Journal: Biochimica et biophysica acta. Molecular basis of disease

Article Title: Npc1 deficiency impairs microglia function via TREM2-mTOR signaling in Niemann-Pick disease type C.

doi: 10.1016/j.bbadis.2024.167478

Figure Lengend Snippet: Fig. 1. Loss of Npc1 impairs brain development in mouse. A Representative photograph of brain from Npc1−/−mice and littermate control at postnatal day 63 (P63). B Brain weight curves of Npc1−/−mice and littermate controls (n = 9; Page x genotype < 0.0001, Page < 0.0001, Pgenotype < 0.0001). C Representative coronal sections of Npc1−/−mice and littermate controls at P42 visualized by hematoxylin and eosin staining. D Quantification of the cortical thickness of mice from P1 to P63 (n = 3). E, F Representative confocal images and quantification of Filipin (white) and DAPI (blue) staining in the cortex of Npc1+/+ and Npc1−/−mice at P42 (n = 3). G, H Representative confocal images and quantification of GFAP (red) and DAPI (blue) staining in the cortex of Npc1+/+ and Npc1−/−mice at P42 (n = 3). I, J Repre sentative confocal images and quantification of MBP (green) and DAPI (blue) staining in the cortex of Npc1+/+ and Npc1−/−mice at P42 (n = 3). K, L Representative confocal images and quantification of cleaved-Caspase3 (c.CASP3, green) staining in the cortex of Npc1+/+ and Npc1−/−mice at P42 (n = 3). M Western blot analysis for c.CASP3, BAX and BCL2 protein expression in the brain of Npc1−/−mice and littermate controls (n = 3). Results are presented as Mean ± SEM. n.s., not sig nificant. *P < 0.05, **P < 0.01 and ***P < 0.001. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: The primary and secondary antibodies were used as follows: cleaved-Caspase3 (Cell Signaling Technology, USA; Cat No. 9664), BAX (Cell Signaling Technology, USA; Cat No. 14796), BCL2 (BOSTER, China; Cat No. BA0412), APOE (Abcam, UK; Cat No. ab183597), CTSD (Abcam, UK; Cat No. ab75852), Npc1 (Abcam, UK; Cat No. ab134113), Npc2 (Abcam, UK; Cat No. ab218192), TREM2 (Cell Signaling Technology, USA; Cat No. 76765), TREM2 (R&D Systems, USA; Cat No. AF1729), TREM2 (Abcam, UK; Cat No. ab305103), SYK (Cell Signaling Technology, USA; Cat No.13198), p-SYK (Cell Signaling Technology, USA; Cat No.2710), AKT (Cell Signaling Technology, USA; Cat No.4691), p-AKT (Cell Signaling Technology, USA; Cat No.13038), GSK3β (Cell Signaling Technology, USA; Cat No. 12456), p-GSK3β (Cell Signaling Technology, USA; Cat No. 5558), mTOR (Cell Signaling Technology, USA; Cat No. 2983), p-mTOR (Cell Signaling Technology, USA; Cat No. 5536), S6K1 (Cell Signaling Technology, USA; Cat No. 34475), p-S6K1 (Cell Signaling Technology, USA; Cat No. 9234), 4EBP1 (Cell Signaling Technology, USA; Cat No. 9644), p-4EBP1 (Cell Signaling Technology, USA; Cat No. 9451), CD63 (Abcam, UK; Cat No. ab217345), GFAP (Cell Signaling Technology, USA; Cat No. 80788), β-Actin (Affinity Biosciences, USA; Cat No. AF7018), β-Tubulin (Absin, China; Cat No. abs830032), Goat anti-Rabbit IgG-HRP (Absin, China; Cat No. abs20040), Goat anti-Mouse IgG-HRP (Absin, China; Cat No. abs20039), Rabbit anti-Sheep IgG-HRP (Abcam, UK; Cat No. ab6747).

Techniques: Control, Staining, Western Blot, Expressing

Journal: BioMed Research International

Article Title: Overexpression of miR-150-5p Alleviates Apoptosis in Sepsis-Induced Myocardial Depression

doi: 10.1155/2020/3023186

Figure Lengend Snippet: Functional study of miR-150-5p in H9C2 cells. (a, b) Apoptosis was detected by Annexin V-FITC/PI staining (a) and TUNEL assay (b) after transfection. Green represents TUNEL positive cells and blue represents cell nucleus. (a, b) Caspase3, Bax, and Bcl-2 mRNA (c) and protein expression (d) were measured by qPCR and western blot after transfection. I: control, II: LPS, III: LPS+NC, IV: LPS+mimics. Data are presented as mean ± standard deviation, repeated for three times. ∗ P < 0.05 compared to the control or the LPS+NC groups.

Article Snippet: GAPDH, akt2, p-akt2, cleaved caspase3, bax, and bcl-2 primary antibodies were incubated with the membranes at 4°C overnight (Akt2; 1 : 200; Santa Cruz Biotechnology, Santa Cruz, CA, USA; p-Akt2; 1 : 500; Abcam, Cambridge, MA, USA; cleaved caspase3; 1 : 500; Cell Signaling Technology, Danvers, MA, USA; bax and bcl-2; 1 : 500; Proteintech; GAPDH; 1 : 10000; Abways).

Techniques: Functional Assay, Staining, TUNEL Assay, Transfection, Expressing, Western Blot, Standard Deviation

Journal: Cell Stress & Chaperones

Article Title: C. sakazakii activates AIM2 pathway accompanying with excessive ER stress response in mammalian mammary gland epithelium

doi: 10.1007/s12192-019-01065-0

Figure Lengend Snippet: C. sakazakii induced apoptosis in mouse model and in bovine mammary epithelial cells. The mammary gland tissues of each group (n ≥ 3) were obtained at 24 h after C. sakazakii and PBS infection. a, c Bcl-2 protein and Bax protein were detected by western blot in mice and in BMECs. b, d The ratio of Bax/Bcl-2 protein was analyzed in mice and BMECs. e The mRNA level of Bcl-2, Bax, and relative mRNA level of β-actin were determined by qRT-PCR in mice. f The ratio mRNA level of Bcl-2/Bax and relative mRNA level of β-actin were determined by qRT-PCR in mice. Data are presented as means ± the standard deviation (SD) (n ≥ 3) (*P<0.05, **P<0.01, ***P<0.001)

Article Snippet: The antibodies used in this study include the following: GAPDH (CST, Boston, MA), Tublin (CST, Boston, MA), Bax (Proteintech Co LTD, Wuhan, Hubei, China), Bcl-2 (Proteintech Co LTD, Wuhan, Hubei, China), AIM2 (CST, Boston, MA), and Cleaved IL-1β (CST, Boston, MA).

Techniques: Infection, Western Blot, Quantitative RT-PCR, Standard Deviation

Journal: American Journal of Translational Research

Article Title: Overexpression of TEM8 promotes ovarian cancer progression via Rac1/Cdc42/JNK and MEK/ERK/STAT3 signaling pathways

doi:

Figure Lengend Snippet: TEM8 promoted proliferation but reduced apoptosis in ovarian cancer cells. A, B. MTT assay showed that TEM8 overexpression increased cell proliferation of ovarian cancer cells, however, these trends were reversed in the TEM8 knockdown groups (n = 9). C-F. Cell cycle assay showed that TEM8 overexpression promoted G0/G1 phase transition of ovarian cancer cells, however, these trends were reversed in the TEM8 knockdown groups (n = 3). G-J. Cell apoptosis assay showed that TEM8 overexpression reduced apoptosis rate of ovarian cancer cells, however, these trends were reversed in the TEM8 knockdown groups (n = 3). K, L. Representative images and quantitation of the western blot showed that the protein expression of Ki-67, cyclin D1, Bcl2, and Bax in the TEM8 overexpression/knockdown groups (n = 3). GAPDH was used as an internal control. Data are presented as mean ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: Proteins were separated using 10% SDS-PAGE, transferred to PVDF membranes and blocked using 5% BSA at room temperature for 2 h. Primary antibodies were incubated at 4°C overnight (dilution ratio of 1:1000): rabbit anti-TEM8 (Abcam, Cambridge, UK), rabbit anti-GATA2 (Abcam, Cambridge, UK), rabbit anti-Rac1 (Abcam, Cambridge, UK), rabbit anti-Cdc42 (Wanleibio, Shenyang, China), rabbit anti-p-Rac1/cdc42 (Cell Signaling Technology, California, USA), rabbit anti-JNK (Cell Signaling Technology, California, USA), mouse anti-p-JNK (Cell Signaling Technology, California, USA), rabbit anti-STAT3 (Wanleibio, Shenyang, China), rabbit anti-MEK (Santa Cruz, USA), rabbit anti-p-MEK (Cell Signaling Technology, California, USA), rabbit anti-ERK (Cell Signaling Technology, California, USA), rabbit anti-p-ERK (Cell Signaling Technology, California, USA), rabbit anti-p-STAT3 (Ser727) (Wanleibio, Shenyang, China), anti-Ki-67 (Wanleibio, Shenyang, China), anti-cyclin D1 (Cell Signaling Technology, California, USA), rabbit anti-MMP2 (Proteintech, Wuhan, China), anti-rabbit MMP9 (Proteintech, Wuhan, China), mouse anti-Bcl2 (Proteintech, Wuhan, China), rabbit anti-Bax (Proteintech, Wuhan, China), rabbit anti-VEGFA (Abcam, Cambridge, UK), mouse anti-GAPDH (1:2000, ZSGB-BIO Technology Co., Ltd., Beijing, China).

Techniques: MTT Assay, Over Expression, Cell Cycle Assay, Sublimation, Apoptosis Assay, Quantitation Assay, Western Blot, Expressing