endothelial cell growth microvascular media egmv2  (PromoCell)

Journal: bioRxiv

Article Title: Urban PM 2.5 at Realistic Environmental Concentrations Impairs Blood–Brain Barrier Integrity and Enhances LOX-1 Expression in Human Brain Endothelial Cells

doi: 10.64898/2026.01.29.702473

Figure Lengend Snippet: A . Physiological rationale: Ambient PM2.5 exposure is epidemiologically linked to increased ischemic stroke risk. This in vitro model simulates the real-life scenario of pre-existing PM2.5 exposure followed by ischemic stroke and subsequent reperfusion. B . Primary adult male HBMEC were exposed to 5, 15, 75, or 300 μg/m 3 PM 2.5 for 48h in total. To compare with the effects of physiological ischemic-like injury, some plates were exposed to hypoxia (1% O 2 ) and glucose deprived media (HGD) for 3h after the initial 24h incubation. Following HGD or normoxia, cells were reperfused with nutrient-enriched media and incubated with PM 2.5 at normoxic (21% O 2 ) conditions as a reference for resolution of ischemia. Barrier integrity, cell viability, reactive oxygen species (ROS), inflammation and LOX-1 expression was assessed. Figure created in BioRender.

Article Snippet: Primary adult male HBMEC were purchased from Innoprot (Spain, Catalog number: P10361, Lot number: 111224CS).

Techniques: In Vitro, Incubation, Expressing

Journal: bioRxiv

Article Title: Urban PM 2.5 at Realistic Environmental Concentrations Impairs Blood–Brain Barrier Integrity and Enhances LOX-1 Expression in Human Brain Endothelial Cells

doi: 10.64898/2026.01.29.702473

Figure Lengend Snippet: Adult male HBMEC were exposed to vehicle or PM 2.5 (5, 15, 75, or 300 μg/m 3 ) for 24h and incubated for 3h in normoxia- or hypoxia and glucose deprivation (HGD) followed by 24h reperfusion. A . Live cell count (CyQUANT nuclear stain) decreased when exposed to ≥75 μg/m 3 PM 2.5 compared to vehicle. HGD treatment reduced live cell count compared to normoxia but did not differ between particle treated groups. B . Reactive oxygen species (ROS) signal (DCHF-DA) normalized to live cell count. Relative ROS levels increased dose-dependently with PM 2.5 concentration, with significant increase observed at PM 2.5 ≥75 μg/m 3 , in comparison to normoxia vehicle. ROS levels were uniformly elevated following HGD across all doses in comparison to normoxia vehicle and significantly higher than untreated HBMEC. (n=12 technical replicates for vehicle and 5, n=8 technical replicates for 15, 75 and 300) C . Analysis of crystal violet-stained HBMEC shows a longer maximum cellular length when treated with ≥15 μg/m 3 PM 2.5 . (n=21-37 individual cells) D . Representative images of crystal violet-stained HBMEC visualizing a differentiated morphology in cells treated with higher PM 2.5 concentration, where cells appear more elongated and expanding towards neighbouring cells. Data presented as mean ± SD. Statistical significance assessed through Kruskal-Wallis test within treatment groups (Normoxia/HGD) and Mann-Whitney test between groups with different treatment (300 normoxia/vehicle HGD). *p<0.05. ***p<0.001. ****p<0.0001.

Article Snippet: Primary adult male HBMEC were purchased from Innoprot (Spain, Catalog number: P10361, Lot number: 111224CS).

Techniques: Incubation, Cell Characterization, CyQUANT Assay, Staining, Concentration Assay, Comparison, MANN-WHITNEY

Journal: bioRxiv

Article Title: Urban PM 2.5 at Realistic Environmental Concentrations Impairs Blood–Brain Barrier Integrity and Enhances LOX-1 Expression in Human Brain Endothelial Cells

doi: 10.64898/2026.01.29.702473

Figure Lengend Snippet: Western Blot assessment of adult male HBMEC exposed to vehicle, 5, 15, 75, or 300 μg/m 3 PM 2.5 during normoxia or ischemic-like injury with hypoxia, glucose deprivation and reperfusion (HGD). A . Representative Western Blot image of IL-6 and β-actin band migration. B . Signal quantification of 25kDa IL-6 shows no difference between PM 2.5 exposure or HGD treated group. C . Signal quantification of 17kDa IL-6 shows dose-dependency with higher IL-6 expression from higher PM 2.5 exposure, with significant increase ≥75 μg/m 3 and from HGD treatment compared to vehicle. D . Representative Western Blot image of LOX-1 and β-actin. E . Signal quantification of LOX-1 displays a dose-dependent increase in LOX-1 with exposure to ≥15 μg/m 3 PM 2.5 or HGD. (n=4-7 technical replicates). Data presented as mean +-SD. Statistical significance assessed by Kruskal-Wallis test. *p<0.05, **p<0.01.

Article Snippet: Primary adult male HBMEC were purchased from Innoprot (Spain, Catalog number: P10361, Lot number: 111224CS).

Techniques: Western Blot, Migration, Expressing

Journal: JACC: Basic to Translational Science

Article Title: Human Left Ventricle circRNA-miRNA-mRNA Network Analyses Reveal a Novel Proangiogenic Role for circNPHP1 Under Ischemic Conditions

doi: 10.1016/j.jacbts.2025.101468

Figure Lengend Snippet: CircNPHP1 Expression in the Human LV and Cultured ECs Cardiac CircNPHP1 levels are higher in patients with IHD with/without associated T2DM and in cultured endothelial cells (human cardiac microvascular endothelial cells and human umbilical vein endothelial cells) exposed to disease-mimicking conditions, such as hypoxia and hypoxia, combined with increased D-glucose level (High glu). (A) Preliminary screening of circRNAs (identified from predicted networks) in endothelial cells: human cardiac microvascular endothelial cells were cultured in normal conditions, hypoxia (1% Oxygen), and hypoxia-high glucose (High Glu) (25 mmol/L D-glucose) conditions, respectively. After 48 hours, cells were harvested for quantitative real-time polymerase chain reaction (qRT-PCR) for the analysis and validation of indicated circRNAs. RNA expression is relative to normoxia. 18S is used as housekeeping gene; n = 3. Data are expressed as mean ± SEM and were assessed by 1-way analysis of variance with Dunnett's post hoc test. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ns = not significant. CircSTX17 was also tested but the levels were undetectable; hence, it is not included in the plot. CircNPHP1 emerges as an interesting candidate for further exploration and validation. (B) Comparison of expression levels of circNPHP1 and its linear counterpart in different patient groups—RNAseq (LV biopsies, cohort 1). Log-transformed normalized (FPM) counts of circNPHP1 and linear NPHP1 are plotted as a boxplot showing the median and IQR. Comparison between patient groups and calculation of P values was done using the R package Limma. CircNPHP1 is significantly up-regulated in IHD+T2DM compared with IHD and non-IHD ( P < 0.05). Linear NPHP1 is expressed at very low levels and does not change between the different patient groups (IHD [n = 12], IHD+T2DM [n = 11], non-IHD [n = 12]). (C) qRT-PCR of circNPHP1 (left) and linear NPHP1 (right) of LV biopsies (cohort 1) from non-IHD (n = 12), IHD (n = 12), and IHD+T2DM (n = 11), and (D) of LV biopsies (cohort 2) from non-IHD (n = 10), IHD (n = 10), IHD+T2DM (n = 10) of patients. RNA expression is relative to non-IHD; 18S is used as housekeeping gene. Data are expressed as mean ± SEM and were assessed by Kruskal-Wallis test followed by Dunn's post hoc test. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ns = not significant. Abbreviations as in .

Article Snippet: Human cardiac microvascular endothelial cells (HCMECs) (Promocell: C-12285 [mono-doner], Lot numbers: 440Z021.5 [male], 440Z021.4 [male], 446Z001.1 [female], 492Z009.4 [female]) and human umbilical vein endothelial cells (HUVECs) (Promocell: C-12208 [poly-donor], Lot numbers: 447Z004, 450Z015, 466Z022, 503Z026) were cultured on 0.2% gelatin (Sigma-Aldrich) in endothelial cell growth medium MV and endothelial cell growth medium 2 (EGM2) (PromoCell).

Techniques: Expressing, Cell Culture, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Biomarker Discovery, RNA Expression, Comparison, Transformation Assay

Journal: JACC: Basic to Translational Science

Article Title: Human Left Ventricle circRNA-miRNA-mRNA Network Analyses Reveal a Novel Proangiogenic Role for circNPHP1 Under Ischemic Conditions

doi: 10.1016/j.jacbts.2025.101468

Figure Lengend Snippet: CircNPHP1 and Linear NPHP1 Expression in Different Cell Types (A) Single-cell and single-nuclei RNAseq data from healthy donor hearts (human heart atlas ) of NPHP1 gene as log2 transformed counts per million (CPM). Each dot indicates the data point as 1 donor (source: human heart atlas ). (B) RNAseq: A preliminary screen was carried out in different cells (sample size = 2 each) to determine absolute levels of circNPHP1 and linear NPHP1 in human umbilical vein endothelial cells (HUVECs or HU), human cardiac microvascular endothelial cells (HCMECs or HC), human AC16 cardiomyocytes (ACs), and cardiac fibroblasts (CFs). (C) HCMEC (left) and HUVEC cells (right) were cultured in either normal conditions, hypoxia (1% oxygen), or hypoxia combined with high glucose (25 Mm D-glucose). After 48 hours, cells were harvested for quantitative real-time polymerase chain reaction for the analysis of circNPHP1 (n = 3). Fold change in RNA expression is relative to normoxia; 18S is used as housekeeping gene. Data are expressed as mean ± SEM and were assessed by 1-way analysis of variance with Dunnett's post hoc test. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ns = not significant. (D) 2 μg of total RNA from HUVEC cells were digested with RNase R enzyme followed by quantitative real-time polymerase chain reaction analysis of circNPHP1 and linear NPHP1 (n = 3). Fold change in RNA expression is relative to undigested RNA; 18S is used as housekeeping gene. Data are expressed as mean ± SEM and were assessed by unpaired Student's t -test. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ns = not significant. Abbreviations as in .

Article Snippet: Human cardiac microvascular endothelial cells (HCMECs) (Promocell: C-12285 [mono-doner], Lot numbers: 440Z021.5 [male], 440Z021.4 [male], 446Z001.1 [female], 492Z009.4 [female]) and human umbilical vein endothelial cells (HUVECs) (Promocell: C-12208 [poly-donor], Lot numbers: 447Z004, 450Z015, 466Z022, 503Z026) were cultured on 0.2% gelatin (Sigma-Aldrich) in endothelial cell growth medium MV and endothelial cell growth medium 2 (EGM2) (PromoCell).

Techniques: Expressing, Transformation Assay, Cell Culture, Real-time Polymerase Chain Reaction, RNA Expression