cell cycle exit  (MedChemExpress)

Journal:

Article Title: BM88/CEND1 coordinates cell cycle exit and differentiation of neuronal precursors

doi: 10.1073/pnas.0610973104

Figure Lengend Snippet: BM88 knockdown in NPCs induces proliferation and impairs neuronal differentiation. (A) Relative expression levels of BM88 mRNA in the developing and postnatal mouse spinal cord and isolated NPCs, measured with quantitative real-time RT-PCR. (B) BM88 immunostaining (green) of dissociated NPCs 72 h a.e. with BM88-specific siRNA (siBM88) or control siRNA (siCTR); DAPI, blue. (Scale bar: 40 μm.) (C) Quantification of BM88 mRNA by real-time RT-PCR (siBM88, 19.6 ± 4.7% of the siCTR mBM88 mRNA levels). (D–G) BM88 knockdown in NPCs enhances proliferation. Proliferation was measured by counting the numbers of DAPI+ cells after equal plating (siBM88, 855 ± 63.2; siCTR, 560 ± 30.2; P < 0.01) (D), the index of Phos-H3+ cells (siBM88, 2.53 ± 0.35; siCTR, 1.51 ± 0.33; P < 0.01) (E), and neurosphere size 96 h a.e. (siBM88, 100 ± 11.1%; siCTR, 52.2 ± 9.1%; P < 0.05) (F and G). (H and I) BM88 knockdown reduced the potential of NPCs for neuronal differentiation. Immunostaining for βIII-tubulin (H; green) and index of βIII-tubulin+ cells (siBM88, 5.85 ± 1.62; siCTR, 10.95 ± 1.56; P < 0.01) (I). (Scale bar: 40 μm.) (J) siBM88 transfected NPCs show increased numbers of Nestin+ cells compared with siCTR-treated cells (siBM88, 49.3 ± 7.1; siCTR, 30.7 ± 6.8; P < 0.01).

Article Snippet: Here, we report the involvement of BM88 [cell cycle exit and neuronal differentiation 1 (Cend1); National Center for Biotechnology Information nomenclature at www.ncbi.nih.gov ] in such cross talk.

Techniques: Knockdown, Expressing, Isolation, Quantitative RT-PCR, Immunostaining, Control, Transfection

Journal:

Article Title: BM88/CEND1 coordinates cell cycle exit and differentiation of neuronal precursors

doi: 10.1073/pnas.0610973104

Figure Lengend Snippet: Forced expression of BM88 reduces the number of cycling progenitors. (A–D) Double GFP fluorescence/BrdU immunostaining 24 h a.e. with BM88/GFP or GFP alone followed by 2-h BrdU pulse. Note the marked reduction in double-labeled cells (yellow) between A and C. (E) Quantitative analysis of BrdU incorporation. The number of BrdU+ transfected cells (GFP+/BrdU+) 24 h a.e. with BM88/GFP or GFP alone is expressed as percentage of the total number of transfected (GFP+) cells [n = 5 embryos; P < 0.001 for GFP vs. BM88; P = 0.4 for BM88 (dorsal) vs. BM88 (ventral)]. (F and G) BM88-electroporated side shows reduced numbers of phos-H3+ cells. Double BM88/phos-H3 immunostaining 24 h a.e. (H–K) BM88 electroporation reduced expression of markers for neuronal precursors such as Pax7 and Cash1 24 h a.e. Note the depletion of dorsal Pax7+ (H and I; double immunofluorescence) and Cash1+ precursors (J and K; in situ hybridization). (L–Q) BM88 overcomes lateral inhibition by down-regulating Notch1. (L–O) In situ hybridization for Notch1 (M and O) and transgene BM88 (L) or GFP immunolabeling (N) in consecutive spinal cord sections 48 h a.e. Arrows in L and M point to Notch1-depleted/BM88+ areas. (P) Double in situ hybridization for BM88 (blue) and Notch1 (red) 48 h a.e. (Q) Quantification of the transfected cells that are Notch1+ by double in situ hybridization. (R) The majority of BM88-transfected cells tend to migrate outside the Notch1+ VZ. Quantification of this effect in comparison with GFP control electroporations (BM88VZ, 23 ± 6.3, vs. GFPVZ, 41.1 ± 1.7; BM88MZ, 77.2 ± 6.1, vs. GFPMZ, 58.9 ± 1.7; n = 5; for both cases, P < 0.01). (S–V) BM88 expression reduced the numbers of Olig2-positive cells 48 h a.e. (S and T) Double in situ hybridization for BM88 (blue) and Olig2 (red). (U and V) GFP and Olig2 in situ hybridization in consecutive sections. [Scale bars: D, G, I, P, and V (for A–U), 40 μm.]

Article Snippet: Here, we report the involvement of BM88 [cell cycle exit and neuronal differentiation 1 (Cend1); National Center for Biotechnology Information nomenclature at www.ncbi.nih.gov ] in such cross talk.

Techniques: Expressing, Fluorescence, Immunostaining, Labeling, BrdU Incorporation Assay, Transfection, Electroporation, Immunofluorescence, In Situ Hybridization, Inhibition, Immunolabeling, Comparison, Control

Journal:

Article Title: BM88/CEND1 coordinates cell cycle exit and differentiation of neuronal precursors

doi: 10.1073/pnas.0610973104

Figure Lengend Snippet: Induction of precocious and ectopic neurogenesis by BM88. Double labeling of spinal cord sections for βIII-tubulin and transgene BM88 from BM88-electroporated (A and B; E and F) and GFP control embryos (C and D; G and H), 24 and 48 h a.e. (I and J) Double in situ hybridization of spinal cord section for BM88 (blue) and SCG10 (red) 48 h after BM88 electroporation. Arrows point to ectopic terminally differentiated SCG10+ neurons in the VZ. (K) A section consecutive to I and J was stained for NF160 by in situ hybridization. (L) Percentage of ventral over dorsal ectopic SCG10+ neurons per embryo (15 sections per embryo; n = 4 embryos). (M–R) BM88 is sufficient to induce MN markers in the VZ. Consecutive sections from BM88-electroporated embryos were stained 48 h a.e. for BM88 transgene (M) and the MN markers ChAT (N), Islet1 (O and P), and Lim3 (Q and R). P is a larger magnification of the area pointed by the arrow in O, and the inset shows at larger magnification an overlay of Islet1/BM88 immunolabeling corresponding to the area marked by the dotted white rectangle in P. (R) Double immunofluorescence for Lim3 and BM88. [Scale bars: B, D, H, and N (for A–N), 40 μm; O (for O, Q, and R), 40 μm; and P, 40 μm.]

Article Snippet: Here, we report the involvement of BM88 [cell cycle exit and neuronal differentiation 1 (Cend1); National Center for Biotechnology Information nomenclature at www.ncbi.nih.gov ] in such cross talk.

Techniques: Labeling, Control, In Situ Hybridization, Electroporation, Staining, Immunolabeling, Immunofluorescence

Journal:

Article Title: BM88/CEND1 coordinates cell cycle exit and differentiation of neuronal precursors

doi: 10.1073/pnas.0610973104

Figure Lengend Snippet: The BM88 effect on neurogenesis is not recapitulated by forced cell cycle exit of neuronal precursors. (A and B) Misexpression of p27Kip1 results in markedly reduced BrdU incorporation as revealed by immunohistochemical detection of BrdU 24 h a.e. (C) Quantification of BrdU incorporation due to GFP control, BM88, and p27Kip1 electroporation. The numbers of GFP+/BrdU+ cells are expressed as percentage of the total number of GFP+ transfected cells (GFP, 38.6 ± 6.64%; BM88, 4.34 ± 2.27%; p27Kip1, 1.82 ± 0.45%; n = 5 embryos; P < 0.001 for GFP vs. BM88 or p27Kip1). (D and E) Forced expression of p27Kip1 in the chick neural tube is not sufficient to reproduce the ectopic neurogenesis effect of BM88 48 h a.e. (D and E) βIII-tubulin immunolabeling/GFP fluorescence. (F–H) In situ hybridization for SCG10 (F), ChAT (G), and Notch1 (H). [Scale bars: B (for A and B and D–H), 40 μm.]

Article Snippet: Here, we report the involvement of BM88 [cell cycle exit and neuronal differentiation 1 (Cend1); National Center for Biotechnology Information nomenclature at www.ncbi.nih.gov ] in such cross talk.

Techniques: BrdU Incorporation Assay, Immunohistochemical staining, Control, Electroporation, Transfection, Expressing, Immunolabeling, Fluorescence, In Situ Hybridization