脂多醣刺激之腹腔巨噬細胞外加介白質-10 可抑制其腫瘤壞死因子-α及介白質-1ß而非介白質-6之分泌   全文下載

Interleukin-10 Administration Inhibits TNF-α and IL-1β, but Not IL-6, Secretion of LPS-Stimulated Peritoneal Macrophages

本研究旨在探討發炎細胞激素介白質-10，額外添加至脂多醣刺激的腹腔巨噬細胞培養基中，對其促發炎細胞激素介白質-1β、介白質-6及腫瘤壞死因子（tumor necrosis factor，TNF）-α分泌之影響。實驗進行首先自BALB/c雌鼠之腹腔取得初代腹腔巨噬細胞，並以脂多醣（ lipopolysaccharide，LPS）刺激以活化巨噬細胞，在48 hr培養期間內的不同時向點，測試培養液中抗發炎及促發炎細胞激素分泌量的變化。結果發現，隨培養時問增加，LPS刺激所產生之介白質-1β、6、10及腫瘤壞死因子-α的濃度亦隨之增加，其分泌濃度分別為介白質-6 ＞ 腫瘤壞死因子-α ＞ 介白質-10 ＞ 介白質-1β。在培養18 hr後，內生性介白質-10含量下降，為探討外加介白質-10之抗發炎作用，因此選擇在脂多醣刺激巨噬細胞後18 h，外加不同濃度（0、75、150及225 pg/mL）抗發炎細胞激素介白質-10至細胞培養液中，並持續培養至48 hr，以觀察外加介白質-10對已活化巨噬細胞分泌細胞激素之影響，結果發現，外加介白質-10 抑制介白質-1β（21.3~38.6%）及腫瘤壞死因子-α（44.7~66.8%）之分泌量，且以添加低劑量（75 pg/mL）介白質-10有最大效果，但對介白質-6之分泌別無顯著影響。另外發現，外加介白質-10，可增加培養液中總介白質-10之濃度（18.4~35.5%），但只有外加低劑量介白質-10可增加內生性介白質-10之分泌量10.4%，外加較高劑量之介白質-10，反而抑制內生性介白質-10之分泌量達31.7至41.9%。綜合本實驗結果推測，添加低劑量介白質-10可經由抑制腫瘤壞死因子-α及介白質-1β，而非介白質-6之分泌，同時刺激內生性介白質-10之分泌，而達到抗發炎之功效。

We hypothesize that exogenous administration of anti-inflammatory cytokine interleukin (IL)-10 would affect the secretion of pro-inflammatory cytokines such as IL-1β, IL-6, or tumor necrosis factor (TNF)–α using lipopolysaccharide (LPS)-stimulated macrophages exuded from the peritoneal cavity of female BALB/c mice. The LPS-stimulated macrophages were incubated for 48 hr to investigate the secretions of pro- and anti-inflammatory cytokine. The results indicated that the secretion levels of IL-1β, IL-6, IL-10, and TNF–α by LPS-stimulated macrophages elevated in a time dependent manner during 48-hr incubation period. The amount of cytokines secretion by LPS-stimulated macrophages varied: IL-6 > TNF-α > IL-10 > IL-1β. However, the secretion of IL-10 started to decrease at 18 hr of incubation. Therefore, the macrophage cultures were extra-administrated with various concentrations (0, 75, 150, and 225 pg/mL) of exogenous IL-10 after 18-hr incubation in order to evaluate the effect of exogenous IL-10 administration on inflammation. IL-10, IL-1β, IL-6, and TNF–α in LPS-stimulated macrophage cultures before and after exogenous IL-10 administration were determined by ELISA. The results demonstrated that exogenous IL-10 administration inhibited IL-1β (21.3-38.6%) and TNF–α (44.7-66.8%) secretion. However, the administration did not inhibit IL-6 secretion. Low dose (75 pg/mL) of exogenous IL-10 exerted maximal effects. IL-10 levels in LPS-stimulated macrophage cultures after exogenous IL-10 administration increased from 18.4% to 35.5%. Furthermore, only low dose of exogenous IL-10 administration increased endogenous IL-10 secretion (by 10.4%), while higher doses of exogenous IL-10 inhibited endogenous IL-10 production (from 31.7% to 41.9%). The results suggest that low dose administration of exogenous IL-10 might exhibit antiinflammatory effects via inhibiting TNF-α and IL-1β, but not IL-6, secretion and increasing endogenous IL-10 production by LPS-stimulated peritoneal macrophages.