Data underlying the research on Protective Effect of miR-214 on Acute Liver Injury in Septic Mice Based on the TLR4/NF-κB Signaling Pathway

This study aims to investigate the protective effect of miR-214 against acute liver injury induced by sepsis. An acute liver injury model in septic mice was established using the CLP method. The mice were randomly assigned to four groups: the sham operation group (sam), the model group (model), the miR-214 inhibitor group (inhibitor), and the miR-214 mimic group (mimic). Liver function indices, including ALT and AST, as well as inflammatory factors such as IL-6, IL-11, IL-1β, and TNF-α, were measured across all groups. Liver histopathological damage was evaluated microscopically following hematoxylin-eosin staining, and the pathological sections were scored accordingly. Additionally, the protein levels of TLR4, NF-κBp65, p-NF-κBp65, and IκB-α in liver tissues were assessed using protein blotting. The expression levels of miR-214 in the liver tissues of mice from each group were quantified by RT-qPCR. The administration of miR-214 mimics resulted in a reduction in ALT and AST levels, indicating improved liver function. Furthermore, miR-214 mimics inhibited the levels of inflammatory factors, including IL-6, IL-11, IL-1β, and TNF-α. Additionally, there was a decrease in the protein expression of TLR4, NF-κBp65, p-NF-κBp65, and IκB-α in septic mice. These effects collectively mitigated liver tissue damage and facilitated the recovery of hepatic histopathological injury.Our findings suggest that the overexpression of miR-214 may confer a protective effect against acute liver injury in a mouse model of sepsis. 1. In comparison to the sham operation group, serum levels of ALT and AST were significantly elevated in both the model and miR-214 inhibitor groups (P<0.001). Additionally, when comparing the model group to the miR-214 inhibitor group, serum levels of ALT and AST were notably lower in the miR-214 inhibitor group (P<0.001). These experimental findings suggest that miR-214 contributes to the recovery of liver function in mice experiencing acute liver injury due to sepsis (see Table 1 and Figure 1). 2. Compared to the sham operation group, serum levels of inflammatory factors (TNF-α, IL-6, IL-11, IL-1β) were significantly elevated in both the model and miR-214 inhibitor groups (P<0.001). Furthermore, when comparing the model group to the miR-214 mimics group, the serum levels of inflammatory factors (TNF-α, IL-6, IL-11, IL-1β) were significantly higher in the miR-214 mimics group, while these levels were significantly reduced in the miR-214 mimics group compared to the model group (P<0.001). These experimental results suggest that miR-214 plays a role in mitigating the inflammatory response in vivo in mice with acute liver injury resulting from sepsis (see Table 2 and Figure 2). 3. Compared to the sham operation group, the pathological scores of hepatic tissue injury were markedly higher in both the model and miR-214 inhibitor groups (P<0.001). Furthermore, when compared to the model group, the pathological scores of hepatic tissue injury in the miR-214 inhibitor group were significantly elevated, while the miR-214 mimics group exhibited significantly lower liver tissue injury pathology scores than the miR-214 inhibitor group (P<0.001). These experimental results suggest that miR-214 mimics may facilitate the recovery of liver tissue injury in mice experiencing acute liver injury due to sepsis (see Table 3 and Figure 3). 4. Hepatic tissue TLR4 pathway protein expression in mice exhibited significant differences among the various groups. Compared to the sham operation group, the protein expression levels of TLR4, NF-κBp65, p-NF-κBp65, and IκB-α were markedly increased in both the model group and the miR-214 inhibitor group (P<0.001). Additionally, when compared to the model group, the hepatic tissue expression of TLR4, NF-κBp65, p-NF-κBp65, and IκB-α was significantly elevated in the miR-214 inhibitor group. In contrast, the expression levels of these proteins significantly decreased in the miR-214 mimics group (P<0.001). The experimental results suggest that miR-214 mimics inhibit the expression of proteins associated with the TLR4 signaling pathway (see Figure 4a and 4b). 5. In comparison to the sham operation group, the expression of miR-214 in the liver tissue of mice in both the model and miR-214 inhibitor groups was significantly decreased. Both groups exhibited more severe symptoms of sepsis and greater liver tissue injury (P<0.001). Furthermore, when compared to the model group, the miR-214 inhibitor group showed a significant reduction in miR-214 expression in liver tissue, which correlated with an improvement in sepsis symptoms and liver tissue injury. Conversely, in the miR-214 mimics group, the expression of miR-214 in liver tissue increased significantly, leading to exacerbated sepsis symptoms and liver tissue injury (P<0.001). These experimental results suggest that overexpression of miR-214 plays a protective role against acute liver injury in the context of sepsis (see Table 4 and Figure 5). 6. Using bivariate Pearson's test, miR-214 exhibited a negative correlation with the severity of acute liver injury in sepsis (P<0.01). Therefore, supplementation with miR-214 seems to promote recovery from acute liver injury in sepsis (see Figure 6).

本研究旨在探讨miR-214对脓毒症诱导的急性肝损伤的保护作用。本研究采用盲肠结扎穿刺（CLP）法构建脓毒症小鼠急性肝损伤模型，将小鼠随机分为四组：假手术组（sam）、模型组（model）、miR-214抑制剂组（inhibitor）以及miR-214模拟物组（mimic）。各组均检测谷丙转氨酶（ALT）、谷草转氨酶（AST）等肝功能指标，以及IL-6、IL-11、IL-1β、TNF-α等炎性因子水平；通过苏木精-伊红（hematoxylin-eosin）染色后镜下评估肝组织病理损伤，并对病理切片进行评分；采用蛋白印迹法检测肝组织中Toll样受体4（TLR4）、核因子κB p65（NF-κBp65）、磷酸化核因子κB p65（p-NF-κBp65）及核因子κB抑制蛋白α（IκB-α）的蛋白水平；通过实时定量聚合酶链反应（RT-qPCR）定量检测各组小鼠肝组织中miR-214的表达水平。

给予miR-214模拟物可降低ALT与AST水平，提示肝功能得到改善；同时可抑制IL-6、IL-11、IL-1β、TNF-α等炎性因子的表达，并下调脓毒症小鼠肝组织中TLR4、NF-κBp65、p-NF-κBp65及IκB-α的蛋白水平。上述效应共同减轻了肝组织损伤，促进肝组织病理损伤的修复。本研究结果显示，miR-214过表达可对脓毒症小鼠急性肝损伤发挥保护作用。

1. 与假手术组相比，模型组与miR-214抑制剂组小鼠的血清ALT、AST水平均显著升高（P<0.001）；相较于模型组，miR-214抑制剂组的血清ALT、AST水平显著降低（P<0.001）。上述实验结果表明，miR-214有助于脓毒症诱导的急性肝损伤小鼠的肝功能恢复（详见表1及图1）。
2. 与假手术组相比，模型组与miR-214抑制剂组小鼠的血清炎性因子（TNF-α、IL-6、IL-11、IL-1β）水平均显著升高（P<0.001）；进一步对比模型组与miR-214模拟物组，miR-214模拟物组的炎性因子水平显著更高，而相较于模型组，miR-214模拟物组的炎性因子水平显著降低（P<0.001）。上述实验结果表明，miR-214可缓解脓毒症诱导的急性肝损伤小鼠体内的炎症反应（详见表2及图2）。
3. 与假手术组相比，模型组与miR-214抑制剂组小鼠的肝组织损伤病理评分均显著升高（P<0.001）；相较于模型组，miR-214抑制剂组的肝组织损伤病理评分进一步升高，而miR-214模拟物组的肝组织损伤病理评分显著低于miR-214抑制剂组（P<0.001）。上述实验结果表明，miR-214模拟物可促进脓毒症诱导的急性肝损伤小鼠的肝组织损伤修复（详见表3及图3）。
4. 各组小鼠肝组织TLR4通路蛋白表达存在显著差异。与假手术组相比，模型组与miR-214抑制剂组小鼠肝组织中TLR4、NF-κBp65、p-NF-κBp65及IκB-α的蛋白表达水平均显著升高（P<0.001）；相较于模型组，miR-214抑制剂组小鼠肝组织中上述蛋白的表达水平进一步升高，而miR-214模拟物组中上述蛋白的表达水平显著降低（P<0.001）。上述实验结果表明，miR-214模拟物可抑制TLR4信号通路相关蛋白的表达（详见图4a及图4b）。
5. 与假手术组相比，模型组与miR-214抑制剂组小鼠肝组织中miR-214的表达水平显著降低，且两组小鼠的脓毒症症状更严重、肝组织损伤程度更高（P<0.001）；相较于模型组，miR-214抑制剂组小鼠肝组织中miR-214的表达水平进一步降低，这与脓毒症症状及肝组织损伤的加重相关；而miR-214模拟物组小鼠肝组织中miR-214的表达水平显著升高，进而导致脓毒症症状及肝组织损伤加重（P<0.001）。上述实验结果表明，miR-214过表达可对脓毒症小鼠的急性肝损伤发挥保护作用（详见表4及图5）。
6. 采用双变量Pearson相关分析显示，miR-214的表达水平与脓毒症急性肝损伤的严重程度呈负相关（P<0.01）。因此，补充miR-214似乎可促进脓毒症急性肝损伤的恢复（详见图6）。