72只泌乳期SD雌性大鼠随机均分成对照组和试验组(n=36),于产后0h试验组每只肌注CpG-DNA200μg,对照组肌注0.01mol·L^-1 PBS(pH7.2)100μL。两组于产后72h均经乳头管灌注2×10^12 CFU·mL^-1金黄色葡萄球菌,每侧各100μL到第4对乳腺内(两侧);分别于灌注前0h,灌注后8、16、24、48和72h(n=6)颈静脉放血处死。组织学观察显示,试验组乳腺腺泡内嗜中性粒细胞(PMN)在感染初期较对照组浸润迅速,试验组在72h时已无PMN浸润,而对照组仍有浸润。与对照组相比,CpG-DNA能显著降低8、16和72h的乳腺组织细菌数。IL-2水平在灌注后有显著下降,但与对照组相比CpG-DNA极显著提高乳腺组织0h IL-2水平,显著提高16、24和48h乳腺组织IL-6水平,显著降低48h和72h乳腺组织中N-乙酰-β-D-氨基葡萄糖苷酶(NAGase)活力。上述结果显示,CpG-DNA可促进炎症初期PMN的快速浸润,促进并加速细胞因子的产生及释放,减少乳腺组织细菌数量,减轻炎症介质对细胞的损伤,表明CpG-DNA对金黄色葡萄球菌感染诱发的大鼠乳腺炎有保护作用。
Seventy two lactating SD rats were randomly divided into control and treatment group. The 100 μL of sterile 0.01 mol·L^-1, pH 7.2 PBS (control group), CpG-DNA 200 μg per rat (treatment group) were injected intramuscularly into tibialis anterior of the left leg after parturition (0 h) respectively. Then 100 μL per rat of bacterial suspension containing 2×10^12 CFU·mL^-1 (both group) of Staphylococcus aureus were inoculated into the fourth (two sides) mammary gland via the teat duct 72 hours after parturition respectively. Before and after 8 h, 16 h, 24 h, 48 h and 72 h (n = 6) of inoculation, all the rats euthanatized and the mammary glands and serum were harvested. The histopathologic evaluations showed CpG-DNA elicited more polymorphonuclear leukocytes (PMN) accumulation in alveoli at the initial stage of the infection. There were no PMN found in alveoli 72 h postinfection in treatment group and however some PMN were still found in control. CpG-DNA induced significant decrease of viable bacteria counts in mammary gland at 8, 16 and 72 h postinfection. Significant decreases in interleukine 2 (IL-2) was observed at different time points in both groups. CpG-DNA induced significant elevated IL-2 before infection compared with control. The peak levels of IL-6 in mammary gland was observed at 48 h postinfection in control and 24 h in treatment. CpG-DNA could induce significant increase of IL-6 in mammary gland at 16, 24 and 48 h postinfection, and could reduce the activity of N-acetyl-β-D-glucose-aminidase (NAGase) significantly in mammary gland at 48 h and 72 h postinfeetion. The result showed that CpG-DNA induced more prompt migration of PMN from blood to mammary gland at the initial stage of S. aureus infection, stimulated the secretion of cytokines, decreased bacteria counts in mammary gland and attenuate the destroy of inflammation-mediator to cell. The study indicated that GpG-DNA could protect mammary gland against mastitis by S. aureus infection in rat.