C. Deutrich

A new mutation in the skeletal ryanodine receptor gene (RYR1) is potentially causative of malignant hyperthermia, central core disease, and severe skeletal malformation

Malignant hyperthermia susceptibility (MHS) and central core disease (CCD) have been shown to result from missense mutations in the ryanodine receptor gene of the skeletal muscle (RYR1). A 15‐year‐old patient who had spondylocostal dysostosis (SCD) developed an MH crisis during general anesthesia. The patient was characterized phenotypically by block vertebrae, vertebral fusion, short neck and thorax, fused ribs, craniofacial abnormalities, spina bifida occulta, and a diaphragmatic defect closed surgically in early infancy. The diagnosis MH susceptible (MHS) was confirmed by the in vitro contracture test (IVCT) on a muscle biopsy. Surprisingly, the histopathological investigation revealed the presence of CCD too. Molecular genetic investigation of the RYR1 gene was performed to search for known MH‐related mutations. Cluster regions of the RYR1 gene, in which mutations have already been found, were examined by direct automated sequencing. In addition to the diagnosis MHS and CCD we were able to identify a novel RYR1 mutation in exon 46: 7358ATC > ACC, resulting in an Ile2453Thr substitution. This mutation was also present in the mother, in whom MH disposition and CCD were determined by muscle investigations. We suggest that the newly identified RYR1 mutation is closely associated with MH and CCD. A probable causative role of the RYR1 gene in SCD patients should be assessed by further genetic investigations.

Aktuelle Aspekte der Diagnostik der malignen Hyperthermie

ZusammenfassungFragestellung. In der vorliegenden Arbeit sollen im Überblick Ergebnisse und Erfahrungen aus 15 Jahren MH-Diagnostik am MH-Zentrum in Leipzig dargestellt werden. Ausführlicher wird als neuer Diagnostikzweig die Molekulargenetik vorgestellt und ihre Aussagekraft im Gesamtkonzept der MH-Dispositionsbestimmung erörtert. Methodik. Seit 1986 wird an unserer Klinik der In-vitro-Kontrakturtest (IVKT) als Standardverfahren zur Bestimmung der MH-Disposition angewandt. Potentielle MH-Zwischenfälle wurden zusätzlich mit der Clinical Grading Scale (CGS) analysiert. Ergänzt wurde die MH-Diagnostik 1999 durch molekulargenetische DNA-Untersuchungsmethoden des Gens für den Ryanodinrezeptor der Skelettmuskelzellen (RYR1). Ergebnisse. Von den insgesamt bei 1.456 Patienten durchgeführten Muskeluntersuchungen lieferten 376 im IVKT die Diagnose MH-susceptible (MHS), 121 MH-equivocal (MHE) und 921 MH-negativ (MHN). Darunter waren 309 Patienten mit klinisch MH-verdächtigen Zwischenfällen, bei denen eine MH-Disposition jedoch mehrheitlich durch den IVKT ausgeschlossen wurde (197 MHN). Eine molekulargenetische Untersuchung des RYR1-Gens erfolgte bei 99 unabhängigen MH-Familien; eine RYR1-Mutation konnten in 46 Fällen identifiziert werden. Es wurden 18 verschiedene Punktmutationen nachgewiesen, von denen 4 (Arg401Cys, Ile2182Phe, Gly2375Ala, Ile2453Thr) bisher noch nicht publiziert wurden. Schlussfolgerungen. Zur Beurteilung einer MH-Veranlagung können der IVKT, die DNA-Analyse und einschränkend die klinische Symptomatik herangezogen werden. Der IVKT ist als hoch sensitives und damit sicheres, die DNA-Analyse als sehr spezifisches Verfahren einzuschätzen. Eine alternative Methodenanwendung ist unter definierten Voraussetzungen möglich. Im Gesamtkonzept der präklinischen MH-Diagnostik sollten die Verfahren jedoch nicht konkurrierend, sondern ausgehend von ihrer spezifischen Aussagekraft gezielt oder ergänzend und vor dem Hintergrund eingesetzt werden, um Zwischenfälle in MH-disponierten Familien mit größtmöglicher Sicherheit zu vermeiden.SummaryObjectives. The aim of this work was to give a survey of experiences and results obtained over a period of 15 years of diagnosis of malignant hyperthermia in the MH centre in Leipzig. The new branch of MH diagnosis, the molecular genetics and its general diagnostic potential will be presented in more detail. Methods. The in vitro contracture test (IVCT), which has been used in our department since 1986, represents the standard method for determining disposition to MH and in addition, suspected MH events were analysed by the clinical grading scale (CGS). In 1999, the diagnosis of MH in our centre was supplemented by molecular genetic examination of the skeletal ryanodine receptor gene (RYR1). Results. A total of 1,456 muscle tests (IVCT) in patients with a potential MH disposition, provided 376 MH susceptible (MHS), 121 MH equivocal (MHE) and 921 MH negative (MHN) results. Out of these 309 persons had a previous clinical MH event, but for the majority of these persons a real MH disposition could be excluded by the IVCT (197 MHN). In 99 independent MH families, the RYR1 was genetically screened identifying a mutation in 46, whereby 18 different RYR1 point mutations were found of which 4 (Arg401Cys, Ile2182Phe, Gly2375Ala, Ile2453Thr) have not yet been published. Conclusions. The disposition to MH may be assessed by the IVCT, DNA analysis and with limitations by the clinical phenotype. The IVCT represents a highly specific method, the DNA analysis appears to be very specific. Under defined conditions an alternative use of the methods is possible. However, these methods should not be regarded as in competition but rather their potential should be complementary or used in specific situations in order to avoid non-detection of MH events in affected families.

Spontaneous Occurrence of the Disposition to Malignant Hyperthermia

ONE characteristic of malignant hyperthermia (MH) is the autosomal-dominant mode of inheritance. This implies that at least one parent of an affected patient should be predisposed to MH. In this report, we describe two families in which MH susceptibility developed spontaneously through a neomutation. These families attracted attention because both parents of an MH-susceptible individual were diagnosed as MH-negative in the in vitro contracture test (IVCT). The IVCT was performed with halothane and caffeine according to the protocol of the European MH Group. This test may provide the following diagnoses: MH-susceptible (MH-positive), MH-negative, or MH-equivocal (possible MH-positive). Cluster regions of the skeletal ryanodine receptor gene (RYR1), in which MH-related mutations have been identified, were amplified by polymerase chain reaction and subsequently analyzed by the direct sequencing technique previously described. Chromosome 19 microsatellite markers were amplified by polymerase chain reaction using labeled fluorescent-tag primers (D19S228, 19S421, RYR.PCR1, D19S422, D19S223, as recommended by the European MH Group, genetic section§) and were checked by fragment gel analysis in the ABI PRISM 377 DNA sequencer (Applied Biosystems, Foster City, CA).

Monitoring the jugular-venous oxyhemoglobin saturation as a decision aid for normothermic cardiopulmonary bypass.

Many etiologies for cerebral complications after open heart surgery have been proposed (e.g., emboli, inadequate cerebral perfusion, etc.). Using a continuous measurement of jugular venous oxygen hemoglobin saturation (Sjo 2), Nakajima et al. (1) described hypoxic phases during cardiopulmonary bypass, delineating the rewarming phase as a critical time for cerebral 0 2 supply. They recommended modifying the perfusion technique to incorporate a considerably slower rewarming through which hypoxic events are diminished and thus better cerebral outcome achieved. However, the displayed form of the cerebrovenous saturation curve can be described as a normal case of hypothermic perfusion (see Figure la); its interpretation can yield additional information. The use of continuous Sjo2 monitoring (catheter U440 with Oximetrix 3 system; Abbott) was confined in our clinic to those patients with a carotid artery stenosis undergoing coronary bypass operation. We consequently obtained Sjo 2 curves that were essentially the same as those reported by Nakajima et al. Nevertheless, the problem with cerebral oxygen deficiency does not occur during the rewarming phase. Moreover, it is during the rewarming period that oxygen deficiency that occurred during the cooling phase disappears. The oxygen deficiency associated with cooling is primarily produced by the impaired conditions of oxygen delivery from hemoglobin, a left shift of the oxygen dissociation curve (in the cooling of blood). Whereas during this period the oxygen consumption is only 30%-50% of the normal level, the oxygen requirement of the stillwarm tissue is considerably higher. If the recommendation of Nakajima et al. is followed and the rewarming phase is lengthened, then increased oxygen uptake is simply spread over a longer period and is not obvious from the Sjo2 measurement. The cerebral oxygen supply can only be secured through the lengthening or complete removal of the cooling phase in a normothermic perfusion technique. As a restilt of these observations and considerations, we switched to normothermic perfusion under the maintenance of the myocardial protection using cold cardioplegia and local cooling. Through the monitoring of normothermic perfusion, an important further factor of the pressure dependency of the cerebral blood circulation, and with it Sjo 2, became evident (Figure 2). The inability to evaluate flow restriction in the extraand intracranial fluid area in the elderly population with increasing carotid artery disease undergoing coronary surgery adds additional importance to the monitoring of Sjo2 independent of whether normoor hypothermic bypass is utilized. lb