| Objective: The present study was conducted to observe the activation of Janus kinase (JAK) 2 and signal transducers and activators of transcription (STAT)-3 in postburn Staphylococcus aureus infection, and the potential effect of treatment with JAK/STAT inhibitors on the development of postburn sepsis. In addition, the influence of mitogen-activated protein kinase (MAPK) and nuclear factor-(B (NF-(B) inhibitors on STAT3 activation was investigated to elucidate the possible crosstalk among these signal transduction pathway in vivo.Material and methods: 97 male Wistar rats were randomly divided into eight groups as follows: (1) normal control group (n=6); (2) scald control group (n=9), rats were subjected to a 20% total body surface area (TBSA) III( scald injury; (3) postburn sepsis group (n=36), rats inflicted with 20% TBSA III( scald followed by SEB-produced Staphylococcus aureus challenge were further divided into 0.5, 1, 2, 6, 12, 24 hours groups, respectively; (4)AG490 (inhibitor of JAK2) treatment group (n=10), intraperitoneal injection of a dose of 8mg/kg AG490 just prior to Staphylococcus aureus challenge, being further divided into 0.5 and 2 hours groups, respectively; (5) RPM (rapamycin, inhibitor of STAT3) treatment group (n=10), intraperitoneal injection of a dose of 0.4 mg/kg prior to Staphylococcus aureus challenge, being further divided into 0.5 and 2 hours groups, respectively. (6) AG126 (inhibitor of MAPK) treatment group (n=10), intraperitoneal injection of a dose of 4mg/kg prior to Staphylococcus aureus challenge, being further divided into 0.5 and 2 hours groups, respectively. (7) PDTC (pyrrolidine dithiocarbamate, inhibitor of NF-(B) treatment group (n=10), intraperitoneal injection of a dose of 50mg/kg prior to Staphylococcus aureus challenge, being further divided into 0.5 and 2 hours groups, respectively. (8) DMSO (dimethyl sulfoxide, solvent of drugs)treatment group (n=6), intraperitoneal injection of a dose of 4ml/kg prior to Staphylococcus aureus challenge. Blood and tissue samples from liver, kidneys and lungs were collected to determine transcription factor, cytokines, as well as organ functional indices. The STAT3 activation was determined by the Electrophoretic Mobility Shift Assay (EMSA). TNF-(, IFN-( and IL-10 protein levels were determined by enzyme-linked immunosorbent assay (ELISA). TNF-α, IFN-α and IL-10 mRNA expression were semi-quantitatively measured by the reverse transcription polymerase chain reaction (RT-PCR) using GADPH as an internal standard. Results: 1. Weak expression of TNF-α, IFN-α and IL-10 mRNA was detected in liver, lungs and kidneys in normal controls, with no marked changes in scald controls at 24 hours. After thermal injury combined with Staphylococcus aureus infection, TNF-(, IFN-( and IL-10 mRNA in liver, lungs and kidneys rapidly increased at 1 hour, peaking at 1-2 hours, and were markedly higher than these in normal or scald controls (P<0.05). Meanwhile, TNF-α, IFN-α and IL-10 protein levels also rapidly increased at 0.5 hour, peaking at 1-6 hours and returning to baseline values 12-24 hours later. Significant positive correlations were found between IL-10/IL-10 mRNA in liver, TNF-(/TNF-( mRNA as well as IFN-(/IFN-( mRNA in lungs, and IFN-(/IFN-( mRNA as well as IL-10/IL-10 mRNA in kidneys. 2. Activation of STAT3 was weak in normal and scald controls, while it was quickly up-regulated at 0.5-1 hour following thermal injury combined with Staphylococcus aureus infection, recovering to normal range at 2 hours in various tissues. But STAT3 was reactivated after 12 hours in liver. Treatment with both AG490 and RPM could inhibit activation of STAT3 at 0.5 hour in liver, lungs and kidneys. PDTC treatment could also inhibit activation of STAT3 at 0.5 hour in liver and lungs. AG126 treatment had no marked effect on activation of STAT3 at various intervals. 3. TNF-(, IFN-( and IL-10 mRNA expression in the liverwere significantly lower in AG490 treated group than postburn sepsis group at 2 hours, with decreases in proteins levels by 29.5(... |
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