Potential role of mesenchymal stem cells in alleviating intestinal ischemia/reperfusion impairment.

BACKGROUND: Transplantation of bone marrow mesenchymal stem cells (MSCs) provides a promising therapeutic efficiency for a variety of disorders caused by ischemia or reperfusion impairment. We have previously demonstrated the efficacy of MSCs in mitigating intestinal ischemia/reperfusion (I/R) injuries in rats, but the mechanism by which MSCs engraft ameliorates I/R injuries has largely been unknown. The present study aimed at investigating probable mechanisms by which MSCs exert their function. METHODS: Male donor derived rat MSCs were implanted into intestine of female recipient rat by direct submucosal injection after superior mesenteric artery clamping and unclamping. The homed MSCs were detected by Y chromosome in situ hybridization probe, and the tumor necrosis factor-α (TNF-α) content in intestinal mucosa was determined by ELISA. Expression of proliferative cell nuclear antigen (PCNA) in bowel mucosa was assayed by real-time PCR and intestinal mucosa expression of phosphorylation extracellular signal-regulated kinase (pERK1/2) and nuclear factor-κB (NF-κB) were evaluated by western blot. RESULTS: Four and seven days after MSCs transplantation, the TNF-α content of bowel mucosa in MSCs group was significantly lower than that in saline group. The PCNA in bowel mucosa showed higher expression in MSCs treated group than the saline group, both at 4 and 7 days after cell transplantation. The expression of intestinal mucosal pERK1/2 in MSCs treated group was markedly higher than that in saline group, and the expression of NF-κB in MSCs treated group was noticeably decreased than that in saline group at 4 and 7 days post MSCs transplantation. CONCLUSION: The present investigation provides novel evidence that MSCs have the potential to reduce intestinal I/R injuries probably due to their ability to accelerate cell proliferation and decrease the inflammatory response within intestinal mucosa after ischemia and reperfusion

Mucosal NF-κB expression in each group at 4 and 7 days after MSCs administered.

<p>(<b>A</b>) Semiquantitative assessment of bands for NF-κB and its internal control protein. (<b>B</b>) Values of the NF-κB band performed densitometrically using the software Quantity One. Each bar represents the mean ± SD of the different groups; values were compared by one-way ANOVA test, <i>**: P<0.01</i>, <i>*: P<0.05</i>. S: sham group; C: control (saline) group; M: MSCs group. 4d and 7d represent 4 days and 7 days postoperatively.</p

Real-Time PCR analysis of PCNA expression in the intestinal mucosa.

<p>Data are means ± SD of each group at different time point and expressed as the ratio of original value mRNA of PCNA to its internal control mRNA of GAPDH. S: sham group; C: control (saline) group; M: MSCs group. 4d and 7d represent 4 days and 7 days postoperatively. ** <i>P<0.01</i>, <i>* P<0.05</i>, compared with control (saline) group, by one-way ANOVA test.</p

ELISA analysis of intestinal mucosal TNF-α content among the three groups.

<p>Data are shown as means ± SD of the three groups, <i>*P < 0.01</i> vs. MSC group; # <i>P <0.05</i> vs. Sham group, values are evaluated by one-way ANOVA test.</p

Expression of pERK1/2 in intestinal mucosa at 4 and 7 days after MSCs transplantation.

<p>(<b>A</b>) Bands of the expression of pERK1/2 and its own tERk1/2 protein were detected by western blot. (<b>B</b>) The ratio value of pERK1/2 to its own tERK1/2 of the bands which was evaluated densitometrically using the software Quantity One for the three groups at different time points. Each bar represents mean ± SD of the ratio value in every group. S: sham group; C: control (saline) group; M: MSCs group. 4d and 7d represent 4 days and 7 days postoperatively. *<i>: P<0.05</i>, compared with control (saline) group, by one-way ANOVA test.</p

MSCs homing photograph (40×).

<p>After cell transplantation, engrafted cells were detected by Y chromosome <i>in </i><i>situ</i> hybridization at 1, 4, 7, and 10 days (the green fluorenscent point represents positive donor derived cells), which corresponded to graphs A, B, C, and D respectively. Most of the homed cells were located at mucosal lamina propria, and the amounts of engrafted cells were more at 4 and 7days than 1 and 10 days postoperatively.</p