Resveratrol mitigates lipopolysaccharide-mediated acute. inflammation in rats by inhibiting the TLR4/NF-κBp65/MAPKs.

Transcription

1 Resveratrol mitigates lipopolysaccharide-mediated acute inflammation in rats by inhibiting the TLR4/NF-κBp65/MAPKs signaling cascade Guangxi Wang a,1, Zhiqiang Hu b,1, Qiuting Fu a,1, Xu Song a,1, Qiankun Cui a, Renyong Jia b*, Yuanfeng Zou a, Changliang He a, Lixia Li a, Zhongqiong Yin a, 1*. anatural Medicine Research Center, College of Veterinary Medic ine, Sichuan Agricultural Univer sity, Chengdu, , China b Key laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Chengdu ,China 1 These authors contribute equally to this work and should be considered as the first author. * Corresponding author: yinzhongqiong Postal address:natural Medicine Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Chengdu, , PR China Tel: ; Fax: address: yinzhongq@163.com (Z.Q. Yin). Ren-Yong Jia, address: cqrc_jry@163.com

2 Supplementary Table S1 The effects of RSV on hematologic determination induced by LPS. Items Blank control LPS DXM+LPS RSV(h)+LPS RSV(m)+LPS RSV(l)+LPS WBC(10 9 /L) ± ±1.083 ** ±2.665 ## 8.700±3.431 ## 9.650±4.003 ## 7.317±2.515 ## RBC(10 12 /L) 6.642± ±1.273 * 6.908±1.307 # 6.913±1.659 # 7.037±1.071 # 6.533±0.665 # HGB ± ±30.22 ** ±27.24 ## ±33.40 ## ±20.90 ## ±13.26 ## PLT(10 9 /L) ± ± * ± ## ± ## ± ## ± # LYM(10 9 /L) 7.833± ±1.432 ** 8.650±1.699 ## 6.300±2.983 ## 7.367±3.222 ## 5.733±2.181 ## GRA(10 9 /L) 3.017± ±0.232 ** 2.733±0.987 ## 2.100±0.890 ## 2.017±0.801 ## 1.367±0.314 # MON(10 9 /L) 0.350± ±0.041 ** 0.383±0.117 ## 0.300±0.110 ## 0.267±0.103 ## 0.217±0.075 ## HCT ± ± ± ± ± ±3.990 MCV ± ± ± ± ± ±1.496 MPV(fl) 5.817± ± ± ± ± ±0.308 MCH ± ± ± ± ± ±0.553 WBC: white blood cell count; RBC: red blood cell count; HGB: hemoglobin concentration; PLT: platelet; GRA: granulocyte; LYM: lymphocyte count; MPV: the average platelet volume; HCT: hematokrit; MCV: mean corpuscular volume; MCH: mean corpuscular hemoglobin and MON: monocytes. DXM+LPS, RSV(h)+LPS, RSV(m)+LPS and RSV(l)+LPS represent the groups which pretreatment with DXM, high, intermediate and low dose of resveratrol before giving them LPS-stimulus, respectively. The values are presented as means±standard deviation (10 rats /group). * P<0.05, ** P<0.01 vs. Blank control group; # P<0.05, ## P<0.01 vs. LPS group.

3 Supplementary Table S2 The effects of RSV on blood biochemical indicators induced by LPS. Groups ALT(U/L) AST(U/L) ALP(U/L) BUN(mmol/L) CRE(µmol/L) CHO(mmol/L) Blank control 48.17± ± ± ± ± ± LPS 55.33±2.42 * ±38.59 * ± * ± ± ± DXM+LPS 49.50±5.32 # ±37.40 # ±86.30 # ± ± ± RSV(h)+LPS 43.83±9.56 # ±69.33 ## ±56.89 # ± ± ± RSV(m)+LPS 49.67±11.73 # ±50.03 # ±70.11 # ± ± ± RSV(l)+LPS 43.50±13.13 # ±60.76 ## ± # ± ± ± ALT: alanine aminotransferase; AST: aspartate transaminase; ALP: alkaline phosphatase; BUN: blood urea nitrogen; CRE: Creatinine; GLU: glucose; CHO: total cholesterol. DXM+LPS, RSV(h)+LPS, RSV(m)+LPS and RSV(l)+LPS represent the groups which pretreatment with DXM, high, intermediate and low dose of resveratrol before giving them LPS-stimulus, respectively. The values are presented as means±standard deviation (10 rats/group). * P<0.05, ** P<0.01 vs. Blank control group; # P<0.05, ## P<0.01 vs. LPS group.

4 Supplementary Table S3 Primer sequences used in RT-PCR analysis. Primer Primer sequences Fragment size (5 >3 ) (forward/reversed) Length(bp) MyD88 TRAF6 IκBα P38 ERK5 ERK1 ERK2 JNK β-actin F-5 TTGCTAGCCTTGTTAGACCGT3 R-5 CTCCTGTTTCTGCTGGTTGCG3 F-5 ACCATCAAATCCGGGAGC3 R-5 TGCCAATCTTCCAAATGTAAATGC3 F-5 CCACCAACTACAACGGCCACAC3 R-5 TTGACATCAGCACCCAAAGTCAC3 F-5 AGTGGCTGACCCTTATGAC3 R-5 CACAGTGAAGTGGGATGGA3 F-5 GGTTGCCTGGTTCTGGT3 R-5 TTGGACAGTTGCTGGGT3 F-5 CTACACGCAGCTGCAGTACATC-3 R-5 GTGCGCTGACAGTAGGTTTGA-3 F-5 GGCACCAACCATTGAGCAGA-3 R-5 -GATCATTGCTGAGGTGCTGTGTC-3 F-5 AGTGTAGAGTGGATGCATGA3 R-5 ATGTGCTTCCTGTGGTTTAC3 F-5 GGAGATTACTGCCCTGGCTCCTAGC3 R-5 GGCCGGACTCATCGTACTCCTGCTT

5 Supplementary Figure S1 Chemical structure of trans-rsv (C 14 H 12 O 3 ).

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7 Supplementary Figure S2 Densitometric analysis of proteins and phosphorylated proteins in liver s and lung s cytoplasm and nucleus. : Mean density of related protein in TLR4 signaling pathway; : Mean density of related proteins and phosphorylated proteins in NF-κBp65 signaling pathway; : Mean density of related proteins and phosphorylated proteins in MAPKs signaling pathway. a to e : Mean density of proteins and phosphorylated proteins in liver s

8 cytoplasm and nucleus; f to j: Mean density of proteins and phosphorylated proteins in lung s cytoplasm and nucleus. DXM+LPS, RSV(h)+LPS, RSV(m)+LPS and RSV(l)+LPS represent the groups which pretreatment with DXM, high, intermediate and low dose of resveratrol before giving them LPS-stimulus, respectively. The values are presented as means ± standard deviation (10 rats /group). * P<0.05, ** P<0.01 vs. Blank control group; # P<0.05, ## P<0.01 vs. LPS group.