Hypertonic saline resuscitation contributes to early accumulation of circulating myeloid-derived suppressor cells in a rat model of hemorrhagic shock

Background Hemorrhagic shock is usually associated with complicated immune and inflammatory responses, which are sometimes crucial for the prognosis. As regulators of the immune and inflammatory system; proliferation, migration, distribution and activation of myeloid-derived suppressor cells （MDSCs） are intimately linked to the inflammation cascade. Methods In a model of severe hemorrhagic shock, thirty-five rats were randomly divided into control, sham, normal saline resuscitation （NS）, hypertonic saline resuscitation （HTS）, and hydroxyethyl starch resuscitation （HES）, with seven in each group. MDSCs were analyzed by flow cytometric staining of CD11b/c＊Gra~ in peripheral blood mononuclear cells （PBMC）, spleen cell suspensions, and bone marrow nucleated cells （BMNC）. Simultaneously, the expressions of arginase-1 （ARG-1） and inducible nitric oxide synthase （iNOS） mRNA in MDSCs were evaluated by quantitative reverse transcription-polymerase chain reaction （qRT-PCR）. Results In the early stage after hemorrhagic shock, fluid resuscitation and emergency treatment, the MDSCs in the PBMC of NS, HTS and HES groups markedly increased, and MDSCs in BMNC of these groups decreased accordingly, significantly different to the control group. In hemorrhagic shock rats infused with HTS at the early resuscitation stage, MDSCs in PBMC increased about 2 and 4 folds, and MDSCs in BMNC decreased about 1.3 and 1.6 folds, as compared to the sham group respectively, with statistically significant difference. Furthermore, compared to the NS and HES groups, the MDSCs in PBMC of HTS group increased 1.6 and 1.8 folds with statistically significant differences; the MDSCs decrease in BMNC was not significant. However, there was no statistically significant difference in MDSCs of spleen among the five groups. In addition, compared to the control, sham, NS and HES groups, the ARG-1 and iNOS mRNA of MDSCs in PBMC, spleen and BMNC in the HTS group had the highest level of expression, but no statistically significant differences were noted. Conclusions In this model of rat with severe and controlled hemorrhagic shock, small volume resuscitation with HTS contributes to dramatically early migration and redistribution of MDSCs from bone marrow to peripheral circulation, compared to resuscitation with NS or HES.

Early changes of CD4＋CD25＋Foxp3＋ regulatory T cells and Th1/Th2, Tc1/Tc2 profiles in the peripheral blood of rats with controlled hemorrhagic shock and no fluid resuscitation

Background Hemorrhagic shock induces immune dysfunction. Regulatory T cells （Tregs）, T-helper （Th） cells, and cytotoxic T-lymphocytes （CTLs） can execute many crucial actions in immune and inflammatory responses. This study was conducted to investigate the early pathophysiological changes of CD4＋CD25＋Foxp3＋ Treg and Th1/Th2, Tc1/Tc2 profiles in the peripheral blood of rats with controlled hemorrhagic shock and no fluid resuscitation. Methods A rat model of controlled hemorrhagic shock with no fluid resuscitation was established. Peripheral blood samples were taken before and four hours after hemorrhagic shock with no fluid resuscitation. Three color flow cytometry was used to detect Tregs, Thl, Th2, Tcl and Tc2 ceils in the samples. Results In the peripheral blood of rats, the percentage of Tregs four hours after hemorrhagic shock was significantly lower than before hemorrhagic shock （P=0.001）. The ratios of Th1/Th2 and Tc1/Tc2 were changed from （23.08±8.98）% to （23.91±15.36）%, and from （40.40±21.56）% to （65.48±23.88）%, respectively. Conclusions At an early stage, the advent of hemorrhagic shock is related to an early decrease of Tregs, and a mild shift in the Th1/Th2, Tc1/Tc2 balance toward Thl and Tcl dominance. These changes are part of a hyper-inflammatory state of the host, and will deteriorate the maintenance of immune balance. Further influences and detailed mechanisms need to be investigated.

Effect of hypertonic saline resuscitation on heme oxygenase-1 mRNA expression and apoptosis of the intestinal mucosa in a rat model of hemorrhagic shock

Background Massive blood loss due to trauma is the leading cause of death in trauma patients and military combatants. The fluid category of resuscitation for hypotensive trauma patients is open to debate. This study was conducted to investigate the early effects of hypertonic and isotonic saline solutions on heme oxygenase-1 （HO-1） mRNA expression and apoptosis in the intestinal mucosa of rats with hemorrhagic shock. Methods A model of severe hemorrhagic shock was established in 21 Sprague-Dawley rats. The rats were randomly divided into sham, normal saline resuscitation （NS）, and hypertonic saline resuscitation （HTS） groups, with 7 in each group. We assessed and compared the HO-1 mRNA expression and apoptosis in the small intestinal mucosa of rats after hemorrhagic shock and resuscitation using the SYBR Green I fluorescence quantitative reverse transcriptase polymerase chain reaction, fluorescein-iso-thiocyanate-annexin V/propidium iodide double staining, and flow cytometry. Results In the early stage of hemorrhagic shock and resuscitation, marked apoptosis occurred in the small intestinal mucosa from both the NS and HTS groups. The apoptotic rate in the NS group was higher than that in the HTS group （P 〈0.01）. Among the three groups, HO-1 mRNA mucosa from the HTS group had the highest level of expression; however, the differences were not significant. There was a significant negative correlation between HO-1 mRNA expression and apoptosis in the small intestinal mucosa from the NS and HTS groups after hemorrhagic shock and resuscitation. Conclusions In this rat model of severe hemorrhagic shock, HTS resuscitation with a small volume is more effective than NS resuscitation in reducing apoptosis of the intestinal mucosa. Further, HO-1 mRNA over-expression in the intestinal mucosa may be one of the molecular mechanisms of HTS in the resuscitation of hemorrhagic shock.