ABSTRACT
ObjectiveFrom the changes of expression of cold-inducible RNA-binding protein (CIRP) in rats with traumatic brain injury under mild hypothermia treatment with Shenfu decoction as a subsidiary, to speculate the mechanism of protective effect of the decoction on the injury.Methods Ninety Sprague-Dawley (SD) rats were divided into three groups by random number table: non-transfection control group, adenovirus mediated immune flourescent reverse transcription virus group (blank AD5-GFP transfection group) and adenovirus mediated immune flourescent reverse transcription virus carrying CIRP silent expression gene group (AD5-GFP-CIRP-SiRNA transfection group), 30 rats in each groups. Then, each group was subdivided into three subgroups: model group, traditional Chinese medicine (TCM) low and high dose groups, 10 rats in each subgroup. After the mild hypothermia treatment for 48 hours, in the TCM low dose group and high dose group, a dose of TCM 1 mL/kg and 5 mL/kg was injected via a tail vein into the rat respectively, while in the model group, 1 mL/kg normal saline was injected into the same vein, once a day for consecutive 2 days in all the groups. Before modeling in the blank AD5-GFP transfection group and AD5-GFP-CIRP-SiRNA transfection group, virus transfection models were reproduced at first by one-time intrathecal injection of 0.1 mL AD5-GFP and 0.1 mL (1×1010 pfu/mL) AD5-GFP-CIRP-SiRNA virus vector respectively, and in model group, 0.1 mL normal saline was given. The rat cortex, hippocampus and hypothalamus part were collected, the brain cell apoptosis was detected by transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL), the CIRP mRNA expression in the cortex, hippocampus and hypothalamus part was measured by reverse transcription-polymerase chain reaction (RT-PCR), the protein expressions of rat sarcoma protein Raf, Ras, extracellular signal-regulated kinase (ERK), phosphorylation ERK (p-ERK), mitogen activated protein kinase (MEK), p-MEK were determined by Western Blot.Results The brain tissue cell apoptosis indexes (AI) in the cortex, hippocampus and hypothalamus part in TCM low, high dose group of non-transfection control and blank AD5-GFP transfection group were lower than those in model group, and the expressions of CIRP mRNA were higher than those in model group, there were no significant differences in AI and CIRP mRNA in the cortex, hippocampus and hypothalamus between model, TCM low and high dose groups of AD5-GFP-CIRP-SiRNA transfection group, but AI was significantly higher and CIRP mRNA was significantly lower than that in corresponding subgroups of AD5-GFP transfection control group and blank AD5-GFP transfection group. Western Blot detection showed that: Raf/Ras, p-MEK/MEK protein expressions revealed no statistical significant differences in different parts of each group (allP > 0.05), the p-ERK/ERK protein expression in the cortex, hippocampus, and hypothalamus part was significantly lower in TCM low and high dose group than that in the model group of non-transfection control group and blank AD5-GFP transfection group, the degree of descent in the TCM high dose group being more significant (the cortex: non-transfection control group was 7.2±1.0 vs. 15.3±1.8, AD5-GFP transfection group was 8.1±0.7 vs. 16.2±1.5; hippocampus part: non-transfection control group was 6.6±0.8 vs. 14.7±2.0, AD5-GFP transfection group was 6.8±1.0 vs. 14.9±1.3; hypothalamus part: non-transfection control group was 9.4±1.1 vs. 12.7±1.7, AD5-GFP transfection group was 10.6±1.3 vs. 9.4±1.1, allP 0.05).Conclusions The Shenfu decoction used in rats with brain trauma under treatment of mild hypothermia is possibly by promoting CIRP over-expression, lowering ERK expression and inhibiting the initiation of signal transduction of the secondary transcription factor phosphorylation, thereby the neural cell apoptosis is decreased and play a subsidiary role of anti-apoptosis of mild hypothermia.
ABSTRACT
The quorum sensing [QS] of pathogens has been found to affect their biofilm forming ability, making it a potential target for anti-microbial therapy. The present research aimed to evaluate the anti-QS activities of different extracts and isolated phytochemicals from Liriodendron hybrid barks and their roles in the inhibition of the growth and biofilm formation of methicillin-resistant Staphylococcus aureus [MRSA]. The assays on the inhibition of QS by the five extracts [n-hexane, dichloromethane, ethyl acetate, acetone, and methanol] and eight isolated compounds were carried out by using both the indicator strains Chromobacrerium violaceum CV026 and C. violaceum ATCC12472. The in vitroeffects of the five extracts and eight isolated compounds on MRSA biofilm were also preliminarily evaluated using crystal violet micro titer plate assays. The results suggested that the dichloromethane extract showed anti-QS and MRSA biofilm inhibitory activities and the n-hexane extract possessed only MRSA biofilm inhibitory effect. The dichloromethane extract could serve as a source for developing bacterial intervention strategies targeting microbial QS system. All eight isolated compounds showed no anti-QS and biofilm formation inhibiting activities. So further researches are still being required to purify and identify the compounds possessing anti-QS and biofilm inhibitory effects from the dichloromethane and n-hexane extracts