, DAPI staining was used to determine the number of nuclei, the mitochondria were stained with Mito Tracker Deep Red either in the absence or presence of 10 μM antimycin A, Texas Red-X phalloidin was used to stain actin filaments for the detection of cell shapes (n = 3); (G–J) representative Western blot analyses and quantitative analyses of the mitochondrial fusion proteins (MFN2, OPA1) and mitochondrial fission proteins (DRP1, MFF) in preadipocytes and differentiated beige cells. β-ACTIN was used as a loading control. Columns represent the mean values ± SD. Statistical analyses were performed using Student’s t-test. n = 5. * p < 0.05. less

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Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.

In Cells on 10 March 2021 by Song, S. B., Park, J. S., et al.

Fig.6.C
Fig.6.C showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
  • WB
NAM-induced mitochondrial fragmentation is in part driven by calcineurin activation through an increase of Ca2+ that is dependent on Drp1. (A,B) Human fibroblasts cultivated with 5 mM NAM for 24 h with 2 μM FK506 for 1 h or 20 μM forskolin for 1 h prior to harvest were lysed and applied to immuno... more
NAM-induced mitochondrial fragmentation is in part driven by calcineurin activation through an increase of Ca2+ that is dependent on Drp1. (A,B) Human fibroblasts cultivated with 5 mM NAM for 24 h with 2 μM FK506 for 1 h or 20 μM forskolin for 1 h prior to harvest were lysed and applied to immunoblotting analysis with antibodies against phosphor-Drp1 or Drp1 (A). Mitochondria were immuno-stained with Tom20 antibody and visualized by confocal microscopy (B). Nuclei were counter-stained with DAPI (blue). Length of mitochondria in at least 20 cells was measured and averaged, and mean values normalized by that of control cells were plotted in the bar graph. ## indicates a significant difference (p < 0.01) between the measured means of the cells untreated and treated with forskolin. (C) Cells transfected with siNeg or siCalcineurin RNA were incubated with 5 mM NAM for the indicated period, and then applied to immunoblotting analysis with antibodies against calcineurin, phosphor-Drp1, and Drp1. In (A,C), protein bands were quantitated, and the relative values of pDrp1 over Drp1 were normalized by those of control cells. Mean values of three different experiments were plotted and presented in bar graphs below. (p < 0.05) between NAM’s effects in the presence and absence of forskolin (A) or calcineurin (C). (D) Cells transfected with siNeg RNA or siDrp1 RNA were cultured in the presence of 5 mM NAM for 48 h, 1 μM SRT1720 for 24 h, or 2μM ionomycin for 5 min. After treatment, cytosolic calcium was stained with Fluo-4 and visualized in situ with fluorescence microscopy. The fluorescence intensities of at least 40 cells in multiple confocal images were quantified by using ImageJ program, and average values were plotted in the bar graph at right after normalization by those of control cells. Black bars and grey bars represent the values of the cells treated with control siRNA and siDrp1 RNA, respectively (*, p < 0.05 and **, p < 0.01 vs. control). less

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Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.

In Cells on 10 March 2021 by Song, S. B., Park, J. S., et al.

Fig.1.B
Fig.1.B showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
  • WB
NAM induces mitochondrial fragmentation and Drp1 activation. (A) Human fibroblasts and MCF-7 cells were cultured in the presence of 5 mM NAM for 48 h. After fixing the cells, mitochondria were labeled with antibodies to OXPHOS (NHF) or Tom20 (MCF-7) and visualized in confocal microscopy. Length o... more
NAM induces mitochondrial fragmentation and Drp1 activation. (A) Human fibroblasts and MCF-7 cells were cultured in the presence of 5 mM NAM for 48 h. After fixing the cells, mitochondria were labeled with antibodies to OXPHOS (NHF) or Tom20 (MCF-7) and visualized in confocal microscopy. Length of mitochondria in 20 different cells was measured and averaged, and the mean value relative to that in untreated cells is presented as a bar in the graph at right (*, p < 0.05 vs. (-)). (B) Fibroblasts cultured in the presence of 5 mM NAM for indicated periods were applied to western blotting against phosphor-Drp1 (S637) or Drp1. Protein bands were quantitated by densitometry. Mean values of the measurement of the bands from three independent experiments are presented in a bar graph below (**, p < 0.01 vs. (-)). (C) Fibroblasts were incubated for 12 h on cover slips in the absence or presence of 5 mM NAM. The cells were then immuno-stained by Tom20 and Drp1 antibodies and applied to confocal imaging for mitochondria (red) and Drp1 protein (green). In (D), two enlarged images are presented to show Drp1 puncta (red) localized on a mitochondrial thread (green). (E) Cells transfected with siNeg RNA or siDrp1 RNA were cultured in the presence of 5 mM NAM for 48 h. After fixing, mitochondria were immune-stained with OXPHOS antibody and visualized. Length of mitochondria in at 10 different cells was measured and averaged, and the mean value relative to that in untreated cells is presented as a bar in the graph at bottom. (*, p < 0.05 vs. (-)). less

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Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.

In Cells on 10 March 2021 by Song, S. B., Park, J. S., et al.

Fig.2.F
Fig.2.F showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
  • WB
NAM-induced mitochondrial fragmentation and Drp1 hypo-phosphorylation are mediated through SIRT1 activation. (A,B) Fibroblasts were transfected with siNeg or siSIRT1 RNA. After 48 h, the cells were incubated with 5 mM NAM for 48 h (upper panels). Fibroblasts were treated with 1 μM SRT1720 for 24 ... more
NAM-induced mitochondrial fragmentation and Drp1 hypo-phosphorylation are mediated through SIRT1 activation. (A,B) Fibroblasts were transfected with siNeg or siSIRT1 RNA. After 48 h, the cells were incubated with 5 mM NAM for 48 h (upper panels). Fibroblasts were treated with 1 μM SRT1720 for 24 h or EX527 for 12 h (lower panels). After the treatments, mitochondria were immuno-stained with OXPHOS antibody and visualized in confocal microscopy (A), and mitochondrial length in at least 10 cells was quantified by ImageJ analysis and graphed (B). (C) Cells treated with 1 μM SRT1720 for indicated periods were lysed and applied to immunoblot analysis for phosphor-Drp1 or Drp1. (D) Cells were incubated on cover slips with 5 mM NAM for 48 h and treated with 1 μM EX527 for the last 12 h of the total period. Mitochondrial length in the confocal microphotographs was determined as in (A). (E) Cells were treated as in (D), and phosphor-Drp1 and Drp1 protein bands were detected by immunoblot analysis. (F) Cells transfected with either siNeg RNA or siSIRT1 RNA were incubated in the absence or presence of 5 mM NAM for 48 h. Then, cells were lysed and applied to immunoblot analysis with antibodies against phosphor-Drp1 or Drp1. For (C,E,F), protein bands were quantitated using a densitometer, and relative intensities (average of three biological repeats) were plotted in bar graphs (*, p < 0.05 and **, p < 0.01 vs. (-)). less

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Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.

In Cells on 10 March 2021 by Song, S. B., Park, J. S., et al.

Fig.2.E
Fig.2.E showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
  • WB
NAM-induced mitochondrial fragmentation and Drp1 hypo-phosphorylation are mediated through SIRT1 activation. (A,B) Fibroblasts were transfected with siNeg or siSIRT1 RNA. After 48 h, the cells were incubated with 5 mM NAM for 48 h (upper panels). Fibroblasts were treated with 1 μM SRT1720 for 24 ... more
NAM-induced mitochondrial fragmentation and Drp1 hypo-phosphorylation are mediated through SIRT1 activation. (A,B) Fibroblasts were transfected with siNeg or siSIRT1 RNA. After 48 h, the cells were incubated with 5 mM NAM for 48 h (upper panels). Fibroblasts were treated with 1 μM SRT1720 for 24 h or EX527 for 12 h (lower panels). After the treatments, mitochondria were immuno-stained with OXPHOS antibody and visualized in confocal microscopy (A), and mitochondrial length in at least 10 cells was quantified by ImageJ analysis and graphed (B). (C) Cells treated with 1 μM SRT1720 for indicated periods were lysed and applied to immunoblot analysis for phosphor-Drp1 or Drp1. (D) Cells were incubated on cover slips with 5 mM NAM for 48 h and treated with 1 μM EX527 for the last 12 h of the total period. Mitochondrial length in the confocal microphotographs was determined as in (A). (E) Cells were treated as in (D), and phosphor-Drp1 and Drp1 protein bands were detected by immunoblot analysis. (F) Cells transfected with either siNeg RNA or siSIRT1 RNA were incubated in the absence or presence of 5 mM NAM for 48 h. Then, cells were lysed and applied to immunoblot analysis with antibodies against phosphor-Drp1 or Drp1. For (C,E,F), protein bands were quantitated using a densitometer, and relative intensities (average of three biological repeats) were plotted in bar graphs (*, p < 0.05 and **, p < 0.01 vs. (-)). less

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Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.

In Cells on 10 March 2021 by Song, S. B., Park, J. S., et al.

Fig.3.L
Fig.3.L showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
  • WB
Direct activation of AMPK decreases phosphorylation of Drp1. (A) Fibroblasts on cover slips were either mock-treated or treated with 5 mM NAM for 48 h. During incubation, cells were further treated with 500 nM AICAR for the last 24 h or 1 μM compound C for the last 3 h. Cells were then fixed, and... more
Direct activation of AMPK decreases phosphorylation of Drp1. (A) Fibroblasts on cover slips were either mock-treated or treated with 5 mM NAM for 48 h. During incubation, cells were further treated with 500 nM AICAR for the last 24 h or 1 μM compound C for the last 3 h. Cells were then fixed, and mitochondria (red) were visualized by confocal microscopy using Tom20 antibody. Length of mitochondria in at least 20 cells from two different slides was measured and averaged, and plotted in the bar graph. Nuclei were counter-stained with DAPI (blue). (*, p < 0.05 and **, p < 0.01 vs. control. #, p < 0.05 between CC and CC + N). (B–F) Cells were cultivated with 5 mM NAM (for the indicated period), 1 µM SRT1720 (for 24 h), 1 µM EX527, 500 nM AICAR (for indicated time period), or 1 µM compound C (for 3 h). Protein extracts were analyzed by immunoblot analysis with antibodies against phosphor-AMPK, AMPK, phosphor-Drp1, Drp1, and β-actin. Protein bands were quantitated through densitometry, and the measured values from three-independent western blots were averaged. Relative values of phosphor-AMPK over AMPK (grey bars) and phosphor-Drp1 over Drp1 (black bars) were normalized by the values of control cells (-), and the relative values were plotted and presented in bar graphs. (*, p < 0.05 and **, p < 0.01 vs. control). less

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Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.

In Cells on 10 March 2021 by Song, S. B., Park, J. S., et al.

Fig.3.D
Fig.3.D showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
  • WB
Direct activation of AMPK decreases phosphorylation of Drp1. (A) Fibroblasts on cover slips were either mock-treated or treated with 5 mM NAM for 48 h. During incubation, cells were further treated with 500 nM AICAR for the last 24 h or 1 μM compound C for the last 3 h. Cells were then fixed, and... more
Direct activation of AMPK decreases phosphorylation of Drp1. (A) Fibroblasts on cover slips were either mock-treated or treated with 5 mM NAM for 48 h. During incubation, cells were further treated with 500 nM AICAR for the last 24 h or 1 μM compound C for the last 3 h. Cells were then fixed, and mitochondria (red) were visualized by confocal microscopy using Tom20 antibody. Length of mitochondria in at least 20 cells from two different slides was measured and averaged, and plotted in the bar graph. Nuclei were counter-stained with DAPI (blue). (*, p < 0.05 and **, p < 0.01 vs. control. #, p < 0.05 between CC and CC + N). (B–F) Cells were cultivated with 5 mM NAM (for the indicated period), 1 µM SRT1720 (for 24 h), 1 µM EX527, 500 nM AICAR (for indicated time period), or 1 µM compound C (for 3 h). Protein extracts were analyzed by immunoblot analysis with antibodies against phosphor-AMPK, AMPK, phosphor-Drp1, Drp1, and β-actin. Protein bands were quantitated through densitometry, and the measured values from three-independent western blots were averaged. Relative values of phosphor-AMPK over AMPK (grey bars) and phosphor-Drp1 over Drp1 (black bars) were normalized by the values of control cells (-), and the relative values were plotted and presented in bar graphs. (*, p < 0.05 and **, p < 0.01 vs. control). less

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Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.

In Cells on 10 March 2021 by Song, S. B., Park, J. S., et al.

Fig.4.C
Fig.4.C showing Western Blotting from the publication: Nicotinamide Treatment Facilitates Mitochondrial Fission through Drp1 Activation Mediated by SIRT1-Induced Changes in Cellular Levels of cAMP and Ca2.
  • WB
NAM-induced Drp1 hypo-phosphorylation is driven by a decrease of cAMP. (A) Human fibroblasts cultivated with 5 mM NAM for 24, 48, or 72 h were lysed and applied to measurement of cellular cAMP level. Amounts of cAMP per cell were normalized by that of untreated control cells, and the relative val... more
NAM-induced Drp1 hypo-phosphorylation is driven by a decrease of cAMP. (A) Human fibroblasts cultivated with 5 mM NAM for 24, 48, or 72 h were lysed and applied to measurement of cellular cAMP level. Amounts of cAMP per cell were normalized by that of untreated control cells, and the relative values were plotted. (B,C) Cells were incubated with 5 mM NAM for 48 h with addition of 100 µM 8-br-cAMP for the last 1 h, and then fixed and stained with antibodies against Tom20 (red) or Drp1 (green) and photographed by confocal microscopy (B), or lysed and applied to western blotting for phosphor-Drp1 (C). (D,E) Cells were either mock-treated (black bars) or treated with 5 mM NAM (grey bars) for 72 h with addition of 1 µM compound C (CC), 1 μM EX527, 500 nM AICAR (A), or 1 µM SRT1720 for the last 3, 24, 24, or 24 h, respectively. Cells were lysed and cellular cAMP levels were quantitated. Values from three independent measurements were averaged, normalized by the values of the control cells, and plotted. (F) Cells were incubated with 5 mM NAM for 48 h with addition of 500 nM IBMX for the last 1 h, lysed, and applied to immunoblot analysis with antibodies against phospho-Drp1 and Drp1. Mean densitometric quantity of protein bands in three independent western blots was normalized by that of untreated cells, and plotted (*, p < 0.05 and **, p < 0.01 vs. control). (G) Cells were incubated with 5 mM NAM for 48 h with addition of 500 nM IBMX for the last 1 h. Cells were fixed and mitochondria (red) were visualized with confocal microscopy using Tom20 antibody. Average mitochondria length was measured using at least 10 cells and values relative to those of untreated control cells were plotted in the graph (*, p < 0.05 vs. control). less

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Caveolin-3 deficiency associated with the dystrophy P104L mutation impairs skeletal muscle mitochondrial form and function.

In J Cachexia Sarcopenia Muscle on 1 June 2020 by Shah, D. S., Nisr, R. B., et al.

Fig.4.A
Fig.4.A showing Western Blotting from the publication: Caveolin-3 deficiency associated with the dystrophy P104L mutation impairs skeletal muscle mitochondrial form and function.
  • WB
Effects of Cav3 deficiency upon proteins linked to mitochondrial function and upon mitochondrial mass in mouse skeletal muscle. Crude muscle homogenates prepared from gastrocnemius muscle of three wild type (WT, +/+) and three Cav3 knockout (Cav3KO, −/−) mice were subject to sodium dodecyl sulfat... more
Effects of Cav3 deficiency upon proteins linked to mitochondrial function and upon mitochondrial mass in mouse skeletal muscle. Crude muscle homogenates prepared from gastrocnemius muscle of three wild type (WT, +/+) and three Cav3 knockout (Cav3KO, −/−) mice were subject to sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotted with antibodies to proteins shown (A) and protein abundance quantified relative to actin (used as a gel loading control) using Image J software (B). The data in (B) (mean ± SEM) is based on analysis of five WT and five Cav3KO mice. Alternatively, muscle tissue from mice was used for analysis of mitochondrial DNA or citrate synthase activity (C) or total muscle protein (D) as described in the methods section. Values in (C and D) are expressed as mean ± SEM from a minimum of three mice. Asterisks indicate a significant change (P < 0.05) to the appropriate WT data or between the indicated bars. Cav3, caveolin‐3; SDHA, succinate dehydrogenase subunit A; MnSOD, manganase superoxide dismutase; WT, wild type; VDAC, voltage‐dependent anion channel. less

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Caveolin-3 deficiency associated with the dystrophy P104L mutation impairs skeletal muscle mitochondrial form and function.

In J Cachexia Sarcopenia Muscle on 1 June 2020 by Shah, D. S., Nisr, R. B., et al.

Fig.7.C
Fig.7.C showing Western Blotting from the publication: Caveolin-3 deficiency associated with the dystrophy P104L mutation impairs skeletal muscle mitochondrial form and function.
  • WB
Effects of Cav3 re‐expression on mitochondrial protein expression, membrane potential, superoxide and mitochondrial respiration. Myoblasts in which Cav3 had been deleted by CRISPR/Cas9 (Cav3KO) were infected with retroviral particles containing an empty vector (Cav3KO + EV) or a vector containing... more
Effects of Cav3 re‐expression on mitochondrial protein expression, membrane potential, superoxide and mitochondrial respiration. Myoblasts in which Cav3 had been deleted by CRISPR/Cas9 (Cav3KO) were infected with retroviral particles containing an empty vector (Cav3KO + EV) or a vector containing HA‐GFP tagged WT‐Cav3 (Cav3KO + Cav3) and stable transformants isolated by antibiotic selection prior to: (A) immunoblotting whole cell lysates (WCL) with antibodies against Cav3 or actin (used as a gel loading control) antibodies; (B) isolation of cytosolic (Cyto) and mitochondrial membranes and immunoblotting using antibodies to proteins indicated; (C) lysis and immunoblotting of WCL with antibodies to proteins shown and quantification of their abundance relative to GAPDH (used as a gel loading control) using Image J software; (D) determination of superoxide content using fluorescence intensity (FI) of MitoSOX and mitochondrial membrane potential monitored by  spectral analysis of JC‐10 aggregate:monomer content from three separate  experiments (for this experiment GFP would interfere with the JC‐10 probe and therefore wild type Cav3 was cloned into pcDNA3.1 using HindIII and XhoI Restriction sites and an empty vector or a vector containing Cav3 were transiently transfected in L6 muscle cells and selected for using G418 before JC‐10 probe incubation); and  (E) analysis of mitochondrial respiration to assess basal oxygen consumption rate using Seahorse technology. Oligomycin (1 μM), FCCP (1 μM) and a mixture of Rotenone (1 μM)/Antimycin (2 μM) were added at the times indicated by dotted lines to determine basal, ATP‐linked and maximal respiration. The Seahorse trace shown in (E) is from a single experiment in which each point represents the mean ± SEM from triplicate analyses. All bar graph data in (E) represent the analysis of three individual experiments (values are mean ± SEM). Asterisks indicate significant change (P < 0.05) to the WT bar value or between indicated bars. Cav3, caveolin‐3; EV, empty vector; SDHA, succinate dehydrogenase subunit A; WT, wild type; VDAC, voltage‐dependent anion channel. less

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Proinflammatory NFkB signalling promotes mitochondrial dysfunction in skeletal muscle in response to cellular fuel overloading.

In Cell Mol Life Sci on 1 December 2019 by Nisr, R. B., Shah, D. S., et al.

Fig.8.A
Fig.8.A showing Western Blotting from the publication: Proinflammatory NFkB signalling promotes mitochondrial dysfunction in skeletal muscle in response to cellular fuel overloading.
  • WB
Effects of cellular fuel overloading on mitophagy and upon proteins linked to mitochondrial dynamics. Wild type L6 myotubes or those stably expressing a retroviral vector encoding a GFP–mCherry–Fis1 mitophagy reporter were incubated with GLC (5 mM), PA (0.4 mM) and BI605906 (10 μM) for 16 h as in... more
Effects of cellular fuel overloading on mitophagy and upon proteins linked to mitochondrial dynamics. Wild type L6 myotubes or those stably expressing a retroviral vector encoding a GFP–mCherry–Fis1 mitophagy reporter were incubated with GLC (5 mM), PA (0.4 mM) and BI605906 (10 μM) for 16 h as indicated prior to: a subcellular fractionation and isolation of a cytosolic and mitochondrial-enriched membrane fraction for immunoblotting with antibodies to proteins shown. b–e Fixing and confocal imaging to visualise and f quantifying (using the Volocity software) mitophagy in L6 myotubes. Data (mean ± SEM) in (f) are from five separate experiments. Asterisks indicate a significant change (P < 0.05) between the indicated bars. Boxed sections in panels b, c, d and e have been expanded to highlight mitophagic particles, some of which are depicted by the white arrow heads less

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GSKIP-Mediated Anchoring Increases Phosphorylation of Tau by PKA but Not by GSK3beta via cAMP/PKA/GSKIP/GSK3/Tau Axis Signaling in Cerebrospinal Fluid and iPS Cells in Alzheimer Disease.

In J Clin Med on 21 October 2019 by Ko, H. J., Chiou, S. J., et al.

Fig.2.A
Fig.2.A showing Western Blotting in a Homo sapiens (Human) sample from the publication: GSKIP-Mediated Anchoring Increases Phosphorylation of Tau by PKA but Not by GSK3beta via cAMP/PKA/GSKIP/GSK3/Tau Axis Signaling in Cerebrospinal Fluid and iPS Cells in Alzheimer Disease.
  • WB
  • Homo sapiens (Human)
GSKIP through GSK3β-mediated anchoring to modulate Tau phosphorylation by PKA kinase. (A) pEGFP-C1 or pEGFP-C1–GSKIP wt and mutants were transiently expressed in SH-SY5Y cells. The cells were untreated (left four lanes) or incubated with 25 μM forskolin (FSK) for 1 h (right four lanes). (B) Kinas... more
GSKIP through GSK3β-mediated anchoring to modulate Tau phosphorylation by PKA kinase. (A) pEGFP-C1 or pEGFP-C1–GSKIP wt and mutants were transiently expressed in SH-SY5Y cells. The cells were untreated (left four lanes) or incubated with 25 μM forskolin (FSK) for 1 h (right four lanes). (B) Kinase-inactive pET32a-HA-GSK3β-K85R or -K85M mutants were transiently expressed in SH-SY5Y cells. pET32a-HA-GSK3β-K85R mutant retained the capacity to bind GSKIP, but such capacity was not evident in pET32a-HA-GSK3β-K85M. Cells were incubated with 25 μM FSK for 2 h and were then analyzed through Western blotting. (C) Knockdown of GSK3β but not GSK3α expression altered phosphorylation of dynamin related protein 1 (Drp1) Ser637 and Tau Ser409. less

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GSKIP-Mediated Anchoring Increases Phosphorylation of Tau by PKA but Not by GSK3beta via cAMP/PKA/GSKIP/GSK3/Tau Axis Signaling in Cerebrospinal Fluid and iPS Cells in Alzheimer Disease.

In J Clin Med on 21 October 2019 by Ko, H. J., Chiou, S. J., et al.

Fig.2.B
Fig.2.B showing Western Blotting in a Homo sapiens (Human) sample from the publication: GSKIP-Mediated Anchoring Increases Phosphorylation of Tau by PKA but Not by GSK3beta via cAMP/PKA/GSKIP/GSK3/Tau Axis Signaling in Cerebrospinal Fluid and iPS Cells in Alzheimer Disease.
  • WB
  • Homo sapiens (Human)
GSKIP through GSK3β-mediated anchoring to modulate Tau phosphorylation by PKA kinase. (A) pEGFP-C1 or pEGFP-C1–GSKIP wt and mutants were transiently expressed in SH-SY5Y cells. The cells were untreated (left four lanes) or incubated with 25 μM forskolin (FSK) for 1 h (right four lanes). (B) Kinas... more
GSKIP through GSK3β-mediated anchoring to modulate Tau phosphorylation by PKA kinase. (A) pEGFP-C1 or pEGFP-C1–GSKIP wt and mutants were transiently expressed in SH-SY5Y cells. The cells were untreated (left four lanes) or incubated with 25 μM forskolin (FSK) for 1 h (right four lanes). (B) Kinase-inactive pET32a-HA-GSK3β-K85R or -K85M mutants were transiently expressed in SH-SY5Y cells. pET32a-HA-GSK3β-K85R mutant retained the capacity to bind GSKIP, but such capacity was not evident in pET32a-HA-GSK3β-K85M. Cells were incubated with 25 μM FSK for 2 h and were then analyzed through Western blotting. (C) Knockdown of GSK3β but not GSK3α expression altered phosphorylation of dynamin related protein 1 (Drp1) Ser637 and Tau Ser409. less

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Amyotrophic Lateral Sclerosis associated FUS mutation shortens mitochondria and induces neurotoxicity.

In Sci Rep on 22 October 2018 by Nakaya, T. & Maragkakis, M.

Fig.4.C
Fig.4.C showing Western Blotting from the publication: Amyotrophic Lateral Sclerosis associated FUS mutation shortens mitochondria and induces neurotoxicity.
  • WB
Differential gene translation analysis for WT- and R495X-expressing neurons. (A) Scatter plot of WT versus control and R495X versus control differential gene translation. The top 2% (280) differentially translated genes that are specific to R495X are shown in red. (B) Gene ontology enriched terms... more
Differential gene translation analysis for WT- and R495X-expressing neurons. (A) Scatter plot of WT versus control and R495X versus control differential gene translation. The top 2% (280) differentially translated genes that are specific to R495X are shown in red. (B) Gene ontology enriched terms for the top 2% (280) differentially translated genes specific to R495X compared to the remaining genes. (C) Immunoblots of differentially translated genes in R495X-expressing neurons (left). Bar plots for gene translational levels quantified from Ribo-Seq (top right) and protein band intensities quantified from immunoblots (bottom right). Protein standard is shown on the right. Band intensities were normalized by TubulinβIII and the average value for control was set to 1. P-values in one-way ANOVA are, 0.01, < 0.0001, 0.001, 0.725 and 0.257 for Kif5b, Dnm1l, Csde1, Mfn2 and Cox4, respectively. *p < 0.05, **p < 0.01 to control in post Tukey’s HSD test. Error bar indicates standard deviation (N = 3). (D) Number of genes that are bound by R495X for gene sets of decreasing differential translation in R495X (top) and R495X binding density in gene exons for the same gene sets (bottom). Error bars indicate standard error. P-value, one-way ANOVA test. less

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Amyotrophic Lateral Sclerosis associated FUS mutation shortens mitochondria and induces neurotoxicity.

In Sci Rep on 22 October 2018 by Nakaya, T. & Maragkakis, M.

Fig.7.A
Fig.7.A showing Western Blotting from the publication: Amyotrophic Lateral Sclerosis associated FUS mutation shortens mitochondria and induces neurotoxicity.
  • WB
R495X4FL partially rescues R495X induced mitochondria shortening and neurotoxicity. (A) Immunoblots for control, R495X- and R495X4FL-expressing neurons (left) and bar plot for normalized protein band intensities (right). Average band intensity for control was set to 1. Protein standard is shown o... more
R495X4FL partially rescues R495X induced mitochondria shortening and neurotoxicity. (A) Immunoblots for control, R495X- and R495X4FL-expressing neurons (left) and bar plot for normalized protein band intensities (right). Average band intensity for control was set to 1. Protein standard is shown on the right. Error bars indicate standard deviation (N = 3). P-values in one-way ANOVA are, 0.015, 0.009, 0.003, 0.731 and 0.093 for Kif5b, Dnm1l, Csde1, Mfn2 and Cox4, respectively and *p < 0.05 and **p < 0.01 in post Tukey’s HSD test. (B) Staining images using MitoTracker (top) and anti-FLAG antibody (middle) with magnified images of MitoTracker stained neuronal processes (bottom) of an R495X4FL-expressing neuron. Scale bar, 10 μm (middle) and 5 μm (bottom). (C) Average mitochondria major-axis length for control, R495X- and R495X4FL-expressing neurons. 880 mitochondria from 10 cells expressing R495X4FL were analyzed. Data for control and R495X are copied from Fig. 5C. Error bars indicate standard error. P-value in one-way ANOVA is < 0.0001 and *p < 0.05 and **p < 0.01 in post Tukey’s HSD test. (D) Cell survival curve for control, WT-, R495X- and R495X4FL-expressing neurons. The number of surviving cells at days 13–15 are expressed as the ratio over those surviving at day 12 (y-axis). The numbers in parenthesis indicate the day after induction. ***p < 0.003, log-rank test. less

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Empagliflozin normalizes the size and number of mitochondria and prevents reduction in mitochondrial size after myocardial infarction in diabetic hearts.

In Physiol Rep on 1 June 2018 by Mizuno, M., Kuno, A., et al.

Fig.5.E
Fig.5.E showing Western Blotting from the publication: Empagliflozin normalizes the size and number of mitochondria and prevents reduction in mitochondrial size after myocardial infarction in diabetic hearts.
  • WB
Effects of myocardial infarction on regulatory factors of mitochondrial fission. (A) Representative immunoblots for Fis1 in the noninfarct remote myocardium from LETO, OLETF treated with a vehicle, and OLETF treated with empagliflozin (Empa) that were subjected to a sham operation or coronary art... more
Effects of myocardial infarction on regulatory factors of mitochondrial fission. (A) Representative immunoblots for Fis1 in the noninfarct remote myocardium from LETO, OLETF treated with a vehicle, and OLETF treated with empagliflozin (Empa) that were subjected to a sham operation or coronary artery ligation (MI). (B) Summary data of Fis1 protein level normalized to vinculin. (C) Representative immunoblots for phospho‐Ser616‐Drp1 and total Drp1 in the noninfarcted myocardium from rats of six groups. (D) Summary data of levels of phospho‐Ser616‐Drp1 and total Drp1. (E) Representative immunoblots for phospho‐Ser637‐Drp1 and total Drp1. (F) Summary data of phospho‐Ser637‐Drp1 and total Drp1 normalized to vinculin level. N = 8 in each group. *P < 0.05. a.u., arbitrary unit. less

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Empagliflozin normalizes the size and number of mitochondria and prevents reduction in mitochondrial size after myocardial infarction in diabetic hearts.

In Physiol Rep on 1 June 2018 by Mizuno, M., Kuno, A., et al.

Fig.5.C
Fig.5.C showing Western Blotting from the publication: Empagliflozin normalizes the size and number of mitochondria and prevents reduction in mitochondrial size after myocardial infarction in diabetic hearts.
  • WB
Effects of myocardial infarction on regulatory factors of mitochondrial fission. (A) Representative immunoblots for Fis1 in the noninfarct remote myocardium from LETO, OLETF treated with a vehicle, and OLETF treated with empagliflozin (Empa) that were subjected to a sham operation or coronary art... more
Effects of myocardial infarction on regulatory factors of mitochondrial fission. (A) Representative immunoblots for Fis1 in the noninfarct remote myocardium from LETO, OLETF treated with a vehicle, and OLETF treated with empagliflozin (Empa) that were subjected to a sham operation or coronary artery ligation (MI). (B) Summary data of Fis1 protein level normalized to vinculin. (C) Representative immunoblots for phospho‐Ser616‐Drp1 and total Drp1 in the noninfarcted myocardium from rats of six groups. (D) Summary data of levels of phospho‐Ser616‐Drp1 and total Drp1. (E) Representative immunoblots for phospho‐Ser637‐Drp1 and total Drp1. (F) Summary data of phospho‐Ser637‐Drp1 and total Drp1 normalized to vinculin level. N = 8 in each group. *P < 0.05. a.u., arbitrary unit. less

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Slower Dynamics and Aged Mitochondria in Sporadic Alzheimer's Disease.

In Oxid Med Cell Longev on 5 December 2017 by Martín-Maestro, P., Gargini, R., et al.

Fig.2.A
Fig.2.A showing Western Blotting in a Mus musculus (House mouse) sample from the publication: Slower Dynamics and Aged Mitochondria in Sporadic Alzheimer's Disease.
  • WB
  • Mus musculus (House mouse)
Decreased fission and fusion protein levels in SAD cells. (a) Representative Western blot analysis and quantification of DLP1 in healthy and SAD fibroblasts under basal conditions. (b) Representative confocal microscopy images showing TOMM20 in green and DLP1 in red in basal conditions. (c) Quant... more
Decreased fission and fusion protein levels in SAD cells. (a) Representative Western blot analysis and quantification of DLP1 in healthy and SAD fibroblasts under basal conditions. (b) Representative confocal microscopy images showing TOMM20 in green and DLP1 in red in basal conditions. (c) Quantification of the amount of DLP1 in mitochondria per cell in the images represented in (b). (d) Quantification of mitochondrial surface measured by TOMM20 label per cell in the images represented in (b). (e)–(g) Representative Western blot analysis and quantification of MFN2/mitofusin 2 (f) and STOML2 (g) in healthy and SAD fibroblasts under basal conditions. Graphs show means and standard deviations of the healthy/AD age- and sex-matched couple samples: AG11020/AG05810 and AG11362/AG05809; AG07310/AG06869; AG05813/AG06263; AG07803/AG06262 (a)–(e) and AG11020/AG05810; AG11362/AG05809; and AG07310/AG06869 (g). Scale bar: 40 μm. ∗p < 0.05. less

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Deep transcriptome annotation enables the discovery and functional characterization of cryptic small proteins.

In Elife on 30 October 2017 by Samandi, S., Roy, A. V., et al.

Fig.13.C
Fig.13.C showing Immunocytochemistry-immunofluorescence from the publication: Deep transcriptome annotation enables the discovery and functional characterization of cryptic small proteins.
  • ICC-IF
AltMiD51-induced mitochondrial fragmentation is dependent on Drp1.(a) Bar graphs show mitochondrial morphologies in HeLa cells treated with non-target or Drp1 siRNAs. Cells were mock-transfected (pcDNA3.1) or transfected with altMiD51Flag. Means of three independent experiments per condition are ... more
AltMiD51-induced mitochondrial fragmentation is dependent on Drp1.(a) Bar graphs show mitochondrial morphologies in HeLa cells treated with non-target or Drp1 siRNAs. Cells were mock-transfected (pcDNA3.1) or transfected with altMiD51Flag. Means of three independent experiments per condition are shown (100 cells for each independent experiment). ***p<0.0005 (Fisher’s exact test) for the three morphologies between altMiD51 and the other experimental conditions. (b) HeLa cells treated with non-target or Drp1 siRNA were transfected with empty vector (pcDNA3.1) or altMiD51Flag, as indicated. Proteins were extracted and analyzed by western blot with antibodies against the Flag tag (altMiD51), Drp1 or actin, as indicated. (c) Confocal microscopy of Drp1 knockdown cells transfected with altMiD51GFP immunostained with anti-TOM20 (blue channel) and anti-Drp1 (red channel) monoclonal antibodies. In each image, boxed areas are shown at higher magnification in the bottom right corner. % of cells with the indicated morphology is indicated on the TOM20 panels. Scale bar, 10 mm. (d) Control Drp1 immunostaining in HeLa cells treated with a non-target siRNA. For (c) and (d), laser parameters for Drp1 and TOM20 immunostaining were identical.10.7554/eLife.27860.048Figure 13—source data 1.Mitochondrial morphologies in HeLa cells treated with non-target or Drp1 siRNAs.Mitochondrial morphologies in HeLa cells treated with non-target or Drp1 siRNAs. less

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Deep transcriptome annotation enables the discovery and functional characterization of cryptic small proteins.

In Elife on 30 October 2017 by Samandi, S., Roy, A. V., et al.

Fig.13.B
Fig.13.B showing Western Blotting from the publication: Deep transcriptome annotation enables the discovery and functional characterization of cryptic small proteins.
  • WB
AltMiD51-induced mitochondrial fragmentation is dependent on Drp1.(a) Bar graphs show mitochondrial morphologies in HeLa cells treated with non-target or Drp1 siRNAs. Cells were mock-transfected (pcDNA3.1) or transfected with altMiD51Flag. Means of three independent experiments per condition are ... more
AltMiD51-induced mitochondrial fragmentation is dependent on Drp1.(a) Bar graphs show mitochondrial morphologies in HeLa cells treated with non-target or Drp1 siRNAs. Cells were mock-transfected (pcDNA3.1) or transfected with altMiD51Flag. Means of three independent experiments per condition are shown (100 cells for each independent experiment). ***p<0.0005 (Fisher’s exact test) for the three morphologies between altMiD51 and the other experimental conditions. (b) HeLa cells treated with non-target or Drp1 siRNA were transfected with empty vector (pcDNA3.1) or altMiD51Flag, as indicated. Proteins were extracted and analyzed by western blot with antibodies against the Flag tag (altMiD51), Drp1 or actin, as indicated. (c) Confocal microscopy of Drp1 knockdown cells transfected with altMiD51GFP immunostained with anti-TOM20 (blue channel) and anti-Drp1 (red channel) monoclonal antibodies. In each image, boxed areas are shown at higher magnification in the bottom right corner. % of cells with the indicated morphology is indicated on the TOM20 panels. Scale bar, 10 mm. (d) Control Drp1 immunostaining in HeLa cells treated with a non-target siRNA. For (c) and (d), laser parameters for Drp1 and TOM20 immunostaining were identical.10.7554/eLife.27860.048Figure 13—source data 1.Mitochondrial morphologies in HeLa cells treated with non-target or Drp1 siRNAs.Mitochondrial morphologies in HeLa cells treated with non-target or Drp1 siRNAs. less

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c-Abl-mediated Drp1 phosphorylation promotes oxidative stress-induced mitochondrial fragmentation and neuronal cell death.

In Cell Death Dis on 12 October 2017 by Zhou, L., Zhang, Q., et al.

Fig.1.C
Fig.1.C showing Western Blotting from the publication: c-Abl-mediated Drp1 phosphorylation promotes oxidative stress-induced mitochondrial fragmentation and neuronal cell death.
  • WB
c-Abl is involved in the regulation of mitochondrial morphology in vivo. (a) WT and c-Abl KO littermate mice were treated with saline or MPTP (four i.p. injections of 20 mg/kg at 2 h intervals). At 2 h after total injections, immuno-EM was performed as described in Materials and Methods section. ... more
c-Abl is involved in the regulation of mitochondrial morphology in vivo. (a) WT and c-Abl KO littermate mice were treated with saline or MPTP (four i.p. injections of 20 mg/kg at 2 h intervals). At 2 h after total injections, immuno-EM was performed as described in Materials and Methods section. Two representative images (upper and lower) were picked for each group. Arrows indicate tyrosine hydroxylase-positive area containing mitochondria in SN. Scale bars, 0.5 μm. (b) Mitochondrial length was measured and divided into different categories of <0.5, 0.5–0.7, 0.7–1.0, 1.0–1.5 and >1.5 μm. Quantification of mitochondrial length is shown as percentages of total mitochondria (at least 50 mitochondria were measured per mouse). (two-way ANOVA, *P<0.05 compared with the WT group, #P<0.05 compared with the WT+MPTP group, n=3 mice.). (c) Striatal tissue or (d) ventral midbrain was collected from WT and c-Abl KO littermate mice 2 h after treatment with saline or MPTP and subjected to immunoblotting using the indicated antibodies. (e) Primary cortical neurons were immunostained with the anti-Tom20 antibody after 0.5 μM DPH treatment for 0, 0.5, 1 and 2 h, respectively, followed by confocal microscopy. Scale bars, 10 μm. (f) Quantification of mitochondrial morphology from panel (e). (two-way ANOVA, fragmented: ****P<0.0001, compared with 0 h group; n=3 slides). (g) Lysates of primary cortical neurons treated with 0.5 μM DPH for the indicated times were immunoblotted with the indicated antibodies less

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c-Abl-mediated Drp1 phosphorylation promotes oxidative stress-induced mitochondrial fragmentation and neuronal cell death.

In Cell Death Dis on 12 October 2017 by Zhou, L., Zhang, Q., et al.

Fig.1.D
Fig.1.D showing Western Blotting from the publication: c-Abl-mediated Drp1 phosphorylation promotes oxidative stress-induced mitochondrial fragmentation and neuronal cell death.
  • WB
c-Abl is involved in the regulation of mitochondrial morphology in vivo. (a) WT and c-Abl KO littermate mice were treated with saline or MPTP (four i.p. injections of 20 mg/kg at 2 h intervals). At 2 h after total injections, immuno-EM was performed as described in Materials and Methods section. ... more
c-Abl is involved in the regulation of mitochondrial morphology in vivo. (a) WT and c-Abl KO littermate mice were treated with saline or MPTP (four i.p. injections of 20 mg/kg at 2 h intervals). At 2 h after total injections, immuno-EM was performed as described in Materials and Methods section. Two representative images (upper and lower) were picked for each group. Arrows indicate tyrosine hydroxylase-positive area containing mitochondria in SN. Scale bars, 0.5 μm. (b) Mitochondrial length was measured and divided into different categories of <0.5, 0.5–0.7, 0.7–1.0, 1.0–1.5 and >1.5 μm. Quantification of mitochondrial length is shown as percentages of total mitochondria (at least 50 mitochondria were measured per mouse). (two-way ANOVA, *P<0.05 compared with the WT group, #P<0.05 compared with the WT+MPTP group, n=3 mice.). (c) Striatal tissue or (d) ventral midbrain was collected from WT and c-Abl KO littermate mice 2 h after treatment with saline or MPTP and subjected to immunoblotting using the indicated antibodies. (e) Primary cortical neurons were immunostained with the anti-Tom20 antibody after 0.5 μM DPH treatment for 0, 0.5, 1 and 2 h, respectively, followed by confocal microscopy. Scale bars, 10 μm. (f) Quantification of mitochondrial morphology from panel (e). (two-way ANOVA, fragmented: ****P<0.0001, compared with 0 h group; n=3 slides). (g) Lysates of primary cortical neurons treated with 0.5 μM DPH for the indicated times were immunoblotted with the indicated antibodies less

Collected and cropped from Cell Death Dis by CiteAb, provided under a CC-BY license

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