Smatlikova Petra, Juhas Stefan, Juhasova Jana, Suchy Tomas, Hubalek Kalbacova Marie, Ellederova Zdenka, Motlik Jan, Klima Jiri
PIGMOD Centre, Laboratory of Cell Regeneration and Plasticity, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences, Libechov, Czech Republic.
Department of Cell Biology, Faculty of Science, Charles University in Prague, Czech Republic.
J Huntingtons Dis. 2019;8(1):33-51. doi: 10.3233/JHD-180303.
Although the highest expression of mutant huntingtin (mtHtt) was observed in the brain, its negative effects were also apparent in other tissues. Specifically, mtHtt impairs metabolic homeostasis and causes transcriptional dysregulation in adipose tissue. Adipogenic differentiation can be induced by the activation of two transcription factors: CCAAT/enhancer-binding protein alpha (CEBPα) and peroxisome proliferator-activated receptor gamma (PPARγ). These same transcription factors were found to be compromised in some tissues of Huntington's disease (HD) mouse models and in lymphocytes of HD patients.
This study investigated the adipogenic potential of mesenchymal stem cells (MSCs) derived from transgenic Huntington's disease (TgHD) minipigs expressing human mtHtt (1-548aa) containing 124 glutamines. Two differentiation conditions were used, employing PPARγ agonist rosiglitazone or indomethacin.
Bone marrow MSCs were isolated from TgHD and WT minipig siblings and compared by their cluster of differentiation using flow cytometry. Their adipogenic potential in vitro was analyzed using quantitative immunofluorescence and western blot analysis of transcription factors and adipogenic markers.
Flow cytometry analysis did not reveal any significant difference between WT and TgHD MSCs. Nevertheless, following differentiation into adipocytes, the expression of CEBPα nuclear, PPARγ and adipogenic marker FABP4/AP2 were significantly lower in TgHD cells compared to WT cells. In addition, we proved both rosiglitazone and indomethacin to be efficient for adipogenic differentiation of porcine MSCs, with rosiglitazone showing a better adipogenic profile.
We demonstrated a negative influence of mtHtt on adipogenic differentiation of porcine MSCs in vitro associated with compromised expression of adipogenic transcription factors.
尽管突变型亨廷顿蛋白（mtHtt）在大脑中的表达最高，但其负面影响在其他组织中也很明显。具体而言，mtHtt会损害代谢稳态并导致脂肪组织中的转录失调。脂肪生成分化可由两种转录因子的激活诱导：CCAAT/增强子结合蛋白α（CEBPα）和过氧化物酶体增殖物激活受体γ（PPARγ）。在亨廷顿舞蹈病（HD）小鼠模型的某些组织以及HD患者的淋巴细胞中，发现这些相同的转录因子受到损害。
本研究调查了源自表达含124个谷氨酰胺的人mtHtt（1-548aa）的转基因亨廷顿舞蹈病（TgHD）小型猪的间充质干细胞（MSC）的脂肪生成潜力。使用了两种分化条件，分别采用PPARγ激动剂罗格列酮或吲哚美辛。
从TgHD和野生型（WT）小型猪同胞中分离出骨髓间充质干细胞，并通过流式细胞术对其分化簇进行比较。使用定量免疫荧光以及转录因子和脂肪生成标志物的蛋白质印迹分析，对它们在体外的脂肪生成潜力进行分析。
流式细胞术分析未发现WT和TgHD间充质干细胞之间存在任何显著差异。然而，在分化为脂肪细胞后，与WT细胞相比，TgHD细胞中CEBPα核、PPARγ和脂肪生成标志物FABP4/AP2的表达显著降低。此外，我们证明罗格列酮和吲哚美辛对猪间充质干细胞的脂肪生成分化均有效，其中罗格列酮显示出更好的脂肪生成情况。
我们证明了mtHtt对猪间充质干细胞体外脂肪生成分化有负面影响，这与脂肪生成转录因子的表达受损有关。
