The Gemini planet imager view of the HD 32297 debris disk

Funding: M.M.B. and J.M. were supported by NASA through Hubble Fellowship grants #51378.01-A and HST-HF2-51414.001, respectively, and I.C. through Hubble Fellowship grant HST-HF2-51405.001-A, awarded by the Space Telescope Science Institute, which is operated by AURA, for NASA, under contract NAS5-26555.We present new H-band scattered light images of the HD 32297 edge-on debris disk obtained with the Gemini Planet Imager. The disk is detected in total and polarized intensity down to a projected angular separation of 0"15, or 20 au. On the other hand, the large-scale swept-back halo remains undetected, likely a consequence of its markedly blue color relative to the parent body belt. We analyze the curvature of the disk spine and estimate a radius of ≍100 au for the parent body belt, smaller than past scattered light studies but consistent with thermal emission maps of the system. We employ three different flux-preserving post-processing methods to suppress the residual starlight and evaluate the surface brightness and polarization profile along the disk spine. Unlike past studies of the system, our high-fidelity images reveal the disk to be highly symmetric and devoid of morphological and surface brightness perturbations. We find the dust scattering properties of the system to be consistent with those observed in other debris disks, with the exception of HR 4796. Finally, we find no direct evidence for the presence of a planetary-mass object in the system.Publisher PDFPeer reviewe

Low Levels of GSTA1 Expression Are Required for Caco-2 Cell Proliferation

The colonic epithelium continuously regenerates with transitions through various cellular phases including proliferation, differentiation and cell death via apoptosis. Human colonic adenocarcinoma (Caco-2) cells in culture undergo spontaneous differentiation into mature enterocytes in association with progressive increases in expression of glutathione S-transferase alpha-1 (GSTA1). We hypothesize that GSTA1 plays a functional role in controlling proliferation, differentiation and apoptosis in Caco-2 cells. We demonstrate increased GSTA1 levels associated with decreased proliferation and increased expression of differentiation markers alkaline phosphatase, villin, dipeptidyl peptidase-4 and E-cadherin in postconfluent Caco-2 cells. Results of MTS assays, BrdU incorporation and flow cytometry indicate that forced expression of GSTA1 significantly reduces cellular proliferation and siRNA-mediated down-regulation of GSTA1 significantly increases cells in S-phase and associated cell proliferation. Sodium butyrate (NaB) at a concentration of 1 mM reduces Caco-2 cell proliferation, increases differentiation and increases GSTA1 activity 4-fold by 72 hours. In contrast, 10 mM NaB causes significant toxicity in preconfluent cells via apoptosis through caspase-3 activation with reduced GSTA1 activity. However, GSTA1 down-regulation by siRNA does not alter NaB-induced differentiation or apoptosis in Caco-2 cells. While 10 mM NaB causes GSTA1-JNK complex dissociation, phosphorylation of JNK is not altered. These findings suggest that GSTA1 levels may play a role in modulating enterocyte proliferation but do not influence differentiation or apoptosis

GSTA1 levels can be modulated in preconfluent Caco-2 cells.

<p>(A) Representative western blot of GSTA1 (∼25 KDa) protein expression in preconfluent Caco-2 cells that were transiently transfected with 40 nM of GSTA1 siRNA or non-specific (NS) siRNA for 72 h. (B) Representative western blot of V5 (∼26 KDa) protein expression in preconfluent Caco-2 cells that were transiently transfected with one µg of GSTA1-V5 or empty vector (EV) for 48 h. β-actin (∼42 KDa) was used as a protein loading control in all panels.</p

Modulation of GSTA1 levels mediate changes in Caco-2 cell growth.

<p>Effect of (A) GSTA1 down-regulation and (B) GSTA1-V5 overexpression on Caco-2 cell viability evaluated by MTS assay over three days. Asterisks depict significant differences between controls and the cells with GSTA1 modulated levels (*, <i>p</i>≤0.05; and **, <i>p</i>≤0.01). (C) Effect of GSTA1-V5 over-expression on cellular proliferation at 72 h as determined by BrdU incorporation in Caco-2 cells. Bars indicated by different letters differ significantly from one another (p≤0.001). Values represent the mean ± S.E. of four independent experiments with three replicates each.</p

Distinct doses of NaB differently affect cell proliferation and AlkP and GSTA1 enzyme activities.

<p>Preconfluent Caco-2 cells were treated with NaB (1 mM and 10 mM) in serum-free media. (A) Cellular proliferation was assessed from 24–96 h. Asterisks depict significant differences between control and NaB treatments (*, p≤0.05; **, p≤0.01 and ***, p≤0.001). (B) Cytotoxicity was determined in preconfluent and postconfluent Caco-2 cells treated with 1 mM and 10 mM NaB at 48 h. Cytotoxicity measured LDH release and presented as % cytotoxicity. (C) AlkP activity (µmol/mg/min) and (D) GSTA1 activity (nmol/mg/min) was determined. Values represent the mean ± S.E. of three independent experiments with six replicates each. Bars indicated by different letters differ significantly from one another (p≤0.001).</p

GSTA1 down-regulation does not affect the sensitivity of Caco-2 cells to NaB-induced apoptosis.

<p>(A) Representative western blots of GSTA1 (∼25 KDa), endogenous caspase-3 (∼35 KDa), activated caspase-3 (∼19 KDa and 17 KDa) in Caco-2 cells. Preconfluent Caco-2 cells were transiently transfected with 40 nM of GSTA1 siRNA or non-specific (NS) siRNA for 72 h and treated with NaB (10 mM) for 48 h. β-actin (∼42 KDa) was used as a protein loading control. Densitometric analysis of (B) GSTA1 levels and (C) caspase-3 activation in GSTA1 down-regulated cells with and without NaB (10 mM) treatment. Values represent the mean ± S.E of three independent experiments with three replicates each. Bars indicated by different letters differ significantly from one another (p≤0.05).</p

GSTA1 levels increase in differentiating Caco-2 cells.

<p>Preconfluent and 10 d postconfluent Caco-2 cells were assessed for: (A) protein expression of GSTA1 (∼25 KDa) and GSTP1 (∼26 KDa). β-actin was used as a protein loading control; (B) GSTA1 enzyme activity (nmol/mg/min); (C) mRNA levels of differentiation markers: AlkP, villin, DPP-4 and E-cadherin by real time RT-PCR; and (D) AlkP enzyme activity (µmol/mg/min). Values represent the mean ± S.E. of three independent experiments with three replicates each. Bars indicated by different letters differ significantly from one another (p≤0.001).</p

GSTA1 down-regulation increases the percentage of Caco-2 cells in the S phase.

<p>(A) Changes of cell cycle phase distribution in GSTA1 down-regulated Caco-2 cells as compared to controls. (B) Graphic representation of percent of cells in G1, S and G2 phase of cell cycle in non-transfected control, GSTA1 siRNA and NS siRNA transfected Caco-2 cells. Asterisks depict significant differences between control and GSTA1 down-regulated cells (*, p≤0.05; and **, p≤0.01).</p

Modulation of GSTA1 does not affect NaB-induced differentiation.

<p>Preconfluent Caco-2 cells were transiently transfected with 40 nM of GSTA1 siRNA or non-specific (NS) siRNA and after 72 h, cells were treated with 1 mM NaB for 72 h; (A) GSTA1 activity (nmol/mg/min) was determined in GSTA1 down-regulated cells. (B) AlkP activity (µmol/mg/min) was measured to determine the effect of GSTA1 down-regulation on NaB-induced differentiation. (C) Preconfluent Caco-2 cells were transiently transfected with one µg of either GSTA1-V5 or empty vector (EV) for 48 h and were treated with 1 mM NaB for 48 h. AlkP activity (µmol/mg/min) was measured to determine the effect of GSTA1 over-expression on NaB-induced differentiation. Values represent the mean ± S.E. of three independent experiments with six replicates each. Bars indicated by different letters differ significantly from one another (p≤0.001).</p

NaB (10 mM) causes GSTA1-JNK complex dissociation without activating JNK in Caco-2 cells.

<p>(A) Representative western blot of GSTA1 (∼25 KDa) and GST Pi (∼26 KDa) protein levels in GSTA1-JNK complexes. Cells were transiently transfected with GSTA1 siRNA and non-specific (NS) siRNA for 72 h and treated with 10 mM NaB. GSTA1-JNK complexes were then pulled-down from cell lysates using c-Jun fusion beads. (B) Representative western blot of phosphorylated JNK (∼54 KDa and 46 KDa) and phosphorylated p38 (∼43 KDa) protein expression in preconfluent Caco-2 cells with the treatment of 10 mM NaB. β-actin (∼42 KDa) was used as a protein loading control.</p