Journal: Cell reports

Article Title: The C9ORF72 repeat expansion alters neurodevelopment

doi: 10.1016/j.celrep.2023.112983

Figure Lengend Snippet: (A) Representative images of brains of 2-month-old mice exposed to vehicle or 2 μg/g VIP antagonist from E9.5–E11.5. (B) Quantification of brain mass in 2-month-old mice that were treated with vehicle or VIP antagonist from E9.5–E11.5. Each data point represents the brain mass of one mouse (n = 9 vehicle and n = 8 VIP antagonist). Mean ± SEM. Unpaired t test. (C and D) Representative images (C) and quantification (D) of cortical thickness in E18.5 control and C9ORF72 -BAC embryos treated with vehicle or 2 μg/g VIP antagonist using immunohistochemistry. Each data point represents the average cortical thickness in one embryo (n = 3 control and 3 C9ORF72 -BAC embryos). The average cortical thickness was determined by measuring the width from layer 1 to layer 6 across 3 different 16-mm sections beginning at approximately bregma + 3.51. Mean ± SEM. One-way ANOVA. Scale bars, 1 mm. (E) Quantification of time to fall in a hanging wire test for 2-month-old control and C9ORF72 -BAC mice treated with vehicle or VIP antagonist from E9.5–E11.5. Each data point represents the average time to fall for one mouse. Control + vehicle, n = 11 mice; control + VIP antagonist, n = 10 mice; C9ORF72 -BAC + vehicle, n = 10 mice; C9ORF72 -BAC + VIP antagonist, n = 16 mice. Mean ± SEM. Kruskal-Wallis test. (F) Quantification of total distance traveled during an open field test for 2-month-old control and C9ORF72 -BAC mice treated with vehicle or VIP antagonist from E9.5–E11.5. Each data point represents the total distance traveled for one mouse. Control + vehicle, n = 11 mice; control + VIP antagonist, n = 10 mice; C9ORF72 -BAC + vehicle, n = 10 mice; C9ORF72 -BAC + VIP antagonist, n = 16 mice. Mean ± SEM. two-way ANOVA.

Article Snippet: Pregnant dams were administered intraperitoneal (IP) injections of 2 μg/g body weight of the VIP receptor antagonist compound (Tocris, cat. no. 3054) once a day at E9.5, E10.5, and E11.5.

Techniques: Control, Immunohistochemistry

Journal: British Journal of Pharmacology

Article Title: Vasoactive intestinal peptide shapes first‐trimester placenta trophoblast, vascular, and immune cell cooperation

doi: 10.1111/bph.14609

Figure Lengend Snippet: Down‐regulation of ECM remodelling gene expression in VIP‐deficient trophoblast cell lines. BeWo cell line at 60% of confluence was transfected with a VIP‐siRNA (siVIP) or with a Scrl sequence, and after 72 hr, the cells were harvested and used for RT‐qPCR. (a) Basal relative expression of VIP/VPACs is shown in BeWo cell line. Two highly positive controls (CT+) were used: the human epithelial colorectal adenocarcinoma cell line, CACO‐2 for VIP expression, and the human neuroblastoma cell line, SH‐SY5Y for VPAC receptors. Results are expressed as mean ± SEM of arbitrary units (A.U.) gene/housekeeping gene expression (n = 5). (b) The VIP expression in BeWo cell line transfected with VIP siRNA (siVIP) or scramble sequence (Scrl) were studied by qPCR. Results are expressed as mean ± SEM of fold change with respect to siVIP. P < 0.05 of seven independent experiments. (c) In the left panel, a double gradient heat map is shown: green for down‐regulated, black for no‐changes, and red for up‐regulated genes. In the right panel, the results are expressed as mean ± SEM of gene fold change (n = 5). In both, panels (b) and (c) the confidence interval was used to determine if the measurement in siVIP was statistically significant compared to Scrl. ECM, extracellular matrix; MMP, metalloproteinase; TIMP, tissue inhibitor of MMP; VIP, vasoactive intestinal peptide

Article Snippet: VIP silencing in trophoblast cell lines To transfect BeWo cells with a VIP siRNA (Santa Cruz Biotechnology, Dallas, TX, USA), cells were grown at 60% of confluence and arrested for 3 hr in Optimem®; 100 nM VIP siRNA: Lipofectamine RNAimax (Life Technologies, Grand Island, NY, USA) complex were made in Optimem and incubated for 15 min before being added to the cells in a drop wise manner; 24‐hr post‐transfection, the media were changed for DMEM:F12 supplemented with 10% FCS for another 48 hr. siRNA with a scramble (Scrl) sequence was used as a negative control.

Techniques: Gene Expression, Transfection, Sequencing, Quantitative RT-PCR, Expressing