rat cardiomyocyte growth medium  (Procell Inc)

Journal: Frontiers in Pharmacology

Article Title: A Polyphenol-Rich Extract of Olive Mill Wastewater Enhances Cancer Chemotherapy Effects, While Mitigating Cardiac Toxicity

doi: 10.3389/fphar.2021.694762

Figure Lengend Snippet: Activities of A009 extracts combined with chemotherapy on tumor cells and cardiomyocytes. A009 (batch L3) deceases the proliferation rate of tumors cells in vitro [ (A) : DU-145 PCa and (B) : HT29 CRC and has additive effects on the cisplatin and the 5-Fluorouracil (5FU) effects. The cardiomyocytes proliferation rate is not affected by A009 alone (C) , and reduced proliferation by 5FU and cisplatin is not further decreased by A009 in vitro . Control: vehicle control. Data are showed as mean ± SEM, one-way ANOVA, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The rat cardiomyocyte cell line H9C2 (PromoCell) was maintained in Myocyte Growth Medium plus Myocyte supplements mix (PromoCell), addition with 10% Fetal Bovine Serum (FBS) (Euroclone), 2 mM l -glutamine (Euroclone), 100 U/ml penicillin and 100 μg/ml streptomycin (Euroclone), at 37°C, 5% CO 2 .

Techniques: In Vitro

Journal: Frontiers in Pharmacology

Article Title: A Polyphenol-Rich Extract of Olive Mill Wastewater Enhances Cancer Chemotherapy Effects, While Mitigating Cardiac Toxicity

doi: 10.3389/fphar.2021.694762

Figure Lengend Snippet: Protective activities of A009 extracts on neonatal murine cardiomyocytes. The cardioprotective effects of the A009 extract on chemotherapy induced cardiotoxicities was assessed, in vitro , on neonatal murine cardiomyocytes. Neonatal murine cardiomyocytes were exposed for 24 h (A) and 48 h (B) to 5-FU (4.6 μM) alone, A009 extracts (dilution 1:800, batches L3 or L4) alone, or the combination of the 5-FU and A009 extracts (dilution 1:800, batches L3 or L4). Number of cardiomyocytes was not affected by 5FU after 24 h treatments, while the OMWW extract improved their number to a slight but significative percent. After 48 h 5FU strongly decreased cardiomyocyte number to 60%. A009 alone did not lower the quantity of cardiomyocytes, and even slightly increased it as in the 24 h (in the L4 formulation). Addition to L3 and L4 together with the cytotoxic drug counteracted 5FU effect at 48 h. Data are showed as mean ± SEM. 5-FU: 5-fluoro-Uracile; L3/L4: A009 batch extract; Control: vehicle control; 5-fluorouracil.

Article Snippet: The rat cardiomyocyte cell line H9C2 (PromoCell) was maintained in Myocyte Growth Medium plus Myocyte supplements mix (PromoCell), addition with 10% Fetal Bovine Serum (FBS) (Euroclone), 2 mM l -glutamine (Euroclone), 100 U/ml penicillin and 100 μg/ml streptomycin (Euroclone), at 37°C, 5% CO 2 .

Techniques: In Vitro

Journal: iScience

Article Title: Hypoxia inhibits the cardiac I K1 current through SUMO targeting Kir2.1 activation by PIP 2

doi: 10.1016/j.isci.2022.104969

Figure Lengend Snippet: Acute hypoxia inhibits I K1 via SUMOylation I K1 in rat ventricular cardiomyocytes (RVCMs) was studied by the whole-cell patch-clamp. Currents were evoked in a high external K + recording buffer and measured at −120 mV, as described in the . Hypoxia was a drop in O 2 from ambient levels to 2% (red), measured at the cell. The time-course of hypoxic inhibition was studied by recording the magnitude of I K1 every second. Cells were studied with the following pipette solutions: control (blue), SUMO1 (1 μM, orange), or SENP1 (2 μM, magenta). CRY2-SENP1 was targeted to the cell membrane by the activation of a 460 nm LED during the time period indicated. I K1 currents were blocked by the addition of 3 mM Ba 2+ (green). (A) Exposure to acute hypoxia (2% O 2 ) inhibits ∼40% of I K1 in RVCMs. Left , representative current sweeps; right , a representative time course showing the kinetics of hypoxic inhibition in RVCMs studied with a control pipette solution. The inhibition is precluded by including SENP1 (2 μM) in the recording pipette. The reverse ramp recording protocol is inset. (B) Including SUMO1 (1 μM) in the recording pipette inhibits I K1 and precludes the effects of hypoxia. SENP1 augments I K1 and protects the current from hypoxia. Data are from 8 to 10 RVCMs per group, ∗∗∗∗p < 0 . 0001 , paired, two-tailed Student’s t test. (C) Hypoxic inhibition of I K1 was reversed by the activation of CRY2-SENP1 with 460 nm light (cyan box) when RVCMs are studied with a control pipette solution. Right , CRY2-SENP1 does not alter current when 2 μM SENP1 is included in the recording pipette. Data are from 6 RVCMs, ∗p < 0 . 05 , ∗∗∗∗p < 0 . 0001 , paired, two-tailed Student’s t test.

Article Snippet: Rat Cardiomyocyte Growth Medium (rCGM) BulletKit , Lonza Biosciences , Cat# CC-4515.

Techniques: Patch Clamp, Inhibition, Transferring, Control, Membrane, Activation Assay, Two Tailed Test

Journal: iScience

Article Title: Hypoxia inhibits the cardiac I K1 current through SUMO targeting Kir2.1 activation by PIP 2

doi: 10.1016/j.isci.2022.104969

Figure Lengend Snippet: PIP 2 opposed hypoxic inhibition of I K1 I K1 was studied in rat ventricular cardiomyocytes (RVCMs) using a whole-cell patch-clamp recording. To the knockdown Kir2.1 expression, RVCMs were transduced with lentiviral particles carrying eGFP and shRNA targeting KCNJ2. Kir2.1 knockdown cells (Kir2.1 kd -CMs) were identified by the expression of eGFP. Cells were studied with a control pipette solution (blue), or with pipette solutions containing purified SUMO1 (1 μM, orange) or SENP1 (2 μM, magenta). Where indicated, the control pipette solution contained diC8 PIP 2 . Paired patch-clamp data were analyzed by Students t -test; ∗∗∗, p < 0.01. The proximity ligation assay (PLA) for native Kir2.1-SUMO1 interactions was performed as described in the and analyzed using an unpaired Mann-Whitney rank test, ∗∗∗∗p < 0.001. See also Figures S7 and . (A) Left , Example sweeps show that I K1 is diminished in Kir2.1 kd -CMs and the remaining current is insensitive to acute hypoxia. Kir2.1 kd -CMs are identified by expression of GFP ( inset , scale bar = 10 μm). Right , summary data from 8 to 10 cells per group show that the regulation of I K1 by hypoxia, SUMO1, and SENP1 is lost in Kir2.1 kd -CMs. (B) The magnitude and the regulation of I K1 by hypoxia, SUMO1, and SENP1 are unaltered when cells are treated with a control, scrambled shRNA; 7-8 cells per group. (C) Left , Example traces to show that including diC8-PIP 2 in the pipette solution reduces the hypoxic inhibition of I K1 in RVCMs in a concentration-dependent manner. The arrow indicates the change in current magnitude between exposure to ambient O 2 and 2% O 2 in the same cell. Right , Concentration-response curve showing that including diC8-PIP 2 in the pipette opposes hypoxic inhibition of I K1 in control RVCMs and RVCMs expressing the scrambled shRNA. The hypoxic-response is diminished in Kir2.1 kd -CMs. Data are mean ± s.d. for 7-8 cells per condition. (D) DiC8-PIP 2 decreases hypoxic inhibition of Kir2.1 channels expressed in HEK293T cells (control) in a concentration-dependent manner using the experimental paradigm described in C, above. Current inhibition is diminished when SENP1 is included in the recording pipette and is not observed when Kir2.1-K49Q channels are studied. Data are mean ± s.d. for 6 cells per condition. (E) PLA shows that native Kir2.1 colocalizes with SUMO1 in RVCMs and that Kir2.1-SUMO interactions are increased by acute hypoxia. Kir2.2 and Kir2.3 do not associate with SUMO1. Representative data are shown with DAPI-labeled nuclei in red and PLA interactions in green for ease of visualization. Summary data are obtained from multiple experiments each with multiple fields of view containing 20-30 nuclei. The scale bar is 10 μm.

Article Snippet: Rat Cardiomyocyte Growth Medium (rCGM) BulletKit , Lonza Biosciences , Cat# CC-4515.

Techniques: Inhibition, Patch Clamp, Knockdown, Expressing, Transduction, shRNA, Control, Transferring, Purification, Proximity Ligation Assay, MANN-WHITNEY, Concentration Assay, Labeling

Journal: iScience

Article Title: Hypoxia inhibits the cardiac I K1 current through SUMO targeting Kir2.1 activation by PIP 2

doi: 10.1016/j.isci.2022.104969

Figure Lengend Snippet:

Article Snippet: Rat Cardiomyocyte Growth Medium (rCGM) BulletKit , Lonza Biosciences , Cat# CC-4515.

Techniques: Recombinant, Virus, shRNA, Control, In Situ, Software, Microscopy