中华心血管病杂志
中華心血管病雜誌
중화심혈관병잡지
Chinese Journal of Cardiology
2014年
5期
428-432
,共5页
孔清%高梦莎%薛贻敏%潘晓芬%赖文盈%伍伟锋
孔清%高夢莎%薛貽敏%潘曉芬%賴文盈%伍偉鋒
공청%고몽사%설이민%반효분%뢰문영%오위봉
心肌炎%白细胞介素-17%白细胞介素-27
心肌炎%白細胞介素-17%白細胞介素-27
심기염%백세포개소-17%백세포개소-27
Myocarditis%Interleukin-17%Interleukin-27
目的 探讨白细胞介素(interleukin,IL)-17在病毒性心肌炎(VMC)小鼠发病中的作用.方法 IL-17A基因敲除(IL-17A-deficient,IL-17A-/-)小鼠和野生型(wild-type,WT) BALB/c小鼠分别腹腔注射柯萨奇病毒B3(CVB3)建立小鼠VMC模型,于注射病毒后第7天,苏木素伊红染色(HE)观察心肌组织形态,实时荧光定量逆转录聚合酶链反应检测心肌组织中IL-27 mRNA的表达水平,酶联免疫吸附试验检测心肌组织IL-27蛋白的表达水平.此外,WT小鼠感染病毒1周后分离提取脾脏淋巴细胞及腹腔巨噬细胞,用0和25 ng/ml重组IL-17(rIL-17)培养淋巴细胞24 h,用0和10 ng/ml rIL-17培养巨噬细胞48 h,实时荧光定量逆转录聚合酶链反应测定培养的淋巴细胞及巨噬细胞中IL-27 mRNA的表达水平,酶联免疫吸附试验检测淋巴细胞、巨噬细胞培养上清液中IL-27的蛋白表达水平.结果 IL-17A-/-小鼠感染病毒后心肌炎症程度轻于WT小鼠(病理积分为0.9±0.3比1.9±0.5,P＜0.05),且IL-27蛋白的表达水平及IL-27亚基p28 mRNA相对表达量均低于WT小鼠[(72±18) pg/ml比(95±25) pg/ml和1.11 ±0.24比3.10±0.80,P均＜0.05].25 ng/ml rIL-17培养的淋巴细胞中IL-27 p28 mRNA相对表达量及蛋白含量与0 ng/ml rIL-17培养的淋巴细胞比较差异均无统计学意义[分别为1.32±0.21比1.02±0.13和(52±11) pg/ml比(49±9)pg/ml,P均＞0.05].10 ng/ml rIL-17培养的巨噬细胞中IL-27 p28 mRNA相对表达量及蛋白表达水平均明显高于0 ng/ml rIL-17培养的巨噬细胞[分别为8.5±3.1比2.2±0.7和(368±95) pg/ml比(150±38)pg/ml,P均＜0.05].结论 在VMC小鼠中,IL-17促进了IL-27的表达,巨噬细胞可能是IL-17调节IL-27产量的重要靶细胞之一,而淋巴细胞可能与此无关.促炎因子IL-17与抑炎因子IL-27可能参与了VMC的发病机制.
目的 探討白細胞介素(interleukin,IL)-17在病毒性心肌炎(VMC)小鼠髮病中的作用.方法 IL-17A基因敲除(IL-17A-deficient,IL-17A-/-)小鼠和野生型(wild-type,WT) BALB/c小鼠分彆腹腔註射柯薩奇病毒B3(CVB3)建立小鼠VMC模型,于註射病毒後第7天,囌木素伊紅染色(HE)觀察心肌組織形態,實時熒光定量逆轉錄聚閤酶鏈反應檢測心肌組織中IL-27 mRNA的錶達水平,酶聯免疫吸附試驗檢測心肌組織IL-27蛋白的錶達水平.此外,WT小鼠感染病毒1週後分離提取脾髒淋巴細胞及腹腔巨噬細胞,用0和25 ng/ml重組IL-17(rIL-17)培養淋巴細胞24 h,用0和10 ng/ml rIL-17培養巨噬細胞48 h,實時熒光定量逆轉錄聚閤酶鏈反應測定培養的淋巴細胞及巨噬細胞中IL-27 mRNA的錶達水平,酶聯免疫吸附試驗檢測淋巴細胞、巨噬細胞培養上清液中IL-27的蛋白錶達水平.結果 IL-17A-/-小鼠感染病毒後心肌炎癥程度輕于WT小鼠(病理積分為0.9±0.3比1.9±0.5,P＜0.05),且IL-27蛋白的錶達水平及IL-27亞基p28 mRNA相對錶達量均低于WT小鼠[(72±18) pg/ml比(95±25) pg/ml和1.11 ±0.24比3.10±0.80,P均＜0.05].25 ng/ml rIL-17培養的淋巴細胞中IL-27 p28 mRNA相對錶達量及蛋白含量與0 ng/ml rIL-17培養的淋巴細胞比較差異均無統計學意義[分彆為1.32±0.21比1.02±0.13和(52±11) pg/ml比(49±9)pg/ml,P均＞0.05].10 ng/ml rIL-17培養的巨噬細胞中IL-27 p28 mRNA相對錶達量及蛋白錶達水平均明顯高于0 ng/ml rIL-17培養的巨噬細胞[分彆為8.5±3.1比2.2±0.7和(368±95) pg/ml比(150±38)pg/ml,P均＜0.05].結論 在VMC小鼠中,IL-17促進瞭IL-27的錶達,巨噬細胞可能是IL-17調節IL-27產量的重要靶細胞之一,而淋巴細胞可能與此無關.促炎因子IL-17與抑炎因子IL-27可能參與瞭VMC的髮病機製.
목적 탐토백세포개소(interleukin,IL)-17재병독성심기염(VMC)소서발병중적작용.방법 IL-17A기인고제(IL-17A-deficient,IL-17A-/-)소서화야생형(wild-type,WT) BALB/c소서분별복강주사가살기병독B3(CVB3)건립소서VMC모형,우주사병독후제7천,소목소이홍염색(HE)관찰심기조직형태,실시형광정량역전록취합매련반응검측심기조직중IL-27 mRNA적표체수평,매련면역흡부시험검측심기조직IL-27단백적표체수평.차외,WT소서감염병독1주후분리제취비장림파세포급복강거서세포,용0화25 ng/ml중조IL-17(rIL-17)배양림파세포24 h,용0화10 ng/ml rIL-17배양거서세포48 h,실시형광정량역전록취합매련반응측정배양적림파세포급거서세포중IL-27 mRNA적표체수평,매련면역흡부시험검측림파세포、거서세포배양상청액중IL-27적단백표체수평.결과 IL-17A-/-소서감염병독후심기염증정도경우WT소서(병리적분위0.9±0.3비1.9±0.5,P＜0.05),차IL-27단백적표체수평급IL-27아기p28 mRNA상대표체량균저우WT소서[(72±18) pg/ml비(95±25) pg/ml화1.11 ±0.24비3.10±0.80,P균＜0.05].25 ng/ml rIL-17배양적림파세포중IL-27 p28 mRNA상대표체량급단백함량여0 ng/ml rIL-17배양적림파세포비교차이균무통계학의의[분별위1.32±0.21비1.02±0.13화(52±11) pg/ml비(49±9)pg/ml,P균＞0.05].10 ng/ml rIL-17배양적거서세포중IL-27 p28 mRNA상대표체량급단백표체수평균명현고우0 ng/ml rIL-17배양적거서세포[분별위8.5±3.1비2.2±0.7화(368±95) pg/ml비(150±38)pg/ml,P균＜0.05].결론 재VMC소서중,IL-17촉진료IL-27적표체,거서세포가능시IL-17조절IL-27산량적중요파세포지일,이림파세포가능여차무관.촉염인자IL-17여억염인자IL-27가능삼여료VMC적발병궤제.
Objective Interleukin-27 (IL-27) has been reported to reduce the levels of interleukin17(IL-17) and alleviate the severity of experimental autoimmune myocarditis.IL-17,an important tissueprotective cytokine in viral myocarditis (VMC),has been reported to increase synovial expression of IL-27 in rheumatoid arthritis.However,the influence of IL-17 on IL-27 expression in murine model of VMC remains unknown.Methods Wild-type (WT) and IL-17A-deficient (IL-17A-/-) mice on the BALB/c background were intraperitoneally (i.p) injected with coxsackievirus B3 (CVB3) for establishing VMC models.Cardiac tissue was obtained on day 7 after CVB3 injection.Myocardial histopathologic changes were observed by hematoxylin-eosin (HE) stained myocardial sections.Expression of IL-27 in heart and serum was measured by real-time reverse transcription-polymerase chain reaction (RT-PCR) and enzyme linked immunosorbent assay (ELISA),respectively.Furthermore,splenic lymphocytes and peritoneal macrophages were purified 1 week after injection from WT mice.Isolated lymphocytes were cultured in the presence of different concentrations (0 and 25 ng/ml) of recombinant IL-17 (rIL-17) for 24 h.Macrophages were cultured with different concentrations of rIL-17 (0 and 10 ng/ml) for 48 h.IL-27 mRNA expression of cultured cells was assayed by RT-PCR,and their protein level in the culture supernatant was measured by ELISA.Results Compared with WT mice,significantly less cardiac inflammation was evidenced in the heart of IL-17A-/-mice (0.9 ± 0.3 vs.1.9 ± 0.5),relative cardiac IL-27 p28 mRNA expressions (1.11 ± 0.24 vs.3.1 ± 0.8) and serum IL-27 protein [(72 ± 18) pg/ml vs.(95 ± 25) pg/ml] were also significantly lower in IL-17A-/-mice (all P ＜ 0.05).In the culture lymphocytes,the relative mRNA (1.02 ± 0.13 vs.1.32 ± 0.21) and protein [(49 ± 9) pg/ml vs.(52 ± 11) pg/ml] expressions of IL-27 p28 and were similar post treatment with 0 and 25 ng/ml rIL-17 (all P ＞0.05).Compared with 0 ng/ml rIL-17 culture with macrophages,higher relative mRNA (8.5 ±3.1 vs.2.2 ±0.7) and protein [(368 ±95) pg/ml vs.(150 ± 38) pg/ml] expressions of IL-27 p28 were detected in 10 ng/ml rIL-17 group (all P ＜0.05).Conclusion Our data indicates that cytokine IL-17 may contribute to the secretion of IL-27 in VMC mice.Furthermore,macrophages but not lymphocytes may be the important IL-27-producing immune cells and major target cells for IL-17.Thus,IL-27 and IL-17 might be actively involved in the pathogenesis of VMC.
