Immune recovery after fluid resuscitation in rats with severe hemorrhagic shock

Abstract

Objective: To investigate the effects of resuscitation with normal saline (NS), hypertonic saline (HTS), and hydroxyethyl starch (HES) on regulatory T cells (Tregs), helper T 1 (Th1)/Th2 and cytotoxic T 1 (Tc1)/Tc2 profiles in the treatment of hemorrhagic shock.

Methods: Rats subjected to severe hemorrhagic shock were resuscitated for 30 min with NS (n=8), HTS (n=8), or HES (n=8); sham (n=8) and naive control (n=8) groups were used for comparison. Following fluid resuscitation, the whole shed blood was reinfused for 30 min, and the rats were observed with continuous hemodynamic monitoring for 120 min. CD4⁺CD25⁺Foxp3⁺ Treg proportions, Th1/Th2 and Tc1/Tc2 profiles in spleen were analyzed by three-color flow cytometry.

Results: The proportion of CD4⁺CD25⁺Foxp3⁺ Tregs and ratios of Th1/Th2 and Tc1/Tc2 did not differ among control, sham, and HTS groups, but were significantly lower in NS and HES groups (both P<0.05 vs. sham); NS and HES levels were similar. The level of Tc1 was significantly increased in HTS (P<0.05 vs. sham), and levels of Tc2 were increased in NS, HES, and HTS groups compared to sham (all P<0.05), but did not differ from each other.

Conclusions: HTS resuscitation has a greater impact on immune system recovery than NS or HES by preserving the proportion of Tregs and maintaining the balance between Th1/Th2 and Tc1/Tc2 cells in the spleen. Thus, HTS resuscitation provides potential immunomodulatory activity in the early stage after hemorrhagic shock.

Keywords: Regulatory T cells; Helper T cells; Cytotoxic T cells; Hemorrhagic shock.

Fig. 1

Mean arterial pressure (MAP) monitoring Sham: sham group (n=8); NS: normal saline group (n=8); HTS: hypertonic saline group (n=8); HES: hydroxyethyl starch group (n=8). Data are represented as mean±SD

Fig. 2

Representative illustration of flow cytometry for Tregs (a) Upper right quadrant represents CD4⁺CD25⁺ cells. (b) Upper right quadrant represents CD4⁺CD25⁺Foxp3⁺ Tregs

Fig. 3

Representative illustration of flow cytometry for Th and Tc cells (a) Upper right quadrants represent CD3⁺interferon (IFN)-γ⁺ (top) and CD3⁺interleukin (IL)-4⁺ (bottom) cells, respectively. (b) Upper right quadrant represents CD3⁺IFN-γ⁺ CD8⁺ Tc1 cells and lower right quadrant represents CD3⁺IFN-γ⁺ CD8⁻ Th1 cells (top); upper right quadrant represents CD3⁺IL-4⁺CD8⁺ Tc2 cells and lower right quadrant represents CD3⁺IL-4⁺CD8⁻ Th2 cells (bottom)

Fig. 4

Percentages of Th and Tc in spleen Control: control group (n=8); Sham: sham group (n=8); NS: normal saline group (n=8); HTS: hypertonic saline group (n=8); HES: hydroxyethyl starch group (n=8). Data are represented as mean±SD. ^* P<0.05, vs. control; ^∆ P<0.05, vs. sham by one-way ANOVA and the least significant difference t-test