Journal: Critical care medicine

Article Title: Inhibition of the Mitochondrial Fission Protein Drp1 Improves Survival in a Murine Cardiac Arrest Model

doi: 10.1097/CCM.0000000000000817

Figure Lengend Snippet: 6-month-old female C57B6 were subjected to an 8-minute asystolic, non-ventilated cardiac arrest induced by KCL. Ventilations and chest compressions were then performed for 90 seconds followed by intravenous epinephrine and Mdivi-1 or DMSO control. CPR was then continued until ROSC or terminated after 5 minutes. Mice achieving ROSC were monitored and ventilated for 2 hours before extubation. Survival and Neurological outcomes were then monitored over 72 hours.

Article Snippet: Mdivi-1 was obtained from Enzo life sciences (Farmingdale, NY).

Techniques:

Journal: Critical care medicine

Article Title: Inhibition of the Mitochondrial Fission Protein Drp1 Improves Survival in a Murine Cardiac Arrest Model

doi: 10.1097/CCM.0000000000000817

Figure Lengend Snippet: A. Mdivi-1, an inhibitor of Drp1 was administered following 8 minutes of cardiac arrest IV. Mdivi-1 dephosphorylation at Serine 637 following cardiac arrest was prevented by Mdivi-1. Relative densitometry to the right expressed as box plots. * (One Way ANOVA p=0.054, pairwise t test p,0.03, N=5 CA vs. CA+Mdivi-1). B. Mitochondrial Protein fractions were probed for Drp1. Increased Drp1 accumulation in the mitochondrial fraction 2 hours post arrest was observed compared to Sham and Mdivi-1 treated mice. VDAC demonstrated mitochondrial purity and equal loading. Rho GDI demonstrates lack of cytosolic protein contaminants (One Way ANOVA p<0.001, N=4, all groups significantly different from one another by pairwise t-test and/or Wilcoxon rank sum test). Error Bars 95% confidence intervals.

Article Snippet: Mdivi-1 was obtained from Enzo life sciences (Farmingdale, NY).

Techniques: De-Phosphorylation Assay

Journal: Critical care medicine

Article Title: Inhibition of the Mitochondrial Fission Protein Drp1 Improves Survival in a Murine Cardiac Arrest Model

doi: 10.1097/CCM.0000000000000817

Figure Lengend Snippet: Representative TEM images and mean values for mitochondria area from mouse hearts two hours post cardiac arrest resuscitation. 2 hours post cardiac resuscitation mitochondria were significantly smaller compared to Sham and Mdivi-1 treated mice suggesting mitochondrial fission. Mdivi-1 and Sham groups were not significantly different from one another (p<0.001, One Way ANOVA, CA different from Sham and CA+Mdivi-1 by pairwise t test, n>600/group from 3 hearts). Scale bar = 1µM. Error Bars ±SEM.

Article Snippet: Mdivi-1 was obtained from Enzo life sciences (Farmingdale, NY).

Techniques:

Journal: Critical care medicine

Article Title: Inhibition of the Mitochondrial Fission Protein Drp1 Improves Survival in a Murine Cardiac Arrest Model

doi: 10.1097/CCM.0000000000000817

Figure Lengend Snippet: A–B. Myocardial measurements of ATP, ADP, and ATP/ADP were not significantly different between Sham, cardiac arrest, and cardiac arrest + Mdivi-1 groups 15 minutes after resuscitation from cardiac arrest. Myocardial tissue Lactate following cardiac arrest (CA) however, was significantly increased compared to Sham and Mdivi-1 treated mice (***One Way ANOVA, p<0.001, pairwise t test Sham vs. CA p=0.009, CA vs. CA+Mdivi-1 p=0.009, CA+Mdivi-1 vs. Sham p=0.057, N=4). Values for Sham and CA+Mdivi-1 mice were not significantly different C. Decreased activity of aconitase is a marker for increased ROS generation and was assessed 15 minutes after resuscitation from cardiac arrest. Aconitase activity following cardiac arrest was significantly decreased compared to Mdivi-1 and Sham treated hearts. Mdivi-1 preserved aconitase activity following resuscitation from cardiac arrest. Mdivi-1, however was not completely preserved aconitase levels and was significantly depressed from Sham (***One Way ANOVA, p <0.001, all pairs significantly different from one another by pairwise t-test p<0.001, N=5). Error bars ±SEM.

Article Snippet: Mdivi-1 was obtained from Enzo life sciences (Farmingdale, NY).

Techniques: Activity Assay, Marker

Journal: Critical care medicine

Article Title: Inhibition of the Mitochondrial Fission Protein Drp1 Improves Survival in a Murine Cardiac Arrest Model

doi: 10.1097/CCM.0000000000000817

Figure Lengend Snippet: Myocardial performance was assessed with a Millar catheter. Maximum Left ventricular maximum systolic pressure (LVP max), dp/dt max, dp/dt min, and stroke volume were depressed at all time points examined following cardiac arrest compared to Sham operated controls and cardiac arrest + Mdivi-1 treated mice. Cardiac output was also significantly worse following cardiac arrest compared to the Mdivi-1 treated group at 90 and 120 minutes following resuscitation from cardiac arrest. Heart rates were significantly higher in Mdivi-1 treated mice compared to cardiac arrest alone animals. N=13, Error bars show 95% confidence intervals. *p<0.05.

Article Snippet: Mdivi-1 was obtained from Enzo life sciences (Farmingdale, NY).

Techniques:

Journal: Critical care medicine

Article Title: Inhibition of the Mitochondrial Fission Protein Drp1 Improves Survival in a Murine Cardiac Arrest Model

doi: 10.1097/CCM.0000000000000817

Figure Lengend Snippet: Left ventricular transthoracic echocardiography was used to measure the percent fractional shortening (%FS) in Sham operated mice, 8 min cardiac arrest (CA), and 8 min cardiac arrest+Mdivi-1 mice. 2 hours following resuscitation, %FS was depressed in CA mice (20%FS), compared to CA+Mdivi-1 (38%FS), and Sham operated mice (37%FS).* (One Way ANOVA, p=0.003, pairwise t test Sham vs. CA p=0.004, Sham vs. CA+ Mdivi p=0.81, CA vs. CA+Mdivi-1 p=0.004. Representative echocardiograms are shown to the left.

Article Snippet: Mdivi-1 was obtained from Enzo life sciences (Farmingdale, NY).

Techniques:

Journal: Critical care medicine

Article Title: Inhibition of the Mitochondrial Fission Protein Drp1 Improves Survival in a Murine Cardiac Arrest Model

doi: 10.1097/CCM.0000000000000817

Figure Lengend Snippet: 24 hours following cardiac arrest survival in the control group was only 43% (9/21) compared to 85% (17/20) in the Mdivi-1 group and 100% of the sham animals. Survival decreased to 38% (8/21) in the controls, 65% (13/20) in the Mdivi-1, and 94% (16/17) of sham animals 72 hours following cardiac arrest. All 3 groups were significantly different. Log Rank (Mantel Cox) Test. p<0.001. N= number of animals at each time point.

Article Snippet: Mdivi-1 was obtained from Enzo life sciences (Farmingdale, NY).

Techniques:

Journal: Critical care medicine

Article Title: Inhibition of the Mitochondrial Fission Protein Drp1 Improves Survival in a Murine Cardiac Arrest Model

doi: 10.1097/CCM.0000000000000817

Figure Lengend Snippet: At 6, 18, 24, 48, and 72 hour post cardiac arrest, Mdivi-1 treated mice had significantly better scores than control cardiac arrest animals. Sham mice scored better than Mdivi-1 treated mice at only 6 and 18 hours post cardiac arrest. Ordinal regression analysis demonstrated significant differences between CA and CA+Mdivi-1 and CA and Sham. *p<0.001

Article Snippet: Mdivi-1 was obtained from Enzo life sciences (Farmingdale, NY).

Techniques:

Journal: Immunology and cell biology

Article Title: Metabolic control of cell fate bifurcations in a hematopoietic progenitor population

doi: 10.1111/imcb.12040

Figure Lengend Snippet: Mitochondrial dynamics and metabolism promote selective DC differentiation. (a) Fatty acid oxidation modulates cDC subset development. Left, frequency of cDC and pDCs following etomoxir treatment. n.s. not significant. Student’s t-test with Holm-Sidak correction. Right, development of IRF8+ and IRF4+ DC subsets following etomoxir treatment. **P < 0.01. Paired t-test. (b) Mitoclearance assay. Progenitors were pulse-labeled with MTGreen at day 0 or day 5 of culture and assessed for decay of fluorescence 3 days later. FACS plots show MTGreen fluorescence by cell division with or without mDivi-1 treatment. Trapezoid gate denotes MTGreen-low population. Plots are representative of 3 independent experiments. (c) Left, unequal levels of aged (pulse-labeled) mitochondria in IRF8-hi and IRF8-int progenitors at day 3 of culture. Right: Quantification of MFI of aged mitochondria in IRF8-hi and IRF8-int progenitors. * P < 0.05. Paired T test. (d) Total mitochondrial content (left) and mitochondrial membrane potential (right) in mature cDC1 and cDC2 subsets at day 9 of culture. Histograms are representative of 3 independent experiments. (e) Upper left, gating of subsets with low or high levels of mitoclearance. Upper right, total mitochondrial content in proliferating progenitors stained at day 0. Lower, total mitochondrial content of populations with low (left) or high (right) levels of retained, pulse-labeled mitochondria, gated as in upper left panel. Vertical statistic denotes MFI of Mitotracker staining. (f) Left, frequency of cDC and pDCs amongst CD11c+ DCs following mDivi-1 (upper row) or M1 (lower row) treatment. *P < 0.05. n.s. not significant. Student’s t-test with Holm-Sidak correction. Right, frequency of IRF8+ and IRF4+ cDC subsets following treatment with mDivi-1 (upper) or M1 (lower). **P < 0.01. ***P < 0.001. Paired t-test. (g) Flt3L induces mROS production in proliferating progenitors. Left, elevated MitoSox levels in cells with high versus low levels of old mitochondria. Histograms are representative of 2 independent experiments. Middle, MitoSox levels versus cell division at day 3 of culture within IRF8-int and IRF8-high progenitors. Right, quantification of MitoSox MFI in IRF8-int and IRF8-high cells. ****P < 0.0001. Student’s t-test. (h) Left, frequency of cDC and pDCs within CD11c+ DCs following treatment with N-Ac. n.s. not significant. Student’s t-test with Holm-Sidak correction. Right, frequency of IRF8+ and IRF4+ cDC subsets following treatment with N-Ac. *P < 0.05. Paired t-test.

Article Snippet: All drugs were added at the beginning of culture and included: mDivi-1 (Cayman Chemical, Ann Arbor, MI, USA, 10µM), M1 (Sigma-Aldrich, St. Louis, MO, USA, 5µM), N-Acetyl cysteine (N-Ac, Sigma, 5mM), 2-deoxyglucose (2-DG, Sigma, 100µM-1mM), and etomoxir (Cayman, 100µM), an inhibitor of the essential fatty acid oxidation enzyme, carnitine palmitoyltransferase I.

Techniques: Labeling, Fluorescence, Staining

Journal: Immunology and cell biology

Article Title: Metabolic control of cell fate bifurcations in a hematopoietic progenitor population

doi: 10.1111/imcb.12040

Figure Lengend Snippet: Nutrient sensor AMPK promotes selective DC differentiation. (a) AMPK signaling in dividing progenitors stimulated by Flt3. Levels of phospho-AMPK, phospho-ACC, and phospho-Raptor in proliferating (black line) and undivided (grey fill) progenitor cells stimulated with Flt3L for 3 days. Plots are representative of 2–3 independent experiments. (b) AMPK-α1 deficiency alters DC development in vitro. Left, development of cDC/pDCs from AMPK-α1 deficient cells and heterozygote littermate (“WT”) controls. n.s. not significant. Student’s t-test with Holm-Sidak correction. Right, differentiation of IRF8+ and IRF4+ cDC subsets from AMPK-α1 deficient progenitors with or without mDivi-1 treatment. *P < 0.05 **P < 0.01 ***P < 0.001. Bar graphs indicate frequency of each subset +/− SEM. Repeated measures one way ANOVA with Tukey’s correction. (c) Steady state in vivo bone marrow DC progenitor (upper), as well as skin draining lymph node (lower) cDC/pDC and cDC subset frequencies of AMPK-α1 knockout mice and heterozygote littermate (WT) controls. In bone marrow, DC progenitors were gated as lineage (CD3, CD19, B220, NK1.1, Ter119, GR1, CD11b) negative, Flt3R+, CX3CR1+ cells. Macrophage dendritic cell progenitors (MDP) were c-kit+ CD11c−. Common dendritic progenitors (CDP) were c-kit− CD11c−. Pre-DCs were c-kit− CD11c+. In lymph nodes, pDCs were lineage- CD11c+ MHCII-int B220+, and within lineage- CD11c+ MHCII+ B220− cDCs, cDC1 were gated as CD8α+ CD11b−, migratory DCs were CD8α- CD11b−, and cDC2 were CD8α− CD11b+. *P < 0.05. ***P < 0.001. n.s. not significant. Paired t-test.

Article Snippet: All drugs were added at the beginning of culture and included: mDivi-1 (Cayman Chemical, Ann Arbor, MI, USA, 10µM), M1 (Sigma-Aldrich, St. Louis, MO, USA, 5µM), N-Acetyl cysteine (N-Ac, Sigma, 5mM), 2-deoxyglucose (2-DG, Sigma, 100µM-1mM), and etomoxir (Cayman, 100µM), an inhibitor of the essential fatty acid oxidation enzyme, carnitine palmitoyltransferase I.

Techniques: In Vitro, In Vivo, Knock-Out

Journal: Scientific Reports

Article Title: Inhibition of Drp1 orchestrates the responsiveness of breast cancer cells to paclitaxel but insignificantly relieves paclitaxel-related ovarian damage in mice

doi: 10.1038/s41598-023-49578-0

Figure Lengend Snippet: The cytotoxic effect of PTX is enhanced by Drp1 deficiency in breast cancer cells. The cytotoxicity of PTX was evaluated in PTX-sensitive and PTX-resistant cells by MTT assay, where the IC 50 of mdivi-1 in PTX-sensitive cells was 59.5 µM ( A ). PTX-sensitive cells (green dashed line) and resistant cells (red dashed line) were incubated with a concentration range of PTX. Also, resistant cells were incubated with different concentrations of PTX combined with the IC 50 of mdivi-1 (blue dashed line) and the percent of viable cells was determined and normalized to the control cells ( B ). Also, the percentage of cell viability was determined in PTX-resistant cells exposed to concentration range of PTX after their transfection Drp1-specific siRNA ( C ). The IC 50 of PTX in PTX-sensitive cells is 85 nM, where for PTX+mdivi-1 is 132 uM. D through F are representative photomicrographs of PTX-sensative cells treated with PTX, PTX-resistant cells treated with PTX, and PTX-resistant cells breated with both PTX and mdivi-1, respectively (Magnification 20X). Dots represent the mean (± SD) of viable cells of n = 5–6 readings. (*) and (#): refer to significant changes in cell viability of PTX+mdivi-1 versus PTX, P < 0.05 and P < 0.001, respectively. “D-F” are representative light micrographs of PTX-regular or resistant cells treated with PTX or cells dually treated with PTX+Mdivi1, respectively.

Article Snippet: Mdivi-1 was obtained from APExBIO Technology LLC, TX, USA, Unitaxel (Paclitaxel) was from Hikma, Egypt (Concentration 300 mg paclitaxel/50 ml), and cisplatin was purchased from Mylan, Viatris, PA, USA.

Techniques: MTT Assay, Incubation, Concentration Assay, Transfection

Journal: Scientific Reports

Article Title: Inhibition of Drp1 orchestrates the responsiveness of breast cancer cells to paclitaxel but insignificantly relieves paclitaxel-related ovarian damage in mice

doi: 10.1038/s41598-023-49578-0

Figure Lengend Snippet: Computational modeling of the molecular interactions of paclitaxel, and mdivi-1 with mitochondrial redox-related enzymes. The 2D interaction diagrams demonstrate the interaction of ligands (PTX or mdivi-1) with mitochondrial ATP synthase, SOD or Thioredoxin reductase 1. The hot spots in protein domains, the superimposition of the docking pose, and the co-crystallized ligands in the active sites in the enzyme active site are shown. The docking score is shown next each protein.

Article Snippet: Mdivi-1 was obtained from APExBIO Technology LLC, TX, USA, Unitaxel (Paclitaxel) was from Hikma, Egypt (Concentration 300 mg paclitaxel/50 ml), and cisplatin was purchased from Mylan, Viatris, PA, USA.

Techniques:

Journal: Scientific Reports

Article Title: Inhibition of Drp1 orchestrates the responsiveness of breast cancer cells to paclitaxel but insignificantly relieves paclitaxel-related ovarian damage in mice

doi: 10.1038/s41598-023-49578-0

Figure Lengend Snippet: Changes in the mitochondrial morphology, and mitochondrial dynamic-related proteins. Electronmicrographs of the mitochondrial morphological changes in PTX-resistant cells ( A ), cells treated with mdivi-1, PTX, or PTX+mdivi-1. ( B ) depicts immunoblotting of the mitochondrial-related protein (Drp1), and fusion proteins (Mfu1 & Mfu2) and the corresponding band intensities ( C ). Drp1 expression was normally expressed in PTX-resistant cells, similar to regular cells. Mdivi-1, alone or combined with PTX did not affect the expression of Drp1, whereas it is downregulated in cells treated with PTX after siRNA transfection. Mfu1&Mfu2 were significantly upregulated in cell treated with mdivi-1 or PTX after siRNA transfection. ( D ) shows the in vitro cell migration of cells treated with PTX, Mdivi-1, mdivi-1+PTX, or cells transfected with siRNA and then were treated with PTX. *, **, *** refer to P < 0.05, P < 0.01, P < 0.001, compared to the untreated cells.

Article Snippet: Mdivi-1 was obtained from APExBIO Technology LLC, TX, USA, Unitaxel (Paclitaxel) was from Hikma, Egypt (Concentration 300 mg paclitaxel/50 ml), and cisplatin was purchased from Mylan, Viatris, PA, USA.

Techniques: Western Blot, Expressing, Transfection, In Vitro, Migration

Journal: Scientific Reports

Article Title: Inhibition of Drp1 orchestrates the responsiveness of breast cancer cells to paclitaxel but insignificantly relieves paclitaxel-related ovarian damage in mice

doi: 10.1038/s41598-023-49578-0

Figure Lengend Snippet: Change in mice body weight, ovarian weight, follicular count, and E2 and AMH levels. Mice body weight ( A ) after various treatments were compared to the initial body weight (Day 0), ovarian weights of treated groups were compared to the corresponding control group ( B ). Differential follicular count demonstrated massive decrease of the count of various types ( C ). Both E2 ( D ) and AMH ( E ) decreased in PTX or cisplatin groups, whereas cotreatment with mdivi-1 did not show significant improvements compared to PTX or cisplatin groups. Data are shown as bars or dots indicating the mean (± SD). Abbreviations : E2: estradiol, AMH: antimullerian hormone.

Article Snippet: Mdivi-1 was obtained from APExBIO Technology LLC, TX, USA, Unitaxel (Paclitaxel) was from Hikma, Egypt (Concentration 300 mg paclitaxel/50 ml), and cisplatin was purchased from Mylan, Viatris, PA, USA.

Techniques:

Journal: Scientific Reports

Article Title: Inhibition of Drp1 orchestrates the responsiveness of breast cancer cells to paclitaxel but insignificantly relieves paclitaxel-related ovarian damage in mice

doi: 10.1038/s41598-023-49578-0

Figure Lengend Snippet: Representative hematoxylin and eosin-stained ovarian sections recovered from healthy mice ( A ), mice intraperitoneally injected with PTX ( B ), Cisplatin ( C ), PTX+mdivi-1 ( D ), or Cis+mdivi-1 ( E ). Both PTX and Cisplatin reduced the size of ovaries, the number of healthy follicles. Cotreatment with mdivi-1did not exert a significant repair.

Article Snippet: Mdivi-1 was obtained from APExBIO Technology LLC, TX, USA, Unitaxel (Paclitaxel) was from Hikma, Egypt (Concentration 300 mg paclitaxel/50 ml), and cisplatin was purchased from Mylan, Viatris, PA, USA.

Techniques: Staining, Injection