Afonso A, Lousada S, Silva J, Ellis A E, Silva M T
Institute of Molecular and Cellular Biology, University of Porto, Portugal.
Dis Aquat Organ. 1998 Sep 11;34(1):27-37. doi: 10.3354/dao034027.
The neutrophil and macrophage responses that accompany inflammation were studied in the peritoneal cavity of rainbow trout Oncorhynchus mykiss using light and electron microscopic cytochemistry. Neutrophils of inflammatory peritoneal exudates were alpha-naphthyl butyrate esterase-negative, peroxidase-positive and rich in cytoplasmic glycogen granules. Macrophages were poor in glycogen, esterase-positive and usually peroxidase-negative. Some peroxidase-positive macrophages were due to the transfer to macrophages of neutrophilic peroxidase. The ultrastructural double labelling for glycogen-peroxidase or esterase/peroxidase was most useful for precisely characterising neutrophils and macrophages in the inflamed peritoneal cavities and for correctly labelling peroxidase-positive macrophages. Intraperitoneal injection of casein, Incomplete Freund's Adjuvant (IFA) and live or formol-killed Yersinia ruckeri resulted in a rapid influx of neutrophils, peaking at 24 to 48 h post-injection and reaching values, in the case of live bacteria, 500 x those in the resting, unstimulated peritoneal cavity. Peritoneal macrophages also increased, but the response was slower (peak at 5 d) and with more modest increases in number (7.5 x). Neutrophil and mononuclear cells returned to normal values after 15 d in the case of casein and bacteria, but continued above base values 30 d after the injection of IFA. Conversely, after the injection of phosphate buffered saline, India ink or with sham-injections, very moderate neutrophil and and macrophage responses subsided in a few hours. Phagocytosis of bacteria was studied by light microscopy of preparations stained for peroxidase by a new method which allows for the simultaneous observation of intracellular bacteria and peroxidase staining. When Y. ruckeri was injected into resting peritoneal cavities, bacteria were ingested by the resident macrophages. When the bacteria were injected into cavities with high numbers of neutrophils (due to the previous injection of casein), more neutrophils than macrophages contained bacteria. Results show that the macrophages are the resident phagocytes of the peritoneal cavity of trout, while neutrophils are present in that body cavity in significant numbers only in situations of inflammation and only as long as the inflammation persists.
利用光学和电子显微镜细胞化学技术，对虹鳟鱼（Oncorhynchus mykiss）腹腔内伴随炎症的中性粒细胞和巨噬细胞反应进行了研究。炎症性腹腔渗出液中的中性粒细胞α-萘丁酸酯酶呈阴性、过氧化物酶呈阳性且富含细胞质糖原颗粒。巨噬细胞糖原含量低、酯酶呈阳性且通常过氧化物酶呈阴性。一些过氧化物酶呈阳性的巨噬细胞是由于中性粒细胞过氧化物酶转移至巨噬细胞所致。糖原-过氧化物酶或酯酶/过氧化物酶的超微结构双重标记对于精确表征炎症腹腔中的中性粒细胞和巨噬细胞以及正确标记过氧化物酶呈阳性的巨噬细胞最为有用。腹腔注射酪蛋白、不完全弗氏佐剂（IFA）以及活的或经甲醛灭活的鲁氏耶尔森菌会导致中性粒细胞迅速涌入，在注射后24至48小时达到峰值，对于活细菌而言，其数量达到静息、未受刺激腹腔中的500倍。腹腔巨噬细胞也会增加，但反应较慢（在第5天达到峰值）且数量增加幅度较小（7.5倍）。在注射酪蛋白和细菌的情况下，中性粒细胞和单核细胞在15天后恢复至正常值，但在注射IFA后30天仍高于基础值。相反，注射磷酸盐缓冲盐水、印度墨水或进行假注射后，非常轻微的中性粒细胞和巨噬细胞反应在数小时内消退。通过一种新方法对制备物进行过氧化物酶染色的光学显微镜观察来研究细菌的吞噬作用，该方法允许同时观察细胞内细菌和过氧化物酶染色。当将鲁氏耶尔森菌注入静息腹腔时，细菌被驻留巨噬细胞摄取。当将细菌注入含有大量中性粒细胞的腹腔（由于先前注射了酪蛋白）时，含有细菌的中性粒细胞比巨噬细胞更多。结果表明，巨噬细胞是鳟鱼腹腔中的驻留吞噬细胞，而中性粒细胞仅在炎症情况下且仅在炎症持续期间才大量存在于该体腔中。
