calpain and Amyotrophic-Lateral-Sclerosis

TAR DNA-binding protein (TDP-43) pathology and reduced expression of adenosine deaminase acting on RNA 2 (ADAR2), which is the RNA editing enzyme responsible for adenosine-to-inosine conversion at the GluA2 glutamine/arginine (Q/R) site, concomitantly occur in the same motor neurons of amyotrophic lateral sclerosis (ALS) patients; this finding suggests a link between these two ALS-specific molecular abnormalities. AMPA receptors containing Q/R site-unedited GluA2 in their subunit assembly are Ca(2+)-permeable, and motor neurons lacking ADAR2 undergo slow death in conditional ADAR2 knockout (AR2) mice, which is a mechanistic ALS model in which the ADAR2 gene is targeted in cholinergic neurons. Moreover, deficient ADAR2 induced mislocalization of TDP-43 similar to TDP-43 pathology seen in the sporadic ALS patients in the motor neurons of AR2 mice. The abnormal mislocalization of TDP-43 specifically resulted from activation of the Ca(2+)-dependent serine protease calpain that specifically cleaved TDP-43 at the C-terminal region, and generated aggregation-prone N-terminal fragments. Notably, the N-terminal fragments of TDP-43 lacking the C-terminus were demonstrated in the brains and spinal cords of ALS patients. Because normalization of either the Ca(2+)-permeability of AMPA receptors or the calpain activity in the motor neurons normalized the subcellular localization of TDP-43 in AR2 mice, it is likely that exaggerated calpain-dependent TDP-43 fragments played a role at least in the initiation of TDP-43 pathology. Elucidation of the molecular cascade of neuronal death induced by ADAR2 downregulation could provide a new specific therapy for sporadic ALS. In this review, we summarized the work from our group on the role of inefficient GluA2 Q/R site-RNA editing and TDP-43 pathology in sporadic ALS, and discussed possible effects of inefficient ADAR2-mediated RNA editing in general.

Topics: Adenosine Deaminase; Amyotrophic Lateral Sclerosis; Animals; Calpain; Cell Death; DNA-Binding Proteins; Humans; Mice; Motor Neurons; Receptors, AMPA; RNA Editing; RNA-Binding Proteins; Spinal Cord

Amyotrophic lateral sclerosis (ALS) is the most common adult-onset motor neuron disease affecting healthy middle-aged individuals. Mislocalization of TAR DNA binding protein of 43 kDa (TDP-43) or TDP-43 pathology observed in the spinal motor neurons is the pathological hallmark of ALS. The mechanism generating TDP-43 pathology remained uncertain. Several reports suggested that cleavage of TDP-43 into aggregation-prone fragments might be the earliest event. Therefore, elucidation of the protease(s) that is responsible for TDP-43 cleavage in the motor neurons is awaited. ALS-specific molecular abnormalities other than TDP-43 pathology in the motor neurons of sporadic ALS patients include inefficient RNA editing at the GluA2 glutamine/arginine (Q/R) site, which is specifically catalyzed by adenosine deaminase acting on RNA 2 (ADAR2). We have developed the conditional ADAR2 knockout (AR2) mice, in which the ADAR2 gene is targeted in motor neurons. We found that Ca(2+)-dependent cysteine protease calpain cleaved TDP-43 into aggregation-prone fragments, which initiated TDP-43 mislocalization in the motor neurons expressing abnormally abundant Ca(2+)-permeable AMPA receptors. Here we summarized the molecular cascade leading to TDP-43 pathology observed in the motor neurons of AR2 mice and discussed possible roles of dysregulation of calpain-dependent cleavage of TDP-43 in TDP-43 pathology observed in neurological diseases in general.

Topics: Amyotrophic Lateral Sclerosis; Animals; Calcium Signaling; Calpain; Disease Models, Animal; DNA-Binding Proteins; Humans; Mice; Mice, Knockout; Motor Neurons; Receptors, AMPA; RNA Editing; Spinal Cord

Calpain is a ubiquitous calcium-sensitive protease that is essential for normal physiologic neuronal function. However, alterations in calcium homeostasis lead to persistent, pathologic activation of calpain in a number of neurodegenerative diseases. Pathologic activation of calpain results in the cleavage of a number of neuronal substrates that negatively affect neuronal structure and function, leading to inhibition of essential neuronal survival mechanisms. In this review, we examine the mechanistic underpinnings of calcium dysregulation resulting in calpain activation in the acute neurodegenerative diseases such as cerebral ischemia and in the chronic neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, Huntington's disease, multiple sclerosis, prion-related encephalopathy, and amylotrophic lateral sclerosis. The premise of this paper is that analysis of the signaling and transcriptional consequences of calpain-mediated cleavage of its various substrates for any neurodegenerative disease can be extrapolated to all of the neurodegenerative diseases vulnerable to calcium dysregulation.

Topics: Alzheimer Disease; Amyotrophic Lateral Sclerosis; Animals; Brain Ischemia; Calpain; Humans; Huntington Disease; Multiple Sclerosis; Neurodegenerative Diseases; Parkinson Disease; Prion Diseases; Signal Transduction; Trauma, Nervous System

Repeated concussion represents a serious health problem as it can result in various brain pathologies, ranging from minor focal tissue injury to severe chronic traumatic encephalopathy. The calcium-dependent protease, calpain, participates in the development of neurodegeneration following concussion, but there is no information regarding the relative contribution of calpain-1 and calpain-2, the major calpain isoforms in the brain. We used a mouse model of repeated concussions, which reproduces most of the behavioral and neuropathological features of the human condition, to address this issue. Deletion of calpain-2 or treatment with a selective calpain-2 inhibitor for 2 weeks prevented most of these neuropathological features. Changes in TAR DNA binding protein 43 (TDP-43) subcellular localization similar to those found in human amyotrophic lateral sclerosis and frontotemporal dementia were also prevented by deletion of calpain-2 or treatment with calpain-2 inhibitor. Our results indicate that a selective calpain-2 inhibitor represents a therapeutic approach for concussion.

Topics: Amyotrophic Lateral Sclerosis; Animals; Brain; Brain Concussion; Calpain; Frontotemporal Dementia; Mice

Cytoplasmic accumulation of TDP-43 in motor neurons is the most prominent pathological feature in amyotrophic lateral sclerosis (ALS). A feedback cycle between nucleocytoplasmic transport (NCT) defect and TDP-43 aggregation was shown to contribute to accumulation of TDP-43 in the cytoplasm. However, little is known about cellular factors that can control the activity of NCT, thereby affecting TDP-43 accumulation in the cytoplasm. Here, we identified via FRAP and optogenetics cytosolic calcium as a key cellular factor controlling NCT of TDP-43. Dynamic and reversible changes in TDP-43 localization were observed in

Topics: Active Transport, Cell Nucleus; alpha Karyopherins; Amyotrophic Lateral Sclerosis; Animals; Calcium; Calpain; Cytoplasm; DNA-Binding Proteins; Drosophila melanogaster; Drosophila Proteins; Neurons

Nuclear dysfunction in motor neurons has been hypothesized to be a principal cause of amyotrophic lateral sclerosis (ALS) pathogenesis. Here, we investigated the mechanism by which the nuclear pore complex (NPC) is disrupted in dying motor neurons in a mechanistic ALS mouse model (adenosine deaminase acting on RNA 2 (ADAR2) conditional knockout (AR2) mice) and in ALS patients. We showed that nucleoporins (Nups) that constituted the NPC were cleaved by activated calpain via a Ca

Topics: Active Transport, Cell Nucleus; Adenosine Deaminase; alpha Karyopherins; Amyotrophic Lateral Sclerosis; Animals; Calcium; Calpain; Disease Models, Animal; DNA-Binding Proteins; Humans; Mice; Mice, Inbred C57BL; Mice, Knockout; Motor Neurons; Nuclear Pore; Nuclear Pore Complex Proteins; Phosphorylation; Receptors, AMPA; RNA Polymerase II; RNA-Binding Proteins; Spinal Cord

TAR DNA-binding protein-43 (TDP-43) pathology, which includes the presence of abnormal TDP-43-containing inclusions with a loss of nuclear TDP-43 in affected neurons, is a pathological hallmark of amyotrophic lateral sclerosis (ALS) and/or frontotemporal lobar degeneration (FTLD). TDP-43 in the pathological brains and spinal cords of ALS/FTLD patients is abnormally fragmented and phosphorylated. It is believed that the generation of aggregation-prone TDP-43 fragments initiates TDP-43 pathology, and we previously reported that calpain has an important role in the generation of such aggregation-prone TDP-43 fragments. However, the role of phosphorylation in TDP-43 pathology has not been largely elucidated, despite previous observations that several kinases and their kinases are involved in TDP-43 phosphorylation. Here, we investigated the role of TDP-43 phosphorylation in the calpain-dependent cleavage of TDP-43 and found that phosphorylated, full-length TDP-43 and calpain-dependent TDP-43 fragments were more resistant to cleavage by calpain than endogenous full-length TDP-43 was. These results suggest that both phosphorylated and calpain-cleaved TDP-43 fragments persist intracellularly for a length of time that is sufficient for self-aggregation, thereby serving as seeds for inclusions.

Topics: Amyotrophic Lateral Sclerosis; Calpain; DNA-Binding Proteins; Frontotemporal Lobar Degeneration; HeLa Cells; Humans; Phosphorylation; Protein Aggregates

Amyotrophic lateral sclerosis (ALS) is a progressive motor neuron disease with a poorly understood cause and no effective treatment. Given that calpains mediate neurodegeneration in other pathological states and are abnormally activated in ALS, we investigated the possible ameliorative effects of inhibiting calpain over-activation in hSOD1(G93A) transgenic (Tg) mice in vivo by neuron-specific over-expression of calpastatin (CAST), the highly selective endogenous inhibitor of calpains. Our data indicate that over-expression of CAST in hSOD1(G93A) mice, which lowered calpain activation to levels comparable to wild-type mice, inhibited the abnormal breakdown of cytoskeletal proteins (spectrin, MAP2 and neurofilaments), and ameliorated motor axon loss. Disease onset in hSOD1(G93A) /CAST mice compared to littermate hSOD1(G93A) mice is delayed, which accounts for their longer time of survival. We also find that neuronal over-expression of CAST in hSOD1(G93A) transgenic mice inhibited production of putative neurotoxic caspase-cleaved tau and activation of Cdk5, which have been implicated in neurodegeneration in ALS models, and also reduced the formation of SOD1 oligomers. Our data indicate that inhibition of calpain with CAST is neuroprotective in an ALS mouse model. CAST (encoding calpastatin) inhibits hyperactivated calpain to prevent motor neuron disease operating through a cascade of events as indicated in the schematic, with relevance to amyotrophic lateral sclerosis (ALS). We propose that over-expression of CAST in motor neurons of hSOD1(G93A) mice inhibits activation of CDK5, breakdown of cytoskeletal proteins (NFs, MAP2 and Tau) and regulatory molecules (Cam Kinase IV, Calcineurin A), and disease-causing proteins (TDP-43, α-Synuclein and Huntingtin) to prevent neuronal loss and delay neurological deficits. In our experiments, CAST could also inhibit cleavage of Bid, Bax, AIF to prevent mitochondrial, ER and lysosome-mediated cell death mechanisms. Similarly, CAST over-expression in neurons attenuated pathological effects of TDP-43, α-synuclein and Huntingtin. These results suggest a potential value of specific small molecule inhibitors of calpains in delaying the development of ALS. Read the Editorial Highlight for this article on page 140.

Topics: Age Factors; Amyotrophic Lateral Sclerosis; Animals; Axons; Calcium-Binding Proteins; Calpain; Cell Death; Cyclin-Dependent Kinase 5; Cysteine Proteinase Inhibitors; Cytoskeletal Proteins; Disease Models, Animal; Disease Progression; Gene Expression Regulation; Humans; Mice; Mice, Transgenic; Motor Activity; Motor Neurons; Nerve Degeneration; Spinal Cord; Superoxide Dismutase

Amyotrophic Lateral Sclerosis (ALS), a severe neurodegenerative disease, affects the upper and lower motor neurons in the brain and spinal cord. In some studies, ALS disease progression has been associated with an increase in calcium-dependent degeneration processes. Motoneurons are specifically vulnerable to sustained membrane depolarization and excessive elevation of intracellular calcium concentration. The present study analyzed intracellular events in embryonic motoneurons and adult spinal cords of the hSOD1G93A ALS mouse model. We observed activation of calpain, a calcium-dependent cysteine protease that degrades a variety of substrates, and a reduction in calcium-calmodulin dependent protein kinase type IV (CaMKIV) levels in protein extracts from spinal cords obtained at several time-points of hSOD1G93A mice disease progression. However, in cultured embryonic motoneurons these differences between controls and hSOD1G93A mutants are not evident. Our results support the hypothesis that age-dependent changes in calcium homeostasis and resulting events, e.g., calpain activation and CaMKIV processing, are involved in ALS pathogenesis.

Topics: Age Factors; Amyotrophic Lateral Sclerosis; Analysis of Variance; Animals; Calcium-Calmodulin-Dependent Protein Kinase Type 4; Calpain; Cells, Cultured; Disease Models, Animal; Embryo, Mammalian; Gene Expression Regulation; Humans; Intercellular Signaling Peptides and Proteins; Mice; Mice, Inbred C57BL; Mice, Transgenic; Motor Neurons; Nerve Tissue Proteins; Potassium; Spinal Cord; Superoxide Dismutase

Elevation in [Ca(2+)]i and activation of calpain-1 occur in central nervous system of SOD1(G93A) transgenic mice model of amyotrophic lateral sclerosis (ALS), but few data are available about the early stage of ALS. We here investigated the level of activation of the Ca(2+)-dependent protease calpain-1 in spinal cord of SOD1(G93A) mice to ascertain a possible role of the protease in the aetiology of ALS. Comparing the events occurring in the 120 day old mice, we found that [Ca(2+)]i and activation of calpain-1 were also increased in the spinal cord of 30 day old mice, as indicated by the digestion of some substrates of the protease such as nNOS, αII-spectrin, and the NR2B subunit of NMDA-R. However, the digestion pattern of these proteins suggests that calpain-1 may play different roles depending on the phase of ALS. In fact, in spinal cord of 30 day old mice, activation of calpain-1 produces high amounts of nNOS active species, while in 120 day old mice enhanced-prolonged activation of calpain-1 inactivates nNOS and down-regulates NR2B. Our data reveal a critical role of calpain-1 in the early phase and during progression of ALS, suggesting new therapeutic approaches to counteract its onset and fatal course.

Topics: Amyotrophic Lateral Sclerosis; Animals; Calcium; Calpain; Disease Models, Animal; Disease Progression; Humans; Mice; Mice, Transgenic; Motor Neurons; Nitric Oxide Synthase Type I; Proteolysis; Receptors, N-Methyl-D-Aspartate; Spinal Cord; Superoxide Dismutase; Superoxide Dismutase-1

We use 1,2-diacetylbenzene (1,2-DAB) to probe molecular mechanisms of proximal giant neurofilamentous axonopathy (PGNA), a pathological hallmark of amyotrophic lateral sclerosis. The spinal cord proteome of rodents displaying 1,2-DAB PGNA suggests a reduction in the abundance of α-II spectrin (Spna2), a key protein in the maintenance of axonal integrity. Protein immunoblotting indicates that this reduction is due to Spna2 degradation. We investigated the importance of such degradation in 1,2-DAB PGNA. Spna2 mutant mice lacking a calpain- and/or caspase-sensitive domain (CSD), thus hypothetically resistant to 1,2-DAB, and wild-type littermates, were treated with 1,2-DAB, 35 mg/kg/day, or saline control, for 3 weeks. 1,2-DAB induced motor weakness and PGNA, irrespective of the genotype. Spna2-calpain breakdown products were not detected in mutant mice, which displayed a normal structure of the nervous system under saline treatment. Intriguingly, treatment with 1,2-DAB reduced the abundance of the caspase-specific 120-kDa Spna2 breakdown products. Our findings indicate that degradation of Spna2 by calpain- and/or caspase is not central to the pathogenesis of 1,2-DAB axonopathy. In addition, the Spna2-CSD seems to be not required for the maintenance of the cytoskeleton integrity. Our conceptual framework offers opportunities to study the role of calpain-caspase cross talk, including that of the protease degradomics, in models of axonal degeneration.

Topics: Amyotrophic Lateral Sclerosis; Animals; Calpain; Carrier Proteins; Caspases; Disease Models, Animal; Genetic Engineering; Giant Axonal Neuropathy; Male; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Microfilament Proteins; Spectrin

Both mislocalization of TDP-43 and downregulation of RNA-editing enzyme ADAR2 co-localize in the motor neurons of amyotrophic lateral sclerosis patients, but how they are linked is not clear. Here we demonstrate that activation of calpain, a Ca2+-dependent cysteine protease, by upregulation of Ca2+-permeable AMPA receptors generates carboxy-terminal-cleaved TDP-43 fragments and causes mislocalization of TDP-43 in the motor neurons expressing glutamine/arginine site-unedited GluA2 of conditional ADAR2 knockout (AR2) mice that mimic the amyotrophic lateral sclerosis pathology. These abnormalities are inhibited in the AR2res mice that express Ca2+-impermeable AMPA receptors in the absence of ADAR2 and in the calpastatin transgenic mice, but are exaggerated in the calpastatin knockout mice. Additional demonstration of calpain-dependent TDP43 fragments in the spinal cord and brain of amyotrophic lateral sclerosis patients, and high vulnerability of amyotrophic lateral sclerosis-linked mutant TDP43 to cleavage by calpain support the crucial role of the calpain-dependent cleavage of TDP43 in the amyotrophic lateral sclerosis pathology.

Topics: Amyotrophic Lateral Sclerosis; Animals; Blotting, Western; Calpain; DNA-Binding Proteins; HeLa Cells; Humans; Mice; Mice, Knockout; Mice, Mutant Strains; Motor Neurons; Up-Regulation

Amyotrophic lateral sclerosis (ALS) is an adult-onset, rapidly progressing, fatal disease occurring in both familial and sporadic forms. Mutations in the gene encoding Cu/Zn superoxide dismutase (SOD1) cause ALS through a gain of toxic function. Calpain activity is increased in mutant SOD1 (SOD1(G93A)) transgenic mice and in models of ischemia because of increased cytosolic calcium, which has been documented in motor neurons in rodent models of familial ALS and in sporadic ALS patients. We report that inhibition of calpain activity using calpastatin prevented the toxicity of SOD1(G93A) in motor neurons of dissociated spinal cord cultures, prolonging viability of and reducing the proportion containing SOD1(G93A) inclusions. The data support the central role of calcium dysregulation in ALS and identify a potential therapeutic pathway.

Topics: Amyotrophic Lateral Sclerosis; Animals; Calcium-Binding Proteins; Calpain; Cell Aggregation; Cell Survival; Cells, Cultured; Cysteine Proteinase Inhibitors; Gene Transfer Techniques; Mice; Motor Neurons; Superoxide Dismutase

Microdialysis perfusion of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) in rat lumbar spinal cord produces severe motoneuron damage and consequently hindlimb paralysis. Here we studied the time course of the AMPA-induced neurodegenerative changes and motor alterations, and the protective effect of leupeptin, an inhibitor of calpain, a Ca(2+)-activated protease. Paralysis occurs at 4-6 h after AMPA perfusion, but cresyl violet staining showed that motoneuron damage starts at about 3 h and progresses until reaching 50% neuronal loss at 6 h and 90% loss at 12 h. In contrast, choline acetyltransferase (ChAT) immunohistochemistry revealed that the enzyme is already decreased at 30 min after AMPA perfusion and practically disappears at 3 h. Microdialysis coperfusion of leupeptin with AMPA prevented the motor alterations and paralysis and remarkably reduced both the decrement in ChAT immunoreactivity and the loss of motoneurons. We conclude that an increased Ca(2+) influx through Ca(2+)-permeable AMPA receptors activates calpain, and as a consequence ChAT content decreases earlier than other Ca(2+)-dependent processes, including the proteolytic activity of calpain, cause the death of motoneurons.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Amyotrophic Lateral Sclerosis; Animals; Calpain; Cysteine Proteinase Inhibitors; Excitatory Amino Acid Agonists; Leupeptins; Male; Motor Neurons; Rats; Rats, Wistar; Rotarod Performance Test; Spinal Cord

Familial amyotrophic lateral sclerosis (fALS) is caused by mutations in Cu/Zn-superoxide dismutase (SOD1), and SOD1 aggregation and calcium toxicity are involved in neuronal death. However, the effect of altered calcium homeostasis on the SOD1 aggregation is unknown. To investigate whether calcium triggers mutant SOD1 aggregation in vitro, human mutant SOD1 (G93A) was transfected into motor neuronal cell line (VSC 4.1 cells). These cells were then treated with calcium ionophore A23187 or agents that induce intracellular calcium release like cyclic ADP ribose, ryanodine or thapsigargin. A23187 was found to increase mutant SOD1 aggregation and neuronal nitric oxide synthase (nNOS) expression. Moreover, the NOS inhibitor (L-NAME) and a NO-dependent cyclic GMP cascade inhibitor (ODQ) reduced SOD1 aggregation, whereas an exogenous NO donor (GSNO) increased mutant SOD1 aggregation, which was also prevented by NOS or cGMP cascade inhibitor. Our data demonstrate that calcium-influx increases SOD1 aggregation by upregulating NO in cultured motor neuronal cells.

Topics: Amyotrophic Lateral Sclerosis; Animals; Calcimycin; Calcium; Calpain; Caspase 3; Cell Line; Humans; Ionophores; Motor Neurons; Multiprotein Complexes; Mutation; Nitric Oxide; Rats; Recombinant Proteins; Superoxide Dismutase; Superoxide Dismutase-1; Transfection

Amyotrophic lateral sclerosis (ALS) is characterized by the selective degeneration of motor neurons. The cause for nerve cell demise is not clear but involves activation of the caspase family of cysteine proteases. We have shown that ER stress and caspase-12 activation occur in ALS transgenic mice carrying the mutant copper/zinc superoxide dismutase (SOD1) gene. In these mice, we found that the antiapoptotic proteins, X-linked Inhibitor of Apoptosis Protein (XIAP) and the related protein, MIAP2 were decreased. To study the role of this, we generated double transgenic mice expressing XIAP in ALS spinal cord neurons using the Thy1 promoter. Overexpression of XIAP inhibited caspase-12 cleavage and reduced calpain activity in the ALS mice. XIAP also reduced the breakdown of calpastatin that is an inhibitor of calpain. In the double transgenic mice, life span was increased by about 12%. These data support the view that XIAP has beneficial effects in ALS and extends survival. The neuroprotective effect of XIAP involves inhibition of caspases and the stabilization of the calpastatin/calpain system that is altered in the ALS mice.

Topics: Amyotrophic Lateral Sclerosis; Animals; Baculoviral IAP Repeat-Containing 3 Protein; Calcium-Binding Proteins; Calpain; Caspase 12; Caspases; Cell Survival; Cysteine Proteinase Inhibitors; Humans; Inhibitor of Apoptosis Proteins; Mice; Mice, Transgenic; Motor Neurons; Spinal Cord; Superoxide Dismutase; Superoxide Dismutase-1; Survival Rate; Ubiquitin-Protein Ligases; X-Linked Inhibitor of Apoptosis Protein

Here, we investigated the effect of calpain inhibitors on apoptosis in organotypic adult spinal cord slices from mice. An increase in calpain I immunoreactivity was found in the nuclei of motor neurons from slices cultured for 30 min. After 4 h, the immunopositive motor neurons exhibited apoptotic changes including nuclear and chromatin condensation. Eight hours after excision, most motor neurons showed nuclear apoptotic features. Two calpain inhibitors, leupeptin and calpain inhibitor XI, inhibited apoptosis in the motor neurons while the caspase inhibitor Z-VAD.fmk had no effect. Leupeptin, but not calpain inhibitor XI and Z-VAD.fmk, also inhibited nucleosomal DNA fragmentation. These results suggest the involvement of calpain I in the induction of apoptosis in motor neurons of adult spinal cord and that apoptosis can be triggered independent of caspase activation.

Topics: Age Factors; Amyotrophic Lateral Sclerosis; Animals; Apoptosis; Calpain; Caspases; Cysteine Proteinase Inhibitors; DNA Fragmentation; Female; Glycoproteins; Immunohistochemistry; Leupeptins; Mice; Motor Neurons; Nerve Degeneration; Organ Culture Techniques; Spinal Cord; Spinal Cord Injuries; Time Factors

Calpain is thought to be involved in muscular degradation in progressive muscular dystrophy (PMD), especially Duchenne and Becker muscular dystrophies. To assess the expression of calpain genes in skeletal muscles of patients with myopathies, we examined mRNA levels of three calpain isoforms by the quantitative reverse transcriptase-polymerase chain reaction method in biopsied muscles from control, PMD and amyotrophic lateral sclerosis (ALS) patients. There was a statistically significant increase in calpain 1 and calpain 2 mRNA levels in PMD and ALS patients as compared to controls. In contrast, there was a decrease in expression of calpain 3 mRNA in PMD, but it was not statistically significant. Expression of calpain 1 and calpain 2 positively correlated with each other, but not with calpain 3. These results indicate that expression of calpain 1 and calpain 2, but not calpain 3, are upregulated in diseased human muscles, likely playing a regulatory role in the process of myofibrillar degradation at the transcriptional as well as posttranslational level.

Topics: Adolescent; Adult; Aged; Amyotrophic Lateral Sclerosis; Calpain; Child; DNA Primers; Female; Humans; Isoenzymes; Male; Middle Aged; Muscle, Skeletal; Muscular Dystrophies; Polymerase Chain Reaction; RNA, Messenger; Up-Regulation