宫内生长受限(IUGR)与由于成肌细胞固有功能缺陷而导致的肌肉质量终身减少有关。但这些缺陷背后的机制尚不清楚。我们的目的是确定缺陷是否与IUGR成肌细胞的炎症和肾上腺素能调节的变化有关。如先前在IUGR肌肉中观察到的。从由高温诱导的胎盘功能不全(PI-IUGR;n=9)产生的IUGR胎羊及其对照(n=9)和由母胎炎症(MI-IUGR;n=6)产生的IUGR胎羊及其对照(n=7)中分离出原代成肌细胞。PI-IUGR成肌细胞的增殖率低于对照组(P<0.05),并且不受IL-6,TNF-α,去甲肾上腺素,或胰岛素。IκB激酶抑制在基础培养基中适度降低(P<0.05)对照成肌细胞的增殖,但在添加TNF-α的培养基中显著降低(P<0.05),在基础培养基和添加TNF-α的培养基中显著降低(P<0.05)PI-IUGR成肌细胞的增殖。MI-IUGR成肌细胞的增殖大于其对照(P<0.05),并且不受与TNF-α孵育的影响。胰岛素增加(P<0.05)MI-IUGR和对照成肌细胞的增殖。分化72小时后,在基础培养基和IL-6添加培养基中,与对照组相比,更少的PI-IUGR成肌细胞(P<0.05)。仅在基础培养基中,PI-IUGR成肌细胞的结蛋白少于对照组(P<0.05)。与去甲肾上腺素一起孵育并不影响肌生成素+或结蛋白+百分比,但胰岛素在对照和PI-IUGR成肌细胞中的标志物均增加(P<0.05)。96小时分化后,无论培养基如何,MI-IUGR成肌细胞均比对照组少(P<0.05),虽然TNF-α降低(P<0.05)结蛋白+成肌细胞。分化的PI-IUGR成肌细胞具有更高的(P<0.05)TNFR1、ULK2和TNF-α刺激的TLR4基因表达,PI-IUGR半腱肌TNFR1和IL6基因表达量较高(P<0.05),更多(P<0.05)c-Fos蛋白,IκBα蛋白较少(P<0.05)。分化的MI-IUGR成肌细胞有更大的(P<0.05)TNFR1和IL6R基因表达,ULK2基因表达倾向于更高(P=0.07),β-catenin蛋白和TNF-α刺激的NFκB磷酸化程度更高(P<0.05)。我们得出的结论是,IUGR成肌细胞中TNF-α/TNFR1/NFκB和其他炎症途径的这些富集成分导致其功能障碍,并有助于解释IUGR胎儿肌肉生长受损。
成肌细胞是干细胞,其功能能力可以限制肌肉生长。然而,紧张的宫内条件导致这些细胞本质上功能失调。这限制了肌肉的生长能力,导致胎儿宫内生长受限(IUGR),低出生体重,出生后肌肉量减少。因此,IUGR出生的食用动物的肉产量降低,IUGR出生的人类的葡萄糖稳态受损,这有助于代谢功能障碍。先前已观察到IUGR成肌细胞的内在功能障碍,但是胎儿编程发生了变化(即,解释不同功能结果的细胞机制发展的永久性变化)尚未确定。这项研究表明,一种机制是TNF-α和其他炎症细胞因子的信号通路增强。这些细胞因子在应激反应和肌肉生长调节中起作用。这些途径的程序性增强意味着IUGR成肌细胞甚至对正常量的循环细胞因子更有反应。不幸的是,成肌细胞对细胞因子的主要反应是分化较慢(即,肌肉生长所需的细胞转化)。这种反应的程序增强直接阻碍成肌细胞依赖性肌肉生长,赤字是终身的。然而,确定这种机制是制定策略以改善低出生体重后代肌肉生长的基本步骤。
Intrauterine growth restriction (IUGR) is linked to lifelong reductions in muscle mass due to intrinsic functional deficits in myoblasts, but the mechanisms underlying these deficits are not known. Our objective was to determine if the deficits were associated with changes in inflammatory and adrenergic regulation of IUGR myoblasts, as was previously observed in IUGR muscle. Primary myoblasts were isolated from IUGR fetal sheep produced by hyperthermia-induced placental insufficiency (PI-IUGR; n = 9) and their controls (n = 9) and from IUGR fetal sheep produced by maternofetal inflammation (MI-IUGR; n = 6) and their controls (n = 7). Proliferation rates were less (P < 0.05) for PI-IUGR myoblasts than their controls and were not affected by incubation with IL-6, TNF-α, norepinephrine, or insulin. IκB kinase inhibition reduced (P < 0.05) proliferation of control myoblasts modestly in basal media but substantially in TNF-α-added media and reduced (P < 0.05) PI-IUGR myoblast proliferation substantially in basal and TNF-α-added media. Proliferation was greater (P < 0.05) for MI-IUGR myoblasts than their controls and was not affected by incubation with TNF-α. Insulin increased (P < 0.05) proliferation in both MI-IUGR and control myoblasts. After 72-h differentiation, fewer (P < 0.05) PI-IUGR myoblasts were myogenin+ than controls in basal and IL-6 added media but not TNF-α-added media. Fewer (P < 0.05) PI-IUGR myoblasts were desmin+ than controls in basal media only. Incubation with norepinephrine did not affect myogenin+ or desmin+ percentages, but insulin increased (P < 0.05) both markers in control and PI-IUGR myoblasts. After 96-h differentiation, fewer (P < 0.05) MI-IUGR myoblasts were myogenin+ and desmin+ than controls regardless of media, although TNF-α reduced (P < 0.05) desmin+ myoblasts for both groups. Differentiated PI-IUGR myoblasts had greater (P < 0.05) TNFR1, ULK2, and TNF-α-stimulated TLR4 gene expression, and PI-IUGR semitendinosus muscle had greater (P < 0.05) TNFR1 and IL6 gene expression, greater (P < 0.05) c-Fos protein, and less (P < 0.05) IκBα protein. Differentiated MI-IUGR myoblasts had greater (P < 0.05) TNFR1 and IL6R gene expression, tended to have greater (P = 0.07) ULK2 gene expression, and had greater (P < 0.05) β-catenin protein and TNF-α-stimulated phosphorylation of NFκB. We conclude that these enriched components of TNF-α/TNFR1/NFκB and other inflammatory pathways in IUGR myoblasts contribute to their dysfunction and help explain impaired muscle growth in the IUGR fetus.
Myoblasts are stems cells whose functional capacity can limit muscle growth. However, stressful intrauterine conditions cause these cells to be intrinsically dysfunctional. This restricts muscle growth capacity, leading to intrauterine growth restriction (IUGR) of the fetus, low birth weight, and less muscle mass after birth. Consequently, meat yield is reduced in IUGR-born food animals and glucose homeostasis is impaired in IUGR-born humans, which contributes to metabolic dysfunction. Intrinsic dysfunction of IUGR myoblasts has been previously observed, but the fetal programming changes (i.e., permanent changes in the development of cellular mechanisms that explains different functional outcomes) have not been identified. This study shows that one mechanism is the enhancement of signaling pathways for TNF-α and other inflammatory cytokines. These cytokines have roles in stress responses and regulation of muscle growth. Programmed enhancement of these pathways means that IUGR myoblasts are more responsive to even normal amounts of circulating cytokines. Unfortunately, the primary response of myoblasts to cytokines is slower differentiation (i.e., cellular transformation necessary for muscle growth). Programmed enhancement of this response directly impedes myoblast-dependent muscle growth, and the deficit is lifelong. However, identifying this mechanism is a fundamental step for developing strategies to improve muscle growth in low birth weight offspring.