Biomedical subjects

Eduardo Bonilla

Publications and source records attributed to Eduardo Bonilla.

At least 19 recordsLinked to original sources

PubMed2007

Caveolin-1(-/-)- and caveolin-2(-/-)-deficient mice both display numerous skeletal muscle abnormalities, with tubular aggregate formation.

Here, we examine the role of "non-muscle" caveolins (Cav-1 and Cav-2) in skeletal muscle biology. Our results indicate that skeletal muscle fibers from male Cav-1(-/-) and Cav-2(-/-) mice show striking abnormalities, such as tubular aggregates, mitochondrial proliferation/aggregation, and increased numbers of M-cadherin-positive satellite cells. Notably, these skeletal muscle defects were more pronounced with increasing age. Because Cav-2-deficient mice displayed normal expression levels of Cav-1, whereas Cav-1-null mice exhibited an almost complete deficiency in Cav-2, these skeletal muscle abnormalities seem to be due to loss of Cav-2. Thus, Cav-2(-/-) mice represent a novel animal model-and the first genetically well-defined mouse model-that can be used to study the pathogenesis of tubular aggregate formation, which remains a poorly understood age-related skeletal muscle abnormality. Finally, because Cav-1 and Cav-2 were not expressed within mature skeletal myofibers, our results indicate that development of these abnormalities probably originates in stem/precursor cells, such as satellite cells or myoblasts. Consistent with this hypothesis, skeletal muscle isolated from male Cav-3(-/-) mice did not show any of these abnormalities. As such, this is the first study linking stem cells with the genesis of these intriguing muscle defects.

Animals↗

PubMed2006

Navajo neurohepatopathy is caused by a mutation in the MPV17 gene.

Navajo neurohepatopathy (NNH) is an autosomal recessive disease that is prevalent among Navajo children in the southwestern United States. The major clinical features are hepatopathy, peripheral neuropathy, corneal anesthesia and scarring, acral mutilation, cerebral leukoencephalopathy, failure to thrive, and recurrent metabolic acidosis with intercurrent infections. Infantile, childhood, and classic forms of NNH have been described. Mitochondrial DNA (mtDNA) depletion was detected in the livers of two patients, suggesting a primary defect in mtDNA maintenance. Homozygosity mapping of two families with NNH suggested linkage to chromosome 2p24. This locus includes the MPV17 gene, which, when mutated, causes a hepatocerebral form of mtDNA depletion. Sequencing of the MPV17 gene in six patients with NNH from five families revealed the homozygous R50Q mutation described elsewhere. Identification of a single missense mutation in patients with NNH confirms that the disease is probably due to a founder effect and extends the phenotypic spectrum associated with MPV17 mutations.

Adult↗

PubMed2006

Aortic rupture in mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes.

BACKGROUND: Microangiopathy has been well described in the brain and muscle of patients with mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS). OBJECTIVE: To describe a patient with the common A3243G/MELAS point mutation who had aortic rupture and whose mother also died of large vessel rupture. DESIGN: Case report. SETTING: Collaboration between a primary care hospital and 2 academic tertiary care hospitals. RESULTS: Histologically, there was marked disarray of the smooth muscle architecture of the aorta, and immunohistochemical staining with antibodies against the mitochondrial DNA-encoded cytochrome-C oxidase I subunit showed uniformly decreased immunostaining of the endothelial and smooth muscle cells of the aorta and vasa vasorum. Polymerase chain reaction and restriction fragment length polymorphism analysis showed that the mutation load was 40.5% in blood but 85.3% in the blood vessels. CONCLUSIONS: The severe vasculopathy in this patient is probably directly related to the high mutation load in the blood vessels. Although aortic rupture is an unusual manifestation of MELAS, it is an important potential complication in patients undergoing minor surgical procedures.

Adolescent↗

PubMed2006

Clinical spectrum of mitochondrial DNA depletion due to mutations in the thymidine kinase 2 gene.

BACKGROUND: Mitochondrial DNA depletion syndrome is an autosomal recessive disorder characterized by decreased mitochondrial DNA copy numbers in affected tissues. It has been linked to 4 genes involved in deoxyribonucleotide triphosphate metabolism: thymidine kinase 2 (TK2), deoxyguanosine kinase (DGUOK), polymerase gamma (POLG), and SUCLA2, the gene encoding the beta-subunit of the adenosine diphosphate-forming succinyl coenzyme A synthetase ligase. OBJECTIVE: To highlight the variability in the clinical spectrum of TK2-related mitochondrial DNA depletion syndrome. DESIGN: Review of patients and the literature. SETTING: Tertiary care university. PATIENTS: Four patients with mitochondrial DNA depletion syndrome and mutations in the TK2 gene. MAIN OUTCOME MEASURES: Definition of clinical variability. RESULTS: Patient 1 had evidence of lower motoneuron disease and was initially diagnosed as having spinal muscular atrophy type 3. Patient 2, who is alive and ambulatory at age 9 years, presented at age 2 years with a slowly progressive mitochondrial myopathy. Patient 3 had a more severe myopathy, with onset in infancy and death at age 6 years of respiratory failure. Patient 4 had a rapidly progressive congenital myopathy with rigid spine syndrome and he died at age 19 months. CONCLUSION: The clinical spectrum of TK2 mutations is not limited to severe infantile myopathy with motor regression and early death but includes spinal muscular atrophy type 3-like presentation, rigid spine syndrome, and subacute myopathy without motor regression and with longer survival.

Adolescent↗

PubMed2006

Rapamycin reduces disease activity and normalizes T cell activation-induced calcium fluxing in patients with systemic lupus erythematosus.

OBJECTIVE: Systemic lupus erythematosus (SLE) is an autoimmune disease of unknown origin. Current treatment options are often ineffective or poorly tolerated. Recent observations have revealed mitochondrial hyperpolarization and enhanced Ca2+ fluxing in T cells from SLE patients. Rapamycin, a lipophilic macrolide antibiotic that regulates mitochondrial transmembrane potential and Ca2+ fluxing, has been used safely and effectively to treat renal transplant rejection since 1999. In addition, rapamycin has been shown to ameliorate T cell function and to prolong survival in lupus-prone MRL/lpr mice. We therefore undertook the present study to investigate whether rapamycin is beneficial in patients with SLE. METHODS: Nine patients with clinically active SLE that had been treated unsuccessfully with other immunosuppressive medications began therapy with rapamycin, 2 mg/day orally. Disease activity was assessed with the British Isles Lupus Assessment Group (BILAG) score, SLE Disease Activity Index (SLEDAI), and requirement for prednisone therapy. Mitochondrial transmembrane potential and Ca2+ fluxing were assessed by flow cytometry. RESULTS: In patients treated with rapamycin, the BILAG score was reduced by a mean +/- SEM of 1.93 +/- 0.9 (P = 0.0218), the SLEDAI by 5.3 +/- 0.8 (P = 0.00002), and concurrent prednisone use by 26.4 +/- 6.7 mg/day (P = 0.0062) compared with pre-rapamycin treatment. While mitochondrial hyperpolarization persisted, pretreatment cytosolic and mitochondrial Ca2+ levels and T cell activation-induced rapid Ca2+ fluxing were normalized in rapamycin-treated patients. CONCLUSION: Rapamycin appears to be a safe and effective therapy for SLE that has been refractory to traditional medications. Mitochondrial dysfunction and Ca2+ fluxing could serve as biomarkers to guide decisions regarding future therapeutic interventions in SLE.

Adolescent↗

PubMed2006

Signaling abnormalities in systemic lupus erythematosus as potential drug targets.

Systemic lupus erythematosus (SLE) is an autoimmune inflammatory disease characterized by T-cell, B-cell, and dendritic cell dysfunction and antinuclear autoantibody production. Much of the knowledge that has been gained about SLE in recent years is related to molecular signaling abnormalities present in the disease. Signaling through the T-cell receptor (TCR) is affected in SLE by alterations in the localization, amount, and activity of numerous protein kinases. TCR stimulation releases calcium from intracellular stores, which triggers an influx of extracellular calcium and activates the transcription of many genes, including interleukin-2. Short-term calcium fluxing is exaggerated in SLE, but long-term calcium fluxing is diminished and may account for sub-optimal interleukin-2 production. SLE T-cells have persistently hyperpolarized mitochondria associated with increased mitochondrial mass, high levels of reactive oxygen species (ROS) and low levels of ATP, which decrease activation-induced apoptosis and instead predispose T cells for necrosis, thus stimulating inflammation in SLE. The pentose phosphate pathway impacts the mitochondrial potential and represents a target for possible intervention. Nitric oxide (NO) is a potential link to tie together the signaling and mitochondrial abnormalities in SLE. NO-induced mitochondrial biogenesis recapitulates the TCR-stimulated calcium fluxing abnormalities of SLE T-cells. Since mitochondria can store calcium, the increase in mitochondrial mass may be implicated in the aberrant calcium fluxing in SLE T cells. The mammalian target of rapamycin senses the mitochondrial potential and regulates calcium release, serving as a novel target of treatment of SLE.

Adenosine Triphosphate↗

PubMed2005

Congenital or late-onset myopathy in patients with the T14709C mtDNA mutation.

Three patients with different clinical phenotypes harbored the same point mutation at nucleotide 14709 (T14709C) in the tRNAGlu gene of mitochondrial DNA (mtDNA). The first patient was a 21-month-old child with severe congenital myopathy, respiratory distress and mild mental retardation. Muscle biopsy showed about 12% cytochrome c oxidase (COX)-negative ragged-red fibers (RRFs), and markedly decreased activities of mitochondrial respiratory chain complexes I, III and IV. The other two patients were 51- and 55-year-old siblings with slowly progressive myopathy and diabetes mellitus. Muscle biopsy showed focal COX-negative RRFs and decreased activities of complexes I, III and IV. In all three patients, the T14709C mutation was abundant in muscle but present at lower levels in accessible tissues. Previously described patients with the same mutation also showed congenital or late-onset myopathy. Diabetes is frequently associated with both phenotypes and is a clinical clue to the molecular diagnosis.

Cysteine↗

PubMed2005

Novel mitochondrial DNA ND5 mutation in a patient with clinical features of MELAS and MERRF.

BACKGROUND: The mitochondrial DNA gene encoding subunit 5 of complex I (ND5) has turned out to be a hot spot for mutations associated with mitochondrial encephalomyopathy with lactic acidosis and strokelike episodes (MELAS) and various overlap syndromes. OBJECTIVE: To describe a novel mutation in the ND5 gene in a young man man with an overlap syndrome of MELAS and myoclonus epilepsy with ragged-red fibers. DESIGN: Case report. PATIENT: A 25-year-old man had recurrent strokes, seizures, and myoclonus. His mother also had multiple strokes. A muscle biopsy specimen showed no ragged-red fibers but several strongly succinate dehydrogenase-reactive blood vessels. RESULTS: Biochemical analysis showed isolated complex I deficiency and molecular analysis revealed a novel heteroplasmic mutation (G13042A) in the ND5 gene. CONCLUSIONS: These data confirm that ND5 is a genetic hot spot for overlap syndromes, including MELAS and strokelike and myoclonus epilepsy with ragged-red fibers.

Adult↗

PubMed2005

Hereditary ferritinopathy: a novel mutation, its cellular pathology, and pathogenetic insights.

We report a family of French Canadian and Dutch ancestry with hereditary ferritinopathy (neuroferritinopathy) and a novel mutation (C insertion at nt646-647 in exon 4) in the ferritin light chain gene, resulting in a longer than normal protein. Our failure to immunostain most of the abnormal ferritin deposits in the proband with a conformation-dependent monoclonal antibody to ferritin light chain supported a previously postulated conformational change of ferritin light chain in this disease. The posterior putamen and cerebellum were the primary pathologic loci in our proband, but asymptomatic hepatocytic intranuclear accumulations of iron and ferritin also were present. Both neurons and glia displayed highly distinctive, if not pathognomonic, swollen to vacuolated nuclei containing ferritin and iron. Hyaline deposits, again staining for both ferritin and iron, were additional morphologic features that may be unique to the ferritinopathies. The iron, at least in putamen where there was a nearly 40-fold increase, appeared to be both in the ferrous (Fe2+) and ferric (Fe3+) form; it was the most likely cause of the observed neuronal and glial apoptosis. We found morphologic evidence of both lipid peroxidation and abnormal nitration of proteins in putaminal neurons and glia, confirming the expected oxidative stress due to this excessive iron. Biochemical and immunohistochemical abnormalities in mitochondria also were demonstrated, probably due to an imbalance in iron homeostasis that had a deleterious effect on the respiratory chain.

Adult↗

PubMed2005

Clinical and genetic features in two families with MELAS and the T3271C mutation in mitochondrial DNA.

The majority of patients with MELAS (mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes) have the A3243G point mutation. The much rarer T3271C mutation has been reported predominantly in Japanese subjects. Our objective was to better define the clinical phenotype and mutation load in patients with MELAS and the T3271C mutation in mitochondrial DNA. We present clinical and molecular genetic data in two pedigrees with the T3271C mutation. The age at onset was 8 years in one proband and 14 years in the other. Both patients had migrainelike headache, seizures, and strokelike episodes. Mutation loads were quantified in multiple tissues from the patients and from family members by polymerase chain reaction-restriction fragment length polymorphism analysis. The symptoms in both probands were typical of MELAS, and, contrary to previous reports, onset was early. Hearing loss was less common than in typical MELAS, and ragged red fibers were absent. The proportion of mutant genomes was consistently and markedly greater in DNA from urinary sediment than from blood. In the mother of one proband, mutant genomes were detected only in DNA from hair follicles and cheek mucosa The phenotype of patients with the T3271C mutation might not be as distinct as that of the A3243G mutation, as previously described. Our data also suggest that urine is a better source of DNA than blood for diagnosis and that multiple tissues should be studied in maternal relatives, especially when the mutation cannot be detected in blood.

Adolescent↗

PubMed2005

Unusual clinical presentations in four cases of Leigh disease, cytochrome C oxidase deficiency, and SURF1 gene mutations.

Mutations in the SURF1 gene are the most frequent causes of Leigh disease with cytochrome c oxidase deficiency. We describe four children with novel SURF1 mutations and unusual features: three had prominent renal symptoms and one had ragged red fibers in the muscle biopsy. We identified five pathogenic mutations in SURF1: two mutations were novel, an in-frame nonsense mutation (834G-->A) and an out-of-frame duplication (820-824dupTACAT). Although renal manifestations have not been described in association with SURF1 mutations, they can be part of the clinical presentation. Likewise, mitochondrial proliferation in muscle (with ragged red fibers) is most unusual in Leigh disease but might be part of an emerging phenotype.

Cytochrome-c Oxidase Deficiency↗

PubMed2004

Studies of COX16, COX19, and PET191 in human cytochrome-c oxidase deficiency.

BACKGROUND: Cytochrome-c oxidase (COX) is the terminal enzyme of the mitochondrial electron transport chain, and COX deficiency is a common cause of mitochondrial diseases. Cytochrome-c oxidase is composed of 13 subunits, of which 3 are encoded by mitochondrial DNA and 10 by nuclear DNA. Mutations have been identified in each of the 3 mitochondrial DNA genes but in none of the nuclear DNA genes. However, COX deficiency has been attributed to mutations in several nuclear DNA-encoded ancillary proteins needed for COX assembly and function. Despite this progress, the molecular basis of COX deficiency remains elusive in many patients, justifying the identification and screening of additional COX assembly genes, such as COX16, COX19, and PET191. OBJECTIVE: To determine if COX16, COX19, and PET191 are implicated in human COX deficiency. METHODS: Mutation screening was performed on 53 patients with isolated COX deficiency by direct sequencing of COX19 and by single-strand conformational polymorphism analysis for COX16 and PET191. RESULTS: No mutations were found in COX16, COX19, or PET191 in these patients. CONCLUSIONS: The COX16, COX19, and PET191 genes are either not involved or very rarely involved in human COX deficiency. Mutations in additional COX assembly genes remain to be identified.

Child↗

PubMed2004

Mitochondrial DNA deletion in a child with megaloblastic anemia and recurrent encephalopathy.

A 3 1/2-year-old boy presented with megaloblastic anemia and recurrent episodes of severe lactic acidosis and coma. At age 4 years, he developed sepsis and died; postmortem examination failed to show any gross abnormality in any tissue. Biochemical analysis of muscle showed decreased activities for all respiratory chain enzymes except complex II. Muscle histochemistry revealed diffuse cytochrome c oxidase deficiency. Southern blot analysis of mitochondrial DNA from muscle, liver, and blood showed a heteroplasmic single mitochindrial DNA deletion of 2.4 kb, which removed the genes for cytochrome c oxidase I and II and the transfer ribonucleic acid genes for serine and aspartic acid. Single large-scale deletions in mitochondrial DNA have been associated with Pearson's syndrome, Kearns-Sayre syndrome, and progressive external ophthalmoplegia. This patient's presentation is unusual and suggests an overlap between Pearson's syndrome and Kearns-Sayre syndrome.

Acidosis, Lactic↗

PubMed2003

Mitochondrial myopathy of childhood associated with mitochondrial DNA depletion and a homozygous mutation (T77M) in the TK2 gene.

BACKGROUND: The mitochondrial DNA depletion syndrome is an autosomal recessive disorder of infancy or childhood characterized by decreased mitochondrial DNA copy number in affected tissues. Mutations in 2 genes involved in deoxyribonucleotide metabolism, the deoxyguanosine kinase gene (DGK) and the thymidine kinase 2 gene (TK2), have been related to this syndrome. OBJECTIVE: To describe 3 siblings with the myopathic form of mitochondrial DNA depletion syndrome and a homozygous mutation in the TK2 gene. PATIENTS AND METHODS: These children developed normally until 12 to 16 months of age, when they started showing difficulty walking, which rapidly progressed to severe limb weakness. They died of respiratory failure between the ages of 23 and 40 months. Histochemical and biochemical studies of respiratory chain complexes were performed in muscle biopsy specimens. The whole coding region of the TK2 gene was sequenced. RESULTS: Muscle biopsy showed ragged-red cytochrome-c oxidase-negative fibers. All affected siblings had markedly decreased activities of respiratory chain complexes. Southern blot analysis showed severe reduction of the mitochondrial DNA-nuclear DNA ratio in muscle biopsy specimens from all patients, indicating 80% to 90% mitochondrial DNA depletion. Sequencing of the TK2 gene showed a homozygous C-->T transition at nucleotide 228 in exon 5, which changes a threonine to a methionine at position 77 (T77M). CONCLUSIONS: These results document the importance of screening the TK2 gene in patients with myopathic mitochondrial DNA depletion syndrome and confirm that exon 5 is a "hot spot" for TK2 mutations.

Blotting, Southern↗

PubMed2003

Clinical and genetic heterogeneity in progressive external ophthalmoplegia due to mutations in polymerase gamma.

BACKGROUND: The mendelian forms of progressive external ophthalmoplegia (PEO) associated with multiple mitochondrial DNA deletions are clinically heterogeneous disorders transmitted as dominant or recessive traits. Autosomal dominant PEO is caused by mutations in at least 3 genes: adenine nucleotide translocator-1 (ANT1), encoding the muscle-specific adenine nucleotide translocator; chromosome 10 open reading frame 2 (C10orf2), encoding Twinkle helicase; and polymerase gamma (POLG), encoding the alpha subunit of polymerase gamma. Mutations in POLG can also cause autosomal recessive PEO, which is often associated with multisystemic disorders. OBJECTIVE AND METHODS: To further investigate the frequency and genotype-phenotype correlations of mutations in the POLG gene, we used single-stranded conformational polymorphism analysis and direct sequencing to screen 30 patients with familial or sporadic PEO and multiple mitochondrial DNA deletions in muscle but without mutations in ANT1 and C10orf2. RESULTS: Four unrelated patients had novel POLG mutations. A woman with PEO and mental retardation had a heterozygous Gly1076Val mutation. Two patients, one with PEO, exercise intolerance, and gastrointestinal dysmotility and the other with PEO, neuropathy, deafness, and hypogonadism, both had a Pro587Leu change. The fourth patient, who was compound heterozygous for Ala889Thr and Arg579Trp mutations, had PEO, gastrointestinal dysmotility, and neuropathy. These mutations were not detected in 120 healthy control alleles. CONCLUSIONS: Our results demonstrate that POLG mutations account for a substantial proportion of patients (13%) with PEO and multiple mitochondrial DNA deletions and cause both clinically and genetically heterogeneous disorders.

Adult↗

PubMed2003

A novel mitochondrial tRNA(Leu(UUR)) mutation in a patient with features of MERRF and Kearns-Sayre syndrome.

In a patient with clinical features of both myoclonus epilepsy ragged-red fibers (MERRF) and Kearns-Sayre syndrome (KSS), we identified a novel guanine-to-adenine mitochondrial DNA (mtDNA) mutation at nucleotide 3255 (G3255A) of the tRNA(Leu(UUR)) gene. Approximately 5% of the skeletal muscle fibers had excessive mitochondria by succinate dehydrogenase histochemistry while a smaller proportion showed cytochrome c oxidase (COX) deficiency. In skeletal muscle, activities of mitochondrial respiratory chain complexes I, I + III, II + III, and IV were reduced. The G3255A transition was heteroplasmic in all tissues tested: muscle (53%), urine sediment (67%), peripheral leukocytes (22%), and cultured skin fibroblasts (< 2%). The mutation was absent in 50 control DNA samples. Single-fiber analysis revealed a higher proportion of mutation in COX-deficient RRF (94% +/- 5, n = 25) compared to COX-positive non-RRF (18% +/- 9, n = 21). The identification of yet another tRNA(Leu(UUR)) mutation reinforces the concept that this gene is a hot-spot for pathogenic mtDNA mutations.

Adenine↗

PubMed2003

Mitochondrial DNA dysfunction in oncocytic hepatocytes.

Hepatic oncocytes with abundant granular, eosinophilic cytoplasm due to mitochondrial hyperplasia are seen in various chronic liver diseases, particularly chronic hepatitis and cirrhosis. Increased mitochondria in oncocytes are thought to be a compensatory mechanism for deficiencies in the hepatocellular respiratory chain, although the pathogenesis of these deficiencies has been uncertain. We selected seven cases of cirrhosis (six with oncocytes, one without) for the following analysis: histoenzymatic and immunohistochemical staining of several mitochondrial DNA (mtDNA)- and nuclear DNA (nDNA)-encoded respiratory chain enzymes; immunostaining using antibodies against double-strand-DNA (anti-DNA) and against Ki-67 (a cell proliferation marker); and Southern blot analysis for mtDNA and nDNA. Eighty percent of oncocytes showed histoenzymatic and immunohistochemical deficiencies of cytochrome c oxidase and the mtDNA-encoded subunit I of complex IV, with preserved expression of nDNA-encoded succinate dehydrogenase and the iron-sulfur subunit of complex III (FeS). Cytoplasmic (but not nuclear) anti-DNA staining was partially or completely absent in approximately 50% of oncocytes. Three cases with oncocytes studied by Southern blot showed mtDNA reductions of 66%, 71%, and 85%. In conclusion, hepatic oncocytes demonstrate significant deficiencies of mtDNA and mtDNA-encoded respiratory chain enzymes. We propose that mtDNA depletion plays an important role in hepatocellular oncocytic transformation.

Adult↗

PubMed2003

Mutation screening in patients with isolated cytochrome c oxidase deficiency.

Cytochrome c oxidase (COX) deficiency has been associated with a variety of clinical conditions and can be due to mutations in nuclear or mitochondrial genes. Despite recent progress in our understanding of the molecular bases of COX deficiency, the genetic defect remains elusive in many cases. We performed mutation screening in 30 patients with biochemical evidence of isolated COX deficiency and heterogeneous clinical phenotypes. Sixteen patients had various forms of encephalomyopathy, and six of these had the neuroradiological features of Leigh syndrome. Four patients had encephalohepatopathy, six had hypertrophic cardiomyopathy, and four had other phenotypes. We studied the three mtDNA genes encoding COX subunits, the 22 mtDNA tRNA genes, and seven COX assembly genes: SCO1, SCO2, SURF1, COX10, COX11, COX15, and COX17. We report two novel pathogenic SURF1 mutations in a patient with Leigh syndrome and one novel SCO2 mutation in a patient with hypertrophic cardiomyopathy. These data show that heterogeneous clinical phenotypes are associated with COX deficiency, that mutations in mtDNA COX genes are rare, and that mutations in additional genes remain to be identified.

Age of Onset↗