Background and purpose
The aim was to assess the value of insoluble PABPN1 muscle fibre nuclei accumulation in the diagnosis of atypical cases of oculopharyngeal muscular dystrophy (OPMD).
Methods
...Muscle biopsies from a selected cohort of 423 adult patients from several Italian neuromuscular centres were analysed by immunofluorescence: 30 muscle biopsies of genetically proven OPMD, 30 biopsies from patients not affected by neuromuscular disorders, 220 from genetically undiagnosed patients presenting ptosis or swallowing disturbances, progressive lower proximal weakness and/or isolated rimmed vacuoles at muscle biopsy and 143 muscle biopsies of patients affected by other neuromuscular diseases.
Results
The detection of insoluble nuclear PABPN1 accumulation is rapid, sensitive (100%) and specific (96%). The revision of our cohort allowed us to discover 23 new OPMD cases out of 220 patients affected with nonspecific muscle diseases.
Conclusions
Oculopharyngeal muscular dystrophy is often misdiagnosed leading to diagnosis delay, causing waste of time and resources. A great number of these cases present symptoms and histological findings frequently overlapping with other muscle diseases, i.e. inclusion body myositis and progressive external ophthalmoplegia. PABPN1 nuclear accumulation is a reliable method for diagnostic purposes and it is safe and useful in helping pathologists and clinicians to direct genetic analysis in the case of suspected OPMD, even when clinical and histological clues are deceptive.
Inherited ataxias are a group of heterogeneous disorders in children or adults but their genetic definition remains still undetermined in almost half of the patients. However, CoQ10 deficiency is a ...rare cause of cerebellar ataxia and ADCK3 is the most frequent gene associated with this defect. We herein report a 48 year old man, who presented with dysarthria and walking difficulties. Brain magnetic resonance imaging showed a marked cerebellar atrophy. Serum lactate was elevated. Tissues obtained by muscle and skin biopsies were studied for biochemical and genetic characterization. Skeletal muscle biochemistry revealed decreased activities of complexes I+III and II+III and a severe reduction of CoQ10, while skin fibroblasts showed normal CoQ10 levels. A mild loss of maximal respiration capacity was also found by high‐resolution respirometry. Molecular studies identified a novel homozygous deletion (c.504del_CT) in ADCK3, causing a premature stop codon. Western blot analysis revealed marked reduction of ADCK3 protein levels. Treatment with CoQ10 was started and, after 1 year follow‐up, patient neurological condition slightly improved. This report suggests the importance of investigating mitochondrial function and, in particular, muscle CoQ10 levels, in patients with adult‐onset cerebellar ataxia. Moreover, clinical stabilization by CoQ10 supplementation emphasizes the importance of an early diagnosis.
Inherited ataxias are a group of heterogeneous disorders in children or adults but their genetic definition remains still undetermined in almost half of the patients. However,
CoQ
10
deficiency is a ...rare cause of cerebellar ataxia and
ADCK3
is the most frequent gene associated with this defect. We herein report a 48 year old man, who presented with dysarthria and walking difficulties. Brain magnetic resonance imaging showed a marked cerebellar atrophy. Serum lactate was elevated. Tissues obtained by muscle and skin biopsies were studied for biochemical and genetic characterization. Skeletal muscle biochemistry revealed decreased activities of complexes I+
III
and
II
+
III
and a severe reduction of
CoQ
10
, while skin fibroblasts showed normal
CoQ
10
levels. A mild loss of maximal respiration capacity was also found by high‐resolution respirometry. Molecular studies identified a novel homozygous deletion (c.504del_
CT
) in
ADCK3
, causing a premature stop codon. Western blot analysis revealed marked reduction of
ADCK3
protein levels. Treatment with
CoQ
10
was started and, after 1 year follow‐up, patient neurological condition slightly improved. This report suggests the importance of investigating mitochondrial function and, in particular, muscle
CoQ
10
levels, in patients with adult‐onset cerebellar ataxia. Moreover, clinical stabilization by
CoQ
10
supplementation emphasizes the importance of an early diagnosis.
Inherited ataxias are a group of heterogeneous disorders in children or adults but their genetic definition remains still undetermined in almost half of the patients. However, CoQ sub(10) deficiency ...is a rare cause of cerebellar ataxia and ADCK3 is the most frequent gene associated with this defect. We herein report a 48 year old man, who presented with dysarthria and walking difficulties. Brain magnetic resonance imaging showed a marked cerebellar atrophy. Serum lactate was elevated. Tissues obtained by muscle and skin biopsies were studied for biochemical and genetic characterization. Skeletal muscle biochemistry revealed decreased activities of complexes I+III and II+III and a severe reduction of CoQ sub(10), while skin fibroblasts showed normal CoQ sub(10) levels. A mild loss of maximal respiration capacity was also found by high-resolution respirometry. Molecular studies identified a novel homozygous deletion (c.504del_CT) in ADCK3 , causing a premature stop codon. Western blot analysis revealed marked reduction of ADCK3 protein levels. Treatment with CoQ sub(10) was started and, after 1year follow-up, patient neurological condition slightly improved. This report suggests the importance of investigating mitochondrial function and, in particular, muscle CoQ sub(10) levels, in patients with adult-onset cerebellar ataxia. Moreover, clinical stabilization by CoQ sub(10) supplementation emphasizes the importance of an early diagnosis.
ContextAlterations in the cAMP signaling pathway are common in hormonally active endocrine tumors. Somatic mutations at GNAS are causative in 30–40% of GH-secreting adenomas. Recently, mutations ...affecting the USP8 and PRKACA gene have been reported in ACTH-secreting pituitary adenomas and cortisol-secreting adrenocortical adenomas respectively. However, the pathogenesis of many GH-secreting adenomas remains unclear.AimComprehensive genetic characterization of sporadic GH-secreting adenomas and identification of new driver mutations.DesignScreening for somatic mutations was performed in 67 GH-secreting adenomas by targeted sequencing for GNAS, PRKACA, and USP8 mutations (n=31) and next-generation exome sequencing (n=36).ResultsBy targeted sequencing, known activating mutations in GNAS were detected in five cases (16.1%), while no somatic mutations were observed in both PRKACA and USP8. Whole-exome sequencing identified 132 protein-altering somatic mutations in 31/36 tumors with a median of three mutations per sample (range: 1–13). The only recurrent mutations have been observed in GNAS (31.4% of cases). However, seven genes involved in cAMP signaling pathway were affected in 14 of 36 samples and eight samples harbored variants in genes involved in the calcium signaling or metabolism. At the enrichment analysis, several altered genes resulted to be associated with developmental processes. No significant correlation between genetic alterations and the clinical data was observed.ConclusionThis study provides a comprehensive analysis of somatic mutations in a large series of GH-secreting adenomas. No novel recurrent genetic alterations have been observed, but the data suggest that beside cAMP pathway, calcium signaling might be involved in the pathogenesis of these tumors.
The role of progenitor/stem cells in pituitary tumorigenesis, resistance to pharmacological treatments and tumor recurrence is still unclear. This study investigated the presence of progenitor/stem ...cells in non‐functioning pituitary tumors (NFPTs) and tested the efficacy of dopamine receptor type 2 (DRD2) and somatostatin receptor type 2 (SSTR2) agonists to inhibit in vitro proliferation. They found that 70% of 46 NFPTs formed spheres co‐expressing stem cell markers, transcription factors (DAX1, SF1, ERG1) and gonadotropins. Analysis of tumor behavior showed that spheres formation was associated with tumor invasiveness (OR = 3,96; IC: 1.05–14.88, p = 0.036). The in vitro reduction of cell proliferation by DRD2 and SSTR2 agonists (31 ± 17% and 35 ± 13% inhibition, respectively, p < 0.01 vs. basal) occurring in about a half of NFPTs cells was conserved in the corresponding spheres. Accordingly, these drugs increased cyclin‐dependent kinase inhibitor p27 and decreased cyclin D3 expression in spheres. In conclusion, they provided further evidence for the existence of cells with a progenitor/stem cells‐like phenotype in the majority of NFPTs, particularly in those with invasive behavior, and demonstrated that the antiproliferative effects of dopaminergic and somatostatinergic drugs were maintained in progenitor/stem‐like cells.
What's new?
One third of pituitary tumors are non‐functioning (NFPTs) producing, but not secreting, gonadotropins, or gonadotropin hormone subunits. Here the authors isolated from the majority of NFPTs, particularly those with invasive behavior, progenitor/stem‐like cells co‐expressing stem cell markers, embryonic pituitary transcription factors and gonadotropins. These cells reproduced anti‐proliferative responses to dopamine and somatostatin analogues observed with bulk tumor cells, underscoring their relevance in the adjuvant medical therapy of NFPTs.
Adrenocortical carcinomas (ACCs) overexpress insulin-like growth factor 2 (IGF2), that drives a proliferative autocrine loop by binding to IGF1R and IR, but IGF1R/IR-targeted therapies failed in ACC ...patients.
The cytoskeleton actin-binding protein filamin A (FLNA) impairs IR signalling in melanoma cells.
Aims of this study were to test FLNA involvement in regulating IGF1R and IR responsiveness to both IGF2 and inhibitors in ACC.
In ACC cells H295R and SW13 and primary cultures (1ACC, 4 adenomas) we found that IGF1R and IR interacted with FLNA, and FLNA silencing increased IGF1R and reduced IR expression, with a downstream effect of increased cell proliferation and ERK phosphorylation.
In addition, FLNA knockdown potentiated antiproliferative effects of IGF1R/IR inhibitor Linsitinib and IGF1R inhibitor NVP-ADW742 in H295R.
Finally, Western blot showed lower FLNA expression in ACCs (n = 10) than in ACAs (n = 10) and an inverse correlation of FLNA/IGF1R ratio with ERK phosphorylation in ACCs only.
In conclusion, we demonstrated that low FLNA levels enhance both IGF2 proliferative effects and IGF1R/IR inhibitors efficacy in ACC cells, suggesting FLNA as a new factor influencing tumor clinical behavior and the response to the therapy with IGF1R/IR-targeted drugs.
•In ACC cell lines FLNA interacts with IGF1R and IR.•FLNA silencing increases IGF1R and decreases IR expression.•Low levels of FLNA enhance IGF2 proliferative effects in ACC cells.•FLNA knockdown potentiates antiproliferative effects of IGF1R inhibitors in H295R.•The loss of FLNA might be a biomarker for Linsitinib responsiveness.
The insulin-like growth factor 2 (IGF2) promotes cell growth by overactivating the IGF system in an autocrine loop in adrenocortical carcinomas (ACCs). The cytoskeleton protein filamin A (FLNA) acts ...as a repressor of IGF2 mitogenic signalling in ACC cells. The aims of this study were to test FLNA expression by immunohistochemistry in 119 ACCs and 26 adrenocortical adenomas (ACAs) and to evaluate its relationship with clinicopathological features and outcome in ACCs. We found that 71.4% of ACCs did not express FLNA, whereas FLNA absence was a rare event in ACAs (15.4%,
< 0.001 vs. ACCs). In addition, the expression of FLNA was associated with a less aggressive tumour behaviour in ACCs. Indeed, the subgroup of ACCs with high FLNA showed a lower ENSAT stage, Weiss score, and S-GRAS score compared to ACCs with low FLNA expression (
< 0.05). Moreover, patients with high FLNA had a longer overall survival than those with low FLNA (
< 0.05). In conclusion, our data suggest that FLNA may represent a "protective" factor in ACCs, and the integration of FLNA immunohistochemical expression in ACC tissues along with other clinical and molecular markers could be helpful to improve diagnostic accuracy and prognosis prediction in ACCs.
Congenital myopathies are a vast group of genetic muscle diseases. Among the causes are mutations in the
gene resulting in truncated type IIa myosin heavy chains (MyHCs). The precise cellular and ...molecular mechanisms by which these mutations induce skeletal muscle symptoms remain obscure. Hence, in the present study, we aimed to explore whether such genetic defects would alter the presence as well as the post-translational modifications of MyHCs and the functionality of myosin molecules. For this, we dissected muscle fibers from four myopathic patients with
truncating mutations and from five human healthy controls. We then assessed
) MyHCs presence/post-translational modifications using LC/MS;
) relaxed myosin conformation and concomitant ATP consumption with a loaded Mant-ATP chase setup;
) myosin activation with an unloaded in vitro motility assay; and
) cellular force production with a myofiber mechanical setup. Interestingly, the type IIa MyHC with one additional acetylated lysine (Lys35-Ac) was present in the patients. This was accompanied by
) a higher ATP demand of myosin heads in the disordered-relaxed conformation;
) faster actomyosin kinetics; and
) reduced muscle fiber force. Overall, our findings indicate that
truncating mutations impact myosin presence/functionality in human adult mature myofibers by disrupting the ATPase activity and actomyosin complex. These are likely important molecular pathological disturbances leading to the myopathic phenotype in patients.
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