Journal: Cell Death Discovery

Article Title: Combination of eribulin and anlotinib exerts synergistic cytotoxicity in retroperitoneal liposarcoma by inducing endoplasmic reticulum stress

doi: 10.1038/s41420-024-02103-2

Figure Lengend Snippet: A – C Dot plots showing the top 3 KEGG pathway enrichment results of anlotinib, eribulin, and combination group. D GSEA plot featuring protein processing in endoplasmic reticulum pathway. E ERS induced by combination therapy was validated through western blot, wherein key ERS markers were upregulated and apoptosis pathway was initiated. F Rescue experiments further validated that a synergistic effect was largely induced by ERS. After applying salubrinal, synergy scores were significantly reduced.

Article Snippet: Salubrinal was purchased from MedChemExpress (Monmouth Junction, NJ, USA).

Techniques: Western Blot

Journal: Journal of Cellular and Molecular Medicine

Article Title: Endoplasmic reticulum stress remodels alveolar bone formation after tooth extraction

doi: 10.1111/jcmm.15753

Figure Lengend Snippet: ER stress plays an important role in bone formation and is regulated by salubrinal (SAL) concentration. A‐B, Osteogenic induction in POBs for 7 d with salubrinal administered at 0.2, 1 or 5 μmol/L in the presence of ER stress inducer tunicamycin (Tm; 100 ng/mL). After 48 h, ALP expression (black, 100×) decreased with Tm treatment, which was reversed by SAL. C, CCK8 assay for POBs showing the same trend. D‐E, Representative Western blots showing SAL’s protection of osteogenesis against ER stress. The expression of Runx2/β‐actin protein increased and the expression of p‐eIF2α/eIF2α decreased with recovery of the ER. Data were presented as the ratio with β‐actin and mean ± SD. The asterisks (*) represent P < .05. VC, vehicle control

Article Snippet: Experimental tooth extraction was performed on 48 Sprague‐Dawley rats which randomly dived into four groups (n = 12/group): (a) M1 extraction, observation for 7 days; (b) M1 extraction, observation for 14 days; (c) M1 extraction + PBS (3 days post‐extraction), observation for 7 days; (d) M1 extraction + Salubrinal (T3045; Target Mol, resolved in DMSO, a selective eIF2 alpha phosphatase inhibitor which protects cells from ER stress and inhibits apoptosis, 1 mg/kg for 3 days post‐extraction), observation for 7 days.

Techniques: Concentration Assay, Expressing, CCK-8 Assay, Western Blot, Control, Colony Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: Endoplasmic reticulum stress remodels alveolar bone formation after tooth extraction

doi: 10.1111/jcmm.15753

Figure Lengend Snippet: Suppressing ER stress could accelerate bone formation. A, Three‐dimensional micro‐CT reconstruction images of the vehicle control (VC) and salubrinal (SAL) groups. B, In the first molar area, bone formation was greater in the SAL group in the sagittal view (alveolar ridge median sagittal plane), occlusal view (root bifurcation surface) and coronal view (maxillary first molar mesial root). C, Micro‐CT assessment of the mesial roots of the maxillary first molars extraction sockets. SAL group BV/TV, Tb.N and Tb.Th were all increased, but Tb.Sp was decreased compared with the VC group. Data were presented as mean ± SD. The asterisks (*) represent P < .05 (n = 3)

Article Snippet: Experimental tooth extraction was performed on 48 Sprague‐Dawley rats which randomly dived into four groups (n = 12/group): (a) M1 extraction, observation for 7 days; (b) M1 extraction, observation for 14 days; (c) M1 extraction + PBS (3 days post‐extraction), observation for 7 days; (d) M1 extraction + Salubrinal (T3045; Target Mol, resolved in DMSO, a selective eIF2 alpha phosphatase inhibitor which protects cells from ER stress and inhibits apoptosis, 1 mg/kg for 3 days post‐extraction), observation for 7 days.

Techniques: Micro-CT, Control, Extraction

Journal: Journal of Cellular and Molecular Medicine

Article Title: Endoplasmic reticulum stress remodels alveolar bone formation after tooth extraction

doi: 10.1111/jcmm.15753

Figure Lengend Snippet: Suppressing ER stress could protect osteogenesis in vivo. A, Representative photomicrographs of tooth extraction sockets in the salubrinal (SAL) and vehicle control (VC) groups. The blue dotted line indicates the tooth extraction socket, and the star indicates irregular woven bone. SAL group significantly increased the number of collagen fibres. Top: HE staining, 50×; bottom: Masson's tricolour staining, 50×, blue fibres in connective tissue represent collagen. B, Representative photomicrographs of ALP staining (200×) in tooth extraction sockets in the SAL and control groups. C, The percentage change in body weight. Body weight increased after using salubrinal. D‐E, Representative Western blots of the protein expression levels of eIF2α, p‐eIF2α and Runx2 in the bone tissue around the tooth extraction wounds with different drug interventions. p‐eIF2α/eIF2α and Runx2/β‐actin protein expression was higher after salubrinal use. F, Transmission electron microscope images showing the ultrastructural changes of the rough ER (red square) and the formation of collagen fibres (blue arrows) in osteoblast around the sockets in the SAL and control groups (top: PBS, bottom: SAL) Left scale bar = 5 μm, middle and right scale bar = 1 μm. Data were presented as the ratio with β‐actin and mean ± SD. The asterisks (*) represent P < .05(n = 9)

Article Snippet: Experimental tooth extraction was performed on 48 Sprague‐Dawley rats which randomly dived into four groups (n = 12/group): (a) M1 extraction, observation for 7 days; (b) M1 extraction, observation for 14 days; (c) M1 extraction + PBS (3 days post‐extraction), observation for 7 days; (d) M1 extraction + Salubrinal (T3045; Target Mol, resolved in DMSO, a selective eIF2 alpha phosphatase inhibitor which protects cells from ER stress and inhibits apoptosis, 1 mg/kg for 3 days post‐extraction), observation for 7 days.

Techniques: In Vivo, Extraction, Control, H&E Stain, Staining, Tissue, Western Blot, Expressing, Transmission Assay, Microscopy, Histology, Transmission Electron Microscopy

Journal: Journal of Cellular and Molecular Medicine

Article Title: Endoplasmic reticulum stress remodels alveolar bone formation after tooth extraction

doi: 10.1111/jcmm.15753

Figure Lengend Snippet: Proposed mechanism of action. ER stress occurred in the alveolar bone after tooth extraction, and excessive ER stress can cause osteoblast apoptosis. Salubrinal promoted osteoblastogenesis and bone healing through regulation of eIF2α

Article Snippet: Experimental tooth extraction was performed on 48 Sprague‐Dawley rats which randomly dived into four groups (n = 12/group): (a) M1 extraction, observation for 7 days; (b) M1 extraction, observation for 14 days; (c) M1 extraction + PBS (3 days post‐extraction), observation for 7 days; (d) M1 extraction + Salubrinal (T3045; Target Mol, resolved in DMSO, a selective eIF2 alpha phosphatase inhibitor which protects cells from ER stress and inhibits apoptosis, 1 mg/kg for 3 days post‐extraction), observation for 7 days.

Techniques: Extraction

Journal: Development (Cambridge, England)

Article Title: Mitochondrial dysfunction in oocytes of obese mothers: transmission to offspring and reversal by pharmacological endoplasmic reticulum stress inhibitors.

doi: 10.1242/dev.114850

Figure Lengend Snippet: Fig. 2. Ovulation rate declines with increasing body weight but is improved by treatment with ER stress inhibitors. (A) Hematoxylin and Eosin-stained ovary sections from 14-week-old gonadotropin-treated lean (+/+ or +/bbb) and obese Blobby (bbb/bbb) mice. Arrows indicate unruptured follicles, which are prevalent in ovaries from obese mice. Representative examples from n=4 mice per genotype are shown. Panels a-c are higher magnifications of the indicated follicles of the Blobby mouse. (B) The number of ovulated oocytes in 14-week-old lean and obese mice treated with saline vehicle (Veh.), salubrinal (Sal) or BGP-15 i.p. once per day for 4 days. Values are means+s.e.m. (lean mice+Veh, n=19; lean+Sal, n=5; lean+BGP-15, n=10; obese mice+Veh, n=22; obese+Sal, n=16; obese+BGP-15, n=14). Different letters indicate significant differences calculated by one-way ANOVA, Tukey’s post hoc test; P<0.0001. Right panel: the number of ovulated oocytes in oviducts is not reduced in bbb/bbb mice prior to the onset of obesity, at 6 weeks of age (+/+; +/bbb: n=11; bbb/bbb n=4; Student’s t-test).

Article Snippet: Salubrinal (Invitrogen) was injected i.p. at 1 mg/kg of bodyweight (Sokka et al., 2007) in approximately 0.1 ml saline daily for four consecutive days starting the day prior to treatment with eCG.

Techniques: Staining, Saline

Journal: Development (Cambridge, England)

Article Title: Mitochondrial dysfunction in oocytes of obese mothers: transmission to offspring and reversal by pharmacological endoplasmic reticulum stress inhibitors.

doi: 10.1242/dev.114850

Figure Lengend Snippet: Fig. 5. Treatment with salubrinal or BGP-15 induces mtDNA replication and normalizes mitochondrial membrane potential (ΔΨm) and autophagy in oocytes from obese mice. Obese mice (bbb/bbb) or lean littermates (+/+ or +/bbb) were treated with vehicle (Veh), salubrinal (Sal) or BGP-15 i.p. once daily for 4 days, and oocytes were collected from the oviducts. (A) Live oocytes stained with JC-1, where red fluorescence indicates high ΔΨm, and green indicates low ΔΨm. (B) Ratio of red to green fluorescence, an indicator of mitochondrial activity. Data are presented as means+s.e.m., n=6-30 oocytes from four mice per group. (C) Live oocytes were assessed for autophagic vacuoles, visualized as green fluorescence. Hoechst 33342 (blue). (D) Autophagy levels were quantified as the sum total of green fluorescence within each oocyte. Data are presented as means+s.e.m., n=8-10 oocytes from three mice per group. (E) mtDNA copy number in individual oocytes collected from treated mice. Data are presented as means+s.e.m.; n=19-38 oocytes from six mice per group over three independent experiments. (F) Representative examples of oocytes from three or four mice per treatment group immunostained for TFAM or DRP1 (green) and Hoechst 33342 (blue). Different letters indicate significant differences calculated by one-way ANOVA, Tukey’s post hoc test, P<0.0001.

Article Snippet: Salubrinal (Invitrogen) was injected i.p. at 1 mg/kg of bodyweight (Sokka et al., 2007) in approximately 0.1 ml saline daily for four consecutive days starting the day prior to treatment with eCG.

Techniques: Membrane, Staining, Fluorescence, Activity Assay

Journal: Development (Cambridge, England)

Article Title: Mitochondrial dysfunction in oocytes of obese mothers: transmission to offspring and reversal by pharmacological endoplasmic reticulum stress inhibitors.

doi: 10.1242/dev.114850

Figure Lengend Snippet: Fig. 6. Oocytes from obese Blobby mice exhibit impaired embryo development that is restored by treatment with salubrinal or BGP-15. Obese Blobby (bbb/bbb) mice or lean littermates (+/+ or +/bbb) at 14 weeks of age (except where indicated) were treated with vehicle (Veh; saline), salubrinal (Sal) or BGP-15 daily for 4 days. Ovulated oocytes were collected from oviducts 16 h following hCG injection and fertilized in vitro. (A) At 4 h after in vitro fertilization, many degenerate oocytes (arrows) from untreated obese mice were observed. (B) The percentage of viable oocytes presented as means+s.e.m. Embryo development was assessed on day 2 and day 5 following IVF and is presented as the mean percentage of embryos exhibiting appropriate (‘on-time’) development+s.e.m. (C) Two-cell embryos developed from viable oocytes by day 2. (D) Blastocysts developed from two-cell embryos by day 5. Lean, n=24; lean+Sal, n=7; lean+BGP- 15, n=6; Blobby, n=15; Blobby+Sal, n=6; Blobby+BGP-15, n=7 pools of oocytes from animals within same group. [C,D, right-hand panels are identical assessments of oocytes from 6-week-old (non-obese) mice. +/+, n=13; Blobby, n=10.] (E) Blastocysts were collected at day 5 and the mtDNA copy number of individual blastocysts was measured by using quantitative PCR. Data are expressed as mean copy number+s.e.m.; lean, n=22 blastocysts; lean+Sal, n=10 blastocysts; lean+BGP-15, n=7 blastocysts; Blobby, n=14 blastocysts; Blobby+Sal, n=10 blastocysts; Blobby+BGP-15, n=7 blastocysts collected from three independent IVF experiments. Different letters indicate significant differences by one-way ANOVA, Tukey’s post hoc test; P<0.0001.

Article Snippet: Salubrinal (Invitrogen) was injected i.p. at 1 mg/kg of bodyweight (Sokka et al., 2007) in approximately 0.1 ml saline daily for four consecutive days starting the day prior to treatment with eCG.

Techniques: Saline, Injection, In Vitro, Real-time Polymerase Chain Reaction

Journal: Development (Cambridge, England)

Article Title: Mitochondrial dysfunction in oocytes of obese mothers: transmission to offspring and reversal by pharmacological endoplasmic reticulum stress inhibitors.

doi: 10.1242/dev.114850

Figure Lengend Snippet: Fig. 7. Altered fetal outcomes of oocytes from obese Blobby mice are alleviated after treatment with salubrinal or BGP-15. Obese mice (bbb/bbb) and lean littermates (+/+) were treated with vehicle (Veh; saline), salubrinal (Sal) or BGP-15 for 4 days, followed by the isolation of oocytes from oviducts and fertilization by using IVF. Blastocysts were collected at day 5 of culture and transferred to non-obese pseudo-pregnant recipient mice with six blastocysts per uterine horn. On embryonic day 14.5, fetuses were collected and weighed (A). Liver (B), heart (C) and kidney (D) were dissected and the relative mtDNA copy number normalized to β-actin nuclear DNA was determined. Data are presented as means+s.e.m.; lean+Veh, n=18 fetuses from six surrogates; Blobby+Veh, n=20 fetuses from six surrogates; Blobby+Sal, n=22 fetuses from six surrogates; Blobby+BGP-15, n=15 fetuses from four surrogates. (E) Liver mtDNA samples (n=4 fetuses from each treatment group) were analyzed by using next generation sequencing, and rearrangements were identified. Different letters indicate significant differences calculated by one-way ANOVA, Tukey’s post hoc test; P<0.01.

Article Snippet: Salubrinal (Invitrogen) was injected i.p. at 1 mg/kg of bodyweight (Sokka et al., 2007) in approximately 0.1 ml saline daily for four consecutive days starting the day prior to treatment with eCG.

Techniques: Saline, Isolation, Next-Generation Sequencing

Journal: Oncotarget

Article Title: Adaptive metabolic rewiring to chronic SFK inhibition

doi: 10.18632/oncotarget.8146

Figure Lengend Snippet: A . Changes in the redox state of parental (BT20 blue line) or resistant (BT20R black line) cells after twelve hours after dasatinib treatment with the (respective IC 50 ) is shown with respect to vehicle treated cells (in red). A representative experiment of three is shown. B . Conversion of LC3-1 to LC3-2 in parental BT20 or resistant BT20R cells was assessed by western blot analysis 0 to 6 hours after treatment with dasatinib IC 50 . Equal protein loading (50μg) was confirmed by re probing with an α-actin antibody. C . Changes in GPR78 expression levels and eIF2 phosphorylation in parental BT20 or resistant BT20R cells was assessed by western blot analysis 0 to 6 hours after treatment with dasatinib (respective IC 50 ). Equal protein loading (50μg) was confirmed by re probing with an α-actin antibody. D . XBP-1 splicing was assessed in BT20 and BT20R cells twelve hours after treatment with vehicle (CTL), thapsigargin (THAP, 500 nM positive control) or dasatinib treatment (DAS, IC 50 concentration). The arrows indicate XBP-1 splice variants. E . BT20R cells were treated with dasatinib (0-50μM) in combination with vehicle, non-toxic concentrations of drugs affecting the ER stress response (i.e. the inhibitor salubrinal or the inducer thapsigargin) or inhibitors of early (3-MA) or later (CQ) autophagy as indicated. Five days after treatment dasatinib IC 50 s in the combinations were obtained utilizing the SRB assay and used to calculate the sensitization values (I value) for each combination. The y-axis represent the mean I values (n=5) plus minus the standard error. The *** indicate a significant difference between the I values with respect to 1 (i.e. dasatinib in combination with vehicle) using ANOVA with p<0.001 followed by t-test. F . BT20 R cells were treated with vehicle (CTL), CQ (1.5 μM), dasatinib IC 50 (1μM) or the combination of CQ and dasatinib for seventy two hours. Annexin V expression was assessed by flow cytometry as described in materials and methods. Increase in cell death by the combination of dasatinib plus CQ respect to the drugs alone was assessed by one way ANOVA (***p<0.001). The bars graph represent the mean values of Annexin V positive cells plus minus the standard error (n=4).

Article Snippet: Dasatinib, buparsilib, (Selleck chemicals), salubrinal, SB431542, PI3-103 (Tocris Biosciences).

Techniques: Western Blot, Expressing, Phospho-proteomics, Positive Control, Concentration Assay, Sulforhodamine B Assay, Flow Cytometry