| Objective: The present study was performed to: (1) Investigate the activation of TLR4/NF-κB signal transduction in sepsis rats and it's influence on secretion of cytokine and organs injury. (2) Evaluate the influence of sexual dimorphism and estrogen on activation of TLR4/NF-κB signal transduction in sepsis. (3) Clarify the effect of estrogen on prevention and treatment of sepsis.Methods: Rats were subjected to lipopolysaccharide (LPS) (5mg/kg) given intraperitoneally to mimic sepsis. Firstly, one hundred male Wistar rats were randomly divided into three groups as follows: (1) normal control group (M) (n=20). (2) LPS group (ML) (n=60): being further divided respectively into lh> 21k 6h subgroups after injection of LPS. (3) LPS+estrogen group (MEL) (n=20): Rats were injected with estradiol benzoate (0.5mg/kg/d) for 5 days, then injected with LPS and 2h later, the rats were euthanized. Secondly, eighty female Wistar rats were randomly divided into four groups as follows: (1) normal control group (F) (n=20). (2) LPS group (FL) (n=20). (3) LPS+OVX group (FOL) (n=20), the rats were injected LPS after 30 days of ovariectomy. (4) LPS+OVX+ estrogen group (FOEL) (n=20), after 30 days of ovariectomized, the rats were injected with estradiol benzoate (0.5mg/kg/d), and after 5 days the rats were injected with LPS. At serial time points in each group, animals were euthanized. Blood and tissue samples from hearts, livers and lungs were harvested. The expression of TLR4/MD-2 protein, TLR4/MD-2 and TNF-α mRNAs, NF-κB activation and serum TNF-α levels were determined by Western blot analysis, reverse transcription polymerase chain reaction (RT-PCR) taking GAPDH as internal standard, electrophoretic mobility shift assay (EMSA) and enzyme-linked immunosorbent assay (ELISA).Results: 1, The regulation of TLR4/ NF-κB signal transduction pathway in multiple organs in male sepsis rats. (1) TLR4/MD-2 mRNA and protein could be detected in hearts, livers and lungs with low values in normal control group, and they were increased at 1h, 2h, 6h after sepsis, especially at 1 ~2h. (2) NF-kB activation in hearts, livers and lungs was detectable at very low levels in normal contonl, and they were increased at 1h, 2h, 6h after sepsis, especially at l-2h (P<0.01). (3) TNF-α gene and protein expression in all tissues as well as in serum samples at very low levels in normal control group and they were enhanced to peak at 1~2h(P<0.01), and decreased at 6h (P<0.01). 2, The influence of sexual difference and estrogen on activation of TLR4/ NF-κB signal transduction pathway in septic rat. The expression of TLR4/MD-2, TNF-a mRNA/protein and NF-κB activation have no difference between normal male and female control groups, but they are decreased in female sepsis group than in male sepsis group(P<0.01). All data increased significantly in FOL group than in FL group(P<0.01). There are no difference of all data between FOEL group and FOL group(P>0.05), and all the index also have no difference between MEL group and ML group(P>0.05). 3, The index of ML, FL, FOL group have significantly negative correlation with serum 17P-estradiol levels(P<0.05). Correspondingly, the index of MEL and FOEL group have negative correlation with serum 17β-estradiol levels(P<0.05), but have no significantly difference (P>0.05).Conclusions: 1, The expression of TLR4/MD-2, TNF-α mRNA and protein and NF-κB activation in hearts, livers and lungs could be markedly up-regulated after injection of LPS and lead to up-regulate the secretion of TNF-α. On the other hand, TNF-α appears to be major cytokine in up-regulation of TLR4/MD-2 mRNA expression and NF-κB activation associated with septic response, implying the interaction between TLR4/NF-κB signal transduction and expression of TNF-a during systemic inflammation. 2, There are no difference in expression of TLR4/MD-2, TNF-α mRNA and protein and NF-κB activation between normal...
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