Journal: Nature

Article Title: Blocking PGE 2 -induced tumour repopulation abrogates bladder cancer chemoresistance

doi: 10.1038/nature14034

Figure Lengend Snippet: a, Kaplan–Meier analysis of bladder-cancer patients with various CK14 staining patterns (enrich/pers denotes an enrichment (increase) or persistence of CK14 staining; resolution denotes an absence) after neoadjuvant chemotherapy (n = 15). b, Relative change in xenograft tumour size after chemotherapy (n = 6 xenograft tumours per group). PDX-3, patient-derived xenograft from patient 3; T24, human bladder cancer cell line-derived xenograft. c, d, Representative immunofluorescence staining (c) and box plots quantifying the percentage of CK14+ bladder cancer cells after chemotherapy (d). DAPI, 4′,6-diamidino-2-phenylindole. e, Schematic demonstrating the transduction of hK14.tdTomato lentiviral reporter into urothelial carcinoma cells. f, Immunofluorescence staining verifying the specific expression of tdTomato (Tm)-positive signal in CK14+ cancer cells. g, Quantitative PCR (qPCR) analysis of KRT14 and UPK1B in cancer cell subpopulations in biological duplicates. h, i, Graphs summarizing viability (h) and corresponding percentage of S phase cells (i) in two cancer subpopulations after chemotherapy in biological duplicates. j, Cell cycle profiles for Tm+ CK14+ (red line) and Tm− CK14− (black line) cancer cells at indicated days after chemotherapy. PI, propidium iodide. Data in b represent mean ± s.e.m.; box plots in c show twenty-fifth to seventy-fifth percentiles, with line indicating the median, and whiskers indicating the smallest and largest values; data in g–i show mean and range. ***P <0.001 (log-rank test (a) and two-tailed Student’s t-test (b)). Scale bars, 100 μm.

Article Snippet: We developed a lentiviral construct that reports on the activity of human KRT14 promoter by sub-cloning a previously characterized and validated promoter region of human KRT14 (ref. 16 ) (obtained from ATCC, MBA-124) using EcoRI and BamHI restriction enzymatic digestion, and inserting this promoter fragment into a promoterless lentiviral vector (Clontech, 631754) tagged with a DD-tdTomato (Tm) red fluorescent protein (Clontech Lenti-X DD-tdTomato vector).

Techniques: Staining, Derivative Assay, Immunofluorescence, Transduction, Expressing, Real-time Polymerase Chain Reaction, Two Tailed Test

Journal: Molecular therapy. Nucleic acids

Article Title: Mechanical stimuli-induced CCL2 restores adult mouse cells to regenerate hair follicles.

doi: 10.1016/j.omtn.2023.03.002

Figure Lengend Snippet: Figure 5. CCL2-CCR4 axis regulates plucking-induced hair regeneration (A) F4/80 immunostaining and statistics show recruitment of M1 macrophage at PPD3 after plucking. (B) CD206 immunostaining and statistics show no obvious recruitment of M2 macrophage at PPD3 after plucking. (C) TNF-a immunostaining and statistics show skin inflammation at PPD3 after plucking. (D) K14 immunostaining and statistics show hair regenerate at PPD3 after plucking. (E) K14 immunostaining and statistics show hair regeneration is blocked after inhibition of CCL2 (iCCL2) or CCR4 (iCCR4) at PPD3. (F) F4/80 immunostaining and statistics show recruitment of M1 macrophage is disrupted after inhibition of CCL2 (iCCL2) or CCR4 (iCCR4) at PPD3. (G) CD206 immunostaining and statistics show no obvious recruitment of M2 macrophage at PPD3 in all three groups. (H) TNF-a immunostaining and statistics show significantly decreased skin inflammation after inhibition of iCCL2 or iCCR4 at PPD3. *p < 0.05, **p < 0.01, ***p < 0.001, #, no significance, n R 3.

Article Snippet: Primary antibodies include AE13 (ab16113, Abcam), AE15 (gift), CD34 (ab81289, Abcam), Sox9 (ab71762, Abcam), K10 (ab76318, Abcam), K14 (A01432, Boster), CD86 (ab53004, Abcam), F4-80 (sc-25830, Santa Cruz), CD163(sc-33560, Santa Cruz), CD206(sc58986, Santa Cruz), TNF-a (AF8208, Beyotime), NCAM (from Dr. Cheng-Ming Chuong laboratory), P16 (ab51243, Abcam), P63 (12143-1-AP, Proteintech), P-cadherin (13-2000Z, Thermo), and PCNA (ab92552, Abcam).

Techniques: Immunostaining, Inhibition

Journal: The Journal of Cell Biology

Article Title: Mice with targeted disruption of the fatty acid transport protein 4 (Fatp 4, Slc27a4) gene show features of lethal restrictive dermopathy

doi: 10.1083/jcb.200207080

Figure Lengend Snippet: Differentiation of neonatal skin of newborn Fatp4 −/− and control Fatp4 +/+ mice, as shown by immunofluorescence microscopy. (A) Immunofluorescence microscopy of keratin K14 in dorsal skin. In the Fatp4 +/+ control mice, K14 is restricted to the stratum basale (sb) and in the Fatp4 −/− mice is additionally detected in the first layers of the stratum spinosum (open arrow). (B) With keratin K10 antibodies, the stratum spinosum and the stratum granulosum (ss+sg) are strongly stained in both wild-type and Fatp4 −/− mice. However, note that both layers are thicker in the mutant. (C) Immunostaining of K6 is negative in the Fatp4 +/+, but intense in the upper suprabasal layers, including the stratum corneum (sc), of the Fatp4 −/− mice. (D) Loricrin as a typical protein of the cornified envelope is detected in the stratum granulosum (sg) and seems to be a bit more prominent in the mutant mice. (E) The presence of occludin in the plasma membranes of the stratum granulosum (bracket; sb, stratum basale) is seen in a somewhat broader appearing zone in the mutant mice, indicating an intact tight junction barrier in both kinds of mice. To the right of each of the immunofluorescence micrographs are the specific phase-contrast images of the same sections. Blue: DAPI nuclear staining. Bars, 50 μm.

Article Snippet: For comparison, antibodies used in immunohistochemistry were as follows: (1) rabbit antibodies to occludin, claudin 1, protein ZO-1 (all from Zytomed), to keratins K16 (gift of Dr. P. Coulombe, Johns Hopkins University School of Medicine, Baltimore, MD) and K10 (gift of Dr. D. Roop, Baylor College, Houston, TX), to periplakin and loricrin (both antisera were gifts of Dr. D. Hohl, Hopital de Beaumont, Lausanne, Switzerland); (2) guinea pig antibodies against keratins K2e, K5 and K14 (all from Progen Biotechnik); and (3) mouse antibodies to keratin K6 (clone KA12; Progen Biotechnik), filaggrin (mouse ascites; from Quartett Co., or from Paesel & Lorei) and to transglutaminase 1, TG-1 (Cell Systems).

Techniques: Control, Immunofluorescence, Microscopy, Staining, Mutagenesis, Immunostaining, Clinical Proteomics