Journal: Nature communications

Article Title: Lysophosphatidylserine induces necrosis in pressure overloaded male mouse hearts via G protein coupled receptor 34.

doi: 10.1038/s41467-023-40201-4

Figure Lengend Snippet: Fig. 1 | Cardiac phenotypes of cardiac-specific iPLA2β-deficient mice after pressure overload. The Pla2g6+/+ and Pla2g6–/– mice were subjected to transverse aortic constriction (TAC) and then analyzed 5 days after the operation. a Survival ratio after TAC. n = 17 (sham-operated Pla2g6+/+), 13 (sham-operated Pla2g6–/–), 51 (TAC-operated Pla2g6+/+), and 51 (TAC-operated Pla2g6–/–). The log-rank test was used for survival analysis. Sham Pla2g6+/+ versus TAC Pla2g6+/+ P = 0.0000001, TAC Pla2g6+/+ versus TAC Pla2g6–/– P = 0.0002. b Representative images of transthoracic M-mode echocardiographic tracing (scale bars, 0.2 s and 5 mm, respectively) and the echocardiographic parameters of the mice. n = 7 (sham-operated Pla2g6+/+), 7 (sham-operated Pla2g6–/–), 14 (TAC-operated Pla2g6+/+) and 10 (TAC-operated Pla2g6–/–). LVIDd and LVIDs, end-diastolic and end-systolic left ventricular (LV) internal dimensions; FS, fractional shortening of LV. c Physiological parameters of the mice. n = 7 (sham-operated Pla2g6+/+), 7 (sham-operated Pla2g6–/–), 14 (TAC- operated Pla2g6+/+), and 10 (TAC-operated Pla2g6–/–), biologically independent samples. d Representative images of the hematoxylin—eosin-stained heart sections. Experiment was repeated five times independently with similar results. Scale bar,

Article Snippet: The following antibodies were used in this study: a monoclonal mouse antibody to iPLA2β (sc376563, Santa Cruz Biotechnology, Product Clone Name; D-4, 1:100 dilution), a monoclonal mouse antibody to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (016-25523, FUJIFILM Wako Pure Chemical Corporation, Product Clone Name; 5A12, 1:1000 dilution), a monoclonal mouse antibody to iPLA2ζ (MAB3210, Abnova, Product Clone Name; AT2G2, 1:1000dilution), a polyclonal goat antibody to iPLA2ε (EB08402, Everest Biotech, 1:1000 dilution), a polyclonal rabbit antibody to iPLA2η (25469-1-AP, Protein Tech, 1:500 dilution), a polyclonal rabbit antibody to iPLA2δ (NBP1-74214, Nobus Bio, 1:500 dilution), a polyclonal rabbit antibody to iPLA2γ (PA5-49993, Thermo Fisher Scientific, 1:1000 dilution), a polyclonal rabbit antibody to GPR132 (17026-1-AP, Protein Tech, 1:1000 dilution), a polyclonal rabbit antibody to GPR34 (PA5-45717, Thermo Fisher Scientific, 1:1000 dilution), a monoclonal rabbit antibody to RIP1 (#3493, Cell Signaling Technology, Product Clone Name; D94C12, 1:1000 dilution), a polyclonal rabbit antibody to phospho-RIP1 (#31122, Cell Signaling Technology, 1:1000 dilution), a polyclonal rabbit antibody to RIP3 (NBP1-77299, Nobus Bio, 1:1000 dilution), and a monoclonal rabbit antibody to phospho-RIP3 (ab195117, Abcam, Product Clone Name; EPR9516(N)−25, 1:1000 dilution).

Techniques: Staining

Journal: Journal of Neuroinflammation

Article Title: iPLA2β loss leads to age-related cognitive decline and neuroinflammation by disrupting neuronal mitophagy

doi: 10.1186/s12974-024-03219-z

Figure Lengend Snippet: Overexpression of iPLA2β in PFC improves cognitive function of old mice. A Scheme of the experimental design. AAV injections were administered 21 days prior to formal behavioral testing. Behavioral test training sessions were conducted two days before the formal testing began. Samples were collected on the seventh day following the initiation of the formal behavioral experiment. B Protein levels of iPLA2β in the 22 M PFC with iPLA2β overexpression (iPLA2β-OE) and control (CON) groups, as assessed by Western blotting and densitometry. C Representative immunofluorescence images of iPLA2β (red) and NeuN (green) in the 22 M PFC with iPLA2β overexpression (iPLA2β-OE) and control (CON) groups. Scale bar: 20 μm. D Representative SA-β-gal staining images in the 22 M PFC with iPLA2β overexpression (iPLA2β-OE) and control (CON) groups. Scale bar: 100 μm. E mRNA expression levels of TGFβ, IL-1β, TNF-α, and CCL2 in the 22 M PFC with iPLA2β overexpression (iPLA2β-OE) and control (CON) groups assessed via qPCR. Normalization was conducted relative to GAPDH expression levels. n = 11. F Representative IHC images of Iba1 in the 22 M PFC with iPLA2β overexpression (iPLA2β-OE) and control (CON) groups. Histogram showing quantification of the Iba1(+) area. Scale bar: 50 μm. G Travel time of the mice to reach the platform during the spatial test. n = 20 per group. H Duration spent by mice in the hidden platform quadrants during the probe trial. n = 20. I Frequency of platform crossings by mice during the probe trial. n = 20 per group. J Recognition index during the Novel object recognition test. Recognition index = time spent exploring a novel object/time spent exploring both objects. n = 20. K Discrimination index during the Novel object recognition test. The discrimination index = (time spent on the novel object − time spent on the familiar object)/ time spent on both objects. n = 20 Data are presented as the mean ± SEM; p values were obtained using the Mann-Whitney U test (B, C, D, and F) and the Kruskal-Wallis test followed by Dunn’s multiple comparisons test (E, G, H, I, J, and K). * p < 0.05. **, p < 0.01; ***, p < 0.001

Article Snippet: Then, brain sections were incubated overnight at 4 °C with the following antibodies: mouse polyclonal iPLA2β (sc-376563, 1:100, Santa Cruz), mouse monoclonal Iba1 (#17198, 1:400, Cell Signaling), or rabbit polyclonal GFAP (ab7260, 1:500, Abcam).

Techniques: Over Expression, Control, Western Blot, Immunofluorescence, Staining, Expressing, MANN-WHITNEY

Journal: Biochimica et biophysica acta. Molecular basis of disease

Article Title: Myeloid- and hepatocyte-specific deletion of group VIA calcium-independent phospholipase A2 leads to dichotomous opposing phenotypes during MCD diet-induced NASH.

doi: 10.1016/j.bbadis.2022.166590

Figure Lengend Snippet: Fig. 6. LPla2g6−/−mice show attenuated hepatic steatosis with altered expression of lipogenic and de novo lipogenesis genes after MCDD feeding. Mice were under the same treatment as in Fig. 5. (A) Representative images of H&E-stained livers (×100 magnification, scale bar = 200 μm). (B) The levels (μg/mg liver) of hepatic triglycerides, NEFA, and cholesterol. (C) RT-qPCR analysis of hepatic PPARs (Pparg, Ppargc1a, and Ppara), fatty-acid uptake and activation genes [Cd36, Cav1 (caveolin1), Fabp4, Slc27a2, and Slc7a5], TG synthesis/lipid droplets genes [Gpat, Agpat2, Dgat2, and Plin2 (perilipin2)], de novo lipogenesis (Srebf1, Fasn, Scd1, Elvol5, and Elovl6), and carbohydrate metabolism (Cebpa, Pck1, G6pc, and Mlxipl). (D) Western blot analysis of hepatic PPARγ (top) and PPARα (bottom), and quantification. (E) Western blot analysis of hepatic ACC, FASN, ELOVL6, and SCD1 expression and quantification of control and LPla2g6−/−mice under MCDD feeding. Data are mean ± SEM, N = 4–9 (B and C) and N = 3–6 (D and E). ***, p < 0.001, **, p < 0.01, and *, p < 0.05 with Mann-Whitney U tests.

Article Snippet: Membranes were incubated with a primary antibody against iPLA2β (D-4, sc-376,563 or T-14, sc-14,463, Santa Cruz Biotechnology, Heidelberg, Germany), CD36 (H-300, sc9154, Santa Cruz), PPARα (H-2, sc-398,394, Santa Cruz), CHOP (B-3, sc-7351, Santa Cruz), ELOVL6 (ab69857, Abcam), SCD1 (ab19862, Abcam), αSMA (ab32575, Abcam), VIMENTIN (ab9254, Abcam), SERPINE1 (#3917-1, Epitomics), ACC (#3662, Cell Signaling), PPARγ (#2435, Cell Signaling), CEBPα (clone EP709Y, cat#1704-1, Epitomics), p-JNK (81E11, #4668, Cell Signaling), FASN (C20G5, #3180, Cell Signaling), βACTIN (AC-5, #5441, Sigma), and GAPDH (#2118, Cell Signaling).

Techniques: Expressing, Staining, Quantitative RT-PCR, Activation Assay, Western Blot, Control, MANN-WHITNEY