Journal: Cell Death Discovery

Article Title: Proinflammatory cytokines sensitise mesenchymal stromal cells to apoptosis

doi: 10.1038/s41420-025-02412-0

Figure Lengend Snippet: A Quantification of Annexin V + cells in Jurkat T lymphoma cells treated with increasing concentrations of anti-FAS antibody (aFAS) for 24 h ( n = 3) and representative Annexin V/PI staining following stimulation with 1 μg/ml of anti-Fas antibody for 24 h. B Quantification of Annexin V + cells in human BM-MSCs treated with increasing concentrations of anti-FAS antibody for 24 h ( n = 3) and representative Annexin V/PI staining following stimulation with 10 μg/ml of anti-Fas antibody for 24 h. C Representative flow cytometric analysis of FAS expression in one of three human BM-MSCs in comparison to peripheral blood mononuclear cells (PBMC). D Quantification of Annexin V + cells in Jurkat cells treated with increasing concentrations of FcFASL for 24 h ( n = 3). E – G Quantification of Annexin V + cells in human BM-MSCs ( E ), mouse BM-MSCs ( F ), and primary MEFs and SV40-immortalised MEFs ( G ) treated with increasing concentrations of FcFASL for 24 h with or without pre-treatment with 50 μM zVAD-FMK for 30 min ( n = 3). H Quantification of Annexin V + cells in human BM-MSCs treated with 10 μg/ml anti-FAS antibody for 24 h in the presence or absence of 500 nM SMAC mimetic (Compound A) with or without pre-treatment with 50 μM zVAD-FMK for 30 min ( n = 3). I Representative Annexin V/PI staining in human BM-MSCs treated with 100 ng/ml TNF for 24 h, and quantification of Annexin V + cells in human BM-MSCs treated with increasing concentrations of TNF ( n = 3). J Quantification of Annexin V + cells in human BM-MSCs (left panel) and mouse BMDMs (right panel) treated with 100 ng/ml TNF, or 25 μg/ml poly(I:C), or 25 μg/ml LPS for 24 h in the presence or absence of 500 nM SMAC mimetic, with or without pre-treatment with the pan-caspase inhibitors Q-VD-OPh or zVAD-FMK for 30 min ( n = 3). K Quantification of Annexin V + cells in human BM-MSCs treated with 100 ng/ml TNF, or 100 μg/ml poly(I:C), or 50 μg/ml LPS for 24 h in the presence or absence of 500 nM SMAC mimetic with or without pre-treatment with increasing concentrations of Q-VD-OPh or zVAD-FMK for 30 min ( n = 3). Data expressed as the mean ± S.E.M. and representative of at least two independent experiments, p values by one-way ANOVA with Tukey’s post hoc test.

Article Snippet: Human adipose-derived MSCs were either purchased from ScienCell or isolated from subcutaneous adipose tissue obtained from outpatient liposuction procedures (Monash human ethics approval #2007/1798; performed with informed patient consent).

Techniques: Staining, Expressing, Comparison

Journal: Cell Death Discovery

Article Title: Proinflammatory cytokines sensitise mesenchymal stromal cells to apoptosis

doi: 10.1038/s41420-025-02412-0

Figure Lengend Snippet: A , B Quantification of Annexin V + cells in Jurkat cells ( A ) and human BM-MSCs ( B ) treated with increasing concentrations of the BH3 mimetic drugs ABT199 (BCL-2 inhibitor, iBCL2), A1331852 (BCL-XL inhibitor, iBCLxL) for 3 h. C Representative Annexin V/PI staining in human BM-MSCs treated with increasing concentrations of BH3 mimetic drugs, and BAK/BAX-deficient (BKX) MSCs treated with 1.25 μM BH3 mimetic drugs for 3 h. D Quantification of Annexin V + cells in human MSCs ( D ) and mouse MSCs ( E ) treated with increasing concentrations of BH3 mimetic drugs for 2 h ( n = 3). F , G Quantification of Annexin V + cells at various time points following treatment of human MSCs with 1.25 μM BH3 mimetic drugs ( F ) and mouse MSCs, MEFs and BAK/BAX deficient MEFs treated with 10 μM BH3 mimetic drugs ( G ) ( n = 3). H Quantification of Annexin V + cells in human MSCs treated with increasing concentrations of various BH3 mimetic drugs combinations for 24 h ( n = 3). I Quantification of Annexin V + cells in mouse MSCs treated with various BH3 mimetic drugs combinations at 10 μM for 24 h ( n = 3). Data expressed as the mean ± S.E.M. and representative of at least two independent experiments.

Article Snippet: Human adipose-derived MSCs were either purchased from ScienCell or isolated from subcutaneous adipose tissue obtained from outpatient liposuction procedures (Monash human ethics approval #2007/1798; performed with informed patient consent).

Techniques: Staining

Journal: Cell Death Discovery

Article Title: Proinflammatory cytokines sensitise mesenchymal stromal cells to apoptosis

doi: 10.1038/s41420-025-02412-0

Figure Lengend Snippet: A Quantification of live cells (Annexin V − ) in human AD-MSCs, UC-MSCs, and BM-MSCs treated with increasing concentrations of BH3 mimetic drugs for 2 h ( n = 3 donors per tissue type). B Relative expression level of the anti-apoptotic genes BCL-XL , BCL-2 , and MCL-1 in cultured AD-MSCs, UC-MSCs, and BM-MSCs ( n = 3 donors per tissue type). Data expressed as the mean ± S.E.M. pooled from two independent experiments, p values by one-way ANOVA with Tukey’s post hoc test.

Article Snippet: Human adipose-derived MSCs were either purchased from ScienCell or isolated from subcutaneous adipose tissue obtained from outpatient liposuction procedures (Monash human ethics approval #2007/1798; performed with informed patient consent).

Techniques: Expressing, Cell Culture

Journal: Cell Death Discovery

Article Title: Proinflammatory cytokines sensitise mesenchymal stromal cells to apoptosis

doi: 10.1038/s41420-025-02412-0

Figure Lengend Snippet: A , B Representative Annexin V/PI staining in unprimed BM-MSCs and BM-MSCs primed with single (TNF or IFN-γ) or dual (TNF and IFN-γ) cytokines at the indicated concentrations for 24 h prior to treatment with vehicle control (DMSO) ( A ) or 1.25 μM BH3 mimetic drugs ( B ) for 2.5 h. C Representative Annexin V/PI staining of two additional BM-MSC donors primed with 10 ng/ml TNF and 100 ng/ml IFN-γ for 24 h prior to treatment with BH3 mimetic drugs for 2.5 h. D Quantification of the proportion of live Annexin V − PI − cells (left panel), early Annexin V + PI − (middle panel) and late Annexin V + PI + apoptotic cells (right panel) in unprimed and primed BM-MSCs from three donors treated with increasing concentrations of BH3 mimetic drugs for 2.5 h ( n = 3). E Representative Annexin V/PI staining in unprimed and primed BM-MSCs treated with 0.125 μM BH3 mimetic drugs for 2.5 h. F Quantification of live (Annexin V − PI − ), early apoptotic (Annexin V + PI - ) and late apoptotic (Annexin V + PI + ) cells from three BM-MSC donors treated with 0.125 μM BH3 mimetic drugs (as shown in E ) for 2.5 h ( n = 3). G Representative Annexin V/PI staining in unprimed and primed MSCs treated with 1.25 μM BH3 mimetic drugs for 30 min. H Representative Annexin V/TO-PRO-3 staining and gating of Annexin V + TO-PRO-3 hi late apoptotic cells in unprimed and primed BM-MSCs treated with BH3 mimetic drugs for 30 min. I , J Representative histograms showing the proportion of TO-PRO-3 int cells after exclusion of Annexin V + TO-PRO-3 hi late apoptotic (as shown in H ) in unprimed and primed BM-MSCs treated with BH3 mimetic drugs (blue histograms) or vehicle (grey histograms) for 30 min. Data expressed as the mean ± S.E.M. and representative of two independent experiments, p values by two-way ANOVA with Dunnett’s multiple comparison test.

Article Snippet: Human adipose-derived MSCs were either purchased from ScienCell or isolated from subcutaneous adipose tissue obtained from outpatient liposuction procedures (Monash human ethics approval #2007/1798; performed with informed patient consent).

Techniques: Staining, Control, Comparison

Journal: Cell Death Discovery

Article Title: Proinflammatory cytokines sensitise mesenchymal stromal cells to apoptosis

doi: 10.1038/s41420-025-02412-0

Figure Lengend Snippet: A Live cell imaging of parental MSCs (top panels; Video ) and apoptosis-deficient BKX-MSCs (bottom panels; Video ) following apoptosis induction with BH3 mimetic drugs. B Live cell imaging of untreated human bone marrow MSCs (top panels; Video ) and BH3 mimetic drug-treated MSCs (bottom panels; Video ) stained with Annexin V, showing formation of apoptotic bodies (arrows). C Representative Annexin V staining (left panel) and quantification of the proportion of apoptotic bodies (right panel) in human MSCs treated with BH3 mimetic drugs ( n = 3). Data are expressed as the mean ± S.E.M and representative of three independent experiments. D Schematic for detection of MSCs and apoptotic bodies within the lungs at various time points after intravenous injection into mice. E Representative staining for CTV and CD45 to detect MSCs in digested lung tissue harvested 10 min post intravenous MSC injection F Representative staining for active caspase 3 within the CTV + cell population, used to identify apoptotic MSCs and apoptotic bodies within digested lung tissue following intravenous injection of CTV-labelled parental MSCs (top panel) or BKX-MSCs (bottom panel). G Quantification of apoptotic bodies detected in the lungs of mice (as shown in E ). Data represent the mean ± S.E.M. of n = 3 mice, p values by one-way ANOVA with Tukey’s post hoc test. Panel D was created with BioRender.com.

Article Snippet: Human adipose-derived MSCs were either purchased from ScienCell or isolated from subcutaneous adipose tissue obtained from outpatient liposuction procedures (Monash human ethics approval #2007/1798; performed with informed patient consent).

Techniques: Live Cell Imaging, Staining, Injection

Journal: Cell Death Discovery

Article Title: Proinflammatory cytokines sensitise mesenchymal stromal cells to apoptosis

doi: 10.1038/s41420-025-02412-0

Figure Lengend Snippet: A Schematic of how the survival of unprimed and primed MSCs was analysed within mouse lung tissue. B Representative staining for CTV and hCD73 to detect MSCs in digested lung tissue harvested 30 min post intravenous MSC injection. C Gating strategy used to identify live MSCs, apoptotic MSCs and apoptotic bodies, based on pooled samples of viable MSCs and BH3-mimetic drug treated MSCs stained with FLICA to detect active caspase 3/7. D Representative FLICA staining within the CTV + cell population used to detect live MSCs, apoptotic MSCs and apoptotic bodies in digested lung tissue. E Quantification of the number of live MSCs, apoptotic MSCs and apoptotic bodies within the lungs (as shown in D ). F Quantification of the proportion of CD45 − (left panel) and CD45 + cells (right panel) within the CTV + FLICA + apoptotic MSC gate. G , H Detection of human CD73 + MSCs within ex vivo cultured lung cells. Digested lung cells from untreated mice, or mice that received intravenous unprimed MSCs or primed MSCs were cultured for six days and the number of human MSCs was quantified. G Representative staining of human CD73 + MSCs within the CD45 − population of cultured lung cells. H Quantification of the number (left panel) and proportion (right panel) of human CD73 + MSCs in cultured lung cells six days after plating. Data represent the mean ± S.E.M. of n = 3 mice, unpaired T -test; ** p ≤ 0.01. Panel A was created with BioRender.com.

Article Snippet: Human adipose-derived MSCs were either purchased from ScienCell or isolated from subcutaneous adipose tissue obtained from outpatient liposuction procedures (Monash human ethics approval #2007/1798; performed with informed patient consent).

Techniques: Staining, Injection, Ex Vivo, Cell Culture