| Objective: The present study was performed to (1) investigate the activation of Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway and its effect on mRNA expression of high mobility group box-1(HMGB1)protein in sepsis induced by cecal ligation and puncture (CLP); (2) primarily study the potential mechanism underlying signal regulation of HMGB1 in rats with sepsis; (3) clarify the relationship between activation of JAK/STAT pathway (STAT1/STAT3) and HMGB1 expression, and their role in the pathogenesis of CLP-induced sepsis and subsequent multiple organ dysfunction syndrome (MODS).Material and methods: 140 male Wistar rats were randomly divided into six groups as follows: (1) normal control group (n=10); (2) sham operation group (n=10); (3) CLP group (n=60): being further divided respectively into 0.5h, 2h, 6h, 12h, 24h and 48h subgroups post-CLP; (4) AG490 treatment group (n=24): subcutaneous injection of AG490 at a dose of 8.0mg/kg at 20mins prior to CLP, being further divided respectively into 2h, 6h, 24h and 48h post-CLP subgroups; (5) RPM treatment group (n=24): subcutaneous injection of RPM at a dose of 0.4mg/kg at 20mins prior to CLP, being further divided respectively into 2h, 6h, 24h, 48h post-CLP subgroups; (6)DMSO treatment group (n=12): subcutaneous injection of DMSO at a dose of 4.0mg/kg at 20mins before CLP, being further divided respectively into 2h and 24h post- sham operation subgroups; At serial time points in each group, animals were sacrificed, and blood and tissue samples from liver, lungs, kidneys and small intestine were harvested. HMGB1 mRNA expression was semi-quantitative determined by the reverse transcription polymerase chain reaction (RT-PCR) taking GAPDH as an internal standard. The major organ functional indices—serum AST, ALT, BUN and Cr contents, were measured with automatic biochemistry analyzer. Activity of STAT1/3 was assayed by electrophoretic mobility shift assay (EMSA).Results: 1. The activation of STAT1 and STAT3 in rats with sepsis and the effect of pretreatment with AG490 and RPM on it There were no or very low activation of STAT1 and STAT3 DNA binding activities in normal controls, and low levels of STAT1 and STAT3 DNA binding activities were detected in sham operation and DMSO groups. STAT1 DNA binding activity in liver, lungs, small intestine and kidneys could be detected early at 0.5h after CLP, peaking at 6h~24h. STAT3 DNA binding activities were detected only in liver and lungs following CLP, while their values were relatively lower compared with STAT1. No marked activation of STAT3 could be observed in small intestine and kidneys at various intervals. Pretreatment with AG490 and RPM reduced both STAT1 and STAT3 DNA binding activities in liver, lungs, as well as small intestine with certain extent at 6, 24 and 48h after CLP respectively, but AG490 treatment only inhibited activation of STAT1 in kidneys at 48h following CLP. 2. (1) The kinetic changes in HMGB1 mRNA expression in CLP rats HMGB1 mRNA expression could be detected in liver, lungs, small intestine and kidneys in normal controls, and it did not markedly elevate in sham group. HMGB1 mRNA in hepatic tissue significantly increased at 6h following CLP, and maintained at relatively high levels up to 48h. It increased rapidly in lungs with a peak value at 2h, and tended to near normal range at 48h. In small intestine, it didn't elevate during early phase after CLP, and increased significantly only at 48h. No marked changes in HMGB1 mRNA expression were found in kidneys throughout the observation period. (2) The effect of pretreatment with AG490 on HMGB1 mRNA expression Compared with corresponding time points in CLP group, AG490 treatment could significantly inhibit the expression of HMGB1 mRNA in liver and small intestine at 24 and 48h, in pulmonary tissue at 2h, but without marked changes in kidneys. (3) The effect of pretreatment with RPM on HMGB1 mRNA expression Compared with corresponding time points in CLP group, pretreatment with RPM significantly reduced the ex...
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