Journal: The Journal of Cell Biology

Article Title: The Retinoblastoma family member p107 regulates the rate of progenitor commitment to a neuronal fate

doi: 10.1083/jcb.200703176

Figure Lengend Snippet: Increased numbers of progenitor cells in the embryonic and adult p107 −/− brains in vivo. (a–c) To assess the size of the neural progenitor population in the adult brain, mice received intraperitoneal injections of BrdU to label proliferating cells over a 10-h period. BrdU-positive cell counts through the lateral ventricles showed a 50% increase in the number of progenitors in p107 −/− brains ( n = 4) in comparison with wild type ( n = 3). (d) The adult progenitor population was also assessed by immunohistochemistry for PCNA. Similar to our BrdU incorporation study, p107-deficient brains had a comparable increase in PCNA + cells along the ventricles. (e) To quantify the number of proliferating progenitors in E10.5 brains, we used an antibody to phosphohistone H3 (PH3), a marker of cells in M phase of the cell cycle. PH3-positive cells were counted along the lateral ventricles in three representative sections of wild-type ( n = 3) and p107 −/− ( n = 3) brains. (f) A 2-h BrdU pulse was used to label progenitors in S phase in E13.5 brains. BrdU-positive cells were counted in four representative regions of the forebrain in wild-type ( n = 4) and p107 −/− ( n = 3) embryos. Note that at adult and embryonic ages, p107 −/− mice had significantly more proliferating cells along the lateral ventricle than wild-type mice. (g and h) To assess whether in vivo cell cycle kinetics were different in p107 −/− neural progenitors, we compared the size of the S-phase fraction by immunolabeling for BrdU and PCNA in adult (g) and E13.5 wild-type and p107 −/− cortices (h). Quantification of BrdU+ cells (S-phase fraction) was compared with the total proliferating population (PCNA + ). After 10.5 h of cumulative BrdU labeling, 78.2% and 78% of the total proliferating populations in wild-type and p107 −/− cortices were BrdU + , respectively, indicating similar cell cycle kinetics. At E13.5 after a 2-h BrdU pulse, 53.6% and 51.2% of the total proliferating populations in wild-type and p107 −/− were BrdU + , respectively. Means were statistically analyzed using a t test, and differences were assessed at *, P < 0.05. Error bars represent SEM. Bar, 100 μm.

Article Snippet: Immunohistochemistry was performed on coronal cryostat sections from embryonic and adult brains with primary antibodies to mouse anti-NeuN (1:100; Chemicon), rabbit antiactive caspase-3 (1:500; BD Biosciences), rabbit anti-PH3 (1:400; Upstate Biotechnology), rat anti-BrdU (1:100; Accurate Chemicals), mouse anti-BrdU (1:100; Becton Dickinson), mouse anti-PCNA (1:300; Vector Laboratories), goat antidoublecortin (1:100; Santa Cruz Biotechnology, Inc.), mouse anti–βIII-tubulin (mouse monoclonal hybridoma supernatant; 1:100; ), and mouse anti-Nestin (1:400; Research Diagnostics).

Techniques: In Vivo, Immunohistochemistry, BrdU Incorporation Assay, Marker, Immunolabeling, Labeling

Journal: The Journal of Cell Biology

Article Title: The Retinoblastoma family member p107 regulates the rate of progenitor commitment to a neuronal fate

doi: 10.1083/jcb.200703176

Figure Lengend Snippet: Defective differentiation in p107 −/− neural precursors. Long-term BrdU birthdating analysis (E13–18) showed a reduction in the number of cells exiting the VZ and arriving in the cortical plate. Pregnant females received an intraperitoneal injection of 20 mg/kg BrdU at gestation day 13.5 (E13.5). Embryos were collected 5 d later at E18.5, and BrdU + cells were counted in the developing cortices. (a–c) p107 −/− embryos had significantly fewer BrdU + cells in the cortex in contrast to wild-type embryos (wild type, n = 4; p107 −/− , n = 4). (d–f) To assess the number of neurons born at E13, short-term BrdU birthdating (E13–14) was performed with pregnant females receiving an intraperitoneal injection of 20 mg/kg BrdU at E13.5, and embryos were collected 24 h later at E14.5. (d and e) Strongly labeled BrdU + cells were primarily located in the IZ and SVZ. (f) Quantification of the number of strongly labeled BrdU + cells revealed a significant reduction in p107-deficient brains. (g–i) Double labeling of BrdU and PCNA was performed to assess whether cells had undergone terminal mitosis and exited the cell cycle (i.e., BrdU + PCNA − cells; arrows). (j–o) Strongly labeled BrdU + cells were colabeled with doublecortin (DCX), a marker of migratory cells (j–l), and Tuj1, an early neuronal marker identifying these cells as newly born neurons (m–o). Arrows in j, k, m, and n identify double-labeled cells. The lower number of BrdU-positive cells in the cortex of p107 −/− embryos at E18.5 and in the SVZ and IZ at E14.5 despite an expanded precursor population demonstrates a defect in the commitment to a neuronal fate. Means for each measurement was assessed with a t test. Error bars represent SEM. cp, cortical plate. *, P < 0.05. Bars: (a–e) 100 μm; (g, h, j, k, m, and n) 25 μm.

Article Snippet: Immunohistochemistry was performed on coronal cryostat sections from embryonic and adult brains with primary antibodies to mouse anti-NeuN (1:100; Chemicon), rabbit antiactive caspase-3 (1:500; BD Biosciences), rabbit anti-PH3 (1:400; Upstate Biotechnology), rat anti-BrdU (1:100; Accurate Chemicals), mouse anti-BrdU (1:100; Becton Dickinson), mouse anti-PCNA (1:300; Vector Laboratories), goat antidoublecortin (1:100; Santa Cruz Biotechnology, Inc.), mouse anti–βIII-tubulin (mouse monoclonal hybridoma supernatant; 1:100; ), and mouse anti-Nestin (1:400; Research Diagnostics).

Techniques: Injection, Labeling, Marker

Journal: Small (Weinheim an der Bergstrasse, Germany)

Article Title: Capsaicin Enhanced the Efficacy of Photodynamic Therapy Against Osteosarcoma via a Pro-Death Strategy by Inducing Ferroptosis and Alleviating Hypoxia.

doi: 10.1002/smll.202306916

Figure Lengend Snippet: Figure 8. Evaluation and biosafety of different treatments. A) H&E staining and images of PCNA, Ki67, TRPV1, Gpx4, and HIF-1𝛼. IHC staining in tumor specimens after various treatments (scale bar = 100 μm). B–F) IOD values in various groups were analyzed by processing IHC staining through ImagePro Plus. G) Triple IF staining of TRPV1 (red), Gpx4 (orange), and HIF-1𝛼(green) in tumor specimens after various treatments (scale bar = 50 μm). H) TUNEL staining of tumor sections after various treatments (scale bar = 50 μm). I) H&E staining of major organs of mice after different treatments (scale bar = 100 μm). J) Hematological and blood biochemical tests of mice after various treatments.

Article Snippet: Mouse anti-human TRPV1 (cat. 66983-1-Ig), mouse anti-human NQO1 (cat. 67240-1-Ig), rabbit anti-human GCLM (cat. 14241-1-AP), rabbit anti-human PCNA (cat. 10205-2-AP), rabbit anti-human KI67 (cat. 27309-1-AP), and rabbit antihuman HIF-1α (cat. 20960-1-AP) were acquired from Proteintech Corp (USA).

Techniques: Staining, Immunohistochemistry, TUNEL Assay