Objective: To explore the mechanism of early pancreatic exocrine function changes in severely scalded rats. Methods: The experimental research methods was used. Eighty male Sprague-Dawley rats aged 7-8 weeks were divided into simple sham injury group (n=8), sham injury+cholecystokinin octapeptide (CCK8) group (n=8), severe scald+CCK8 group (n=32), and extremely severe scald+CCK8 group (n=32) by the random number table, which were treated accordingly. Immediately after injury of rats in the 2 sham injury groups and 1, 2, 3, and 7 days after injury of rats in the 2 scald groups, the improved methods including pancreatic duct puncture and catheterization were used to dynamically collect the pancreatic-bile juice (PBJ) of rats. The PBJ secretory volume within 1 h was recorded, and the content of pancreatic lipase, α-amylase, and trypsin in PBJ was detected by enzyme-linked immunosorbent assay (ELISA), and the number of samples was 8. The femoral venous blood was collected, and the concentrations of pancreatic lipase and α-amylase in serum were detected by standard colorimetry to reflect their activity (n=8). The pancreatic tissue was extracted, and the levels of interleukin-1β (IL-1β) and IL-6 in pancreatic tissue were detected by ELISA (n=8), the expression of hypoxia-inducible factor 1α (HIF-1α) in pancreatic tissue was detected by immunofluorescence method, and the histopathological changes in pancreatic tissue were observed by hematoxylin-eosin staining, the severity of pancreatic tissue injury in the 2 scald groups was evaluated by modified Schmidt method (n=6), and the ultrastructure of acinar cells in pancreatic tissue was observed by transmission electron microscopy. Data were statistically analyzed with analysis of variance for factorial design, Tukey test, independent sample t test, and least significant difference test. Results: Compared with the PBJ secretory volume (0.740±0.030) mL in the pancreatic tissue of rats in simple sham injury group within 1 h immediately after injury, the (0.823±0.033) mL in sham injury+CCK8 group was significantly increased (t=4.92, P<0.05). Compared with that of rats in sham injury+CCK8 group immediately after injury, the PBJ secretory volume of rats within 1 h in severe scald+CCK8 group ((0.681±0.024), (0.608±0.056), (0.525±0.025), and (0.720±0.044) mL) and extremely severe scald+CCK8 group ((0.540±0.025), (0.406±0.021), (0.475±0.036), and (0.690±0.018) mL) was significantly decreased on 1, 2, 3, and 7 days after injury (P<0.05). Compared with that in severe scald+CCK8 group, the PBJ secretory volume of rats within 1 h in extremely severe scald+CCK8 group was significantly decreased on 1 and 2 days after injury (P<0.05). Compared with that of rats in simple sham injury group immediately after injury, the content of pancreatic lipase, α-amylase, and trypsin in PBJ of rats in sham injury+CCK8 group immediately after injury was significantly increased (with t values of 4.56, 3.30, and 4.99, respectively, P<0.05). Compared with that of rats in sham injury+CCK8 group immediately after injury, the content of pancreatic lipase and α-amylase in PBJ of rats in severe scald+CCK8 group and extremely severe scald+CCK8 group was significantly decreased on 1, 2, 3, and 7 days after injury (P<0.05), the trypsin content in PBJ of rats in extremely severe scald+CCK8 group was significantly decreased on 2 days after injury (P<0.05). Compared with that in severe scald+CCK8 group, the content of pancreatic lipase in PBJ of rats in extremely severe scald+CCK8 group was significantly decreased on 1, 2, and 3 days after injury (P<0.05), and the content of α-amylase and trypsin in PBJ was significantly decreased on 1 and 2 days after injury (P<0.05). There were no statistically significant differences in the activities of pancreatic lipase and α-amylase in serum of rats among the 4 groups at various time points after injury (P>0.05). Compared with that of rats in sham injury+CCK8 group immediately after injury, the levels of IL-1β in pancreatic tissue of rats in severe scald+CCK8 group on 1, 2, and 3 days after injury and in extremely severe scald+CCK8 group on 1, 2, 3, and 7 days after injury were significantly increased (P<0.05), and the levels of IL-6 in pancreatic tissue of rats in severe scald+CCK8 group and extremely severe scald+CCK8 group were significantly increased on 1, 2, 3, and 7 days after injury (P<0.05). Compared with that in severe scald+CCK8 group, the IL-1β level in pancreatic tissue of rats in extremely severe scald+CCK8 group was significantly increased on 2 and 3 days after injury (P<0.05), and IL-6 level in pancreatic tissue was significantly increased on 2 days after injury (P<0.05). The expression levels of HIF-1α in pancreatic tissue of rats in simple sham injury group and sham injury+CCK8 group immediately after injury were lower; and compared with that in sham injury+CCK8 group immediately after injury, the expression levels of HIF-1α in pancreatic tissue of rats in the 2 scald groups increased to a certain extent at different time points after injury, and the expression position was transited from the edge of the pancreatic tissue to the whole pancreas, the expression levels of HIF-1α in pancreatic tissue of rats in the 2 scald groups tended to be normal on 7 days after injury. Compared with that in simple sham injury group immediately after injury, the proportion of acinar cell cytoplasm in pancreatic tissue of rats in sham injury+CCK8 group was increased; and with the increase of time after injury, edema, hemorrhage, necrosis, and inflammatory infiltration appeared in pancreatic tissue of rats in the 2 scald groups. Compared with that in severe scald+CCK8 group, the scores of edema, inflammatory cell infiltration, bleeding, and necrosis in pancreatic tissue of rats in extremely severe scald+CCK8 group were increased to varying degrees at various time points after injury, and the scores of pancreatic tissue of rats in the 2 scald groups basically recovered to normal on 7 days after injury. Compared with that in simple sham injury group immediately after injury, the number of enzyme granules in acinar cells of pancreatic tissue of rats in sham injury+CCK8 group was increased, and with the increase of time after injury, the enzyme granules in acinar cells of rats in the 2 scald groups were gradually reduced basically. Conclusions: The exocrine functions of pancreas, such as synthesis and secretion of pancreatic enzymes, are decreased in the early stage in severely scalded rats. And the greater the scalded area, the more significant the decline of pancreatic exocrine function. This change may be related to hypoxic injury and inflammation in pancreatic tissue after severe scald.
目的： 探讨严重烫伤大鼠早期胰腺外分泌功能变化的机制。 方法： 采用实验研究方法。取80只7~8周龄雄性SD大鼠，按随机数字表法分为单纯假伤组（8只）、假伤+胆囊收缩素八肽（CCK8）组（8只）、严重烫伤+CCK8组（32只）和特重烫伤+CCK8组（32只），分别作相应处理。2组假伤大鼠伤后即刻及2组烫伤大鼠伤后1、2、3、7 d，采用改进的胰管穿刺置管法动态收集大鼠胆胰混合液（PBJ），记录1 h内分泌PBJ的体积，采用酶联免疫吸附测定（ELISA）法检测并计算PBJ中胰脂肪酶、α-淀粉酶和胰蛋白酶的含量（样本数为8）；收集股静脉血，采用标准比色法检测血清中胰脂肪酶和α-淀粉酶的浓度，反映其活力（样本数为8）；提取胰腺组织，采用ELISA法检测胰腺组织中白细胞介素1β（IL-1β）和IL-6水平（样本数为8），采用免疫荧光法检测胰腺组织中缺氧诱导因子1α（HIF-1α）的表达，采用苏木精-伊红染色观察胰腺组织病理学变化，采用改良Schmidt法评估（仅2组烫伤大鼠）胰腺组织损伤的严重程度（样本数为6），采用透射电子显微镜观察胰腺组织腺泡细胞超微结构。对数据行析因设计方差分析、Tukey检验、独立样本t检验和LSD检验。 结果： 与单纯假伤组大鼠伤后即刻胰腺组织1 h内的PBJ分泌体积（0.740±0.030）mL相比，假伤+CCK8组的（0.823±0.033）mL显著升高（t=4.92，P<0.05）。与假伤+CCK8组伤后即刻相比，严重烫伤+CCK8组［（0.681±0.024）、（0.608±0.056）、（0.525±0.025）、（0.720±0.044）mL］、特重烫伤+CCK8组［（0.540±0.025）、（0.406±0.021）、（0.475±0.036）、（0.690±0.018）mL］大鼠伤后1、2、3、7 d胰腺组织1 h内的PBJ分泌体积均显著下降（P<0.05）。与严重烫伤+CCK8组相比，特重烫伤+CCK8组大鼠伤后1、2 d胰腺组织1 h内的PBJ分泌体积均显著下降（P<0.05）。相较于单纯假伤组伤后即刻，假伤+CCK8组大鼠伤后即刻PBJ中胰脂肪酶、α-淀粉酶和胰蛋白酶含量均显著升高（t值分别为4.56、3.30、4.99，P<0.05）。与假伤+CCK8组大鼠伤后即刻相比，严重烫伤+CCK8组和特重烫伤+CCK8组大鼠伤后1、2、3、7 d的PBJ中胰脂肪酶和α-淀粉酶含量均显著降低（P<0.05），特重烫伤+CCK8组大鼠伤后2 d的PBJ中胰蛋白酶含量显著降低（P<0.05）。与严重烫伤+CCK8组相比，特重烫伤+CCK8组大鼠伤后1、2、3 d的PBJ中胰脂肪酶含量均显著降低（P<0.05），伤后1、2 d的PBJ中α-淀粉酶和胰蛋白酶含量均显著降低（P<0.05）。4组大鼠伤后各时间点血清中胰脂肪酶、α-淀粉酶活力比较，差异均无统计学意义（P>0.05）。与假伤+CCK8组大鼠伤后即刻相比，严重烫伤+CCK8组大鼠伤后1、2、3 d和特重烫伤+CCK8组大鼠伤后1、2、3、7 d胰腺组织中的IL-1β水平均显著升高（P<0.05），严重烫伤+CCK8组和特重烫伤+CCK8组大鼠伤后1、2、3、7 d胰腺组织中的IL-6水平均显著升高（P<0.05）。与严重烫伤+CCK8组相比，特重烫伤+CCK8组大鼠伤后2、3 d胰腺组织中的IL-1β水平均显著升高（P<0.05），伤后2 d胰腺组织中的IL-6水平均显著升高（P<0.05）。单纯假伤组和假伤+CCK8组大鼠伤后即刻胰腺组织中HIF-1α表达水平均较低；相较于假伤+CCK8组伤后即刻，2组烫伤大鼠胰腺组织中HIF-1α表达水平于伤后不同时间点均呈一定程度的升高，其表达位置由胰腺组织边缘向全胰腺过渡，2组烫伤大鼠胰腺组织中HIF-1α表达水平于伤后7 d趋于正常。相较于单纯假伤组伤后即刻，假伤+CCK8组大鼠胰腺组织中腺泡细胞胞质占比增大；2组烫伤大鼠随伤后时间的延长，胰腺组织出现不同程度的水肿、出血、坏死及炎症浸润。与严重烫伤+CCK8组相比较，特重烫伤+CCK8组大鼠伤后各时间点胰腺组织水肿、炎症细胞浸润、出血和坏死得分表现出不同程度的升高，且2组烫伤大鼠胰腺组织各项目评分均于伤后7 d基本恢复正常。相较于单纯假伤组伤后即刻，假伤+CCK8组大鼠胰腺组织腺泡细胞内酶原颗粒数量增加；2组烫伤大鼠随伤后时间的延长，腺泡细胞内酶原颗粒基本呈逐渐减少趋势。 结论： 严重烫伤大鼠早期胰酶合成和分泌等胰腺外分泌功能下降；且烫伤面积越大，外分泌功能下降程度越显著。这一变化可能与严重烫伤后胰腺组织发生缺氧损伤和炎症反应有关。.