Myosin Heavy Chain Converter Domain Mutations Drive Early-Stage Changes in Extracellular Matrix Dynamics in Hypertrophic Cardiomyopathy.

More than 60% of hypertrophic cardiomyopathy (HCM)-causing mutations are found in the gene loci encoding cardiac myosin-associated proteins including myosin heavy chain (MHC) and myosin binding protein C (MyBP-C). Moreover, patients with more than one independent HCM mutation may be at increased risk for more severe disease expression and adverse outcomes. However detailed mechanistic understanding, especially at early stages of disease progression, is limited. To identify early-stage HCM triggers, we generated single (MYH7 c.2167C > T [R723C] with a known pathogenic significance in the MHC converter domain) and double (MYH7 c.2167C > T [R723C]; MYH6 c.2173C > T [R725C] with unknown significance) myosin gene mutations in human induced pluripotent stem cells (hiPSCs) using a base-editing strategy. Cardiomyocytes (CMs) derived from hiPSCs with either single or double mutation exhibited phenotypic characteristics consistent with later-stage HCM including hypertrophy, multinucleation, altered calcium handling, metabolism, and arrhythmia. We then probed mutant CMs at time points prior to the detection of known HCM characteristics. We found MYH7/MYH6 dual mutation dysregulated extracellular matrix (ECM) remodeling, altered integrin expression, and interrupted cell-ECM adhesion by limiting the formation of focal adhesions. These results point to a new phenotypic feature of early-stage HCM and reveal novel therapeutic avenues aimed to delay or prohibit disease onset.

Anti-Excitotoxic Effects of N-Butylidenephthalide Revealed by Chemically Insulted Purkinje Progenitor Cells Derived from SCA3 iPSCs.

Spinocerebellar ataxia type 3 (SCA3) is characterized by the over-repetitive CAG codon in the ataxin-3 gene (ATXN3), which encodes the mutant ATXN3 protein. The pathological defects of SCA3 such as the impaired aggresomes, autophagy, and the proteasome have been reported previously. To date, no effective treatment is available for SCA3 disease. This study aimed to study anti-excitotoxic effects of n-butylidenephthalide by chemically insulted Purkinje progenitor cells derived from SCA3 iPSCs. We successfully generated Purkinje progenitor cells (PPs) from SCA3 patient-derived iPSCs. The PPs, expressing both neural and Purkinje progenitor's markers, were acquired after 35 days of differentiation. In comparison with the PPs derived from control iPSCs, SCA3 iPSCs-derived PPs were more sensitive to the excitotoxicity induced by quinolinic acid (QA). The observations of QA-treated SCA3 PPs showing neural degeneration including neurite shrinkage and cell number decrease could be used to quickly and efficiently identify drug candidates. Given that the QA-induced neural cell death of SCA3 PPs was established, the activity of calpain in SCA3 PPs was revealed. Furthermore, the expression of cleaved poly (ADP-ribose) polymerase 1 (PARP1), a marker of apoptotic pathway, and the accumulation of ATXN3 proteolytic fragments were observed. When SCA3 PPs were treated with n-butylidenephthalide (n-BP), upregulated expression of calpain 2 and concurrent decreased level of calpastatin could be reversed, and the overall calpain activity was accordingly suppressed. Such findings reveal that n-BP could not only inhibit the cleavage of ATXN3 but also protect the QA-induced excitotoxicity from the Purkinje progenitor loss.

Human Olfactory Ensheathing Cell Transplantation Improves Motor Function in a Mouse Model of Type 3 Spinocerebellar Ataxia.

Spinocerebellar ataxia (SCA) is a progressive neurodegenerative disease that affects the cerebellum and spinal cord. Among the 40 types of SCA, SCA type 3 (SCA3), also referred to as Machado-Joseph disease, is the most common. In the present study, we investigated the therapeutic effects of intracranial transplantation of human olfactory ensheathing cells (hOECs) in the ATXN3-84Q mouse model of SCA3. Motor function begins to decline in ATXN3-84Q transgenic mice at approximately 13 weeks of age. ATXN3-84Q mice that received intracranial hOEC transplantation into the dorsal raphe nucleus of the brain exhibited significant improvements in motor function, as measured by the rotarod performance test and footprint pattern analysis. In addition, intracranial hOEC transplantation alleviated cerebellar inflammation, prohibited Purkinje cells from dying, and enhanced the neuroplasticity of cerebellar Purkinje cells. The protein levels of tryptophan hydroxylase 2, the rate-limiting enzyme for serotonin synthesis in the cerebellum, and ryanodine receptor (RYR) increased in mice that received intracranial hOEC transplantation. Because both serotonin and RYR can enhance Purkinje cell maturation, these effects may account for the therapeutic benefits mediated by intracranial hOEC transplantation in SCA3 mice. These results indicate that intracranial hOEC transplantation has potential value as a novel strategy for treating SCA3.

The antisenescence effect of trans-cinnamaldehyde on adipose-derived stem cells.

As assuring cell quality is an essential parameter for the success of stem cell therapy, the impact of various senescence-inducing stress signals, and strategies to circumvent them, has been an important area of focus in stem cell research. The aim of this study was to demonstrate the capacity of Trans-cinnamaldehyde (TC) in reversing stress-induced senescence and maintaining the quality of stem cells in a chemically (H2O2)-induced cell senescence model. Because of the availability and the promising application potential in regenerative medicine, adipose-derived stem cells (ADSCs) were chosen for the study. We found that H2O2 treatment resulted in the expression of senescence characteristics in the ADSCs, including decreased proliferation rate, increased senescence-associated ß-galactosidase (SA-ß-gal) activity, decreased silent mating type information regulation 2 homolog (SIRT1) expression, and decreased telomerase activity. However, TC treatment was sufficient to rescue or reduce the effects of H2O2 induction, ultimately leading to an increased proliferation rate, a decrease in the percentage of SA-ß-gal-positive cells, upregulation of SIRT1 expression, and increased telomerase activity of the senescent ADSCs at the cellular level. Moreover, a chemically induced liver fibrosis animal model was used to evaluate the functionality of these rescued cells in vivo. Liver dysfunction was established by injecting 200 mg/kg thioacetamide (TAA) intraperitoneally into Wistar rats every third day for 60 days. The experimental rats were separated into groups: normal group (rats without TAA induction), sham group (without ADSC transplantation), positive control group (transplanted with normal ADSCs), H2O2 group (transplanted with H2O2-induced senescent ADSCs), and H2O2 + TC group (transplanted with ADSCs pretreated with H2O2 and then further treated with TC). In the transplantation group, 1 × 10(6) human ADSCs were introduced into each rat via direct liver injection. Based on the biochemical analysis and immunohistochemical staining results, it was determined that the therapeutic effects on liver fibrosis by the induced senescent ADSCs (H2O2 group) were not as significant as those exerted by the normal ADSCs (the positive control group). However, the H2O2 + TC group showed significant reversal of liver damage when compared to the H2O2 group 1 week posttransplantation. These data confirmed that the TC treatment had the potential to reduce the effects of H2O2-induced senescence and to restore in vivo functionality of the induced senescent ADSCs. It is therefore suggested that TC has potential applications in maintaining the quality of stem cells and could aid in treating senescence-related disorders.