Relationship between lymphocyte apoptosis and endotoxin translocation after thermal injury in rats.

AIM

To investigate the relationship between lymphocyte apoptosis in peripheral blood, spleen and mesenteric lymph nodes(MLN) and endotoxin translocation after thermal injury in rats.

METHODS

In a Wistar rat model inflicted with 30% TBSA III degree scalding, serum LPS levels in portal vein and vena cava were quantified by tachypleus amebocyte lysate (TAL) technique. The analysis of peripheral blood lymphocyte was employed in in situ Cell Death Detection Kit and evaluated by flow cytometry. Apoptotic lymphocytes in paraffin-embedded spleen and MLN sections were examined by histologic analysis, in situ deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) and peroxidase (POD) staining. The images were taken by Cooldccd camera system, and the count and optical density value (transmission light) of apoptotic lymphocytes were analyzed with software Spot and Imagine proplus 4.10a(IPP4.10a).

RESULTS

In the period of 3 to 48 postburn hours (PBHs) serum LPS level (x10(3) EU.L(-1)) in portal vein (2.11+/-0.02, 5.66+/-0.20, 3.70+/-0.22, 2.56+/-0.28, 0.90+/-0.11) was higher than that in vena cava (0.63+/-0.01, 1.53+/-0.18, 0.83+/-0.32, 0.52+/-0.12, 0.23+/-0.02, P<0.01), but both increased sharply in postburn rats (P<0.01) and reached a peak at 6 PBH. Analysis of apoptotic lymphocytes showed that the proportion (%) of postburn apoptotic cells was much higher than that in healthy rats (8.34+/-1.53, 8.13+/-1.81, 20.77+/-3.94, 23.90+/-3.92, 11.23+/-1.35 and 13.26+/-2.09 at 3, 6, 12, 24, 48 and 72 PBH, respectively, vs 3.99+/-1.72, P<0.01), especially after 6 PBH. The concentrations of lymphocytic apoptosis at 12 and 24 PBH were markedly higher than that at other time points. Meantime, few apoptotic lymphocytes were found in normal MLN, but increased postburn obviously (3+/-1 vs 546+/-83, 285+/-39, 149+/-30, 58+/-10, 36+/-11 and 33+/-9 in turn, P<0.01), especially at 3 PBH, whereas apoptotic lymphocytes were concentrated in splenic cortex before the burn and decreased obviously during 72 PBHs (499+/-186 vs 12+/-8, 19+/-15, 12+/-7, 100+/-15, 123+/-25 and 226+/-26 in turn, P<0.01) though a slight rise was found in the medulla after 24 PBH. Optical density of apoptotic lymphocytes was significantly reduced in spleen in the 24 PBHs and raised in MLN during 48 PBHs than that prior to the burn, respectively.

CONCLUSION

Gut-origin LPS is a major cause of endotoxemia taken place early in rats following severe thermal injury and could induce extensive lymphocyte apoptosis in blood and MLN, which suggests an immunosuppression state could follow the initial injury and favores a septic state based on apoptotic mechanism.