No abstract

One puzzling observation in patients affected with Hutchinson-Gilford progeria syndrome (HGPS), who overall exhibit systemic and dramatic premature aging, is the absence of any conspicuous cognitive impairment. Recent studies based on induced pluripotent stem cells derived from HGPS patient cells have revealed a lack of expression in neural derivatives of lamin A, a major isoform of LMNA that is initially produced as a precursor called prelamin A. In HGPS, defective maturation of a mutated prelamin A induces the accumulation of toxic progerin in patient cells. Here, we show that a microRNA, miR-9, negatively controls lamin A and progerin expression in neural cells. This may bear major functional correlates, as alleviation of nuclear blebbing is observed in nonneural cells after miR-9 overexpression. Our results support the hypothesis, recently proposed from analyses in mice, that protection of neural cells from progerin accumulation in HGPS is due to the physiologically restricted expression of miR-9 to that cell lineage.

SUMMARYThe role of microRNAs (miRNAs) as coordinators of stem cell fate has emerged over the last decade. We have used human embryonic stem cells to identify miRNAs involved in neural lineage commitment induced by the inhibition of TGF-like moleculemediated pathways. Among several candidate miRNAs expressed in the fetal brain, the two isoforms of miR-125 alone were detected in a time window compatible with a role in neural commitment in vitro. Functional analysis indicated that miR-125 isoforms were actively involved in the promotion of pluripotent cell conversion into SOX1-positive neural precursors. miR-125 promotes neural conversion by avoiding the persistence of non-differentiated stem cells and repressing alternative fate choices. This was associated with the regulation by miR-125 of SMAD4, a key regulator of pluripotent stem cell lineage commitment. Activation of miR-125 was directly responsive to the levels of TGF-like molecules, placing miR-125 at the core of mechanisms that lead to the irreversible neural lineage commitment of pluripotent stem cells in response to external stimuli.

Hutchinson–Gilford progeria syndrome (HGPS) is a lethal premature and accelerated aging disease caused by a de novo point mutation in LMNA encoding A‐type lamins. Progerin, a truncated and toxic prelamin A issued from aberrant splicing, accumulates in HGPS cells' nuclei and is a hallmark of the disease. Small amounts of progerin are also produced during normal aging. We show that progerin is sequestered into abnormally shaped promyelocytic nuclear bodies, identified as novel biomarkers in late passage HGPS cell lines. We found that the proteasome inhibitor MG132 induces progerin degradation through macroautophagy and strongly reduces progerin production through downregulation of SRSF‐1 and SRSF‐5 accumulation, controlling prelamin A mRNA aberrant splicing. MG132 treatment improves cellular HGPS phenotypes. MG132 injection in skeletal muscle of Lmna G609G/G609G mice locally reduces SRSF‐1 expression and progerin levels. Altogether, we demonstrate progerin reduction based on MG132 dual action and shed light on a promising class of molecules toward a potential therapy for children with HGPS.

Organs-on-a-chip platforms seek to recapitulate the complex microenvironment of human organs using miniaturized microfluidic devices. Besides modeling healthy organs, these devices have been used to model diseases, yielding new insights into pathophysiology. Hutchinson-Gilford progeria syndrome (HGPS) is a premature aging disease showing accelerated vascular aging, leading to the death of patients due to cardiovascular diseases. HGPS targets primarily vascular cells, which reside in mechanically active tissues. Here, we developed a progeria-on-a-chip model and examined the effects of biomechanical strain in the context of vascular aging and disease. Physiological strain induced a contractile phenotype in primary smooth muscle cells (SMCs), while a pathological strain induced a hypertensive phenotype similar to that of angiotensin II treatment. Interestingly, SMCs derived from human induced pluripotent stem cells of HGPS donors (HGPS iPS-SMCs), but not from healthy donors, showed an exacerbated inflammatory response to strain. In particular, we observed increased levels of inflammation markers as well as DNA damage. Pharmacological intervention reversed the strain-induced damage by shifting gene expression profile away from inflammation. The progeria-on-a-chip is a relevant platform to study biomechanics in vascular biology, particularly in the setting of vascular disease and aging, while simultaneously facilitating the discovery of new drugs and/or therapeutic targets.

Major physiological changes are governed by alternative splicing of RNA, and its misregulation may lead to specific diseases. With the use of a genome-wide approach, we show here that this splicing step can be modified by medication and demonstrate the effects of the biguanide metformin, on alternative splicing. The mechanism of action involves AMPK activation and downregulation of the RBM3 RNA-binding protein. The effects of metformin treatment were tested on myotonic dystrophy type I (DM1), a multisystemic disease considered to be a spliceopathy. We show that this drug promotes a corrective effect on several splicing defects associated with DM1 in derivatives of human embryonic stem cells carrying the causal mutation of DM1 as well as in primary myoblasts derived from patients. The biological effects of metformin were shown to be compatible with typical therapeutic dosages in a clinical investigation involving diabetic patients. The drug appears to act as a modifier of alternative splicing of a subset of genes and may therefore have novel therapeutic potential for many more diseases besides those directly linked to defective alternative splicing.

Melanocytes are essential for skin homeostasis and protection, and their defects in humans lead to a wide array of diseases that are potentially extremely severe. To date, the analysis of molecular mechanisms and the function of human melanocytes have been limited because of the difficulties in accessing large numbers of cells with the specific phenotypes. This issue can now be addressed via a differentiation protocol that allows melanocytes to be obtained from pluripotent stem cell lines, either induced or of embryonic origin, based on the use of moderate concentrations of a single cytokine, bone morphogenic protein 4. Human melanocytes derived from pluripotent stem cells exhibit all the characteristic features of their adult counterparts. This includes the enzymatic machinery required for the production and functional delivery of melanin to keratinocytes. Melanocytes also integrate appropriately into organotypic epidermis reconstructed in vitro. The availability of human cells committed to the melanocytic lineage in vitro will enable the investigation of those mechanisms that guide the developmental processes and will facilitate analysis of the molecular mechanisms responsible for genetic diseases. Access to an unlimited resource may also prove a vital tool for the treatment of hypopigmentation disorders when donors with matching haplotypes become available in clinically relevant banks of pluripotent stem cell lines.

Hutchinson‐Gilford progeria syndrome is a rare congenital disease characterized by premature aging in children. The results of this systematic comparative study of the three main treatments currently administered or proposed to progeria‐affected children reveal the complexity of the modes of action of different drugs and underscore the use of induced pluripotent stem cell derivatives as a critical and powerful tool for standardized, comparative pharmacological studies.