浸泡免疫迟钝爱德华氏菌疫苗诱导牙鲆黏液细胞的动态变化

Dynamic changes of mucous cells in Paralichthys olivaceus induced by immersion immunization with inactivated Edwardsiella tarda

为探明鱼类黏膜相关淋巴组织中黏液细胞对疫苗免疫的应答特性,本研究利用组织学和组织化学技术,研究了浸泡免疫灭活迟钝爱德华氏菌前后牙鲆皮肤、鳃、前肠、中肠和后肠中黏液细胞数量和特性的时序变化。苏木精-曙红(HE)、阿尔新蓝-过碘酸雪夫氏(AB-PAS)染色结果显示,免疫前皮肤主要分布含中性黏多糖的Ⅰ型黏液细胞以及含中性黏多糖和少量酸性黏多糖的Ⅲ型黏液细胞,鳃中可观察到Ⅰ、Ⅲ型以及含酸性黏多糖和少量中性黏多糖的Ⅳ型黏液细胞,前肠中主要分布Ⅰ型黏液细胞,中肠、后肠上皮中可观察到Ⅰ、Ⅲ、Ⅳ型和含酸性黏多糖的Ⅱ型黏液细胞。免疫后,牙鲆黏膜免疫相关淋巴组织中黏液细胞总量随时间呈现先增多后减少趋势,后肠在第5天达到峰值,其他于第3天达到峰值;免疫后各组织中Ⅰ型黏液细胞数量减少,含酸性黏多糖成分的Ⅱ、Ⅲ及Ⅳ型黏液细胞数量显著增多,表明疫苗免疫诱导黏液细胞成分由中性黏多糖向酸性黏多糖转变。p H 2.5和p H 1.0条件下AB染色结果显示,免疫后黏膜相关淋巴组织中酸性黏液细胞数量增多,且多数为硫酸化酸性黏液细胞。本研究结果为鱼类黏液的免疫防御功能及黏液细胞在鱼类黏膜免疫中的作用提供了理论依据。

The mucosal surfaces of fish, including skin, gill and gastrointestinal tract, contain numerous poorly studied immune substances that act as the first line of defence against invading pathogens.The fish body＇s mucosal surfaces are protected from pathogens, physical and chemical attacks by the gel-like extracellular matrix（mucus）which is synthesized, stored and secreted by mucous cells. In order to better understand the vaccine-induced immune responses of mucous cells, histological and histochemical techniques were used to investigate the dynamic changes of the numbers and types of mucous cells in mucosa-associated lymphoid tissues, including skin, gills,foregut, midgut and hindgut, in Paralichthys olivaceus after immersion immunization with inactivated Edwardsiella tarda. The results of hematoxylin and eosin（HE） staining, alcian blue and periodic acid Schiff reaction（AB-PAS）, indicated that the type Ⅰ mucous cells containing PAS-positive neutral mucopolysaccharide and type Ⅲ cells containing neutral mucopolysaccharide and a little AB-positive acidic mucopolysaccharide in skin, type Ⅰ, Ⅲ, and Ⅳwhich contained acidic mucopolysaccharide and a little neutral mucopolysaccharide in gill, type Ⅰ in foregut mainly, and type Ⅰ, Ⅲ, Ⅳ and Ⅱ which contained acidic mucopolysaccharide in midgut and hindgut, were observed before immunization. After immunization with inactivated E. tarda, the mucous cell numbers were increased over time and were peaked on day 3 post immunization（p.i.） in skin, gill, foregut and midgut and on day 5 p.i. in hindgut; and then were decreased to the level of control groups; Additionally, the amount of typeⅠmucous cells was reduced while the amount of type Ⅱ, Ⅲ and Ⅳ mucous cells was increased,revealing that the components of mucous cells have been changed from neutral mucopolysaccharide to acidic mucopolysaccharide due to the immunization with E. tarda. The results of AB staining showed that the mucous cells containing acidic mucopolysaccharide were increased after immunization, which were mostly sulfated acid mucopolysaccharide-containing mucous cells. These results will provide valuable information for the researches on the mucosal immunity in fish.