PTRH2 is Necessary for Purkinje Cell Differentiation and Survival and its Loss Recapitulates Progressive Cerebellar Atrophy and Ataxia Seen in IMNEPD Patients

Hom ozygous variants in the peptidyl-tRNA hydrolase 2 gene (PTRH2) cause infantile-onset multisystem neurologic, endocrine, and pancreatic disease. The objective is to delineate the mechanisms underlying the core cerebellar phenotype in this disease. For this, we generated constitutive (Ptrh2LoxPxhCMVCre, Ptrh2−/− mice) and Purkinje cell (PC) specific (Ptrh2LoxPxPcp2Cre, Ptrh2ΔPCmice) Ptrh2 mutant mouse models and investigated the effect of the loss of Ptrh2 on cerebellar development. We show that Ptrh2−/− knockout mice had severe postnatal runting and lethality by postnatal day 14. Ptrh2ΔPC PC specific knockout mice survived until adult age; however, they showed progressive cerebellar atrophy and functional cerebellar deficits with abnormal gait and ataxia. PCs of Ptrh2ΔPC mice had reduced cell size and density, stunted dendrites, and lower levels of ribosomal protein S6, a readout of the mammalian target of rapamycin pathway. By adulthood, there was a marked loss of PCs. Thus, we identify a cell autonomous requirement for PTRH2 in PC maturation and survival. Loss of PTRH2 in PCs leads to downregulation of the mTOR pathway and PC atrophy. This suggests a molecular mechanism underlying the ataxia and cerebellar atrophy seen in patients with PTRH2 mutations leading to infantile-onset multisystem neurologic, endocrine, and pancreatic disease

Redefining the MED13L syndrome

Congenital cardiac and neurodevelopmental deficits have been recently linked to the mediator complex subunit 13-like protein MED13L, a subunit of the CDK8-associated mediator complex that functions in transcriptional regulation through DNA-binding transcription factors and RNA polymerase II. Heterozygous MED13L variants cause transposition of the great arteries and intellectual disability (ID). Here, we report eight patients with predominantly novel MED13L variants who lack such complex congenital heart malformations. Rather, they depict a syndromic form of ID characterized by facial dysmorphism, ID, speech impairment, motor developmental delay with muscular hypotonia and behavioral difficulties. We thereby define a novel syndrome and significantly broaden the clinical spectrum associated with MED13L variants. A prominent feature of the MED13L neurocognitive presentation is profound language impairment, often in combination with articulatory deficits

Effects of Simvastatin and Daptomycin alone and in combination on L6 myocytes

Statine inhibieren die Cholesterolbiosynthese und sind die Medikamentengruppe der ersten Wahl für die Behandlung einer Hypercholesterinämie. Fallberichte suggerieren eine verstärkte Muskelschädigung durch eine Kombination von Statinen mit dem Antibiotikum Daptomycin, welches bei Infektionen durch grampositive Erreger, einschließlich MRSA und VRE, eingesetzt wird. Bislang existieren kaum Daten bezüglich einer gemeinsamen Wirkung von Statinen und Daptomycin auf das Muskelgewebe. In der vorliegenden In-vitro-Arbeit wurde daher modellhaft die alleinige und kombinierte Wirkung von Simvastatin und Daptomycin auf die Zellmorphologie, die Expression intrazellulärer Proteine und die Apoptose/Nekrose von murinen L6-Myozyten und auf die Proliferation von L6-Myoblasten analysiert. Lichtmikroskopische Untersuchungen der L6-Myozyten mit Bisbenzimid H33258 und Propidiumiodid zeigten nach 72-stündiger Exposition gegenüber 10 µM Simvastatin (ca. 4,2 mg/l Medium) und/oder 300 mg Daptomycin/l Medium einen Verlust der Zelladhärenz und eine Karyo und Zytolyse. Die Ergebnisse wiesen auf einen toxischen Effekt von Simvastatin und Daptomycin allein und auf eine verstärkte Zytotoxizität durch die Kombination beider Substanzen hin. Die zellschädigende Wirkung von Simvastatin und Daptomycin wurde durchflusszytometrisch mit Annexin V-FITC und Propidiumiodid-Färbung und im MTT-Zytotoxizitätstest bestätigt und konnte zusammenfassend als zeit- und konzentrationsabhängig charakterisiert werden. Eine Zytotoxizität wurde nach zehntägiger Exposition für therapeutische Konzentrationen beider Substanzen gezeigt. Ein additiver bis synergistischer Effekt mit Simvastatin bestand erst in einer supratherapeutischen Dosierung von 300 mg Daptomycin/l Medium. Ein Anstieg der Caspase 3 in L6-Myozyten wurde immunhistochemisch nach 24 stündiger Exposition mit Simvastatin und/oder Daptomycin in keiner Versuchsreihe nachgewiesen. Simvastatin führte aber ab einer Konzentration von 1 µM nach 24 stündiger Exposition zu einer reduzierten Expression des kleinen G-Proteins Rho und des Zytoskelettproteins Vimentin in L6-Myozyten. Dies suggeriert übereinstimmend mit In-vitro-Arbeiten weiterer Autoren eine Dysprenylierung und eine Veränderung des Zytoskeletts als pathophysiologische Elemente der Muskelschädigung. Daptomycin allein und in Kombination mit Simvastatin hatte keine (zusätzliche) Wirkung. Dies könnte auf einen alternativen Mechanismus der Muskelschädigung durch Daptomycin hinweisen. Für L6-Myoblasten wurde eine Proliferationshemmung durch 10 µM Simvastatin durchflusszytometrisch mit CFSE-Färbung identifiziert. Daptomycin in einer maximalen Konzentration von 300 mg/l Medium zeigte keine antiproliferative Wirkung und keinen adjuvanten Effekt in Kombination mit Simvastatin. Zusammenfassend wurde in dieser In-vitro-Arbeit ein zytotoxischer Effekt für Simvastatin und Daptomycin allein und eine additive bis synergistische Zytotoxizität für die Kombination beider Substanzen identifiziert. Letztere bestand für Kombinationen mit Daptomycin in supratherapeutischen Dosierungen. Die Arbeit lieferte Hinweise darauf, dass der Zellschädigung jeweils unterschiedliche Mechanismen zugrundeliegen. Ein weitergehendes Verständnis der pathophysiologischen Zusammenhänge der Muskelschädigung könnte dazu beitragen, das Risikoprofil besser zu charakterisieren und Ansatzpunkte für eine ursächliche Therapie zu identifizieren.Statins inhibit the endogenous cholesterol synthesis and are the first-line treatment for hypercholesterolaemia. Case reports suggest an enhanced myotoxicity of statins in combination with daptomycin, an antibiotic used for the treatment of gram-positive bacteria including MRSA and VRE. Data regarding the combined effects of statins and daptomycin on muscle cells are rare. This in vitro study examined the sole and common effect of simvastatin and daptomycin on cell morphology, intracellular protein expression, apoptosis/necrosis and proliferation of murine L6 myoblasts or myocytes respectively. Microscopic analysis with bisbenzimide H33258 and propidiumiodide after 72 h of exposure with 10 µM simvastatin (= 4.2 mg/l) and/or 300 mg daptomycin/l medium revealed loss of cell adherence and karyo- and cytolysis of L6 myocytes. The results indicated a cytotoxicity of simvastatin and daptomycin alone, which was augmented by combination of both substances. Flow cytometry analysis with Annexin V-FITC and propidiumiodide and MTT assay confirmed a concentration- and time-dependant cytotoxicity of both agents. After 10 days of exposure therapeutic concentrations of each substance elicited a cytopathic effect. An additive or mildly synergistic effect in combination with simvastatin was identified at supratherapeutic doses of 300 mg daptomycin/l medium. Caspase 3 elevation was not detected with immunohistochemical staining after 24 h of incubation with simvastatin and/or daptomycin. However, simvastatin at a minimal concentration of 1 µM reduced the expression of the small g-protein Rho and the cytoskeletal protein vimentin. Consistent with other in vitro studies, these results suggest dysprenylation and cytoskeletal disruption as pathogenetic elements of simvastatin muscletoxicity. Daptomycin alone or combined with simvasatin showed no (additional) effect, implying that daptomycin may elicit a different pathomechanism. In L6 myoblasts, simvastatin at a concentration of 10 µM impaired proliferation after 24 h of exposure. Daptomycin alone did not exert an antiproliferative effect and had no additional influence in combination with simvastatin. In summary, a cytotoxic effect of simvastatin and daptomycin alone and an additive or mildly synergistic cytotoxicity by combination of both substances were identified. Augmented toxicity was demonstrated for combinations of simvastatin at therapeutic concentrations and daptomycin at dosages above the therapeutic range. This study implicated different underlying pathomechanisms of myopathy for simvastatin and daptomycin. Further understanding of the pathophysiologic mechanisms may improve risk evaluation and help to establish a therapeutic approach for the treatment of medication- induced myopathy

Recessive mutation in EXOSC3 associates with mitochondrial dysfunction and pontocerebellar hypoplasia

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PTRH2 is Necessary for Purkinje Cell Differentiation and Survival and its Loss Recapitulates Progressive Cerebellar Atrophy and Ataxia Seen in IMNEPD Patients

Homozygous variants in the peptidyl-tRNA hydrolase 2 gene (PTRH2) cause infantile-onset multisystem neurologic, endocrine, and pancreatic disease. The objective is to delineate the mechanisms underlying the core cerebellar phenotype in this disease. For this, we generated constitutive (Ptrh2LoxPxhCMVCre, Ptrh2−/− mice) and Purkinje cell (PC) specific (Ptrh2LoxPxPcp2Cre, Ptrh2ΔPCmice) Ptrh2 mutant mouse models and investigated the effect of the loss of Ptrh2 on cerebellar development. We show that Ptrh2−/− knockout mice had severe postnatal runting and lethality by postnatal day 14. Ptrh2ΔPC PC specific knockout mice survived until adult age; however, they showed progressive cerebellar atrophy and functional cerebellar deficits with abnormal gait and ataxia. PCs of Ptrh2ΔPC mice had reduced cell size and density, stunted dendrites, and lower levels of ribosomal protein S6, a readout of the mammalian target of rapamycin pathway. By adulthood, there was a marked loss of PCs. Thus, we identify a cell autonomous requirement for PTRH2 in PC maturation and survival. Loss of PTRH2 in PCs leads to downregulation of the mTOR pathway and PC atrophy. This suggests a molecular mechanism underlying the ataxia and cerebellar atrophy seen in patients with PTRH2 mutations leading to infantile-onset multisystem neurologic, endocrine, and pancreatic disease.Open Access funding enabled and organized by Projekt DEAL. Our research was financially supported by the German Research Foundation (SFB1315, FOR3004) and the Charité

Pontine Tegmental Cap Dysplasia in an Extremely Preterm Infant and Review of the Literature

Pontine tegmental cap dysplasia is a rare hindbrain malformation syndrome with a hypoplastic pons, a tissue protrusion into the fourth ventricle, and cranial nerve dysfunction. We here report clinical, imaging, and genetic findings of the first extremely low-birth-weight preterm infant with pontine tegmental cap dysplasia born at 25 weeks of gestation and provide an overview of 29 sporadic cases. A prenatally diagnosed hypoplastic and rostrally shifted cerebellum was indicative of a hindbrain defect and later identified as an early sign of pontine tegmental cap dysplasia in our patient. The neonate exhibited severe muscle hypotonia, persistent thermolability, and clinical signs of an involvement of facial, cochlear, and hypoglossal nerves. Furthermore, paroxysmal episodes of agonizing pain with facial tics, tonic and clonic muscle contractions, blepharospasm, and singultus are highlighted as new phenotypic features of pontine tegmental cap dysplasia. With our report, we present a severe case of pontine tegmental cap dysplasia and provide a brief overview of current knowledge on this rare disease

Additional file 2: Figure S1. of Phenotype variability of infantile-onset multisystem neurologic, endocrine, and pancreatic disease IMNEPD

PTRH2 protein and mRNA levels in IMNEPD patients with a PTRH2 missense mutation c.254A > C (p.Q85P). (TIF 26 kb

Expanding the phenotype of NUP85 mutations beyond nephrotic syndrome to primary autosomal recessive microcephaly and Seckel syndrome spectrum disorders

Primary autosomal recessive microcephaly and Seckel syndrome spectrum disorders (MCPH-SCKS) include a heterogeneous group of autosomal recessive inherited diseases characterized by primary (congenital) microcephaly, the absence of visceral abnormalities, and a variable degree of cognitive impairment, short stature and facial dysmorphism. Recently, biallelic variants in the nuclear pore complex (NPC) component nucleoporin 85 gene (NUP85) were reported to cause steroid-resistant nephrotic syndrome (SRNS). Here, we report biallelic variants in NUP85 in two pedigrees with an MCPH-SCKS phenotype spectrum without SRNS, thereby expanding the phenotypic spectrum of NUP85-linked diseases. Structural analysis predicts the identified NUP85 variants cause conformational changes that could have an effect on NPC architecture or on its interaction with other NUPs. We show that mutant NUP85 is, however, associated with a reduced number of NPCs but unaltered nucleocytoplasmic compartmentalization, abnormal mitotic spindle morphology, and decreased cell viability and proliferation in one patient's cells. Our results also indicate the link of common cellular mechanisms involved in MCPH-SCKS spectrum disorders and NUP85-associated diseases. In addition to the previous studies, our results broaden the phenotypic spectrum of NUP85-linked human disease and propose a role for NUP85 in nervous system development.SCOPUS: ar.jDecretOANoAutActifinfo:eu-repo/semantics/publishe

Redefining the MED13L syndrome.

Congenital cardiac and neurodevelopmental deficits have been recently linked to the mediator complex subunit 13-like protein MED13L, a subunit of the CDK8-associated mediator complex that functions in transcriptional regulation through DNA-binding transcription factors and RNA polymerase II. Heterozygous MED13L variants cause transposition of the great arteries and intellectual disability (ID). Here, we report eight patients with predominantly novel MED13L variants who lack such complex congenital heart malformations. Rather, they depict a syndromic form of ID characterized by facial dysmorphism, ID, speech impairment, motor developmental delay with muscular hypotonia and behavioral difficulties. We thereby define a novel syndrome and significantly broaden the clinical spectrum associated with MED13L variants. A prominent feature of the MED13L neurocognitive presentation is profound language impairment, often in combination with articulatory deficits