Almazov National Medical Research Centre, 2 Akkuratova Str., St-Petersburg 197341, Russia.
St-Petersburg State University, 7-9, Universitetskaya nab., St-Petersburg 199034, Russia.
Cells. 2019 Mar 21;8(3):266. doi: 10.3390/cells8030266.
Lamin A is involved in many cellular functions due to its ability to bind chromatin and transcription factors and affect their properties. Mutations of gene encoding lamin A affect the differentiation capacity of stem cells, but the mechanisms of this influence remain largely unclear. We and others have reported recently an interaction of lamin A with Notch pathway, which is among the main developmental regulators of cellular identity. The aim of this study was to explore the influence of mutations on the proosteogenic response of human cells of mesenchymal origin and to further explore the interaction of LMNA with Notch pathway. Mutations R527C and R471C in are associated with mandibuloacral dysplasia type A, a highly penetrant disease with a variety of abnormalities involving bone development. We used lentiviral constructs bearing mutations R527C and R471C and explored its influence on proosteogenic phenotype expression and Notch pathway activity in four types of human cells: umbilical vein endothelial cells (HUVEC), cardiac mesenchymal cells (HCMC), aortic smooth muscle cells (HASMC), and aortic valve interstitial cells (HAVIC). The proosteogenic response of the cells was induced by the addition of either LPS or specific effectors of osteogenic differentiation to the culture medium; phenotype was estimated by the expression of osteogenic markers by qPCR; activation of Notch was assessed by expression of Notch-related and Notch-responsive genes by qPCR and by activation of a luciferase CSL-reporter construct. Overall, we observed different reactivity of all four cell lineages to the stimulation with either LPS or osteogenic factors. R527C had a stronger influence on the proosteogenic phenotype. We observed the inhibiting action of R527C on osteogenic differentiation in HCMC in the presence of activated Notch signaling, while R527C caused the activation of osteogenic differentiation in HAVIC in the presence of activated Notch signaling. Our results suggest that the effect of a mutation is strongly dependent not only on a specific mutation itself, but also might be influenced by the intrinsic molecular context of a cell lineage.
核纤层蛋白 A 由于能够结合染色质和转录因子并影响其性质，因此参与许多细胞功能。编码核纤层蛋白 A 的基因突变会影响干细胞的分化能力，但这种影响的机制在很大程度上仍不清楚。我们和其他人最近报道了核纤层蛋白 A 与 Notch 通路的相互作用， Notch 通路是细胞身份的主要发育调节剂之一。本研究的目的是探讨突变对间充质来源的人类细胞成骨前体反应的影响，并进一步探讨 LMNA 与 Notch 通路的相互作用。突变 R527C 和 R471C 位于 基因中，与 A 型下颌面骨发育不良有关，这是一种高外显率疾病，涉及骨骼发育的多种异常。我们使用携带突变 R527C 和 R471C 的慢病毒构建体，并在四种类型的人类细胞中探索其对成骨前体表型表达和 Notch 通路活性的影响：脐静脉内皮细胞（HUVEC）、心脏间充质细胞（HCMC）、主动脉平滑肌细胞（HASMC）和主动脉瓣间质细胞（HAVIC）。通过向培养基中添加 LPS 或成骨分化的特定效应物来诱导细胞的成骨前体反应；通过 qPCR 检测成骨标志物的表达来估计表型；通过 qPCR 检测 Notch 相关和 Notch 反应基因的表达以及激活荧光素酶 CSL 报告基因构建体来评估 Notch 的激活。总体而言，我们观察到所有四种细胞系对 LPS 或成骨因子刺激的反应不同。R527C 对成骨前体表型的影响更强。我们观察到在激活的 Notch 信号存在下，R527C 对 HCMC 中成骨分化有抑制作用，而在激活的 Notch 信号存在下，R527C 导致 HAVIC 中成骨分化的激活。我们的结果表明，突变的影响不仅强烈依赖于特定的突变本身，还可能受到细胞谱系固有分子环境的影响。
