Journal: Lasers in Surgery and Medicine

Article Title: Effects of Nonablative Er‐ YAG Laser on Human Endometrial Stromal Cells (hESCs): A Pilot Study

doi: 10.1002/lsm.70020

Figure Lengend Snippet: The boxplots of the IGFBP‐1 secretions in the cultured media of human endometrial stromal cells (hESCs). Confluent hESC cultures were treated with E 2 (10 −8 M) (group E 2 ), E 2 (10 −8 M) + ethanol (0.1%) (group E 2 + S), E 2 (10 −8 M) + P 4 (10 −7 M) (group E 2 + P 4 ), Er‐YAG laser+ E 2 (10 −8 M) (group E 2 + L), Er‐YAG laser + E 2 (10 −8 M) + P 4 (10 −7 M) (group E 2 + P 4 + L) for 12, 24, 48, and 72 h. Only hESCs (incubated in serum‐free DMEM/F12) and hESCs + Er‐YAG laser (hESCs + L) groups were incubated for 12 and 72 h. IGFBP‐1 levels were quantified by ELISA in culture media and normalized to total cell protein ( n = 3, median (Q1–Q3). DMEM, Dulbecco modified Eagle medium; E 2 , estradiol; ELISA, enzyme‐linked immunosorbent assay; Er‐YAG, nonablative Erbium YAG; ESC, endometrial stromal cell; IGFBP‐1, insulin‐like growth factor‐binding protein‐1; P 4 , progesterone.

Article Snippet: To investigate the effect of the Er‐YAG laser on endometrial decidualization, we measured insulin‐like growth factor‐binding protein‐1 (IGFBP‐1) levels in conditioned media using ELISA kits (Elabscience; E‐EL‐H0442, Houston, TX, USA) [ ] with a sensitivity of 0.10 ng/mL and no reported cross‐reactivity or interference.

Techniques: Cell Culture, Incubation, Enzyme-linked Immunosorbent Assay, Modification, Binding Assay

Journal: The EMBO Journal

Article Title: IGFBP 1 increases β‐cell regeneration by promoting α‐ to β‐cell transdifferentiation

doi: 10.15252/embj.201592903

Figure Lengend Snippet: A Schema of the analysis of gene expression in islets isolated from control larvae and larvae subjected to β‐cell ablation. β cells were ablated by exposing nitroreductase (NTR)‐expressing transgenic larvae to metronidazole (MTZ) from 3 to 4 dpf. Islets were then isolated, and their RNA extracted and analyzed by microarray. Out of the 470 genes that were upregulated more than 50%, 33 genes encoded proteins that harbored a signal peptide for secretion (according to the algorithm of SignalP). Excluding genes that encode enzymes or proteins with a transmembrane (TM) domain, we selected 11 genes for overexpression studies in zebrafish larvae (C–E). B Microarray heat map showing the upregulation in expression of the 11 candidate genes in β‐cell‐ablated versus control islets. Igfbp1a and b were the genes whose expression increased the most after β‐cell ablation. C Schema of the construct used for overexpression of the candidate genes (under the control of the beta‐actin promoter), and expression of GFP in the heart (as an internal control for genomic integration). D, E Representative images at 6 dpf of Tg ( ins:kaede ) ;Tg ( ins:CFP‐NTR ) transgenic larvae that had been injected at the 1–2 cell stage with transposase mRNA (control) or transposase mRNA + bactin:igfbp1a ( bactin:igfbp1a ), subjected to β‐cell ablation by metronidazole (MTZ) during 3–4 dpf, and subsequently allowed to regenerate for 2 days. The GFP + heart (arrowhead) visualizes successful integration of the construct. Islets are indicated by white arrows. Scale bars: 100 μm. F Quantification of β‐cell regeneration at 6 dpf in control ( n = 23), bactin:igfbp1a ‐overexpressing ( n = 13), and bactin:igfbp1b ‐overexpressing ( n = 8) Tg ( ins:kaede ) ;Tg ( ins:CFP‐NTR ) larvae; *** P = 0.0002, ns = non‐significant ( P = 0.3106). G Immunohistochemistry showing Igfbp1 protein expression in 6 dpf Tg ( ins:GFP ) following β‐cell ablation between 3 and 4 dpf. Scale bar: 50 μm. H Both liver‐specific ( lfabp promoter; n = 46) and widespread ( bactin promoter; n = 18) overexpression of igfbp1a increase β‐cell regeneration when compared to control ( n = 77); *** P < 0.001. I–M Quantification of β cells with or without β‐cell ablation and igfbp1a overexpression by confocal microscopy, which detects even weakly insulin‐expressing β cells; **** P < 0.0001. Scale bars: 15 μm. Data information: Results are presented as mean values ± SEM and analyzed with one‐way ANOVA (F, H) or two‐way ANOVA (I). See also Fig .

Article Snippet: The islets were then treated with PBS (vehicle control) or 1 μg/ml of recombinant human IGFBP1 protein (R&D Systems) for 72 h. Thereafter, the islets were snap‐frozen in Tissue‐Tek TM (Sakura) and sectioned.

Techniques: Gene Expression, Isolation, Control, Expressing, Transgenic Assay, Microarray, Over Expression, Construct, Injection, Immunohistochemistry, Confocal Microscopy

Journal: The EMBO Journal

Article Title: IGFBP 1 increases β‐cell regeneration by promoting α‐ to β‐cell transdifferentiation

doi: 10.15252/embj.201592903

Figure Lengend Snippet: A–C Igfbp1a promotes β‐cell regeneration, rather than β‐cell survival. To trace β cells, we exposed Tg ( ins:kaede ) larvae to UV light (which causes the existing Kaede protein to switch from emitting green fluorescence to emitting red fluorescence) just before ablating the β cells by MTZ treatment from 3 to 4 dpf. After regeneration, the newly formed β cells are green, whereas the β cells that survived ablation are yellow (overlap of green and red). Representative confocal images (A, B) at 6 dpf of control and bactin:igfbp1a ‐overexpressing Tg ( ins:kaede ) ;Tg ( ins:CFP‐NTR ) transgenic larvae; arrows indicate surviving (yellow) β cells. Scale bars: 10 μm. (C) Quantification of β‐cell regeneration (green bars) and β‐cell survival (yellow bars) per larva at 6 dpf. *** P < 0.001; n = 20 larvae in the control group, n = 27 larvae in the bactin:igfbp1a ‐overexpressing group. D–F Igfbp1a does not promote δ‐cell regeneration. We treated control and bactin:igfbp1a ‐overexpressing Tg ( sst:flag‐NTR ) ;Tg ( sst:dsRed ) larvae with MTZ from 3 to 4 dpf to ablate δ cells, and then allowed them to regenerate until 6 dpf. Representative confocal images (D, E) at 6 dpf of control and bactin:igfbp1a ‐overexpressing larvae showing comparable number of δ cells after 2 days of regeneration. Scale bars: 15 μm. (F) Quantification of the total number of δ cells per δ‐cell‐ablated larva at 6 dpf compared to the baseline number of δ cells in non‐ablated control larvae. ns, P = 0.2325; n = 13 in the control group, n = 7 in the bactin:igfbp1a group. G Other Igfbps do not promote β‐cell regeneration. We injected Tg ( ins:kaede ) ;Tg ( ins:CFP‐NTR ) transgenics with either transposase mRNA (control; n = 31) or transposase mRNA + one of six different igfbps , igfbp2a ( n = 25), igfbp2b ( n = 25), igfbp3 ( n = 26), igfbp5a ( n = 17), igfbp6b ( n = 34), and igfbp7 ( n = 17), treated them with MTZ from 3 to 4 dpf to ablate their β cells, and then quantified their β cells after 2 days of regeneration, at 6 dpf; ns=non‐significant (igfbp2a P = 0.986, igfbp3 P = 0.979, igfbp5a P = 0.999, igfbp6b P = 0.997, and igfbp7 P = 0.999); igfbp2b * P = 0.037. H Tg ( pcsk1:GFP ) is expressed in regenerating β cells within the islet (arrowheads), but not outside the islet (arrow), at 6 dpf in larvae overexpressing igfbp1a . Scale bar: 15 μm. I, J Free‐glucose levels during β‐cell regeneration in control ( bactin:mCherry ) and bactin : igfbp1a ‐overexpressing Tg ( ins:kaede ) ;Tg ( ins:CFP‐NTR ) larvae (I), as well as in Tg ( ins:kaede ) ;Tg ( ins:CFP‐NTR ) larvae injected in the pericardial sac at 4 dpf with 1 ng of recombinant mouse Igfbp1 (J). We treated larvae with MTZ from 3 to 4 dpf to ablate their β cells and monitored their free‐glucose levels at 3–7 dpf. Free‐glucose levels were significantly lower after genetic igfbp1a overexpression or Igfbp1‐protein injection (red lines) than in controls (black lines) at 7 and 6 dpf, respectively. Baseline reference levels of free glucose throughout development are shown for a different set of larvae without β‐cell ablation. n = 24 larvae (four pools of six larvae) per data point; *** P < 0.001, * P < 0.05. Data information: Results are presented as mean values ± SEM and analyzed with t ‐test (C), Mann‐Whitney test (F), one‐way ANOVA (G), or two‐way ANOVA (I, J).

Article Snippet: The islets were then treated with PBS (vehicle control) or 1 μg/ml of recombinant human IGFBP1 protein (R&D Systems) for 72 h. Thereafter, the islets were snap‐frozen in Tissue‐Tek TM (Sakura) and sectioned.

Techniques: Fluorescence, Control, Transgenic Assay, Injection, Recombinant, Over Expression, MANN-WHITNEY

Journal: The EMBO Journal

Article Title: IGFBP 1 increases β‐cell regeneration by promoting α‐ to β‐cell transdifferentiation

doi: 10.15252/embj.201592903

Figure Lengend Snippet: Isolated islets from mice and from human donors were treated for 3 days with recombinant mouse or human IGFBP 1 protein, respectively. A, B Representative confocal images of control and Igfbp1‐treated mouse islets. gcg + ins + cells are indicated by arrows. Scale bars: 25 μm. C Quantification of the percentage of gcg + ins + bihormonal cells per ins + cells in mouse islets after treatment with increasing concentrations of Igfbp1. P = 0.7304 (Igfbp1 0.3 μg/ml), ** P = 0.0023 (1 μg/ml), ** P = 0.0013 (10 μg/ml), * P = 0.0410 (25 μg/ml), respectively. Multiple optical sections from 10–50 islets per group, from 4–8 different mice, were used for the quantifications. D Quantification of β‐cell proliferation in mouse islets after treatment with Igfbp1; data displayed as relative values so that the proliferation markers EdU and Ki67 can be compared. Multiple optical sections from 14–22 islets, from 3 different mice, were used for the quantifications. E, F Representative confocal images of control or IGFBP1‐treated human islets. A gcg + ins + cell is indicated by an arrow. For clarity, individual colors for denoting insulin (green), glucagon (red), and DAPI (blue) are also shown separately. Scale bars: 15 μm. G Quantification of the percentage of gcg + ins + bihormonal cells per ins + cells in human islets from five donors. ** P = 0.0036. Co‐expression of insulin and glucagon is indicative of transdifferentiation, which could reflect either α cells transdifferentiating to β cells or vice versa. ** P < 0.0028; n = 115 sections in the control group, n = 147 sections in the IGFBP1 group. H Our proposed mechanism by which the extracellular signals insulin, IGFs, and IGFBP1 can regulate α‐ to β‐cell transdifferentiation. Data information: Results are presented as mean values ± SEM and analyzed with Kruskal‐Wallis (C) or Mann‐Whitney tests (D, G). Source data are available online for this figure.

Article Snippet: The islets were then treated with PBS (vehicle control) or 1 μg/ml of recombinant human IGFBP1 protein (R&D Systems) for 72 h. Thereafter, the islets were snap‐frozen in Tissue‐Tek TM (Sakura) and sectioned.

Techniques: Isolation, Recombinant, Control, Expressing, MANN-WHITNEY

Journal: The EMBO Journal

Article Title: IGFBP 1 increases β‐cell regeneration by promoting α‐ to β‐cell transdifferentiation

doi: 10.15252/embj.201592903

Figure Lengend Snippet: A–D Correlations between baseline levels of fasting IGFBP1 levels and BMI (A, C) or fasting insulin levels (B, D) in men (A, B) and women (C, D). The correlations between these baseline values are significant both in subjects who developed T2D (red squares) and in controls with a normal glucose tolerance (NGT) (blue circles) at follow‐up after 8–10 years. Results are presented as individual values with linear regression. Related to Table .

Article Snippet: The islets were then treated with PBS (vehicle control) or 1 μg/ml of recombinant human IGFBP1 protein (R&D Systems) for 72 h. Thereafter, the islets were snap‐frozen in Tissue‐Tek TM (Sakura) and sectioned.

Techniques:

Journal: The EMBO Journal

Article Title: IGFBP 1 increases β‐cell regeneration by promoting α‐ to β‐cell transdifferentiation

doi: 10.15252/embj.201592903

Figure Lengend Snippet: High levels of IGFBP1 are associated with a lower risk of developing type‐2 diabetes

Article Snippet: The islets were then treated with PBS (vehicle control) or 1 μg/ml of recombinant human IGFBP1 protein (R&D Systems) for 72 h. Thereafter, the islets were snap‐frozen in Tissue‐Tek TM (Sakura) and sectioned.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Hepatic Oleate Regulates Insulin-like Growth Factor-Binding Protein 1 Partially through the mTORC1-FGF21 Axis during High-Carbohydrate Feeding

doi: 10.3390/ijms232314671

Figure Lengend Snippet: Hepatic SCD1 deficiency up-regulates IGFBP1. ( A ) Relative hepatic Igfbp1 mRNA levels in 8–10 weeks old male WT (n = 4) and GKO (n = 6) mice fed HCD for 10 days. ( B ) Relative hepatic Igfbp1 mRNA levels in 8–13 weeks old male LOX (n = 13) and LKO (n = 12) mice fed HCD for 7–10 days. ( C ) Plasma IGFBP1 protein concentration in 8–10 weeks old male LOX (n = 7) and LKO (n = 7) mice fed HCD for 7–10 days. ( D ) Relative IGFBP1 mRNA levels in HepG2 cells treated for 24 h with BSA (n = 10) or BSA plus 4 μM (n = 12) of SCD inhibitor. Values are mean ± SEM; * p < 0.05 vs. WT, LOX, or BSA, *** p < 0.005 vs. WT, LOX or BSA by Student’s two-tailed t -test.

Article Snippet: Lastly, secreted IGFBP1 protein was measured using the Human IGFBP-1 ELISA Kit PicoKineTM (Boster Biological Technology, Pleasanton, CA, USA, Catalog #EK0382).

Techniques: Clinical Proteomics, Protein Concentration, Two Tailed Test

Journal: International Journal of Molecular Sciences

Article Title: Hepatic Oleate Regulates Insulin-like Growth Factor-Binding Protein 1 Partially through the mTORC1-FGF21 Axis during High-Carbohydrate Feeding

doi: 10.3390/ijms232314671

Figure Lengend Snippet: Oleic acid blunts IGFBP1 up-regulation in vivo and in vitro. ( A ) Relative hepatic Igfbp1 mRNA levels in 12–14 weeks old male LOX and LKO mice fed either HCD supplemented with 15% by weight tristearin (n = 4 LOX and n = 4 LKO) or 15% by weight triolein (n = 5 LOX; 7 LKO) for 10 days. ( B ) Relative hepatic Igfbp1 mRNA levels in 8–10 weeks old male WT (n = 4), GKO (n = 6), and SCD5tg (n = 5) mice fed HCD for 10 days. ( C ) Relative Igfbp1 mRNA level in HepG2 cells treated for 24 h with either BSA (n = 10), BSA plus 4 μM SCD inhibitor (n = 12), BSA plus 4μM SCD inhibitor, and 500 μM oleic acid (n = 12), or BSA plus SCD inhibitor and 500 μM steric acid (n = 12). ( D ) Secreted IGFBP1 protein concentration in HepG2 cells treated for 24 h with either BSA (n = 7), BSA plus 4 μM SCD inhibitor (n = 8), BSA plus 4 μM SCD inhibitor and 500 μM oleic acid (n = 7), or BSA plus SCD inhibitor and 500 μM steric acid (n = 8). Values are mean ± SEM; * p < 0.05 vs. WT, LOX, or BSA; ** p < 0.01 vs. WT, LOX, or BSA; *** p < 0.005 vs. WT, LOX triolein or BSA by Student′s two-tailed t -test.

Article Snippet: Lastly, secreted IGFBP1 protein was measured using the Human IGFBP-1 ELISA Kit PicoKineTM (Boster Biological Technology, Pleasanton, CA, USA, Catalog #EK0382).

Techniques: In Vivo, In Vitro, Protein Concentration, Two Tailed Test

Journal: International Journal of Molecular Sciences

Article Title: Hepatic Oleate Regulates Insulin-like Growth Factor-Binding Protein 1 Partially through the mTORC1-FGF21 Axis during High-Carbohydrate Feeding

doi: 10.3390/ijms232314671

Figure Lengend Snippet: Inhibition of mTORC1 by rapamycin suppresses IGFBP1 induction. ( A ) Relative hepatic Igfbp1 mRNA levels in 10–12 weeks old male LOX and LKO mice fed HCD diet for 10 days treated daily with either vehicle (n = 3 LOX; 3 LKO) or rapamycin (n = 5 LOX + R; 5 LKO + R). ( B ) Relative Igfbp1 mRNA levels of HepG2 cells treated with either BSA (n = 5), BSA plus 1μM SCD inhibitor (n = 5), or BSA plus 4 μM SCD inhibitor and rapamycin (n = 5). Values are mean ± SEM; * p < 0.05 vs. LOX or BSA; ** p < 0.01 vs. LOX+V or BSA; *** p < 0.005 vs. LOX or BSA by Student′s two-tailed t -test.

Article Snippet: Lastly, secreted IGFBP1 protein was measured using the Human IGFBP-1 ELISA Kit PicoKineTM (Boster Biological Technology, Pleasanton, CA, USA, Catalog #EK0382).

Techniques: Inhibition, Two Tailed Test

Journal: International Journal of Molecular Sciences

Article Title: Hepatic Oleate Regulates Insulin-like Growth Factor-Binding Protein 1 Partially through the mTORC1-FGF21 Axis during High-Carbohydrate Feeding

doi: 10.3390/ijms232314671

Figure Lengend Snippet: IGFBP1 is regulated by SCD1 through the mTORC1-FGF21 axis. ( A ) Relative hepatic Scd1 and Fgf21 mRNA levels in LOX/LOX (n = 6, 3 male; 3 female) and DLKO (n = 6, 3 male; 3 female) mice fed HCD for 10 days. ( B ) Relative hepatic Igfbp1 mRNA level in LOX/LOX (n = 6, 3 male; 3 female) and DLKO mice fed HCD (n = 6, 3 male; 3 female) for 10 days. ( C ) Relative Fgf21 mRNA level in HepG2 cells treated with BSA (n = 6), BSA plus 1 μM SCD inhibitor (n = 6), BSA plus 1 μM SCD inhibitor and 100 μM oleic acid (n = 5), BSA plus 1 μM SCD inhibitor and 100 μM steric acid (n = 5), BSA plus 1 μM SCD inhibitor and rapamycin (n = 5). ( D ) Relative FGF21 mRNA level in HepG2 cells treated with control lentiviral particles (n = 6), control lentiviral particles plus SCD inhibitor (n = 6), or shFGF21 lentiviral particles plus SCD inhibitor (n = 6). ( E ) Relative IGFBP1 mRNA level in HepG2 cells treated with control lentiviral particles (n = 6), control lentiviral particles plus SCD inhibitor (n = 5), or shFGF21 lentiviral particles plus SCD inhibitor (n = 5). Values are mean ± SEM; ns = not significant; * p < 0.05 vs. LOX/LOX, BSA, or control; ** p < 0.01 vs. LOX/LOX, BSA, or control; *** p < 0.005 vs. LOX/LOX, BSA, or control by Student′s two-tailed t -test.

Article Snippet: Lastly, secreted IGFBP1 protein was measured using the Human IGFBP-1 ELISA Kit PicoKineTM (Boster Biological Technology, Pleasanton, CA, USA, Catalog #EK0382).

Techniques: Control, Two Tailed Test

Journal: International Journal of Molecular Sciences

Article Title: Hepatic Oleate Regulates Insulin-like Growth Factor-Binding Protein 1 Partially through the mTORC1-FGF21 Axis during High-Carbohydrate Feeding

doi: 10.3390/ijms232314671

Figure Lengend Snippet: Proposed model of IGFBP1 induction through the mTORC1-FGF21 axis.

Article Snippet: Lastly, secreted IGFBP1 protein was measured using the Human IGFBP-1 ELISA Kit PicoKineTM (Boster Biological Technology, Pleasanton, CA, USA, Catalog #EK0382).

Techniques:

Journal: Molecular human reproduction

Article Title: Decreased expression of NR4A nuclear receptors in adenomyosis impairs endometrial decidualization.

doi: 10.1093/molehr/gaw042

Figure Lengend Snippet: Figure 2 NR4A regulation of human endometrial stromal cell (hESC) decidualization in vitro. hESCs were transduced with Ad-LacZ or Ad-NR4A at a multiplicity of infection (MOI) of 25 or were treated with 0.5 mM 8-Br-cAMP and 1 µM MPA for 48 h. PRL (A) and IGFBP1 (B) mRNA levels were measured by real-time PCR. The results are presented as the mean ± SEM; n = 3 samples. * P < 0.05, ** P < 0.01, and *** P < 0.001 compared with Ad-LacZ. hESCs were infected with the indicated adenoviruses at a MOI of 50, followed by treatment with or without 8-Br-cAMP and MPA for an add- itional 4 days. Prolactin (C) and IGFBP-1 (D) released into the medium was measured by Enzyme Linked Fluorescent Assay (ELFA) and ELISA. The results are presented as the mean ± SEM; n = 5 samples. * P < 0.05, ** P < 0.01, and *** P < 0.001 compared with Ad-LacZ. hESCs were transfected with 50 nM of the indicated specific NR4A siRNA for 72 h, followed by treatment with or without 8-Br-cAMP and MPA for an additional 4 days. Prolactin (E) and IGFBP-1 (F) released into the medium were measured by ELFA and ELISA. The results are presented as the mean ± SEM; n = 4 sam- ples. *** P < 0.001 compared with siCTL. # P < 0.05 and ## P < 0.01 compared with siCTL plus 8-Br-cAMP and MPA. (G and H) Fluorescein isothio- cyanate labeled phalloidin was used to label actin filaments, and immunofluorescence was used to analyze the morphological transformation of hESCs.

Article Snippet: Detection of prolactin and IGFBP-1 levels in cultured supernatants Prolactin and IGFBP-1 levels in supernatants were measured using a Vidas prolactin kit (bioMérieux) and a human IGFBP1 ELISA kit (BOSTER).

Techniques: In Vitro, Transduction, Infection, Real-time Polymerase Chain Reaction, Fluorescence, Enzyme-linked Immunosorbent Assay, Transfection, Labeling, Transformation Assay