先天性肠闭锁患儿组织STAT3和血清STAT3 mRNA,IL-12p40,IL-13Rα2水平表达及与预后的相关性研究

Study on the Expression of Tissue STAT3 and Serum STAT3 mRNA,IL-12p40 and IL-13Rα2 Levels in Children with Congenital Intestinal Atresia and Their Correlation with Prognosis

目的探究先天性肠闭锁(congenital intestinal atresia)患儿组织信号传导与转录激活因子3(signal transducerand activator of transcription 3,STAT3)和血清STAT3 mRNA,IL-12p40及IL-13Rα2水平表达及与预后的相关性。方法收集2020年1月~2023年1月在河北省儿童医院进行治疗的先天性肠闭锁患儿术中切除的100例肠闭锁病变组织、正常肠管组织及术前血清样本,根据Grosfeld分型标准将患儿分为Ⅰ型39例,Ⅱ型22例,Ⅲ型30例和Ⅳ型9例。根据术后6个月的恢复情况,将患儿分为预后良好组(n=78)和预后不良组(n=22);并选取93例同期体检健康儿童的血清样本作为对照。免疫组织化学检测STAT3在组织中阳性表达及定位情况;Western blot检测组织中STAT3蛋白表达;荧光定量PCR法(qPCR)检测血清中STAT3 mRNA的表达水平。采用Pearson相关分析先天性肠闭锁患儿血清STAT3与炎症因子水平的相关性。采用Logistic回归分析影响先天性肠闭锁患儿预后的因素。利用受试者工作特征(ROC)曲线分析血清STAT3水平对先天性肠闭锁患儿预后的预测效能。结果免疫组织化学结果显示,STAT3阳性表达主要定位于细胞质和细胞核,先天性肠闭锁组织中的阳性表达率(86%)显著高于正常肠管组织(18%),差异具有统计学意义(χ^(2)=92.628,P<0.05)。Western blot结果显示STAT3在先天性肠闭锁组织中的相对表达量(1.59±0.21)显著高于正常肠管组织中的相对表达水平(0.81±0.12),差异具有统计学意义(t=30.567,P<0.05)。qPCR法结果显示先天性肠闭锁组患儿血清STAT3 mRNA(2.13±0.56),IL-12p40(0.89±0.13ng/ml)以及IL-13Rα2(6.42±1.86ng/ml)水平均显著高于对照组(1.06±0.11,0.37±0.08ng/ml,1.35±0.41ng/ml),差异具有统计学意义(t=18.101,33.170,25.708,均P<0.05)。随着患儿分型的增加,STAT3 mRNA及IL-12p40,IL-13Rα2水平逐渐增加,差异具有统计学意义(F=52.666,160.300,25.82,均P<0.05)。Pearson相关分析显示先天性肠闭锁患儿血清STAT3 mRNA与炎症因子IL-12p40,IL-13Rα2水平均呈显著正相关(r=0.496,0.564,均P<0.001)。先天性肠闭锁患儿预后不良组血清STAT3 mRNA(3.01±0.75)表达水平显著高于预后良好组(1.88±0.51),差异具有统计学意义(t=8.212,P<0.05)。Logistic回归分析结果显示,STAT3 mRNA,IL-12p40,IL-13Rα2水平以及低出生质量均是先天性肠闭锁患儿预后不良的独立危险因素(均P<0.05)。ROC曲线可知血清STAT3评估先天性肠闭锁患儿预后的曲线下面积(AUC)为0.916,敏感度和特异度分别为81.82%,88.46%,当血清STAT3 mRNA水平高于2.47时先天性肠闭锁患儿发生预后不良的几率较高。结论STAT3在先天性肠闭锁患儿组织和血清中的表达显著升高,血清STAT3对患儿预后情况具有一定的预测价值。

Objective To investigate the expression of tissue signal transducer and activator of transcription 3(STAT3)and serum STAT3 mRNA,IL-12p40 and IL-13Rα2 levels in children with congenital intestinal atresia and their correlation with prognosis.Methods From January 2020 to January 2023,100 cases of intestinal atresia lesion tissues,normal intestinal tissues and preoperative serum samples were collected from children with congenital intestinal atresia who underwent treatment in Hebei Children’s Hospital.According to the Grosfeld typing criteria,these children were categorized into 39 cases of type I,22 cases of type II,30 cases of type III and 9 cases of type IV.Based on the recovery situation at 6 months after surgery,these children were separated into a good prognosis group(n=78)and a poor prognosis group(n=22).Serum samples from 93 cases of healthy children undergoing medical examinations during the same period were regarded as control samples.Immunohistochemistry was applied to detect the positive expression and localization of STAT3 in tissues.Western blot was applied to detect the expression of STAT3 protein in tissues,and quantitative polymerase chain reaction(qPCR)was applied to detect the expression level of STAT3 mRNA in serum.Pearson correlation was applied to analyze the correlation between serum STAT3 and inflammatory factor levels in children with congenital intestinal atresia.Logistic regression was used to analyze the factors affecting the prognosis of children with congenital intestinal atresia.Receiver operating characteristic(ROC)curve was applied to analyze the predictive efficacy of serum STAT3 level on the prognosis of children with congenital intestinal atresia.Results Immunohistochemical results showed that STAT3 positive expression was mainly localized in the cytoplasm and nucleus.The positive expression rate in congenital intestinal atresia tissue(86%)was higher than that in normal intestinal tissue(18%),and the difference was significant(χ^(2)=92.628,P<0.05).Western blot results showed that the relative expression level of STAT3 in congenital intestinal atresia tissue(1.59±0.21)was higher than that in normal intestinal tissue(0.81±0.12),and the difference was significant(t=30.567,P<0.05).The results of qPCR showed that serum STAT3(2.13±0.56),IL-12p40(0.89±0.13 ng/ml),and IL-13Rα2 levels(6.42±1.86ng/ml)in the congenital intestinal atresia group were higher than those in the control groups(1.06±0.11,0.37±0.08ng/ml,1.35±0.41ng/ml),and the differences were significant(t=18.101,33.170,25.708,all P<0.05).The levels of STAT3 and IL-12p40,IL-13Rα2 were gradually increased with the increase of the children’s subtypes,and the differences were significant(F=52.666,160.300,25.82,all P<0.05).Pearson correlation analysis showed a positive correlation between serum STAT3,IL-12p40,and IL-13Rα2 levels in children with congenital intestinal atresia(r=0.496,0.564,all P<0.001).The expression level of serum STAT3 in poor prognosis group(3.01±0.75)was higher than that in good prognosis group(1.88±0.51),and the differences was statistically significant(t=8.212,P<0.05).Logistic regression showed that STAT3,IL-12p40,IL-13Rα2,and low birth quality were all independent risk factors for poor prognosis in children with congenital intestinal atresia(all P<0.05).The ROC curve showed that the area under the curve(AUC)for evaluating the prognosis of children with congenital intestinal atresia by serum STAT3 expression was 0.916,with a sensitivity of 81.82%and a specificity of 88.46%,respectively.When the serum STAT3 mRNA level was higher than 2.47,children with congenital intestinal atresia had a higher probability of poor prognosis.Conclusion The expression of STAT3 is increased in the tissues and serum of children with congenital intestinal atresia.Serum STAT3 may have a predictive value for the prognosis of affected children.