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1.
Cell ; 183(6): 1617-1633.e22, 2020 12 10.
Article in English | MEDLINE | ID: mdl-33259802

ABSTRACT

Histone H3.3 glycine 34 to arginine/valine (G34R/V) mutations drive deadly gliomas and show exquisite regional and temporal specificity, suggesting a developmental context permissive to their effects. Here we show that 50% of G34R/V tumors (n = 95) bear activating PDGFRA mutations that display strong selection pressure at recurrence. Although considered gliomas, G34R/V tumors actually arise in GSX2/DLX-expressing interneuron progenitors, where G34R/V mutations impair neuronal differentiation. The lineage of origin may facilitate PDGFRA co-option through a chromatin loop connecting PDGFRA to GSX2 regulatory elements, promoting PDGFRA overexpression and mutation. At the single-cell level, G34R/V tumors harbor dual neuronal/astroglial identity and lack oligodendroglial programs, actively repressed by GSX2/DLX-mediated cell fate specification. G34R/V may become dispensable for tumor maintenance, whereas mutant-PDGFRA is potently oncogenic. Collectively, our results open novel research avenues in deadly tumors. G34R/V gliomas are neuronal malignancies where interneuron progenitors are stalled in differentiation by G34R/V mutations and malignant gliogenesis is promoted by co-option of a potentially targetable pathway, PDGFRA signaling.


Subject(s)
Brain Neoplasms/genetics , Carcinogenesis/genetics , Glioma/genetics , Histones/genetics , Interneurons/metabolism , Mutation/genetics , Neural Stem Cells/metabolism , Receptor, Platelet-Derived Growth Factor alpha/genetics , Animals , Astrocytes/metabolism , Astrocytes/pathology , Brain Neoplasms/pathology , Carcinogenesis/pathology , Cell Lineage , Cellular Reprogramming/genetics , Chromatin/metabolism , Embryo, Mammalian/metabolism , Epigenesis, Genetic , Gene Expression Regulation, Neoplastic , Gene Silencing , Glioma/pathology , Histones/metabolism , Lysine/metabolism , Mice, Inbred C57BL , Models, Biological , Neoplasm Grading , Oligodendroglia/metabolism , Promoter Regions, Genetic/genetics , Prosencephalon/embryology , Receptor, Platelet-Derived Growth Factor alpha/metabolism , Transcription, Genetic , Transcriptome/genetics
2.
J Med Genet ; 61(9): 908-913, 2024 Aug 29.
Article in English | MEDLINE | ID: mdl-38955476

ABSTRACT

BACKGROUND: Transport protein particle (TRAPP) is a multiprotein complex that functions in localising proteins to the Golgi compartment. The TRAPPC11 subunit has been implicated in diseases affecting muscle, brain, eye and to some extent liver. We present three patients who are compound heterozygotes for a missense variant and a structural variant in the TRAPPC11 gene. TRAPPC11 structural variants have not yet been described in association with a disease. In order to reveal the estimated genesis of identified structural variants, we performed sequencing of individual breakpoint junctions and analysed the extent of homology and the presence of repetitive elements in and around the breakpoints. METHODS: Biochemical methods including isoelectric focusing on serum transferrin and apolipoprotein C-III, as well as mitochondrial respiratory chain complex activity measurements, were used. Muscle biopsy samples underwent histochemical analysis. Next-generation sequencing was employed for identifying sequence variants associated with neuromuscular disorders, and Sanger sequencing was used to confirm findings. RESULTS: We suppose that non-homologous end joining is a possible mechanism of deletion origin in two patients and non-allelic homologous recombination in one patient. Analyses of mitochondrial function performed in patients' skeletal muscles revealed an imbalance of mitochondrial metabolism, which worsens with age and disease progression. CONCLUSION: Our results contribute to further knowledge in the field of neuromuscular diseases and mutational mechanisms. This knowledge is important for understanding the molecular nature of human diseases and allows us to improve strategies for identifying disease-causing mutations.


Subject(s)
Muscular Dystrophies , Adult , Child , Female , Humans , Male , Gene Deletion , High-Throughput Nucleotide Sequencing , Muscle, Skeletal/pathology , Muscle, Skeletal/metabolism , Muscular Dystrophies/genetics , Muscular Dystrophies/pathology , Mutation, Missense/genetics
3.
Acta Neuropathol ; 145(1): 49-69, 2023 01.
Article in English | MEDLINE | ID: mdl-36437415

ABSTRACT

Pediatric central nervous system (CNS) tumors represent the most common cause of cancer-related death in children aged 0-14 years. They differ from their adult counterparts, showing extensive clinical and molecular heterogeneity as well as a challenging histopathological spectrum that often impairs accurate diagnosis. Here, we use DNA methylation-based CNS tumor classification in combination with copy number, RNA-seq, and ChIP-seq analysis to characterize a newly identified CNS tumor type. In addition, we report histology, patient characteristics, and survival data in this tumor type. We describe a biologically distinct pediatric CNS tumor type (n = 31 cases) that is characterized by focal high-level amplification and resultant overexpression of either PLAGL1 or PLAGL2, and an absence of recurrent genetic alterations characteristic of other pediatric CNS tumor types. Both genes act as transcription factors for a regulatory subset of imprinted genes (IGs), components of the Wnt/ß-Catenin pathway, and the potential drug targets RET and CYP2W1, which are also specifically overexpressed in this tumor type. A derived PLAGL-specific gene expression signature indicates dysregulation of imprinting control and differentiation/development. These tumors occurred throughout the neuroaxis including the cerebral hemispheres, cerebellum, and brainstem, and were predominantly composed of primitive embryonal-like cells lacking robust expression of markers of glial or neuronal differentiation (e.g., GFAP, OLIG2, and synaptophysin). Tumors with PLAGL1 amplification were typically diagnosed during adolescence (median age 10.5 years), whereas those with PLAGL2 amplification were diagnosed during early childhood (median age 2 years). The 10-year overall survival was 66% for PLAGL1-amplified tumors, 25% for PLAGL2-amplified tumors, 18% for male patients, and 82% for female patients. In summary, we describe a new type of biologically distinct CNS tumor characterized by PLAGL1/2 amplification that occurs predominantly in infants and toddlers (PLAGL2) or adolescents (PLAGL1) which we consider best classified as a CNS embryonal tumor and which is associated with intermediate survival. The cell of origin and optimal treatment strategies remain to be defined.


Subject(s)
Central Nervous System Neoplasms , Neuroectodermal Tumors, Primitive , Child , Child, Preschool , Female , Humans , Infant , Male , Cell Cycle Proteins/genetics , Central Nervous System Neoplasms/genetics , DNA Methylation , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Neuroectodermal Tumors, Primitive/genetics , RNA-Binding Proteins/genetics , Transcription Factors/genetics , Transcription Factors/metabolism , Tumor Suppressor Proteins/genetics , Wnt Signaling Pathway/genetics
4.
Acta Neurochir (Wien) ; 165(1): 177-186, 2023 01.
Article in English | MEDLINE | ID: mdl-36437400

ABSTRACT

PURPOSE: Intracranial aneurysm (IA) rupture results in one of the most severe forms of stroke, with severe neurological sequelae. Inflammation appears to drive aneurysm formation and progression with macrophages playing a key role in this process. However, less is known about their involvement in aneurysm rupture. This study is aimed at demonstrating how relationship between the M1 (pro-inflammatory) and M2 (reparative) macrophage subtypes affect an aneurysm's structure resulting in its rupture. METHODS: Forty-one saccular aneurysm wall samples were collected during surgery including 13 ruptured and 28 unruptured aneurysm sacs. Structural changes were evaluated using histological staining. Macrophages in the aneurysm wall were quantified and defined as M1 and M2 using HLA-DR and CD163 antibodies. Aneurysm samples were divided into four groups according to the structural changes and the M2/1 ratio. Data were analyzed using the Mann-Whitney U test. RESULTS: This study has demonstrated an association between the severity of structural changes of an aneurysm with inflammatory cell infiltration within its wall and subsequent aneurysm rupture. More severe morphological changes and a significantly higher number of inflammatory cells were observed in ruptured IAs (p < 0.001). There was a prevalence of M2 macrophage subtypes within the wall of ruptured aneurysms (p < 0.001). A subgroup of unruptured IAs with morphological and inflammatory changes similar to ruptured IAs was observed. The common feature of this subgroup was the presence of an intraluminal thrombus. CONCLUSIONS: The degree of inflammatory cell infiltration associated with a shift in macrophage phenotype towards M2 macrophages could play an important role in structural changes of the aneurysm wall leading to its rupture.


Subject(s)
Aneurysm, Ruptured , Intracranial Aneurysm , Macrophages , Humans , Aneurysm, Ruptured/complications , Aneurysm, Ruptured/pathology , Inflammation/complications , Intracranial Aneurysm/complications , Intracranial Aneurysm/pathology , Macrophages/pathology , Thrombosis/complications
5.
Acta Obstet Gynecol Scand ; 101(1): 37-45, 2022 Jan.
Article in English | MEDLINE | ID: mdl-34693986

ABSTRACT

INTRODUCTION: Uterus transplantation is a causal treatment for absolute uterine factor infertility. Assessing rejection signs using a histopathological examination of the ectocervical biopsy from the transplanted uterus is common practice in all human uterus transplants worldwide to date. A provisional scoring system was used for the histopathological assessment of subclinical rejection signs in uterus recipients. Here we hypothesized that histopathological and immunohistochemical findings in the normal uteri would differ from the borderline category of subclinical rejection in uterine transplants. MATERIAL AND METHODS: This prospective observational study included ectocervical biopsies of 54 women who underwent hysterectomy for benign reasons. All biopsy samples were assessed histopathologically and immunohistochemically. RESULTS: Most of the ectocervical biopsies showed clustering lymphocytic infiltrates affecting the stromal-epithelial interface with the epithelial influx of lymphocytes, primarily CD45RO-positive activated T-cells with CD8 T-lymphocyte predominance. CD4-positive T-lymphocytes and B-cells were rarely detected in the ectocervix. These morphological findings and immunoprofiles of lymphocytic populations overlapped with the so-called borderline changes defined in the provisional scoring system for rejection in the transplanted uteri. The immunoprofiles of ectocervical and endocervical lymphocytic populations differed, with strikingly prominent B-cell participation in the endocervix vs the rare detection of B-cells in the ectocervix. CONCLUSIONS: The histopathological and immunohistochemical findings in the uteri of premenopausal women were similar to the borderline category of the currently used provisional scoring system of subclinical uterine rejection utilized in all uterine transplant studies. However, future similar studies are required to validate our findings.


Subject(s)
Cervix Uteri/pathology , Graft Rejection/pathology , Infertility, Female/surgery , Uterus/transplantation , Adult , Biopsy , Female , Humans , Hysterectomy , Middle Aged , Prospective Studies , Research Design
6.
Acta Neurochir (Wien) ; 164(6): 1459-1472, 2022 06.
Article in English | MEDLINE | ID: mdl-35043265

ABSTRACT

BACKGROUND: Childhood thalamopeduncular gliomas arise at the interface of the thalamus and cerebral peduncle. The optimal treatment is total resection but not at the cost of neurological function. We present long-term clinical and oncological outcomes of maximal safe resection. METHODS: Retrospective review of prospectively collected data: demography, symptomatology, imaging, extent of resection, surgical complications, histology, functional and oncological outcome. RESULTS: During 16-year period (2005-2020), 21 patients were treated at our institution. These were 13 girls and 8 boys (mean age 7.6 years). Presentation included progressive hemiparesis in 9 patients, raised intracranial pressure in 9 patients and cerebellar symptomatology in 3 patients. The tumour was confined to the thalamus in 6 cases. Extent of resection was judged on postoperative imaging as total (6), near-total (6) and less extensive (9). Surgical complications included progression of baseline neurological status in 6 patients, and 5 of these gradually improved to preoperative status. All tumours were classified as low-grade gliomas. Disease progression was observed in 9 patients (median progression-free survival 7.3 years). At last follow-up (median 6.1 years), all patients were alive, median Lansky score of 90. Seven patients were without evidence of disease, 6 had stable disease, 7 stable following progression and 1 had progressive disease managed expectantly. CONCLUSION: Paediatric patients with low-grade thalamopeduncular gliomas have excellent long-term functional and oncological outcomes when gross total resection is not achievable. Surgery should aim at total resection; however, neurological function should not be endangered due to excellent chance for long-term survival.


Subject(s)
Brain Neoplasms , Glioma , Brain Neoplasms/complications , Brain Neoplasms/diagnostic imaging , Brain Neoplasms/surgery , Child , Female , Glioma/complications , Glioma/diagnostic imaging , Glioma/surgery , Humans , Magnetic Resonance Imaging , Male , Neurosurgical Procedures/methods , Retrospective Studies , Thalamus/diagnostic imaging , Thalamus/pathology , Thalamus/surgery , Treatment Outcome
7.
Int J Mol Sci ; 23(19)2022 Sep 28.
Article in English | MEDLINE | ID: mdl-36232755

ABSTRACT

Compelling evidence supports the health benefits of physical exercise on the immune system, possibly through the molecules secreted by the skeletal muscles known as myokines. Herein, we assessed the impact of exercise interventions on plasma Heat shock protein 90 (Hsp90) levels in 27 patients with idiopathic inflammatory myopathies (IIM) compared with 23 IIM patients treated with standard-of-care immunosuppressive therapy only, and in 18 healthy subjects undergoing strenuous eccentric exercise, and their associations with the traditional serum markers of muscle damage and inflammation. In contrast to IIM patients treated with pharmacotherapy only, in whom we demonstrated a significant decrease in Hsp90 over 24 weeks, the 24-week exercise program resulted in a stabilization of Hsp90 levels. These changes in Hsp90 levels were associated with changes in several inflammatory cytokines/chemokines involved in the pathogenesis of IIM or muscle regeneration in general. Strenuous eccentric exercise in healthy volunteers induced a brief increase in Hsp90 levels with a subsequent return to baseline levels at 14 days after the exercise, with less pronounced correlations to systemic inflammation. In this study, we identified Hsp90 as a potential myokine and mediator for exercise-induced immune response and as a potential biomarker predicting improvement after physiotherapy in muscle endurance in IIM.


Subject(s)
Exercise Therapy , HSP90 Heat-Shock Proteins , Inflammation , Muscle, Skeletal , Myositis , Biomarkers/blood , Biomarkers/metabolism , Chemokines/blood , Chemokines/metabolism , Cytokines/blood , Cytokines/metabolism , HSP90 Heat-Shock Proteins/blood , HSP90 Heat-Shock Proteins/metabolism , Healthy Volunteers , Humans , Immunosuppressive Agents/therapeutic use , Inflammation/blood , Inflammation/drug therapy , Inflammation/metabolism , Inflammation/therapy , Muscle, Skeletal/metabolism , Myositis/blood , Myositis/drug therapy , Myositis/metabolism , Myositis/therapy
8.
Cesk Patol ; 58(1): 57-60, 2022.
Article in English | MEDLINE | ID: mdl-35387458

ABSTRACT

Uterus transplantation is a new experimental treatment method of absolute uterine factor infertility which affects 3-5% of infertile women. Absolute uterine factor infertility includes infertile women with agenesis or severe malformation of the uterus, several acquired uterine diseases causing infertility, and patients of fertile age after hysterectomy because of various causes. Uterus transplantation is considered a new method of assisted reproduction which allows women with absolute uterine factor infertility to have own biological offspring. However, uterus transplantation is considered a radical method of reproduction by some ethicists. Nevertheless, recent analysis of newborns from transplanted uterus has shown high level of childbirths of mature and near-to-term newborns and did not confirm increased risk for both babies and mothers. Therefore, together with gestational surrogacy and adoption, uterus transplantation is nowadays considered promising and unique solution for women with absolute uterine factor infertility. Similarly to other solid organ transplants, the pathologist should be an integral part of the multidisciplinary uterus transplantation research teams. The primary role of the pathologist is histopathological evaluation of rejection changes in the biopsy samples from the ectocervix of the uterine allografts that is based on the provisional scoring system suggested by Swedish pioneers in uterus transplantation research. As the word provisional suggests, this scoring system is continuously studied and the principles of the evaluation of rejection after uterus transplantation could be adjusted in the future.


Subject(s)
Infertility, Female , Organ Transplantation , Female , Humans , Infant, Newborn , Infertility, Female/etiology , Infertility, Female/surgery , Organ Transplantation/adverse effects , Organ Transplantation/methods , Pathologists , Uterus/abnormalities , Uterus/transplantation
9.
Cesk Patol ; 58(3): 126-134, 2022.
Article in English | MEDLINE | ID: mdl-36224034

ABSTRACT

The aim of the presented communication is to clearly inform the general professional public about the newly approved modifications in this classification, including the newly approved types of tumours. A significant change is the new grading system for these tumours, including the innovative involvement of tumour profiling at the molecular level in the system for determining the degree of tumour differentiation and the application of the principle of integrated diagnostics, i. e. the synthesis of available histopathological and molecular findings in CNS tumors.


Subject(s)
Central Nervous System Neoplasms , Central Nervous System Neoplasms/diagnosis , Humans , World Health Organization
10.
Cesk Patol ; 58(3): 135-137, 2022.
Article in English | MEDLINE | ID: mdl-36224035

ABSTRACT

The new WHO classification of CNS tumors is largely based on molecular diagnostic. Without molecular methods some entities can no longer be diagnosed. We are trying to show a rational approach to the CNS tumors diagnostics, which is based on conventional molecular methods such as RT-PCR, Sanger sequencing, MLPA, extended by the next generation sequencing (NGS) and methylation SNP array.


Subject(s)
Central Nervous System Neoplasms , High-Throughput Nucleotide Sequencing , Central Nervous System Neoplasms/diagnosis , Central Nervous System Neoplasms/genetics , Humans , Mutation
11.
Neuropathol Appl Neurobiol ; 47(6): 796-811, 2021 10.
Article in English | MEDLINE | ID: mdl-33942341

ABSTRACT

AIMS: Tuberous sclerosis complex (TSC) is a genetic disorder associated with dysregulation of the mechanistic target of rapamycin complex 1 (mTORC1) signalling pathway. Neurodevelopmental disorders, frequently present in TSC, are linked to cortical tubers in the brain. We previously reported microRNA-34a (miR-34a) among the most upregulated miRs in tubers. Here, we characterised miR-34a expression in tubers with the focus on the early brain development and assessed the regulation of mTORC1 pathway and corticogenesis by miR-34a. METHODS: We analysed the expression of miR-34a in resected cortical tubers (n = 37) compared with autopsy-derived control tissue (n = 27). The effect of miR-34a overexpression on corticogenesis was assessed in mice at E18. The regulation of the mTORC1 pathway and the expression of the bioinformatically predicted target genes were assessed in primary astrocyte cultures from three patients with TSC and in SH-SY5Y cells following miR-34a transfection. RESULTS: The peak of miR-34a overexpression in tubers was observed during infancy, concomitant with the presence of pathological markers, particularly in giant cells and dysmorphic neurons. miR-34a was also strongly expressed in foetal TSC cortex. Overexpression of miR-34a in mouse embryos decreased the percentage of cells migrated to the cortical plate. The transfection of miR-34a mimic in TSC astrocytes negatively regulated mTORC1 and decreased the expression of the target genes RAS related (RRAS) and NOTCH1. CONCLUSIONS: MicroRNA-34a is most highly overexpressed in tubers during foetal and early postnatal brain development. miR-34a can negatively regulate mTORC1; however, it may also contribute to abnormal corticogenesis in TSC.


Subject(s)
Astrocytes/metabolism , Brain/growth & development , MicroRNAs/genetics , Tuberous Sclerosis/genetics , Adolescent , Adult , Animals , Brain/pathology , Cerebral Cortex/pathology , Child , Child, Preschool , Female , Humans , Infant , Male , Mice, Inbred C57BL , MicroRNAs/metabolism , Neurons/pathology , Signal Transduction/genetics , Tuberous Sclerosis/complications , Tuberous Sclerosis/pathology , Young Adult
12.
Cytokine ; 137: 155350, 2021 01.
Article in English | MEDLINE | ID: mdl-33128920

ABSTRACT

BACKGROUND: Interleukin-35 (IL-35) is a recently described heterodimeric cytokine that belongs to the IL-12 family and consists of p35 (IL-12a) and EBI3 (IL-27b) subunits. The expression of IL-35 in humans is inducible in response to inflammatory stimuli. Increased IL-35 levels were documented in several autoimmune inflammatory diseases, suggesting a possible immunomodulatory role in their pathogenesis. OBJECTIVES: The aim of this study was to explore a potential role of IL-35 in the pathogenesis of idiopathic inflammatory myopathies (IIM) by studying the expression of IL-35 subunits in muscle biopsy samples and by evaluating serum levels of IL-35 and their association with disease activity in IIM patients. METHODS: The expression of IL-35 subunits was studied in serial sections of 9 muscle biopsy samples [4 polymyositis (PM), 5 dermatomyositis (DM)] and in 7 non-inflammatory control muscle biopsies. Serum levels of IL-35 were measured in 23 PM, 28 DM and 15 cancer associated myositis (CAM) patients as well as in 40 healthy controls. Disease activity was evaluated using the Myositis Disease Activity Assessment Tool (MDAAT) and by serum muscle enzymes. RESULTS: Expression of both IL-35 subunits was evident in the inflammatory infiltrates in IIM muscle biopsies, while no IL-35 expression was observed in control muscle samples. IL-35 serum levels were increased in all IIM patients compared to healthy controls [median 119.5 (range 32.1-1074.5) vs 36.2 (range 1.5-86.5) pg/ml, P < 0.001]. There were no differences in IL-35 serum levels between myositis subgroups (DM, PM or CAM). Serum IL-35 levels correlated significantly with physician's assessment of global (r = 0.29, p = 0.021), muscle (r = 0.30, p = 0.017) and extramuscular (r = 0.30, p = 0.016) disease activity as well as creatine kinase (r = 0.26, p = 0.044) and lactate dehydrogenase (r = 0.40, p = 0.003) levels. There was a significant correlation with pulmonary activity in patients with interstitial lung disease (r = 0.39, p = 0.037). Serum IL-35 correlated negatively with duration of treatment (r = -34, p = 0.009). CONCLUSIONS: IL-35 is overexpressed in inflammatory infiltrates in muscle tissue and serum in IIM patients and there is correlation with several disease activity parameters. These data suggest potential role of locally produced IL-35 in the pathogenesis of inflammatory myopathies.


Subject(s)
Interleukins/metabolism , Muscles/metabolism , Myositis/metabolism , Polymyositis/metabolism , Adolescent , Adult , Aged , Biopsy , Child , Female , Humans , Interleukins/blood , Male , Middle Aged , Muscles/pathology , Myositis/blood , Myositis/pathology , Polymyositis/blood , Polymyositis/pathology , Up-Regulation , Young Adult
13.
Clin Exp Rheumatol ; 39(5): 1021-1032, 2021.
Article in English | MEDLINE | ID: mdl-33124569

ABSTRACT

OBJECTIVES: The aim of this cross-sectional study was to explore the circulating and skeletal muscle expression of clusterin (CLU) in inflammatory myopathies (IIM) and its potential implication in pathogenetic mechanisms of the disease. METHODS: A total of 85 IIM patients and 86 healthy controls (HC) were recruited. In addition, 20 IIM patients and 21 HC underwent a muscle biopsy. Circulating CLU was measured by ELISA. Serum cytokine profile of patients and HC was assessed by Cytokine 27-plex Assay. Immunohistochemical localisation of CLU was assessed in 10 IIM and 4 control muscle tissue specimens. The expression of CLU and myositis related cytokines in muscle was determined by qPCR. RESULTS: Serum levels of CLU were significantly increased in IIM patients compared to controls (86.2 (71.6-99.0) vs. 59.6 (52.6-68.4) µg/mL, p<0.0001) and positively correlated with myositis disease activity assessment (MYOACT) (r=0.337, p=0.008), myositis intention-to-treat activity index (MITAX) (r=0.357, p=0.004) and global disease assessment evaluated by physician (r=0.309, p=0.015). Moreover, serum CLU correlated with cytokines and chemokines involved in IIM and their combined effect on disease activity was revealed by multivariate redundancy analysis. In muscle tissue, CLU mRNA was increased in IIM patients compared to controls (p=0.032) and CLU accumulated in the cytoplasm of regenerating myofibres. CONCLUSIONS: We suggest that the up-regulation of clusterin in circulation and skeletal muscle of IIM patients may be an inflammation and atrophy induced response of the organism intended to limit the environment, favouring further muscle damage.


Subject(s)
Clusterin , Myositis , Clusterin/genetics , Cross-Sectional Studies , Cytokines , Humans , Muscle, Skeletal
14.
Brain ; 143(1): 131-149, 2020 01 01.
Article in English | MEDLINE | ID: mdl-31834371

ABSTRACT

Tuberous sclerosis complex (TSC) is an autosomal dominantly inherited neurocutaneous disorder caused by inactivating mutations in TSC1 or TSC2, key regulators of the mechanistic target of rapamycin complex 1 (mTORC1) pathway. In the CNS, TSC is characterized by cortical tubers, subependymal nodules and subependymal giant cell astrocytomas (SEGAs). SEGAs may lead to impaired circulation of CSF resulting in hydrocephalus and raised intracranial pressure in patients with TSC. Currently, surgical resection and mTORC1 inhibitors are the recommended treatment options for patients with SEGA. In the present study, high-throughput RNA-sequencing (SEGAs n = 19, periventricular control n = 8) was used in combination with computational approaches to unravel the complexity of SEGA development. We identified 9400 mRNAs and 94 microRNAs differentially expressed in SEGAs compared to control tissue. The SEGA transcriptome profile was enriched for the mitogen-activated protein kinase (MAPK) pathway, a major regulator of cell proliferation and survival. Analysis at the protein level confirmed that extracellular signal-regulated kinase (ERK) is activated in SEGAs. Subsequently, the inhibition of ERK independently of mTORC1 blockade decreased efficiently the proliferation of primary patient-derived SEGA cultures. Furthermore, we found that LAMTOR1, LAMTOR2, LAMTOR3, LAMTOR4 and LAMTOR5 were overexpressed at both gene and protein levels in SEGA compared to control tissue. Taken together LAMTOR1-5 can form a complex, known as the 'Ragulator' complex, which is known to activate both mTORC1 and MAPK/ERK pathways. Overall, this study shows that the MAPK/ERK pathway could be used as a target for treatment independent of, or in combination with mTORC1 inhibitors for TSC patients. Moreover, our study provides initial evidence of a possible link between the constitutive activated mTORC1 pathway and a secondary driver pathway of tumour growth.


Subject(s)
Astrocytoma/genetics , Brain Neoplasms/genetics , Extracellular Signal-Regulated MAP Kinases/genetics , MAP Kinase Signaling System/genetics , MicroRNAs/metabolism , RNA, Messenger/metabolism , Tuberous Sclerosis/genetics , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Adolescent , Adult , Astrocytes/drug effects , Astrocytes/metabolism , Astrocytoma/etiology , Astrocytoma/metabolism , Brain Neoplasms/complications , Brain Neoplasms/metabolism , Butadienes/pharmacology , Child , Child, Preschool , Enzyme Inhibitors/pharmacology , Extracellular Signal-Regulated MAP Kinases/antagonists & inhibitors , Extracellular Signal-Regulated MAP Kinases/metabolism , Female , Gene Expression Profiling , Guanine Nucleotide Exchange Factors/genetics , Guanine Nucleotide Exchange Factors/metabolism , High-Throughput Nucleotide Sequencing , Humans , Infant , Intracellular Signaling Peptides and Proteins/genetics , Intracellular Signaling Peptides and Proteins/metabolism , Male , Mechanistic Target of Rapamycin Complex 1 , Nitriles/pharmacology , RNA-Seq , Sequence Analysis, RNA , Tuberous Sclerosis/complications , Tuberous Sclerosis Complex 1 Protein/genetics , Tuberous Sclerosis Complex 2 Protein/genetics , Tumor Cells, Cultured , Young Adult
15.
Cesk Patol ; 57(4): 221-225, 2021.
Article in English | MEDLINE | ID: mdl-35042355

ABSTRACT

Symplastic haemangioma is a rare vascular tumor presented with regressive and degenerative atypia in stromal cells. Its morphology represents a challenge in classification of vascular tumors, regarding their biological behaviour in particular. We present a case report of a 47-years-old female with a history of left-sided breast adenocarcinoma treated by resection followed by adjuvant chemotherapy and radiotherapy. Three years after the primary diagnosis a tumorous mass appeared in the region of upper margin of left scapula, in subcutaneous tissues and the trapezius muscle. Histologically, the tumor was formed by multiple blood vessels of varied diameter and wall thickness. Endothelial lining was bland, without atypia; thromboses were observed in vascular spaces. In the interstitium, a population of spindle and pleomorphic cells with distinctive atypia and bizarre nuclei was found. These cells showed positivity in immunohistochemical expression of smooth muscle actin, further extensive immunohistochemistry including cytokeratines was negative. Mitoses were absent, proliferating activity was minimal. Signs of infiltrative growth pattern were not found and the tumor lacked hallmarks of malignant behaviour. A diagnosis of symplastic haemangioma was established. Above mentioned atypical stromal cells show myofibroblastic and sporadically smooth muscle differentiation. Their atypical appearence is associated with degenerative alterations similar to changes in leiomyomas with bizarre nuclei or ancient schwannomas. Etiopathogenesis of these changes is not clear, there are hypotheses considering long-lasting persistence of the lesion, regression of ischaemic or postinflammatory origin, or, like in our case, postirradiative degeneration. Differential diagnosis of symplastic haemangioma is widespred and contains many histological entities of variant histogenesis and biological potential. For proper classification, an extensive investigation including immunohistochemistry, clinical and anamnestic data and imaging methods is necessary.


Subject(s)
Hemangioma , Leiomyoma , Skin Neoplasms , Diagnosis, Differential , Female , Hemangioma/radiotherapy , Humans , Immunohistochemistry , Middle Aged
16.
Cesk Patol ; 57(3): 136-143, 2021.
Article in English | MEDLINE | ID: mdl-34551560

ABSTRACT

Molecular assays for translocation detection in different tumors have gradually been incorporated into routine diagnostics. However, conventional methods such as fluorescence in situ hybridization (FISH) and reverse transcriptase-PCR come with several drawbacks. Next-generation sequencing (NGS) can provide in-depth detection of numerous gene alterations. The anchored multiplex PCR assay proved to be a fast and easy-to-analyze approach for routine diagnostics laboratories. Next-generation sequencing-based anchored multiplex PCR technique (Archer FusionPlex Panels) is beneficial in both diagnosis for patient care and in identification of a novel fusion breakpoint in tumors. NGS is useful in identifying targetable molecular changes (point mutations, fusion genes, etc.) in tumors that can serve as a rationale for inclusion of patients with advanced disease in ongoing clinical trials and allow for better risk stratification.


Subject(s)
High-Throughput Nucleotide Sequencing , Neoplasms , Humans , In Situ Hybridization, Fluorescence , Multiplex Polymerase Chain Reaction , Neoplasms/diagnosis , Neoplasms/genetics , Translocation, Genetic
17.
Cesk Patol ; 57(3): 154-160, 2021.
Article in English | MEDLINE | ID: mdl-34551564

ABSTRACT

Examination of changes in the methylation profile of DNA in cancer is currently used to determine the diagnosis or prognostic and predictive biomarkers. It complements histological or molecular biological examinations. At the same time, it helps to identify new diagnostic groups and subgroups. Currently, this diagnosis is most common in brain tumors, where it has become a routine examination. The established methylation profile may help even where the diagnosis or subgroup classification of the disease cannot be determined in any other way, as is the case with medulloblastoma.


Subject(s)
Brain Neoplasms , Cerebellar Neoplasms , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Brain Neoplasms/diagnosis , Brain Neoplasms/genetics , Cerebellar Neoplasms/genetics , DNA Methylation , Humans , Prognosis
18.
BMC Neurol ; 20(1): 359, 2020 Sep 24.
Article in English | MEDLINE | ID: mdl-32972372

ABSTRACT

BACKGROUND: Immune-mediated mechanisms substantially contribute to the Rasmussen encephalitis (RE) pathology, but for unknown reasons, immunotherapy is generally ineffective in patients who have already developed intractable epilepsy; overall laboratory data regarding the effect of immunotherapy on patients with RE are limited. We analyzed multiple samples from seven differently treated children with RE and evaluated the effects of immunotherapies on neuroinflammation. Immunotherapy was introduced to all patients at the time of intractable epilepsy and they all had to undergo hemispherothomy. METHODS: Immunohistochemistry, flow cytometry, Luminex multiplex bead and enzyme-linked immunosorbent assay techniques were combined to determine: 1) inflammatory changes and lymphocyte subpopulations in 45 brain tissues; 2) lymphocyte subpopulations and the levels of 12 chemokines/cytokines in 24 cerebrospinal fluid (CSF) samples and 30 blood samples; and 3) the dynamics of these parameters in four RE patients from whom multiple samples were collected. RESULTS: Sustained T cell-targeted therapy with cyclophosphamide, natalizumab, alemtuzumab, and intrathecal methotrexate (ITMTX), but not with azathioprine, substantially reduced inflammation in brain tissues. Despite the therapy, the distributions of CD8+ T cells and the levels of C-X-C motif ligand (CXCL) 10, CXCL13, and B cell activating factor (BAFF) in patients' CSF remained increased compared to controls. A therapeutic approach combining alemtuzumab and ITMTX was the most effective in producing simultaneous reductions in histopathological inflammatory findings and in the numbers of activated CD8+ T cells in the brain tissue, as well as in the overall CD8+ T cell population and chemokine/cytokine production in the CSF. CONCLUSIONS: We provide evidence that various T cell-targeted immunotherapies reduced inflammation in the brains of RE patients. The observation that intractable epilepsy persisted in all of the patients suggests a relative independence of seizure activity on the presence of T cells in the brain later in the disease course. Thus, new therapeutic targets must be identified. CXCL10, CXCL13 and BAFF levels were substantially increased in CSF from all patients and their significance in RE pathology remains to be addressed.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , Encephalitis/therapy , Immunotherapy/methods , Brain/pathology , Child , Child, Preschool , Cytokines/immunology , Encephalitis/pathology , Enzyme-Linked Immunosorbent Assay , Female , Humans , Inflammation/therapy , Male
19.
Cesk Patol ; 56(2): 95-98, 2020.
Article in English | MEDLINE | ID: mdl-32493026

ABSTRACT

Amoebic colitis represents a common parasitic infection in developing countries. In western world, it is encountered only sporadically. The clinical presentation is usually non-specific, non-invasive laboratory tests are often false negative and endoscopic and histopathological appearance may mimic other illnesses, especially Crohns disease. The disease therefore harbours a huge risk of misdiagnosing and a proper diagnosis is usually challenging. We present a case of an amoebic colitis with Crohn-like features and negative parasitological testing in a 53-years-old woman, in which the final diagnosis was established on the basis of its histopathological examination.


Subject(s)
Crohn Disease , Dysentery, Amebic , Crohn Disease/complications , Crohn Disease/diagnosis , Crohn Disease/pathology , Diagnosis, Differential , Dysentery, Amebic/complications , Dysentery, Amebic/diagnosis , Dysentery, Amebic/pathology , Female , Humans , Middle Aged
20.
Prostate ; 79(2): 126-139, 2019 02.
Article in English | MEDLINE | ID: mdl-30256431

ABSTRACT

BACKGROUND: Prostate-specific membrane antigen (PSMA), also known as glutamate carboxypeptidase II (GCPII), is an important diagnostic and therapeutic target in prostate cancer. PSMA/GCPII is also expressed in many healthy tissues, but its function has only been established in the brain and small intestine. Several research groups have attempted to produce PSMA/GCPII-deficient mice to study the physiological role of PSMA/GCPII in detail. The outcomes of these studies differ dramatically, ranging from embryonic lethality to production of viable PSMA/GCPII-deficient mice without any obvious phenotype. METHODS: We produced PSMA/GCPII-deficient mice (hereafter also referred as Folh1-/- mice) by TALEN-mediated mutagenesis on a C57BL/6NCrl background. Using Western blot and an enzyme activity assay, we confirmed the absence of PSMA/GCPII in our Folh1-/- mice. We performed anatomical and histopathological examination of selected tissues with a focus on urogenital system. We also examined the PSMA/GCPII expression profile within the mouse urogenital system using an enzyme activity assay and confirmed the presence of PSMA/GCPII in selected tissues by immunohistochemistry. RESULTS: Our Folh1-/- mice are viable, breed normally, and do not show any obvious phenotype. Nevertheless, aged Folh1-/- mice of 69-72 weeks exhibit seminal vesicle dilation, which is caused by accumulation of luminal fluid. This phenotype was also observed in Folh1+/- mice; the overall difference between our three cohorts (Folh1-/- , Folh1+/- , and Folh1+/+ ) was highly significant (P < 0.002). Of all studied tissues of the mouse urogenital system, only the epididymis appeared to have a physiologically relevant level of PSMA/GCPII expression. Additional experiments demonstrated that PSMA/GCPII is also present in the human epididymis. CONCLUSIONS: In this study, we provide the first evidence characterizing the reproductive tissue phenotype of PSMA/GCPII-deficient mice. These findings will help lay the groundwork for future studies to reveal PSMA/GCPII function in human reproduction.


Subject(s)
Glutamate Carboxypeptidase II/deficiency , Membrane Glycoproteins/deficiency , Seminal Vesicles/enzymology , Seminal Vesicles/pathology , Aging/metabolism , Aging/pathology , Animals , Antigens, Surface/genetics , Antigens, Surface/metabolism , Glutamate Carboxypeptidase II/genetics , Glutamate Carboxypeptidase II/metabolism , Humans , Immunohistochemistry , Male , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Mice , Mice, Inbred C57BL
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