Mindin is upregulated during colitis and may activate NF-κB in a TLR-9 mediated manner

AIM:To investigate the regulation of mindin expression and the signaling pathway involved during inflammation.METHODS:C57BL/6 mice were treated with 3% dextran sulfate sodium (DSS) in drinking water for 6 d to induce acute colitis,and then the colon was harvested for histological analysis or for RNA isolation.mRNA expression of mindin and nuclear factor (NF)-κB p65 was analyzed by quantitative real time polymerase chain reaction (RT-PCR) and mindin expression construct was conf irmed by Western blotting.Mouse macrophage and intestinal epithelial lineage cells were stimulated with different cytokines and toll-like receptor (TLR) ligands,before pNF-κB-luciferase activity was assessed using the Dual-Luciferase reporter assay system.RESULTS:mRNA expression of mindin was upregulated 4.7 ± 1.1 fold compared with the baseline during DSS-induced intestinal inflammation in the mice.Stimulation with CpG-ODN (a known TLR-9 ligand) induced 4.2 ± 0.3 fold upregulation of mindin expression in RAW 264.7 cells.Full-length of mindin was cloned from cDNA of mouse mesenteric lymph node,then the pCMV-Mindin-Flag expression vector was established and the protein expression level was confi rmed.Transfection of the mindin construct and stimulation with CpG-ODN signifi cantly increased the NF-κB-luciferase activity by 2.5 ± 0.3 and 4.5 ± 0.5 fold in RAW264.7 and CMT93 cells,respectively (P < 0.01).CONCLUSION:Mindin expression is upregulated during intestinal inflammation and may induce NF-κB promoter activation in a TLR-9 mediated manner.

Study on the Significance of TLR-9 Expression in Rats with Acute Lung Injury Induced by Thoracic Trauma

Objective:To investigate the significance of toll receptor-9(TLR-9)expression in rats with acute lung injury(ALI)induced by thoracic trauma.Methods:Fifty healthy male SD rats were selected for the study,and 40 of them were randomly selected to establish the acute lung injury model induced by thoracic trauma,and 8 rats were randomly selected at 5,24,48 and 72 h after the impact,which were killed by cervical dislocation after arterial bleeding,and the lung tissue sections were made.Randomly select 8 rats from the remaining 10 rats as the control group,take blood and kill them to make lung tissue sections for reference.The lung dry/wet weight ratio(W/D)of the five groups were compared,and the levels of TLR-9 in bronchoalveolar lavage fluid(BALF)and serum,and the levels of TLR-9 mR NA,IL-6 mR NA,TLR-9 protein and IL-6protein in lung tissue were detected.Results:There was no significant difference in W/D levels among the control group,experimental group 1 and experimental group 4(P>0.05),while the W/D level of experimental group 3 was higher than that of the other four groups,and the W/D level of experimental group 2 was higher than that of the control group(P<0.05).The levels of ILR-9 in serum and BALF of rats in experimental group 1,experimental group 2 and experimental group 4 had no significant difference(P>0.05),which were higher than those in control group and lower than those in experimental group 3(P<0.05).The levels of TLR-9 mR NA,IL-6 mR NA,TLR-9 protein and IL-6 protein were the highest in the experimental group 3,and the lowest in the control group(P<0.05).The levels of TLR-9 mR NA and IL-6 mR NA in lung tissue of rats in experimental group 1,experimental group 2 and experimental group 4 had no significant difference(P>0.05).There was no significant difference in the levels of TLR-9 protein and IL-6 protein between experimental group 2 and experimental group 4(P>0.05),which were higher than those in experimental group 1(P<0.05).Conclusion:The expression level of TLR-9 is significantly increased in rats after thoracic trauma,and the expression of TLR-9 can reflect the degree of lung injury in rats after trauma to a certain extent,which is inferred to be related to the inflammatory response after lung injury.

血浆游离mtDNA及脑组织TLR-9/MAPK的表达在蛛网膜下腔出血中的意义

目的探讨蛛网膜下腔出血(SAH)模型大鼠血浆中游离线粒体DNA(mtDNA)的水平及脑组织中Toll样受体-9/丝裂原蛋白激酶(TLR-9/MAPK)的表达及意义。方法将40只大鼠随机分为SAH组和假手术(Sham)组,每组20只。SAH组采用血管内穿刺法建立SD大鼠SAH动物模型,Sham组仅穿刺但不造成蛛网膜下腔出血。模型建立成功24h后,采用荧光定量实时PCR检测大鼠外周血中血浆游离mtDNA的表达量;采用逆转录PCR(RT-PCR)和免疫组化法分别检测大鼠脑组织中TLR-9、P38 MAPK的表达。结果 SAH组大鼠在建模24h后血浆中游离mtDNA表达量高于Sham组(P<0.05);RT-PCR及免疫组化检测均显示脑组织中TLR9、P38 MAPK表达较Sham组增多(P<0.05)。结论血浆游离mtDNA与脑组织中TLR-9/MAPK的表达存在协同升高的现象,mtDNA可能通过TLR-9/MAPK通路介导脑组织局部及全身性炎症反应的发生。

CpG-ODN对肾小球肾炎进展的影响

目的观察CpG-ODN对肾小球肾炎进展的影响。方法将Wistar雄性大鼠随机分为N组(正常对照)、M组(模型对照)、CpG组和GpC组。N组大鼠经尾静脉注射生理盐水2.5ml/(kg·d),其余3组大鼠经尾静脉注射抗-thy1.1单克隆抗体2.5ml/(kg·d),隔日1次,共3次。注射抗-thy1.1单克隆抗体后,隔天,CpG组经尾静脉注射CpG-ODN,300μg/只,GpC组经尾静脉注射GpC-ODN,300μg/只,隔日注射,共3次。各组大鼠分别于用药前和第1针注射后1周测定尿蛋白;用药后第8周留取大鼠24h尿、血清及肾脏组织,检测24h尿蛋白定量、血清白蛋白和肾功;肾脏组织用于肾脏病理检查以及RT-PCR法检测TLR-9mRNA的表达。结果CpG组与M组和GpC组相比,血清白蛋白含量明显降低,且差异有显著意义;M组、CpG组和GpC组在给药后1周均出现尿蛋白,在给药后8周,24h尿蛋白含量明显增多;CpG组与M组相比,TLR-9mRNA在肾脏的表达水平明显增加,且差异有显著意义;GpC组与M组相比,差异无显著意义;光镜下可见CpG组肾组织病理改变明显加重。结论天然CpG-ODN可促进肾小球肾炎病情加重,TLR-9可能在其发生机理中起重要作用。

镜下清理富血小板血浆治疗轻中度膝骨关节炎

[目的]探究关节镜下清理术联合富血小板血浆(platelet-rich plasma,PRP)治疗轻中度膝骨关节炎(knee osteoarthritis,KOA)的疗效及可能机制。[方法]2021年1月—2022年4月收治的轻中度KOA患者110例,抽签法随机将其分为两组,每组55例。联合组接受关节镜下清理术联合PRP治疗,清理组仅关节镜下清理术治疗。比较两组围手术期、随访及辅助检查结果。[结果]两组手术时间、切口总长度、术中失血量、下地行走时间、切口愈合等级及住院时间的差异均无统计学意义(P>0.05)。术后7 d联合组肿胀VAS评分[(4.0±1.1)vs(4.5±1.0),P<0.05]显著低于清理组。随访12个月以上,联合组完全负重活动时间显著早于清理组[(45.2±6.2)d vs(50.1±7.2)d,P<0.05]。随时间推移,两组VAS、WOMAC和Lysholm膝关节评分均显著改善(P<0.05)。术后12个月联合组的VAS[(1.3±0.4)vs(2.1±0.6),P<0.05]、WOMAC[(25.0±5.2)vs(31.5±5.0),P<0.05]、Lysholm评分[(90.5±8.0)vs(85.3±8.1),P<0.05]均显著优于清理组。辅助检查方面,与治疗前相比,治疗后1个月两组患者血清TLR-4、MMP-9、SDF-1水平均显著下降(P<0.05);治疗后1个月,联合组的血清TLR-4[(7.0±2.0)ng/ml vs(8.4±2.2)ng/ml,P<0.05]、MMP-9[(26.0±7.0)pg/ml vs(30.1±7.3)pg/ml,P<0.05]和SDF-1[(453.4±47.2)μg/L vs(495.8±52.2)μg/L,P<0.05]显著优于清理组。[结论]关节镜下清理联合PRP治疗可减轻中度KOA患者疼痛,改善其膝关节功能,其机理可能与降低血液TLR-4、MMP-9、SDF-1水平有关。