Diebel Mark E, Diebel Lawrence N, Liberati David M
Department of Surgery, Wayne State University, Detroit, Michigan 48201, USA.
J Trauma. 2011 Aug;71(2):474-9. doi: 10.1097/TA.0b013e318228239d.
Laboratory studies demonstrate gender dimorphism following trauma/hemorrhagic shock (T/HS). These differences have been attributed to estrogen (E2) levels. Maintenance of gut barrier function by E2 following T/HS has been recently described. However, the mechanisms are not clear. The principle humoral defense mechanism of the gut is provided by secretory immunoglobulin IgA. It is transported across intestinal epithelial cells (IEC) by a specific transmembrane protein receptor (polyimmunoglobulin receptor, pIgR). Transport of IgA (transcytosis) may be influenced by a number of factors. We postulated that there may be differences in IgA transcytosis and IEC pIgR expression in response to sex hormones. We studied this in vitro.
Confluent HT-29 IEC monolayers were established in a two-chamber cell culture system. E2 or dihydrotestosterone (DHT) was added for 72 hours; then dimeric IgA (dIgA) was added to the basal chamber (4°C, to obtain maximal pIgR binding of dIgA). Apical media were sampled at intervals and recovery of secretory immunoglobulin IgA quantitated by enzyme-linked immunosorbent assay. PIgR expression in HT-29 cells was quantitated as mean fluorescence intensity using flow cytometry. Monolayer integrity was confirmed by serial measurement of transepithelial electrical resistance.
IgA transcytosis increased fourfold in 12-hour versus 3-hour culture periods in the control experiments. A similar finding was noted in the DHT experiments on IgA transcytosis. There were dramatic increases in IgA transcytosis across HT-29 cells exposed to E2.This was apparent at both 3- and 12-hour experimental time points and exhibited a dose-response effect. HT-29 cells cocultured with E2 increased pIgR expression in a time- and dose-dependent fashion. The greatest pIgR expression was noted following coculture of HT-29 cells with E2 for 6 days at the 1.0 μmol/L E2 concentration. The integrity of HT-29 monolayers in both the E2 and DHT treatment groups at T = 0 and 72 hours was assessed and showed no significant differences versus control cells.
IgA transcytosis was augmented by E2 in a dose-response fashion. This effect was due to augmented intracellular trafficking of IgA and later partly due to increased pIgR expression. The dose-related effects of E2 on IgA transport confirm the findings in animal studies that improved outcomes in females can be related to the estrus cycle.
实验室研究表明，创伤/失血性休克（T/HS）后存在性别差异。这些差异归因于雌激素（E2）水平。最近有研究描述了T/HS后E2对肠道屏障功能的维持作用。然而，其机制尚不清楚。肠道的主要体液防御机制由分泌型免疫球蛋白IgA提供。它通过一种特定的跨膜蛋白受体（多聚免疫球蛋白受体，pIgR）穿过肠上皮细胞（IEC）。IgA的转运（转胞吞作用）可能受多种因素影响。我们推测，性激素可能导致IgA转胞吞作用和IEC pIgR表达存在差异。我们对此进行了体外研究。
在双室细胞培养系统中建立融合的HT-29 IEC单层。加入E2或二氢睾酮（DHT）72小时；然后将二聚体IgA（dIgA）加入基底室（4°C，以获得dIgA与pIgR的最大结合）。定期采集顶侧培养基样本，通过酶联免疫吸附测定法定量分泌型免疫球蛋白IgA的回收率。使用流式细胞术将HT-29细胞中pIgR的表达定量为平均荧光强度。通过连续测量跨上皮电阻来确认单层的完整性。
在对照实验中，IgA转胞吞作用在12小时培养期内比3小时培养期增加了四倍。在DHT实验中，IgA转胞吞作用也有类似发现。暴露于E2的HT-29细胞的IgA转胞吞作用显著增加。在3小时和12小时的实验时间点均明显可见，且呈现剂量反应效应。与E2共培养的HT-29细胞以时间和剂量依赖的方式增加pIgR表达。在1.0 μmol/L E2浓度下，HT-29细胞与E2共培养6天后，pIgR表达最高。评估了E2和DHT处理组在T = 0和72小时时HT-29单层的完整性，与对照细胞相比无显著差异。
E2以剂量反应方式增强IgA转胞吞作用。这种效应是由于IgA细胞内转运增加，随后部分是由于pIgR表达增加。E2对IgA转运的剂量相关效应证实了动物研究中的发现，即雌性动物预后改善可能与发情周期有关。
