Journal: Nature Communications

Article Title: Inhibition of lysyl oxidases synergizes with 5-azacytidine to restore erythropoiesis in myelodysplastic and myeloid malignancies

doi: 10.1038/s41467-023-37175-8

Figure Lengend Snippet: a The LOXL2 gene expression in BM cellular fractions of n = 20 MN patients (RT-qPCR), including n = 14 MSCs, n = 15 MNCs, n = 16 CD71 + cells and n = 12 CD34 + cells; median±interquartile ranges (IQR); Two-sided Mann-Whitney U test. b The LOX/LOXL gene expression in ex-vivo expanded MSCs (RT-qPCR). Individual data points for MN patients versus age-matched HY are presented; median±IQR; Two-sided Mann-Whitney U test. c Pan-LOX/LOXL activity was measured in BM plasma of MN and HY using Lysyl Oxidase Activity Assay Kit. The background level of pan-LOX/LOXL activity (signal-to-noise) was controlled by the treatment with a pan-LOX/LOXL inhibitor BAPN. The activity was normalized to the total protein content in BM plasma showing relative fluorescence units (RFU) per µg/ml; Two-sided Mann-Whitney U -test at different dilutions between MN and HY cohorts using mean data from duplicates for each dilution. d , e LOXL2 protein concentration and activity in HY ( n = 15) vs MN patients ( n = 109). The background level of LOXL2 activity (signal-to-noise) was controlled by BAPN treatment. f , g LOXL2 protein concentrations and activity are shown in MN patients stratified by bone marrow fibrosis (BMF) grade (BMF ≤ 1, n = 76 vs BMF > 1, n = 33). For d – g , the data are displayed as median ± IQR. Two-sided Mann-Whitney U -test ( d , e ) and Kruskal-Wallis test with Dunn’s multiple comparisons ( f , g ) were performed. h , i BM MSCs ( h ) and HSPCs ( i ) from n = 11 MN and n = 3 HY were treated daily with the indicated concentrations of PXS-5505 for 11 days (MSCs) or 4 days (HSPCs), and cell viability was assessed using CellTiter-Glo assay; mean ± SD of 5 or 6 cell culture replicates. j Ex-vivo expanded MSCs were treated with indicated concentrations of PXS-5505 or BAPN every 24 h and pan-LOX/LOXL activity in supernatants of MSC culture was measured using the Lysyl Oxidase Activity Assay Kit. Combined data for n = 5 MN patients are presented as mean of fold changes ±SD; RM one-way ANOVA with Tukey’s multiple comparisons for each time-point. k Production of cross-linked collagen was assessed in MSC-derived fibroblasts of n = 2 MDS-BMF samples treated for 7 days using PXS-5505 or BAPN. The data are mean of fibroblast culture quadruplicates (top) and triplicates (bottom)±SD; Ordinary one-way ANOVA with Tukey’s multiple comparisons. P < 0.05 indicated statistical significance. Source data are provided as a Source Data file.

Article Snippet: After 24 h of culture, PBS (vehicle control) or PXS-5505 or 5-AZA or BAPN (TCI, A0796) was added at indicated concentrations and supernatants were harvested after 0 h, 3 h, 6 h, 12 h and 24 h. Cells were treated with inhibitors again and supernatants were harvested at 27 h, 30 h, 36 h and 48 h for pan-LOX/LOXL activity measurement using the Lysyl Oxidase Activity Assay Kit (Fluorometric).

Techniques: Gene Expression, Quantitative RT-PCR, MANN-WHITNEY, Ex Vivo, Activity Assay, Clinical Proteomics, Fluorescence, Protein Concentration, Glo Assay, Cell Culture, Derivative Assay

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: Survival curves of (A) male C57BL/6J (3-4 weeks old) mice administered BAPN 0.1%, 0.3%, or 0.5% wt/vol (N=10/concentration) for 12 weeks, (B) five mouse strains (male, 3-4 weeks old) administered BAPN 0.5% (N=19-21/mouse strain), (C) male C57BL/6J mice started BAPN 0.5% at 3 (N=38), 4 (N=15), or 26 (N=15) weeks of age, and (D) male (N=24) and female (N=23) C57BL/6J mice administered BAPN 0.5% at 4 weeks of age. P values were determined by Log-Rank analysis. P=0.008 for BAPN 0.1% vs 0.3% and P<0.001 for BAPN 0.1% vs 0.5% (A); P values for (B) are detailed in ; P=0.003 for 4- and 26-week comparison, and P<0.001 for 3- vs 4-week comparison and 3- vs 26- week comparison (C).

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: Concentration Assay, Comparison

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: BAPN (0.5%) was administered in drinking water for 12 weeks in 3- week-old male mice. In situ images of thoracic aortas for all survived mice with C57BL/6J (A) , C57BL/6N (B) , or B6/129SF1 (C) mouse strain after 12 weeks of BAPN administration.

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: In Situ

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: BAPN (0.5% wt/vol) was administered in drinking water for 12 weeks. Necropsy was performed for mice that died during BAPN administration within 12 weeks. (A) Examples of ex vivo images showing ascending or descending aortic rupture. (B) Death rate attributed to aortic rupture in the ascending versus descending aortic regions of male and female mice. Necropsy was performed on 78 male mice and 21 female mice. Incidence based on aortic rupture location was analyzed by Chi-square in male mice (P<0.001), and by Fisher-exact test in female mice (P<0.001).

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: Ex Vivo

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: Ultrasound of ascending aortas in young (4-week-old) C57BL/6J mice administered vehicle (drinking water only) versus BAPN (A) and comparing dilatation when BAPN administration was initiated in young (4-week-old) versus mature (26-week-old) mice (B) . Data in A and B were analyzed using R mixed effects model. BAPN versus vehicle at week 12 in male and female mice: P=0.014 and P=0.012, respectively (A) . Young versus Mature within males: P<0.001; and Young versus Mature within females: P=0.01 (B) . In situ measurements of maximal diameters of ascending/arch regions (C) and descending thoracic regions (D) in 4-week-old C57BL/6J mice administered vehicle versus BAPN. In situ measurements of maximal diameters of ascending/arch regions (E) and descending thoracic regions (F) in 4- versus 26-week-old C57BL/6J mice administered BAPN. Data in C-F were analyzed using two-way ANOVA with the Bonferroni correction for the post hoc test.

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: In Situ

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: Ex vivo measurements of maximal diameters of suprarenal aortas in 4-week-old C57BL/6J mice administered vehicle versus BAPN (A) and in 4- versus 26-week-old C57BL/6J mice administered BAPN (B) .

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: Ex Vivo

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: BAPN was administered in drinking water for 1, 2, 3, or 4 weeks in 3-4-week-old male C57BL/6J mice. (A) Examples of in situ images of thoracic aortas. (B) Hematoxylin-eosin (H&E) and Verhoff staining images of ascending aortic dissection and descending thoracic dissection.

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: In Situ, Staining, Dissection

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: Representative images of hematoxylin-eosin (H&E), Verhoeff iron hematoxylin, Movat’s pentachrome, α-SMA (smooth muscle actin), and CD68 staining in ascending and descending aortas of mice with BAPN administration for 12 weeks. The approximate location of the tissue section from the ascending and descending region is indicated by the blue line. * indicates false lumen.

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: Staining

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: Representative images of Verhoeff staining in ascending and descending aortas from mice administered for 12 weeks with BAPN.

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: Staining

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: Ascending and proximal descending thoracic aortas were harvested from 4-week-old male C57BL/6J mice administered either vehicle or BAPN for 7 days. (A) The cartoon shows the aortic regions where tissue was acquired for contractility analysis. The cartoon was created with BioRender.com. The contractile force in response to 5-HT (5-hydroxytryptamine) was measured in aortic rings harvested from (B) ascending and (C) descending thoracic regions. N=4 for vehicle and N=8 for BAPN group. Data were analyzed using Sigmoid function of R. P<0.001 and P=0.04 for comparing the inflection point values (distance from origin) between vehicle and BAPN groups in the ascending and descending thoracic aortic regions, respectively.

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques:

Journal: bioRxiv

Article Title: β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

doi: 10.1101/2023.10.22.563474

Figure Lengend Snippet: (A) Principle component analysis for unfiltered transcriptomes of ascending and proximal descending thoracic aortas harvested from 4- week-old male C57BL/6J mice administered either vehicle or BAPN for 7 days. (B) Z- scored heatmap of differentially expressed genes (DEGs) in the interaction analysis between sexes and administration of two-way ANOVA. Box plots for genes related to (C) smooth muscle cell contraction, (D) cell proliferation, and (E) extracellular maturation (ECM). n=5 biological replicates per group.

Article Snippet: BAPN (0.1%, 0.3%, or 0.5% wt/vol; CAS: 2079-89-2; Cat # A3134-25G, Millipore-Sigma or Cat # A0796-500G, TCI Co.) was administered via drinking water when mice were 3- 4 or 26 weeks of age.

Techniques: