Journal: Neurology Research International

Article Title: Subarachnoid Transplant of the Human Neuronal hNT2.19 Serotonergic Cell Line Attenuates Behavioral Hypersensitivity without Affecting Motor Dysfunction after Severe Contusive Spinal Cord Injury

doi: 10.1155/2011/891605

Figure Lengend Snippet: Neural and human markers with differentiation of the hNT2.19 cell line in vitro. The hNT2.19 cell line was treated for two weeks with retinoic acid and mitotic inhibitors and lifted to substrate-coated 8-well plastic TC slides for differentiation and immunohistochemistry for neuron-specific markers. As soon as 4 days in vitro, a variety of neural markers appeared, which remained strong until at least 6 wks of differentiation: TuJ1 (a), hNSE (b), NFL (c), NFM (d), and NFH (e). For comparison, the negative control hNT2.6 cell line was cultured similarly as the hNT2.19 cells and is here stained for TuJ1 (f). Magnification bar = 20 nm, (a–f).

Article Snippet: For immunohistochemistry of sectioned spinal cord tissues, the polyclonal antibody anti-5HT (ab10385; dilution 1/100 (in vivo)) was purchased from Abcam Inc, Cambridge, MA, and the antihuman TuJ1 antibody (Neuron-specific class III beta-tubulin) was purchased from Neuromics, Edina, MN (MO15013; dilution 1/100 (in vivo).

Techniques: In Vitro, Immunohistochemistry, Comparison, Negative Control, Cell Culture, Staining

Journal: Neurology Research International

Article Title: Subarachnoid Transplant of the Human Neuronal hNT2.19 Serotonergic Cell Line Attenuates Behavioral Hypersensitivity without Affecting Motor Dysfunction after Severe Contusive Spinal Cord Injury

doi: 10.1155/2011/891605

Figure Lengend Snippet: Transplant of hNT2.19 and hNT2.6 cell lines in the severe contusive SCI model: TuJ1 and 5HT immunohistochemistry. Rats were injured with severe contusive SCI followed at two weeks by hNT2.6 (a, b) or hNT2.19 (c, d) cell grafts. Sagittal spinal cord sections were examined at 8 wks after SCI for evidence of surviving lumbar subarachnoid hNT2.6 (a, b) or hNT2.19 (c, d) cell line grafts, utilizing TuJ1 (a, c) or 5HT (b, d) immunohistochemistry. The hNT2.19 and control hNT2.6 (10 6 cells/injection), which had been differentiated for two weeks in vitro, were injected into the subarachnoid space two weeks after the SCI. Cell graft sites were colocalized with 5HT (b, d) and the human-specific marker TUJ1 (neuron-specific class III β -tubulin; (a, c)). There are many surviving hNT2.19 (c) and hNT2.6 (a) grafted cells visible on the pial surface, which stain for TuJ1 (arrows) at the end of the experiment, 56 days after SCI and about 6 weeks after cell transplant. Adjacent sections with the same grafted hNT2.19 (d) and hNT2.6 cells (b) are stained for 5HT, but only the hNT2.19 cells (d) are labeled for 5HT (arrows).

Article Snippet: For immunohistochemistry of sectioned spinal cord tissues, the polyclonal antibody anti-5HT (ab10385; dilution 1/100 (in vivo)) was purchased from Abcam Inc, Cambridge, MA, and the antihuman TuJ1 antibody (Neuron-specific class III beta-tubulin) was purchased from Neuromics, Edina, MN (MO15013; dilution 1/100 (in vivo).

Techniques: Immunohistochemistry, Control, Injection, In Vitro, Marker, Staining, Labeling

Journal: bioRxiv

Article Title: Loss of p27 Kip1 causes metabolic reprogramming and is sufficient to induce a Warburg effect and glutamine addiction in untransformed cells

doi: 10.64898/2026.02.06.703945

Figure Lengend Snippet: (a) Graphical representation of pyruvate fate after glycolysis. (b, c) Lactate levels in the exometabolome of p27 +/+ (n = 8) and p27 -/- (n = 4) MEFs grown in full medium (b) or glucose starvation medium (c) for 24 h. Data is presented as box plot, means are represented by yellow squares. ns: p > 0.05; ***: p < 0.001. (d) LDHA gene expression in p27 +/+ (n = 8) and p27 -/- (n = 4) MEFs grown in full medium for 24 h from RNA-Seq data expressed as Count Per Million, normalized to p27 +/+ levels. Graph shows means ± SEM. ****: p < 0.0001. ( e ) LDHA mRNA levels in p27 +/+ and p27 -/- MEFs by RT-qPCR, normalized to p27 +/+ levels. Graph shows means ± SEM from five independent experiments. ***: p < 0.001. ( f ) Immunoblots for LDHA, p27 and actin in p27 +/+ and p27 -/- MEFs (left panel). Graph shows means ± SEM of the quantification of LDHA levels normalized to actin (n = 5) (right panel). *: p < 0.05. ( g ) LDHB gene expression in p27 +/+ (n = 8) and p27 -/- (n = 4) MEFs grown in full medium for 24 h from RNA-Seq data expressed as Count Per Million, normalized to p27 +/+ levels. Graph shows means ± SEM. ****: p < 0.0001. ( h ) LDHB mRNA levels in p27 +/+ and p27 -/- MEFs by RT-qPCR, normalized to p27 +/+ levels. Graph shows means ± SEM from five independent experiments. **: p < 0.01. ( i ) Immunoblots for LDHB, p27, and β-tubulin in p27 +/+ and p27 -/- MEFs (left panel). Graph shows means ± SEM of the quantification of LDHB levels normalized to β-tubulin or actin (n = 8) (right panel). ****: p < 0.0001. (j) PDK1 gene expression in p27 +/+ (n = 8) and p27 -/- (n = 4) MEFs grown in full medium for 24 h from RNA-Seq data expressed as Count Per Million, normalized to p27 +/+ levels. Graph shows means ± SEM. ****: p < 0.0001. (k) PDK1 mRNA level in p27 +/+ and p27 -/- MEFs by RT-qPCR, normalized to p27 +/+ levels. Graph shows means ± SEM from 5 independent experiments. **: p < 0.01. ( l ) Immunoblots for PDK1, p27 and actin in p27 +/+ and p27 -/- MEFs (left panel). Graph shows means ± SEM of the quantification of PDK1 levels normalized to actin (n = 6) (right panel). *: p < 0.05. (m) Immunoblots for Lys18-lactylated Histone H3 (HH3-K18-La), p27 and actin in p27 +/+ and p27 -/- MEFs (left panel). Graph shows means ± SEM of the quantification of HH3-K18-La levels normalized to actin (n = 6) (right panel). **: p < 0.01. (n) ATP quantification in p27 +/+ and p27 -/- MEFs treated with either vehicle or Sodium Oxamate (50 mM) for 1 h. Graph shows means ± SEM from four independent experiments. *: p < 0.05; ***: p < 0.001.

Article Snippet: Rabbit anti-Hexokinase 1 (C35C4; WB 1/1000), GAPDH (D16H11; WB 1/1000), PKM1/2 (C103A3; WB 1/1000), LDHA (2012S, WB 1/1000), GLUT1 (E4S6I; WB 1/1000; IF 1/100), PDK1 (3820S; WB 1/1000), β-Tubulin (2128S, WB 1/1000) were purchased from Cell Signalling Technology.

Techniques: Gene Expression, RNA Sequencing, Quantitative RT-PCR, Western Blot