Ghrelin restores CD4 T cell proliferation after sepsis.

<p>Mice were subjected to sham operation or CLP with vehicle (normal saline) or ghrelin (2 nm/mouse) injections at 5 and 20 h following CLP operation. Spleens were harvested at day 3 after CLP. Isolated splenocytes were labeled with CFSE (5 μM) and cultured ex vivo in anti-CD3/CD28 (1 μg/ml each) coated wells for 72 h, followed by the staining of cells with APC- anti-mouse CD4 Abs and acquision of the data using flow cytometry. The cell proliferation percentages were calculated and sham group was normalized as 100%. Data are expressed as mean ± SEM (n = 4–5 mice/group). *P < 0.05 vs. sham and ^#P < 0.05 vs. vehicle-treated animal. CLP, cecal ligation and puncture; CFSE, carboxyfluorescein succinimidyl ester; CD, cluster of differentiation.</p

Ghrelin increases AKT phosphorylation in sepsis.

<p>Mice were subjected to sham or CLP operation and treated with vehicle (normal saline) or ghrelin (2 nm/mouse) at 5 and 20 h following CLP operation. Spleens were harvested at day 3 after CLP. The levels of phosphorylated AKT were measured by Western blotting. Total AKT served as control. Data are expressed as mean ± SEM (n = 4–5 mice/group). *P < 0.05 vs. sham and ^#P < 0.05 vs. vehicle-treated animal. The dotted lines on the representative Western blot reflect the corresponding groups as shown in the bar diagram bellow. CLP, cecal ligation and puncture.</p

Ghrelin modulates the expression of cell cycle regulatory proteins in sepsis.

<p>Mice were subjected to sham or CLP operation and treated with vehicle (normal saline) or ghrelin (2 nm/mouse) at 5 and 20 h following CLP operation. Spleens were harvested at day 3 after CLP. The levels of (A) cyclin D1, (B) cyclin B1, and (C) p57 were measured by western blotting. Data are expressed as mean ± SEM (n = 4–5 mice/group). *P < 0.05 vs. sham and ^#P < 0.05 vs. vehicle-treated animal. <i>T</i>he dotted lines on the representative Western blot reflect the corresponding groups as shown in the bar diagram bellow. CLP, cecal ligation and puncture.</p

Sepsis alters the expression of cell cycle proteins in the spleen.

<p>Mice were subjected to CLP or sham operation and spleens were harvested at day 1, 2 and 3 after CLP. Total protein was extracted from spleen and the expression of (A) cyclin D1, (B) cyclin B1, and <b>(C)</b> p57 were measured by western blotting. Data are expressed as mean ± SEM (n = 4–5 mice/group). *P < 0.05 vs. sham and ^#P < 0.05 vs. day 1 after CLP. The dotted lines on the representative Western blot reflect the corresponding groups as shown in the bar diagram bellow. CLP, cecal ligation and puncture.</p

Sepsis induces the expression of apoptosis-related proteins in the spleen.

<p>Mice were subjected to CLP or sham operation and spleens were harvested at day 1, 2 and 3 after CLP. Total proteins were extracted from spleen and the expression of (A) Fas, (B) Fas-L and (C) cleaved caspase-8 were measured by western blotting. Data are expressed as mean ± SEM (n = 4–5 mice/group). *P < 0.05 vs. sham and ^#P < 0.05 vs. day 1 after CLP. The dotted lines on the representative Western blot reflect the corresponding groups as shown in the bar diagram bellow. CLP, cecal ligation and puncture.</p

Sepsis decreases the frequencies of CD4 T cells.

<p>Mice were subjected to CLP or sham operation and spleens were harvested at day 1, 2 and 3 after CLP. Isolated splenocytes were stained for APC- anti-mouse CD4 Abs. (A) The percentages of CD4 T cells were assessed by flow cytometry and (B) total CD4 T cells in the spleen were calculated by multiplying total splenic cells. Data are expressed as mean ± SEM (n = 4–5 mice/group). *P < 0.05 vs. sham mice. CLP, cecal ligation and puncture; CD, cluster of differentiation.</p

Sepsis impairs the proliferation of CD4 T cells <i>ex vivo</i>.

<p>Mice were subjected to CLP or sham operation and spleens were harvested at day 1, 2 and 3 after CLP. Isolated splenocytes were labeled with CFSE (5 μM) and stained with APC- anti-mouse CD4 Abs after the culture. The frequencies of proliferated cells were calculated and the sham group was normalized as 100%. Data are expressed as mean ± SEM (n = 4–5 mice/group). *P < 0.05 vs. sham and ^#P < 0.05 vs. day 1 after CLP. CLP, cecal ligation and puncture; CFSE, carboxyfluorescein succinimidyl ester; CD, cluster of differentiation.</p

Ghrelin prevents the loss of CD4 T cells after sepsis.

<p>Mice were subjected to sham operation or CLP with vehicle (normal saline) or ghrelin (2 nm/mouse) injections at 5 and 20 h after CLP. Spleen was harvested at day 3 after CLP. Isolated splenocytes were stained for APC- anti-mouse CD4 Abs. (A) The percentages of CD4 T cells in the splenocytes were assessed by flow cytometry and (B) total CD4 T cells were calculated by multiplying total splenic cells. Data are expressed as mean ± SEM (n = 4–5 mice/group). *P < 0.05 vs. sham mice. CLP, cecal ligation and puncture.</p