Kirti Bhatt

Skeletal muscle gene expression profiles in 20-29 year old and 65-71 year old women.

Gene expression profiling may provide leads for investigations of the molecular basis of functional declines associated with aging. In this study, high-density oligonucleotide arrays were used to probe the patterns of gene expression in skeletal muscle of seven young women (20-29 years old) and eight healthy older women (65-71 years old). The older subjects had reduced muscle mass, strength, and peak oxygen consumption relative to young women. There were approximately 1000 probe sets that suggested differential gene expression in younger and older muscle according to statistical criteria. The most highly overexpressed genes (>3-fold) in older muscle were p21 (cyclin-dependent kinase inhibitor 1A), which might reflect increased DNA damage, perinatal myosin heavy chain, which might reflect increased muscle fiber regeneration, and tomoregulin, which does not have a defined function in muscle. More than 40 genes encoding proteins that bind to pre-mRNAs or mRNAs were expressed at higher levels in older muscle. More than 100 genes involved in energy metabolism were expressed at lower levels in older muscle. In general, these results support previous observations on the differences in gene expression profiles between younger and older men.

Insulin-like growth factor-1 and myostatin mRNA expression in muscle: comparison between 62-77 and 21-31 yr old men.

The present study was done to determine the effect of age on muscle concentrations of mRNAs encoding two growth factors that are thought to be important regulators of muscle mass: insulin-like growth factor-1 (IGF-1) and myostatin. Quantitative RT-PCR assays indicated that the mean IGF-1 mRNA concentration in older muscle (62-77 yr, n=15 men) was approximately 25% less, per ng total RNA (P<0.005), than in young adult muscle (21-31 yr, n=12 men). One third of the older men had IGF-1 mRNA levels below the lowest concentration observed in young muscle. Myostatin mRNA concentrations were similar in young and old muscle. Muscle mass and myofibrillar protein synthesis rates among eight older men did not correlate with either IGF-1 or myostatin mRNA levels. We conclude that IGF-1 gene expression in muscle tends to decline with normal aging. The functional significance is uncertain.

TWEAK/Fn14, a pathway and novel therapeutic target in myotonic dystrophy

Myotonic dystrophy type 1 (DM1), the most prevalent muscular dystrophy in adults, is characterized by progressive muscle wasting and multi-systemic complications. DM1 is the prototype for disorders caused by RNA toxicity. Currently, no therapies exist. Here, we identify that fibroblast growth factor-inducible 14 (Fn14), a member of the tumor necrosis factor receptor super-family, is induced in skeletal muscles and hearts of mouse models of RNA toxicity and in tissues from DM1 patients, and that its expression correlates with severity of muscle pathology. This is associated with downstream signaling through the NF-κB pathways. In mice with RNA toxicity, genetic deletion of Fn14 results in reduced muscle pathology and better function. Importantly, blocking TWEAK/Fn14 signaling with an anti-TWEAK antibody likewise improves muscle histopathology and functional outcomes in affected mice. These results reveal new avenues for therapeutic development and provide proof of concept for a novel therapeutic target for which clinically available therapy exists to potentially treat muscular dystrophy in DM1.

Intron retention induced by microsatellite expansions as a disease biomarker

Significance A number of hereditary neurological and neuromuscular diseases are caused by the abnormal expansion of short tandem repeats, or microsatellites, resulting in the expression of repeat expansion RNAs and proteins with pathological properties. Although these microsatellite expansions may occur in either the coding or noncoding regions of the genome, trinucleotide CNG repeats predominate in exonic coding and untranslated regions while intron mutations vary from trinucleotide to hexanucleotide GC-rich, and A/AT-rich, repeats. Here, we use transcriptome analysis combined with complementary experimental approaches to demonstrate that GC-rich intronic expansions are selectively associated with host intron retention. Since these intron retention events are detectable in both affected tissues and peripheral blood, they provide a sensitive and disease-specific diagnostic biomarker. Expansions of simple sequence repeats, or microsatellites, have been linked to ∼30 neurological–neuromuscular diseases. While these expansions occur in coding and noncoding regions, microsatellite sequence and repeat length diversity is more prominent in introns with eight different trinucleotide to hexanucleotide repeats, causing hereditary diseases such as myotonic dystrophy type 2 (DM2), Fuchs endothelial corneal dystrophy (FECD), and C9orf72 amyotrophic lateral sclerosis and frontotemporal dementia (C9-ALS/FTD). Here, we test the hypothesis that these GC-rich intronic microsatellite expansions selectively trigger host intron retention (IR). Using DM2, FECD, and C9-ALS/FTD as examples, we demonstrate that retention is readily detectable in affected tissues and peripheral blood lymphocytes and conclude that IR screening constitutes a rapid and inexpensive biomarker for intronic repeat expansion disease.

Design of Bivalent Nucleic Acid Ligands for Recognition of RNA-Repeated Expansion Associated with Huntington’s Disease

We report the development of a new class of nucleic acid ligands that is comprised of Janus bases and the MPγPNA backbone and is capable of binding rCAG repeats in a sequence-specific and selective manner via, inference, bivalent H-bonding interactions. Individually, the interactions between ligands and RNA are weak and transient. However, upon the installation of a C-terminal thioester and an N-terminal cystine and the reduction of disulfide bond, they undergo template-directed native chemical ligation to form concatenated oligomeric products that bind tightly to the RNA template. In the absence of an RNA target, they self-deactivate by undergoing an intramolecular reaction to form cyclic products, rendering them inactive for further binding. The work has implications for the design of ultrashort nucleic acid ligands for targeting rCAG-repeat expansion associated with Huntingtons disease and a number of other related neuromuscular and neurodegenerative disorders.