人脐带来源间充质干细胞对佐剂性关节炎大鼠巨噬细胞功能的影响

目的:研究人脐带来源间充质干细胞(hUC-MSCs)对佐剂性关节炎大鼠巨噬细胞（macrophage,Mφ）功能影响的初步机制。方法:完全弗氏佐剂法建立佐剂性关节炎大鼠模型(adjuvant arthritis,AA),分离AA大鼠炎症高峰期腹腔中Mφ,体外以Transwell小室法与hUC-MSCs不同比例（1∶1,1∶5,1∶10,1∶50）共培养48 h,流式细胞术检测Mφ的极化情况、中性红法检测Mφ吞噬功能,ELISA法检测共培养上清中细胞因子TNF-α、IL-1β、IL-6和IL-10水平。结果:hUC-MSCs(1∶10,1∶5,1∶1)与Mφ共培养后,与模型组比较,M2型Mφ百分比均值明显增加［(34.0±2.6)%、(33.0±1.9)%、(47.0±7.3)% vs. (24.0±1.9)%］(P<0.01),M1型Mφ的百分比显著降低［(31.0±2.5)%、(30.0±2.7)%、(30.0±1.9)% vs. (39.0±7.1)%］(P<0.01),M2/M1的比值均值相应随共培养hUC-MSCs细胞数的增多而增大(1.2±0.1、1.2±0.2、1.5±0.2 vs. 0.8±0.1)(P<0.05);与模型组比较,hUC-MSCs 不同比例(1∶50,1∶10,1∶5,1∶1)与Mφ共培养还可明显降低Mφ的吞噬指数(2.10±0.18、2.00±0.16、1.50±0.11、1.40±0.11 vs. 2.40±0.20) (P<0.05),明显降低Mφ共培养上清中TNF-α［（143.0±15.0）、(128.0±12.0)、(95.0±8.5)、(44.0±5.9) pg/mL vs. (157.0±14.0) pg/mL］(P<0.01)、IL-1β［(70.0±9.9)、(66.0±5.9)、(54.0±5.2)、(41.0±6.4) pg/mL vs. (84.0±12.0) pg/mL］(P<0.01)和IL-6［(95.0±5.8)、(73.0±4.5)、(65.0±5.1)、(52.0±4.7) pg/mL vs. (109.0±7.6) pg/mL］(P<0.01)的水平,促进IL-10的分泌［(228.0±34.0)、(445.0±55.0)、(568.0±49.0)、(858.0±77.0) pg/mL vs. (85.0±14.0) pg/mL)］(P<0.01)。结论:hUC-MSCs体外可抑制AA大鼠Mφ从M1型向M2型极化,抑制Mφ吞噬功能,降低TNF-α、IL-1β和IL-6水平,升高IL-10水平,对AA大鼠Mφ功能有调节作用。

Human umbilical cord-derived mesenchymal stem cells alleviate adjuvant arthritis by regulating macrophage in vitro

AIM: To study the effect and mechanism of the human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) on the function of rat macrophages in vitro. METHODS: Peritoneal macrophages were isolated from rats with adjuvant arthritis (AA) at 28-30th days and co-cultured with hUC-MSCs（1∶1,1∶5,1∶10,1∶50）for 48 hours.The phagocytic ability of rat macrophages was detected by phagocytosis neutral red method;the percentage of macrophage subtype was detected by flow cytometry;the TNF-α, IL-1β,IL-6 and IL-10 levels in culture supernatant were detected by ELISA.RESULTS:After co-culturing hUC-MSCs (1∶10,1∶5,1∶1) with Mφ,the percentage of M2 macrophages increased significantly ［(34.0±2.6)%,(33.0±1.9)%,(47.0±7.3)% vs. (24.0±1.9)%］(P<0.01),the percentage of M1 macrophages decreased significantly［(31.0±2.5)%,(30.0±2.7)%,(30.0±1.9)% vs. (39.0±7.1)%］(P<0.01),and the average M2/M1 ratio increased with the number of co-cultured hUC-MSCs cells (1.2±0.1,1.2±0.2,1.5±0.2 vs. 0.8±0.1) (P<0.05);Compared with the model group,co-cultivation of hUC-MSCs with different ratios (1∶50, 1∶10,1∶5,1∶1) and Mφ can also significantly reduce the phagocytic index of Mφ(2.10±0.18,2.00±0.16, 1.50±0.11,1.40±0.11 vs. 2.4±0.20) (P<0.05),significantly reduced the levels of TNF-α((143.0±15.0),(128.0±12.0),(95.0±8.5), (44.0±5.9) pg/mL vs. (157.0±14.0) pg/mL)) (P<0.01),IL-1β((70.0±9.9), (66.0±5.9),(54.0±5.2),(41.0±6.4) pg/mL vs. (84.0±12.0) pg/mL)(P<0.01) and IL-6 ((95.0±5.8),(73.0±4.5), (65±5.1),(52.0±4.7)pg/mL vs. (109.0±7.6) pg/mL)(P<0.01),promoting IL-10 secretion ((228.0±34.0), (445.0±55.0),(568.0±49.0),(858.0±77.0) pg/mL vs. (85.0±14.0) pg/mL)(P<0.01).CONCLUSION: hUC-MSCs could regulate the function of AA rats macrophage in vitro with inhibition of the polarization and the phagocytosis, restoration the balance of pro-inflammatory and anti-inflammatory cytokines level.