caspase 9 inhibitor z lehd fmk  (R&D Systems)

Journal: bioRxiv

Article Title: Energy Flux Regulates Cell Death Induced by California Serogroup Orthobunyaviruses

doi: 10.64898/2026.03.13.711679

Figure Lengend Snippet: (A-B) BE(2)-C wild type cells were treated with nothing, DMSO vehicle, or 5 µM oligomycin ATP synthase inhibitor immediately prior to inoculation with LACV (MOI = 1) or INKV (MOI = 1). At 48 (LACV) or 96 (INKV) HPI, the percent survival was determined as compared to uninfected controls. (C) BE(2)-C WT cells were treated with nothing, DMSO vehicle, or 10 µM Z-LEHD-FMK caspase 9 inhibitor immediately prior to inoculation with LACV (MOI = 1). At 48 (LACV) HPI, the percent survival was determined as compared to uninfected controls. (D) BE(2)-C WT cells were treated with nothing, DMSO vehicle, or 10 µM Q-VD-Oph pan caspase inhibitor immediately prior to inoculation with LACV (MOI = 1). At 48 (LACV) HPI, the percent survival was determined as compared to uninfected controls. (A-D) Asterisks represent significance values from one-way ANOVA as follows: *p ≤0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001; n = 3.

Article Snippet: Caspase 9 inhibitor Z-LEHD-FMK (R&D Systems, catalog #FMK008) was dissolved in DMSO to create a 20 mM stock solution.

Techniques:

Journal: bioRxiv

Article Title: Energy Flux Regulates Cell Death Induced by California Serogroup Orthobunyaviruses

doi: 10.64898/2026.03.13.711679

Figure Lengend Snippet: (A&D) The effect of interventions on apoptosis was interrogated by measuring activated caspase 9 approximately 24 hours post treatment via flow cytometry. Camptothecin (4 ug/mL) was utilized to induce apoptosis via caspase 9 action. Activated caspase 9 was detected using a fluorochrome inhibiter of caspases (FLICA) conjugated to a green FAM fluorescent probe. Samples were run on a CytoFLEX LX cytometer and the percent change in median fluorescence intensity (MFI) from BE(2)-C wild type, untreated cells was reported. (A) 10 µM Q-VD-Oph pan caspase inhibitor was challenged with LACV MOI = 1 or camptothecin (4 ug/mL). Each bar represents the MFI from a single run of flow cytometry; traces are available in Supplemental Figure 6. (B) BE(2)-C wild type cells were treated with DMSO vehicle or 10 µM Q-VD-Oph pan caspase inhibitor immediately prior to inoculation with LACV (MOI = 1, 5) then incubated at either 37°C or at 33°C for 48 HPI. Percent survival was determined as compared to uninfected controls. (C) BE(2)-C wild type cells were pre-grown for at least three days and subsequently maintained in media with either 10 mM galactose or 10 mM glucose as the primary carbon source. Cells were treated with either DMSO vehicle or 10 µM Q-VD-Oph pan caspase inhibitor immediately prior to inoculation with LACV (MOI = 1, 5). At 48 HPI, percent survival was determined as compared to uninfected controls. (B-C) Asterisks represent significance values from unpaired t-tests as follows: ns = not significant, *p ≤0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001; n=3. (D) 10 mM galactose as the primary carbon source (initiated at least 3 days prior to and maintained through inoculation and incubation), 5 µM oligomycin, and 33°C incubation temperature were challenged with camptothecin (4 ug/mL). Each bar represents the MFI from a single run of flow cytometry; traces are available in Supplemental Figure 6.

Article Snippet: Caspase 9 inhibitor Z-LEHD-FMK (R&D Systems, catalog #FMK008) was dissolved in DMSO to create a 20 mM stock solution.

Techniques: Flow Cytometry, Cytometry, Fluorescence, Incubation

Journal: bioRxiv

Article Title: Mitochondrial Respiratory Chain Function is crucial for Muscle Toxicity in Facioscapulohumeral Muscular Dystrophy

doi: 10.1101/2025.11.25.690559

Figure Lengend Snippet: (A) Left: longitudinal quantitation of DUX4 and ZSCAN4-tdTomato containing nuclei in iDUX4 ZSCAN4-tdT reporter myoblasts at 2h intervals after DOX (62.5 ng/mL). Right: immunofluorescence micrographs of iDUX4 ZSCAN4-tdT reporter myoblasts before (0h), and 4h and 16h after exposure to DOX (62.5 ng/mL; green=DUX4, red=ZSCAN4-tdTomato, blue=nuclei; scale bar=100μm). (B) Normalised oxygen consumption rate (OCR) curves of iDUX4 control (CTRL) and DOX-induced (62.5 ng/mL DOX for 4h and 16h) myoblasts. (C) Respirometric assessment of mitochondrial respiration reveals reduction of basal, maximal and ATP-linked respiration in iDUX4 myoblasts 4h after DOX (62.5 ng/mL). (D) Normalised extracellular acidification rate (ECAR) curves of iDUX4 control (CTRL) and DOX-induced (62.5 ng/mL DOX for 4h and 16h) myoblasts. (E) ATP production rates demonstrating reduction of ATP in iDUX4 myoblasts 4h after DOX (62.5 ng/mL), mainly arising from defective oxidative phosphorylation (OXPHOS; mitoATP), while glycoATP is unaffected before 16h. (F) Assaying of Casp9 (red; onset 4h after DOX) and Casp3/7 activation (grey; onset 8h after DOX), measured at 2h intervals over 16h. (G) Assaying Annexin V as a marker of apoptosis in iDUX4 myoblasts after DOX (62.5 ng/mL), measured at 2h intervals over 16h. (H) Casp3/7 activation in iDUX4 myoblasts 8h after DOX (62.5 ng/mL) is prevented by a Casp9 inhibitor (Z-LEHD-FMK TFA; 10 μM). (I) Casp3/7 and Casp9 activation in iDUX4 myoblasts 8h after DOX (62.5 ng/mL) with non-targeted (Q10; 20 μM) or mitochondria-targeted (mitoQ10; 20μM) Coenzyme Q10. [n=3-6, data is mean ± s.d., where * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001].

Article Snippet: Caspase 9 inhibitor Z-LEHD-FMK TFA (S7313) and mitoQ10 (mesylate; S8978) were from Selleckchem (Houston, TX, USA).

Techniques: Quantitation Assay, Immunofluorescence, Control, Phospho-proteomics, Activation Assay, Marker