Frank Hanisch

Heterozygous mutations of the kinesin KIF21A in congenital fibrosis of the extraocular muscles type 1 (CFEOM1).

Congenital fibrosis of the extraocular muscles type 1 (CFEOM1; OMIM #135700) is an autosomal dominant strabismus disorder associated with defects of the oculomotor nerve. We show that individuals with CFEOM1 harbor heterozygous missense mutations in a kinesin motor protein encoded by KIF21A. We identified six different mutations in 44 of 45 probands. The primary mutational hotspots are in the stalk domain, highlighting an important new role for KIF21A and its stalk in the formation of the oculomotor axis.

Frequent low penetrance mutations in the Lamin A/C gene, causing Emery Dreifuss muscular dystrophy.

Emery Dreifuss muscular dystrophy is a genetically heterogeneous disorder characterized by the clinical triad of early onset contractures, progressive muscular wasting and weakness with humeroperoneal distribution and cardiac conduction defects. Mutations in the Lamin A/C (LMNA) gene are responsible for the autosomal dominant and the autosomal recessive forms. Familiar and sporadic patients carrying mutations in the LMNA gene show high variability in the clinical symptomatology and age of onset. In this report, we describe four families harboring missense mutations in the LMNA gene and we show that the effect of mutations ranges from silent to fully penetrant. We suggest that incomplete penetrance of dominant mutations in the LMNA gene is a common feature and we emphasize the significance of mutational analysis in relatives of sporadic cases of laminopathies, as asymptomatic carriers face high risk of sudden cardiac death.

Three novel mutations in KIF21A highlight the importance of the third coiled-coil stalk domain in the etiology of CFEOM1

BackgroundCongenital fibrosis of the extraocular muscles types 1 and 3 (CFEOM1/CFEOM3) are autosomal dominant strabismus disorders that appear to result from maldevelopment of ocular nuclei and nerves. We previously reported that most individuals with CFEOM1 and rare individuals with CFEOM3 harbor heterozygous mutations in KIF21A. KIF21A encodes a kinesin motor involved in anterograde axonal transport, and the familial and de novo mutations reported to date predictably alter one of only a few KIF21A amino acids – three within the third coiled-coil region of the stalk and one in the distal motor domain, suggesting they result in altered KIF21A function. To further define the spectrum of KIF21A mutations in CFEOM we have now identified all CFEOM probands newly enrolled in our study and determined if they harbor mutations in KIF21A.ResultsSixteen CFEOM1 and 29 CFEOM3 probands were studied. Three previously unreported de novo KIF21A mutations were identified in three CFEOM1 probands, all located in the same coiled-coil region of the stalk that contains all but one of the previously reported mutations. Eight additional CFEOM1 probands harbored three of the mutations previously reported in KIF21A; seven had one of the two most common mutations, while one harbored the mutation in the distal motor domain. No mutation was detected in 5 CFEOM1 or any CFEOM3 probands.ConclusionAnalysis of sixteen CFEOM1 probands revealed three novel KIF21A mutations and confirmed three reported mutations, bringing the total number of reported KIF21A mutations in CFEOM1 to 11 mutations among 70 mutation positive probands. All three new mutations alter amino acids in heptad repeats within the third coiled-coil region of the KIF21A stalk, further highlighting the importance of alterations in this domain in the etiology of CFEOM1.

Phenotype of matrin-3-related distal myopathy in 16 German patients.

To characterize the phenotype of patients with distal myopathy with vocal cord and pharyngeal weakness due to the p.S85C mutation in the matrin‐3 gene (MATR3, Mendelian Inheritance in Man 164015). Recently, it has been suggested that patients with this mutation may suffer from familial amyotrophic lateral sclerosis.

Influence of moderate and profound hyperventilation on cerebral blood flow, oxygenation and metabolism

OBJECTIVE The aim of the present study was to examine the impact of moderate and profound hyperventilation on regional cerebral blood flow (rCBF), oxygenation and metabolism. MATERIALS AND METHODS Twelve anesthetized pigs were subjected to moderate (mHV) and profound (pHV) hyperventilation (target arterial pO(2): 30 and 20 mmHg, respectively) for 30 min each, after baseline normoventilation (BL) for 1 h. Local cerebral extracellular fluid (ECF) concentrations of glucose, lactate, pyruvate and glutamate as well as brain tissue oxygenation (p(ti)O(2)) were monitored using microdialysis and a Licox oxygen sensor, respectively. In nine pigs, regional cerebral blood flow (rCBF) was also continuously measured via a thermal diffusion system. RESULTS Both moderate and profound hyperventilation resulted in a significant decrease in rCBF (BL: 37.9+/-4.3 ml/100 g/min; mHV: 29.4+/-3.6 ml/100 g/min; pHV: 23.6+/-4.7 ml/100 g/min; p<0.05) and p(ti)O(2) (BL: 22.7+/-4.1 mmHg; mHV: 18.9+/-4.9 mmHg; pHV: 13.0+/-2.2 mmHg; p<0.05). A p(ti)O(2) decrease below the critical threshold of 10 mmHg was induced in three animals by moderate hyperventilation and in five animals by profound hyperventilation. Furthermore, significant increases in lactate (BL: 1.06+/-0.18 mmol/l; mHV: 1.36+/-0.20 mmol/l; pHV: 1.67+/-0.17 mmol/l; p<0.005), pyruvate (BL: 46.4+/-7.8 micromol/l; mHV: 58.0+/-10.3 micromol/l; pHV: 66.1+/-12.7 micromol/l; p<0.05), and lactate/glucose ratio were observed during hyperventilation. (Data are presented as mean+/-S.E.M.) CONCLUSIONS Both moderate and profound hyperventilation may result in insufficient regional oxygen supply and anaerobic metabolism, even in the uninjured brain. Therefore, the use of hyperventilation cannot be considered as a safe procedure and should either be avoided or used with extreme caution.

Lactate increase and oxygen desaturation in mitochondrial disorders – Evaluation of two diagnostic screening protocols

BackgroundMitochondrial disorders are characterized by an accumulation of lactate and an insufficient oxygen extraction from blood during exercise. Therefore, both parameters (lactate and oxygen saturation) can be used as screening tests in mitochondrial disorders. However, conflicting results regarding sensitivities and specifities of both tests have been reported.MethodWe examined 27 patients with genetically defined mitochondrial disorders (single deletions n = 15,multiple deletions n = 5, A3243G mutation n = 7), patients with other neuromuscular disorders, and healthy controls. In the first test subjects performed intermittent isometric handgrip exercise (0.5 Hz) at 80 % (3 minutes) and 30 % (3 and 15 minutes) of maximal contraction force (MCF). Oxygen saturation and partial pressure in cubital venous blood from the exercising arm were measured. In the second test subjects underwent cycle ergometry at 30 W for 15 minutes. Venous lactate at rest, during and 15 minutes postexercise was determined.ResultBoth tests showed specificities of 92–96%. Sensitivities for changes of venous oxygen partial pressure and oxygen saturation ranged from 21–26% at 80% MCF for 3 minutes to 47–58% at 30% MCF for 15 minutes. Sensitivities for venous resting, peak, and post–exercise lactate was 33%, 58%, and 67%, respectively. The degree of deoxygenation, however,was independent of the intensity and duration of the applied forces. Oxygen desaturation and lactate increase in patients with mitochondrial disorders were not different in patients with and without clinical symptoms of myopathy. There were significant correlations between the heteroplasmy and both the degree of oxygen desaturation and lactate increase in patients with single deletions. In patients who performed both protocols (n = 16) a combination of both tests increased sensitivity up to 87%.ConclusionOxygen desaturation in forearm exercise tests and lactate increase in cycle ergometry tests show a high specifity but only moderate sensitivity. Combination of the two screening test clearly increases the sensitivity.

MuSK–antibody positive pure ocular myasthenia gravis

Sirs: Antibodies (Ab) against the muscle-specific receptor tyrosine kinase (MuSK) were found in 38–58 % of patients with so-called seronegative, (that means acetylcholine receptor (AChR) Ab negative) generalized myasthenia gravis (MG) [1–3]. Ocular symptoms are frequently the initial findings in MuSK-Ab-positive MG [4]. In two series, however, MuSK-Abs could not be detected in 18 and 13 patients with pure ocular MG, respectively [4, 5]. Therefore, it was concluded that patients with MuSK-Ab carry a high risk of early generalization. We present a case with MuSK-Ab positive MG and isolated bilateral ptosis for three years. The 57-year old German female patient had noticed increasing bilateral ptosis, worse in the evening, which eventually impaired vision. The symptoms had lasted for at least two years. Clinical examination showed bilateral symmetrical ptosis with a palpebral fissure of 5 mm. Ptosis did not improve after i. v. application of edrophonium chloride. There was no evidence of further ocular, bulbar, or generalized involvement. There was no pathological decrease in the orbicularis oris muscle power after repetitive 3/s stimulation of the facial nerve. AChR-, titin-, MAK-, TAK-, and TRAK antibodies were not found. The MuSK-antibody ratio was 13 (normal ratio < 10, Laboratory Prof. Seelig & Kollegen, Karlsruhe, Germany) [6]. Ptosis responded well upon treatment with up to 330 mg/d pyridostigmine and 150 mg/d azathioprine (palpebral fissure became 8–10 mm wide). No generalized symptoms have developed during treatment for twelve months. During that time the MuSK-antibody ratio decreased to a ratio of 8.5. Out of our five patients with MuSK-Ab two others showed initially isolated ocular involvement: One female patient’s symptoms became generalized within three months. The other female patient has been showing isolated ptosis for 15 months (being untreated for two months, treated for 13 months). Initially isolated ocular symptoms occur in about 50 % of AChRAb positive MG. The progression from pure ocular MG into generalized MG is very rare after a disease course of > 2 years. Therefore, most of the studies on ocular MG used an observation period of two years [7–9]. However, neither the Ossermann Classification nor the MGFA Clinical Classification refers to a temporal category to distinguish between MG with initial ocular symptoms and pure ocular MG [10, 11]. Within two years 34–86 % of untreated AChR-Ab positive patients with ocular involvement, but only 7–17 % of patients treated with immunosuppressive drugs progressed into a generalized MG [7–9]. Initial ocular symptoms, either isolated or associated with other symptoms were found in 6/10 patients with MuSK-Ab positive generalized MG [4]. Recently, one MuSK-Ab positive female with ptosis and diplopia was reported, for whom immunosuppressive treatment was started three months after the onset of ocular symptoms and which did not generalize during a course of more than twelve months [12]. Since our patient remained with only ocular symptoms for at least two years without treatment her disorder can clearly be classified as ocular myasthenia. In conclusion our case indicates that a small proportion of cases with MuSK-Ab stay as a pure ocular MG over a period exceeding two years.

Characteristics of pain in amyotrophic lateral sclerosis.

Pain is an often underestimated and neglected symptom in amyotrophic lateral sclerosis (ALS).

AMP deaminase deficiency in skeletal muscle is unlikely to be of clinical relevance

BackgroundThe homozygous c.34C>T mutation in the AMPD1 gene encoding the muscle-specific isoform of AMP deaminase (AMPD) accounts for the vast majority of inherited skeletal muscle AMPD deficiencies. It is controversial (i) whether AMPD deficiency is associated with exercise-induced complaints and (ii) whether an acquired form exists in which an underlying neuromuscular disorder additionally lowers the AMPD activity.Designc.34C>T mutation (homozygous- TT, heterozygous-CT,wildtype-CC) was screened in 107 healthy blood donors and 294 patients with skeletal muscle biopsy including 200 with exercise-induced complaints. Additional screening for c.468G > T and c.860A > T mutations was performed in all CT.AMPD was analysed histochemically and biochemically.ResultsThe mutant allele frequency (MAF) was not different in blood donors, patients with muscle biopsy, and the subgroup with exercise-induced complaints (12.9–16.8 %). CT was found in 63 and TT in 13 patients. The c.468G>T mutation was not detected. The c.860A > T mutation was found in 2.8 % blood donors and 1.6% CT. AMPD activities showed a substantial overlap in CC and CT, but not with TT. AMPD activities in TT, CT, and CC did not differ in patients with a defined neuromuscular disorder and those with only exercise-induced complaints. In TT, CT, and CC there were no differences in the frequency of signs and symptoms, hyperCKemia, or myopathological biopsy pattern.ConclusionMAF did not differ in normal and diseased populations. The frequency of exertion-induced complaints was not higher in TT than CC. There was no specific phenotype of TT and no evidence for an acquired form of AMPD deficiency.

Pain in adult patients with Pompe disease A cross-sectional survey

BACKGROUND Pompe disease is a rare hereditary metabolic myopathy caused by a deficiency of acid-α-glucosidase. We investigated the presence and severity of pain and its interference with daily activities in a large group of adults with Pompe disease, who we compared with an age-matched control group. METHODS Data were collected in a cross-sectional survey in Germany and The Netherlands. Pain was assessed using the short-form brief pain inventory (BPI). Patients also completed the Short Form-36 item (SF-36v2), the Hospital Anxiety and Depression Scale (HADS) and the Rotterdam Handicap Scale (RHS). RESULTS Forty-five percent of the 124 adult Pompe patients reported having had pain in the previous 24h, against 27% of the 111 controls (p=0.004). The median pain severity score in Pompe patients reporting pain was 3.1 (on a scale from 0 to 10), indicating mild pain; against 2.6 amongst controls (p=0.06). The median score of pain interference with daily activities in patients who reported pain was 3.3, against 1.3 in controls (p=0.001). Relative to patients without pain, those with pain had lower RHS scores (p=0.02), lower SF-36 Physical and Mental component summary scores (p<0.001 and p=0.049), and higher levels of depression and anxiety (p=0.005 and p=0.003). CONCLUSIONS To date, this is one of the largest studies on pain in a specific neuromuscular disorder. Nearly one in two Pompe patients had experienced pain in the previous 24h. Although pain severity and its interference with daily life were mild, pain was related to a reduced quality of life, less participation in daily life, and greater depression and anxiety. Its management should therefore be seen as part of clinical practice involving Pompe patients.