中国组织工程研究
中國組織工程研究
중국조직공정연구
Journal of Clinical Rehabilitative Tissue Engineering Research
2015年
10期
1556-1561
,共6页
张继峰%张紫琦%雒晓甜%侯林义%姜琴%吕洁萍%张文凯
張繼峰%張紫琦%雒曉甜%侯林義%薑琴%呂潔萍%張文凱
장계봉%장자기%락효첨%후림의%강금%려길평%장문개
干细胞%移植%肺损伤%骨髓间充质干细胞%脓毒症%NF-κB%巨噬细胞炎性蛋白2
榦細胞%移植%肺損傷%骨髓間充質榦細胞%膿毒癥%NF-κB%巨噬細胞炎性蛋白2
간세포%이식%폐손상%골수간충질간세포%농독증%NF-κB%거서세포염성단백2
Bone Marrow%Mesenchymal Stem Cel Transplantation%Sepsis%Lung Injury%NF-kappa B%Macrophage Inflammatory Proteins
背景：骨髓间充质干细胞对急性肺损伤具有治疗作用，但其机制尚不清楚，探究其机制可使广大急性肺损伤患者获益。目的：探讨骨髓间充质干细胞治疗大鼠脓毒症急性肺损伤的可能机制。方法：①36只成年Wistar大鼠随机均分为3组：假手术组、脓毒症组和骨髓间充质干细胞治疗组，后2组采用盲肠结扎穿孔术建立脓毒症急性肺损伤模型，假手术组大鼠仅翻动盲肠，不结扎穿孔。骨髓间充质干细胞治疗组术后经股静脉注射骨髓间充质干细胞悬液1 mL(1×109L-1)，其他2组同法注射生理盐水1 mL，6 h后测定3组大鼠血清中白细胞介素10、巨噬细胞炎性蛋白2水平，取肺组织进行苏木精-伊红染色，光镜下观察肺组织病理结构的改变。②支气管肺泡灌洗法获取大鼠肺泡巨噬细胞，所获肺泡巨噬细胞接种于24孔培养板，分为3组：对照组加入生理盐水，脓毒症模型组加入脓毒症大鼠血浆，骨髓间充质干细胞干预组加入脓毒症大鼠血浆和骨髓间充质干细胞共同干预，于37℃、体积分数为5%CO 2培养箱孵育1 h。留取肺泡巨噬细胞，采用激光扫描共聚焦显微镜检测肺泡巨噬细胞NF-κB(P65)蛋白入核情况。结果与结论：①动物实验：与假手术组相比，脓毒症组、骨髓间充质干细胞治疗组血清巨噬细胞炎性蛋白2水平升高(P <0.05)，但骨髓间充质干细胞治疗组巨噬细胞炎性蛋白2水平明显低于脓毒症组(P <0.05)；各组血清白细胞介素10水平差异无显著性意义(P>0.05)；脓毒症组和骨髓间充质干细胞治疗组肺内大量炎性细胞浸润、肺间质水肿、肺内出血，但骨髓间充质干细胞治疗组症状较脓毒症组有所减轻。②细胞实验：与对照组相比，脓毒症模型组、骨髓间充质干细胞干预组肺泡巨噬细胞的NF-κB(P65)蛋白入核明显升高(P <0.05)，但骨髓间充质干细胞干预组明显低于脓毒症模型组(P <0.05)。结果表明骨髓间充质干细胞在脓毒症急性肺损伤时可能调控肺泡巨噬细胞NF-κB(P65)蛋白入核，使其减少促炎细胞因子巨噬细胞炎性蛋白2表达，进而减少中性粒细胞浸润起到肺保护作用，暂不能说明骨髓间充质干细胞对白细胞介素10有影响。
揹景：骨髓間充質榦細胞對急性肺損傷具有治療作用，但其機製尚不清楚，探究其機製可使廣大急性肺損傷患者穫益。目的：探討骨髓間充質榦細胞治療大鼠膿毒癥急性肺損傷的可能機製。方法：①36隻成年Wistar大鼠隨機均分為3組：假手術組、膿毒癥組和骨髓間充質榦細胞治療組，後2組採用盲腸結扎穿孔術建立膿毒癥急性肺損傷模型，假手術組大鼠僅翻動盲腸，不結扎穿孔。骨髓間充質榦細胞治療組術後經股靜脈註射骨髓間充質榦細胞懸液1 mL(1×109L-1)，其他2組同法註射生理鹽水1 mL，6 h後測定3組大鼠血清中白細胞介素10、巨噬細胞炎性蛋白2水平，取肺組織進行囌木精-伊紅染色，光鏡下觀察肺組織病理結構的改變。②支氣管肺泡灌洗法穫取大鼠肺泡巨噬細胞，所穫肺泡巨噬細胞接種于24孔培養闆，分為3組：對照組加入生理鹽水，膿毒癥模型組加入膿毒癥大鼠血漿，骨髓間充質榦細胞榦預組加入膿毒癥大鼠血漿和骨髓間充質榦細胞共同榦預，于37℃、體積分數為5%CO 2培養箱孵育1 h。留取肺泡巨噬細胞，採用激光掃描共聚焦顯微鏡檢測肺泡巨噬細胞NF-κB(P65)蛋白入覈情況。結果與結論：①動物實驗：與假手術組相比，膿毒癥組、骨髓間充質榦細胞治療組血清巨噬細胞炎性蛋白2水平升高(P <0.05)，但骨髓間充質榦細胞治療組巨噬細胞炎性蛋白2水平明顯低于膿毒癥組(P <0.05)；各組血清白細胞介素10水平差異無顯著性意義(P>0.05)；膿毒癥組和骨髓間充質榦細胞治療組肺內大量炎性細胞浸潤、肺間質水腫、肺內齣血，但骨髓間充質榦細胞治療組癥狀較膿毒癥組有所減輕。②細胞實驗：與對照組相比，膿毒癥模型組、骨髓間充質榦細胞榦預組肺泡巨噬細胞的NF-κB(P65)蛋白入覈明顯升高(P <0.05)，但骨髓間充質榦細胞榦預組明顯低于膿毒癥模型組(P <0.05)。結果錶明骨髓間充質榦細胞在膿毒癥急性肺損傷時可能調控肺泡巨噬細胞NF-κB(P65)蛋白入覈，使其減少促炎細胞因子巨噬細胞炎性蛋白2錶達，進而減少中性粒細胞浸潤起到肺保護作用，暫不能說明骨髓間充質榦細胞對白細胞介素10有影響。
배경：골수간충질간세포대급성폐손상구유치료작용，단기궤제상불청초，탐구기궤제가사엄대급성폐손상환자획익。목적：탐토골수간충질간세포치료대서농독증급성폐손상적가능궤제。방법：①36지성년Wistar대서수궤균분위3조：가수술조、농독증조화골수간충질간세포치료조，후2조채용맹장결찰천공술건립농독증급성폐손상모형，가수술조대서부번동맹장，불결찰천공。골수간충질간세포치료조술후경고정맥주사골수간충질간세포현액1 mL(1×109L-1)，기타2조동법주사생리염수1 mL，6 h후측정3조대서혈청중백세포개소10、거서세포염성단백2수평，취폐조직진행소목정-이홍염색，광경하관찰폐조직병리결구적개변。②지기관폐포관세법획취대서폐포거서세포，소획폐포거서세포접충우24공배양판，분위3조：대조조가입생리염수，농독증모형조가입농독증대서혈장，골수간충질간세포간예조가입농독증대서혈장화골수간충질간세포공동간예，우37℃、체적분수위5%CO 2배양상부육1 h。류취폐포거서세포，채용격광소묘공취초현미경검측폐포거서세포NF-κB(P65)단백입핵정황。결과여결론：①동물실험：여가수술조상비，농독증조、골수간충질간세포치료조혈청거서세포염성단백2수평승고(P <0.05)，단골수간충질간세포치료조거서세포염성단백2수평명현저우농독증조(P <0.05)；각조혈청백세포개소10수평차이무현저성의의(P>0.05)；농독증조화골수간충질간세포치료조폐내대량염성세포침윤、폐간질수종、폐내출혈，단골수간충질간세포치료조증상교농독증조유소감경。②세포실험：여대조조상비，농독증모형조、골수간충질간세포간예조폐포거서세포적NF-κB(P65)단백입핵명현승고(P <0.05)，단골수간충질간세포간예조명현저우농독증모형조(P <0.05)。결과표명골수간충질간세포재농독증급성폐손상시가능조공폐포거서세포NF-κB(P65)단백입핵，사기감소촉염세포인자거서세포염성단백2표체，진이감소중성립세포침윤기도폐보호작용，잠불능설명골수간충질간세포대백세포개소10유영향。
BACKGROUND:Bone marrow mesenchymal stem cels have a therapeutic effect on acute lung injury, but the mechanism is unclear. If the mechanism is understood, the majority of patients with acute lung injury can obtain a benefit. OBJECTIVE:To explore the possible mechanism underlying bone marrow mesenchymal stem cels in the treatment of acute lung injury with sepsis in rats. METHODS: (1) Thirty-six adult Wistar rats were randomly divided into three groups, sham operation group (sham group), sepsis group and bone marrow mesenchymal stem cels group (cel treatment group). In the sepsis and cel treatment groups, animal models of sepsis with acute lung injury were established by cecal ligation and puncture, while in the sham group, the cecum was not ligated and punctured. Then, 1 mL normal saline was injected via the femoral vein in the sepsis and sham groups, and 1 mL bone marrow mesenchymal stem cel suspension (1×109/L) was injected into the cel treatment group. After 6 hours, interleukin 10 and macrophage inflammatory protein-2 levels in serum were measured in the three groups. Lung tissues were taken for pathological observation using hematoxylin-eosin staining. (2) Rat alveolar macrophages were obtained by bronchoalveolar lavage, seeded into 24-wel culture plates, and divided into three groups: control group (group A), sepsis model group (group B) and intervention group of bone marrow mesenchymal stem cels (group C). Normal saline, septic plasma, and co-intervention of septic plasma and mesenchymal stem cels were used in the groups A, B, C, respectively. Then, cels in the three groups were cultured in a 5% CO2 incubator at 37℃ for 1 hour. After that, alveolar macrophages were taken to detect whether nuclear factor-κB (P65) protein entered into the nucleus using laser scanning confocal microscopy. RESULTS AND CONCLUSION: (1) The results of animal experiments showed that compared with the sham group, the macrophage inflammatory protein-2 levels in the sepsis group and cel treatment group were significantly increased (P < 0.05), but the macrophage inflammatory protein-2 level in the cel treatment group was significantly lower than that in the sepsis group (P < 0.05); there were no significant differences in serum interleukin 10 levels among the three groups (P > 0.05); inflammatory cel infiltration, interstitial pulmonary edema and pulmonary hemorrhage existed in the sepsis and cel treatment groups, but these symptoms were significantly reduced in the cel treatment group compared with the sepsis group. (2) Results from cel experiments showed that compared with the group A, in group B and group C, the number of nuclear factor-κB (P65) proteins into the nucleus was significantly higher (P < 0.05), but it was lower in the group C than the group B (P < 0.05). These findings indicate that bone marrow mesenchymal stem cels in acute lung injury with sepsis can regulate nuclear factor-κB (P65) protein of alveolar macrophages into the nucleus, reduce expression of macrophage inflammatory protein-2, and thereby play a protective role in the lungvia reducing neutrophil infiltration. Temporarily, this study cannot explain whether bone marrow mesenchymal stem cels have an effect on interleukin 10.
