Journal: Cardio-oncology

Article Title: Radiation- and age-related vascular dysfunction as an early indicator of cardiovascular risk: a long-term study in the ApoE −/− mouse model of atherosclerosis

doi: 10.1186/s40959-025-00395-6

Figure Lengend Snippet: Experimental set-up for irradiation and OCT-imaging of the murine Arteria saphena. A For irradiation in the X-ray device (1), anesthetized animals were positioned on their left side and secured on a Plexiglas holder. The bent right leg and lower abdomen were shielded with lead to protect them from radiation, ensuring that only the inner side of the left lower leg remained within the irradiation field. The exposure area beneath the irradiation window was defined by a collimator plate made of a bismuth-lead-tin alloy (MCP-96) with copper cutouts (2). Up to five animals were irradiated simultaneously on an underlying Plexiglas plate (3). The mesures are given in centimeters. B The OCT system for vascular imaging of the A. saphena operates using near-infrared light emitted by a diode (1), which is transmitted to the scanner head (2) via fiber optic cables (3). Within the scanner head the incoming light is collimated to a beam of 2.4 mm in diameter through a collimator (4) (focal length = 12 mm) and subsequently divided into a reference and probe beam of equal diameter with a beam splitter. To scan the arterial surface, the probe beam is diffracted via two galvanometric scanners (5) (Cambridge Technologies, Planegg) and focused through an achromatic lense (6) (focal length = 25.4 mm, diameter = 15 mm). The light reflected by the arterial surface and the reference beam that has been reflected by a mirror are then recombined by the beam splitter. Fiber optic cables lead the resulting interference signal through a collimator (focal length = 40 mm) and to a spectrometer to be spectrally analyzed with a diffraction grating (1200 lines/mm). The interference spectrum is then focused through an achromatic lense (focal length = 75 mm) and detected with a silicon detector (LIS-1024, pixel size: 7.8 μm × 125 μm × 1024 px, Photon Vision Systems Inc., Homer, USA). A Fast Fourier Transform of the interference signal provides depth-resolved information about the arterial tissue. C Representative recording of the A. saphena (white arrows) and Vena saphena medialis (grey arrows) of a C57BL/6 mouse aged 8 weeks, one day after irradiation with 2 Gy. The upper picture row in the foreground represents 2-D cross sectional OCT-images. The picture row below in the background are video-recordings to orientate on the tissue. Left: Vessel diameter at rest after application of physiological buffer solution. Middle: Arterial vasoconstriction (VC) after application of buffer solution with high potassium concentration (K+). Right: Arterial vasodilation (VD) induced by sodium nitroprusside (SNP). The diameter of the saphenous vein was unaffected. Below: Time course of inner diameter changes of A. saphena with fitted sigmoid function (black line). d0: initial diameter, dVC: minimal diameter during VC. dVD: maximal diameter during VD. t 1/2 : time of half VC or VD

Article Snippet: To assess the arterial diameter at baseline the exposed A. saphena was moistened with a physiological buffer solution (NaCl: 119 mmol/l, Merck, Darmstadt, Germany; KCl: 4.7 mmol/l, Merck; MgSO 4 : 1.17 mmol/l, Sigma-Aldrich, Taufkirchen, Germany; NaHCO 3 : 25 mmol/l, Merck; KH 2 PO 4 : 1.18 mmol/l, Merck; Glucose: 5.5 mmol/l, Merck; EDTA: 0.027 mmol/l, Prolabo, VWR International, Darmstadt) right before starting OCT. Acquisition of the baseline diameter stopped automatically after 30 initial B-scans (equivalent to a recording time of 7.5 s).

Techniques: Irradiation, Imaging, Concentration Assay

Journal: bioRxiv

Article Title: Comprehensive profiling of transcription factors for reprogramming human astrocytes to neuronal cells through endogenous CRISPR-based gene activation

doi: 10.1101/2025.10.11.681828

Figure Lengend Snippet: A. ORF overexpression of annotated INSM1, LHX6, and ZNF276 isoforms. RNA levels of neuron marker genes shown. *p < 0.05, ***p < 0.001 by global one-way ANOVA with Dunnett’s post hoc test comparing all groups to mCherry. B. Immunofluorescence staining of reprogrammed cells to assess MAP2, NeuN, and SLC17A7 expression 25 days after transduction. C. Number of spikes (neuronal firing events) in 5-minute multi-electrode array recording 32 days post-transduction. D. Flow analysis of TUBB3-2A-mCherry expression expression as proxy for early neuronal differentiation 5 days post-transduction of VP64 dSpCas9 VP iPSCs with gRNA. E. Summary of mouse gRNA sublibrary. F. Significance (P adj ) versus fold change in gRNA abundance between MAP2-high and MAP2-low populations in mouse CRISPRa screen. G. Top enriched biological processes for upregulated DEGs (L2FC >1, p adj <0.01 determined by DESeq2 vs mCherry, n=121 genes) from RNA-seq 10 days after INSM1 ORF overexpression. Statistical significance of term enrichment was determined using a two-tailed Fisher’s exact test followed by Benjamini–Hochberg correction.

Article Snippet: SLC17A7 screen: To screen for factors that cooperate with INSM1 to enhance glutamatergic subtype specification, cells were sorted based on abundance of SLC17A7 RNA using HCR-FlowFISH according to the method described in Reilly et al. 2021 with the following modifications: SLC17A7 probeset and buffers were ordered from Molecular Instruments ( https://www.molecularinstruments.com ), and SLC17A7 probeset was used at a final concentration of 8nM overnight.

Techniques: Over Expression, Marker, Immunofluorescence, Staining, Expressing, Transduction, RNA Sequencing, Two Tailed Test

Journal: bioRxiv

Article Title: Comprehensive profiling of transcription factors for reprogramming human astrocytes to neuronal cells through endogenous CRISPR-based gene activation

doi: 10.1101/2025.10.11.681828

Figure Lengend Snippet: A. Schematic of paired CRISPRa screens. B. Summary of TFpaired screening library. C. Scatter plot of z-score of gRNA abundance in the INSM1 MAP2 paired screen and the INSM1 SLC17A7 paired screen. D. Euler diagrams of differentially accessible peaks. Differential peaks (p adj <.01) for each sample were determined by DESeq2 vs. non-targeting gRNA. E. Top enriched biological processes for genes nearest top 1000 differentially accessible peaks by z-score for INSM1 (I), IKZF1 (IK), or peaks unique in only the combination of INSM1-IKZF1 (I+IK). Statistical significance of term enrichment was determined using a two-tailed Fisher’s exact test followed by Benjamini–Hochberg correction. F. Browser tracks of ATAC-seq (reads per kilobase per million mapped reads [RPKM]-normalized BigWig, bin size = 25bp. ‘Diff.peaks’ denotes peak significance between IKZF1-INSM1 and non-targeting using DESeq2. G. ANKS1B and KALRN peak accessibility and RNA expression indicate lack of significance after reprogramming with individual factors but significance after reprogramming with combination. H. Expression level ofANKS1B and KALRN in neural cell types in the Human Protein Atlas Single Cell Type data .

Article Snippet: SLC17A7 screen: To screen for factors that cooperate with INSM1 to enhance glutamatergic subtype specification, cells were sorted based on abundance of SLC17A7 RNA using HCR-FlowFISH according to the method described in Reilly et al. 2021 with the following modifications: SLC17A7 probeset and buffers were ordered from Molecular Instruments ( https://www.molecularinstruments.com ), and SLC17A7 probeset was used at a final concentration of 8nM overnight.

Techniques: Two Tailed Test, RNA Expression, Expressing