Anja Lisbeth Frederiksen

Tissue specific distribution of the 3243A→G mtDNA mutation

Background: The 3243A→G is a common pathogenic mitochondrial DNA (mtDNA) point mutation causing a variety of different phenotypes. Segregation of this mutation to different tissues during embryonic life and postnatally is still enigmatic. Objective: To investigate the tissue distribution of this mutation. Methods: In 65 individuals from nine families segregating the 3243A→G mutation, the mutation load (% mutated mtDNA) was determined in various tissues. Mutation load was measured in two to four cell types—blood leucocytes, buccal cells, skeletal muscle cells, and urine epithelial cells (UEC)—derived from all three embryogenic germ layers. Results: There was a significant correlation among mutation loads in the four tissues (r = 0.80–0.89, p<0.0001). With blood serving as reference, the mutation load was increased by 16% in buccal mucosa, by 31% in UEC, and by 37% in muscle. There were significant differences between the mitotic tissues blood, buccal mucosa, and UEC (p<0.0001), but no difference between UEC and muscle. Using the present data as a cross sectional investigation, a negative correlation of age with the mutation load was found in blood, while the mutation load in muscle did not change with time; 75% of the children presented with higher mutation loads than their mothers in mitotic tissues but not in the post-mitotic muscle. Conclusions: There appears to be a uniform distribution of mutant mtDNA throughout the three germ layers in embryogenesis. The significant differences between mutation loads of the individual tissue types indicate tissue specific segregation of the 3243A→G mtDNA later in embryogenesis.

Limited diagnostic value of enzyme analysis in patients with mitochondrial tRNA mutations.

We evaluated the diagnostic value of respiratory chain (RC) enzyme analysis of muscle in adult patients with mitochondrial myopathy (MM). RC enzyme activity was measured in muscle biopsies from 39 patients who carry either the 3243A>G mutation, other tRNA point mutations, or single, large‐scale deletions of mtDNA. Findings were compared with those obtained from asymptomatic relatives with the 3243A>G mutation, myotonic dystrophy patients, and healthy subjects. Plasma lactate concentration, maximal oxygen uptake, and ragged‐red fibers/cytochrome c–negative fibers in muscle were also determined. Only 10% of patients with the 3243A>G point mutation had decreased enzyme activity of one or more RC complexes, whereas this was the case for 83% of patients with other point mutations and 62% of patients with deletions. Abnormal muscle histochemistry was found in 65%, 100%, and 85% of patients, respectively, in these three groups. The results indicate that RC enzyme analysis in muscle is not a sensitive test for MM in adults. In these patients, abnormal muscle histochemistry appears to be a better predictor ofMM. Muscle Nerve, 2010

Prevalence of migraine in persons with the 3243A>G mutation in mitochondrial DNA.

Over the last three decades mitochondrial dysfunction has been postulated to be a potential mechanism in migraine pathogenesis. The lifetime prevalence of migraine in persons carrying the 3243A>G mutation in mitochondrial DNA was investigated.

Distribution of RET mutations in multiple endocrine neoplasia 2 in Denmark 1994-2014: a nationwide study

Background: Germline mutations of the REarranged during Transfection (RET) proto-oncogene cause multiple endocrine neoplasia 2 (MEN2). It is unclear whether the distribution of RET mutations varies among populations. The first nationwide study of the distribution of RET mutations was conducted, and the results were compared to those of other populations. Methods: This retrospective cohort study included 1583 patients who underwent RET gene testing in one of three centers covering all of Denmark between September 1994 and December 2014. Primary testing method was Sanger sequencing, which included exons 8–11 and 13–16. Mutations were defined according to the ARUP database July 1, 2016. Results: RET mutations were identified in 163 patients from 36 apparently unrelated families. Among the 36 families 13 (36.1%) carried mutations in codon 611, four (11.1%) in codon 618, three (8.3%) in codon 620, one (2.8%) in codon 631, six (16.7%) in codon 634, one (2.8%) in codon 790, one (2.8%) in codon 804, one (2.8%) in codon 852, one (2.8%) in codon 883, and five (13.9%) in codon 918. Among the 13 families with codon 611 mutations, 12 had the p.C611Y mutation. Conclusions: The distribution of RET mutations in Denmark appears to differ from that of other populations. Mutations in codon 611 were the most prevalent, followed by more frequently reported mutations. This might be due to a possible founder effect for the p.C611Y mutation. However, further studies are needed to find possible explanations for the skewed mutational spectrum in Denmark.

Incidence and prevalence of multiple endocrine neoplasia 2B in Denmark: a nationwide study

Multiple endocrine neoplasia 2B (MEN2B) is an autosomal dominant inherited cancer syndrome associating medullary thyroid carcinoma (MTC), pheochromocytoma (PHEO), ganglioneuromatosis of the aerodigestive tract and facial, ophthalmologic and skeletal abnormalities. MEN2B is caused by the M918T and A883F mutation of the REarranged during Transfection (RET) proto-oncogene in approximately 95% and <5% of cases, respectively. Only very few other mutations have been reported to cause MEN2B. In approximately 75% of MEN2B patients, mutations occur as de novo (Wells et al. 2015). The epidemiology of MEN2B is poorly defined. A nationwide study from Northern Ireland reported of three MEN2B patients and 1,824,000 inhabitants alive at April 21, 2012, yielding a point prevalence of 1.65 per million (Znaczko et al. 2014). However, it is unclear if this is representative of larger populations. A German study reported an MEN2B incidence (M918T carriers only) of 1.4 per million live births per year from 1991 to 2000 and estimated that at least half of all German RET carriers were captured (Machens et al. 2013). Meanwhile, the incidence of MEN2B and M918T carriers in a complete population is undisclosed. We conducted a nationwide study of the incidence and prevalence of MEN2B in Denmark from 1941 to 2014. This retrospective cohort study included 12 unique MEN2B patients identified through the following sources:

Neonatal High Bone Mass With First Mutation of the NF-κB Complex: Heterozygous De Novo Missense (p.Asp512Ser) RELA (Rela/p65)

Heritable disorders that feature high bone mass (HBM) are rare. The etiology is typically a mutation(s) within a gene that regulates the differentiation and function of osteoblasts (OBs) or osteoclasts (OCs). Nevertheless, the molecular basis is unknown for approximately one‐fifth of such entities. NF‐κB signaling is a key regulator of bone remodeling and acts by enhancing OC survival while impairing OB maturation and function. The NF‐κB transcription complex comprises five subunits. In mice, deletion of the p50 and p52 subunits together causes osteopetrosis (OPT). In humans, however, mutations within the genes that encode the NF‐κB complex, including the Rela/p65 subunit, have not been reported. We describe a neonate who died suddenly and unexpectedly and was found at postmortem to have HBM documented radiographically and by skeletal histopathology. Serum was not available for study. Radiographic changes resembled malignant OPT, but histopathological investigation showed morphologically normal OCs and evidence of intact bone resorption excluding OPT. Furthermore, mutation analysis was negative for eight genes associated with OPT or HBM. Instead, accelerated bone formation appeared to account for the HBM. Subsequently, trio‐based whole exome sequencing revealed a heterozygous de novo missense mutation (c.1534_1535delinsAG, p.Asp512Ser) in exon 11 of RELA encoding Rela/p65. The mutation was then verified using bidirectional Sanger sequencing. Lipopolysaccharide stimulation of patient fibroblasts elicited impaired NF‐κB responses compared with healthy control fibroblasts. Five unrelated patients with unexplained HBM did not show a RELA defect. Ours is apparently the first report of a mutation within the NF‐κB complex in humans. The missense change is associated with neonatal osteosclerosis from in utero increased OB function rather than failed OC action. These findings demonstrate the importance of the Rela/p65 subunit within the NF‐κB pathway for human skeletal homeostasis and represent a new genetic cause of HBM.

Neonatal High Bone Mass With First Mutation Of The NF-κB Complex

Heritable disorders that feature high bone mass (HBM) are rare. The etiology is typically a mutation(s) within a gene that regulates the differentiation and function of osteoblasts (OBs) or osteoclasts (OCs). Nevertheless, the molecular basis is unknown for approximately one‐fifth of such entities. NF‐κB signaling is a key regulator of bone remodeling and acts by enhancing OC survival while impairing OB maturation and function. The NF‐κB transcription complex comprises five subunits. In mice, deletion of the p50 and p52 subunits together causes osteopetrosis (OPT). In humans, however, mutations within the genes that encode the NF‐κB complex, including the Rela/p65 subunit, have not been reported. We describe a neonate who died suddenly and unexpectedly and was found at postmortem to have HBM documented radiographically and by skeletal histopathology. Serum was not available for study. Radiographic changes resembled malignant OPT, but histopathological investigation showed morphologically normal OCs and evidence of intact bone resorption excluding OPT. Furthermore, mutation analysis was negative for eight genes associated with OPT or HBM. Instead, accelerated bone formation appeared to account for the HBM. Subsequently, trio‐based whole exome sequencing revealed a heterozygous de novo missense mutation (c.1534_1535delinsAG, p.Asp512Ser) in exon 11 of RELA encoding Rela/p65. The mutation was then verified using bidirectional Sanger sequencing. Lipopolysaccharide stimulation of patient fibroblasts elicited impaired NF‐κB responses compared with healthy control fibroblasts. Five unrelated patients with unexplained HBM did not show a RELA defect. Ours is apparently the first report of a mutation within the NF‐κB complex in humans. The missense change is associated with neonatal osteosclerosis from in utero increased OB function rather than failed OC action. These findings demonstrate the importance of the Rela/p65 subunit within the NF‐κB pathway for human skeletal homeostasis and represent a new genetic cause of HBM.

Case report: vitamin D-dependent rickets type 1 caused by a novel CYP27B1 mutation.

Vitamin D‐dependent rickets type 1 VDDR‐1 is a recessive inherited disorder with impaired activation of vitamin D, caused by mutations in CYP27B1. We present long‐time follow‐up of a case with a novel mutation including high‐resolution peripheral quantitative computed tomography of the bone. Adequate treatment resulted in a normalized phenotype.

Novel Somatic RET Mutation Questioning the Causality of the RET I852M Germline Sequence Variant in Multiple Endocrine Neoplasia 2A

Author order, full names, highest earned academic degrees, complete affiliations and contact information: Jes Sloth Mathiesen Department of ORL Head & Neck Surgery, Odense University Hospital, Sdr. Boulevard 29, DK‐5000 Odense, Denmark 2 Institute of Clinical Research, University of Southern Denmark, Winsløwparken 19, DK‐ 5000 Odense, Denmark E‐mail address: jes_mathiesen@yahoo.dk Phone: +45 2252 3083

Bone structure in two adult subjects with impaired minor spliceosome function resulting from RNU4ATAC mutations causing microcephalic osteodysplastic primordial dwarfism type 1 (MOPD1)

Microcephalic osteodysplastic primordial dwarfism type 1 (MOPD1), or Taybi-Linder syndrome is characterized by distinctive skeletal dysplasia, severe intrauterine and postnatal growth retardation, microcephaly, dysmorphic features, and neurological malformations. It is an autosomal recessive disorder caused by homozygous or compound heterozygous mutations in the RNU4ATAC gene resulting in impaired function of the minor spliceosome. Here, we present the first report on bone morphology, bone density and bone microstructure in two adult MOPD1 patients and applied radiographs, dual energy X-ray absorptiometry, high-resolution peripheral quantitative computed tomography and biochemical evaluation. The MOPD1 patients presented with short stature, low BMI but normal macroscopic bone configuration. Bone mineral density was low. Compared to Danish reference data, total bone area, cortical bone area, cortical thickness, total bone density, cortical bone density, trabecular bone density and trabecular bone volume per tissue volume (BV/TV) were all low. These findings may correlate to the short stature and low body weight of the MOPD1 patients. Our findings suggest that minor spliceosome malfunction may be associated with altered bone modelling.