Journal: bioRxiv

Article Title: Oncogenic Ppm1d mutations deregulate the p53 pathway in primary mouse gliomas

doi: 10.1101/2025.11.25.690483

Figure Lengend Snippet: a) Intracranial injection of DF1 producing cells carrying RCAS vectors encoding PDGFB, Cre, and luciferase to enable conditional targeting of neural stem cells. b) Mice were injected at postnatal day 3-5 and monitored for tumor development through symptom assessment and bioluminescence imaging starting at week 4 to detect tumor-associated signals. Kaplan-Meier survival estimates and and log-rank (Mantel-Cox) test p= 0.0277, Ppm1d flex- /wt ; p53 fl/wt n=21, Ppm1d wt/wt ; p53 fl/wt n=19 c) Immunohistochemistry (IHC) with Ki67, Olig2, and GFAP for tumor-bearing brains from wild-type Ppm1d (Ppm1d wt/wt ; p53 fl/wt ) and truncated Ppm1d (Ppm1d flex- /wt ; p53 fl/wt ) mice. Tumor and normal regions were denoted in GFAP staining; GFAP exhibited cytoplasmic localization. Scale bar: 100 μm. d) Quantification of Ki67 and Olig2 staining. GFAP expression was excluded from quantification due to its cytoplasmic distribution. Data are presented as mean ± SEM; n = 3. Student paired t-test, * p< 0.05 and ** p< 0.01 e) IHC staining for Myc-tag was performed on brain sections from wild-type Ppm1d (Ppm1d wt/wt ; p53 fl/wt ) and truncated Ppm1d (Ppm1d flex- /wt ; p53 fl/wt ) mice. Whole-mount sections (scale bar 2 mm) and tumor-region sections (scale bar 100 μm). Red boxes represent tumor regions. f) Quantification of Myc Tag IHC staining. Data are shown as mean ± SEM; n = 3. Student unpaired t-test, ** p< 0.01

Article Snippet: IHC staining for Ki67 (Cat #Bethyl IHC-00375), Olig2 (Cat #ab109186), GFAP (Cat #NB300-141) and MYC (Cat #ab32072) was conducted by HistoWiz.

Techniques: Injection, Luciferase, Imaging, Immunohistochemistry, Staining, Expressing

Journal: Molecular Therapy. Methods & Clinical Development

Article Title: AAV8-based gene replacement therapy for hereditary spastic paraplegia type 5

doi: 10.1016/j.omtm.2025.101531

Figure Lengend Snippet: In vitro validation of generated constructs (A) Overview of in vitro experiments. (B) Gene expression of CYP7B1 in HepG2 cells after transfection with plasmids for GFP (control), hCYP7B1, and CYP7B1-FLAG assessed by qPCR. CYP7B1 expression after transfection with the respective plasmids is upregulated 10,000-fold compared to untreated and GFP controls. (C) Western blot analysis confirms strong expression of CYP7B1 (VCL, vinculin, loading control). (D) Immunofluorescence of HepG2 cells fixed 24 h after transfection with the CYP7B1-FLAG construct, stained against the FLAG tag, the ER marker CLIMP63, and DNA (scale bars, 20 μm).

Article Snippet: Immunohistochemical staining for CYP7B1 (Proteintech, 24889-1-AP) was performed on 4 μm deparaffinized sections.

Techniques: In Vitro, Biomarker Discovery, Generated, Construct, Gene Expression, Transfection, Control, Expressing, Western Blot, Immunofluorescence, Staining, FLAG-tag, Marker

Journal: Molecular Therapy. Methods & Clinical Development

Article Title: AAV8-based gene replacement therapy for hereditary spastic paraplegia type 5

doi: 10.1016/j.omtm.2025.101531

Figure Lengend Snippet: Liver transduction efficacy and liver oxysterol levels (A) Immunohistochemistry of liver sections against CYP7B1 from wild-type (WT) animals, Cyp7B1 −/− animals injected with different doses of AAV8-TTR-hCYP7B1, AAV8-TTR-eGFP (vehicle), and untreated KO animals (KO). (B) mRNA levels of human CYP7B1 and endogenous murine Cyp7b1 for the six experimental groups normalized to Eif2α and Actβ. Strong expression of human CYP7B1 is observed with all doses of AAV-TTR-hCYP7B1. (C) Western blot of liver homogenates (VCL, vinculin, loading control). CYP7B1 expression in all groups receiving AAV-TTR-CYP7B1 injections exceeded WT expression and highest doses led to oversaturation of the membrane, while untreated and vehicle-injected KO animals show no signal for CYP7B1. (D) Liver oxysterol levels at the endpoint of the study (D49). Both 25-HC and 27-HC were significantly reduced compared to the GFP vehicle-treated group. Levels of the cerebral metabolite 24-HC decreased slightly but remained within the range of WT animals. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; and ∗∗∗∗ p < 0.0001.

Article Snippet: Immunohistochemical staining for CYP7B1 (Proteintech, 24889-1-AP) was performed on 4 μm deparaffinized sections.

Techniques: Transduction, Immunohistochemistry, Injection, Expressing, Western Blot, Control, Membrane