Rejuvenating the aged cell through epigenetic modulation: defining culture conditions to reverse mesenchymal stromal cell (MSC) ageing

项目原理和重要性：我们最近发现了可能参与MSC衰老的表观遗传调控因子。我们使用短发夹RNA （shRNA）文库对398种表观遗传酶进行了未发表的研究，结果表明，在衰老/非增殖的人间充质干细胞（hMSCs）中，沉默特定的表观遗传酶具有逆转生长停滞和重建细胞增殖的能力。有趣的是，大部分已确定的表观遗传酶具有相关的小分子（化学/化合物）调节剂，这为开发具有逆转hMSC衰老能力的体外培养条件提供了独特的机会。事实上，体细胞重编程为多能性已经告诉我们，在细胞水平上，将年龄重置为“基态”是可能的。仔细操纵表观遗传状态也可能使细胞在不消除其特性的情况下“再次年轻”(4)。这将最终导致如何逆转细胞衰老和/或保持细胞处于年轻状态的新见解，这将对再生医学产生重大影响和应用。目的和目标：本研究将明确表观遗传学在骨髓间充质干细胞老化过程中的作用，并为建立体外逆转细胞老化的培养条件创造新的技术进步。目的1：建立基于表观遗传酶活性调控的hMSC衰老调控条件。目的2：了解hMSC衰老的表观遗传控制机制，从而确定骨骼抗衰老靶点。目的3通过单细胞RNA测序（scRNASeq）了解表观遗传调控对年龄逆转反应的异质性。

Project rationale and importance: We have recently discovered epigenetic regulators that may be involved in MSC ageing. Our unpublished data using a short hairpin RNA (shRNA) library targeting 398 epigenetic enzymes indicates that silencing of specific epigenetic enzymes in aged/non-proliferating human MSCs (hMSCs) has the capacity to reverse growth arrest and re-establish cell proliferation. Interestingly, a large proportion of the identified epigenetic enzymes have associated small molecule (chemical/compound) modulators, which presents a unique opportunity to develop in vitro culture conditions that have the capacity to reverse hMSC ageing. Indeed, the reprogramming of somatic cells to pluripotency has taught us that at the cellular level it is possible to reset age to a 'ground state'. Careful manipulation of epigenetic state is also likely to make cells 'young again' without erasing their identity(4). This will ultimately lead to novel insights in how to reverse cell ageing and/or maintain cells in a youthful state, which will have significant impact and applications in regenerative medicine.Aim and objectives: This studentship will define the role of epigenetics in the ageing process of hMSCs, and create new technological advances towards the establishment of culture conditions to reverse cell ageing in vitro.Objective 1 To develop in vitro conditions with the capacity to modify hMSC ageing based on the control of epigenetic enzyme activity. Objective 2 To deliver a mechanistic understanding of the epigenetic control of hMSC ageing, hence identifying skeletal anti-ageing targets. Objective 3 To understand the heterogeneity of response to age reversal through epigenetic modulation by single cell RNA sequencing (scRNASeq).