Biology subjects

Machiraju, P.

Publications and source records attributed to Machiraju, P..

4 recordsLinked to original sources

bioRxiv2020

A novel synthetic peptide provides neuroprotection and rescues rat cortical neurons from anesthetic-induced cell death and modulation of growth and synaptic assembly

Anesthetics, although necessary for many modern procedures, exert neurotoxic effects in various experimental models; the underlying mechanisms, however, remain unknown. In the absence of this information, any in vivo mitigation strategy would be challenging. To help address these gaps, we sought to determine whether preserving mitochondrial network integrity with a non-toxic, short-life synthetic peptide, P110, could protect cortical neurons against both inhalational and intravenous anesthetic-induced neurotoxicity. This study provides the first direct comparative account of three key general anesthetics (desflurane, propofol, and ketamine) under identical conditions and demonstrates their impact on cellular viability, neurite outgrowth, and synaptic network assembly on neonatal rat cortical neurons. Further, we discovered that the inhibition of Fis1 receptor-mediated fission reverses anesthetic-induced aberrations in an agent-specific manner. Our data thus underscore the importance of a reductionist approach, offering unique opportunities to explore non-toxic, mitochondria-based therapeutic tools for mitigating anesthetic-induced harm to the developing brain.

neuroscience↗

bioRxiv2019

Early Inhibition of Retinoic Acid Signaling Rapidly Generates Cardiomyocytes Expressing Ventricular Markers from Human Induced Pluripotent Stem Cells

SUMMARYCurrent protocols for the differentiation of cardiomyocytes from human induced pluripotent stem cells (iPSCs) generally require prolonged time in culture and result in heterogeneous cellular populations. We present a method for the generation of beating cardiomyocytes expressing specific ventricular markers after just 14 days. Addition of the pan-retinoic acid receptor inverse agonist BMS 493 to human iPSCs for the first 8 days of differentiation resulted in increased protein expression of the ventricular isoform of myosin regulatory light chain (MLC2V) from 18.7% {+/-} 1.72% to 55.8% {+/-} 11.4% (p <0.0001) in cells co-expressing the cardiac muscle protein troponin T (TNNT2). Increased MLC2V expression was also accompanied by a slower beating rate (49.4 {+/-} 1.53 vs. 93.0 {+/-} 2.81 beats per minute, p <0.0001) and increased contraction amplitude (201% {+/-} 8.33% vs. 100% {+/-} 10.85%, p <0.0001) compared to untreated cells. Improved directed differentiation will improve in vitro cardiac modeling.

cell biology↗

bioRxiv2019

Reversible Mitochondrial Fragmentation in iPSC-Derived Cardiomyocytes from Children with DCMA, a Mitochondrial Cardiomyopathy

BackgroundThe dilated cardiomyopathy with ataxia syndrome (DCMA) is an understudied autosomal recessive disease caused by loss-of-function mutations in the poorly characterized gene DNAJC19. Clinically, DCMA is commonly associated with heart failure and early death in affected children through an unknown mechanism. DCMA has been linked to Barth syndrome, a rare but well-studied disorder caused by deficient maturation of cardiolipin (CL), a key mitochondrial membrane phospholipid.\n\nMethodsPeripheral blood mononuclear cells from two children with DCMA and severe cardiac dysfunction were reprogrammed into induced pluripotent stem cells (iPSCs). Patient and control iPSCs were differentiated into beating cardiomyocytes (iPSC-CMs) using a metabolic selection strategy and mitochondrial structure and CL content before and after incubation with the mitochondrially-targeted peptide SS-31 were quantified.\n\nResultsPatient iPSCs carry the causative DNAJC19 mutation (rs137854888) found in the Hutterite population and the iPSC-CMs demonstrated highly fragmented and abnormally-shaped mitochondria associated with an imbalanced isoform ratio of the mitochondrial protein OPA1, an important regulator of mitochondrial fusion. These abnormalities were reversible by incubation with SS-31 for 24 hours. Differentiation of iPSCs into iPSC-CMs increased the number of CL species observed but consistent, significant differences in CL content were not seen between patients and control.\n\nConclusionsWe describe a unique and novel cellular model that provides insight into the mitochondrial abnormalities present in DCMA and identifies SS-31 as a potential therapeutic for this devastating disease.

cell biology↗

bioRxiv2019

SS-31 Reverses Mitochondrial Fragmentation in Fibroblasts from Patients with DCMA, a Mitochondrial Cardiomyopathy

ObjectivesWe used patient dermal fibroblasts to characterize the mitochondrial abnormalities associated with the dilated cardiomyopathy with ataxia syndrome (DCMA) and to study the effect of the mitochondrially-targeted peptide SS-31 as a potential novel therapeutic.\n\nBackgroundDCMA is an understudied autosomal recessive disorder thought to be related to Barth syndrome but caused by mutations in DNAJC19, a protein of unknown function localized to the mitochondria. The clinical disease is characterized by 3-methylglutaconic aciduria, dilated cardiomyopathy, abnormal neurological development and other heterogeneous features. Until recently no effective therapies had been identified and affected patients frequently died in early childhood from intractable heart failure.\n\nMethodsDermal fibroblasts from four pediatric patients with DCMA were used to establish parameters of mitochondrial dysfunction. Mitochondrial structure, reactive oxygen species (ROS) production, cardiolipin composition and gene expression were evaluated.\n\nResultsImmunocytochemistry with semi-automated quantification of mitochondrial structural metrics and transmission electron microscopy demonstrated mitochondria to be highly fragmented in DCMA fibroblasts compared to healthy control cells. Live-cell imaging demonstrated significantly increased ROS production in patient cells. These structural and functional abnormalities were reversed by treating DCMA fibroblasts with SS-31, a synthetic peptide that localizes to the inner mitochondrial membrane. Levels of cardiolipin were not significantly different between control and DCMA cells and were unaffected by SS-31 treatment.\n\nConclusionsOur results demonstrate the abnormal mitochondrial structure and function in fibroblasts from patients with DCMA and suggest that SS-31 may represent a potential therapy for this devastating disease.

cell biology↗