Department of Neonatology, The Affiliated Wuxi Maternity and Child Health Care Hospital of Nanjing Medical University, Wuxi, China.
J Cell Physiol. 2019 Aug;234(8):13431-13438. doi: 10.1002/jcp.28022. Epub 2019 Jan 7.
To explore the relationship between high-mobility group box 1 (HMGB1) and NLR pyrin domain containing 3 (NLRP3) in the development of necrotizing enterocolitis (NEC).
NEC rat models were constructed and treated with HMGB1 inhibitor glycyrrhizin (GL) with different concentration. An inflammatory condition of intestinal tissue in newborn NEC rats was observed by hematoxylin and eosin staining. The messenger RNA (mRNA) and protein expression of HMGB1, NLRP3, toll-like receptor 4 (TLR4), nuclear factor-κB (NF-κB), and caspase 1 were determined by real-time polymerase chain reaction and western blot analysis, respectively. The content of interleukin (IL)-1β and tumor necrosis factor-α (TNF-α) was determined by enzyme-linked immunosorbent assay. Human intestinal epithelial cell lines were induced to NEC by lipopolysaccharides (LPSs). LPS-induced cells were transfected with small interfering RNA-HMGB1 and NLRP3 plasmid vector. The mRNA and protein expression of HMGB1, NLRP3, TLR4, NF-κB, caspase 1, IL-1β, and TNF-α were determined by real-time PCR and western blot analysis, respectively.
The mRNA and protein expression of HMGB1 and NLRP3 in the NEC group was significantly higher than the control group. Inhibition of HMGB1 expression improved intestinal inflammation in newborn NEC rats. The expression of HMGB1, NLRP3, TLR4, NF-κB, and caspase 1 was upregulated in NEC and was weakened after treating with GL. LPS induction to intestinal epithelial cells markedly increased the expression of HMGB1, NLRP3, TLR4, NF-κB, caspase 1, IL-1β, and TNF-α. The knockdown of HMGB1 abolished the increase of expression, whereas further transfection with NLRP3 plasmid vector recovered the increase.
HMGB1 and NLRP3 were all upregulated in the development of NEC. Inhibition on HMGB1 could improve the intestinal inflammation in NEC by inhibiting NLRP3 via TLR4 and NF-κB signaling pathways.
探讨高迁移率族蛋白 B1（HMGB1）与富含吡喃结构域的 NOD 样受体蛋白 3（NLRP3）在坏死性小肠结肠炎（NEC）发展中的关系。
构建 NEC 大鼠模型，并使用不同浓度的 HMGB1 抑制剂甘草酸（GL）进行处理。通过苏木精和伊红染色观察新生 NEC 大鼠肠道组织的炎症情况。通过实时聚合酶链反应和蛋白质印迹分析分别测定 HMGB1、NLRP3、Toll 样受体 4（TLR4）、核因子-κB（NF-κB）和半胱天冬酶 1 的信使 RNA（mRNA）和蛋白表达。通过酶联免疫吸附试验测定白细胞介素（IL）-1β和肿瘤坏死因子-α（TNF-α）的含量。用人肠上皮细胞系通过脂多糖（LPS）诱导 NEC。用小干扰 RNA-HMGB1 和 NLRP3 质粒载体转染 LPS 诱导的细胞。通过实时 PCR 和蛋白质印迹分析分别测定 HMGB1、NLRP3、TLR4、NF-κB、半胱天冬酶 1、IL-1β和 TNF-α的 mRNA 和蛋白表达。
NEC 组的 HMGB1 和 NLRP3 的 mRNA 和蛋白表达均明显高于对照组。抑制 HMGB1 表达可改善新生 NEC 大鼠的肠道炎症。HMGB1、NLRP3、TLR4、NF-κB 和半胱天冬酶 1 的表达在 NEC 中上调，并用 GL 处理后减弱。LPS 诱导肠上皮细胞明显增加 HMGB1、NLRP3、TLR4、NF-κB、半胱天冬酶 1、IL-1β 和 TNF-α的表达。HMGB1 的敲低消除了这种增加，而进一步转染 NLRP3 质粒载体恢复了增加。
HMGB1 和 NLRP3 在 NEC 的发展中均上调。抑制 HMGB1 可通过 TLR4 和 NF-κB 信号通路抑制 NLRP3，从而改善 NEC 中的肠道炎症。
