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(A-C) Bar plots showing differential regulation of pericyte markers (RGS5, ABCC9), VSMC markers <t>(CNN1,</t> MYH11), and pan-mural cell markers (ACTA2, SMTN, TAGLN) in fibroblasts treated with (A) NGF, (B) BDNF, or (C) NT3 compared with untreated fibroblasts. Bar heights represent log₂ fold change, and bar color indicates the corresponding P values. (D) Schematic of the collagen gel contraction assay in which synovial fibroblasts were embedded in collagen I gels and treatment (E) Representative images of collagen I gel contraction following treatment with NGF, BDNF, or NT3 (F) Quantification of collagen I gel contraction expressed as percent contraction. Values represent mean ± standard deviation (SD). Individual data points indicate biological replicates. Statistical analysis was performed using one-way ANOVA, with P values shown. (G) Schematic illustrating the synovial organoid/explant experimental design. (H) Representative H&E image of synovial explant culture after 3 days of culture. (I) Whole-mount immunoluorescence images showing preservation of endothelial cells after 3 days in culture, marked by CD31 (magenta), and vascular smooth muscle cells marked by MYH11, surrounding CD31 vascular structures. (J) Whole-mount staining showing expression of neurotrophin receptors NGFR, NTRK1, NTRK2, and NTRK3 (green) around vascularature, together with CD31 (magenta) and DAPI (blue). (K) Synovial organoids embedded in Matrigel and treated with NGF, BDNF, or NT3, followed by fixation and staining for a-smooth muscle actin (aSMA) (brown) to visualize mural cells. All 20X images were cropped at similar scale. Scale bar, 150 µm.

Cnn1, supplied by Proteintech, used in various techniques. Bioz Stars score: 94/100, based on 97 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "Fibroblasts neurotrophin signaling sustains pathological vascular maturation in rheumatoid arthritis"

Article Title: Fibroblasts neurotrophin signaling sustains pathological vascular maturation in rheumatoid arthritis

Journal: bioRxiv

doi: 10.64898/2026.03.12.711120

(A-C) Bar plots showing differential regulation of pericyte markers (RGS5, ABCC9), VSMC markers (CNN1, MYH11), and pan-mural cell markers (ACTA2, SMTN, TAGLN) in fibroblasts treated with (A) NGF, (B) BDNF, or (C) NT3 compared with untreated fibroblasts. Bar heights represent log₂ fold change, and bar color indicates the corresponding P values. (D) Schematic of the collagen gel contraction assay in which synovial fibroblasts were embedded in collagen I gels and treatment (E) Representative images of collagen I gel contraction following treatment with NGF, BDNF, or NT3 (F) Quantification of collagen I gel contraction expressed as percent contraction. Values represent mean ± standard deviation (SD). Individual data points indicate biological replicates. Statistical analysis was performed using one-way ANOVA, with P values shown. (G) Schematic illustrating the synovial organoid/explant experimental design. (H) Representative H&E image of synovial explant culture after 3 days of culture. (I) Whole-mount immunoluorescence images showing preservation of endothelial cells after 3 days in culture, marked by CD31 (magenta), and vascular smooth muscle cells marked by MYH11, surrounding CD31 vascular structures. (J) Whole-mount staining showing expression of neurotrophin receptors NGFR, NTRK1, NTRK2, and NTRK3 (green) around vascularature, together with CD31 (magenta) and DAPI (blue). (K) Synovial organoids embedded in Matrigel and treated with NGF, BDNF, or NT3, followed by fixation and staining for a-smooth muscle actin (aSMA) (brown) to visualize mural cells. All 20X images were cropped at similar scale. Scale bar, 150 µm.

Figure Legend Snippet: (A-C) Bar plots showing differential regulation of pericyte markers (RGS5, ABCC9), VSMC markers (CNN1, MYH11), and pan-mural cell markers (ACTA2, SMTN, TAGLN) in fibroblasts treated with (A) NGF, (B) BDNF, or (C) NT3 compared with untreated fibroblasts. Bar heights represent log₂ fold change, and bar color indicates the corresponding P values. (D) Schematic of the collagen gel contraction assay in which synovial fibroblasts were embedded in collagen I gels and treatment (E) Representative images of collagen I gel contraction following treatment with NGF, BDNF, or NT3 (F) Quantification of collagen I gel contraction expressed as percent contraction. Values represent mean ± standard deviation (SD). Individual data points indicate biological replicates. Statistical analysis was performed using one-way ANOVA, with P values shown. (G) Schematic illustrating the synovial organoid/explant experimental design. (H) Representative H&E image of synovial explant culture after 3 days of culture. (I) Whole-mount immunoluorescence images showing preservation of endothelial cells after 3 days in culture, marked by CD31 (magenta), and vascular smooth muscle cells marked by MYH11, surrounding CD31 vascular structures. (J) Whole-mount staining showing expression of neurotrophin receptors NGFR, NTRK1, NTRK2, and NTRK3 (green) around vascularature, together with CD31 (magenta) and DAPI (blue). (K) Synovial organoids embedded in Matrigel and treated with NGF, BDNF, or NT3, followed by fixation and staining for a-smooth muscle actin (aSMA) (brown) to visualize mural cells. All 20X images were cropped at similar scale. Scale bar, 150 µm.

Techniques Used: Collagen Gel Contraction Assay, Standard Deviation, Preserving, Staining, Expressing

(A) RNAscope images showing transcript expression of pericyte marker RGS5 (yellow) and VSMC marker MYH11 (red) in fibroblasts treated with different neurotrophins, overlaid with DAPI (blue). Scale bar, 150 µm. All images were acquired at 20x magnification and cropped to the same scale.(B) Quantification of total numbers of RGS5+, MYH11+, and RGS5+/MYH11+ double-positive cells following treatment with the indicated neurotrophins.(C) mRNA expression of the pericyte marker RGS5 in fibroblasts transfected with siRNAs targeting NGFR or NTRK1, in the presence or absence of NGF stimulation.(D) mRNA expression of the VSMC marker MYH11 in fibroblasts transfected with siRNAs targeting NTRK2 or NTRK3, in the presence or absence of BDNF or NT3 stimulation, respectively.(E-F) mRNA expression of the mural cell marker CNN1 (calponin) in fibroblasts stimulated with NGF (E) in the presence or absence of a TRKA inhibitor, or stimulated with BDNF (F) in the presence or absence of an NTRK2 inhibitor.

Figure Legend Snippet: (A) RNAscope images showing transcript expression of pericyte marker RGS5 (yellow) and VSMC marker MYH11 (red) in fibroblasts treated with different neurotrophins, overlaid with DAPI (blue). Scale bar, 150 µm. All images were acquired at 20x magnification and cropped to the same scale.(B) Quantification of total numbers of RGS5+, MYH11+, and RGS5+/MYH11+ double-positive cells following treatment with the indicated neurotrophins.(C) mRNA expression of the pericyte marker RGS5 in fibroblasts transfected with siRNAs targeting NGFR or NTRK1, in the presence or absence of NGF stimulation.(D) mRNA expression of the VSMC marker MYH11 in fibroblasts transfected with siRNAs targeting NTRK2 or NTRK3, in the presence or absence of BDNF or NT3 stimulation, respectively.(E-F) mRNA expression of the mural cell marker CNN1 (calponin) in fibroblasts stimulated with NGF (E) in the presence or absence of a TRKA inhibitor, or stimulated with BDNF (F) in the presence or absence of an NTRK2 inhibitor.

Techniques Used: RNAscope, Expressing, Marker, Transfection

Blocking Assay:

Article Title: Fibroblasts neurotrophin signaling sustains pathological vascular maturation in rheumatoid arthritis
Article Snippet: Using the Bio-Rad TRANS-BLOT TURBO transfer system (Bio-Rad, #690BR324) with a dry transfer protocol for 30 minutes according to the manufacturer’s instructions (Bio-Rad). .. Following transfer, membranes were blocked for 20 minutes in Everyblot blocking buffer (Bio-Rad # 12010020) and incubated overnight at 4°C with primary antibodies from TrkA and TrkB antibody sampler kit against TrkA, TrkB, p-TrkA/TrkB(Cell Signaling Technology, #4638, 1:500), CNN1 (Proteintech-24855-1-AP), MYH11 (Proteintech-21404-1-AP), NGF (Abcam-ab52918) and GAPDH (Thermo Fisher Scientific, #MA5-15738),or beta-actin (Cell Signaling Technology, #3700). .. Protein concentration was determined using the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific, #23227), and 30-50 μg of protein was subjected to SDS-PAGE on mini-protein TGX precast Gels (Bio-Rad#45610954) and transferred to PVDF membranes (Bio-Rad#1620174).

Incubation:

other:

Article Title: Tirzepatide mitigates thoracic aortic aneurysm and dissection by alleviating the loss of the contractile phenotype in vascular smooth muscle cells and reducing vascular inflammation.
Article Snippet: Background: Thoracic aortic aneurysm and dissection (TAAD) is a kind of life-threatening cardiovascular condition with a poor prognosis, currently lacking effective drug therapies that can halt the progression of TAAD.. Tirzepatide, a dual GIP and GLP-1 receptor agonist used for type 2 diabetes and weight management, has shown cardioprotective potential, yet its effect on TAAD remains unknown.. Methods: A TAAD model in mice was developed by providing 0.5% β-aminopropionitrile (BAPN) for 28 days.Mice were categorized into control, tirzepatide-only, BAPN, and BAPN combined with tirzepatide.Tirzepatide (10 nmol/kg) was administered daily via intraperitoneal injection.

Image Search Results

Aerobic exercise preserves the contractile phenotype of vascular smooth muscle cells (VSMCs) in the mouse aorta. A , Quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis of MYH11 , ACTA2 , MYL9 , CNN1 , TAGLN , and SPP1 mRNA levels in distal nonlesion tissue (AD-distal non-lesion; n=5) and lesion tissue (AD-lesion; n=5) from patients with AD. B and C , Western blot analysis ( B ) and quantification ( C ) of MYH11 (myosin heavy chain 11), CNN1 (calponin 1), α-SMA (α‐smooth muscle actin), and OPN (osteopontin) protein expression in human aortic specimens (n=8). D through G , Three-week-old mice underwent 1 week of acclimation training followed by administration of β-aminopropionitrile (BAPN; 1 g/kg per day) or normal drinking water, combined with 12 weeks of either forced treadmill exercise training (Run) or sedentary (Sed) conditions. D , Schematic of the experimental timeline. E , qRT-PCR analysis of Myh11 , Myl9 , Acta2 , Cnn1 , Tagln , and Spp1 mRNA levels in mouse aortic tissues (n=6). F and G , Western blot analysis ( F ) and quantification ( G ) of MYH11, α-SMA, CNN1, and OPN protein levels in mouse aortic tissues (n=6). Data are presented as mean±SEM. An unpaired Student t test was performed in A and C , 2-way ANOVA followed by a Tukey multiple-comparison test was used in E and G . Panel D created in BioRender. Mao, J. (2026) https://BioRender.com/qlycz8m .

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Exercise Attenuates Aortic Dissection Via PDE5A-Mediated Inhibition of Vascular Smooth Muscle Cell Phenotypic Switch

doi: 10.1161/ATVBAHA.125.324293

Figure Lengend Snippet: Aerobic exercise preserves the contractile phenotype of vascular smooth muscle cells (VSMCs) in the mouse aorta. A , Quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis of MYH11 , ACTA2 , MYL9 , CNN1 , TAGLN , and SPP1 mRNA levels in distal nonlesion tissue (AD-distal non-lesion; n=5) and lesion tissue (AD-lesion; n=5) from patients with AD. B and C , Western blot analysis ( B ) and quantification ( C ) of MYH11 (myosin heavy chain 11), CNN1 (calponin 1), α-SMA (α‐smooth muscle actin), and OPN (osteopontin) protein expression in human aortic specimens (n=8). D through G , Three-week-old mice underwent 1 week of acclimation training followed by administration of β-aminopropionitrile (BAPN; 1 g/kg per day) or normal drinking water, combined with 12 weeks of either forced treadmill exercise training (Run) or sedentary (Sed) conditions. D , Schematic of the experimental timeline. E , qRT-PCR analysis of Myh11 , Myl9 , Acta2 , Cnn1 , Tagln , and Spp1 mRNA levels in mouse aortic tissues (n=6). F and G , Western blot analysis ( F ) and quantification ( G ) of MYH11, α-SMA, CNN1, and OPN protein levels in mouse aortic tissues (n=6). Data are presented as mean±SEM. An unpaired Student t test was performed in A and C , 2-way ANOVA followed by a Tukey multiple-comparison test was used in E and G . Panel D created in BioRender. Mao, J. (2026) https://BioRender.com/qlycz8m .

Article Snippet: Subsequently, the membrane was blocked with 5% skimmed milk at room temperature for 2 hours and incubated at 4 °C overnight with the following primary antibodies: MYH11 (myosin heavy chain 11; catalog no: A10827; Polyclonal, ABclonal Technology, Wuhan, China), CNN1 (calponin 1; catalog no: A16638; Polyclonal, ABclonal Technology, Wuhan, China), α-SMA (alpha-smooth muscle actin; catalog no: A2547; clone no: 1A4, Sigma-Aldrich, St Louis, United States), OPN (osteopontin; catalog no: 83341-1-RR, clone no: 240206A2, Proteintech, Wuhan, China), PDE5A (catalog no: A6831, Polyclonal, ABclonal Technology, Wuhan, China), RUNX1 (catalog no: 25315-1-AP, Polyclonal, Proteintech, Wuhan, China), α-tubulin (catalog no: AC012, Clone No: AMC0479, ABclonal Technology, Wuhan, China), and GAPDH (glyceraldehyde-3-phosphate dehydrogenase; catalog no: AC002, Clone No: AMC0062R, ABclonal Technology, Wuhan, China).

Techniques: Reverse Transcription, Polymerase Chain Reaction, Quantitative RT-PCR, Western Blot, Expressing, Comparison

Pharmacological inhibition of PDE5A (phosphodiesterase 5A) abrogates the protective effect of exercise in β-aminopropionitrile (BAPN)–induced mice. Mice receiving BAPN water were subjected to 12 weeks of either treadmill exercise (Run) or sedentary (Sed) conditions, while concurrently receiving intraperitoneal injection of sildenafil (Sild) or vehicle (Veh) treatment for 12 weeks (n=23–28). A , Schematic illustration of the experimental timeline. B , Survival curves for BAPN-induced mice undergoing Run or Sed with or without sildenafil treatment. Survival data were analyzed by the Kaplan-Meier method and compared using log-rank tests (n=23–28). C , Representative images of aortas from BAPN-induced mice undergoing Run or Sed combined with sildenafil treatment or not. D , Maximal thoracic aorta diameter in mice receiving BAPN water (n=8). E , The incidence of aortic dissection formation in mice from the indicated experimental groups (n=23–28). F , The incidence of aortic rupture in mice from indicated experimental groups (n=23–28). G and H , Representative hematoxylin and eosin (H&E), elastic Verhoeff-Van Gieson (EVG), and Masson trichrome staining of the thoracic aortas in mice receiving BAPN-supplemented water ( G ), and quantification of elastin degradation based on a 4‐point scoring system ( H ; n=6). Scale bar=100 µm. I and J , Western blot analysis ( I ) and quantification ( J ) of MYH11 (myosin heavy chain 11), CNN1 (calponin 1), α-SMA (α‐smooth muscle actin), and OPN (osteopontin) protein levels in BAPN-induced mice undergoing Run or Sed combined with sildenafil treatment or not (n=6). Data are presented as mean±SEM. Two-way ANOVA followed by Tukey multiple-comparison test was used in D , H , and J , Fisher exact test was performed in E and F . Created in BioRender. Mao, J. (2026) https://BioRender.com/qlycz8m .

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Exercise Attenuates Aortic Dissection Via PDE5A-Mediated Inhibition of Vascular Smooth Muscle Cell Phenotypic Switch

doi: 10.1161/ATVBAHA.125.324293

Figure Lengend Snippet: Pharmacological inhibition of PDE5A (phosphodiesterase 5A) abrogates the protective effect of exercise in β-aminopropionitrile (BAPN)–induced mice. Mice receiving BAPN water were subjected to 12 weeks of either treadmill exercise (Run) or sedentary (Sed) conditions, while concurrently receiving intraperitoneal injection of sildenafil (Sild) or vehicle (Veh) treatment for 12 weeks (n=23–28). A , Schematic illustration of the experimental timeline. B , Survival curves for BAPN-induced mice undergoing Run or Sed with or without sildenafil treatment. Survival data were analyzed by the Kaplan-Meier method and compared using log-rank tests (n=23–28). C , Representative images of aortas from BAPN-induced mice undergoing Run or Sed combined with sildenafil treatment or not. D , Maximal thoracic aorta diameter in mice receiving BAPN water (n=8). E , The incidence of aortic dissection formation in mice from the indicated experimental groups (n=23–28). F , The incidence of aortic rupture in mice from indicated experimental groups (n=23–28). G and H , Representative hematoxylin and eosin (H&E), elastic Verhoeff-Van Gieson (EVG), and Masson trichrome staining of the thoracic aortas in mice receiving BAPN-supplemented water ( G ), and quantification of elastin degradation based on a 4‐point scoring system ( H ; n=6). Scale bar=100 µm. I and J , Western blot analysis ( I ) and quantification ( J ) of MYH11 (myosin heavy chain 11), CNN1 (calponin 1), α-SMA (α‐smooth muscle actin), and OPN (osteopontin) protein levels in BAPN-induced mice undergoing Run or Sed combined with sildenafil treatment or not (n=6). Data are presented as mean±SEM. Two-way ANOVA followed by Tukey multiple-comparison test was used in D , H , and J , Fisher exact test was performed in E and F . Created in BioRender. Mao, J. (2026) https://BioRender.com/qlycz8m .

Techniques: Inhibition, Injection, Dissection, Staining, Western Blot, Comparison

Overexpression of PDE5A (phosphodiesterase 5A) in vascular smooth muscle cells (VSMCs) rescues β-aminopropionitrile (BAPN)–induced aortic dissection in mice. Two-week-old male mice were intravenously injected with recombinant adeno-associated virus (AAV) serotype 9 (AAV9) carrying either a control vector or Pde5a -encoding construct. After 1 week, mice received BAPN water for 28 days. A , Schematic illustration of the experimental timeline. B , Survival curves for mice receiving normal water or BAPN water after injection with adeno-associated virus (AAV)-Pde5a or control AAV (AAV-null). Survival data were analyzed by the Kaplan-Meier method and compared using log-rank tests (n=10–25). C , Representative images of aortas from the indicated experimental groups. D , Maximal thoracic aorta diameter in mice from the indicated experimental groups (n=8). E , The incidence of aortic dissection formation in mice from the indicated experimental groups (n=10–25). F , The incidence of aortic rupture in mice from indicated experimental groups (n=10–25). G and H , Representative Hematoxylin and Eosin (H&E), elastic Verhoeff-Van Gieson (EVG), and Masson trichrome staining of the thoracic aortas in mice with injection of AAV-Pde5a or AAV-null ( G ), and quantification of elastin degradation based on a 4‐point scoring system ( H ; n=6). Scale bar=100 µm. I and J , Western blot analysis ( I ) and quantification ( J ) of MYH11 (myosin heavy chain 11), CNN1 (calponin 1), α-SMA (α‐smooth muscle actin), and OPN (osteopontin) protein levels in mice injected with AAV-Pde5a or AAV-null (n=6). Data are presented as mean±SEM. Two-way ANOVA followed by Tukey multiple-comparison test was used in D , H , and J , Fisher exact test was performed in E and F . Panel A created in BioRender. Mao, J. (2026) https://BioRender.com/qlycz8m .

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Exercise Attenuates Aortic Dissection Via PDE5A-Mediated Inhibition of Vascular Smooth Muscle Cell Phenotypic Switch

doi: 10.1161/ATVBAHA.125.324293

Figure Lengend Snippet: Overexpression of PDE5A (phosphodiesterase 5A) in vascular smooth muscle cells (VSMCs) rescues β-aminopropionitrile (BAPN)–induced aortic dissection in mice. Two-week-old male mice were intravenously injected with recombinant adeno-associated virus (AAV) serotype 9 (AAV9) carrying either a control vector or Pde5a -encoding construct. After 1 week, mice received BAPN water for 28 days. A , Schematic illustration of the experimental timeline. B , Survival curves for mice receiving normal water or BAPN water after injection with adeno-associated virus (AAV)-Pde5a or control AAV (AAV-null). Survival data were analyzed by the Kaplan-Meier method and compared using log-rank tests (n=10–25). C , Representative images of aortas from the indicated experimental groups. D , Maximal thoracic aorta diameter in mice from the indicated experimental groups (n=8). E , The incidence of aortic dissection formation in mice from the indicated experimental groups (n=10–25). F , The incidence of aortic rupture in mice from indicated experimental groups (n=10–25). G and H , Representative Hematoxylin and Eosin (H&E), elastic Verhoeff-Van Gieson (EVG), and Masson trichrome staining of the thoracic aortas in mice with injection of AAV-Pde5a or AAV-null ( G ), and quantification of elastin degradation based on a 4‐point scoring system ( H ; n=6). Scale bar=100 µm. I and J , Western blot analysis ( I ) and quantification ( J ) of MYH11 (myosin heavy chain 11), CNN1 (calponin 1), α-SMA (α‐smooth muscle actin), and OPN (osteopontin) protein levels in mice injected with AAV-Pde5a or AAV-null (n=6). Data are presented as mean±SEM. Two-way ANOVA followed by Tukey multiple-comparison test was used in D , H , and J , Fisher exact test was performed in E and F . Panel A created in BioRender. Mao, J. (2026) https://BioRender.com/qlycz8m .

Techniques: Over Expression, Dissection, Injection, Recombinant, Virus, Control, Plasmid Preparation, Construct, Staining, Western Blot, Comparison

Pharmacological inhibition of RUNX1 (runt-related transcription factor 1) with Ro5-3335 attenuates aortic dissection via PDE5A (phosphodiesterase 5A). Three-week-old male mice received β-aminopropionitrile (BAPN) water (1 g/kg per day) or normal drinking water and were subcutaneously injected with Ro5-3335 (20 mg/kg) or vehicle for 28 days. A , Schematic illustration of the experimental timeline. B , Survival curves for normal standard drinking water or BAPN water mice administered with Ro5-3335 or vehicle. Survival data were analyzed by the Kaplan-Meier method and compared using log-rank tests (n=10–27). C , Representative images of aortas in mice across experimental groups. D , Maximal thoracic aorta diameter in mice administered with Ro5-3335 or vehicle (n=8). E , The incidence of aortic dissection formation in mice from indicated experimental groups (n=10–27). F , The incidence of aortic rupture in mice from indicated experimental groups (n=10–27). G and H , Representative hematoxylin and eosin (H&E), elastic Verhoeff-Van Gieson (EVG), and Masson trichrome staining of the thoracic aortas in mice across experimental groups ( G ), and quantification of elastin degradation based on a 4‐point scoring system ( H ; n=6). Scale bar=100 µm. I and J , Western blot analysis ( I ) and quantification ( J ) of PDE5A, MYH11 (myosin heavy chain 11), CNN1 (calponin 1), α-SMA (α‐smooth muscle actin), and OPN (osteopontin) protein levels in mice across experimental groups (n=6). Data are presented as mean±SEM. Two-way ANOVA followed by Tukey multiple-comparison test was used in D , H , and J , Fisher exact test was performed in E and F . Panel A created in BioRender. Mao, J. (2026) https://BioRender.com/qlycz8m .

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Exercise Attenuates Aortic Dissection Via PDE5A-Mediated Inhibition of Vascular Smooth Muscle Cell Phenotypic Switch

doi: 10.1161/ATVBAHA.125.324293

Figure Lengend Snippet: Pharmacological inhibition of RUNX1 (runt-related transcription factor 1) with Ro5-3335 attenuates aortic dissection via PDE5A (phosphodiesterase 5A). Three-week-old male mice received β-aminopropionitrile (BAPN) water (1 g/kg per day) or normal drinking water and were subcutaneously injected with Ro5-3335 (20 mg/kg) or vehicle for 28 days. A , Schematic illustration of the experimental timeline. B , Survival curves for normal standard drinking water or BAPN water mice administered with Ro5-3335 or vehicle. Survival data were analyzed by the Kaplan-Meier method and compared using log-rank tests (n=10–27). C , Representative images of aortas in mice across experimental groups. D , Maximal thoracic aorta diameter in mice administered with Ro5-3335 or vehicle (n=8). E , The incidence of aortic dissection formation in mice from indicated experimental groups (n=10–27). F , The incidence of aortic rupture in mice from indicated experimental groups (n=10–27). G and H , Representative hematoxylin and eosin (H&E), elastic Verhoeff-Van Gieson (EVG), and Masson trichrome staining of the thoracic aortas in mice across experimental groups ( G ), and quantification of elastin degradation based on a 4‐point scoring system ( H ; n=6). Scale bar=100 µm. I and J , Western blot analysis ( I ) and quantification ( J ) of PDE5A, MYH11 (myosin heavy chain 11), CNN1 (calponin 1), α-SMA (α‐smooth muscle actin), and OPN (osteopontin) protein levels in mice across experimental groups (n=6). Data are presented as mean±SEM. Two-way ANOVA followed by Tukey multiple-comparison test was used in D , H , and J , Fisher exact test was performed in E and F . Panel A created in BioRender. Mao, J. (2026) https://BioRender.com/qlycz8m .

Techniques: Inhibition, Dissection, Injection, Staining, Western Blot, Comparison

Journal: bioRxiv

Article Title: Fibroblasts neurotrophin signaling sustains pathological vascular maturation in rheumatoid arthritis

doi: 10.64898/2026.03.12.711120

Figure Lengend Snippet: (A-C) Bar plots showing differential regulation of pericyte markers (RGS5, ABCC9), VSMC markers (CNN1, MYH11), and pan-mural cell markers (ACTA2, SMTN, TAGLN) in fibroblasts treated with (A) NGF, (B) BDNF, or (C) NT3 compared with untreated fibroblasts. Bar heights represent log₂ fold change, and bar color indicates the corresponding P values. (D) Schematic of the collagen gel contraction assay in which synovial fibroblasts were embedded in collagen I gels and treatment (E) Representative images of collagen I gel contraction following treatment with NGF, BDNF, or NT3 (F) Quantification of collagen I gel contraction expressed as percent contraction. Values represent mean ± standard deviation (SD). Individual data points indicate biological replicates. Statistical analysis was performed using one-way ANOVA, with P values shown. (G) Schematic illustrating the synovial organoid/explant experimental design. (H) Representative H&E image of synovial explant culture after 3 days of culture. (I) Whole-mount immunoluorescence images showing preservation of endothelial cells after 3 days in culture, marked by CD31 (magenta), and vascular smooth muscle cells marked by MYH11, surrounding CD31 vascular structures. (J) Whole-mount staining showing expression of neurotrophin receptors NGFR, NTRK1, NTRK2, and NTRK3 (green) around vascularature, together with CD31 (magenta) and DAPI (blue). (K) Synovial organoids embedded in Matrigel and treated with NGF, BDNF, or NT3, followed by fixation and staining for a-smooth muscle actin (aSMA) (brown) to visualize mural cells. All 20X images were cropped at similar scale. Scale bar, 150 µm.

Article Snippet: Following transfer, membranes were blocked for 20 minutes in Everyblot blocking buffer (Bio-Rad # 12010020) and incubated overnight at 4°C with primary antibodies from TrkA and TrkB antibody sampler kit against TrkA, TrkB, p-TrkA/TrkB(Cell Signaling Technology, #4638, 1:500), CNN1 (Proteintech-24855-1-AP), MYH11 (Proteintech-21404-1-AP), NGF (Abcam-ab52918) and GAPDH (Thermo Fisher Scientific, #MA5-15738),or beta-actin (Cell Signaling Technology, #3700).

Techniques: Collagen Gel Contraction Assay, Standard Deviation, Preserving, Staining, Expressing

Journal: bioRxiv

Article Title: Fibroblasts neurotrophin signaling sustains pathological vascular maturation in rheumatoid arthritis

doi: 10.64898/2026.03.12.711120

Figure Lengend Snippet: (A) RNAscope images showing transcript expression of pericyte marker RGS5 (yellow) and VSMC marker MYH11 (red) in fibroblasts treated with different neurotrophins, overlaid with DAPI (blue). Scale bar, 150 µm. All images were acquired at 20x magnification and cropped to the same scale.(B) Quantification of total numbers of RGS5+, MYH11+, and RGS5+/MYH11+ double-positive cells following treatment with the indicated neurotrophins.(C) mRNA expression of the pericyte marker RGS5 in fibroblasts transfected with siRNAs targeting NGFR or NTRK1, in the presence or absence of NGF stimulation.(D) mRNA expression of the VSMC marker MYH11 in fibroblasts transfected with siRNAs targeting NTRK2 or NTRK3, in the presence or absence of BDNF or NT3 stimulation, respectively.(E-F) mRNA expression of the mural cell marker CNN1 (calponin) in fibroblasts stimulated with NGF (E) in the presence or absence of a TRKA inhibitor, or stimulated with BDNF (F) in the presence or absence of an NTRK2 inhibitor.

Techniques: RNAscope, Expressing, Marker, Transfection

(A) Schematic illustration of chemically defined differentiation protocol from iPSCs to iSMCs. (B) Differentiation efficacy determined by flow cytometric analysis of CD140b positive cells. (Data is presented as mean ± SD). (C) Immunofluorescence analysis for SMC markers confirms SMC identity of iSMCs (alpha-smooth muscle actin (Actin) and calponin (CNN1) (both green), myosin heavy chain 11 (MHY11) (red), nuclei (DAPI, blue), scale bar 200 µm. (D) SMC score calculated as the mean expression of Travaglini SMC genes (MSigDB: M41672) in human pulmonary artery smooth muscle cells (GSE144274), human iPSC lines (HipSci resource) and merged expression values of untreated iSMCs from all lines and clones. (E) PCA analysis of untreated iSMCs based on 2000 most variable genes (Log2(TPM+1)) of three independent WT cell lines (dark green dots) and three clones per BMPR2-mutant of, whereas BMPR2 MutExDo (light green dots) and BMPR2 MutKiDo (medium green dots); average of each experimental groups depicted as cross. (F) Quantification of cell numbers relative to the respective vehicle control after 8d treatment. Data is presented as mean ± SD, n= 3 independent biological replicates per column indicated as dots. Exploratory two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A, and Activin A in combination with sotatercept, within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. (G) Quantification of apoptotic signal measured as caspase3/7 activity and normalization to respective cell number after 5d treatment. Data is presented as mean ± SD, n= 6-9 independent biological replicates per column indicated as dots. Two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A and Activin A in combination with sotatercept within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001.

Journal: bioRxiv

Article Title: iPSC modeling of pulmonary arterial hypertension to uncover pathomechanisms and unrecognized modes of action of sotatercept

doi: 10.64898/2026.03.12.711267

Figure Lengend Snippet: (A) Schematic illustration of chemically defined differentiation protocol from iPSCs to iSMCs. (B) Differentiation efficacy determined by flow cytometric analysis of CD140b positive cells. (Data is presented as mean ± SD). (C) Immunofluorescence analysis for SMC markers confirms SMC identity of iSMCs (alpha-smooth muscle actin (Actin) and calponin (CNN1) (both green), myosin heavy chain 11 (MHY11) (red), nuclei (DAPI, blue), scale bar 200 µm. (D) SMC score calculated as the mean expression of Travaglini SMC genes (MSigDB: M41672) in human pulmonary artery smooth muscle cells (GSE144274), human iPSC lines (HipSci resource) and merged expression values of untreated iSMCs from all lines and clones. (E) PCA analysis of untreated iSMCs based on 2000 most variable genes (Log2(TPM+1)) of three independent WT cell lines (dark green dots) and three clones per BMPR2-mutant of, whereas BMPR2 MutExDo (light green dots) and BMPR2 MutKiDo (medium green dots); average of each experimental groups depicted as cross. (F) Quantification of cell numbers relative to the respective vehicle control after 8d treatment. Data is presented as mean ± SD, n= 3 independent biological replicates per column indicated as dots. Exploratory two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A, and Activin A in combination with sotatercept, within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. (G) Quantification of apoptotic signal measured as caspase3/7 activity and normalization to respective cell number after 5d treatment. Data is presented as mean ± SD, n= 6-9 independent biological replicates per column indicated as dots. Two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A and Activin A in combination with sotatercept within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001.

Article Snippet: On day 12, for quantification of HPAH phenotypic intracellular or transmembrane markers, cells were fixed with 4% paraformaldehyde (Sigma-Aldrich, cat. no.158127) for 10 min, ice-cold 90% methanol (JT Baker, cat. no. 8045) for 15 min, permeabilized with 1% tritonX-100 (Sigma-Aldrich, cat. no. 93443) PBS pH7.4 (1X) (Gibco, cat. no. 70011-036) supplemented with 1 % BSA (Sigma-Aldrich, cat. no. A9418) for 20 min and stained with the primary antibody CNN1 (Santa Cruz Biotechnology, cat. no. sc-53136, RRID:AB_793529, mouse, 1:50) or ITGA2 (Abcam, cat. no. ab181548, RRID:AB_2847852, rabbit, 1:100) in PBS pH7.4 (1X) (Gibco, cat. no. 70011-036) supplemented with 1 % BSA (Sigma-Aldrich, cat. no. A9418) and 1% tritonX-100 (Sigma-Aldrich, cat. no. 93443) over night at 4 °C.

Techniques: Immunofluorescence, Expressing, Clone Assay, Mutagenesis, Control, Activity Assay

( A ) Functional contraction assay with iSMCs derived from representative WT cell line or BMPR2 Mut clones. Double arrow indicates well diameter as starting position for contraction. ( B ) Contraction was determined as the area of the gel disc after 72 h relative to the initial area of the iSMC-gel-discs. Data is presented as mean ± SD, n= 3 independent biological replicates per column indicated as dots. Exploratory two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A and Activin A in combination with sotatercept within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. ( C ) Quantification of the ratio of phosphorylated myosin light chain (pMLC) to total MLC by automated capillary-based western blot analysis. Data is presented as mean ± SD, n= 3 independent biological replicates per column indicated as dots. Exploratory two-way ANOVA with Sidak’s multiple-comparisons was used to compare genotype differences within Activin A or Activin A in combination with sotatercept, followed by two-tailed unpaired Student’s t-tests for treatment comparisons within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. ( D, F ) Immunofluorescence staining of components of the contractile apparatus in iSMCs of one iPSC line / clone representative for the respective genotype (BMPR2 WT , BMPR2 MutExDo , BMPR2 MutKiDo ). Actin and calponin (CNN1) (green), nuclei DAPI (blue), scale bar 200 µm. ( E ) Quantification of actin from immunofluorescence images expressed as mean fluorescence intensity (MFI) normalized to cell number (DAPI). Data is presented as mean ± SD, n= 6 independent biological replicates per column indicated as dots. Two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A and Activin A in combination with sotatercept within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. ( G ) Quantification of CNN1 from flow cytometry analysis (MFI). Data is presented as mean ± SD, n= 3 independent biological replicates per column indicated as dots. Exploratory two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A and Activin A in combination with sotatercept within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. ( H ) Heatmaps showing differentially expressed genes with pro-contractile function in Activin A vs. Activin A + sotatercept treated iSMCs each condition normalized to the untreated control. For contraction gene set scoring (MSigDB: M1429), bulk RNA-seq expression values were analysed using a gene-by-sample FPKM matrix (mean FPKM of three independent biological replicates per group, Activin A and Activin A + sotatercept). Stars (*) highlight mentioned genes in the respective text.

Journal: bioRxiv

Article Title: iPSC modeling of pulmonary arterial hypertension to uncover pathomechanisms and unrecognized modes of action of sotatercept

doi: 10.64898/2026.03.12.711267

Figure Lengend Snippet: ( A ) Functional contraction assay with iSMCs derived from representative WT cell line or BMPR2 Mut clones. Double arrow indicates well diameter as starting position for contraction. ( B ) Contraction was determined as the area of the gel disc after 72 h relative to the initial area of the iSMC-gel-discs. Data is presented as mean ± SD, n= 3 independent biological replicates per column indicated as dots. Exploratory two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A and Activin A in combination with sotatercept within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. ( C ) Quantification of the ratio of phosphorylated myosin light chain (pMLC) to total MLC by automated capillary-based western blot analysis. Data is presented as mean ± SD, n= 3 independent biological replicates per column indicated as dots. Exploratory two-way ANOVA with Sidak’s multiple-comparisons was used to compare genotype differences within Activin A or Activin A in combination with sotatercept, followed by two-tailed unpaired Student’s t-tests for treatment comparisons within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. ( D, F ) Immunofluorescence staining of components of the contractile apparatus in iSMCs of one iPSC line / clone representative for the respective genotype (BMPR2 WT , BMPR2 MutExDo , BMPR2 MutKiDo ). Actin and calponin (CNN1) (green), nuclei DAPI (blue), scale bar 200 µm. ( E ) Quantification of actin from immunofluorescence images expressed as mean fluorescence intensity (MFI) normalized to cell number (DAPI). Data is presented as mean ± SD, n= 6 independent biological replicates per column indicated as dots. Two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A and Activin A in combination with sotatercept within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. ( G ) Quantification of CNN1 from flow cytometry analysis (MFI). Data is presented as mean ± SD, n= 3 independent biological replicates per column indicated as dots. Exploratory two-way ANOVA with genotype and treatment as factors was followed by Sidak’s multiple-comparisons test to compare genotype differences within Activin A treatment and to compare effects of Activin A and Activin A in combination with sotatercept within each genotype. ns, not significant; *p < 0.05, **p < 0.01, ***p < 0.001. ( H ) Heatmaps showing differentially expressed genes with pro-contractile function in Activin A vs. Activin A + sotatercept treated iSMCs each condition normalized to the untreated control. For contraction gene set scoring (MSigDB: M1429), bulk RNA-seq expression values were analysed using a gene-by-sample FPKM matrix (mean FPKM of three independent biological replicates per group, Activin A and Activin A + sotatercept). Stars (*) highlight mentioned genes in the respective text.

Techniques: Functional Assay, Contraction Assay, Derivative Assay, Clone Assay, Western Blot, Two Tailed Test, Immunofluorescence, Staining, Fluorescence, Flow Cytometry, Control, RNA Sequencing, Expressing

$A , B , Flow cytometry analysis of single‐cell suspension from the abdominal aorta of mice treated with Ang II. A , presents the quantification of MYH11 + cells; B , shows the quantification of CD45 + CD68 + CD11b + cells. The data represent the cell fraction of all aortic cells (mean±SEM, n=6). C, Flow cytometry analysis and quantification of abdominal aorta single‐cell suspension from Ang II‐treated mice showing the proportion of MYH11 + cells expressing macrophage markers. These data show the fraction of CD68 + CD11b + cells as a percentage of MYH11 + cells (mean±SEM, n=6). D , A t‐SNE clustering analysis of flow cytometry data from the abdominal aorta of Ang II‐treated mice was performed. MYH11 + cells were clustered based on the expression of CNN1, ACTA2, TAGLN, CD45, CD68, CD11b, CD38, TNFα, IL1b, and IL6, resulting in distinct clusters presented on the t‐SNE plot. E , A heat map representation illustrates the relative expression of SMC, macrophage, and inflammatory markers based on the mean fluorescence intensity for each cluster. F – H , The relative abundance of distinct clusters as a proportion of MYH11 + cells (mean±SEM, n=6). ACTA2 indicates actin alpha 2; Ang II, angiotensin II; Apoe −/− , apolipoprotein E knockout mice; MYH11, myosin heavy chain 11; cGAS, cyclic GMP‐AMP synthase; CNN, Calponin 1; IL‐1b, interleukin 1 beta; IL‐6, interleukin 6; MYH11, myosin heavy chain 11; Nox4 TG, Nox4 transgenic mice; Nox4 −/− , Nox4 knockout mice; SMC, smooth muscle cell; STING, stimulator of interferon gene; TAGLN, transgelin; TNFα, tumor necrosis factor alpha; t‐SNE, t‐distributed stochastic neighbor embedding; and TUBB, tubulin beta.$

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: Aging‐Associated Nox4 ‐Mediated Mitochondrial Reactive Oxygen Species and DNA Damage Promote Vascular Cell Reprogramming and Aortic Remodeling in Abdominal Aneurysms

doi: 10.1161/JAHA.125.044949

Figure Lengend Snippet: A , B , Flow cytometry analysis of single‐cell suspension from the abdominal aorta of mice treated with Ang II. A , presents the quantification of MYH11 + cells; B , shows the quantification of CD45 + CD68 + CD11b + cells. The data represent the cell fraction of all aortic cells (mean±SEM, n=6). C, Flow cytometry analysis and quantification of abdominal aorta single‐cell suspension from Ang II‐treated mice showing the proportion of MYH11 + cells expressing macrophage markers. These data show the fraction of CD68 + CD11b + cells as a percentage of MYH11 + cells (mean±SEM, n=6). D , A t‐SNE clustering analysis of flow cytometry data from the abdominal aorta of Ang II‐treated mice was performed. MYH11 + cells were clustered based on the expression of CNN1, ACTA2, TAGLN, CD45, CD68, CD11b, CD38, TNFα, IL1b, and IL6, resulting in distinct clusters presented on the t‐SNE plot. E , A heat map representation illustrates the relative expression of SMC, macrophage, and inflammatory markers based on the mean fluorescence intensity for each cluster. F – H , The relative abundance of distinct clusters as a proportion of MYH11 + cells (mean±SEM, n=6). ACTA2 indicates actin alpha 2; Ang II, angiotensin II; Apoe −/− , apolipoprotein E knockout mice; MYH11, myosin heavy chain 11; cGAS, cyclic GMP‐AMP synthase; CNN, Calponin 1; IL‐1b, interleukin 1 beta; IL‐6, interleukin 6; MYH11, myosin heavy chain 11; Nox4 TG, Nox4 transgenic mice; Nox4 −/− , Nox4 knockout mice; SMC, smooth muscle cell; STING, stimulator of interferon gene; TAGLN, transgelin; TNFα, tumor necrosis factor alpha; t‐SNE, t‐distributed stochastic neighbor embedding; and TUBB, tubulin beta.

Article Snippet: Samples were incubated in 200 μL FACS buffer with a mix of intracellular marker antibodies: MYH11‐FITC (Biorbyt, Durham, NC; orb13894), CNN1 (calponin‐1)‐AlexaFluor 555 (Bioss; bs‐0095R‐A555), ACTA2‐DyLight 405, TAGLN‐PerCP (Novus Biologicals, Centennial, CO; NBP2‐34522V, NBP2‐47689PCP), TNFα (tumor necrosis factor alpha)‐Brilliant Violet 510 (BioLegend; 506 339), IL1b‐APC‐Cy7 (LifeSpan Bio, Newark, CA; LS‐C730762‐100), and IL6‐PerCP‐eFluor 710 (Thermo Fisher; 46–7061‐80).

Techniques: Flow Cytometry, Single Cell, Suspension, Expressing, Fluorescence, Knock-Out, Transgenic Assay

Smooth muscle cells of great arteries (A) Featureplots demonstrating distinct distributions and specificity of contractile SMC markers. (B) Immunofluorescence (IF) staining for CNN1 (green) of coronary tissue showing medial and adventitial (Adv)population. (C) Featureplot of RGS5 , a commonly used pericyte marker delineating multiple populations. (D) Differentially expressed genes (DEGs) ranking between the two major smooth muscle cell clusters. (E) Dot plot summarizing Gene Ontology (GO) analysis of DEGs between the two clusters containing SMCs. (F) Featureplot and RNAscope of microvascular specific marker. (G) Featureplot of intimal/medial SMC markers demonstrating spatial or functional heterogeneity among SMC subtypes. (H) Spatial transcriptomics maps and RNAscope demonstrating distinct spatial localization intimal/medial SMC clusters/markers from (G). See also and .

Journal: Cell Genomics

Article Title: A cell and transcriptome atlas of human arterial vasculature

doi: 10.1016/j.xgen.2025.101034

Figure Lengend Snippet: Smooth muscle cells of great arteries (A) Featureplots demonstrating distinct distributions and specificity of contractile SMC markers. (B) Immunofluorescence (IF) staining for CNN1 (green) of coronary tissue showing medial and adventitial (Adv)population. (C) Featureplot of RGS5 , a commonly used pericyte marker delineating multiple populations. (D) Differentially expressed genes (DEGs) ranking between the two major smooth muscle cell clusters. (E) Dot plot summarizing Gene Ontology (GO) analysis of DEGs between the two clusters containing SMCs. (F) Featureplot and RNAscope of microvascular specific marker. (G) Featureplot of intimal/medial SMC markers demonstrating spatial or functional heterogeneity among SMC subtypes. (H) Spatial transcriptomics maps and RNAscope demonstrating distinct spatial localization intimal/medial SMC clusters/markers from (G). See also and .

Article Snippet: CNN1 , OriGene , SKU: TA327614; RRID: AB_3716482.

Techniques: Immunofluorescence, Staining, Marker, RNAscope, Functional Assay

Review

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Structured Review

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1) Product Images from "Fibroblasts neurotrophin signaling sustains pathological vascular maturation in rheumatoid arthritis"

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