目的 探讨雪貂食管黏膜下腺体(SMG)在胃食管生理稳态下清酸的作用机制。方法 选取无锡珊瑚礁公司6~8个月成年雪貂食管和出生1 d、3 d、5 d、1周和2周幼年雪貂食管,于宁夏医科大学总医院病理科制成冷冻和石蜡组织。苏木精-伊红染色法(HE染色)、过碘酸雪夫染色(PAS)对成年雪貂食管SMG形态学分析;LECTIN染色检测不同的凝集素在雪貂食管中的SMG结合情况;免疫组化及免疫荧光检测MUC5B和MU5AC的表达情况;组织免疫原位杂交检测囊性纤维化跨膜电导调节器(CFTR)在成年雪貂食管SMG中表达情况;免疫荧光检测新生雪貂1、3、5、7、14 d SMG中淋巴细胞结合增强因子(LEF1)表达情况,分析幼年雪貂食管上皮细胞LEF1在不同发育阶段的表达情况。结果 HE染色结果显示成年雪貂有角质化的多层鳞状食管上皮,食管SMG位于食管黏膜下层,是一个大小不等的小腔。PAS染色结果显示SMG具有分泌功能,正面腺体部分暗紫色的腺泡,产生的黏多糖分泌黏液性细胞。在成年雪貂食管SMG中不同类型细胞膜可特异性结合不同凝集素,黏膜细胞中结合双花扁豆凝集素(DBA)、鸡冠珊瑚树凝集素(ECL)、非偶联西非单叶豆凝集素Ⅰ (GSL Ⅰ) 、非偶联西非单叶豆凝集素Ⅱ(GSL Ⅱ)、大豆凝集素(SBA)、他卡林生物化合物凝集素(tacalin biotylated)、荆豆凝集素(ULEX)、麦胚凝集素(WGA),导管细胞结合凝集素GSL Ⅰ、GSL Ⅱ,上皮细胞结合凝集素ECL、花生凝集素(PNA)和WGA,浆液细胞结合凝集素伴刀豆凝集素(ConA)、红腰果E型凝集素(PHA-E) 和红腰果L型凝集素(PHA-L)。Mucin染色结果显示MUC5B、MU5AC在成年雪貂食管腺体黏液细胞胞质及腺体管腔内染色均呈阳性;原位杂交结果提示,在成年雪貂食管SMG中可见CFTR呈阳性表达;随着腺体的发育,在雪貂出生后3 d至1周可见发育腺体中LEF1的表达,当腺体分化成熟时LEF1表达消失。结论 雪貂食管SMG在稳态下表达CFTR调节HCO3^-的分泌,同时SMG具有分泌含有黏液蛋白MUC5B、MU5AC的黏液,这有助于食管的清酸和对黏膜的保护。通过对发育情况下SMG中LEF1表达的变化, 说明SMG可能通过WNT信号LEF1调控对稳态的维持。

Objective To investigate the mechanism of acid clearing in ferret esophageal submucosal gland under gastroesophageal physiological homeostasis.Methods The esophagus of adult ferrets aged 6-8 months and juvenile ferrets aged 1, 3, 5, 1 and 2 weeks after birth were selected and made into frozen and paraffin tissues in the Department of Pathology of General Hospital of Ningxia Medical University. Hematoxylin and eosin staining (HE staining) and periodized SCHFF staining (PAS) were used to analyze the morphology of esophageal submucosal glands (SMG) in adult ferrets. SMG binding of ferret esophagus was detected by LECTIN ; the expressions of MUC5B and MU5AC were detected by immunohistochemistry and immunofluorescence. The expression of cystic fibrosis transmembrane conductance regulator (CFTR) in adult ferret esophageal SMG was detected by tissue immunofluorescence in situ hybridization. Immunofluorescence was used to detect the expression of lymphocyte binding enhancement factor (LEF1) in SMG of neonatal ferrets on day 1, 3, 5, 7, and 14, and to analyze the expression of LEF1 in esophageal epithelial cells of juvenile ferrets at different developmental stages.Results HE staining showed that the SMG of the submucosal glands of the adult ferret esophagus was the connective tissue beneath the mucosal muscle layer of the esophagus, and PAS staining showed that the SMG had a secretory function; different cell types in the SMG of the adult ferret esophagus specifically bound different lectins, and the mucosal cells specifically bound Dolichos biflorus (DBA), ECL, GSL Ⅰ, GSL Ⅱ, SOYBEAN, tacalin biotylated, ULEX, Triticum vulgare Wheat germ agglutinin (WGA); specific binding of lectins GSL I and GSL Ⅱ in ductal cell membranes and positive expression of lectins ECL, PNA and WGA in epithelial cell membranes; lectins Mucin staining showed positive staining for MUC5B MU5AC in the cytoplasm of adult ferret esophageal glandular mucus cells and in the lumen of the gland. in situ hybridisation showed that CFTR was positively expressed in the SMG of the adult ferret esophagus; as the gland developed, LEF₁ was expressed in the developing gland from 3 days to 1 week after birth, but disappeared when the gland differentiated and matured.Conclusions In ferret SMG, CFTR expression can regulate HCO3^- secretion in steady state, and SMG can secrete mucus containing mucins MUC5B and MU5AC, which is helpful for esophageal acid clearance and mucosal protection. The changes of LEF₁ expression in SMG under developmental conditions suggest that SMG may regulate the maintenance of homeostasis through WNT signaling LEF₁.