Department of Anatomy and Cell Biology, Martin Luther University Faculty of Medicine, Halle (Saale), Germany.
Hum Reprod. 2017 Jul 1;32(7):1382-1392. doi: 10.1093/humrep/dex087.
How does a maternal diabetic hyperadiponectineamia affect signal transduction and lipid metabolism in rabbit preimplantation blastocysts?
In a diabetic pregnancy increased levels of adiponectin led to a switch in embryonic metabolism towards a fatty acid-dependent energy metabolism, mainly affecting genes that are responsible for fatty acid uptake and turnover.
Although studies in cell culture experiments have shown that adiponectin is able to regulate lipid metabolism via 5'-AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor α (PPARα), data on the effects of adiponectin on embryonic lipid metabolism are not available. In a diabetic pregnancy in rabbits, maternal adiponectin levels are elevated fourfold and are accompanied by an increase in intracellular lipid droplets in blastocysts, implying consequences for the embryonic hormonal and metabolic environment.
STUDY DESIGN, SIZE, DURATION: Rabbit blastocysts were cultured in vitro with adiponectin (1 μg/ml) and with the specific AMPK-inhibitor Compound C for 15 min, 1 h and 4 h (N ≥ 3 independent experiments: for RNA analysis, n ≥ 4 blastocysts per treatment group; for protein analysis three blastocysts pooled per sample and three samples used per experiment). Adiponectin signalling was verified in blastocysts grown in vivo from diabetic rabbits with a hyperadiponectinaemia (N ≥ 3 independent experiments, n ≥ 4 samples per treatment group, eight blastocysts pooled per sample).
PARTICIPANTS/MATERIALS, SETTING, METHODS: In these blastocysts, expression of molecules involved in adiponectin signalling [adaptor protein 1 (APPL1), AMPK, acetyl-CoA carboxylase (ACC), p38 mitogen-activated protein kinases (p38 MAPK)], lipid metabolism [PPARα, cluster of differentiation 36 (CD36), fatty acid transport protein 4 (FATP4), fatty acid binding protein (FABP4), carnitine palmityl transferase 1 (CPT1), hormone-senstive lipase (HSL), lipoprotein lipase (LPL)] and members of the insulin/insulin-like growth factor (IGF)-system [IGF1, IGF2, insulin receptor (InsR), IGF1 receptor (IGF1R)] were analyzed by quantitative RT-PCR and western blot. Analyses were performed in both models, i.e. adiponectin stimulated blastocysts (in vitro) and in blastocysts grown in vivo under increased adiponectin levels caused by a maternal diabetes mellitus.
In both in vitro and in vivo models adiponectin increased AMPK and ACC phosphorylation, followed by an activation of the transcription factor PPARα, and CPT1, the key enzyme of β-oxidation (all P < 0.05 versus control). Moreover, mRNA levels of the fatty acid transporters CD36, FATP4 and FABP4, and HSL were upregulated by adiponectin/AMPK signalling (all P < 0.05 versus control). Under diabetic developmental conditions the amount of p38 MAPK was upregulated (P < 0.01 versus non-diabetic), which was not observed in blastocysts cultured in vitro with adiponectin, indicating that the elevated p38 MAPK was not related to adiponectin. However, a second effect of adiponectin has to be noted: its intensification of insulin sensitivity, by regulating IGF availability and InsR/IGF1R expression.
Not applicable.
There are two main limitations for our study. First, human and rabbit embryogenesis can only be compared during blastocyst development. Therefore, the inferences from our findings are limited to the embryonic stages investigated here. Second, the increased adiponectin levels and lack of maternal insulin is only typical for a diabetes mellitus type one model.
This is the first mechanistic study demonstrating a direct influence of adiponectin on lipid metabolism in preimplantation embryos. The numbers of young women with a diabetes mellitus type one are increasing steadily. We have shown that preimplantation embryos are able to adapt to changes in the uterine milieu, which is mediated by the adiponectin/AMPK signalling. A tightly hormonal control during pregnancy is essential for survival and proper development. In this control process, adiponectin plays a more important role than known so far.
STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the German Research Council (DFG RTG ProMoAge 2155), the EU (FP7 Epihealth No. 278418, FP7-EpiHealthNet N°317146), COST Action EpiConcept FA 1201 and SALAAM BM 1308. The authors have no conflict(s) of interest to disclose.
母体糖尿病引起的高 adiponectineamia 如何影响兔胚泡前体信号转导和脂质代谢？
在糖尿病妊娠中，adiponectin 水平升高导致胚胎代谢向脂肪酸依赖性能量代谢转变，主要影响负责脂肪酸摄取和周转的基因。
尽管细胞培养实验研究表明 adiponectin 能够通过 5'-AMP 激活蛋白激酶 (AMPK) 和过氧化物酶体增殖物激活受体 α (PPARα) 调节脂质代谢，但关于 adiponectin 对胚胎脂质代谢影响的数据尚不清楚。在兔的糖尿病妊娠中，母体 adiponectin 水平升高四倍，并伴有胚胎内脂滴增加，这意味着对胚胎激素和代谢环境有影响。
研究设计、规模、持续时间：将兔胚泡在体外与 adiponectin（1μg/ml）和特定的 AMPK 抑制剂 Compound C 培养 15 分钟、1 小时和 4 小时（N≥3 个独立实验：用于 RNA 分析，n≥4 个每个处理组的胚泡；用于蛋白质分析，三个胚泡混合为一个样本，每个实验使用三个样本）。用来自糖尿病高 adiponectinaemia 兔的体内生长的胚泡验证 adiponectin 信号（N≥3 个独立实验，n≥3 个每个处理组的样本，每个样本使用 8 个胚泡）。
参与者/材料、设置、方法：在这些胚泡中，分析了参与 adiponectin 信号转导的分子[衔接蛋白 1 (APPL1)、AMPK、乙酰辅酶 A 羧化酶 (ACC)、p38 丝裂原激活蛋白激酶 (p38 MAPK)]、脂质代谢[过氧化物酶体增殖物激活受体 α (PPARα)、分化簇 36 (CD36)、脂肪酸转运蛋白 4 (FATP4)、脂肪酸结合蛋白 4 (FABP4)、肉碱棕榈酰转移酶 1 (CPT1)、激素敏感脂肪酶 (HSL)、脂蛋白脂肪酶 (LPL)]和胰岛素/胰岛素样生长因子 (IGF)系统的成员[IGF1、IGF2、胰岛素受体 (InsR)、IGF1 受体 (IGF1R)]通过定量 RT-PCR 和 Western blot 进行分析。在两种模型中均进行了分析，即在体外刺激的胚泡和因母体糖尿病引起的体内生长的高 adiponectinaemia 下的胚泡。
在体内和体外模型中，adiponectin 增加了 AMPK 和 ACC 的磷酸化，随后激活了转录因子 PPARα 和β-氧化的关键酶 CPT1（所有 P<0.05 与对照组相比）。此外，脂肪酸转运蛋白 CD36、FATP4 和 FABP4 的 mRNA 水平以及 HSL 也被 adiponectin/AMPK 信号上调（所有 P<0.05 与对照组相比）。在糖尿病发育条件下，p38 MAPK 的量增加（P<0.01 与非糖尿病组相比），但在体外用 adiponectin 培养的胚泡中未观察到，这表明升高的 p38 MAPK 与 adiponectin 无关。然而，adiponectin 还有第二个作用：通过调节 IGF 的可用性和 InsR/IGF1R 的表达来增强胰岛素敏感性。
不适用。
我们的研究有两个主要限制。首先，人和兔的胚胎发生只能在胚泡发育期间进行比较。因此，我们的发现的推论仅限于此处研究的胚胎阶段。其次，增加的 adiponectin 水平和缺乏母体胰岛素仅典型于 1 型糖尿病模型。
这是第一项关于 adiponectin 直接影响兔前体胚胎脂质代谢的机制研究。患有 1 型糖尿病的年轻女性人数稳步增加。我们已经表明，胚胎能够适应子宫内环境的变化，这是由 adiponectin/AMPK 信号介导的。在妊娠期间进行严格的激素控制对于生存和正常发育至关重要。在这个控制过程中，adiponectin 比已知的发挥更重要的作用。
研究资金/竞争利益：这项工作得到了德国研究委员会（DFG RTG ProMoAge 2155）、欧盟（FP7 Epihealth No. 278418、FP7-EpiHealthNet N°317146）、COST 行动 EpiConcept FA 1201 和 SALAAM BM 1308 的支持。作者没有利益冲突。
