Journal: Annals of Translational Medicine

Article Title: Identifying potential therapeutic targets of Tang-Yi-Ping for the treatment of impaired glucose tolerance: a tandem mass tag-labeled quantitative proteomic analysis

doi: 10.21037/atm-21-4257

Figure Lengend Snippet: Specific information on 16 protein targets in the treatment of IGT with TYP

Article Snippet: For example, the expression levels of Rbp4 (boster, #PB0368, China) and Flot2 (boster, #PB0663, China) in the 3 groups were calculated with GAPDH (boster, #BM1623, China) as an internal reference.

Techniques:

Journal: Annals of Translational Medicine

Article Title: Identifying potential therapeutic targets of Tang-Yi-Ping for the treatment of impaired glucose tolerance: a tandem mass tag-labeled quantitative proteomic analysis

doi: 10.21037/atm-21-4257

Figure Lengend Snippet: Relative expression levels of Rbp4 and Flot2 proteins in pancreatic samples of the 3 groups. Rbp4: *, P<0.05, vs. Control; #, P<0.05, vs. IGT. Flot: *, P<0.05, vs. Control; #, P<0.05, vs. IGT. IGT, impaired glucose tolerance.

Article Snippet: For example, the expression levels of Rbp4 (boster, #PB0368, China) and Flot2 (boster, #PB0663, China) in the 3 groups were calculated with GAPDH (boster, #BM1623, China) as an internal reference.

Techniques: Expressing, Control

Journal:

Article Title: Dynamic Partitioning into Lipid Rafts Controls the Endo-Exocytic Cycle of the ?L/? 2 Integrin, LFA-1, during Leukocyte Chemotaxis D⃞

doi: 10.1091/mbc.E05-05-0413

Figure Lengend Snippet: αL/β2 partitions into lipid rafts upon chemokine stimulation in PMN. (A) Freshly isolated PMN were incubated in the absence or presence of 100 nM fMLF for 5 min at 37°C, followed by lysis in 1% Triton X-100 and floatation on a discontinuous step sucrose gradient. The relative distribution of αL and the raft marker flotillin-2 were analyzed by Western blotting. (B) Densitometric analysis of a representative experiment showing the distribution of αL and flotillin-2 over a sucrose gradient in resting (control) or activated (100 nM fMLF) PMN.

Article Snippet: Monoclonal antibody (mAb) TS1.22 (anti-αL) was kindly provided by T. A. Springer (Harvard Medical School, Boston, MA). mAbs anti-αL clone 27, anti-flotillin-2 clone 29, anti-LAMP-1, and anti-Rab11 clone 47 were from BD Biosciences (San Jose, CA). mAb anti-rat β 2 clone WT3 was purchased from American Type Culture Collection (Manassas, VA).

Techniques: Isolation, Incubation, Lysis, Marker, Western Blot

Journal: The Journal of Neuroscience

Article Title: Reggies/Flotillins Regulate Retinal Axon Regeneration in the Zebrafish Optic Nerve and Differentiation of Hippocampal and N2a Neurons

doi: 10.1523/JNEUROSCI.0870-09.2009

Figure Lengend Snippet: Reggie downregulation reduces axon outgrowth. A, Mo antisense oligonucleotides labeled with lissamine were applied immediately after ONS. A maximum-intensity projection of a deconvoluted Z-stack from a retina, 3 d after ONS, illustrates that RGCs and axons are labeled by retrograde Mo transport. B, C, Reggie-2 Ab immunostaining is present in control Mo-treated retinas which did not alter reggie expression (B). Reggie Mos reduced reggie-2 Ab staining of RGCs 7 d after ONS (C, images were taken with the same microscope settings). Scale bars, 50 μm. D, Regeneration was assessed by quantifying axons from mini-explants isolated 4 d after ONS. Retina pairs (21) (control- and reggie–Mo-treated eyes from the same fish) were analyzed. The mean number of axons per explant was normalized to the control retina, and relative outgrowth efficiency is shown.

Article Snippet: Anti-Reggie-1 [ESA (epidermal surface antigen)], anti-cdc42, and anti-Rac1 monoclonal antibodies (mAbs) were from BD Transduction Laboratories, anti-RhoA mAb, anti-Arp3 polyclonal antibody (pAB), and anti N-WASP mAb from Santa Cruz, anti-Ras mAb from Oncogene/Calbiochem.

Techniques: Labeling, Immunostaining, Expressing, Staining, Microscopy, Isolation

Journal: The Journal of Neuroscience

Article Title: Reggies/Flotillins Regulate Retinal Axon Regeneration in the Zebrafish Optic Nerve and Differentiation of Hippocampal and N2a Neurons

doi: 10.1523/JNEUROSCI.0870-09.2009

Figure Lengend Snippet: Reggie-1 knockdown impairs process formation in N2a cells. Cells transfected with either GL2 control siRNAs or a mix of reggie-1-specific siRNAs were stimulated with IGF-1, fixed 24 h after stimulation, and stained with phalloidin to visualize F-actin and cell morphology. Untreated and GL2 siRNA-transfected cells produced numerous filopodia (A, B), whereas reggie-1 downregulation (R1 siRNA) led to the formation of large lamellipodia (F). Significantly fewer cells formed neurites after reggie-1 siRNA transfection (E), which efficiently downregulated reggie-1 and reggie-2 expression compared with GL2 siRNA cells in Western blot experiments (G). D, N2a cells were simultaneously transfected with reggie-1 siRNA and the reggie-1 construct (R1–EGFP rescue) without siRNA-binding sites. A large proportion of the cells no longer exhibited the lamellipodia-rich phenotype but had instead many filopodia and neurites (D–F) much as control cells (A, B). Mean ± SEM; *p < 0.05, **p < 0.01, ***p < 0.001, Student's t test. Scale bars, 20 μm.

Article Snippet: Anti-Reggie-1 [ESA (epidermal surface antigen)], anti-cdc42, and anti-Rac1 monoclonal antibodies (mAbs) were from BD Transduction Laboratories, anti-RhoA mAb, anti-Arp3 polyclonal antibody (pAB), and anti N-WASP mAb from Santa Cruz, anti-Ras mAb from Oncogene/Calbiochem.

Techniques: Transfection, Staining, Produced, Expressing, Western Blot, Construct, Binding Assay

Journal: The Journal of Neuroscience

Article Title: Reggies/Flotillins Regulate Retinal Axon Regeneration in the Zebrafish Optic Nerve and Differentiation of Hippocampal and N2a Neurons

doi: 10.1523/JNEUROSCI.0870-09.2009

Figure Lengend Snippet: Reggie-1 is required for hippocampal neuron differentiation. Representative wide-field images of transfected hippocampal neurons. A, Cells transfected with control (GL2) siRNA differentiated normally (insert) having long thin axons (to the right) and dendritic arborizations. B, C, Note the short and thick protrusions, large lamellipodia, and broad dendrites of reggie-1 (R1) siRNA-treated cells. D, Cells simultaneously transfected with the reggie-1 rescue construct (R1–EGFP rescue) and reggie-1 siRNAs differentiated normally. Scale bars, 20 μm. Histogram in E shows the quantification of the cells transfected with control GL2 siRNA, with reggie-1 siRNAs, and simultaneously transfected with R1–EGFP rescue construct and reggie-1 siRNAs.

Article Snippet: Anti-Reggie-1 [ESA (epidermal surface antigen)], anti-cdc42, and anti-Rac1 monoclonal antibodies (mAbs) were from BD Transduction Laboratories, anti-RhoA mAb, anti-Arp3 polyclonal antibody (pAB), and anti N-WASP mAb from Santa Cruz, anti-Ras mAb from Oncogene/Calbiochem.

Techniques: Transfection, Construct

Journal: The Journal of Neuroscience

Article Title: Reggies/Flotillins Regulate Retinal Axon Regeneration in the Zebrafish Optic Nerve and Differentiation of Hippocampal and N2a Neurons

doi: 10.1523/JNEUROSCI.0870-09.2009

Figure Lengend Snippet: Reggie-1 downregulation affects Rho GTPase activation in N2a cells. N2a cells starved overnight, stimulated with 50 ng/ml IGF-1 for 5 min, and assayed for GTP-loading of small GTPases. The histograms, A–D, show changes in the intensity (relative to controls) in the respective Western blots (n = 4), which are exemplified below each histogram. Mean ± SEM; *p < 0.05, **p < 0.01, ***p < 0.001, Student's t test. Total Rac, cdc 42, RhoA, and Ras of crude lysates were used as loading control. A, B, In reggie-1 siRNA-treated cells, the activation patterns of Rac1 and cdc42 were significantly altered. IGF-1 application increased the activation of Rac1 in control-transfected cells, whereas siRNA-treated cells showed an elevated level of activated Rac1 without IGF-1, which increased further after IGF application. In contrast, cdc42 activity decreased during IGF application in control-transfected cells and remained at low levels in siRNA-transfected cells with or without IGF-1. C, RhoA activation increased during IGF application in control cells and further increased in reggie-1 siRNA-treated cells. D, Ras stimulation by IGF in control transfectants was absent in reggie–siRNA-treated cells.

Article Snippet: Anti-Reggie-1 [ESA (epidermal surface antigen)], anti-cdc42, and anti-Rac1 monoclonal antibodies (mAbs) were from BD Transduction Laboratories, anti-RhoA mAb, anti-Arp3 polyclonal antibody (pAB), and anti N-WASP mAb from Santa Cruz, anti-Ras mAb from Oncogene/Calbiochem.

Techniques: Activation Assay, Western Blot, Transfection, Activity Assay

Journal: The Journal of Neuroscience

Article Title: Reggies/Flotillins Regulate Retinal Axon Regeneration in the Zebrafish Optic Nerve and Differentiation of Hippocampal and N2a Neurons

doi: 10.1523/JNEUROSCI.0870-09.2009

Figure Lengend Snippet: Coimmunoprecipitation assays for the analysis of the complex of Arp3, N-WASP, and cortactin and phosphorylation of cofilin and cortactin in reggie siRNA-treated N2a cells. A, In IGF-1 stimulated control cells, Abs against N-WASP and cortactin coprecipitate Arp3. In reggie-1 siRNA-treated cells, the Arp3, N-WASP, cortactin complex is present at lower levels and is almost disrupted during IGF stimulation. B, The phosphorylation of cortactin at Tyr466 is unchanged after IGF-1 treatment of control cells but is strongly reduced during IGF-1 treatment of reggie-1 siRNA cells. C, The phosphorylation of cofilin at Ser3 slightly decreases during IGF stimulation but is increased in reggie-1 siRNA-treated cells with or without IGF-1.

Article Snippet: Anti-Reggie-1 [ESA (epidermal surface antigen)], anti-cdc42, and anti-Rac1 monoclonal antibodies (mAbs) were from BD Transduction Laboratories, anti-RhoA mAb, anti-Arp3 polyclonal antibody (pAB), and anti N-WASP mAb from Santa Cruz, anti-Ras mAb from Oncogene/Calbiochem.

Techniques:

Journal: The Journal of Neuroscience

Article Title: Reggies/Flotillins Regulate Retinal Axon Regeneration in the Zebrafish Optic Nerve and Differentiation of Hippocampal and N2a Neurons

doi: 10.1523/JNEUROSCI.0870-09.2009

Figure Lengend Snippet: Reggie-1 downregulation affects signaling in N2a cells. A, Activation of p38 was reduced in reggie siRNA-treated cells independently of IGF-1 stimulation in comparison with control transfectants. B, ERK1/2 activation was decreased in response to reggie siRNA. C, pFAK showed a significant activation during IGF-1 stimulation, and this stimulation was reduced in reggie-1 siRNA-treated cells. Total FAK was decreased in reggie-1 siRNA-treated cells. Total p38 (A), ERK1/2 (B), and GAPDH (C) were used as loading controls. Mean ± SEM; n = 4; *p < 0.05, **p < 0.01, ***p < 0.001, Student's t test. D, JNK, PKC, and PKB phosphorylation was not affected by reggie-1 siRNA-treatment.

Article Snippet: Anti-Reggie-1 [ESA (epidermal surface antigen)], anti-cdc42, and anti-Rac1 monoclonal antibodies (mAbs) were from BD Transduction Laboratories, anti-RhoA mAb, anti-Arp3 polyclonal antibody (pAB), and anti N-WASP mAb from Santa Cruz, anti-Ras mAb from Oncogene/Calbiochem.

Techniques: Activation Assay