Journal: Scientific Reports

Article Title: Follistatin like-1 aggravates silica-induced mouse lung injury

doi: 10.1038/s41598-017-00478-0

Figure Lengend Snippet: Up-regulation of Fstl1 in silica-injured mice and patients with silicosis. ( a ) Fstl1 mRNA expression in lung tissues of C57BL/6J mice at the different time-point after silica injury was determined by qRT-PCR analysis (n = 6 per group; * P < 0.05; ** P < 0.01 by one-way ANOVA followed by Student’s t test). ( b ) FSTL1 protein in lung tissues of C57BL/6J mice at the different time-point after silica injury was determined by western blot analysis. β-tubulin was used as a loading control. ( c ) Immunohistochemistry (IHC) of FSTL1 in lung sections of C57BL/6J mice 21 days after saline or silica injury. Representative images of the staining are shown. NA stands for normal area, FA stands for fibrotic area; both are shown at higher magnification. (n = 6 per group; scale bars, 200 μm). ( d ) Fstl1 mRNA expression in primary alveolar macrophages (AMs), alveolar epithelial cells (AECs) and fibroblasts (Fb) at day 21 after saline or silica injury was determined by qRT-PCR analysis (n = 6 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( e ) FSTL1 protein in serum of C57BL/6J mice at the different time-point after silica injury was determined by ELISA analysis (n = 6 per group; * P < 0.05; ** P < 0.01 by one-way ANOVA followed by Student’s t test). ( f ) Representative images of lung fibrotic area of patient with silicosis with H&E staining and immunohistochemical staining for FSTL1 protein (NA stands for normal area, FA stands for fibrotic area; Scale bars, 200 μm). ( g ) FSTL1 levels in serum of patients with silicosis and normal control individuals were determined by ELISA (* P < 0.05 by one-way ANOVA followed by Student’s t test).

Article Snippet: The plate was incubated at 4 °C overnight, then 2.5 μg/mL biotinylated human FSTL1 detection antibody (R&D system, MAB1694) was added followed by washing with wash buffer and the plate was incubated for 1 hour at room temperature.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Control, Immunohistochemistry, Saline, Staining, Enzyme-linked Immunosorbent Assay, Immunohistochemical staining

Journal: Scientific Reports

Article Title: Follistatin like-1 aggravates silica-induced mouse lung injury

doi: 10.1038/s41598-017-00478-0

Figure Lengend Snippet: Fstl1 +/− mice have an attenuated pulmonary fibrosis phenotype. ( a ) Western blot analysis of FSTL1 expression in lung tissues of Fstl1 +/− and WT mice 21 days after saline or silica injury. β-tubulin was used as a loading control. ( b ) H&E staining of lung sections of Fstl1 +/− and WT mice 21 days after silica injury. Representative images of the staining are shown. Arrows show inflammatory cells. (n = 6 per group; scale bars, 200 μm). ( c ) Lung fibrotic score analysis of the lung sections from Fstl1 +/− and WT mice 21 days after silica injury (n = 6 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). The fibrotic area is presented as a percentage. ( d ) Hydroxyproline contents in lung tissues from Fstl1 +/− and WT mice were measured 21 days after saline or silica injury (n = 6 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( e ) Masson trichrome staining of lung sections of Fstl1 +/− and WT mice 21 days after silica treatment. Representative images of the staining are shown (n = 6 per group; scale bars, 200 μm). ( f ) qRT-PCR analysis of Col1a1 mRNA expression in lung tissues from Fstl1 +/− and WT mice 21 days after saline or silica injury (n = 3 per group; *** P < 0.001 by one-way ANOVA followed by Student’s t test). ( g ) Western blot analysis of type I collagen (Col1) expression in lung tissues from Fstl1 +/− and WT mice 21 days after saline or silica injury (n = 3 per group). β-tubulin was used as a loading control. ( h ) qRT-PCR analysis of fibronectin ( Fn1 ) mRNA expression in lung tissues from Fstl1 +/− and WT mice 21 days after saline or silica injury (n = 3 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test).

Article Snippet: The plate was incubated at 4 °C overnight, then 2.5 μg/mL biotinylated human FSTL1 detection antibody (R&D system, MAB1694) was added followed by washing with wash buffer and the plate was incubated for 1 hour at room temperature.

Techniques: Western Blot, Expressing, Saline, Control, Staining, Quantitative RT-PCR

Journal: Scientific Reports

Article Title: Follistatin like-1 aggravates silica-induced mouse lung injury

doi: 10.1038/s41598-017-00478-0

Figure Lengend Snippet: Fstl1 +/− mice have an attenuated pulmonary inflammatory response. ( a–e ) Fstl1 +/− and their WT littermate mice were intratracheally exposured to saline or 200 mg/Kg silica, bronchoalveolar lavage fluid (BALF) were collected from Fstl1 +/− and WT mice 7 days after administration of saline or silica. ( a ) The number of total BALF cells was determined by hemocytometer (n = 7 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( b–d ) The differential cell counts in BALF were determined according to standard morphologic criteria. ( b ) Macrophages (n = 7 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( c ) PMNs (Polymorphonuclear neutrophils, n = 7 per group). ( d ) Lymphocytes (n = 7 per group). ( e ) The level of cytokine IL-1β in BALF was detected by ELISA assay (n = 7 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( f–g ) Fstl1 +/− and their WT littermate mice were intratracheally exposured to saline or silica, lung tissues were collected from Fstl1 +/− and WT mice 7 days after administration of saline or silica. ( f ) The level of cytokine IL-1β in lung tissues was detected by ELISA assay (n = 4 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( g ) The levels of NLRP3 and caspase-1 (p20) in lung tissues were determined by western blot analysis. β-tubulin was used as a loading control.

Article Snippet: The plate was incubated at 4 °C overnight, then 2.5 μg/mL biotinylated human FSTL1 detection antibody (R&D system, MAB1694) was added followed by washing with wash buffer and the plate was incubated for 1 hour at room temperature.

Techniques: Saline, Enzyme-linked Immunosorbent Assay, Western Blot, Control

Journal: Scientific Reports

Article Title: Follistatin like-1 aggravates silica-induced mouse lung injury

doi: 10.1038/s41598-017-00478-0

Figure Lengend Snippet: Fstl1 +/− mice have less myofibroblast accumulation after silica exposure. ( a ) Immunofluorescence analysis of α-SMA expression in lung sections of Fstl1 +/− and WT mice 21 days after saline or silica exposure. Representative images of the staining are shown. (α-SMA, green; Endomucin, red; nucleus, blue; scale bars, 200 μm). ( b ) qRT-PCR analysis of α-SMA mRNA expression in lung tissues from Fstl1 +/− and WT mice 21 days after saline or silica treatment (n = 3 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( c ) Western blot analysis of α-SMA expression in lung tissues from Fstl1 +/− and WT mice 21 days after saline or silica treatment. β-tubulin was used as a loading control.

Article Snippet: The plate was incubated at 4 °C overnight, then 2.5 μg/mL biotinylated human FSTL1 detection antibody (R&D system, MAB1694) was added followed by washing with wash buffer and the plate was incubated for 1 hour at room temperature.

Techniques: Immunofluorescence, Expressing, Saline, Staining, Quantitative RT-PCR, Western Blot, Control

Journal: Scientific Reports

Article Title: Follistatin like-1 aggravates silica-induced mouse lung injury

doi: 10.1038/s41598-017-00478-0

Figure Lengend Snippet: Fstl1 modulates myofibroblast differentiation via facilitating TGF-β1 signaling. ( a ) qRT-PCR analysis of TGF-β1 mRNA expression in lung tissues of Fstl1 +/− and WT mice at indicated time after saline or silica exposure (n = 4 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( b ) ELISA analysis of active form of TGF-β1 protein in lung tissues of Fstl1 +/− and WT mice at indicated time after saline or silica exposure (n = 4 per group; * P < 0.05 by one-way ANOVA followed by Student’s t test). ( c ) The levels of phosphorylation of Smad2/3 (p-Smad2/3), total Smad2/3 (T-Smad2/3), phosphorylation of Smad1/5 (p-Smad1/5) and total Smad1/5 (T-Smad1/5) in lung tissues of of Fstl1 +/− and WT mice 21 days after saline or silica exposure were determined by western blot analysis. β-tubulin was used as a loading control. ( d ) Primary lung fibroblasts from Fstl1 +/− and WT mice were treated with 5 ng/ml TGF-β1. Protein expressions of α-SMA in cell extracts and type I collagen (Col1) in medium 24 h after TGF-β1 treatment were determined by western blot analysis. β-tubulin was used as a loading control.

Article Snippet: The plate was incubated at 4 °C overnight, then 2.5 μg/mL biotinylated human FSTL1 detection antibody (R&D system, MAB1694) was added followed by washing with wash buffer and the plate was incubated for 1 hour at room temperature.

Techniques: Quantitative RT-PCR, Expressing, Saline, Enzyme-linked Immunosorbent Assay, Phospho-proteomics, Western Blot, Control

Journal: Scientific Reports

Article Title: Follistatin like-1 aggravates silica-induced mouse lung injury

doi: 10.1038/s41598-017-00478-0

Figure Lengend Snippet: FSTL1-neutralizing antibody attenuates silica-induced lung inflammation and subsequent pulmonary fibrosis in mice. ( a ) In a FSTL1 blockage experiment, C57BL/6 mice were intraperitoneally injected with 22B6 mAb or IgG1 (n = 6 per group) every other day from 1 day after silica challenge till the mice were sacrificed on day 7 for inflammation analysis or day 21 for fibrosis analysis. ( b–f ) For inflammation analysis, ( b ) the number of total BALF cells was determined by hemocytometer (* P < 0.05 by one-way ANOVA followed by Student’s t test). ( c–e ) The differential cell counts in BALF were determined according to standard morphologic criteria. ( c ) Macrophages (* P < 0.05 by one-way ANOVA followed by Student’s t test). ( d ) PMNs. ( e ) Lymphocytes. ( f ) The level of cytokine IL-1β was detected by ELISA assay (* P < 0.05 by one-way ANOVA followed by Student’s t test). ( g – i ) For fibrosis analysis, ( g ) hydroxyproline contents in lung tissues were measured (* P < 0.05, *** P < 0.001 by one-way ANOVA followed by Student’s t test). ( h ) Representative images of the H&E staining of lung sections are shown (Scale bars, 200 μm). ( i ) Lung fibrotic score analysis of the lung sections. The fibrotic area is presented as a percentage (* P < 0.05 by one-way ANOVA followed by Student’s t test). ( j ) Interventional dosing regimen of lung fibrosis model. C57BL/6J mice were intraperitoneally injected with 22B6 mAb or IgG1 (n = 10 per group) at indicated time after silica exposure, and lungs were harvested on day 21. ( k ) Representative images of the H&E staining of lung sections are shown (Scale bars, 200 μm). ( l ) Lung fibrotic score analysis of the lung sections. The fibrotic area is presented as a percentage (* P < 0.05 by one-way ANOVA followed by Student’s t test). ( m ) Hydroxyproline contents in lung tissues (*** P < 0.001 by one-way ANOVA followed by Student’s t test).

Article Snippet: The plate was incubated at 4 °C overnight, then 2.5 μg/mL biotinylated human FSTL1 detection antibody (R&D system, MAB1694) was added followed by washing with wash buffer and the plate was incubated for 1 hour at room temperature.

Techniques: Injection, Enzyme-linked Immunosorbent Assay, Staining

Journal: Scientific Reports

Article Title: Follistatin like-1 aggravates silica-induced mouse lung injury

doi: 10.1038/s41598-017-00478-0

Figure Lengend Snippet: Clinical characteristics of the study groups.

Article Snippet: The plate was incubated at 4 °C overnight, then 2.5 μg/mL biotinylated human FSTL1 detection antibody (R&D system, MAB1694) was added followed by washing with wash buffer and the plate was incubated for 1 hour at room temperature.

Techniques:

Journal: The Journal of Craniofacial Surgery

Article Title: FSTL1 Can Be a Promising Target in TMJ Osteoarthritis via Regulating Chondrocyte Mitophagy and Apoptosis

doi: 10.1097/SCS.0000000000010906

Figure Lengend Snippet: (A) Typical CBCT scan of TMD patients without OA. The red arrow showed the relatively normal bone margins. (B) Typical CBCT scan of TMD patients with OA. The red arrow showed the bone resorption in condyle. (C) The ELISA results of FSTL1 in TMJ auricular synovium fluid. (D) The ROC curve of the diagnosis value FSTL1 concentration in TMJ OA progress. CBCT indicates cone beam computed tomography; FSTL1, follistatin-like protein 1; OA, osteoarthritis; ROC, receiver operating characteristic; TMD, temporomandibular joint disorder; TMJ, temporomandibular joint.

Article Snippet: The used antibodies were as followed, FSTL1 (1:200, Proteintech), ACAN (1:200, Proteintech), COL2A1(1:200, Millipore), and MMP13 (1:200, Abclonal).

Techniques: Enzyme-linked Immunosorbent Assay, Biomarker Discovery, Concentration Assay, Computed Tomography

Journal: The Journal of Craniofacial Surgery

Article Title: FSTL1 Can Be a Promising Target in TMJ Osteoarthritis via Regulating Chondrocyte Mitophagy and Apoptosis

doi: 10.1097/SCS.0000000000010906

Figure Lengend Snippet: (A) The Rt-qPCR results of FSTL1 with the targeting plasmid transfection. (B) The western blot results of FSTL1 with the targeting plasmid transfection. (C) The Rt-qPCR results of the mRNA fold change of related typical markers to chondrocyte synthesis and catabolism as well as under the inflammatory stimulation. (D) The western blot results of changes of related typical markers to chondrocyte synthesis and catabolism as well as under the inflammatory stimulation. (E) The safranine O staining of chondrogenic cells with / without FSTL1 overexpression. FSTL1 indicates follistatin-like protein 1; Rt-qPCR, quantitative reverse transcription polymerase chain reaction.

Article Snippet: The used antibodies were as followed, FSTL1 (1:200, Proteintech), ACAN (1:200, Proteintech), COL2A1(1:200, Millipore), and MMP13 (1:200, Abclonal).

Techniques: Quantitative RT-PCR, Plasmid Preparation, Transfection, Western Blot, Staining, Over Expression, Reverse Transcription, Polymerase Chain Reaction

Journal: The Journal of Craniofacial Surgery

Article Title: FSTL1 Can Be a Promising Target in TMJ Osteoarthritis via Regulating Chondrocyte Mitophagy and Apoptosis

doi: 10.1097/SCS.0000000000010906

Figure Lengend Snippet: (A, B) The mitosox probe staining and the quantified analysis of the mitochondrial morphology and function with / without FSTL1 overexpression. (C, D) The western blot and the quantified analysis results of mitochondrial function with / without FSTL1 overexpression and inflammatory stimulation. (E, F) Flow cytometry analysis of the chondrocytes apoptosis rate with / without FSTL1 overexpression. (G, H) Flow cytometry analysis of the chondrocytes cell cycle with / without FSTL1 overexpression. FSTL1 indicates follistatin-like protein 1.

Article Snippet: The used antibodies were as followed, FSTL1 (1:200, Proteintech), ACAN (1:200, Proteintech), COL2A1(1:200, Millipore), and MMP13 (1:200, Abclonal).

Techniques: Staining, Over Expression, Western Blot, Flow Cytometry

Journal: The Journal of Craniofacial Surgery

Article Title: FSTL1 Can Be a Promising Target in TMJ Osteoarthritis via Regulating Chondrocyte Mitophagy and Apoptosis

doi: 10.1097/SCS.0000000000010906

Figure Lengend Snippet: (A) The Rt-qPCR results mRNA fold change with the targeted si-FSTL1. (B) The western blot results of FSTL1 with the targeting si-RNA and its impact of typical markers to chondrocyte synthesis and catabolism. (C, D) The mitosox probe and IF staining of mt-COⅡ and the quantified analysis of the mitochondrial morphology and function with / without downregulation of FSTL1 under inflammatory stimulation. (E, F) The western blot and the quantified analysis results of mitochondrial function with / without downregulation of FSTL1 and inflammatory stimulation. (G, H) Flow cytometry analysis of the chondrocytes apoptosis rate with / without FSTL1 downregulation. FSTL1 indicates follistatin-like protein 1; Rt-qPCR, quantitative reverse transcription polymerase chain reaction

Article Snippet: The used antibodies were as followed, FSTL1 (1:200, Proteintech), ACAN (1:200, Proteintech), COL2A1(1:200, Millipore), and MMP13 (1:200, Abclonal).

Techniques: Quantitative RT-PCR, Western Blot, Staining, Flow Cytometry, Reverse Transcription, Polymerase Chain Reaction

Journal: The Journal of Craniofacial Surgery

Article Title: FSTL1 Can Be a Promising Target in TMJ Osteoarthritis via Regulating Chondrocyte Mitophagy and Apoptosis

doi: 10.1097/SCS.0000000000010906

Figure Lengend Snippet: (A, B) HE and safranin O/Fast green staining of the cartilage in the sham group and UAC-induced OA rats with si-NC or si-FSTL1 injected group. Scale bar=100 μm. (C) IHC staining of the cartilage in the sham group and UAC-induced OA rats with si-NC or si-FSTL1 injected group. Scale bar=100 μm in the larger field of view and 50 μm in the smaller field of view. (D) The gross OARSI score of the cartilage condition in the sham group and UAC-induced OA rats with si-NC or si-FSTL1 injected group. (E) The statistical analysis of the cartilage IHC staining in the sham group and UAC-induced OA rats with si-NC or si-FSTL1 injected group. FSTL1 indicates follistatin-like protein 1; HE, hematoxylin-eosin; IHC, immunohistochemical; OA, osteoarthritis; UAC, unilateral anterior crossbite.

Article Snippet: The used antibodies were as followed, FSTL1 (1:200, Proteintech), ACAN (1:200, Proteintech), COL2A1(1:200, Millipore), and MMP13 (1:200, Abclonal).

Techniques: Staining, Injection, Immunohistochemistry, Immunohistochemical staining

Journal: Journal of Biological Chemistry

Article Title: Nucleolar Follistatin Promotes Cancer Cell Survival under Glucose-deprived Conditions through Inhibiting Cellular rRNA Synthesis

doi: 10.1074/jbc.m110.144477

Figure Lengend Snippet: FIGURE 1. Glucose deprivation enhances FST expression and nucleolar localization. HeLa cells were incubated in glucose-free medium and harvested at the indicated time. A, the FST mRNA level was measured with real time qPCR. B, FST protein level was detected by immunoblotting. The RT-qPCR data are shown as arbitrary values after normalizing to -actin mRNA level, and the data are presented as the means S.D. of three independent experiments. **, p 0.01 versus normal glucose controls. C, HeLa cells were incubated with (Glu) or without (Glu) glucose for 24 h and then stained with FST monoclonal antibod- ies.Nucleolinwasusedasthenucleolarmarker.Thearrowsindicatethenucleolar FST (left panel). The relative fluorescence intensities in the entire cell (columns T), cytoplasm(columnsC),andnucleus(columnsN)weremeasured.Thedatashown arethemeansS.D.of50cells(rightpanel).**,p0.01.D,FSTproteinexpres- sions in total (lanes T), cytoplasmic (lanes C), and nuclear (lanes N) fractions were analyzed by immunoblotting. Lamin B and -tubulin were used as nuclear and cytoplasm protein marker, respectively.

Article Snippet: The membrane was blocked with 3% bone serum album in TBS-T buffer (20 mM Tris-HCl, pH 8.0, 150 mM NaCl, 0.05% Tween 20), probed with antibodies targeting to FST (Santa Cruz Biotechnology, Santa Cruz, CA), -tubulin (Cell Signaling Technology, Beverly, MA), Lamin B (Santa Cruz Biotechnology), or PARP (Cell Signaling Technology), incubated with horseradish-conjugated secondary antibodies, detected with the SuperSignal West Pico chemiluminescence substrate (Thermo Fisher Scientific), and finally exposed to an x-ray film.

Techniques: Expressing, Incubation, Western Blot, Quantitative RT-PCR, Staining, Fluorescence, Marker

Journal: Journal of Biological Chemistry

Article Title: Nucleolar Follistatin Promotes Cancer Cell Survival under Glucose-deprived Conditions through Inhibiting Cellular rRNA Synthesis

doi: 10.1074/jbc.m110.144477

Figure Lengend Snippet: FIGURE 4. FST influences histone modifications at rDNA region. HeLa cells transfected with siRNA targeting FST (FSTi-1) or control siRNA (NC) were cultured with (Glu) or without (Glu) glucose for 24 h. ChIP analysis was performed with antibodies against H3K4me2 (A) or acetyl-H3 (B) and analyzed by qPCR. Cell lines stably expressing FST, FST/NLS, or control vector were cultured with (Glu) or without (Glu) glucose for 24 h. ChIP analysis was then performed with antibodies against H3K4me2 (C) or acetyl-H3 (D) and analyzed by qPCR. The occupancies of H3K4me2 and acetyl-H3 at rDNA regions were normalized to GAPDH. The values are the means S.D. of three independent experiments. *, p 0.05; **, p 0.01.

Article Snippet: The membrane was blocked with 3% bone serum album in TBS-T buffer (20 mM Tris-HCl, pH 8.0, 150 mM NaCl, 0.05% Tween 20), probed with antibodies targeting to FST (Santa Cruz Biotechnology, Santa Cruz, CA), -tubulin (Cell Signaling Technology, Beverly, MA), Lamin B (Santa Cruz Biotechnology), or PARP (Cell Signaling Technology), incubated with horseradish-conjugated secondary antibodies, detected with the SuperSignal West Pico chemiluminescence substrate (Thermo Fisher Scientific), and finally exposed to an x-ray film.

Techniques: Transfection, Control, Cell Culture, Stable Transfection, Expressing, Plasmid Preparation

Journal: Scientific Reports

Article Title: Follistatin-like 1 protects against hypoxia-induced pulmonary hypertension in mice

doi: 10.1038/srep45820

Figure Lengend Snippet: ( a ) Serum concentration of FSTL1 protein by ELISA in patients with COPD only (n = 8), COPD combined with PH (n = 8) and healthy controls (CTL, n = 7). ( b ) Effect of chronic hypoxia on RVSP and RVHI ( c ) in C57BL/6 mice. n = 8. ( d ) QRT-PCR analysis of Fstl1 mRNA in lung tissue of C57BL/6 mice under hypoxia as normalized by Gapdh mRNA. n = 10. ( e ) Representative cropped western blots and statistical analysis of FSTL1 protein in lung tissue of C57BL/6 mice under hypoxia as normalized by GAPDH. n = 10. ( f ) Serum concentration of FSTL1 protein by ELISA in C57BL/6 mice under hypoxia. n = 7–11. ( g ) Representative immunofluorescence images showing FSTL1 (green) and α-SMA (red) staining of pulmonary arterioles from lung sections in hypoxia-treated mice and untreated ones. Nuclei were stained with DAPI (blue). n = 4–5. Bar = 50 μm. Data are presented as mean ± SEM. * P < 0.05, ** P < 0.01. COPD = chronic obstructive pulmonary diseases. PH = pulmonary hypertension. RVSP = right ventricular systolic pressure. RVHI = right ventricular hypertrophy index. W = week. ELISA = enzyme-linked immunosorbent assay. GAPDH = glyceraldehyde-3-phosphate dehydrogenase. α-SMA = α-smooth muscle actin. DAPI = 4′,6-diamidino-2-phenylindole.

Article Snippet: For systemic delivery of FSTL1 protein, mice were randomly subjected to tail-vein injection with recombinant human FSTL1 protein (100 ng/g mouse , R & D Systems, USA) or with equal dosage of vehicle (PBS), immediately before the beginning of hypoxia or after 2 weeks of hypoxia, and repeated every 3 days until the end.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Western Blot, Immunofluorescence, Staining

Journal: Scientific Reports

Article Title: Follistatin-like 1 protects against hypoxia-induced pulmonary hypertension in mice

doi: 10.1038/srep45820

Figure Lengend Snippet: ( a ) Representative cropped western blots of FSTL1 protein in Fstl1 +/− and WT mice as normalized by GAPDH. n = 6. ( b ) RVSP in Fstl1 +/− and WT mice under hypoxia. n = 7–12. ( c ) RVHI in Fstl1 +/− and WT mice under hypoxia. n = 8–10. ( d ) Representative images showing hematoxylin and eosin staining of pulmonary arterioles from lung sections in Fstl1 +/− and WT mice under hypoxia. n = 4–5. Bar = 20 μm. ( e ) Representative immunofluorescence images showing α-SMA staining (red) of pulmonary arterioles from lung sections in Fstl1 +/− and WT mice under hypoxia. n = 4–5. Bar = 50 μm. ( f ) MT% of pulmonary arteries grouped by 0–50 μm and 50–100 μm in outer diameter from lung sections in Fstl1 +/− and WT mice under hypoxia. n = 5. ( g ) Numbers of completely muscularized arterioles (0–50 μm in outer diameter) per 10 fields from lung sections in Fstl1 +/− and WT mice under hypoxia. n = 5. Data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001. HPH = hypoxia-induced PH. WT = wide type. RVSP = right ventricular systolic pressure. RVHI = right ventricular hypertrophy index. MT = media thickness. W = week. GAPDH = glyceraldehyde-3-phosphate dehydrogenase. α-SMA = α-smooth muscle actin.

Article Snippet: For systemic delivery of FSTL1 protein, mice were randomly subjected to tail-vein injection with recombinant human FSTL1 protein (100 ng/g mouse , R & D Systems, USA) or with equal dosage of vehicle (PBS), immediately before the beginning of hypoxia or after 2 weeks of hypoxia, and repeated every 3 days until the end.

Techniques: Western Blot, Staining, Immunofluorescence

Journal: Scientific Reports

Article Title: Follistatin-like 1 protects against hypoxia-induced pulmonary hypertension in mice

doi: 10.1038/srep45820

Figure Lengend Snippet: ( a ) FSTL1 treatment regimen in HPH model of mice. ( b ) Representative cropped western blots of serum FSTL1 protein in mice intravenously administrated with FSTL1 or PBS under hypoxia. n = 4. RVSP ( c ) and RVHI ( d ) in mice intravenously administrated with FSTL1 or PBS under hypoxia. n = 5. ( e ) Representative images showing hematoxylin and eosin staining of pulmonary arterioles from lung sections in mice intravenously administrated with FSTL1 or PBS under hypoxia. n = 4–5. Bar = 20 μm. (f) Representative immunofluorescence images showing α-SMA staining (red) of pulmonary arterioles from lung sections in mice intravenously administrated with FSTL1 or PBS under hypoxia. n = 4–5. Bar = 50 μm. (g) MT% of pulmonary arteries grouped by 0–50 μm and 50–100 μm in outer diameter from lung sections in mice intravenously administrated with FSTL1 or PBS under hypoxia. n = 5. ( h ) Numbers of completely muscularized arterioles (0–50 μm in outer diameter) per 10 fields from lung sections in mice intravenously administrated with FSTL1 or PBS under hypoxia. n = 5. Data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001. HPH = hypoxia-induced PH. PBS = phosphate buffer saline. RVSP = right ventricular systolic pressure. RVHI = right ventricular hypertrophy index. α-SMA = α-smooth muscle actin. MT = media thickness. W = week.

Article Snippet: For systemic delivery of FSTL1 protein, mice were randomly subjected to tail-vein injection with recombinant human FSTL1 protein (100 ng/g mouse , R & D Systems, USA) or with equal dosage of vehicle (PBS), immediately before the beginning of hypoxia or after 2 weeks of hypoxia, and repeated every 3 days until the end.

Techniques: Western Blot, Staining, Immunofluorescence, Saline

Journal: Scientific Reports

Article Title: Follistatin-like 1 protects against hypoxia-induced pulmonary hypertension in mice

doi: 10.1038/srep45820

Figure Lengend Snippet: ( a ) Effect of FSTL1 on cellular viability under hypoxia and PDGF-BB ( b ) in MTT assay. n = 3. ( c ) Effect of FSTL1 on DNA synthesis under hypoxia and PDGF-BB ( d ) in BrdU assay for flow cytometer analysis. Cells in synthesis phrase (S, P4) at a cell cycle was calculated as the percent of P4/(P3 + P4 + P5). n = 3. ( e ) Effect of FSTL1 on cellular migration in transwell chamber. Nuclei of trans-membrane cells were stained with DAPI (blue). n = 3. Bar = 50 μm. Data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001. HPASMCs = human pulmonary artery smooth muscle cells. P3 = G0/G1 phrase. P4 = S phase. P5 = G2 phrase. P6 = apoptosis phrase. MTT = 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide. BrdU = 5-bromo-2-deoxyuridine. DAPI = 4’,6-diamidino-2-phenylindole. N = normoxia. H = hypoxia. V = vehicle. P = PDGF-BB. PDGF = platelet derived growth factor. FSTL1 = ng/mL.

Article Snippet: For systemic delivery of FSTL1 protein, mice were randomly subjected to tail-vein injection with recombinant human FSTL1 protein (100 ng/g mouse , R & D Systems, USA) or with equal dosage of vehicle (PBS), immediately before the beginning of hypoxia or after 2 weeks of hypoxia, and repeated every 3 days until the end.

Techniques: MTT Assay, DNA Synthesis, BrdU Staining, Flow Cytometry, Migration, Membrane, Staining, Derivative Assay

Journal: Scientific Reports

Article Title: Follistatin-like 1 protects against hypoxia-induced pulmonary hypertension in mice

doi: 10.1038/srep45820

Figure Lengend Snippet: ( a ) QRT-PCR analysis of FSTL1 mRNA in HPASMCs transfected with siRNA or N.C., as normalized by GAPDH mRNA. n = 4. ( b ) Representative cropped western blots of FSTL1 protein in HPASMCs transfected with siRNA or N.C. n = 3. ( c ) Effect of FSTL1 siRNA transfection on cellular viability in MTT assay. n = 3. ( d ) Effect of FSTL1 siRNA transfection on DNA synthesis in BrdU assay for flow cytometer analysis. Cells in synthesis phrase (S, P4) at a cell cycle was calculated as the percent of P4/(P3 + P4 + P5). n = 3. ( e ) Effect of FSTL1 siRNA transfection on cellular migration in transwell chamber. Nuclei of trans-membrane cells were stained with DAPI (blue). n = 3. Bar = 50 μm. Data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001. SiRNA = small interfering RNA. N.C. = negative control. P3 = G0/G1 phrase. P4 = S phase. P5 = G2 phrase. P6 = apoptosis phrase. N = normoxia. H = hypoxia. GAPDH = glyceraldehyde-3-phosphate dehydrogenase. DAPI = 4′, 6-diamidino-2-phenylindole. MTT = 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide. BrdU = 5-bromo-2-deoxyuridine.

Article Snippet: For systemic delivery of FSTL1 protein, mice were randomly subjected to tail-vein injection with recombinant human FSTL1 protein (100 ng/g mouse , R & D Systems, USA) or with equal dosage of vehicle (PBS), immediately before the beginning of hypoxia or after 2 weeks of hypoxia, and repeated every 3 days until the end.

Techniques: Quantitative RT-PCR, Transfection, Western Blot, MTT Assay, DNA Synthesis, BrdU Staining, Flow Cytometry, Migration, Membrane, Staining, Small Interfering RNA, Negative Control

Journal: Scientific Reports

Article Title: Follistatin-like 1 protects against hypoxia-induced pulmonary hypertension in mice

doi: 10.1038/srep45820

Figure Lengend Snippet: ( a ) Representative cropped western blots and statistical analysis of phosphorylations of AMPK (p-AMPK, n = 5) and ERK (p-ERK, n = 6) in lung tissue of Fstl1 +/− mice and WT controls under hypoxia. ( b ) Representative cropped western blots and statistical analysis of p-AMPK (n = 5) and p-ERK (n = 5) in lung tissue of mice treated with FSTL1 or PBS under hypoxia. Representative cropped western blots and statistical analysis of p-ERK ( c ) and p-AMPK ( d ) in HPASMCs exposed to hypoxia or normoxia for 24 h. n = 3. ( e ) Representative cropped western blots and statistical analysis of p-ERK in HPASMCs transfected with FSTL1 siRNA exposed to hypoxia or normoxia for 24 h. n = 3. Data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001. HPH = hypoxia-induced PH. ERK = extracellular regulated kinase. AMPK = AMP-activated protein kinase. PBS = phosphate buffer saline. FSTL1 = 250 ng/mL. SiRNA = small interfering RNA. W = week.

Article Snippet: For systemic delivery of FSTL1 protein, mice were randomly subjected to tail-vein injection with recombinant human FSTL1 protein (100 ng/g mouse , R & D Systems, USA) or with equal dosage of vehicle (PBS), immediately before the beginning of hypoxia or after 2 weeks of hypoxia, and repeated every 3 days until the end.

Techniques: Western Blot, Transfection, Saline, Small Interfering RNA

Journal: Scientific Reports

Article Title: Follistatin-like 1 protects against hypoxia-induced pulmonary hypertension in mice

doi: 10.1038/srep45820

Figure Lengend Snippet: ( a ) Effect of ERK inactivation on FSTL1 knockdown-induced cellular viability under hypoxia in MTT assay. n = 3. ( b ) Effect of ERK inactivation on FSTL1 knockdown-induced DNA synthesis under hypoxia in BrdU assay for flow cytometer analysis. Cells in synthesis phrase (S, P4) at a cell cycle was calculated as the percent of P4/(P3 + P4 + P5). n = 3. ( c ) Effect of ERK inactivation on FSTL1 knockdown-induced cellular migration under hypoxia in transwell chamber. Nuclei of trans-membrane cells were stained with DAPI (blue). n = 3. Bar = 50 μm. Data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001. ERK = extracellular regulated kinase. P3 = G0/G1 phrase. P4 = S phase. P5 = G2 phrase. P6 = apoptosis phrase. DAPI = 4′,6-diamidino-2-phenylindole. BrdU = 5-bromo-2-deoxyuridine. SiRNA = small interfering RNA. MTT = 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide.

Article Snippet: For systemic delivery of FSTL1 protein, mice were randomly subjected to tail-vein injection with recombinant human FSTL1 protein (100 ng/g mouse , R & D Systems, USA) or with equal dosage of vehicle (PBS), immediately before the beginning of hypoxia or after 2 weeks of hypoxia, and repeated every 3 days until the end.

Techniques: Knockdown, MTT Assay, DNA Synthesis, BrdU Staining, Flow Cytometry, Migration, Membrane, Staining, Small Interfering RNA

Journal: European Journal of Nuclear Medicine and Molecular Imaging

Article Title: [ 68 Ga]Ga-NODAGA-RGD post MI reflects activated fibroblasts rather than angiogenesis

doi: 10.1007/s00259-025-07489-4

Figure Lengend Snippet: Violin-plot visualization of gene markers for injury-activated fibroblasts at day 5 post MI (Fstl1, Col1a1, Col3a1, Col5a1) comparing relative gene expression between the experimental groups and cell types. Median values are visualized by a black horizontal bar. ( n = 5 per group)

Article Snippet: Primary antibodies targeting FSTL1 (R&D Systems, AF1738, dilution 1:100), Integrin (Abcam, ab179473, dilution 1:100) CD31 (R&D Systems, AF3628, dilution 1:100) were applied overnight at 4°.

Techniques: Gene Expression

Journal: European Journal of Nuclear Medicine and Molecular Imaging

Article Title: [ 68 Ga]Ga-NODAGA-RGD post MI reflects activated fibroblasts rather than angiogenesis

doi: 10.1007/s00259-025-07489-4

Figure Lengend Snippet: Expression pattern of α v β 3 -integrin, Fstl1 and CD31 in hearts at day 5 after permanent ligation of the LAD in both remote and infarcted myocardium, and the SHAM group. α v β 3 -integrin and Fstl1 are highly expressed within the infarct area ( A ), while profound less expression was detected in the remote area, comparable to the SHAM group ( B , C ). CD31 is expressed within the scar ( D ), but shows basal expression levels in the remote area ( E ) – similar to the SHAM group ( F ). Co-localization of Fstl1 and α v β 3 -integrin was quantified in co-labelled heart sections ( G ). Regions of Interest (ROI) were analyzed according to a positive fluorescence signal for both proteins ( H ). All α v β 3 -integrin positive ROIs were contained in the Fstl1-positive ROI population ( I ). Scale bar: 100 μm

Article Snippet: Primary antibodies targeting FSTL1 (R&D Systems, AF1738, dilution 1:100), Integrin (Abcam, ab179473, dilution 1:100) CD31 (R&D Systems, AF3628, dilution 1:100) were applied overnight at 4°.

Techniques: Expressing, Ligation, Fluorescence