Copper Toxicity Associated With an ATP7A-Related Complex Phenotype.

BACKGROUND: The ATP7A gene encodes a copper transporter whose mutations cause Menkes disease, occipital horn syndrome (OHS), and, less frequently, ATP7A-related distal hereditary motor neuropathy (dHMN). Here we describe a family with OHS caused by a novel mutation in the ATP7A gene, including a patient with a comorbid dHMN that worsened markedly after being treated with copper histidinate. METHODS: We studied in detail the clinical features of the patients and performed a genomic analysis by using TruSight One Expanded Sequencing Panel. Subsequently, we determined the ATP7A and ATP7B expression levels, mitochondrial membrane potential, and redox balance in cultured fibroblasts of Patient 1. RESULTS: We found a novel ATP7A late truncated mutation p.Lys1412AsnfsX15 in the two affected members of this family. The co-occurrence of OHS and dHMN in Patient 1 reveals the variable phenotypic expressivity of the variant. A severe clinical and neurophysiologic worsening was observed in the dHMN of Patient 1 when he was treated with copper replacement therapy, with a subsequent fast recovery after the copper histidinate was withdrawn. Functional studies revealed that the patient had low levels of both ATP7A and ATP7B, the other copper transporter, and high levels of superoxide ion in the mitochondria. CONCLUSIONS: Our findings broaden the clinical spectrum of ATP7A-related disorders and demonstrate that two clinical phenotypes can occur in the same patient. The copper-induced toxicity and low levels of both ATP7A and ATP7B in our patient suggest that copper accumulation in motor neurons is the pathogenic mechanism in ATP7A-related dHMN.

Severe elastolysis in hereditary gelsolin (AGel) amyloidosis.

AGel amyloidosis is a dominantly inherited systemic amyloidosis caused by mutations p.D214N or p.D214Y resulting in gelsolin amyloid (AGel) formation. AGel accumulates extracellularly in many tissues and alongside elastic fibres. AGel deposition associates with elastic fibre degradation leading to severe clinical manifestations, such as cutis laxa and angiopathic complications. We analysed elastic fibre pathology in dermal and vascular tissue and plasma samples from 35 patients with AGel amyloidosis and 40 control subjects by transmission electron microscopy, immunohistochemistry and ELISA methods. To clarify the pathomechanism(s) of AGel-related elastolysis, we studied the roles of MMP-2, -7, -9, -12 and -14, TIMP-1 and TGFß. We found massive accumulation of amyloid fibrils along elastic fibres as well as fragmentation and loss of elastic fibres in all dermal and vascular samples of AGel patients. Fibrils of distinct types formed fibrous matrix. The degradation pattern of elastic fibres in AGel patients was different from the age-related degradation in controls. The elastin of elastic fibres in AGel patients was strongly decreased compared to controls. MMP-9 was expressed at lower and TGFß at higher levels in AGel patients than in controls. The accumulation of amyloid fibrils with severe elastolysis characterises both dermal and vascular derangement in AGel amyloidosis.

Small amounts of functional ATP7A protein permit mild phenotype.

Mutations in ATP7A lead to at least three allelic disorders: Menkes disease (MD), Occipital horn syndrome and X-linked distal motor neuropathy. These disorders are mainly seen in male individuals, but a few affected females have been described. More than 400 different mutations have been identified in the ATP7A gene. We have conducted several studies in the hope of uncovering the relationship between genotype and phenotype. We have examined the X-inactivation pattern in affected females, the effect of exon-deletions and--duplications, and splice-site mutations on the composition and amount of ATP7A transcript, and we have examined the structural location of missense mutations. The X-inactivation pattern did not fully explain the manifestation of MD in a small fraction of carriers. Most of the affected females had preferential inactivation of the X-chromosome with the normal ATP7A gene, but a few individuals exhibited preferential inactivation of the X-chromosome with the mutated ATP7A gene. The observed mild phenotype in some patients with mutations that effect the composition of the ATP7A transcript, seems to be explained by the presence of a small amount of normal ATP7A transcript. The location of missense mutations on structural models of the ATP7A protein suggests that affected conserved residues generally lead to a severe phenotype. The ATP7A protein traffics within the cells. At low copper levels, ATP7A locates to the Trans-Golgi Network (TGN) to load cuproenzymes with copper, whereas at higher concentrations, ATP7A shifts to the post-Golgi compartments or to the plasma membrane to export copper out of the cell. Impaired copper-regulation trafficking has been observed for ATP7A mutants, but its impact on the clinical outcome is not clear. The major problem in patients with MD seems to be insufficient amounts of copper in the brain. In fact, prenatal treatment of mottled mice as a model for human MD with a combination of chelator and copper, produces a slight increase in copper levels in the brain which perhaps leads to longer survival and more active behavior. In conclusion, small amounts of copper at the right location seem to relieve the symptoms.

Golgi pH, its regulation and roles in human disease.

Most organelles within the exocytic and endocytic pathways typically acidify their interiors, a phenomenon that is known to be crucial for their optimal functioning in eukaryotic cells. This review highlights recent advances in our understanding of how Golgi acidity is maintained and regulated, and how its misregulation contributes to organelle dysfunction and disease. Both its biosynthetic products (glycans) and protein-sorting events are highly sensitive to changes in Golgi luminal pH and are affected in certain human disease states such as cancers and cutis laxa. Other potential disease states that are caused by, or are associated with, Golgi pH misregulation will also be discussed.

Inherited copper transport disorders: biochemical mechanisms, diagnosis, and treatment.

Copper is an essential trace element required by all living organisms. Excess amounts of copper, however, results in cellular damage. Disruptions to normal copper homeostasis are hallmarks of three genetic disorders: Menkes disease, occipital horn syndrome, and Wilson's disease. Menkes disease and occipital horn syndrome are characterized by copper deficiency. Typical features of Menkes disease result from low copper-dependent enzyme activity. Standard treatment involves parenteral administration of copper-histidine. If treatment is initiated before 2 months of age, neurodegeneration can be prevented, while delayed treatment is utterly ineffective. Thus, neonatal mass screening should be implemented. Meanwhile, connective tissue disorders cannot be improved by copper-histidine treatment. Combination therapy with copper-histidine injections and oral administration of disulfiram is being investigated. Occipital horn syndrome characterized by connective tissue abnormalities is the mildest form of Menkes disease. Treatment has not been conducted for this syndrome. Wilson's disease is characterized by copper toxicity that typically affects the hepatic and nervous systems severely. Various other symptoms are observed as well, yet its early diagnosis is sometimes difficult. Chelating agents and zinc are effective treatments, but are inefficient in most patients with fulminant hepatic failure. In addition, some patients with neurological Wilson's disease worsen or show poor response to chelating agents. Since early treatment is critical, a screening system for Wilson's disease should be implemented in infants. Patients with Wilson's disease may be at risk of developing hepatocellular carcinoma. Understanding the link between Wilson's disease and hepatocellular carcinoma will be beneficial for disease treatment and prevention.

Loss-of-function mutations in ATP6V0A2 impair vesicular trafficking, tropoelastin secretion and cell survival.

Autosomal recessive cutis laxa type 2 (ARCL2), a syndrome of growth and developmental delay and redundant, inelastic skin, is caused by mutations in the a2 subunit of the vesicular ATPase H+-pump (ATP6V0A2). The goal of this study was to define the disease mechanisms that lead to connective tissue lesions in ARCL2. In a new cohort of 17 patients, DNA sequencing of ATP6V0A2 detected either homozygous or compound heterozygous mutations. Considerable allelic and phenotypic heterogeneity was observed, with a missense mutation of a moderately conserved residue p.P87L leading to unusually mild disease. Abnormal N- and/or mucin type O-glycosylation was observed in all patients tested. Premature stop codon mutations led to decreased ATP6V0A2 mRNA levels by destabilizing the mutant mRNA via the nonsense-mediated decay pathway. Loss of ATP6V0A2 either by siRNA knockdown or in ARCL2 cells resulted in distended Golgi cisternae, accumulation of abnormal lysosomes and multivesicular bodies. Immunostaining of ARCL2 cells showed the accumulation of tropoelastin (TE) in the Golgi and in large, abnormal intracellular and extracellular aggregates. Pulse-chase studies confirmed impaired secretion and increased intracellular retention of TE, and insoluble elastin assays showed significantly reduced extracellular deposition of mature elastin. Fibrillin-1 microfibril assembly and secreted lysyl oxidase activity were normal in ARCL2 cells. TUNEL staining demonstrated increased rates of apoptosis in ARCL2 cell cultures. We conclude that loss-of-function mutations in ATP6V0A2 lead to TE aggregation in the Golgi, impaired clearance of TE aggregates and increased apoptosis of elastogenic cells.

Vacuolar H+-ATPase meets glycosylation in patients with cutis laxa.

Glycosylation of proteins is one of the most important post-translational modifications. Defects in the glycan biosynthesis result in congenital malformation syndromes, also known as congenital disorders of glycosylation (CDG). Based on the iso-electric focusing patterns of plasma transferrin and apolipoprotein C-III a combined defect in N- and O-glycosylation was identified in patients with autosomal recessive cutis laxa type II (ARCL II). Disease-causing mutations were identified in the ATP6V0A2 gene, encoding the a2 subunit of the vacuolar H(+)-ATPase (V-ATPase). The V-ATPases are multi-subunit, ATP-dependent proton pumps located in membranes of cells and organels. In this article, we describe the structure, function and regulation of the V-ATPase and the phenotypes currently known to result from V-ATPase mutations. A clinical overview of cutis laxa syndromes is presented with a focus on ARCL II. Finally, the relationship between ATP6V0A2 mutations, the glycosylation defect and the ARCLII phenotype is discussed.

Stretching of the Mueller muscle results in involuntary contraction of the levator muscle.

PURPOSE: Since the levator muscle involuntarily and tonically contracts against the weight and elastic resistance of the upper eyelid to maintain an adequate visual field, a mechanoreceptor such as a muscle spindle or a periodontal mechanoreceptor is thought to be essential for its functioning. It was surmised that the Mueller muscle might act as a serial kind of muscle spindle of the levator muscle. METHODS: The response of the bilateral levator muscles evoked by stretching the Mueller muscle of each eyelid of 87 patients with dermatochalasis or aponeurotic blepharoptosis was electromyographically and photographically recorded. RESULTS: Stretching of the unilateral Mueller muscle evoked contraction of the ipsilateral levator muscle in 18 and of the bilateral levator muscle in 69 of the 87 patients. CONCLUSIONS: The Mueller muscle can be thought of as a large, serial kind of muscle spindle, so that stretching by voluntary phasic contraction of the levator muscle for initial eye opening may evoke an afferent impulse to the mesencephalic trigeminal nucleus. Subsequently, this nucleus may stimulate the central caudal nucleus of the oculomotor nuclear complex, leading to involuntary tonic contraction of the ipsilateral or bilateral levator muscles, in the form of a continuous stretch reflex, to maintain an adequate visual field.

Eyelid kinematics following blepharoplasty.

PURPOSE: This study characterizes the effects of blepharoplasty on blink dynamics in subjects with dermatochalasis. The authors evaluate the hypothesis that orbicularis oculi removal and the consequent alterations in blink are potentially harmful consequences of blepharoplasty. METHODS: Sixteen patients were studied, before and after laser blepharoplasty, by a modified scleral search coil technique. Changes in lid position during blinks were recorded before surgery as well as 2 months, and 1 year postoperatively. Off-line analyses assessed blink down-phase amplitude, peak velocity, duration, and main sequence (peak velocity versus amplitude) relationships. RESULTS: Despite muscle resection, there was no significant compromise of mean blink down-phase amplitude, peak velocity, or main sequence following blepharoplasty. Mean blink duration was likewise unchanged at either follow-up session from the preoperative state. Our data show that upper lid blepharoplasty does not cause any lasting decrement in lid function in blinking. CONCLUSIONS: Blepharoplasty includes resection of a portion of the orbicularis oculi. It appears unlikely that the purposeful resection of preseptal portion of the orbicularis oculi that accompanies blepharoplasty is responsible for any functional complications such as dry eye.

Cutis laxa: etiology, pathophysiology, characteristics, and management.

Cutis laxa causes premature aging of the skin, which results in an abnormal appearance. Corrective, not aesthetic, surgery is one treatment option. A blepharoplasty and facelift would be two procedures that can be performed.

Dermatochalasis and dry eye.

The medical records of 141 patients with dermatochalasis seen during the 32-month period of January 1989 through August 1991 were reviewed. Patients were classified on the basis of symptoms and mode of treatment and were examined for effectiveness of blepharoplasty in ameliorating these symptoms. Seventy-three patients (51.8%) had symptoms similar to those found in keratoconjunctivitis sicca, including mattering, burning, itching, redness, epiphora, foreign-body sensation, and photophobia. Of these 73 patients, 38 (52.1%) underwent upper eyelid blepharoplasty. Subjective improvement in symptoms was achieved in 33 of these patients (86.8%) postoperatively. Upper eyelid blepharoplasty may represent an effective component in the treatment of patients with dermatochalasis and dry-eye symptoms.

Cutaneous extensibility in health and disease.

The results are recorded of a number of studies performed over the last few years using a simple and reproducible method for investigating the extensibility of intact skin. A major advantage of this method lies in the fact that it causes the patient no discomfort and leaves his skin without blemish. We have found abnormal behaviour of skin in response to applied stress in the heritable connective tissue disorders: Ehlers-Danlos Syndrome, cutis laxa, and pseudoxanthoma elasticum, and in the acquired disorders of acromegaly, hypopituitarism and scleroderma. An abnormal elastic modulus, however, was found only in pseudoxanthoma elasticum and the growth-hormone disorders.