recombinant mouse igf1 protein  (MedChemExpress)

Journal: BMC Medicine

Article Title: IGF1-mediated mesenchymal-endothelial transition as a potential regulatory target in calcific aortic valve disease

doi: 10.1186/s12916-025-04433-z

Figure Lengend Snippet: IGF1 identified as a key molecule mediating MEndT. A Conduct protein-protein interaction network analysis (PPI) on the overlapping differentially expressed genes between pVECs vs. pVICs and pVICs-OM8d vs. pVICs. B Analyze the FPKM values of SOD2, CCL2, CCN2, IGF1, and COL3A1 in the transcriptome sequencing of pVICs-OM8d vs. pVICs, normalized to the control pVICs group. Values are mean ± SD of 4 biological replicates. Statistical tests used: ANOVA. C , D WB ( C ) and qPCR ( D ) were used to detect the protein expression of IGF1, IGF1R, P-IGF1R, and the mRNA expression levels of IGF1, IGF1R in pVICs cultured for 8 days in GM or OM. D Normalized to pVICs and GAPDH. Values are mean ± SD of 3 independent experiments. Statistical tests used: ANOVA. E , F Statistical analysis of tube formation assays of pVICs with exogenous addition of IGF1 and BMS-536924 during OM induction ( E ), and after knockdown of IGF1 expression. Normalized to GM group ( F ). Normalized to pVICs. Values are mean ± SD. 3 biological replicates, with 3 random measurements within each replicate, n =9. Statistical tests used: ANOVA. G , H Assessed the changes in protein expression ( G ) of CDH5, CD31, α-SMA, PI3K, Akt, P-Akt, and HIF-1α in pVICs after the exogenous addition of recombinant IGF1 and inhibitor BMS-536924 in GM and OM, and changes in mRNA expression levels ( H ) of CDH5, CD31, α-SMA. H Normalized to pVICs and GAPDH. Values are mean ± SD of 3 independent experiments. Statistical tests used: ANOVA. I , J Detect the protein ( J ) and mRNA ( I ) expression of CDH5, CD31, α-SMA in pVICs after knockdown of IGF1 expression during OM induction. I Normalized to pVICs and GAPDH. Values are mean ± SD of 3 independent experiments. Statistical tests used: ANOVA. K , L Tube formation assays in pVICs with exogenous addition of IGF1 and BMS-536924 during OM induction, and after knockdown of IGF1 expression, scale bars: 200 µm

Article Snippet: Each group received intraperitoneal injections every 2 days with either saline (100 μl), recombinant mouse IGF1 protein (1 μM, 100 μl; MCE, HY-P7070), or BMS-536924 (1 mg; MCE, HY-10262).

Techniques: Sequencing, Control, Expressing, Cell Culture, Knockdown, Recombinant

Journal: BMC Medicine

Article Title: IGF1-mediated mesenchymal-endothelial transition as a potential regulatory target in calcific aortic valve disease

doi: 10.1186/s12916-025-04433-z

Figure Lengend Snippet: IGF1-mediated MEndT and disease progression in the AVWI mouse mode. A Schematic diagram of the animal experiment procedure. B Echocardiographic evaluation of the sham surgery group and the AVWI mouse groups with intraperitoneal injection of saline, IGF1, or BMS-536924. Parameters assessed included: aortic valve annulus diameter (mm) and transaortic peak velocity (mm/s). C , D Statistical analysis of the aortic valve annulus diameter (mm) and transaortic peak velocity (mm/s) in each group of mice. n =8. Values are mean ± SD. Statistical tests used: ANOVA. E HE staining (scale bars: 200 µm) and multiplex immunofluorescence histology (scale bars: 100 µm, 50 µm, or 20 µm) for CD31, tdTomato, and DAPI of mouse aortic valve paraffin sections. Yellow arrows indicate CD31-positive cells labeled with tdTomato. F , G Statistical analysis of the aortic valve thickness (µm) (HE staining, E ) and tdTomato labeled cells expressing CD31 within the aortic valve region (immunofluorescence staining, G ) in each group of mice. n =8. Values are mean ± SD. Statistical tests used: ANOVA

Article Snippet: Each group received intraperitoneal injections every 2 days with either saline (100 μl), recombinant mouse IGF1 protein (1 μM, 100 μl; MCE, HY-P7070), or BMS-536924 (1 mg; MCE, HY-10262).

Techniques: Biomarker Discovery, Injection, Saline, Staining, Multiplex Assay, Immunofluorescence, Labeling, Expressing

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: ( A-B ) Schematic illustrating derivation of FOXG1⁺ neural progenitor cells (NPCs) from human pluripotent stem cells using a 2D culture system. Immunostaining confirms co-expression of FOXG1 (red) and Nestin (green), with DAPI (blue) marking nuclei. ( C ) Gene Ontology (GO) enrichment analysis of FOXG1⁺ NPCs highlights processes including forebrain development, neuronal differentiation, and cell fate commitment. Dot size reflects the number of associated genes; color indicates false discovery rate (FDR). ( D ) STRING network analysis identifies enriched clusters involved in immune signaling, cell cycle regulation, and metabolic networks. ( E ) Pathway enrichment analysis shows strong activation of Insulin/IGF, mTOR, Notch, Neurotrophin, and MAPK signaling pathways. ( F ) Expression analysis reveals high levels of IGF1R and IGF2R, absent IGF1 ligand, and selective expression of Relaxin receptors, suggesting reliance on paracrine IGF1 signaling. ( G ) Expression of IGFBP2, IGFBPL1, IRS1–3, and regulators of IGF bioavailability and insulin signaling is detected, favoring IGF axis competence. ( H ) Schematic of rosette-derived neural aggregate (RONA) culture system showing 3D tissue organization supported by Matrigel and nutrient exchange. ( I-L ) Immunostaining of RONAs reveals central IGF1 expression (green), peripheral CK8⁺ epithelial cells (red), and FOXG1⁺ NPCs co-expressing IGF1R at the plasma membrane. ( M ) Quantitative co-localization analysis shows substantial overlap between FOXG1 and IGF1R (Manders’ coefficients ∼0.6). ( N ) Working model proposing that IGF1⁺ niche cells secrete IGF1 to regulate neighboring FOXG1⁺ progenitors in RONAs.

Article Snippet: Recombinant human IGF1 (PeproTech, Cat# 100-11) at 100 ng/mL, IGF1-neutralizing antibody (R&D Systems, Cat# AF-291-NA), or IGF1R antibody (Abcam, Cat# ab9572) were added to designated cultures to assess ligand dependency.

Techniques: Immunostaining, Expressing, Activation Assay, Protein-Protein interactions, Derivative Assay, Clinical Proteomics, Membrane

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: (A-B) Brightfield imaging of RONAs shows regionalized progenitor (S1) and niche (S2) zones under control, IGF1 supplementation, IGF1 neutralization, and IGF1R blockade conditions. IGF1 maintains or enhances the S1/S2 ratio, while IGF1 or IGF1R blockade significantly reduces it (*p < 0.05). (C-D) Top and side-view images reveal that IGF1 promotes vertical tissue expansion, whereas IGF1 or IGF1R neutralization leads to thinner aggregates. (E) Quantification of aggregate thickness shows IGF1 significantly increases vertical growth compared to control, while neutralization treatments reduce it. (F-G) Neurosphere assays demonstrate that IGF1 enhances the size but not the total number of neurospheres, supporting its role in promoting clonal expansion. (H-I) BrdU incorporation assays show that IGF1 increases proliferation of FOXG1⁺ NPCs, while IGF1 or IGF1R blockade decreases BrdU⁺ cell fractions (*p < 0.05). (J-K) Developmental analysis reveals high IGF1R expression during early human fetal brain stages, correlating strongly with FOXG1 expression (r = 0.81). (L-O) Western blot and densitometry analyses demonstrate that IGF1 supplementation sustains FOXG1, Pax6, and Hes1 expression over time, supporting progenitor maintenance and survival.

Article Snippet: Recombinant human IGF1 (PeproTech, Cat# 100-11) at 100 ng/mL, IGF1-neutralizing antibody (R&D Systems, Cat# AF-291-NA), or IGF1R antibody (Abcam, Cat# ab9572) were added to designated cultures to assess ligand dependency.

Techniques: Imaging, Control, Neutralization, BrdU Incorporation Assay, Expressing, Western Blot

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: (A-C) Western blot analysis shows that IGF1 rapidly and robustly induces ERK phosphorylation within 10 minutes, sustained up to 60 minutes. Total ERK levels remain stable. (D-F) IGF1 stimulation also induces Akt phosphorylation, peaking between 10–60 minutes, while total Akt levels modestly decline. (G-I) IGF1 enhances phosphorylation of S6, a downstream mTORC1 target, with peak activation at 30–60 minutes and stable total S6 levels. (J-P) BrdU incorporation assays combined with pathway-specific inhibitors demonstrate that PI3K, AKT, and mTOR pathways are essential for IGF1-driven proliferation. MEK/ERK inhibition partially reduces proliferation, indicating a secondary but supportive role.

Article Snippet: Recombinant human IGF1 (PeproTech, Cat# 100-11) at 100 ng/mL, IGF1-neutralizing antibody (R&D Systems, Cat# AF-291-NA), or IGF1R antibody (Abcam, Cat# ab9572) were added to designated cultures to assess ligand dependency.

Techniques: Western Blot, Phospho-proteomics, Activation Assay, BrdU Incorporation Assay, Inhibition

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: (A) Workflow illustrating ribosome profiling (Ribo-Seq) in cultured FOXG1⁺ neural progenitors, including cycloheximide treatment, ribosome-protected fragment (RPF) isolation, and sequencing. (B-C) Cumulative distribution plots show that IGF1 broadly enhances translation efficiency transcriptome-wide, while Torin1 attenuates this effect, particularly for known mTOR targets. (D) KEGG pathway enrichment identifies ribosome biogenesis, oxidative phosphorylation, proteasome function, and neurodegenerative pathways (e.g., Parkinson’s, Huntington’s) among IGF1-upregulated translational targets. (E-F) GO biological processes enriched among IGF1-upregulated transcripts include cytoplasmic translation, ribonucleoprotein biogenesis, and macromolecule biosynthesis. (G) IGF1-responsive translational programs prominently involve proteostasis mechanisms such as ubiquitin-dependent protein catabolism and multivesicular body transport. (H) IGF1 stimulation enhances translation of mitochondrial pathways, including the electron transport chain and oxidative phosphorylation. (I) IGF1 promotes translation of genes involved in genome stability, including DNA repair, chromatin remodeling, and cell cycle checkpoints.

Article Snippet: Recombinant human IGF1 (PeproTech, Cat# 100-11) at 100 ng/mL, IGF1-neutralizing antibody (R&D Systems, Cat# AF-291-NA), or IGF1R antibody (Abcam, Cat# ab9572) were added to designated cultures to assess ligand dependency.

Techniques: Cell Culture, Isolation, Sequencing, Phospho-proteomics, Ubiquitin Proteomics

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: (A) Gene Ontology and pathway enrichment analysis identifies translational programs regulated by IGF1, including nervous system development, axon guidance, ribosome biogenesis, cytoplasmic translation, lipoprotein metabolism, and neuronal maturation pathways (e.g., SLIT– ROBO signaling). (B-C) Polysome fractionation analysis shows that IGF1 promotes GSX1 and ACTB mRNA association with heavier polysome fractions, indicating enhanced translational engagement, while Torin1 treatment shifts transcripts toward lighter fractions. (D) Bar graph quantifying polysome-to-monosome ratios reveals that IGF1 significantly enhances global translational output, while mTOR inhibition (Torin1) suppresses this effect. (E) 5’UTR reporter assays demonstrate that luciferase reporters bearing the GSX1, eEF2, or ACTB 5’UTRs exhibit reduced translation upon Torin1 treatment, with GSX1 5’UTR showing the greatest sensitivity, indicating 5’UTR-dependent mTOR control. (F-G) Immunofluorescence images and quantification show that expression of a GSX1 5’UTR– luciferase reporter reduces BrdU incorporation compared to control reporters, suggesting translational repression impairs progenitor proliferation. (H-I) Brightfield and GFP imaging reveal that GSX1 5’UTR–GFP constructs significantly reduce GFP intensity compared to control, confirming strong 5’UTR-mediated translational repression affecting protein output in neural progenitor colonies.

Article Snippet: Recombinant human IGF1 (PeproTech, Cat# 100-11) at 100 ng/mL, IGF1-neutralizing antibody (R&D Systems, Cat# AF-291-NA), or IGF1R antibody (Abcam, Cat# ab9572) were added to designated cultures to assess ligand dependency.

Techniques: Fractionation, Inhibition, Luciferase, Control, Immunofluorescence, Expressing, BrdU Incorporation Assay, Imaging, Construct

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: ( A-B ) Schematic illustrating derivation of FOXG1⁺ neural progenitor cells (NPCs) from human pluripotent stem cells using a 2D culture system. Immunostaining confirms co-expression of FOXG1 (red) and Nestin (green), with DAPI (blue) marking nuclei. ( C ) Gene Ontology (GO) enrichment analysis of FOXG1⁺ NPCs highlights processes including forebrain development, neuronal differentiation, and cell fate commitment. Dot size reflects the number of associated genes; color indicates false discovery rate (FDR). ( D ) STRING network analysis identifies enriched clusters involved in immune signaling, cell cycle regulation, and metabolic networks. ( E ) Pathway enrichment analysis shows strong activation of Insulin/IGF, mTOR, Notch, Neurotrophin, and MAPK signaling pathways. ( F ) Expression analysis reveals high levels of IGF1R and IGF2R, absent IGF1 ligand, and selective expression of Relaxin receptors, suggesting reliance on paracrine IGF1 signaling. ( G ) Expression of IGFBP2, IGFBPL1, IRS1–3, and regulators of IGF bioavailability and insulin signaling is detected, favoring IGF axis competence. ( H ) Schematic of rosette-derived neural aggregate (RONA) culture system showing 3D tissue organization supported by Matrigel and nutrient exchange. ( I-L ) Immunostaining of RONAs reveals central IGF1 expression (green), peripheral CK8⁺ epithelial cells (red), and FOXG1⁺ NPCs co-expressing IGF1R at the plasma membrane. ( M ) Quantitative co-localization analysis shows substantial overlap between FOXG1 and IGF1R (Manders’ coefficients ∼0.6). ( N ) Working model proposing that IGF1⁺ niche cells secrete IGF1 to regulate neighboring FOXG1⁺ progenitors in RONAs.

Article Snippet: Human FOXG1+ NPCs were cultured under growth factor/insulin-deprived conditions for 16 hours before treatment with 20 ng/mL recombinant human IGF1 (PeproTech, #100-11) for 30 minutes, or pre-treated with 250 nM Torin 1 (Tocris, #4247) for 30 minutes followed by 30 minutes of IGF1 in the continued presence of Torin 1.

Techniques: Immunostaining, Expressing, Activation Assay, Protein-Protein interactions, Derivative Assay, Clinical Proteomics, Membrane

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: (A-B) Brightfield imaging of RONAs shows regionalized progenitor (S1) and niche (S2) zones under control, IGF1 supplementation, IGF1 neutralization, and IGF1R blockade conditions. IGF1 maintains or enhances the S1/S2 ratio, while IGF1 or IGF1R blockade significantly reduces it (*p < 0.05). (C-D) Top and side-view images reveal that IGF1 promotes vertical tissue expansion, whereas IGF1 or IGF1R neutralization leads to thinner aggregates. (E) Quantification of aggregate thickness shows IGF1 significantly increases vertical growth compared to control, while neutralization treatments reduce it. (F-G) Neurosphere assays demonstrate that IGF1 enhances the size but not the total number of neurospheres, supporting its role in promoting clonal expansion. (H-I) BrdU incorporation assays show that IGF1 increases proliferation of FOXG1⁺ NPCs, while IGF1 or IGF1R blockade decreases BrdU⁺ cell fractions (*p < 0.05). (J-K) Developmental analysis reveals high IGF1R expression during early human fetal brain stages, correlating strongly with FOXG1 expression (r = 0.81). (L-O) Western blot and densitometry analyses demonstrate that IGF1 supplementation sustains FOXG1, Pax6, and Hes1 expression over time, supporting progenitor maintenance and survival.

Article Snippet: Human FOXG1+ NPCs were cultured under growth factor/insulin-deprived conditions for 16 hours before treatment with 20 ng/mL recombinant human IGF1 (PeproTech, #100-11) for 30 minutes, or pre-treated with 250 nM Torin 1 (Tocris, #4247) for 30 minutes followed by 30 minutes of IGF1 in the continued presence of Torin 1.

Techniques: Imaging, Control, Neutralization, BrdU Incorporation Assay, Expressing, Western Blot

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: (A-C) Western blot analysis shows that IGF1 rapidly and robustly induces ERK phosphorylation within 10 minutes, sustained up to 60 minutes. Total ERK levels remain stable. (D-F) IGF1 stimulation also induces Akt phosphorylation, peaking between 10–60 minutes, while total Akt levels modestly decline. (G-I) IGF1 enhances phosphorylation of S6, a downstream mTORC1 target, with peak activation at 30–60 minutes and stable total S6 levels. (J-P) BrdU incorporation assays combined with pathway-specific inhibitors demonstrate that PI3K, AKT, and mTOR pathways are essential for IGF1-driven proliferation. MEK/ERK inhibition partially reduces proliferation, indicating a secondary but supportive role.

Article Snippet: Human FOXG1+ NPCs were cultured under growth factor/insulin-deprived conditions for 16 hours before treatment with 20 ng/mL recombinant human IGF1 (PeproTech, #100-11) for 30 minutes, or pre-treated with 250 nM Torin 1 (Tocris, #4247) for 30 minutes followed by 30 minutes of IGF1 in the continued presence of Torin 1.

Techniques: Western Blot, Phospho-proteomics, Activation Assay, BrdU Incorporation Assay, Inhibition

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: (A) Workflow illustrating ribosome profiling (Ribo-Seq) in cultured FOXG1⁺ neural progenitors, including cycloheximide treatment, ribosome-protected fragment (RPF) isolation, and sequencing. (B-C) Cumulative distribution plots show that IGF1 broadly enhances translation efficiency transcriptome-wide, while Torin1 attenuates this effect, particularly for known mTOR targets. (D) KEGG pathway enrichment identifies ribosome biogenesis, oxidative phosphorylation, proteasome function, and neurodegenerative pathways (e.g., Parkinson’s, Huntington’s) among IGF1-upregulated translational targets. (E-F) GO biological processes enriched among IGF1-upregulated transcripts include cytoplasmic translation, ribonucleoprotein biogenesis, and macromolecule biosynthesis. (G) IGF1-responsive translational programs prominently involve proteostasis mechanisms such as ubiquitin-dependent protein catabolism and multivesicular body transport. (H) IGF1 stimulation enhances translation of mitochondrial pathways, including the electron transport chain and oxidative phosphorylation. (I) IGF1 promotes translation of genes involved in genome stability, including DNA repair, chromatin remodeling, and cell cycle checkpoints.

Article Snippet: Human FOXG1+ NPCs were cultured under growth factor/insulin-deprived conditions for 16 hours before treatment with 20 ng/mL recombinant human IGF1 (PeproTech, #100-11) for 30 minutes, or pre-treated with 250 nM Torin 1 (Tocris, #4247) for 30 minutes followed by 30 minutes of IGF1 in the continued presence of Torin 1.

Techniques: Cell Culture, Isolation, Sequencing, Phospho-proteomics, Ubiquitin Proteomics

Journal: bioRxiv

Article Title: Spatially Organized IGF1-mTOR Signaling Controls Human Forebrain Progenitor Fate Through Coordinated Transcriptional and Translational Programs

doi: 10.1101/2025.05.08.652851

Figure Lengend Snippet: (A) Gene Ontology and pathway enrichment analysis identifies translational programs regulated by IGF1, including nervous system development, axon guidance, ribosome biogenesis, cytoplasmic translation, lipoprotein metabolism, and neuronal maturation pathways (e.g., SLIT– ROBO signaling). (B-C) Polysome fractionation analysis shows that IGF1 promotes GSX1 and ACTB mRNA association with heavier polysome fractions, indicating enhanced translational engagement, while Torin1 treatment shifts transcripts toward lighter fractions. (D) Bar graph quantifying polysome-to-monosome ratios reveals that IGF1 significantly enhances global translational output, while mTOR inhibition (Torin1) suppresses this effect. (E) 5’UTR reporter assays demonstrate that luciferase reporters bearing the GSX1, eEF2, or ACTB 5’UTRs exhibit reduced translation upon Torin1 treatment, with GSX1 5’UTR showing the greatest sensitivity, indicating 5’UTR-dependent mTOR control. (F-G) Immunofluorescence images and quantification show that expression of a GSX1 5’UTR– luciferase reporter reduces BrdU incorporation compared to control reporters, suggesting translational repression impairs progenitor proliferation. (H-I) Brightfield and GFP imaging reveal that GSX1 5’UTR–GFP constructs significantly reduce GFP intensity compared to control, confirming strong 5’UTR-mediated translational repression affecting protein output in neural progenitor colonies.

Article Snippet: Human FOXG1+ NPCs were cultured under growth factor/insulin-deprived conditions for 16 hours before treatment with 20 ng/mL recombinant human IGF1 (PeproTech, #100-11) for 30 minutes, or pre-treated with 250 nM Torin 1 (Tocris, #4247) for 30 minutes followed by 30 minutes of IGF1 in the continued presence of Torin 1.

Techniques: Fractionation, Inhibition, Luciferase, Control, Immunofluorescence, Expressing, BrdU Incorporation Assay, Imaging, Construct

Journal: Science Advances

Article Title: Hedgehog-interacting protein orchestrates alveologenesis and protects against bronchopulmonary dysplasia and emphysema

doi: 10.1126/sciadv.adu2958

Figure Lengend Snippet: ( A ) IF analysis of SMA in the alveoli of Hhip -deleted (HHIP CKO) and control mice at P14. ( B ) Number of myofibroblasts per unit alveolar area of Hhip -deleted and control mice. ( C and D ) UMAP showing cell clusters in the lung fibroblasts of Hhip -deleted and control mice at P14. ( E ) Violin plots showing the expression of Pdgfra , Acta2 , Hhip , and Cdh4 in alveolar myofibroblasts (ALMF), ductal myofibroblasts (DMF), peribronchial fibroblasts (Perib), adventitial fibroblasts (Adv), and alveolar fibroblasts (Alv). ( F ) Expression of Acta2 , Myh11 , Tagln , and Igf1 in ALMFs and DMFs of Hhip -deleted and control mice. ( G ) qPCR analysis of Igf1 expression in the lung stromal cells isolated from Hhip -deleted and control mice. ( H ) Analysis of Igf1 (RNA in situ ) and SMA expression in the alveoli. ( I ) Number of SMA + Igf1 + cells per unit alveolar area of Hhip -deleted and control mice at P14. ( J ) Percentage of SMA + Igf1 + cells in total Igf1 + cells. ( K ) Top 10 activated pathways in Hhip -deleted, relative to control myofibroblasts, analyzed with IPA. ( L ) qPCR analysis of Gli1 , Igf1 , and Acta2 expression in the lung stromal cells treated with PBS, SHH, and SHH plus HHIP. ( M ) qPCR analysis of Acta2 expression in SHH-stimulated lung stromal cells treated with vehicle or IGF1R inhibitor. ( N ) IF analysis of SMA expression in the alveoli of Hhip -deleted mice administered with vehicle or IGF1R inhibitor. ( O ) Number of myofibroblasts per unit alveolar area of Hhip -deleted mice administered with vehicle or IGF1R inhibitor. All in vitro experiments have been repeated at least one time with consistent results for validation. Each data point represents one mouse [(B), (G), (I), (J), and (O)] of an individual experiment. Data are expressed as mean ± SD. * P < 0.05, ** P < 0.005, *** P < 0.0005, and **** P < 0.0001.

Article Snippet: For IGF-1 treatment, the confluent fibroblasts were cultured in 0.5% FBS with 1% antibiotic-antimycotic, and recombinant murine IGF1 (catalog no. 250-19; PeproTech) was added at 50 ng/ml.

Techniques: Control, Expressing, Isolation, In Situ, In Vitro, Biomarker Discovery

Journal: Science Advances

Article Title: Hedgehog-interacting protein orchestrates alveologenesis and protects against bronchopulmonary dysplasia and emphysema

doi: 10.1126/sciadv.adu2958

Figure Lengend Snippet: ( A ) Violin plots showing the expression of Cdkn1a in ALMFs and DMFs of Hhip -deleted and control mice. ( B ) Senescence β-galactosidase staining of Hhip -deleted and control lungs. ( C ) IF analysis of SMA, p21, and CDH4 in Hhip -deleted and control lungs. Arrow: p21 + DMFs; arrowhead: p21 + ALMFs. ( D and E ) Number of p21 + ALMFs (D) and DMFs (E) per unit alveolar area of Hhip -deleted and control mice at P14. ( F ) IF analysis of SPC and p21 in Hhip -deleted and control lungs. Arrow: p21 + SPC + cells. ( G ) Percentage of p21 + cells in AT2s. ( H ) Activation of senescence pathways in the AT2s of Hhip -deleted mice, relative to control AT2s, and analyzed with IPA. ( I ) Top 5 upstream regulators in the AT2s of Hhip -deleted mice, relative to control AT2s, analyzed with IPA. ( J ) AT2 organoids cocultured with lung stromal cells ( R26R SmoM2/+ ) pre-infected with adenovirus-empty and adenovirus-Cre, treated with anti-IGF1 antibody and IgG. ( K and L ) Quantification of colony-forming efficiency (CFE) and organoid size. ( M ) IF analysis of SPC and p21 in AT2 organoids. Arrow: p21 + SPC + cells. ( N ) Percentage of p21 + cells in AT2s in the organoid assay. All in vitro experiments have been repeated at least one time with consistent results for validation. Each data point represents one mouse [(D), (E), and (G)] of an individual experiment. Data are expressed as Mean ± SD. * P < 0.05, ** P < 0.005, *** P < 0.0005, and **** P < 0.0001.

Article Snippet: For IGF-1 treatment, the confluent fibroblasts were cultured in 0.5% FBS with 1% antibiotic-antimycotic, and recombinant murine IGF1 (catalog no. 250-19; PeproTech) was added at 50 ng/ml.

Techniques: Expressing, Control, Staining, Activation Assay, Infection, In Vitro, Biomarker Discovery

Journal: Science Advances

Article Title: Hedgehog-interacting protein orchestrates alveologenesis and protects against bronchopulmonary dysplasia and emphysema

doi: 10.1126/sciadv.adu2958

Figure Lengend Snippet: ( A ) snRNA-seq analysis of GLI1 , PATCH1 , IGF1 , and ACTA2 expression in ALMFs and DMFs of human BPD. ( B ) H&E images of the lungs of hyperoxia-treated and control mice. ( C ) Quantification of MLI of hyperoxia-treated and control lungs. ( D ) qPCR analysis of Hhip , Gli1 , and Igf1 expression in the lung stromal cells isolated from hyperoxia-treated and control mice. ( E ) IF analysis of SMA in the lungs of hyperoxia-treated and control mice. ( F ) Histology quantification of the number of myofibroblasts of hyperoxia-treated and control mice. ( G ) qPCR analysis of Acta2 expression in the lung stromal cells isolated from hyperoxia-treated and control mice. ( H ) qPCR analysis of Cdkn1a expression in the lung epithelial cells isolated from hyperoxia-treated and control mice. ( I ) IF analysis of p21 and SPC in the lungs of hyperoxia-treated and control mice. Arrow: p21 + SPC + cells. ( J and K ) Quantification of the percentage of p21 + AT2s and number of AT2s in hyperoxia-treated and control mice. Each data point represents one mouse [(C), (D), (F) to (H), (J), and (K)] of an individual experiment. Data are expressed as mean ± SD. * P < 0.05, ** P < 0.005, and **** P < 0.0001.

Article Snippet: For IGF-1 treatment, the confluent fibroblasts were cultured in 0.5% FBS with 1% antibiotic-antimycotic, and recombinant murine IGF1 (catalog no. 250-19; PeproTech) was added at 50 ng/ml.

Techniques: Expressing, Control, Isolation

Journal: Science Advances

Article Title: Hedgehog-interacting protein orchestrates alveologenesis and protects against bronchopulmonary dysplasia and emphysema

doi: 10.1126/sciadv.adu2958

Figure Lengend Snippet: ( A ) Design strategy for HHIP-Fc recombinant protein and its analysis by Western blotting. ( B ) qPCR analysis of Gli1 expression in the lung stromal cells isolated from neonatal mice 3 and 5 days after one dose of HHIP or HHIP-Fc treatment. ( C ) H&E images of hyperoxia-exposed lungs, treated with HHIP-Fc or Fc fragment control. ( D ) MLI quantification of hyperoxia-exposed lungs, treated with HHIP-Fc or Fc. ( E and F ) IF analysis and quantification of myofibroblasts in hyperoxia-exposed lungs, treated with HHIP-Fc or Fc. ( G to I ) IF analysis and quantification of AT2 number and p21 + AT2 percentage in hyperoxia-exposed lungs, treated with HHIP-Fc or Fc. Arrow: p21 + SPC + cells. ( J ) qPCR analysis of Igf1 expression in the lung stromal cells isolated from hyperoxia-exposed mice, treated with HHIP-Fc or Fc. Each data point represents one mouse [(B), (D), (F), and (H) to (J)] of an individual experiment. Data are expressed as mean ± SD. * P < 0.05, ** P < 0.005, *** P < 0.0005, and **** P < 0.0001.

Article Snippet: For IGF-1 treatment, the confluent fibroblasts were cultured in 0.5% FBS with 1% antibiotic-antimycotic, and recombinant murine IGF1 (catalog no. 250-19; PeproTech) was added at 50 ng/ml.

Techniques: Recombinant, Western Blot, Expressing, Isolation, Control