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pancreatic β-cell line mouse insulinoma 6 (min6) (National Centre for Cell Science)

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National Centre for Cell Science pancreatic β-cell line mouse insulinoma 6 (min6)
Pancreatic β Cell Line Mouse Insulinoma 6 (Min6), supplied by National Centre for Cell Science, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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pancreatic β-cell line mouse insulinoma 6 (min6) - by Bioz Stars, 2026-10

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Article Title: Antihyperglycemic and Insulin Secretagogue Activities of Abrus precatorius Leaf Extract
Article Snippet: MIN6-β cells were procured from the National Centre for Cell Science, Pune, India.

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Taurine supplement alleviates doxorubicin‐induced β‐cell inflammation and senescence. MIN6 cells were pre‐treated with 100 μM taurine for 24 h, followed by 200 nM doxorubicin (DOXO) treatment for 24 h. Cells were cultured in FBS‐free medium to avoid possible contamination of taurine. (A) QPCR analysis of the genes related to inflammation, senescence, and apoptosis in each group of doxorubicin‐induced senescence model. ( n = 3) Relative mRNA levels were normalized to β‐actin. (B) Immunoblotting analysis of p53 and p21 and densitometric quantification. ( n = 3). (C) Immunofluorescence staining of DNA damage marker γ–H2AX in each group (scale bar: 100 μm). ( n = 5). (D) FACS analysis of β‐gal+ PI‐(senescent) and PI+ (dead) MIN6 cells. All results are presented as mean ± SEM. Significance was determined using two‐way ANOVA with Tukey correction. * p < 0.05, ** p < 0.01, *** p < 0.001.

Journal: Journal of Diabetes

Article Title: Taurine Alleviates Pancreatic β‐Cell Senescence by Inhibition of p53 Pathway

doi: 10.1111/1753-0407.70100

Figure Lengend Snippet: Taurine supplement alleviates doxorubicin‐induced β‐cell inflammation and senescence. MIN6 cells were pre‐treated with 100 μM taurine for 24 h, followed by 200 nM doxorubicin (DOXO) treatment for 24 h. Cells were cultured in FBS‐free medium to avoid possible contamination of taurine. (A) QPCR analysis of the genes related to inflammation, senescence, and apoptosis in each group of doxorubicin‐induced senescence model. ( n = 3) Relative mRNA levels were normalized to β‐actin. (B) Immunoblotting analysis of p53 and p21 and densitometric quantification. ( n = 3). (C) Immunofluorescence staining of DNA damage marker γ–H2AX in each group (scale bar: 100 μm). ( n = 5). (D) FACS analysis of β‐gal+ PI‐(senescent) and PI+ (dead) MIN6 cells. All results are presented as mean ± SEM. Significance was determined using two‐way ANOVA with Tukey correction. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Mouse pancreatic β‐cell line MIN6 (AddexBio Technologies, Cat#C0018008) and rat insulinoma cell INS‐1E (AddexBio Technologies, Cat#C0018009) were cultured in DMEM (Gibco, Cat#12800082) or RPMI 1640 supplemented with 15% FBS, 1% penicillin–streptomycin, and 50 μM β‐mercaptoethanol.

Techniques: Cell Culture, Western Blot, Immunofluorescence, Staining, Marker

Taurine supplementation alleviates TNF‐α‐induced β‐cell inflammation and senescence. MIN6 cells were pre‐treated with 100 μM taurine for 24 h, followed by 20 ng/mL TNF‐α treatment for 24 h. Cells were cultured in FBS‐free medium to avoid possible contamination of taurine. (A) QPCR analysis of the genes related to senescence in each group of TNF‐α‐induced senescence model. ( n = 4). Relative mRNA levels were normalized to β‐actin. (B) QPCR analysis of the genes related to inflammation and apoptosis in each group. ( n = 4). Relative mRNA levels were normalized to β‐actin. (C) Immunoblotting analysis of p53 and p21 in each group and densitometric quantification. ( n = 3). All results are presented as mean ± SEM. Significance was determined using two‐way ANOVA with Tukey correction. * p < 0.05, ** p < 0.01, *** p < 0.001.

Journal: Journal of Diabetes

Article Title: Taurine Alleviates Pancreatic β‐Cell Senescence by Inhibition of p53 Pathway

doi: 10.1111/1753-0407.70100

Figure Lengend Snippet: Taurine supplementation alleviates TNF‐α‐induced β‐cell inflammation and senescence. MIN6 cells were pre‐treated with 100 μM taurine for 24 h, followed by 20 ng/mL TNF‐α treatment for 24 h. Cells were cultured in FBS‐free medium to avoid possible contamination of taurine. (A) QPCR analysis of the genes related to senescence in each group of TNF‐α‐induced senescence model. ( n = 4). Relative mRNA levels were normalized to β‐actin. (B) QPCR analysis of the genes related to inflammation and apoptosis in each group. ( n = 4). Relative mRNA levels were normalized to β‐actin. (C) Immunoblotting analysis of p53 and p21 in each group and densitometric quantification. ( n = 3). All results are presented as mean ± SEM. Significance was determined using two‐way ANOVA with Tukey correction. * p < 0.05, ** p < 0.01, *** p < 0.001.

Techniques: Cell Culture, Western Blot

β‐cells acquire taurine through Slc6a6‐mediated uptake. (A) QPCR analysis of taurine biosynthesis related genes and its transporter Slc6a6 in MIN6 cells and mouse hepatocytes. The results are presented as relative levels over respective gene expression in mouse hepatocytes. ( n = 4). (B, C) MIN6 cells were transfected with siRNA against Scramble or Slc6a6 for 24 h, followed by treatment with taurine (100 μM) or vehicle for 24 h. (B) Immunoblotting analysis of SLC6A6 protein level in each group. ( n = 3). (C) Intracellular taurine levels in the transfected MIN6 cells. ( n = 4). (D) MIN6 cells were pre‐treated with non‐FBS culture medium. The cells were then treated with taurine (100 μM) for 24 h, followed by treatment with SLC6A6 inhibitor (SLC6A6i) (100 μM) or vehicle for 30 min. Intracellular taurine concentration was measured by LC–MS/MS. ( n = 3). All results are presented as mean ± SEM. Significance was determined using two‐tailed independent student's t ‐test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Journal: Journal of Diabetes

Article Title: Taurine Alleviates Pancreatic β‐Cell Senescence by Inhibition of p53 Pathway

doi: 10.1111/1753-0407.70100

Figure Lengend Snippet: β‐cells acquire taurine through Slc6a6‐mediated uptake. (A) QPCR analysis of taurine biosynthesis related genes and its transporter Slc6a6 in MIN6 cells and mouse hepatocytes. The results are presented as relative levels over respective gene expression in mouse hepatocytes. ( n = 4). (B, C) MIN6 cells were transfected with siRNA against Scramble or Slc6a6 for 24 h, followed by treatment with taurine (100 μM) or vehicle for 24 h. (B) Immunoblotting analysis of SLC6A6 protein level in each group. ( n = 3). (C) Intracellular taurine levels in the transfected MIN6 cells. ( n = 4). (D) MIN6 cells were pre‐treated with non‐FBS culture medium. The cells were then treated with taurine (100 μM) for 24 h, followed by treatment with SLC6A6 inhibitor (SLC6A6i) (100 μM) or vehicle for 30 min. Intracellular taurine concentration was measured by LC–MS/MS. ( n = 3). All results are presented as mean ± SEM. Significance was determined using two‐tailed independent student's t ‐test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Techniques: Gene Expression, Transfection, Western Blot, Concentration Assay, Liquid Chromatography with Mass Spectroscopy, Two Tailed Test

The protective effects of taurine against β‐cell senescence depend on its transporter SLC6A6. (A, B) MIN6 cells were pre‐treated with the SLC6A6 inhibitor (SLC6A6i) (100 μM) or vehicle for 30 min, followed by treatment with taurine (100 μM) and doxorubicin (200 nM) or vehicle for 24 h in non‐FBS culture medium. The intracellular taurine concentration was then measured by LC–MS/MS. ( n = 3). (B) Immunoblotting analysis of p53 and p21 in each group. (C–F) MIN6 cells were pre‐treated with doxorubicin (200 nM). The cells were then transfected with siRNA against Scramble or Slc6a6 for 24 h, followed by treatment with taurine (100 μM) or vehicle for 24 h. (C) Immunoblotting analysis of SLC6A6, p53, and p21 in each group. ( n = 3). (D) QPCR analysis of gene expressions related to senescence in each group ( n = 4). (E) QPCR analysis of the genes related to β‐cell specific SASP in each group. ( n = 4). (F) QPCR analysis of genes related to inflammation and apoptosis. ( n = 4). All results are presented as mean ± SEM. Significance was determined using two‐way ANOVA with Tukey correction. * p < 0.05, ** p < 0.01, *** p < 0.001.

Journal: Journal of Diabetes

Article Title: Taurine Alleviates Pancreatic β‐Cell Senescence by Inhibition of p53 Pathway

doi: 10.1111/1753-0407.70100

Figure Lengend Snippet: The protective effects of taurine against β‐cell senescence depend on its transporter SLC6A6. (A, B) MIN6 cells were pre‐treated with the SLC6A6 inhibitor (SLC6A6i) (100 μM) or vehicle for 30 min, followed by treatment with taurine (100 μM) and doxorubicin (200 nM) or vehicle for 24 h in non‐FBS culture medium. The intracellular taurine concentration was then measured by LC–MS/MS. ( n = 3). (B) Immunoblotting analysis of p53 and p21 in each group. (C–F) MIN6 cells were pre‐treated with doxorubicin (200 nM). The cells were then transfected with siRNA against Scramble or Slc6a6 for 24 h, followed by treatment with taurine (100 μM) or vehicle for 24 h. (C) Immunoblotting analysis of SLC6A6, p53, and p21 in each group. ( n = 3). (D) QPCR analysis of gene expressions related to senescence in each group ( n = 4). (E) QPCR analysis of the genes related to β‐cell specific SASP in each group. ( n = 4). (F) QPCR analysis of genes related to inflammation and apoptosis. ( n = 4). All results are presented as mean ± SEM. Significance was determined using two‐way ANOVA with Tukey correction. * p < 0.05, ** p < 0.01, *** p < 0.001.

Techniques: Concentration Assay, Liquid Chromatography with Mass Spectroscopy, Western Blot, Transfection

Taurine mitigates senescence, inflammation, and oxidative stress via a p53‐dependent pathway while preserving mitochondrial function independently of p53. (A–C) MIN6 cells were pre‐treated with DOXO (200 nM). The cells were then transfected with siRNA against scramble or p53 for 24 h, followed by treatment with taurine (100 μM) or vehicle for 24 h. Cells were cultured in FBS‐free medium to avoid possible contamination of taurine. (A) QPCR analysis of the genes related to senescence and inflammation in each group. ( n = 4) Relative mRNA levels were normalized to β‐actin. (B) Cellular content of malondialdehyde (MDA) in each group. ( n = 4). (C) Mitochondrial membrane potential was measured using TMRE mitochondrial membrane potential assay. ( n = 7). All results are presented as mean ± SEM. Significance was determined using two‐way ANOVA with Tukey correction. * p < 0.05, ** p < 0.005, *** p < 0.001.

Journal: Journal of Diabetes

Article Title: Taurine Alleviates Pancreatic β‐Cell Senescence by Inhibition of p53 Pathway

doi: 10.1111/1753-0407.70100

Figure Lengend Snippet: Taurine mitigates senescence, inflammation, and oxidative stress via a p53‐dependent pathway while preserving mitochondrial function independently of p53. (A–C) MIN6 cells were pre‐treated with DOXO (200 nM). The cells were then transfected with siRNA against scramble or p53 for 24 h, followed by treatment with taurine (100 μM) or vehicle for 24 h. Cells were cultured in FBS‐free medium to avoid possible contamination of taurine. (A) QPCR analysis of the genes related to senescence and inflammation in each group. ( n = 4) Relative mRNA levels were normalized to β‐actin. (B) Cellular content of malondialdehyde (MDA) in each group. ( n = 4). (C) Mitochondrial membrane potential was measured using TMRE mitochondrial membrane potential assay. ( n = 7). All results are presented as mean ± SEM. Significance was determined using two‐way ANOVA with Tukey correction. * p < 0.05, ** p < 0.005, *** p < 0.001.

Techniques: Preserving, Transfection, Cell Culture, Membrane

Identification of Taurine‐CDKN2AIP binding in pancreatic β cells. (A, B) Limited proteolysis‐mass spectrometry (LiP‐MS) was used to screen for taurine interacting proteins in the INS1E β‐cell proteome. Heatmap shows potential taurine binding targets identified by LiP‐MS. Vehicle: N = 3. Taurine: N = 3. (B) p53 pathway related proteins levels between two groups and their binding scores with taurine. (C) Three‐dimensional diagram of the binding modes between human CDKN2AIP and taurine. Taurine potentially binds to CDKN2AIP via residues PRO484, LEU485, LYS486. (D) DARTS analysis using MIN6 cell lysates incubated with taurine. (E) DARTS analysis using INS1E cell lysates incubated with taurine. (F) 500 ng of CDKN2AIP recombinant protein were subjected to SDS‐PAGE and silver staining to assess purity. (G) DARTS analysis using CDKN2AIP recombinant protein incubated with taurine. (H) MIN6 cells treated with taurine (100 μM, 24 h) or vehicle were subjected to immunoprecipitation against CDKN2AIP.

Journal: Journal of Diabetes

Article Title: Taurine Alleviates Pancreatic β‐Cell Senescence by Inhibition of p53 Pathway

doi: 10.1111/1753-0407.70100

Figure Lengend Snippet: Identification of Taurine‐CDKN2AIP binding in pancreatic β cells. (A, B) Limited proteolysis‐mass spectrometry (LiP‐MS) was used to screen for taurine interacting proteins in the INS1E β‐cell proteome. Heatmap shows potential taurine binding targets identified by LiP‐MS. Vehicle: N = 3. Taurine: N = 3. (B) p53 pathway related proteins levels between two groups and their binding scores with taurine. (C) Three‐dimensional diagram of the binding modes between human CDKN2AIP and taurine. Taurine potentially binds to CDKN2AIP via residues PRO484, LEU485, LYS486. (D) DARTS analysis using MIN6 cell lysates incubated with taurine. (E) DARTS analysis using INS1E cell lysates incubated with taurine. (F) 500 ng of CDKN2AIP recombinant protein were subjected to SDS‐PAGE and silver staining to assess purity. (G) DARTS analysis using CDKN2AIP recombinant protein incubated with taurine. (H) MIN6 cells treated with taurine (100 μM, 24 h) or vehicle were subjected to immunoprecipitation against CDKN2AIP.

Techniques: Binding Assay, Mass Spectrometry, Incubation, Recombinant, SDS Page, Silver Staining, Immunoprecipitation

Taurine treatment accelerates p53 degradation by binding to CDKN2AIP. (A) p53 protein degradation was detected using cycloheximide (CHX, 10 μM) chase assay. (B) HEK 293 cells were transfected with plasmids encoding GFP‐tagged CDKN2AIP (WT) and CDKN2AIP‐triple mutant (MT) for 48 h. DARTS analysis was performed using cell lysates incubated with taurine, followed by immunoblotting analysis as indicated. (C) INS‐1E cells were transfected with plasmids encoding GFP control, GFP‐tagged CDKN2AIP, and its triple mutant for 24 h, followed by taurine treatment for 24 h. Immunoblotting analysis of CDKN2AIP and p53 in each group.

Journal: Journal of Diabetes

Article Title: Taurine Alleviates Pancreatic β‐Cell Senescence by Inhibition of p53 Pathway

doi: 10.1111/1753-0407.70100

Figure Lengend Snippet: Taurine treatment accelerates p53 degradation by binding to CDKN2AIP. (A) p53 protein degradation was detected using cycloheximide (CHX, 10 μM) chase assay. (B) HEK 293 cells were transfected with plasmids encoding GFP‐tagged CDKN2AIP (WT) and CDKN2AIP‐triple mutant (MT) for 48 h. DARTS analysis was performed using cell lysates incubated with taurine, followed by immunoblotting analysis as indicated. (C) INS‐1E cells were transfected with plasmids encoding GFP control, GFP‐tagged CDKN2AIP, and its triple mutant for 24 h, followed by taurine treatment for 24 h. Immunoblotting analysis of CDKN2AIP and p53 in each group.

Techniques: Binding Assay, Transfection, Mutagenesis, Incubation, Western Blot, Control

RSPO1 induces β cell replication in MIN6 cells, in isolated islets, and in adult WT mice (A) Assessment of MIN6 cell number upon 24-h incubation with increasing doses of native RSPO1 or saline. (B) Representative photographs of pancreatic islets isolated from WT adult mice incubated with either RSPO1 (at 0.4, 1, or 2 μM) or saline. Aiming to label replicating cells, isolated islets were co-incubated with BrdU during the last 24 h and then stained for insulin (red) and BrdU (green). (C) Quantification of the percentage of BrdU + β cells in control islets and islets incubated for 72 h with increasing doses of RSPO1. (D) Pancreatic sections obtained from adult WT mice administered intraperitoneally for 5 consecutive days with different doses of RSPO1 and stained for insulin (red) and Ki67 (green). (E) Quantitative assessment of Ki67 + cells per islet after 5 consecutive RSPO1 administrations. All data shown represent mean ± SEM of n = 5. Results were considered significant if p < 0.0001 (∗∗∗∗), p < 0.001 (∗∗∗), p < 0.01 (∗∗), and p < 0.05 (∗) using one-way ANOVA. See also .

Journal: Cell Reports Medicine

Article Title: RSPO1, a potent inducer of pancreatic β cell neogenesis

doi: 10.1016/j.xcrm.2025.102126

Figure Lengend Snippet: RSPO1 induces β cell replication in MIN6 cells, in isolated islets, and in adult WT mice (A) Assessment of MIN6 cell number upon 24-h incubation with increasing doses of native RSPO1 or saline. (B) Representative photographs of pancreatic islets isolated from WT adult mice incubated with either RSPO1 (at 0.4, 1, or 2 μM) or saline. Aiming to label replicating cells, isolated islets were co-incubated with BrdU during the last 24 h and then stained for insulin (red) and BrdU (green). (C) Quantification of the percentage of BrdU + β cells in control islets and islets incubated for 72 h with increasing doses of RSPO1. (D) Pancreatic sections obtained from adult WT mice administered intraperitoneally for 5 consecutive days with different doses of RSPO1 and stained for insulin (red) and Ki67 (green). (E) Quantitative assessment of Ki67 + cells per islet after 5 consecutive RSPO1 administrations. All data shown represent mean ± SEM of n = 5. Results were considered significant if p < 0.0001 (∗∗∗∗), p < 0.001 (∗∗∗), p < 0.01 (∗∗), and p < 0.05 (∗) using one-way ANOVA. See also .

Article Snippet: The pancreatic immortalized β-cell lines MIN6 (AddexBio) and Ins1 (ATCC), the α-cell-derived αTC1 (ATCC) and the acinar cell line 266-6 (ATCC) were cultured in 100mm Petri dishes (Eppendorf) in cell line-specific culture media indicated by manufacturer’s instructions and maintained in an humified incubator at 37°C and 5% CO2.

Techniques: Isolation, Incubation, Saline, Staining, Control, IF-P

RSPO1 stimulates β cell replication via the activation of Wnt signaling The putative activation of the canonical Wnt signaling pathway upon RSPO1 incubation was assessed in vitro . (A) MIN6 cell number quantification upon incubation for 24 h with increasing doses of two RSPO1 mutants Lgr4 M and Znrf3 M . (B) MIN6 cell number assessment upon a 24-h treatment with saline, 0.1% DMSO, 100 nM MSAB, or 500 nM cardamonin and co-incubation of MIN6 cells with either MSAB or cardamonin and native RSPO1 at 400 nM. (C) Quantification of BrdU + cells in ex vivo murine islets incubated for 72 h with saline, 100 nM MSAB, 1 μM RSPO1, or a combination of the 2. (D) β-catenin protein levels upon RSPO1 treatment assessed by ELISA following a 3-h incubation with saline, native RSPO1, RSPO1 Lgr4 M , or RSPO1 Znrf3 M at 400 nM. Data shown represent mean ± SEM of n = 5. Results were considered significant if p < 0.0001 (∗∗∗∗), p < 0.001 (∗∗∗), p < 0.01 (∗∗), and p < 0.05 (∗) using one-way ANOVA. See also .

Journal: Cell Reports Medicine

Article Title: RSPO1, a potent inducer of pancreatic β cell neogenesis

doi: 10.1016/j.xcrm.2025.102126

Figure Lengend Snippet: RSPO1 stimulates β cell replication via the activation of Wnt signaling The putative activation of the canonical Wnt signaling pathway upon RSPO1 incubation was assessed in vitro . (A) MIN6 cell number quantification upon incubation for 24 h with increasing doses of two RSPO1 mutants Lgr4 M and Znrf3 M . (B) MIN6 cell number assessment upon a 24-h treatment with saline, 0.1% DMSO, 100 nM MSAB, or 500 nM cardamonin and co-incubation of MIN6 cells with either MSAB or cardamonin and native RSPO1 at 400 nM. (C) Quantification of BrdU + cells in ex vivo murine islets incubated for 72 h with saline, 100 nM MSAB, 1 μM RSPO1, or a combination of the 2. (D) β-catenin protein levels upon RSPO1 treatment assessed by ELISA following a 3-h incubation with saline, native RSPO1, RSPO1 Lgr4 M , or RSPO1 Znrf3 M at 400 nM. Data shown represent mean ± SEM of n = 5. Results were considered significant if p < 0.0001 (∗∗∗∗), p < 0.001 (∗∗∗), p < 0.01 (∗∗), and p < 0.05 (∗) using one-way ANOVA. See also .

Techniques: Activation Assay, Incubation, In Vitro, Saline, Ex Vivo, Enzyme-linked Immunosorbent Assay, IF-P

An FC-coupled RSPO1 induces pancreatic β cell neogenesis in vitro , ex vivo , and in vivo (A) Assessment of MIN6 cell number upon 24-h incubation with saline or increasing doses of FC-coupled RSPO1 protein ( n = 5). (B) Quantification of the percentage of BrdU+ cells in saline-treated islets and islets incubated for 72 h with 200 nM or 1 or 3 μM FC-RSPO1 ( n = 5). (C) Weekly monitoring of random glycemia in 10-week-old NOD females injected intraperitoneally weekly for 18 weeks with either saline or 2.4 mg/kg of FC-RSPO1 ( n = 12 in control group, n = 10 in treated group). (D) Quantification of the whole β cell mass in NOD mice weekly administered with either saline or 2.4 mg/kg of FC-coupled RSPO1. The β cell mass of 10-week-old NOD females was used to evaluate the insulin + area at the beginning of the study ( n = 12 in control group, n = 10 in treated group). (E–H) Pancreatic sections from FC-RSPO1-treated NOD mice stained for the β cell markers Pdx1 (E), PC1/3 (F), Nkx6.1 (G), and Glut2 (H). All data shown represent mean ± SEM. Results were considered significant if p < 0.0001 (∗∗∗∗), p < 0.001 (∗∗∗), p < 0.01 (∗∗), and p < 0.05 (∗) following a one-way ANOVA (A and B); a one-way ANOVA, a Mann-Whitney test, or a Kruskal-Wallis test (C); or an unpaired Student’s t test (D).

Journal: Cell Reports Medicine

Article Title: RSPO1, a potent inducer of pancreatic β cell neogenesis

doi: 10.1016/j.xcrm.2025.102126

Figure Lengend Snippet: An FC-coupled RSPO1 induces pancreatic β cell neogenesis in vitro , ex vivo , and in vivo (A) Assessment of MIN6 cell number upon 24-h incubation with saline or increasing doses of FC-coupled RSPO1 protein ( n = 5). (B) Quantification of the percentage of BrdU+ cells in saline-treated islets and islets incubated for 72 h with 200 nM or 1 or 3 μM FC-RSPO1 ( n = 5). (C) Weekly monitoring of random glycemia in 10-week-old NOD females injected intraperitoneally weekly for 18 weeks with either saline or 2.4 mg/kg of FC-RSPO1 ( n = 12 in control group, n = 10 in treated group). (D) Quantification of the whole β cell mass in NOD mice weekly administered with either saline or 2.4 mg/kg of FC-coupled RSPO1. The β cell mass of 10-week-old NOD females was used to evaluate the insulin + area at the beginning of the study ( n = 12 in control group, n = 10 in treated group). (E–H) Pancreatic sections from FC-RSPO1-treated NOD mice stained for the β cell markers Pdx1 (E), PC1/3 (F), Nkx6.1 (G), and Glut2 (H). All data shown represent mean ± SEM. Results were considered significant if p < 0.0001 (∗∗∗∗), p < 0.001 (∗∗∗), p < 0.01 (∗∗), and p < 0.05 (∗) following a one-way ANOVA (A and B); a one-way ANOVA, a Mann-Whitney test, or a Kruskal-Wallis test (C); or an unpaired Student’s t test (D).

Techniques: In Vitro, Ex Vivo, In Vivo, Incubation, Saline, Injection, Control, Staining, IF-P, MANN-WHITNEY

Characterization of lipotoxicity-induced MIN6 mouse β-cell small EVs. A, Mouse MIN6 β-cell line was treated with palmitate (PAL) or control (BSA) for 24 hours and sEV were isolated from the conditioned media. Overlap of a representative Nanoparticle Tracking Analysis (NTA) graph depicting untreated (BSA) control (CTL) EV and palmitate (PAL) EV with the mode for each (120 and 110 nm, respectively). B and C, Overall quantification of particles released from CTL− (n = 9 independent isolations) and PAL EV (n = 15 independent isolations) and average mode of those particles (C). D, Zeta potential was acquired from CTL EV (n = 4) and PAL EV (n = 5) using ZetaView (Particle Matrix). E, Western blot analysis of CTL EV, PAL EV, and MIN6 lysate (control) for sEV biogenesis markers TSG101, CD9, and CD63, with Calnexin as a negative control (representative example from n = 4-10 EV blots. F, Transmission electron microscopy (TEM) of isolated PAL EV (representative example from n = 3 EV isolations); scale bar represents 100 nm. G, Venn diagram depicting unique and overlapping lipid specifies from lipidomic analysis of MIN6 EVs (n = 4/condition) vs MIN6 lysate (n = 3/condition). H, Differential expression of lipid species defined as a molecular percentage of total lipids from PAL EV vs CTL EV. Values are a mean ± SEM. Statistical significance among groups is indicated by *, P < .05.

Journal: Endocrinology

Article Title: Lipotoxicity Induces β-cell Small Extracellular Vesicle–Mediated β-cell Dysfunction in Male Mice

doi: 10.1210/endocr/bqaf067

Figure Lengend Snippet: Characterization of lipotoxicity-induced MIN6 mouse β-cell small EVs. A, Mouse MIN6 β-cell line was treated with palmitate (PAL) or control (BSA) for 24 hours and sEV were isolated from the conditioned media. Overlap of a representative Nanoparticle Tracking Analysis (NTA) graph depicting untreated (BSA) control (CTL) EV and palmitate (PAL) EV with the mode for each (120 and 110 nm, respectively). B and C, Overall quantification of particles released from CTL− (n = 9 independent isolations) and PAL EV (n = 15 independent isolations) and average mode of those particles (C). D, Zeta potential was acquired from CTL EV (n = 4) and PAL EV (n = 5) using ZetaView (Particle Matrix). E, Western blot analysis of CTL EV, PAL EV, and MIN6 lysate (control) for sEV biogenesis markers TSG101, CD9, and CD63, with Calnexin as a negative control (representative example from n = 4-10 EV blots. F, Transmission electron microscopy (TEM) of isolated PAL EV (representative example from n = 3 EV isolations); scale bar represents 100 nm. G, Venn diagram depicting unique and overlapping lipid specifies from lipidomic analysis of MIN6 EVs (n = 4/condition) vs MIN6 lysate (n = 3/condition). H, Differential expression of lipid species defined as a molecular percentage of total lipids from PAL EV vs CTL EV. Values are a mean ± SEM. Statistical significance among groups is indicated by *, P < .05.

Article Snippet: MIN6 β-cell line was obtained from AddexBio and previously authenticated in our laboratory ( ).

Techniques: Control, Isolation, Zeta Potential Analyzer, Western Blot, Negative Control, Transmission Assay, Electron Microscopy, Quantitative Proteomics

Proteomic analysis of lipotoxic β-cell small EV. A, PAL EV and CTL EV were isolated upon PAL or BSA (control) treatment on MIN6 β-cell line for 24 hours. Final sEV pellets were subjected to proteomic analysis. Volcano plot depicting differentially expressed proteins in PAL EV vs CTL EV (n = 3/condition; FC > 1.5; P < .05). B, Venn diagram revealed 70 uniquely enriched proteins in both CTL EV and PAL EV with ∼1400 overlapping proteins. C, Heatmap shows top 10 upregulated and 10 downregulated proteins found in PAL EVs vs CTL EV. D, Panther Go Slim analysis of protein classes that were enriched in PAL EV are depicted in the pie chart. Inserts reveal differentially expressed proteins for “intercellular signal molecule” and “protein-binding activity modulator along with FC and P value. E and F, Western blot confirmation of differentially expressed proteins, IAPP and APP relating to β-cell function and identity in PAL EV (vs CTL EV an MIN6 lysate).

Journal: Endocrinology

Article Title: Lipotoxicity Induces β-cell Small Extracellular Vesicle–Mediated β-cell Dysfunction in Male Mice

doi: 10.1210/endocr/bqaf067

Figure Lengend Snippet: Proteomic analysis of lipotoxic β-cell small EV. A, PAL EV and CTL EV were isolated upon PAL or BSA (control) treatment on MIN6 β-cell line for 24 hours. Final sEV pellets were subjected to proteomic analysis. Volcano plot depicting differentially expressed proteins in PAL EV vs CTL EV (n = 3/condition; FC > 1.5; P < .05). B, Venn diagram revealed 70 uniquely enriched proteins in both CTL EV and PAL EV with ∼1400 overlapping proteins. C, Heatmap shows top 10 upregulated and 10 downregulated proteins found in PAL EVs vs CTL EV. D, Panther Go Slim analysis of protein classes that were enriched in PAL EV are depicted in the pie chart. Inserts reveal differentially expressed proteins for “intercellular signal molecule” and “protein-binding activity modulator along with FC and P value. E and F, Western blot confirmation of differentially expressed proteins, IAPP and APP relating to β-cell function and identity in PAL EV (vs CTL EV an MIN6 lysate).

Article Snippet: MIN6 β-cell line was obtained from AddexBio and previously authenticated in our laboratory ( ).

Techniques: Isolation, Control, Protein Binding, Activity Assay, Western Blot, Cell Function Assay

Small EV generation contributes to lipotoxic-mediated β-cell dysfunction. A and B, MIN6 cells were treated with PAL or PAL + GW4869 (5 μM; 24 hours) vs Control (BSA) EV and EV particle concentrations (A) and mode (B) were assessed using NTA (n = 5-11 independent EV isolations). C and D, Healthy human cadaveric islets were treated with PAL or PAL + GW4869 (5 μM; 24 hours) vs Control (BSA) EV. Particle concentration and mode were assessed using NTA (n = 6 individual videos/treatment). E and F, C57BL/6L mouse islets were treated with palmitate (PAL; 0.5 mM) ± GW4869 for 24 hours. Static glucose stimulated insulin secretion (GSIS) was assessed at 4 mM basal and 16 mM stimulatory glucose concentrations and insulin stimulation index is expressed as 16 mM glucose divided by 4 mM basal concentrations (n = 10-14 independent experiments per condition). G and H, Healthy human islets were treated with 0.5 mM PAL ± GW4869 (5 μM) for 24 hours and static GSIS was conducted. Insulin stimulation index is depicted as 16 mM stimulatory values divided by 4 mM basal values (n = 6 independent experiments per condition).

Journal: Endocrinology

Article Title: Lipotoxicity Induces β-cell Small Extracellular Vesicle–Mediated β-cell Dysfunction in Male Mice

doi: 10.1210/endocr/bqaf067

Figure Lengend Snippet: Small EV generation contributes to lipotoxic-mediated β-cell dysfunction. A and B, MIN6 cells were treated with PAL or PAL + GW4869 (5 μM; 24 hours) vs Control (BSA) EV and EV particle concentrations (A) and mode (B) were assessed using NTA (n = 5-11 independent EV isolations). C and D, Healthy human cadaveric islets were treated with PAL or PAL + GW4869 (5 μM; 24 hours) vs Control (BSA) EV. Particle concentration and mode were assessed using NTA (n = 6 individual videos/treatment). E and F, C57BL/6L mouse islets were treated with palmitate (PAL; 0.5 mM) ± GW4869 for 24 hours. Static glucose stimulated insulin secretion (GSIS) was assessed at 4 mM basal and 16 mM stimulatory glucose concentrations and insulin stimulation index is expressed as 16 mM glucose divided by 4 mM basal concentrations (n = 10-14 independent experiments per condition). G and H, Healthy human islets were treated with 0.5 mM PAL ± GW4869 (5 μM) for 24 hours and static GSIS was conducted. Insulin stimulation index is depicted as 16 mM stimulatory values divided by 4 mM basal values (n = 6 independent experiments per condition).

Article Snippet: MIN6 β-cell line was obtained from AddexBio and previously authenticated in our laboratory ( ).

Techniques: Control, Concentration Assay

Effect of PBM on the viability of MIN6 cells and rat islets. MIN6 cells and rat islets are exposed to 2.8 mW/cm² PBM illumination. a Results of a representative flow cytometry experiment on the viability of MIN6 cells after PBM. Live cells are in Q3 (annexin-negative and PI-negative). Annexin and PI conditions correspond to death control. b Effect of PBM on MIN6 cells’ viability. Results are normalized to the control value. c Representative images of islet viability experiment by confocal microscopy after PBM. Live cells are labeled with Syto13 (green) and dead cells are labeled with PI (red). d Effect of PBM on rat islets’ viability. Results are normalized to the control value. PBM: photobiomodulation, PI: propidium iodide. Results are expressed as the mean ± SEM,n = 4, Scale bares = 100 μm, One-way ANOVA Kruskal–Wallis’s test with pairwise comparison.

Journal: Scientific Reports

Article Title: Long-term safety of photobiomodulation exposure to beta cell line and rat islets in vitro and in vivo

doi: 10.1038/s41598-024-77660-8

Figure Lengend Snippet: Effect of PBM on the viability of MIN6 cells and rat islets. MIN6 cells and rat islets are exposed to 2.8 mW/cm² PBM illumination. a Results of a representative flow cytometry experiment on the viability of MIN6 cells after PBM. Live cells are in Q3 (annexin-negative and PI-negative). Annexin and PI conditions correspond to death control. b Effect of PBM on MIN6 cells’ viability. Results are normalized to the control value. c Representative images of islet viability experiment by confocal microscopy after PBM. Live cells are labeled with Syto13 (green) and dead cells are labeled with PI (red). d Effect of PBM on rat islets’ viability. Results are normalized to the control value. PBM: photobiomodulation, PI: propidium iodide. Results are expressed as the mean ± SEM,n = 4, Scale bares = 100 μm, One-way ANOVA Kruskal–Wallis’s test with pairwise comparison.

Article Snippet: MIN6 cells (immortalized murine pancreatic insulin-secreting β-cells) (AddexBio, San Diego, California, USA) were used and cultured in DMEM 24.8 mM glucose (Gibco, Thermo Fisher Scientific, Waltham, Massachusetts, USA) supplemented with 15% fetal bovine serum (FBS), 1% (v/v) penicillin-streptomycin, and 50μM 2-β mercapto-ethanol.

Techniques: Flow Cytometry, Control, Confocal Microscopy, Labeling, Comparison

Effect of PBM on insulin secretion of MIN6 cells and rat islets. MIN6 cells and rat islets are exposed to 2.8 mW/cm² PBM illumination. a Effect of PBM on MIN6 cells’ insulin secretion (normalized to total insulin content) in response to glucose (n = 8). Results are normalized by total insulin content. b Effects of PBM on islets’ insulin secretion (normalized to total insulin content) in response to glucose (n = 5). Results are normalized by total insulin content. c Effect of PBM on MIN6 cells’ insulin stimulation index (high glucose insulin secretion/low glucose insulin secretion)). d Effect of PBM on islets’ insulin stimulation index. PBM: Photobiomodulation. Results are expressed as mean ± SEM, One-way ANOVA Welch’s with Games Howell post-hoc test, ** p < 0.01, *** p < 0.001.

Journal: Scientific Reports

Article Title: Long-term safety of photobiomodulation exposure to beta cell line and rat islets in vitro and in vivo

doi: 10.1038/s41598-024-77660-8

Figure Lengend Snippet: Effect of PBM on insulin secretion of MIN6 cells and rat islets. MIN6 cells and rat islets are exposed to 2.8 mW/cm² PBM illumination. a Effect of PBM on MIN6 cells’ insulin secretion (normalized to total insulin content) in response to glucose (n = 8). Results are normalized by total insulin content. b Effects of PBM on islets’ insulin secretion (normalized to total insulin content) in response to glucose (n = 5). Results are normalized by total insulin content. c Effect of PBM on MIN6 cells’ insulin stimulation index (high glucose insulin secretion/low glucose insulin secretion)). d Effect of PBM on islets’ insulin stimulation index. PBM: Photobiomodulation. Results are expressed as mean ± SEM, One-way ANOVA Welch’s with Games Howell post-hoc test, ** p < 0.01, *** p < 0.001.

Techniques:

Effect of PBM on superoxide content and mitochondrial membrane potential of MIN6 cells. The MIN6 cells are exposed to 2.8 mW/cm² PBM illumination. a Effect of PBM on MIN6 cells’ superoxide content. Data are expressed as the mean fluorescence intensity. (n = 3 for PBM 2 h,n = 6 for PBM 24 h). b Effect of PBM on the MIN6 cells’ mitochondrial masse. c Effect of PBM on the MIN6 cells’ mitochondrial membrane potential (n = 3 for PBM 2 h,n = 5 for PBM 24 h). Data obtained by flow cytometer and expressed as the mean fluorescence intensity. The mitochondrial membrane potential is (TMRM-CCCp)/MTG. AU: Arbitrary unit; MTG: Mitotracker Green™; PBM: photobiomodulation. Results expressed as the mean ± SEM, One-way ANOVA Welch’s with Games-Howell post-hoc test, * p < 0.05, ** p < 0.01.

Journal: Scientific Reports

Article Title: Long-term safety of photobiomodulation exposure to beta cell line and rat islets in vitro and in vivo

doi: 10.1038/s41598-024-77660-8

Figure Lengend Snippet: Effect of PBM on superoxide content and mitochondrial membrane potential of MIN6 cells. The MIN6 cells are exposed to 2.8 mW/cm² PBM illumination. a Effect of PBM on MIN6 cells’ superoxide content. Data are expressed as the mean fluorescence intensity. (n = 3 for PBM 2 h,n = 6 for PBM 24 h). b Effect of PBM on the MIN6 cells’ mitochondrial masse. c Effect of PBM on the MIN6 cells’ mitochondrial membrane potential (n = 3 for PBM 2 h,n = 5 for PBM 24 h). Data obtained by flow cytometer and expressed as the mean fluorescence intensity. The mitochondrial membrane potential is (TMRM-CCCp)/MTG. AU: Arbitrary unit; MTG: Mitotracker Green™; PBM: photobiomodulation. Results expressed as the mean ± SEM, One-way ANOVA Welch’s with Games-Howell post-hoc test, * p < 0.05, ** p < 0.01.

Techniques: Membrane, Fluorescence, Flow Cytometry

Effect of PBM on ATP/ADP of MIN6 cells. The MIN6 are exposed to 2.8 mW/cm² PBM illumination. PBM: photobiomodulation. Results are obtained by HPLC and expressed as the mean ± SEM,n = 7, One-way ANOVA Welch’s with Games-Howell post-hoc test.

Journal: Scientific Reports

Article Title: Long-term safety of photobiomodulation exposure to beta cell line and rat islets in vitro and in vivo

doi: 10.1038/s41598-024-77660-8

Figure Lengend Snippet: Effect of PBM on ATP/ADP of MIN6 cells. The MIN6 are exposed to 2.8 mW/cm² PBM illumination. PBM: photobiomodulation. Results are obtained by HPLC and expressed as the mean ± SEM,n = 7, One-way ANOVA Welch’s with Games-Howell post-hoc test.

Techniques:

Effect of PBM on mitochondrial and cell respiration of MIN6 cells. The MIN6 cells are exposed to 2.8 mW/cm² PBM illumination. a Effect of PBM on the MIN6 cells’ complex I (glutamate malate) activity. b Effect of PBM on the MIN6 cells’ complex II (succinate) activity. c Effect of PBM on the MIN6 cells’ complex IV (TMPD) activity. d Effect of PBM on respiration of the whole MIN6 cells. AA: antimycin A; Oligo: Oligomycine; PBM: photobiomodulation; RCR: respiratory control ratio (state 3/state 4) graduation is in the right y-axis; ROUT: routine respiration of intact cells. Results are obtained by oxygraphy and expressed as nmol of oxygen consumed per minute per 5 million live cells. Results are expressed as the mean ± SEM,n = 4, One-way ANOVA Fisher’s with Tukey post-hoc test , * p < 0.05, ** p < 0.01.

Journal: Scientific Reports

Article Title: Long-term safety of photobiomodulation exposure to beta cell line and rat islets in vitro and in vivo

doi: 10.1038/s41598-024-77660-8

Figure Lengend Snippet: Effect of PBM on mitochondrial and cell respiration of MIN6 cells. The MIN6 cells are exposed to 2.8 mW/cm² PBM illumination. a Effect of PBM on the MIN6 cells’ complex I (glutamate malate) activity. b Effect of PBM on the MIN6 cells’ complex II (succinate) activity. c Effect of PBM on the MIN6 cells’ complex IV (TMPD) activity. d Effect of PBM on respiration of the whole MIN6 cells. AA: antimycin A; Oligo: Oligomycine; PBM: photobiomodulation; RCR: respiratory control ratio (state 3/state 4) graduation is in the right y-axis; ROUT: routine respiration of intact cells. Results are obtained by oxygraphy and expressed as nmol of oxygen consumed per minute per 5 million live cells. Results are expressed as the mean ± SEM,n = 4, One-way ANOVA Fisher’s with Tukey post-hoc test , * p < 0.05, ** p < 0.01.

Techniques: Activity Assay, Control

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pancreatic β-cell line mouse insulinoma 6 (min6) (National Centre for Cell Science)

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