Journal: Neurogastroenterology and motility : the official journal of the European Gastrointestinal Motility Society

Article Title: Zic2 is required for enteric nervous system development and neurite outgrowth: a mouse model of enteric hyperplasia and dysplasia

doi: 10.1111/nmo.12101

Figure Lengend Snippet: Abnormal ENS development and neurite outgrowth in Zic2m1Nisw/m1Nisw embryos (A, B) Material is retained in the intestines of E18.5 mutants (B) presenting as green intestines vs the normal yellow intestines of wild-type embryos (A); N > 50. (C–H) Whole-mount NADPH diaphorase staining of E18.5 wild type (C, E, G) and Zic2m1Nisw/m1Nisw (D, F, H) gut regions; N = 5. (I) Statistically significant increase in NADPH diaphorase-stained cells in the mutant gut compared with wild type (E18.5; N = 4). (J–O) Whole-mount immunostaining for the indicated markers of E18.5 wild type (J–J”, L, N) or Zic2m1Nisw/m1Nisw (K–K”, M, O) midguts showing abnormal neurite pattern; N = 5. (P) Statistically significant decrease in the distance between internodal strands in the mutant gut compared with wild type (E18.5). (Q–R) Segments of E13.5 proximal midgut were cultured in 3D collagen matrix with GDNF for 3 days and stained with p75NTR antibody (green) to detect ENCC. Zic2m1Nisw/m1Nisw mutant ENCCs (R) showed extensive outgrowth of neurites compared with wild type (Q); N = 4. (S) Statistically significant increase in neurite outgrowth in the mutant gut compared with wild type. Neurite length was measured from the confluent edge of the migrating ENCC to the distal tip of the neurite (highlighted by an arrow). Data expressed as mean ± standard deviation (SD) (*P < 0.05, ***P < 0.001, Student’s t-test).

Article Snippet: Gut explant culture For collagen gel gut explant cultures, E13.5 proximal hindgut segments were placed on a glass bottom dish (MatTek, Ashland, MA, USA) coated with 1 mg mL −1 3D collagen gel (R&D, Minneapolis, MN, USA) with 10 ng mL −1 GDNF (USBiological, Marblehead, MA, USA) and cultured for 3 days.

Techniques: Staining, Mutagenesis, Immunostaining, Cell Culture, Standard Deviation

Journal: Human Molecular Genetics

Article Title: Minimally invasive transplantation of iPSC-derived ALDHhiSSCloVLA4+ neural stem cells effectively improves the phenotype of an amyotrophic lateral sclerosis model

doi: 10.1093/hmg/ddt425

Figure Lengend Snippet: NSCs produce neurotrophins and reduce micro- and macrogliosis in SOD1G93A mice. The amounts of ( A ) GDNF, ( B ) BDNF, ( C ) NT3 and ( D ) TGF-α secreted by the NSCs were determined by ELISA, which demonstrated a significant production of these neuroprotective substances with respect to fibroblasts. ( E ) Expression of GFAP (red) was reduced in the spinal cord of transplanted SOD1 mice. Nuclei are stained with DAPI (blue). Scale bar: 70 µm. NSC transplantation significantly reduced the presence of microglial cells ( F ) and astrocytes ( G ) in the spinal cord of intrathecally and systemically injected mice (data represent the mean ± SD, *significant, P < 0.001, ANOVA).

Article Snippet: On day 24, we added the growth factors BDNF, GDNF and insulin-like growth factor-1 (10 ng/ml each; PeproTech).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Staining, Transplantation Assay, Injection

Journal: Nature protocols

Article Title: Generation of human brain region–specific organoids using a miniaturized spinning bioreactor

doi: 10.1038/nprot.2017.152

Figure Lengend Snippet: Culture media.

Article Snippet: Mouse embryonic fibroblast feeder cells isolated from E13.5 CF1 mouse embryos (Charles River, strain code 023) that have been growth-arrested by γ-irradiation using a Cs-137 irradiator Growth media components and supplements DMEM (Corning, cat. no. 10–013) DMEM/F-12, HEPES (Gibco, cat. no. 11330032) Neurobasal medium (Gibco, cat. no. 21103049) KnockOut Serum Replacement (KOSR; Gibco, cat. no. 10828028) GlutaMAX supplement (Gibco, cat. no. 35050061) MEM non-essential amino acids solution (MEM-NEAA; Gibco, cat. no. 11140050) Penicillin–streptomycin (Pen/Strep; Gibco, cat. no. 15140122) Amphotericin B (Thermo Fisher, cat. no. 15290026 2-Mercaptoethanol (Gibco, cat. no. 21985023) N-2 supplement (Gibco, cat. no. 17502048) B-27 supplement (Gibco, cat. no. 17504044) Matrigel growth factor-reduced basement membrane matrix (Corning, cat. no. 354230) Insulin solution (Sigma-Aldrich, cat. no. I9278–5ML) FBS (Corning, cat. no. 35–010) Small molecules and growth factors A83–01 (Stem Cell Technologies, cat. no. 72022) Dorsomorphin (Stem Cell Technologies, cat. no. 72102) SB-431542 (Stem Cell Technologies, cat. no. 72232) CHIR-99021 (Stem Cell Technologies, cat. no. 72052) Purmorphamine (Stem Cell Technologies, cat. no. 72202) LDN-193189 (Stem Cell Technologies, cat. no. 72146) FGF-2 (Peprotech, cat. no. 100–18B) FGF-8 (Peprotech, cat. no. 100–25A) WNT-3A (R&D Systems, cat. no. 5036-WN-010) SHH (Peprotech, cat. no. 100–45) CNTF (Peprotech, cat. no. 450–13) BDNF (Peprotech, cat. no. 450–02) GDNF (Peprotech, cat. no. 450–10) Ascorbic acid (Sigma-Aldrich, cat. no. 1043003) cAMP (Sigma-Aldrich, cat. no. A9501) Enzymes and other reagents Collagenase, type IV (Thermo Fisher, cat. no. 17104019) BSA (Sigma-Aldrich, cat. no. A9418) Dulbecco’s PBS without calcium and magnesium (DPBS; Corning, cat. no. 21–031) Donkey serum (Millipore, cat. no. S30) 0.1% (wt/vol) Gelatin in water (Stem Cell Technologies, cat. no. 7903) Ethyl alcohol, 95% (vol/vol), 190 proof, pure (Pharmco-Aaper, cat. no. 111000190CSGL) Sterile deionized water DMSO (Sigma-Aldrich, cat. no. 472301) 1-Thioglycerol (Sigma-Aldrich, cat. no. M1753) 10% (vol/vol) Bleach 16% (vol/vol) Formaldehyde, methanol free (Polysciences, cat. no. 18814–10) Sucrose (Sigma-Aldrich, cat. no. S5016) Tissue-freezing medium (General Data, cat. no. TFM-5) Triton X-100 (Sigma-Aldrich, cat. no. T9284) Tween 20 (Sigma-Aldrich, cat. no. P7949) Slide-mounting solution (Fisher Scientific, cat. no. 14-390-5)

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