Journal: Science Advances

Article Title: Altered X-chromosome inactivation predisposes to autoimmunity

doi: 10.1126/sciadv.adn6537

Figure Lengend Snippet: ( A ) Representative flow cytometry analysis of spontaneously activated B220 + CD69 + B cells in spleen from 1-year-old WT and Ftx −/− females. Percentages in leucocytes are shown on the graphs beneath. Each triangle represents a mouse. Median values are shown (Mann-Whitney test, * P < 0.05 and ** P < 0.01). ( B ) Same as (A) for spontaneously activated CD4 + CD69 + T cells (Mann-Whitney test, * P < 0.05 and ** P < 0.01). ( C ) Total IgG, IgM, IgG2b, and IgG2c natural antibody levels in sera of 3-month-, 1-year, and >1.5-year-old WT or Ftx −/− females measured by ELISA. Each circle represents a mouse. Mean values are shown (Mann-Whitney test, * P < 0.05 and ** P < 0.01). ( D ) Cytokines levels in the blood analyzed with cytometric bead array assays on sera from 3-month-, 1-year-, or 2-year-old WT and Ftx −/− females. Each triangle represents a mouse. Median values are shown ( t test, * P < 0.05).

Article Snippet: BM, spleen, blood, and peritoneal cavity cells were stained using the following antibodies: CD3 PerCP-Vio770 (130-119-656, Miltenyi Biotec), CD4-allophycocyanin (APC) (130-123-207, Miltenyi Biotec), CD5-APC-Vio770 (130-120-165, Miltenyi Biotec), CD8-fluorescein isothiocyanate (FITC) (130-118-468, Miltenyi Biotec), CD11b APC (553312, BD Pharmingen), CD11c phycoerythrin (PE)–Vio770 (130-110-840, Miltenyi Biotec), CD19-FITC (557398, BD Pharmingen), CD21-APC-Vio770 (130-111-733, Miltenyi Biotec), CD23-PE-Vio770 (130-118-764, Miltenyi Biotec), CD38-PE (130-123-571, Miltenyi Biotec), CD43-PE (130-112-887, Miltenyi Biotec), CD69-PE (130-115-575, Miltenyi Biotec), CD138 PE-Vio615 (130-108-989, Miltenyi Biotec), F4/80 FITC (130-117-509, Miltenyi Biotec), Ter119 PE (130-112-909, Miltenyi Biotec), SiglecH APC-Vio770 (130-112-299, Miltenyi Biotec), B220-APC (130-110-847, Miltenyi Biotec), B220 VioBlue (130-110-851, Miltenyi Biotec), IgM-VioBlue (130-116-318, Miltenyi Biotec), IgD-PE (130-111-496, Miltenyi Biotec), GL7-PE-Cy7 (144619, BioLegend), Ly6C-FITC (130111-915, Miltenyi Biotec), streptavidin FITC (554060, BD Biosciences), CD138 BV605 (563147, BD-Horizon), CD23 BV605 (101637, BioLegend), I-A/I-E BV711 (107643, BioLegend), CD19 BV786 (563333, BD Horizon), T and B cell activation antigen (GL7) PE (561530, BD Pharmingen), CD95 PE-Cy7 (557653, BD Pharmingen), IgM APC-eFluor 780 (47-5790-82, Invitrogen), CD45R/B220 APC/cyanine7 (103224, BioLegend), CD11b eFluor 450 (48-0112-82, eBioscience), CD267 (TACI) BV421 (742840, BD Biosciences), IgD BUV395 (564274, BD Horizon), streptavidin APC (4317-82, eBioscience), CD3 Biotin (100304, BioLegend), Biotin CD11c (568970, BD Biosciences), CD21 PercP Cy5.5 (562797, BD Biosciences), and Fixable Viability Dye eFluor 506 (65-0866-18, Invitrogen) following the recommendations of the manufacturers.

Techniques: Flow Cytometry, MANN-WHITNEY, Enzyme-linked Immunosorbent Assay

Journal: Cell reports

Article Title: Decreased KAT5 Expression Impairs DNA Repair and Induces Altered DNA Methylation in Kidney Podocytes.

doi: 10.1016/j.celrep.2019.01.005

Figure Lengend Snippet: Figure 7. KAT5 Increases Nephrin Expression through Reductions in DNA DSBs and DNA Methylation at the Promoter Region in Cultured Human Podocytes (A) (Left) Real-time RT-PCR analysis and (right) western blots of nephrin expression in cultured human podocytes with normal (6 mM) or high (30 mM) glucose treatment. (B) Methylation-specific PCR (MSP) analysis of the nephrin promoter region in cultured human podocytes with normal (6 mM) or high (30 mM) glucose treatment. (C) (Left) Real-time RT-PCR analysis and (right) western blots of nephrin expression in KAT5-overexpressed cultured human podocytes. (D) Methylation-specific PCR (MSP) analysis of the nephrin promoter region in KAT5 overexpressed cultured human podocytes. (E and F) Chromatin immunoprecipitation (ChIP) assay for DNMT1 (E) and DNMT3B (F) in the promoter region of nephrin in KAT5-overexpressed cultured human podocytes. (G) gH2AX foci was decreased in the nephrin promoter region by ChIP assay. Control, podocytes transfected with control vector. KAT5, KAT5-overexpressing podocytes. Experiments were performed in podocytes cultured with 30 mM of D-glucose (C–G). n = 6 in each group. Data represent mean ± SEM. *p < 0.05 and **p < 0.01 versus controls by two-tailed Student’s t test.

Article Snippet: pCMV-SPORT6-KAT5 and pCMV-SPORT6 were purchased from the RIKEN Center for Life Science Technologies (Hyogo, Japan), DNMT1 cDNA clone (SC325419) was purchased from Origene and transient transfection was performed using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s instructions.

Techniques: Expressing, DNA Methylation Assay, Cell Culture, Quantitative RT-PCR, Western Blot, Methylation, Chromatin Immunoprecipitation, Control, Transfection, Plasmid Preparation, Two Tailed Test

Journal: Cell Death & Disease

Article Title: PLK1 and AURKB phosphorylate survivin differentially to affect proliferation in racially distinct triple-negative breast cancer

doi: 10.1038/s41419-022-05539-5

Figure Lengend Snippet: List of reagents and antibodies.

Article Snippet: 45 , Aurora B (AURKB) (NM_004217) Human Tagged ORF Clone , , RC210288 , OriGene.

Techniques: Concentration Assay, cDNA Synthesis, SYBR Green Assay, Protease Inhibitor, Saline, Plasmid Preparation, Magnetic Beads, Membrane

Journal: Cell Death & Disease

Article Title: PLK1 and AURKB phosphorylate survivin differentially to affect proliferation in racially distinct triple-negative breast cancer

doi: 10.1038/s41419-022-05539-5

Figure Lengend Snippet: Bar graphs showing mitosis scores in the Emory ( A ) and Dekalb ( B ) cohorts. C Heatmap showing the expression levels of various kinases in the TCGA BC dataset. D , E Bar graphs showing the expression levels of PLK1 ( D ) and AURKB ( E ) in AA ( n = 3) and EA ( n = 3) TNBC cell lines. F – H Representative IHC images of PLK1 and AURKB ( F ) and quantification bar graphs showing PLK1 ( G ) and AURKB ( H ) levels in grade- and stage-matched AA and EA patients with TNBC (Dekalb cohort). I Immunoblot showing PLK1 and AURKB protein levels in AA and EA TNBC cell lines ( n = 3 each). FPKM fragments per kilobase of transcript per million mapped reads. Bars indicate mean ± SEM. Unpaired two-tailed Student’s t -test with Welch’s correction was used to determine statistical significance (* P < 0.05, *** P < 0.0005, ns = non-significant). The scale bar represents 100 µm.

Article Snippet: 45 , Aurora B (AURKB) (NM_004217) Human Tagged ORF Clone , , RC210288 , OriGene.

Techniques: Expressing, Western Blot, Two Tailed Test

Journal: Cell Death & Disease

Article Title: PLK1 and AURKB phosphorylate survivin differentially to affect proliferation in racially distinct triple-negative breast cancer

doi: 10.1038/s41419-022-05539-5

Figure Lengend Snippet: Expression of various kinases in AA and EA patients with TNBC (TCGA dataset).

Article Snippet: 45 , Aurora B (AURKB) (NM_004217) Human Tagged ORF Clone , , RC210288 , OriGene.

Techniques: Expressing

Journal: Cell Death & Disease

Article Title: PLK1 and AURKB phosphorylate survivin differentially to affect proliferation in racially distinct triple-negative breast cancer

doi: 10.1038/s41419-022-05539-5

Figure Lengend Snippet: A , B Immunoblots showing the levels of PLK1, survivin, p-survivin (S20), and β-actin after PLK1 silencing ( A , B ) or inhibition ( C ) in AA and EA TNBC cell lines. D – F Immunoblots showing the levels of AURKB, survivin, p-survivin (T117), and β-actin after AURKB silencing ( D , E ), or inhibition ( F ) in AA and EA TNBC cell lines.

Article Snippet: 45 , Aurora B (AURKB) (NM_004217) Human Tagged ORF Clone , , RC210288 , OriGene.

Techniques: Western Blot, Inhibition

Journal: Cell Death & Disease

Article Title: PLK1 and AURKB phosphorylate survivin differentially to affect proliferation in racially distinct triple-negative breast cancer

doi: 10.1038/s41419-022-05539-5

Figure Lengend Snippet: A – F Representative IHC images ( A , B ) and bar graphs ( C – F ) showing Ki-67 and survivin levels in AA ( C , E ) and EA ( D , F ) TNBC xenografts under various treatment conditions. G , H Immunoblots showing the levels of p-survivin (T117), p-survivin (S20), total survivin, AURKB, PLK1, and β-actin in AA ( G ) and EA ( H ) fresh-frozen xenograft tumor lysates from mice treated with volasertib, barasertib, or their combination ( n = 12 per treatment group). Bars represent mean ± SEM. Unpaired two-tailed Student’s t -test with Welch’s correction was used to determine statistical significance (**** P < 0.00005, ns = non-significant). The scale bar represents 100 µm.

Article Snippet: 45 , Aurora B (AURKB) (NM_004217) Human Tagged ORF Clone , , RC210288 , OriGene.

Techniques: Western Blot, Two Tailed Test