Journal: The FASEB Journal

Article Title: Cathepsin B as a potential cystatin M/E target in the mouse hair follicle

doi: 10.1096/fj.201700267R

Figure Lengend Snippet: Regulation of epidermal protease activity by CST6. Model of the regulatory role of CST6 in processes that control epidermal cornification, desquamation and HF maintenance. 1 ) Inhibition of CTSL activity by CST6 is important in the cornification process, as CTSL is the elusive processing and activating enzyme for (TGM)-3. CTSL is also able to process CTSD, which in turn can activate TGM-1. 2 ) Inhibition of CTSV regulates desquamation, as CTSV is able to degrade desmosomal and corneodesmosomal proteins, such as desmoglein-1, desmocollin-1, and corneodesmosin. As CTSV is expressed only in humans, murine CTSL probably controls the specific functional enzymatic activities of both human CTSL and CTSV. 3 ) The findings in the present study suggest that inhibition of CtsB by Cst6 protects HF maintenance in mice. 4 ) Inhibition of human LGMN regulates the processing of (pro)-cathepsins; however mouse LGMN is not inhibited by mouse Cst6.

Article Snippet: Subsequently, standards, controls, and samples (undiluted up to 32× diluted) were incubated for 1 h, followed by incubation with monoclonal rat anti-mouse Cst6 (R&D Systems) in PBS/1% normal rabbit serum/0.1% bovine serum albumin/0.05% Tween-20 for 30 min. Next, wells were incubated with goat anti-rat biotinylated antibody (Vector Laboratories) for 30 min, followed by a final incubation with avidin-biotinylated horseradish peroxidase complex (Vector Laboratories) for 30 min.

Techniques: Activity Assay, Control, Inhibition, Functional Assay

Journal: PLoS ONE

Article Title: The Transcription Factor Myt3 Acts as a Pro-Survival Factor in β-cells

doi: 10.1371/journal.pone.0051501

Figure Lengend Snippet: Saggital sections of E14.5, E16.5 and E18.5 pancreata were analysed for expression of Insulin, Glucagon or Pdx1, as indicated (red), and Myt3 (green). Nuclei were stained with Hoechst (blue). Arrows indicate co-localisation of Myt3 with indicated endocrine markers.

Article Snippet: Sections were co-stained with rabbit anti-Myt3 (1/250) and guinea pig anti-Insulin (1/1000; Linco), guinea pig anti-Glucagon (1/1000; Linco), guinea pig anti-PP (1/100; Linco), goat anti-Somatostatin (1/1000; Santa Cruz) or mouse anti-Pdx1 (1/500; DSHB).

Techniques: Expressing, Staining

Journal: PLoS ONE

Article Title: The Transcription Factor Myt3 Acts as a Pro-Survival Factor in β-cells

doi: 10.1371/journal.pone.0051501

Figure Lengend Snippet: A) A screenshot of the Myt3 promoter region in the UCSC genome browser showing Foxa2, Pdx1, Mafa and Neurod1 ChIP-seq data from islets. Peaks indicate binding sites. ChIP-qPCR was used to validate B) Foxa2, C) Pdx1 and D) Neurod1 binding within the Myt3 promoter region. Nkx2.2 , Pdx1 and Abcc8 are positive controls for Foxa2, Pdx1 and Neurod1 binding respectively. E–G) Relative luciferase activity of the indicated luciferase reporter vectors co-transfected with empty vector or with Foxa2, Pdx1 or Neurod1 expressing vectors. Mutant vectors had the indicated transcription factor binding sites altered by site-directed mutagenesis. Wild type and mutant binding site sequences are as indicated. H) Myt3 expression relative to β-actin following treatment of mPAC cells with pAdV- Ngn3 . ChIP-qPCR was used to determine I) H3K4me1, J) H3K4me3, K) H3K27ac and L) H3K27me3 histone modifications at the Myt3 promoter. * indicates a statistically significant difference at p≤0.05, ** at p≤0.01, and *** at p≤0.001 based on student’s t-test for luciferase data and a Kruskal-Wallis test with a Dunn’s multiple comparison for ChIP-qPCR data.

Article Snippet: Sections were co-stained with rabbit anti-Myt3 (1/250) and guinea pig anti-Insulin (1/1000; Linco), guinea pig anti-Glucagon (1/1000; Linco), guinea pig anti-PP (1/100; Linco), goat anti-Somatostatin (1/1000; Santa Cruz) or mouse anti-Pdx1 (1/500; DSHB).

Techniques: ChIP-sequencing, Binding Assay, Luciferase, Activity Assay, Transfection, Plasmid Preparation, Expressing, Mutagenesis

Journal: PLoS ONE

Article Title: The Transcription Factor Myt3 Acts as a Pro-Survival Factor in β-cells

doi: 10.1371/journal.pone.0051501

Figure Lengend Snippet: Islets were transduced with adenoviruses expressing shRNA’s targeting Myt3 or a scrambled sequence. A) Myt3 expression was determined relative to β-actin. Gapdh was used as a control for off target effects of the virus. B) Western blot analysis of Myt3 and β-actin protein levels in islets. C) Results of the densitometry of triplicate western blot analyses from B relative to β-actin. Ex vivo islets were transduced as above and D) cellular insulin content and insulin secretion induced with E) 16.7 mM Glucose, F) 30 mM KCl or G) 10 mM Arginine were determined 48 hrs later by Insulin ELISA. qPCR was used to determine the relative expression of H) transcription factors and cofactors and I) Genes involved in β-cell function/physiology as compared to β-actin . J) Western blot analysis of Mafa, Pdx1, and β-actin protein levels. K) Results of the densitometry of triplicate western blot analyses from J relative to β-actin. * indicates a statistically significant difference at p≤0.05, ** at p≤0.01, *** at p≤0.001 based on students t-test.

Article Snippet: Sections were co-stained with rabbit anti-Myt3 (1/250) and guinea pig anti-Insulin (1/1000; Linco), guinea pig anti-Glucagon (1/1000; Linco), guinea pig anti-PP (1/100; Linco), goat anti-Somatostatin (1/1000; Santa Cruz) or mouse anti-Pdx1 (1/500; DSHB).

Techniques: Transduction, Expressing, Sequencing, Western Blot, Ex Vivo, Enzyme-linked Immunosorbent Assay, Cell Function Assay

Journal: International Journal of Molecular Sciences

Article Title: Expression and Regulation of a Novel Decidual Cells-Derived Estrogen Target during Decidualization

doi: 10.3390/ijms24010302

Figure Lengend Snippet: Cstb expression is associated with angiogenesis in mouse uterine stromal cells Angptl7 . ( A ) Gene ontology (GO) functional classification of the DEGs. ( B ) The expressions of Cstb and ZO1 were detected by immunofluorescence in the uterus on day 6. Bar = 300 μm. ( C ) The expression of Angptl7 in stromal cells of the con NC group, dc NC group, and dc si Cstb group was detected by RT-qPCR. ( D ) The expression of Angptl7 in mouse uteri from days 5 to 8 of pregnancy was detected by in situ hybridization. Bar = 300 μm. * p < 0.05. NC, negative control; si Cstb , siRNA of Cstb ; dc, in vitro decidualization.

Article Snippet: Briefly, frozen sections were fixed in 4% paraformaldehyde solution for 10 min and then soaked in 0.1% Triton X-100 in PBS for 15 min. After blocking with 5% donkey serum (Zhongshan Jinqiao, Beijing, China) in a 37 °C oven for 60 min, the sections were incubated with mouse anti- Cstb (1:500, Santa Cruz, sc-166561), ZO1 (1:200, Cell Signaling Technology, Boston, MA, USA, #13663) diluted in PBS overnight in a 4 °C refrigerator.

Techniques: Expressing, Functional Assay, Immunofluorescence, Quantitative RT-PCR, In Situ Hybridization, Negative Control, In Vitro

Journal: Bioengineering

Article Title: Simultaneous High-Frame-Rate Acoustic Plane-Wave and Optical Imaging of Intracranial Cavitation in Polyacrylamide Brain Phantoms during Blunt Force Impact

doi: 10.3390/bioengineering11020132

Figure Lengend Snippet: Cavitation mapping entailed the ( a ) creation of a spectral map by transforming the raw signal of a signal channel through Matlab’s built-in STFT function, ( b ) the fabrication of a baseline and comparison to a frame of interest by extracting data based on selected user-defined frequencies, and ( c ) a comparison of baseline and a selected acquisition for mapping potential regions of stable and inertial cavitation onto a reconstructed plane wave frame.

Article Snippet: Cavitation mapping involved transforming the raw time-domain signal of an individual channel into Matlab’s built-in short-time Fourier transform (STFT) function ( a) with inputs shown in mapped with an amplitude logarithmic scale.

Techniques: Comparison

Journal: Bioengineering

Article Title: Simultaneous High-Frame-Rate Acoustic Plane-Wave and Optical Imaging of Intracranial Cavitation in Polyacrylamide Brain Phantoms during Blunt Force Impact

doi: 10.3390/bioengineering11020132

Figure Lengend Snippet: Matlab STFT inputs for the acoustic spectral analysis.

Article Snippet: Cavitation mapping involved transforming the raw time-domain signal of an individual channel into Matlab’s built-in short-time Fourier transform (STFT) function ( a) with inputs shown in mapped with an amplitude logarithmic scale.

Techniques: Plasmid Preparation

Journal: The Journal of Biological Chemistry

Article Title: Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion

doi: 10.1016/j.jbc.2025.108187

Figure Lengend Snippet: Loss of CPT1A function in the pancreas of male mice promotes increased pancreatic and islet triglyceride accumulation with no change in body composition, energy expenditure, respiratory quotient, or insulin sensitivity. A , expression of Cpt1a, Cpt1b, and Cpt1c were measured by RT-PCR using RNA isolated from islets, ( B ) CPT1A abundance in formalin-fixed pancreatic tissue, ( C ) CPT1 enzymatic activity in mitochondrial suspensions isolated from whole pancreatic tissue, ( D ) triglyceride content measured from whole pancreatic tissue, ( E ) islet triglyceride scores, ( F ) body mass, ( G ) fat mass, ( H ), lean mass, ( I ) fluid mass, ( J ) energy expenditure (EE) across the light ( white bars ) and dark ( gray bars ) cycle over a 7 day period, ( K ) respiratory quotient (RQ) across the light ( white bars ) and dark ( gray bars ) cycle over a 7 day period, ( L ) total activity quantified by the total number of X and Y beam breaks (XT + XY) over a 7 day period, and ( M ) an insulin tolerance test in 20 week old mice. All experiments were performed in male littermate Cpt1a CON versus Cpt1a Pdx1−/− mice. n = 8 to 10 per group ( A – D and F – M ), n = 4 ( E ). A representative islet in ( B ) was chosen from each genotype. ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001. CPT1A, carnitine palmitoyltransferase 1A.

Article Snippet: Mice with a pancreas-specific deletion of Cpt1a (Cpt1a Pdx1−/− ) were generated by crossing Cpt1a fl/fl mice (C57BL/6- Cpt1a tm1.1mrl Model # 9759 from Taconic) with Pdx1-Cre mice (Stock # 014647; Jackson Laboratory, Bar Harbor, ME).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Isolation, Activity Assay

Journal: The Journal of Biological Chemistry

Article Title: Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion

doi: 10.1016/j.jbc.2025.108187

Figure Lengend Snippet: Decreased Cpt1a expression in the pancreas of female mice correlates with increased pancreatic triglyceride accrual but no change in insulin sensitivity. A , expression of Cpt1a was measured by RT-PCR using RNA isolated from islets, ( B ) CPT1A abundance in formalin-fixed pancreatic tissue, ( C ) triglyceride content from whole pancreatic tissue, ( D ) body mass, ( E ) fat mass, ( F ), lean mass, ( G ) fluid mass, ( H ) an insulin tolerance test in 25 week old mice. All experiments were performed in female littermate Cpt1a CON versus Cpt1a Pdx1−/− mice (n = 9–10 per group). A representative islet in ( B ) was chosen from each genotype. ∗∗ p < 0.01 and ∗∗∗ p < 0.001. CPT1A, carnitine palmitoyltransferase 1A.

Article Snippet: Mice with a pancreas-specific deletion of Cpt1a (Cpt1a Pdx1−/− ) were generated by crossing Cpt1a fl/fl mice (C57BL/6- Cpt1a tm1.1mrl Model # 9759 from Taconic) with Pdx1-Cre mice (Stock # 014647; Jackson Laboratory, Bar Harbor, ME).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Isolation

Journal: The Journal of Biological Chemistry

Article Title: Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion

doi: 10.1016/j.jbc.2025.108187

Figure Lengend Snippet: Deletion of CPT1A in pancreatic tissue reduces glucose tolerance and β-cell function in male mice, but the ability of β-cells to proliferate in response to metabolic adaptations is retained. Glucose tolerance tests in 12 week old ( A ) and 24 week old ( C ) male Cpt1a CON versus Cpt1a Pdx1−/− mice with respective area under the curve (AUC) calculations ( B , D ). E , glucose- and KCl-stimulated insulin secretion in islets isolated from 12 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice with respective AUC calculations for glucose- ( F ) and KCl-stimulated time points ( G ), that is, time points highlighted in gray in panel E . H , insulin content in the same islets as shown in panel E . I , insulin positive area and ( J ) islet fraction calculated from formalin-fixed pancreatic tissue in 15 week old, 30 week old, and 52 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice. K , β-cell mass and ( L ) pancreas mass in 30 week old and 52 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice. Quantification of the number of Ki-67 + nuclei per islet in 30 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice treated with Vehicle or Cort for 1 week ( M ) or following a 1 week exposure to 10% or 45% fat purified diet ( N ). n = 10 to 12 ( A , B ), n = 15 to 28 ( C , D ), n = 5 ( E – H ), n = 14 ( I – L ), n = 10 ( M , N ); ns = not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001. CPT1A, carnitine palmitoyltransferase 1A.

Article Snippet: Mice with a pancreas-specific deletion of Cpt1a (Cpt1a Pdx1−/− ) were generated by crossing Cpt1a fl/fl mice (C57BL/6- Cpt1a tm1.1mrl Model # 9759 from Taconic) with Pdx1-Cre mice (Stock # 014647; Jackson Laboratory, Bar Harbor, ME).

Techniques: Cell Function Assay, Isolation, Purification

Journal: The Journal of Biological Chemistry

Article Title: Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion

doi: 10.1016/j.jbc.2025.108187

Figure Lengend Snippet: CPT1a activity is required for the lipid-mediated potentiation of GSIS, and activity is not altered by dietary fat content. A and B , serum triglycerides (TGs) were assessed following oral gavage of a 20% intralipid emulsion in 15 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice. C and D , glucose tolerance and ( E , F ) serum insulin were determined in 15 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice given an oral gavage of saline or 20% intralipid emulsion at −120 min, followed by intraperitoneal injection of glucose at 0 min. Time points of lipid and glucose administration are indicated by blue arrows . G , body mass, ( H ) fat mass, ( I ) serum insulin, and ( J ) pancreatic triglyceride content were assessed in male Cpt1a CON versus Cpt1a Pdx1−/− mice on low- or high-fat diet for 8 weeks. K and L , glucose tolerance and ( M ) in vivo GSIS were assessed in male Cpt1a CON versus Cpt1a Pdx1−/− mice on low-fat diet for 8 weeks. N and O , glucose tolerance and ( P ) in vivo GSIS were assessed in male Cpt1a CON versus Cpt1a Pdx1−/− mice on high-fat diet for 8 weeks. n = 10 to 14 ( A – F ), n = 4 to 6 ( G – P ). ns = not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. LFD, low fat diet, HFD, high-fat diet; CPT1A, carnitine palmitoyltransferase 1A; GSIS, glucose-stimulated insulin secretion.

Article Snippet: Mice with a pancreas-specific deletion of Cpt1a (Cpt1a Pdx1−/− ) were generated by crossing Cpt1a fl/fl mice (C57BL/6- Cpt1a tm1.1mrl Model # 9759 from Taconic) with Pdx1-Cre mice (Stock # 014647; Jackson Laboratory, Bar Harbor, ME).

Techniques: Activity Assay, Emulsion, Saline, Injection, In Vivo

Journal: The Journal of Biological Chemistry

Article Title: Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion

doi: 10.1016/j.jbc.2025.108187

Figure Lengend Snippet: CPT1 A activity is dispensable for maintenance of β-cell function and glucose tolerance in female mice. A , A glucose tolerance test in 16 to 28 week old female Cpt1a CON versus Cpt1a Pdx1−/− mice with respective AUC calculation ( B ). C , glucose- and KCl-stimulated insulin secretion in islets isolated from 30 week old female Cpt1a CON versus Cpt1a Pdx1−/− mice with respective AUC calculations for glucose- and KCl-stimulated time points ( D ), that is, time points highlighted in gray in panel C . E , insulin content in the same islets as shown in panel C . F , fasting serum insulin in 15 week old and 30 week old female Cpt1a CON versus Cpt1a Pdx1−/− mice. G , insulin-positive area and ( H ) islet fraction calculated from formalin-fixed paraffin-embedded pancreatic tissue in 30 week old female Cpt1a CON versus Cpt1a Pdx1−/− mice. I , Ins1, Ins2, Mafa, and Nkx6.1 mRNA levels in isolated islets from 30 week old female Cpt1a CON versus Cpt1a Pdx1−/− mice. n = 23 to 28 ( A and B ), n = 3 to 5 ( C – E ), n = 7 ( F – I ). ns = not significant, ∗ p < 0.05 and ∗∗ p < 0.01. CPT1A, carnitine palmitoyltransferase 1A; AUC, area under the curve.

Article Snippet: Mice with a pancreas-specific deletion of Cpt1a (Cpt1a Pdx1−/− ) were generated by crossing Cpt1a fl/fl mice (C57BL/6- Cpt1a tm1.1mrl Model # 9759 from Taconic) with Pdx1-Cre mice (Stock # 014647; Jackson Laboratory, Bar Harbor, ME).

Techniques: Activity Assay, Cell Function Assay, Isolation, Formalin-fixed Paraffin-Embedded

Journal: The Journal of Biological Chemistry

Article Title: Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion

doi: 10.1016/j.jbc.2025.108187

Figure Lengend Snippet: Markers of β-cell maturity are unchanged in male mice with a pancreatic reduction in CPT1A activity. A , Ins1, Ins2, Mafa, Nkx6.1, Pdx1, and Slc2a2 mRNA levels in isolated islets from 30 week male Cpt1a CON versus Cpt1a Pdx1−/− mice. B , immunofluorescence staining for insulin ( green ) costaining with MafA ( red ), Nkx6.1 ( red ; C ), and glucagon ( red ; F ) in formalin-fixed paraffin-embedded (FFPE) pancreatic tissue from 30 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice. D , fasting serum insulin in 12 week old, 30 week old, and 52 week old, ( E ) fasting serum proinsulin:C-peptide ratio, and ( G ) fasting serum glucagon in 30 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice. H , glucose-stimulated glucagon secretion, and ( I ) fold change in glucose-stimulated glucagon secretion, in islets isolated from 12 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice. n = 7 to 8 ( A ), n = 5; the scale bar represents 100 μm ( B , C , and F ), n = 14 to 24 ( D ), n = 6 ( E ), n = 10 to 15 ( G ), and n = 4 ( H and I ). ns = not significant, ∗∗ p < 0.01, ∗∗∗ p < 0.001. CPT1A, carnitine palmitoyltransferase 1A.

Article Snippet: Mice with a pancreas-specific deletion of Cpt1a (Cpt1a Pdx1−/− ) were generated by crossing Cpt1a fl/fl mice (C57BL/6- Cpt1a tm1.1mrl Model # 9759 from Taconic) with Pdx1-Cre mice (Stock # 014647; Jackson Laboratory, Bar Harbor, ME).

Techniques: Activity Assay, Isolation, Immunofluorescence, Staining, Formalin-fixed Paraffin-Embedded

Journal: The Journal of Biological Chemistry

Article Title: Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion

doi: 10.1016/j.jbc.2025.108187

Figure Lengend Snippet: GLUT2 abundance is unchanged, but the dedifferentiation marker, ALDH1A3, is elevated in β-cells from male, but not female, mice deficient in CPT1a. A , immunofluorescence staining for insulin ( green ) costaining with GLUT2 in red in FFPE tissue from 30 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice and ( B ) 30 week old male db/+ versus db/db mice. C , GLUT2 abundance was determined in islets isolated from Cpt1a CON versus Cpt1a Pdx1−/− and db/+ versus db/db mice. Quantifications are shown underneath immunoblots using a nonspecific protein detected by the GLUT2 antibody as the loading control. Islets from 2 to 3 mice were pooled and run in triplicate ( top panel ) or duplicate ( bottom panel ). D , immunofluorescence staining for insulin ( green ) costaining with ALDH1A3 ( red ) in FFPE tissue from 15, 30, and 52 week old male and 30 week old female Cpt1a CON versus Cpt1a Pdx1−/− mice. E , 832/13 insulinoma cells either untreated (No Tx) or treated for 8 h with 2 μM camptothecin (Tx). Cells were fixed in 2% formaldehyde followed by immunofluorescence staining for cleaved caspase 3 (CC3). F , immunofluorescence staining for insulin ( green ) and cleaved caspase 3 ( red ) in FFPE tissue from 52 week old male Cpt1a CON versus Cpt1a Pdx1−/− mice. n = 5 per genotype (A-B and D, F). Scale bars represent 100 μm, inset scale bar represents 20 μm. M = male and F = female. ns = not significant, ∗ p < 0.05. CPT1A, carnitine palmitoyltransferase 1A; FFPE, formalin-fixed, paraffin-embedded; GLUT2, glucose transporter 2.

Article Snippet: Mice with a pancreas-specific deletion of Cpt1a (Cpt1a Pdx1−/− ) were generated by crossing Cpt1a fl/fl mice (C57BL/6- Cpt1a tm1.1mrl Model # 9759 from Taconic) with Pdx1-Cre mice (Stock # 014647; Jackson Laboratory, Bar Harbor, ME).

Techniques: Marker, Immunofluorescence, Staining, Isolation, Western Blot, Control, Formalin-fixed Paraffin-Embedded

Journal: The Journal of Biological Chemistry

Article Title: Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion

doi: 10.1016/j.jbc.2025.108187

Figure Lengend Snippet: Minimal changes in global transcripts or mitochondrial content, but reduced ATP production, is observed in male Cpt1a Pdx1−/− mice. A , volcano plot showing differentially expressed genes in isolated islets from male Cpt1a CON versus Cpt1a Pdx1−/− mice, n = 3. B , mitochondrial number was calculated using the ratio of mitochondrial DNA (MT-CO1 or MT-ND1) relative to nuclear DNA (Sdha or Ndufa2) in genomic DNA isolated from male Cpt1a CON versus Cpt1a Pdx1−/− mice, n = 8. C , immunoblot analysis of total OXPHOS in isolated islets from male Cpt1a CON versus Cpt1a Pdx1−/− mice. Each band represents islets pooled from two animals. Blot repeated on three separate occasions. D , intracellular measurement of ATP levels using 50 islets from male and female Cpt1a CON versus Cpt1a Pdx1−/− mice between 15 and 30 weeks of age, n = 7 to 10. ns = not significant, ∗∗ p < 0.01. CPT1A, carnitine palmitoyltransferase 1A.

Article Snippet: Mice with a pancreas-specific deletion of Cpt1a (Cpt1a Pdx1−/− ) were generated by crossing Cpt1a fl/fl mice (C57BL/6- Cpt1a tm1.1mrl Model # 9759 from Taconic) with Pdx1-Cre mice (Stock # 014647; Jackson Laboratory, Bar Harbor, ME).

Techniques: Isolation, Western Blot

Journal: Molecular Metabolism

Article Title: Genome-wide analysis of PDX1 target genes in human pancreatic progenitors

doi: 10.1016/j.molmet.2018.01.011

Figure Lengend Snippet: Efficient differentiation of XM001 iPSCs into pancreatic progenitors . (A) Schematic of iPSC-derived pancreatic progenitor differentiation protocol. (B) Immunostaining for FOXA2 and SOX17 on day 3. Scale bar indicates 50 μm. (C) Representative FACS plot of SOX17 + cells at DE stage. A differentiated sample stained with only the secondary antibody and XM001 iPSCs were used as negative controls. (D) FACS quantification of the percentage of SOX17 + cells at DE stage (n = 3). (E) Immunostaining for PDX1 on day 10. Scale bar indicates 50 μm. (F) Representative FACS plot of PDX1 + cells at the PP stage. A differentiated sample stained with only the secondary antibody and XM001 iPSCs were used as negative controls. (G) FACS quantification of the percentage of PDX1 + cells at PP stage (n = 3).

Article Snippet: The cells were incubated with goat anti-SOX17 antibody (1:500, #GT15094, Acris/Novus) and goat anti-PDX1 antibody (1:500, #AF2419, R&D Systems) for 30 min at room temperature and then stained with Alexa Fluor 555-conjugated donkey antibody directed against goat (1:500, #A21432, Invitrogen) for 30 min at room temperature.

Techniques: Derivative Assay, Immunostaining, Staining

Journal: Molecular Metabolism

Article Title: Genome-wide analysis of PDX1 target genes in human pancreatic progenitors

doi: 10.1016/j.molmet.2018.01.011

Figure Lengend Snippet: Characterization of PDX1 binding in XM001 PP cells . (A) ChIP-seq data tracks showing the enrichment of H3K27ac (blue) and PDX1 (red) at the loci of important pancreatic genes. (B) Average ChIP-seq Signal of H3K27ac (blue) and PDX1 (red) at PDX1 binding sites shows enrichment of H3K27ac at PDX1-bound sites. (C) Distribution of PDX1 binding sites among genomic features. PDX1 binds predominantly to intergenic, intronic, and promoter regions. (D) Meta-genomic plot of the enrichment of PDX1 at the transcriptional start sites (TSS) of its target genes displayed as binding sites per base pair (bp) per gene over the genomic regions of all RefSeq genes. (E) Most enriched motif discovered by de novo motif analysis resembles the known PDX1 consensus sequence and is identified in 62.2% of all PDX1-bound sequences.

Article Snippet: The cells were incubated with goat anti-SOX17 antibody (1:500, #GT15094, Acris/Novus) and goat anti-PDX1 antibody (1:500, #AF2419, R&D Systems) for 30 min at room temperature and then stained with Alexa Fluor 555-conjugated donkey antibody directed against goat (1:500, #A21432, Invitrogen) for 30 min at room temperature.

Techniques: Binding Assay, ChIP-sequencing, Sequencing

Journal: Molecular Metabolism

Article Title: Genome-wide analysis of PDX1 target genes in human pancreatic progenitors

doi: 10.1016/j.molmet.2018.01.011

Figure Lengend Snippet: Functional characterization of PDX1-bound genes . (A) Venn diagram depicting the overlap of PDX1-bound genes and the differentially expressed genes identified by microarray analysis. (B) MA plot showing the log2 fold change over the mean log2 expression of PDX1-bound genes. Differentially expressed genes are displayed in color. Green indicates enrichment in PPs, whereas brown indicates enrichment in iPSCs. (C) Bar chart of log10 p-values from enriched GO terms and KEGG and Reactome pathways of PDX1-bound genes upregulated in PPs. (D) Venn diagram showing the overlap of high confidence PDX1 binding sites from adult human islets and PPs and some annotated genes. (E) Bar chart of log10 p-values from enriched gene ontology and KEGG/Reactome pathways from genes bound in adult islets and/or PPs.

Article Snippet: The cells were incubated with goat anti-SOX17 antibody (1:500, #GT15094, Acris/Novus) and goat anti-PDX1 antibody (1:500, #AF2419, R&D Systems) for 30 min at room temperature and then stained with Alexa Fluor 555-conjugated donkey antibody directed against goat (1:500, #A21432, Invitrogen) for 30 min at room temperature.

Techniques: Functional Assay, Microarray, Expressing, Binding Assay

Journal: Molecular Metabolism

Article Title: Genome-wide analysis of PDX1 target genes in human pancreatic progenitors

doi: 10.1016/j.molmet.2018.01.011

Figure Lengend Snippet: Analysis of T2DM SNPs in XM001 PPs and adult islets . (A) Venn diagram depicting the overlap of T2DM SNPs found in active regions of adult islets and XM001 PPs. (B) P-values of T2DM SNPs near known T2DM-associated genes found in active regions of adult islets , and XM001 PPs. P-values of SNPs bound by PDX1 in PPs and islets are shown as black circles and triangles, respectively. (C) ChIP-seq data tracks showing H3K27ac and PDX1 from PP and islet , , cells at the HNF1B locus. The SNP rs11263763 is located in a PP specific PDX1 binding site in the first intron of HNF1B. The PP specific enhancer regions are shaded in gray.

Article Snippet: The cells were incubated with goat anti-SOX17 antibody (1:500, #GT15094, Acris/Novus) and goat anti-PDX1 antibody (1:500, #AF2419, R&D Systems) for 30 min at room temperature and then stained with Alexa Fluor 555-conjugated donkey antibody directed against goat (1:500, #A21432, Invitrogen) for 30 min at room temperature.

Techniques: ChIP-sequencing, Binding Assay