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1.
Hum Mutat ; 43(4): 477-486, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35112411

RESUMO

The synthesis of cytochrome c oxidase 2 (SCO2 ) gene encodes for a mitochondrial located metallochaperone essential for the synthesis of the cytochrome c oxidase (COX) subunit 2. Recessive mutations in SCO2 have been reported in several cases with fatal infantile cardioencephalomyopathy with COX deficiency and in only four cases with axonal neuropathy. Here, we identified a homozygous pathogenic variant (c.361G > C; p.[Gly121Arg]) in SCO2 in two brothers with isolated axonal motor neuropathy. To address pathogenicity of the amino acid substitution, biochemical studies were performed and revealed increased level of the mutant SCO2 -protein and dysregulation of COX subunits in leukocytes and moreover unraveled decrease of proteins involved in the manifestation of neuropathies. Hence, our combined data strengthen the concept of SCO2 being causative for a very rare form of axonal neuropathy, expand its molecular genetic spectrum and provide first biochemical insights into the underlying pathophysiology.


Assuntos
Doença de Charcot-Marie-Tooth , Proteínas de Transporte/genética , Doença de Charcot-Marie-Tooth/genética , Doença de Charcot-Marie-Tooth/patologia , Complexo IV da Cadeia de Transporte de Elétrons/genética , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Humanos , Masculino , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Chaperonas Moleculares/genética , Mutação , Oxirredutases/genética , Oxirredutases/metabolismo , Irmãos
2.
Neuropathol Appl Neurobiol ; 47(6): 840-855, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-33428302

RESUMO

AIMS: MICU1 encodes the gatekeeper of the mitochondrial Ca2+ uniporter, MICU1 and biallelic loss-of-function mutations cause a complex, neuromuscular disorder in children. Although the role of the protein is well understood, the precise molecular pathophysiology leading to this neuropaediatric phenotype has not been fully elucidated. Here we aimed to obtain novel insights into MICU1 pathophysiology. METHODS: Molecular genetic studies along with proteomic profiling, electron-, light- and Coherent anti-Stokes Raman scattering microscopy and immuno-based studies of protein abundances and Ca2+ transport studies were employed to examine the pathophysiology of MICU1 deficiency in humans. RESULTS: We describe two patients carrying MICU1 mutations, two nonsense (c.52C>T; p.(Arg18*) and c.553C>T; p.(Arg185*)) and an intragenic exon 2-deletion presenting with ataxia, developmental delay and early onset myopathy, clinodactyly, attention deficits, insomnia and impaired cognitive pain perception. Muscle biopsies revealed signs of dystrophy and neurogenic atrophy, severe mitochondrial perturbations, altered Golgi structure, vacuoles and altered lipid homeostasis. Comparative mitochondrial Ca2+ transport and proteomic studies on lymphoblastoid cells revealed that the [Ca2+ ] threshold and the cooperative activation of mitochondrial Ca2+ uptake were lost in MICU1-deficient cells and that 39 proteins were altered in abundance. Several of those proteins are linked to mitochondrial dysfunction and/or perturbed Ca2+ homeostasis, also impacting on regular cytoskeleton (affecting Spectrin) and Golgi architecture, as well as cellular survival mechanisms. CONCLUSIONS: Our findings (i) link dysregulation of mitochondrial Ca2+ uptake with muscle pathology (including perturbed lipid homeostasis and ER-Golgi morphology), (ii) support the concept of a functional interplay of ER-Golgi and mitochondria in lipid homeostasis and (iii) reveal the vulnerability of the cellular proteome as part of the MICU1-related pathophysiology.


Assuntos
Proteínas de Ligação ao Cálcio/deficiência , Cálcio/metabolismo , Proteínas de Transporte de Cátions/deficiência , Proteínas de Transporte da Membrana Mitocondrial/genética , Doenças Musculares/genética , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Transporte de Cátions/metabolismo , Humanos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/deficiência , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Doenças Musculares/patologia , Proteômica
3.
J Inherit Metab Dis ; 43(2): 297-308, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31339582

RESUMO

Transport And Golgi Organization protein 2 (TANGO2) deficiency has recently been identified as a rare metabolic disorder with a distinct clinical and biochemical phenotype of recurrent metabolic crises, hypoglycemia, lactic acidosis, rhabdomyolysis, arrhythmias, and encephalopathy with cognitive decline. We report nine subjects from seven independent families, and we studied muscle histology, respiratory chain enzyme activities in skeletal muscle and proteomic signature of fibroblasts. All nine subjects carried autosomal recessive TANGO2 mutations. Two carried the reported deletion of exons 3 to 9, one homozygous, one heterozygous with a 22q11.21 microdeletion inherited in trans. The other subjects carried three novel homozygous (c.262C>T/p.Arg88*; c.220A>C/p.Thr74Pro; c.380+1G>A), and two further novel heterozygous (c.6_9del/p.Phe6del); c.11-13delTCT/p.Phe5del mutations. Immunoblot analysis detected a significant decrease of TANGO2 protein. Muscle histology showed mild variation of fiber diameter, no ragged-red/cytochrome c oxidase-negative fibers and a defect of multiple respiratory chain enzymes and coenzyme Q10 (CoQ10 ) in two cases, suggesting a possible secondary defect of oxidative phosphorylation. Proteomic analysis in fibroblasts revealed significant changes in components of the mitochondrial fatty acid oxidation, plasma membrane, endoplasmic reticulum-Golgi network and secretory pathways. Clinical presentation of TANGO2 mutations is homogeneous and clinically recognizable. The hemizygous mutations in two patients suggest that some mutations leading to allele loss are difficult to detect. A combined defect of the respiratory chain enzymes and CoQ10 with altered levels of several membrane proteins provides molecular insights into the underlying pathophysiology and may guide rational new therapeutic interventions.


Assuntos
Encefalopatias Metabólicas/genética , Doenças Mitocondriais/genética , Debilidade Muscular/genética , Mutação , Proteômica/métodos , Rabdomiólise/genética , Encefalopatias Metabólicas/diagnóstico , Ácidos Graxos/metabolismo , Feminino , Complexo de Golgi/genética , Complexo de Golgi/metabolismo , Homozigoto , Humanos , Lactente , Masculino , Doenças Mitocondriais/diagnóstico , Fosforilação Oxidativa , Fenótipo , Rabdomiólise/diagnóstico , Sequenciamento Completo do Genoma
4.
Int J Mol Sci ; 20(21)2019 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-31684150

RESUMO

Transforming growth factor beta 3 (TGFß3) promotes tenogenic differentiation and may enhance tendon regeneration in vivo. This study aimed to apply TGFß3 absorbed in decellularized equine superficial digital flexor tendon scaffolds, and to investigate the bioactivity of scaffold-associated TGFß3 in an in vitro model. TGFß3 could effectively be loaded onto tendon scaffolds so that at least 88% of the applied TGFß3 were not detected in the rinsing fluid of the TGFß3-loaded scaffolds. Equine adipose tissue-derived multipotent mesenchymal stromal cells (MSC) were then seeded on scaffolds loaded with 300 ng TGFß3 to assess its bioactivity. Both scaffold-associated TGFß3 and TGFß3 dissolved in the cell culture medium, the latter serving as control group, promoted elongation of cell shapes and scaffold contraction (p < 0.05). Furthermore, scaffold-associated and dissolved TGFß3 affected MSC musculoskeletal gene expression in a similar manner, with an upregulation of tenascin c and downregulation of other matrix molecules, most markedly decorin (p < 0.05). These results demonstrate that the bioactivity of scaffold-associated TGFß3 is preserved, thus TGFß3 application via absorption in decellularized tendon scaffolds is a feasible approach.


Assuntos
Matriz Extracelular/metabolismo , Células-Tronco Mesenquimais/citologia , Tendões/fisiologia , Engenharia Tecidual/métodos , Alicerces Teciduais , Fator de Crescimento Transformador beta3/metabolismo , Animais , Diferenciação Celular , Células Cultivadas , Decorina/genética , Decorina/metabolismo , Regulação da Expressão Gênica , Cavalos , Humanos , Células-Tronco Mesenquimais/metabolismo , Sistema Musculoesquelético/metabolismo , Tenascina/genética , Tenascina/metabolismo , Tendões/citologia
5.
Mod Pathol ; 31(5): 829-839, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29327707

RESUMO

The appendix gives rise to goblet cell carcinoids, which represent special carcinomas with distinct biological and histological features. Their genetic background and molecular relationship to colorectal adenocarcinoma is largely unknown. We therefore performed a next-generation sequencing analysis of 25 appendiceal carcinomas including 11 goblet cell carcinoids, 7 adenocarcinomas ex-goblet cell carcinoid, and 7 primary colorectal-type adenocarcinomas, using a modified Colorectal Cancer specific Panel comprising 32 genes linked to colorectal and neuroendocrine tumorigenesis. The mutational profiles of these neoplasms were compared with those of conventional adenocarcinomas, mixed adenoneuroendocrine carcinomas, and neuroendocrine carcinomas of the colorectum. In addition, a large-scale pan-cancer sequencing panel covering 409 genes was applied to selected cases of goblet cell carcinoid/adenocarcinoma ex-goblet cell carcinoid (n=2, respectively). Mutations in colorectal cancer-related genes (eg, TP53, KRAS, APC) were rare to absent in both, goblet cell carcinoids and adenocarcinomas ex-goblet cell carcinoid, but frequent in primary colorectal-type adenocarcinomas of the appendix. Additional large-scale sequencing of selected goblet cell carcinoids and adenocarcinomas ex-goblet cell carcinoid revealed mutations in Wnt-signaling-associated genes (USP9X, NOTCH1, CTNNA1, CTNNB1, TRRAP). These data suggest that appendiceal goblet cell carcinoids and adenocarcinomas ex-goblet cell carcinoid constitute a morphomolecular entity, histologically and genetically distinct from appendiceal colorectal-type adenocarcinomas and its colorectal counterparts. Altered Wnt-signaling associated genes, apart from APC, may act as potential drivers of these neoplasms. The absence of KRAS/NRAS mutations might render some of these tumors eligible for anti-EGFR directed therapy regimens.


Assuntos
Neoplasias do Apêndice/genética , Tumor Carcinoide/genética , Neoplasias Colorretais/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias do Apêndice/metabolismo , Neoplasias do Apêndice/patologia , Tumor Carcinoide/metabolismo , Tumor Carcinoide/patologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Feminino , Perfilação da Expressão Gênica , Células Caliciformes/metabolismo , Células Caliciformes/patologia , Humanos , Masculino , Instabilidade de Microssatélites , Pessoa de Meia-Idade , Mutação , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas Wnt/genética , Adulto Jovem
6.
Int J Mol Sci ; 19(9)2018 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-30154348

RESUMO

Transplantation of multipotent mesenchymal progenitor cells is a valuable option for treating tendon disease. Tenogenic differentiation leading to cell replacement and subsequent matrix modulation may contribute to the regenerative effects of these cells, but it is unclear whether this occurs in the inflammatory environment of acute tendon disease. Equine adipose-derived stromal cells (ASC) were cultured as monolayers or on decellularized tendon scaffolds in static or dynamic conditions, the latter represented by cyclic stretching. The impact of different inflammatory conditions, as represented by supplementation with interleukin-1ß and/or tumor necrosis factor-α or by co-culture with allogeneic peripheral blood leukocytes, on ASC functional properties was investigated. High cytokine concentrations increased ASC proliferation and osteogenic differentiation, but decreased chondrogenic differentiation and ASC viability in scaffold culture, as well as tendon scaffold repopulation, and strongly influenced musculoskeletal gene expression. Effects regarding the latter differed between the monolayer and scaffold cultures. Leukocytes rather decreased ASC proliferation, but had similar effects on viability and musculoskeletal gene expression. This included decreased expression of the tenogenic transcription factor scleraxis by an inflammatory environment throughout culture conditions. The data demonstrate that ASC tenogenic properties are compromised in an inflammatory environment, with relevance to their possible mechanisms of action in acute tendon disease.


Assuntos
Diferenciação Celular , Inflamação/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Adipogenia , Animais , Biomarcadores , Sobrevivência Celular , Células Cultivadas , Microambiente Celular , Condrogênese , Técnicas de Cocultura , Citocinas/metabolismo , Matriz Extracelular/metabolismo , Cavalos , Humanos , Inflamação/etiologia , Inflamação/patologia , Mediadores da Inflamação/metabolismo , Tendões , Alicerces Teciduais
7.
NMR Biomed ; 29(7): 952-60, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27195474

RESUMO

Individual tumor characterization and treatment response monitoring based on current medical imaging methods remain challenging. This work investigates hyperpolarized (13) C compounds in an orthotopic rat hepatocellular carcinoma (HCC) model system before and after transcatheter arterial embolization (TAE). HCC ranks amongst the top six most common cancer types in humans and accounts for one-third of cancer-related deaths worldwide. Early therapy response monitoring could aid in the development of personalized therapy approaches and novel therapeutic concepts. Measurements with selectively (13) C-labeled and hyperpolarized urea, pyruvate and fumarate were performed in tumor-bearing rats before and after TAE. Two-dimensional, slice-selective MRSI was used to obtain spatially resolved maps of tumor perfusion, cell energy metabolic conversion rates and necrosis, which were additionally correlated with immunohistochemistry. All three injected compounds, taken together with their respective metabolites, exhibited similar signal distributions. TAE induced a decrease in blood flow into the tumor and thus a decrease in tumor to muscle and tumor to liver ratios of urea, pyruvate and its metabolites, alanine and lactate, whereas conversion rates remained stable or increased on TAE in tumor, muscle and liver tissue. Conversion from fumarate to malate successfully indicated individual levels of necrosis, and global malate signals after TAE suggested the washout of fumarase or malate itself on necrosis. This study presents a combination of three (13) C compounds as novel candidate biomarkers for a comprehensive characterization of genetically and molecularly diverse HCC using hyperpolarized MRSI, enabling the simultaneous detection of differences in tumor perfusion, metabolism and necrosis. If, as in this study, bolus dynamics are not required and qualitative perfusion information is sufficient, the desired information could be extracted from hyperpolarized fumarate and pyruvate alone, acquired at higher fields with better spectral separation. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Espectroscopia de Ressonância Magnética Nuclear de Carbono-13/métodos , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/terapia , Embolização Terapêutica/métodos , Imagem Molecular/métodos , Compostos Orgânicos/metabolismo , Animais , Carcinoma Hepatocelular/diagnóstico , Linhagem Celular Tumoral , Feminino , Imageamento por Ressonância Magnética/métodos , Ratos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Resultado do Tratamento
8.
Transl Sports Med ; 2023: 4488334, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-38654917

RESUMO

Background: Extensive physical activity (PA; ≥18 MET∗h/week, MET metabolic equivalent of tasks hours) postcancer diagnosis has shown favorable effects on colorectal cancer disease-free survival. However, the feasibility of introducing this high volume of PA in this patient group is unclear. Therefore, the aim of the F-PROTECT study was to evaluate the feasibility of extensive and prolonged PA (≥18 MET∗h/week over 12 months) in colorectal cancer patients with the primary objectives to (1) recruit 50 patients within 12 months and (2) reach an attendance rate of ≥70%. Methods: Single-armed, bicentric, prospective intervention study in colorectal cancer patients (≤80 years; UICC II/III Union for International Cancer Control) after histopathological confirmed R0-resection who were consecutively recruited from visceral surgery units of 10 clinics in Germany. Recruitment rates were calculated using screening logs. Intervention was a 12-month endurance-focused exercise program with supervised and home-based training. Attendance rates defined as ≥70% participation in training sessions were calculated by training diaries. Results: Out of 521 patients who were screened for eligibility, 50 (23 female; 59 ± 10 years, UICC 44% II, 56% III; adjuvant chemotherapy 60%) were recruited within 15 months. Mean duration between surgery and first training was 103 ± 57 days. Training attendance rate was 64% (including 9 dropouts). Six (12%) participants reached ≥18 MET∗h/week in ≥70% of training sessions between 4-12 months. 28 adverse events (n = 9 serious) occurred, however, were not assessed as training related. Conclusions: The present intervention involving a combination of supervised and home-based exercise training in postsurgical colorectal cancer patients was not feasible. Strategies specifically designed for this patient group must be developed and investigated to motivate long-term PA. Registration. The study was prospectively registered at clinicaltrials.gov (NCT01991847).

9.
Mol Neurobiol ; 60(5): 2602-2618, 2023 May.
Artigo em Inglês | MEDLINE | ID: mdl-36692708

RESUMO

PPP1R21 acts as a co-factor for protein phosphatase 1 (PP1), an important serine/threonine phosphatase known to be essential for cell division, control of glycogen metabolism, protein synthesis, and muscle contractility. Bi-allelic pathogenic variants in PPP1R21 were linked to a neurodevelopmental disorder with hypotonia, facial dysmorphism, and brain abnormalities (NEDHFBA) with pediatric onset. Functional studies unraveled impaired vesicular transport as being part of PPP1R21-related pathomechanism. To decipher further the pathophysiological processes leading to the clinical manifestation of NEDHFBA, we investigated the proteomic signature of fibroblasts derived from the first NEDHFBA patient harboring a splice-site mutation in PPP1R21 and presenting with a milder phenotype. Proteomic findings and further functional studies demonstrate a profound activation of the ubiquitin-proteasome system with presence of protein aggregates and impact on cellular fitness and moreover suggest a cross-link between activation of the proteolytic system and cytoskeletal architecture (including filopodia) as exemplified on paradigmatic proteins including actin, thus extending the pathophysiological spectrum of the disease. In addition, the proteomic signature of PPP1R21-mutant fibroblasts displayed a dysregulation of a variety of proteins of neurological relevance. This includes increase proteins which might act toward antagonization of cellular stress burden in terms of pro-survival, a molecular finding which might accord with the presentation of a milder phenotype of our NEDHFBA patient.


Assuntos
Complexo de Endopeptidases do Proteassoma , Fala , Humanos , Actinas , Debilidade Muscular , Mutação/genética , Fenótipo , Proteína Fosfatase 1/genética , Proteômica
10.
J Neurol ; 270(6): 3138-3158, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-36892629

RESUMO

BACKGROUND AND PURPOSE: Myotonic dystrophy type 1 (DM1) is the most common form of adult-onset muscular dystrophy and is caused by an repeat expansion [r(CUG)exp] located in the 3' untranslated region of the DMPK gene. Symptoms include skeletal and cardiac muscle dysfunction and fibrosis. In DM1, there is a lack of established biomarkers in routine clinical practice. Thus, we aimed to identify a blood biomarker with relevance for DM1-pathophysiology and clinical presentation. METHODS: We collected fibroblasts from 11, skeletal muscles from 27, and blood samples from 158 DM1 patients. Moreover, serum, cardiac, and skeletal muscle samples from DMSXL mice were included. We employed proteomics, immunostaining, qPCR and ELISA. Periostin level were correlated with CMRI-data available for some patients. RESULTS: Our studies identified Periostin, a modulator of fibrosis, as a novel biomarker candidate for DM1: proteomic profiling of human fibroblasts and murine skeletal muscles showed significant dysregulation of Periostin. Immunostaining on skeletal and cardiac muscles from DM1 patients and DMSXL mice showed an extracellular increase of Periostin, indicating fibrosis. qPCR studies indicated increased POSTN expression in fibroblasts and muscle. Quantification of Periostin in blood samples from DMSXL mice and two large validation cohorts of DM1 patients showed decreased levels in animals and diseased individuals correlating with repeat expansion and disease severity and presence of cardiac symptoms identified by MRI. Analyses of longitudinal blood samples revealed no correlation with disease progression. CONCLUSIONS: Periostin might serve as a novel stratification biomarker for DM1 correlating with disease severity, presence of cardiac malfunction and fibrosis.


Assuntos
Cardiomiopatias , Distrofia Miotônica , Adulto , Humanos , Camundongos , Animais , Distrofia Miotônica/genética , Expansão das Repetições de Trinucleotídeos , Proteômica , Músculo Esquelético , Células Musculares/metabolismo , Cardiomiopatias/genética , Cardiomiopatias/metabolismo , Gravidade do Paciente , Miotonina Proteína Quinase/genética
11.
Orphanet J Rare Dis ; 17(1): 29, 2022 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-35101074

RESUMO

BACKGROUND: Goltz syndrome (GS) is a X-linked disorder defined by defects of mesodermal- and ectodermal-derived structures and caused by PORCN mutations. Features include striated skin-pigmentation, ocular and skeletal malformations and supernumerary or hypoplastic nipples. Generally, GS is associated with in utero lethality in males and most of the reported male patients show mosaicism (only three non-mosaic surviving males have been described so far). Also, precise descriptions of neurological deficits in GS are rare and less severe phenotypes might not only be caused by mosaicism but also by less pathogenic mutations suggesting the need of a molecular genetics and functional work-up of these rare variants. RESULTS: We report two cases: one girl suffering from typical skin and skeletal abnormalities, developmental delay, microcephaly, thin corpus callosum, periventricular gliosis and drug-resistant epilepsy caused by a PORCN nonsense-mutation (c.283C > T, p.Arg95Ter). Presence of these combined neurological features indicates that CNS-vulnerability might be a guiding symptom in the diagnosis of GS patients. The other patient is a boy with a supernumerary nipple and skeletal anomalies but also, developmental delay, microcephaly, cerebral atrophy with delayed myelination and drug-resistant epilepsy as predominant features. Skin abnormalities were not observed. Genotyping revealed a novel PORCN missense-mutation (c.847G > C, p.Asp283His) absent in the Genome Aggregation Database (gnomAD) but also identified in his asymptomatic mother. Given that non-random X-chromosome inactivation was excluded in the mother, fibroblasts of the index had been analyzed for PORCN protein-abundance and -distribution, vulnerability against additional ER-stress burden as well as for protein secretion revealing changes. CONCLUSIONS: Our combined findings may suggest incomplete penetrance for the p.Asp283His variant and provide novel insights into the molecular etiology of GS by adding impaired ER-function and altered protein secretion to the list of pathophysiological processes resulting in the clinical manifestation of GS.


Assuntos
Aciltransferases , Hipoplasia Dérmica Focal , Proteínas de Membrana , Aciltransferases/genética , Feminino , Hipoplasia Dérmica Focal/complicações , Hipoplasia Dérmica Focal/genética , Hipoplasia Dérmica Focal/patologia , Humanos , Masculino , Proteínas de Membrana/genética , Mutação , Fenótipo
12.
Cancers (Basel) ; 13(11)2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-34205964

RESUMO

The molecular carcinogenesis of intraductal tubulopapillary neoplasms (ITPN), recently described as rare neoplasms in the pancreato-biliary tract with a favorable prognosis despite a high incidence of associated pancreato-biliary adenocarcinoma, is still poorly understood. To identify driver genes, chromosomal gains and losses, mutational signatures, key signaling pathways, and potential therapeutic targets, the molecular profile of 11 biliary and 6 pancreatic ITPNs, associated with invasive adenocarcinoma in 14/17 cases, are studied by whole exome sequencing (WES). The WES of 17 ITPNs reveals common copy number variants (CNVs) broadly distributed across the genome, with recurrent chromosomal deletions primarily in 1p36 and 9p21 affecting the tumor suppressors CHD5 and CDKN2A, respectively, and gains in 1q affecting the prominent oncogene AKT3. The identified somatic nucleotide variants (SNVs) involve few core signaling pathways despite high genetic heterogeneity with diverse mutational spectra: Chromatin remodeling, the cell cycle, and DNA damage/repair. An OncoKB search identifies putative actionable genomic targets in 35% of the cases (6/17), including recurrent missense mutations of the FGFR2 gene in biliary ITPNs (2/11, 18%). Our results show that somatic SNV in classical cancer genes, typically associated with pancreato-biliary carcinogenesis, were absent (KRAS, IDH1/2, GNAS, and others) to rare (TP53 and SMAD4, 6%, respectively) in ITPNs. Mutational signature pattern analysis reveals a predominance of an age-related pattern. Our findings highlight that biliary ITPN and classical cholangiocarcinoma display commonalities, in particular mutations in genes of the chromatin remodeling pathway, and appear, therefore, more closely related than pancreatic ITPN and classical pancreatic ductal adenocarcinoma.

13.
J Equine Vet Sci ; 94: 103227, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-33077088

RESUMO

In magnetic resonance imaging (MRI) examinations, moderate to severe changes of the distal sesamoidean impar ligament (DSIL) were found in horses with lameness localized to their feet. Histologic abnormalities were detected more commonly in lame horses. Because of its heterogeneity and small thickness, evaluation of the DSIL in MRI can be challenging. The aim of the study was to determine the optimal sequence and the ideal transverse perpendicular angle for visualization of the DSIL before and after arthrography of the distal interphalangeal joint (DIPJ). Twenty-five cadaver forelimbs were examined with low-field MRI. Sagittal, frontal, and three different angled transverse planes were obtained before and after arthrography of the DIPJ. All planes were acquired in T1w (weighted) Gradient Recall Echo (GRE), T2∗w GRE, T2w Fast Spin Echo (FSE), and Short Tau Inversion Recovery (STIR) FSE and visualization of the DSIL was scored by two observers. Visualization of the DSIL was best on sagittal T2w FSE and STIR FSE images. All transverse planes were inferior compared with sagittal sequences. After arthrography of the DIPJ, visualization of the DSIL origin improved in sagittal T2w FSE sequences, and agreement between observers increased for sagittal T2w FSE and STIR FSE images. Sagittal T2w FSE and STIR FSE images allowed good visualization of the DSIL in low-field MRI. Visualization of the DSIL did not improve for altered angled transverse sequences but increased with arthrography of the DIPJ. Subjective influence between different observers was found but decreased with DIPJ arthrography.


Assuntos
Artrografia , Imageamento por Ressonância Magnética , Animais , Membro Anterior , Cavalos , Ligamentos , Variações Dependentes do Observador
14.
PLoS One ; 14(5): e0217576, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31150471

RESUMO

Doxorubicin (DOX) is a widely used chemotherapeutic anticancer drug. Its intrinsic fluorescence properties enable investigation of tumor response, drug distribution and metabolism. First phantom studies in vitro showed optoacoustic property of DOX. We therefore aimed to further investigate the optoacoustic properties of DOX in biological tissue in order to explore its potential as theranostic agent. We analysed doxorubicin hydrochloride (Dox·HCl) and liposomal encapsulated doxorubicin hydrochloride (Dox·Lipo), two common drugs for anti-cancer treatment in clinical medicine. Optoacoustic measurements revealed a strong signal of both doxorubicin substrates at 488 nm excitation wavelength. Post mortem analysis of intra-tumoral injections of DOX revealed a detectable optoacoustic signal even at three days after the injection. We thereby demonstrate the general feasibility of doxorubicin detection in biological tissue by means of optoacoustic tomography, which could be applied for high resolution imaging at mesoscopic depths dictated by effective penetration of visible light into the biological tissues.


Assuntos
Doxorrubicina/análogos & derivados , Neoplasias/diagnóstico por imagem , Técnicas Fotoacústicas/métodos , Nanomedicina Teranóstica/métodos , Tomografia/métodos , Animais , Linhagem Celular Tumoral/transplante , Modelos Animais de Doenças , Doxorrubicina/administração & dosagem , Doxorrubicina/química , Estudos de Viabilidade , Feminino , Humanos , Injeções Intralesionais , Camundongos , Projetos Piloto , Polietilenoglicóis/administração & dosagem , Polietilenoglicóis/química
15.
Cell Transplant ; 27(10): 1434-1450, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30251565

RESUMO

Age-related degenerative changes in tendon tissue represent a common cause for acute tendon pathologies. Although the regenerative potential of multipotent mesenchymal stromal cells (MSC) was reported to restore functionality in injured tendon tissue, cellular mechanisms of action remain partly unclear. Potential tenogenic differentiation of applied MSC is affected by various intrinsic and extrinsic factors. The current study presents an in vitro model to evaluate the combined extrinsic effects of decellularized equine tendon matrix, transforming growth factor beta 3 (TGFß3) and bone morphogenetic protein 12 (BMP12) on the tenogenic fate of equine adipose tissue-derived MSC. Monolayer MSC cultures supplemented with TGFß3 and BMP12 as well as MSC cultured on tendon matrix scaffolds preloaded with the growth factors were incubated for 3 and 5 days. Histological evaluation and real time reverse transcription polymerase chain reaction (RT-PCR) revealed that growth factor-mediated tenogenic induction of MSC was modified by the conditions of the surrounding microenvironment. While the gene expression pattern in monolayer cultures supplemented with TGFß3 or TGFß3 and BMP12 revealed an upregulation for collagen 1A2, collagen 3A1, tenascin c, scleraxis and mohawk ( p < 0.05 ), the presence of tendon matrix led to an upregulation of decorin and osteopontin as well as to a downregulation of smad8 ( p < 0.05). Preloading of scaffolds with either TGFß3, or with TGFß3 and BMP12 promoted a tenocyte-like phenotype and improved cell alignment. Furthermore, gene expression in scaffold culture was modulated by TGFß3 and/or BMP12, with downregulation of collagen 1A2, collagen 3A1, decorin, scleraxis, smad8 and osteopontin, whereas gene expression of tenascin c was increased. This study shows that growth factor-induced tenogenic differentiation of equine MSC is markedly altered by topographical constraints of decellularized tendon tissue in vitro. While TGFß3 represents an effective mediator for tenogenic induction, the role of BMP12 in tenogenesis may be of modulatory character and needs further evaluation.


Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Células-Tronco Mesenquimais/citologia , Tendões/química , Tendões/citologia , Alicerces Teciduais/química , Fator de Crescimento Transformador beta3/metabolismo , Animais , Diferenciação Celular , Sobrevivência Celular , Células Cultivadas , Expressão Gênica , Cavalos , Células-Tronco Mesenquimais/metabolismo , Tendões/ultraestrutura , Engenharia Tecidual/métodos
17.
Brain Res ; 1155: 24-33, 2007 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-17512917

RESUMO

Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder linked to a mutation in the huntingtin gene leading to protein aggregation in neurons. The generation of new neurons in neurogenic regions, such as the subventricular zone of the lateral ventricle and the dentate gyrus of the hippocampus, is affected by these aggregation processes. In particular, hippocampal neurogenesis is reduced in the R6/2 transgenic mouse model of HD. Since physical activity stimulates adult hippocampal neurogenesis, we examined whether running is capable to rescue the impaired hippocampal neurogenesis in R6/2 mice. Proliferation of hippocampal cells measured by proliferating cell nuclear antigen (PCNA) marker was reduced in R6/2 animals by 64% compared to wild type mice. Accordingly, newly generated neurons labeled with doublecortin (DCX) were diminished by 60% in the hippocampus of R6/2 mice. Furthermore, the number of newly generated mature neurons was decreased by 76%. Within the hippocampus of wild type animals, a four-week running period resulted in a doubling of PCNA-, DCX-, and bromo-deoxyuridine (BrdU)-labeled cells. However, physical exercise failed to stimulate proliferation and survival of newly generated neurons in R6/2 transgenic mouse model of HD. These findings suggest that mutant huntingtin alters the hippocampal microenvironment thus resulting in an impaired neurogenesis. Importantly, this adverse microenvironment impeded neurogenesis upregulation such as induced by physical exercise. Future studies need to decipher the molecular pathways involved in repressing the generation of new neurons after physical activity in huntingtin transgenic rodents.


Assuntos
Hipocampo/fisiopatologia , Doença de Huntington/terapia , Atividade Motora/fisiologia , Condicionamento Físico Animal/fisiologia , Animais , Divisão Celular , Modelos Animais de Doenças , Proteína Duplacortina , Hipocampo/patologia , Humanos , Proteína Huntingtina , Doença de Huntington/patologia , Doença de Huntington/fisiopatologia , Masculino , Camundongos , Camundongos Transgênicos , Regeneração Nervosa , Proteínas do Tecido Nervoso/genética , Proteínas Nucleares/genética , Células-Tronco/patologia , Células-Tronco/fisiologia
18.
Eur J Paediatr Neurol ; 11(1): 46-9, 2007 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17161965

RESUMO

Walker--Warburg syndrome (WWS), the most severe alpha-dystroglycanopathy, is characterized by brain and eye anomalies, and congenital muscular dystrophy (CMD). So far at least four genes (POMT1, POMT2, Fukutin, and FKRP gene) have been implicated in WWS, accounting for about 30% of all cases. We report a male patient with WWS resulting from a homozygous nonsense mutation (R514X) in the POMT1 gene. The patient had congenital hydrocephalus which was detected at 29 weeks of gestation. A brain MRI obtained after birth revealed type II lissencephaly, hydrocephalus, and pontocerebellar hypoplasia. The case also exhibited severe ocular malformations and muscular hypotonia due to CMD.


Assuntos
Códon sem Sentido/genética , Hidrocefalia/genética , Manosiltransferases/genética , Distrofias Musculares/congênito , Distrofias Musculares/genética , Humanos , Recém-Nascido , Masculino , Síndrome
19.
Clin Neurol Neurosurg ; 108(7): 692-8, 2006 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-16102895

RESUMO

We report the clinical, structural, functional and genetic characterization of a 37-year-old Caucasian female, presenting as a sporadic case of complicated spastic paraplegia with thin corpus callosum (CC), cognitive impairment, amyotrophy of the hand muscles and a sensorimotor neuropathy and review the literature for spastic paraplegia with thin CC. Magnetic resonance imaging (MRI) examination revealed a thin CC with fronto-parietal cortical atrophy. 18Fluordesoxyglucose positron emission tomography (FDG-PET) showed reduced cortical and thalamic metabolism. By transcranial magnetic stimulation, we delineated a severe impairment of transcallosal inhibition. Sequence analysis did not reveal disease causing mutations in the genes SLC12A6 (Andermann), Spastin (SPG 4), BSCL2 (SPG 17) and Spartin (SPG 20). We reviewed the literature for HSP with thin CC and found 113 HSP patients with thin CC previously described (35 with linkage to chromosome 15q13-15). Thin CC and peripheral neuropathy often appear together in spastic paraplegia and might be indicative for combined degeneration mechanism of central and peripheral axons.


Assuntos
Corpo Caloso/patologia , Atrofia Muscular/complicações , Malformações do Sistema Nervoso/complicações , Paraplegia/complicações , Doenças do Sistema Nervoso Periférico/complicações , Adulto , Córtex Cerebral/diagnóstico por imagem , Córtex Cerebral/patologia , Córtex Cerebral/fisiopatologia , Cromossomos Humanos Par 15/genética , Transtornos Cognitivos/diagnóstico por imagem , Transtornos Cognitivos/patologia , Transtornos Cognitivos/fisiopatologia , Corpo Caloso/diagnóstico por imagem , Corpo Caloso/fisiopatologia , Análise Mutacional de DNA , Metabolismo Energético/fisiologia , Feminino , Mãos/patologia , Mãos/fisiopatologia , Humanos , Padrões de Herança/genética , Imageamento por Ressonância Magnética , Músculo Esquelético/patologia , Músculo Esquelético/fisiopatologia , Atrofia Muscular/genética , Atrofia Muscular/fisiopatologia , Mutação , Malformações do Sistema Nervoso/genética , Malformações do Sistema Nervoso/fisiopatologia , Paraplegia/genética , Paraplegia/fisiopatologia , Linhagem , Nervos Periféricos/patologia , Nervos Periféricos/fisiopatologia , Doenças do Sistema Nervoso Periférico/genética , Doenças do Sistema Nervoso Periférico/fisiopatologia , Tomografia por Emissão de Pósitrons , Síndrome
20.
Neuromuscul Disord ; 15(4): 271-5, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15792865

RESUMO

Mutations of the protein O-mannosyltransferase (POMT1) gene affect glycosylation of alpha-dystroglycan, leading to Walker-Warburg syndrome, a lethal disorder in early life with severe congenital muscular dystrophy, and brain and eye malformations. Recently, we described a novel form of recessive limb girdle muscular dystrophy with mild mental retardation, associated with an abnormal alpha-dystroglycan pattern in the muscle, suggesting a glycosylation defect. Here, we present evidence that this distinct phenotype results from a common mutation (A200P) in the POMT1 gene. Our findings further expand the phenotype of glycosylation disorders linked to POMT1 mutations. Furthermore, the A200P mutation is part of a conserved core haplotype, indicating an ancestral founder mutation.


Assuntos
Deficiência Intelectual/genética , Manosiltransferases/genética , Distrofia Muscular do Cíngulo dos Membros/genética , Mutação/genética , Adolescente , Adulto , Alanina/genética , Alelos , Criança , Análise Mutacional de DNA/métodos , Feminino , Humanos , Deficiência Intelectual/fisiopatologia , Masculino , Modelos Moleculares , Distrofia Muscular do Cíngulo dos Membros/fisiopatologia , Fenótipo , Polimorfismo de Nucleotídeo Único , Prolina/genética , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos
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